Book of abstracts · Topic: Theme A: β-Amyloid Diseases / A1.a. Disease Mechanisms,...

P001 / #1447

Topic: Theme A: β-Amyloid Diseases / A1.a. Disease Mechanisms, Pathophysiology: Abeta aggregation, protein misfolding

GENOME-WIDE ASSOCIATION STUDIES, FUNCTIONAL GENOMICS AND CELL BIOLOGY IDENTIFY TBC1D5 AS AN ALZHEIMER’S DISEASE-RISK GENE AND UNCOVER ITS ROLE IN OAΒ CLEARANCE.

Lecture Title:

N. Albargothy1, S.-Y. Won2, T. Gunasekaran3, B. Choi2, S.H. Jang2, C. Lim2, K.S. Cho2, K. Lee4, M. Hallbeck1 1Linkoping University, Department Of Clinical And Experimental Medicine, Division Of Neurobiology And Clinical Pathology, Linkoping, Sweden, 2Konkuk University, Department Of Biological Sciences, Seoul, Korea, Republic of, 3Chosun University, Gwangju Alzheimer's Disease And Related Dementias Cohort Cente, Gwangju, Korea, Republic of, 4Chosun University, Department Of Biomedical Science, Seoseok-dong, Korea, Republic of

Aims: The prevalence of Alzheimer’s disease (AD) along with its social and economic burden continues to rise worldwide. Increasing evidence points towards smaller, pre-fibrillary aggregates, known as beta-amyloid oligomers (oAβ), as the primary cause of disease. We identified TBC1D5 as an AD risk gene from multiple resampled GWAS in the Korean population followed by confirmational functional genomic screening in a Drosophila AD model. Further, we investigated the role of this key regulator of intracellular trafficking and vesicle transport in the clearance of oAβ in differentiated neuron-like cells. Methods: We established a stable cell line lacking TBC1D5 by selectively inhibiting the human orthologue of this endocytic gene using siRNA in differentiated neuron-like SH-SY5Y cells. We then investigated the effect of TBC1D5 knockdown on oAβ uptake, processing and clearance after 3, 24 and 48 hours post-treatment with oAβ. Results: The levels of TBC1D5 are significantly reduced in the cortex of human AD patients’ brains compared to that of age-matched controls and is associated with temporal lobe atrophy. Knockdown of TBC1D5 also increases oAβ accumulation in neuronal-like cells, decreases clearance of oAβ internalised from extracellular milieu and increases Aβ-induced cytotoxicity. Conclusions: Our results suggest that endocytosis is a major cellular pathway associated with AD, and that down-regulation of molecular components of endocytic machinery may be involved in the pathological mechanisms underlying AD. Modulators of endocytic proteins, like TBC1D5, may serve as effective therapeutic targets to enhance the proper recycling and clearance of toxic oAβ to slow down disease progression.

P002 / #655


HYDROXYPROPYL-METHYLCELLULOSE DERIVATIVE PREVENTS AMYLOID-BETA ACCUMULATION, NEUROINFLAMMATION, AND APOPTOTIC NEURODEGENERATION IN A MOUSE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

T. Ali1, A. Klein1, A. Vu1, K. Doh-Ura2, S. Gilch1 1University of Calgary, Comparative Biology And Experimental Medicine, CALGARY, Canada, 2Tohoku University Graduate School of Medicine, Neurochemistry, Sendai, Japan

Aims: Polysaccharides such as cellulose ether (CE) formulations are emerging as potential therapeutics for neurodegenerative diseases associated with protein misfolding. The main objective of our study was to test the therapeutic effects of the CE (TC-5RW), a modified hydroxypropyl-methylcellulose in the 5xFAD transgenic mouse model of Alzheimer’s disease (AD). Methods: Two groups of 5xFAD mice were used, one group was treated with only a single subcutaneous dose of TC-5RW (4g/kg) at the age of 6 weeks. Mice of both groups were sacrificed at 10 months and brains were collected for immunoblotting and confocal microscopy analyses of Aβ, GFAP and caspase-3. Additionally, we applied Thioflavin-T assay to analyse Ab aggregation kinetics, and tested Ab toxicity on mouse neuroblastoma (N2a) cells by MTT assay. Results: The immunoblotting results revealed that TC-5RW reduced the Aβ level in the brain homogenates of TC-5RW-treated 5xFAD mice compared to control 5xFAD mice. Interestingly, the immunofluorescence assay using Aβ6E10 antibody further indicated that TC-5RW reduced the Aβ plaque burden in the hippocampi and cortical tissues. TC-5RW attenuated astrogliosis in hippocampi and cortical tissues, shown by analysis of GFAP reactivity. Furthermore, reduced levels of caspase-3 were found by immunoblot in brain homogenates of TC-5RW treated mice, indicating alleviation of apoptotic neurodegeneration compared to control 5xFAD mice. The in vitro Thioflavin-T and MMT results indicated that TC-5RW inhibited Aβ aggregation and toxicity. Conclusions: Overall, our in vivo and in vitro results suggest that CE based compounds might be valuable and emerging therapeutics for the prevention and treatment of AD.

P003 / #778


APP ACCUMULATIONS AROUND DENSE-CORE AMYLOID PLAQUES AS A NOVEL HALLMARK OF ALZHEIMER’S DISEASE

Lecture Title:

G. Barthet1, T. Jorda1, M. Petrel1, V. Kouskoff1, F. Cordelières1, S. Frykman2, U. Müller3, C. Mulle1 1IINS, Cnrs - University Of Bordeaux, Bordeaux, France, 2Karolinska Institutet, Dept Of Neurobiology, Care Sciences And Society, Stockholm, Sweden, 3Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Bioinformatics And Functional Genomics, Heidelberg, Germany

Aims: In Alzheimer’s disease (AD), a central role is given to the extracellular deposition of Aβ peptides, remotely produced by the proteolysis of the amyloid precursor protein (APP). This contrasts with other neurodegenerative diseases which are characterized by the intraneuronal aggregation of full-length proteins such as huntingtin, α-synuclein or TDP-43. Here, we aimed at investigating whether full-length APP accumulates intracellularly in AD. Methods: We performed immunofluorescent co-labelings in human hippocampal sections followed by quantitative analyses of hallmarks density, intensity and colocalization. Results: We report that in the human AD hippocampus, APP remarkably accumulates in the surrounding of dense-core amyloid plaques. These APP accumulations contain APP secretases necessary to produce Aβ peptides and are enriched in presynaptic proteins (Syt1, VAMP2). The Nter domain, but not the Cter domain of APP is enriched in the core of amyloid plaques uncovering a potential pathological role of the secreted APP-Nter in dense-core plaques. Ultrastructural analysis of APP accumulations reveals abundant multivesicular bodies containing presynaptic vesicles proteins and autophagosomal built-up of APP. Conclusions: Altogether, our data reveals a key role of intraneuronal full-length APP in AD mechanism and highlights APP accumulations as potential sources of Aβ and Nter peptides to fuel amyloid plaques.

P004 / #779


THE GENETIC LANDSCAPE FOR AMYLOID BETA NUCLEATION ACCURATELY DISCRIMINATES FAMILIAL ALZHEIMER’S DISEASE MUTATIONS

Lecture Title:

B. Bolognesi1,2, M. Seuma3, M. Badia3, A. Faure4, B. Lehner5 1Institute for Bioengineering of Catalunya, Phase Transitions In Health & Disease, Barcelona, Spain, 2IBEC, Phase Transitions In Health & Disease, barcelona, Spain, 3Institute for Bioengineering of Catalunya, Phase Transitions In Health & Disease, barcelona, Spain, 4Center for Genomic Regulation, Systems Biology, barcelona, Spain, 5Center for Genomic Regulation, Systems Biology, Barcelona, Spain

Aims: Amyloid plaques of the amyloid beta (Aß) peptide are a universal pathological hallmark of Alzheimer's disease (AD) and mutations in Aß cause familial forms of AD (fAD) . However, the molecular mechanism by which these mutations cause fAD remains unclear and the vast majority of mutations in Aß are variants of uncertain clinical significance. Here, we set out to identify all of the mutations in Aß that are likely to cause these rare forms of AD, by measuring their ability to nucleate amyloids in vivo. Methods: To do so, we have recently developed a deep mutagenesis method that quantifies the ability of thousands of protein variants to nucleate the formation of amyloid fibrils in vivo. By this means, we could quantify amyloid nucleation for more than 14,000 variants of Aß. Results: As a result, we provide the first description of how mutations alter the nucleation of any amyloid fibril. Our results reveal a modular organisation of mutational effects along the Aß sequence and uncover the role of charge and specific gatekeeper residues in the disordered N-terminus in preventing the nucleation of amyloids. Strikingly, the in vivo nucleation scores, unlike computational predictors and previous measurements, accurately discriminate all the known dominant fAD mutations. This suggests that accelerated nucleation is the fundamental molecular mechanism by which mutations cause fAD. Conclusions: Taken together, these results provide the first global picture of how sequence changes prevent and promote the nucleation of amyloid fibrils and provide a clinically-validated resource for the future interpretation of genetic variation in Aß.

P005 / #805


EFFECT OF ALZHEIMER’S DISEASE-RELATED Β-AMYLOID PEPTIDE CONFORMATIONS IN TRANSGENIC MOUSE MODELS OF Β-AMYLOIDOSIS

Lecture Title:

M. Célestine1, M. Jacquier-Sarlin2, E. Borel2, A.-S. Hérard1, A. Buisson2, M. Dhenain1 1Université Paris-Saclay, CEA, CNRS, Laboratoire des Maladies Neurodégénératives : mécanismes, thérapies, imagerie, Molecular Imaging Research Center, Fontenay-aux-Roses, France, 2Université Grenoble Alpes, Inserm, U1216, Grenoble Institut Neurosciences (gin), Grenoble, France

Aims: Early brain dysfunctions found in Alzheimer's disease (AD) are due to soluble pathological forms of b-amyloid peptide (Aβ). Although familial mutation-induced phenotypes are identified in AD patients, the in vivo effects of different Aβ variants are poorly understood. Here we characterized the effects of different Aβ variants on amyloid induction in a transgenic mouse model. Methods: We produced well-characterized amyloid variants: Aβ1-42, Aβ bearing the Icelandic mutation (A2T) or the Osaka mutation (E22D[1]). Then, two-month-old APP/PS1dE9 mice were inoculated with these variants in the dentate gyrus (n=6/per group). At 4 months post-inoculation, behavioral tests and functional brain connectivity (FC) study was performed. We assessed Aβ oligomerization profiles and cerebral amyloid load. [1] Tomiyama et al., Ann Neurol, 2008 Results: Wild type and Osaka-Aβ induced memory impairment (Fig. A) and loss of FC in mice hippocampus (Fig. B). Also, it led to increased 6-mer and 12-mer amyloid forms at inoculation site (Fig. C) but also at distance in the cortex. This suggests that Aβ variants differently modulate Aβ metabolism and aggregation. Amyloid plaque assessment showed an Osaka-Aβ-induced increase in amyloid load in the hippocampus (Fig. D) and its connected regions.

Conclusions: A single inoculation of different Aβ variants in APP/PS1dE9 mice expressing wild type Aβ is sufficient to modulate cognitive decline, Aβ forms occurrence and amyloid load. This suggests that Aβ variants specifically interact with the cerebral environments to induce different toxic downstream events.

P006 / #1144


THE LUMINESCENT CONJUGATED OLIGOTHIOPHENE H-FTAA ATTENUATES TOXICITY OF THE AMYLOID-BETA PEPTIDE WITH THE ARCTIC MUTATION

Lecture Title:

L. Sandin1, S. Sjödin1, A.-C. Brorsson2, K. Kågedal1, L. Civitelli3 1Linkoping University, Department Of Clinical And Experimental Medicine, Linkoping, Sweden, 2Linkoping University, Department Of Physics, Chemistry And Biology, Linkoping, Sweden, 3University of Oxford, Nuffield Department Of Clinical Neurosciences, Oxford, United Kingdom

Aims: The prevailing opinion is that prefibrillar β-amyloid (Aβ) species, rather than end-stage amyloid fibrils cause neuronal dysfunction in Alzheimer’s disease, although the mechanisms behind Aβ neurotoxicity remain to be elucidated. Luminescent conjugated oligothiophenes (LCOs) have spectral properties upon binding to amyloid proteins and have previously been reported to change the toxicity of Aβ1-42 and prion protein. The aim of this study was to investigate whether the LCOs h-FTAA and p-FTAA could change the toxicity of Aβ with the Arctic mutation (AβArc). Methods: A panel of cell biology techniques as well as biophysical methods i.e. Thioflavin T (ThT) assay, transmission electron microscopy and binding assays were used. Results: Cell viability assays demonstrated that AβArc toxicity on neuroblastoma cells declined with increased time of aggregation. Interestingly, h-FTAA but not p-FTAA, rescued the AβArc-mediated toxicity. Aggregation kinetics of AβArc using ThT, h-FTAA and p-FTAA showed differences after 5 h, which suggest that ThT, h-FTAA and p-FTAA may affect the aggregation process in different ways. Furthermore, a binding assay of AβArc bound to either h-FTAA or p-FTAA demonstrated different binding affinity of h-FTAA and p-FTAA to AβArc which might explain the diverse protective ability of h-FTAA and p-FTAA to AβArc-mediated toxicity. Conclusions: In conclusion, we show that h-FTAA but not p-FTAA attenuates toxicity of AβArc. This protection probably depends on specific interactions between h-FTAA and AβArc that cannot be formed by p-FTAA. These results indicate that h-FTAA might have a therapeutic potential for familial Alzheimer’s disease caused by the Arctic mutation.

P007 / #1406


MIR-277 TARGETS HID TO AMELIORATE AΒ42-MEDIATED NEURODEGENERATION IN DROSOPHILA EYE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

P. Deshpande1, C.-Y. Chen2, C. Yeates1, C.-H. Chen2,3, M. Kango-Singh1,4,5,6, A. Singh1,4,5,6,7 1University of Dayton, Department Of Biology, Dayton, United States of America, 2Institution of Molecular and Cellular Biology , National Taiwan University, Molecular And Cellular Biology, Taipei, Taiwan, 3National Institute of Infectious Diseases and Vaccinology, National Health Research Institutes, Infectious Diseases And Vaccinology, Miaoli, Taiwan, 4University of Dayton, Premedical Program, Dayton, United States of America, 5University of Dayton, Center For Tissue Regeneration & Engineering (trend), Dayton, United States of America, 6University of Dayton, Integrative Science And Engineering (ise), Dayton, United States of America, 7Indiana State University, Center For Genomic Advocacy (tcga), Terre Haute, United States of America

Aims: Alzheimer’s disease (AD), an age-related progressive neurodegenerative disorder, exhibits reduced cognitive functions with no cure to date. One of the reasons for AD is the extracellular accumulation of Amyloid-beta 42 (Aβ42) plaques. Accumulation of Aβ42 plaque(s) triggers aberrant signaling resulting in neuronal cell death by an unknown mechanism(s). microRNAs regulate the gene expression of the components which are involved in signaling pathways. We wanted to look for microRNAs involved in neurodegeneration Methods: We misexpressed human Aβ42 in the developing retina of Drosophila, which exhibits AD-like neuropathology. We screened for microRNA which post-transcriptionally regulates expression of genes by degrading mRNA of the target genes. In a forward genetic screen with candidate miRNAs, we identified mir-277 as a genetic modifier of Aβ42-mediated neurodegeneration. Results: Gain-of-function of mir-277 rescues Aβ42 mediated neurodegeneration whereas loss-of-function of mir-277 enhances Aβ42 mediated neurodegeneration. Moreover, misexpression of higher levels of mir-277 in the GMR>Aβ42 background restores the retinal axonal targeting indicating functional rescue. Furthermore, we have identified head involution defective (Hid) as one of the targets of mir-277 by Fly TargetScan and validated by luciferase assay and qPCR. The hid transcript levels are decreased by one third when mir-277 is misexpressed in the GMR>Aβ42 background in comparison to the GMR>Aβ42 fly model. Conclusions: Hence, here we provide a mechanism of how mir-277 modulates Aβ42 mediated neurodegeneration by regulating hid transcript levels and demonstrate its neuroprotective role in Aβ42-mediated neuropathology.

P008 / #1439


AΒ PEPTIDES FORM ION CHANNELS IN PHOSPHOLIPID BILAYER MEMBRANES BY THE MEDIATION OF GANGLIOSIDE AT LOW CONCENTRATION

Lecture Title:

N. Dokholyan, D. Zhang, J. Wang, M. Swulius Penn State College of Medicine, Pharmacology, Hummelstown, United States of America

Aims: Alzheimer’s disease (AD) is the most common neurodegenerative disorder, mainly affecting the elderly. The pathophysiology of the disease is characterized by accumulation of extracellular plaques, predominantly comprised of amyloid beta (Aβ), and cytoplasmic neurofibrillary tangles, mostly comprised of tau protein. The Aβ aggregation process is found strongly dependent on the concentration. Despite mounting evidence for the aggregation process of high Aβ concentration, little is known about the Aβoligomer formation at physiological concentrations, especially under the regulation of the cell membrane. Methods: Here, we use liposomes to probe the interaction between diverse membrane environments and the Aβ aggregation process. We find that Aβ interacts with membrane that is rich in monosialotetrahexosylganglioside (GM1), and GM1 clusters maintain Aβ oligomers’ stability, and promote the amyloid fibril formation. We also determine the regulation of membrane constituents on Aβ aggregation and investigate the formation of Aβ ion channels at low concentrations under the regulation of GM1. Results: Finally, we characterize the structure of Aβ ion channels by circular dichroism and mass spectrometry for computational modeling of the structure. Conclusions: The interrogation of the formation and regulation of Aβ channel structures on membranes provide new insight into the understanding of AD pathogenesis.

P009 / #1781


PROTEOMIC ANALYSIS OF NEUROTROPHIC AND NEUROPROTECTIVE EFFECTS OF PREGNENOLONE SULFATE IN ALZHEIMER’S DISEASE

Lecture Title:

F. El Bitar1, S. Abdulaziz1, A. Ayodele2 1King Faisal Specialist Hospital and Research Center, Department Of Genetics, Riyadh, Saudi Arabia, 2King Faisal Specialist Hospital and Research Center, Stem Cell And Tissue Re-engineering Program, Riyadh, Saudi Arabia

Aims: Background: Alzheimer’s disease (AD) is the most common cause of dementia in the elderly. The toxicity of b-amyloid (Ab) peptides is thought to be involved in neuronal damage and cognitive decline in this pathology. Multiple studies focused on role of neuroprotective molecules including Pregnenlone Sulfate (PREGS) to attenuate the toxic consequences of Ab peptides in the development of AD. Aims: Our objective is to investigate the proteomic analysis of neurotrophic and neuroprotective effects of PREGS on in vitro model of AD. Methods: Proteins were extracted from neuroblastoma B104 cells pretreated either with PREGS alone or PREGS before Aβ25–35. Extracted proteins were subjected to proteomic analysis using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) to generate expression proteomics data based on both qualitative and quantitative differential expression changes between the sample groups. Results: The proteomic analysis revealed 260 significantly differentially expressed proteins (ANOVA test p< 0.05, >1.5-fold change (FC)) in 2 groups including Aβ25–35 treated cells compared to PREGS and Aβ25–35 treated ones (186 up and 74 down). As to control compared to PREGS treated cells, we found 38 significantly differentially expressed proteins (26 up and 12 down). Functional network analysis using ingenuity pathway analysis (IPA) revealed presence of AD related proteins such as NCAM1, CDC42, RAC1, RHOG, HSPA5, GNB2l1 and TPM1. Conclusions: Conclusion: Thus, PREGS is neuroprotective and neurotrophic drug that could be promising in Alzheimer’s disease treatment. The continuity in examining the mechanism(s) underlying PREGS activity is promising towards identifying efficient therapy for AD.

P010 / #1443


FERRITIN IRON IS CHEMICALLY REDUCED IN THE PRESENCE OF AGGREGATING Β-AMYLOID

Lecture Title:

J. Everett1, J. Brooks2, F. Lermyte2, P. O'Connor3, P. Sadler3, J. Dobson4, J. Collingwood2, N. Telling1 1Keele University, School Of Pharmacy And Bioengineering, Stoke-on-Trent, United Kingdom, 2University of Warwick, School Of Engineering, Coventry, United Kingdom, 3University of Warwick, Department Of Chemistry, Coventry, United Kingdom, 4University of Florida, Department Of Materials Science And Engineering, Gainesville, United States of America

Aims: Low-oxidation-state (< 3+) iron phases associated with amyloid plaque pathology are implicated in Alzheimer’s disease pathogenesis, as they promote elevated redox activity and toxicity. The origin and mechanism for the formation of this iron remains unresolved, but may involve the interaction of the disease-related peptide β-amyloid with the iron storage protein ferritin. Here we have used synchrotron x-ray spectromicroscopy coupled with electron microscopy to establish the chemistry of β-amyloid/ferritin interactions. Methods: β-amyloid (1-42) was incubated with ferritin in a buffer medium modelled on cerebral spinal fluid. The resulting β-amyloid/ferritin aggregates were examined using scanning transmission x-ray microscopy (STXM) at Diamond Light Source beamline I08 and the Swiss Light Source PolLux beamline. Measurements were performed at the carbon K-edge to detect organic constituents, and the calcium and iron L-edges to determine the distribution and chemical state of these metals. The spatial resolution for x-ray spectromicroscopy was 30-50 nm. Results: STXM showed that the co-aggregation of β-amyloid and ferritin resulted in the conversion of ferritin’s inert ferric core into more reactive low-oxidation-states. The chemical composition of the in vitro aggregates formed in this study was shown to be analogous to primary components of amyloid plaque material extracted from the grey matter of Alzheimer's disease subjects. Conclusions: These findings implicate β-amyloid in the altered iron metabolism and increased oxidative stress observed in Alzheimer’s disease. Amyloid-associated iron phases have biomarker potential to assist with disease diagnosis and staging, and may represent targets for therapies designed to lower oxidative stress in Alzheimer’s tissue.

P011 / #1101


A POTENTIAL DEFENSE MECHANISM AGAINST AMYLOID DEPOSITION IN CEREBELLUM

Lecture Title:

S. Funamoto1, A. Shahnur1, M. Nakano2, S. Ishihara1, T. Miyasaka1, N. Kakuda1, T. Saito3, T. Saido4, M. Nishimura2 1Doshisha University, Neuropathology, Kyotanabe, Japan, 2Shiga Unversity of Medical Science,

Neurology, Otsu, Japan, 3Nagoya City University, Department Of Neurocognitive Science, Kawasumi−１,

Japan, 4RIKEN, Center For Brain Science, Wako-shi, Japan

Aims: Senile plaques are observed mainly in cerebral cortex (CX), but less in cerebellum (CB). Tg2576 mice also exhibit less Aβ deposition in CB, although APP is overexpressed under the control of prion promoter. APPNL-G-F KI mice also show less Aβ deposition in CB, despite the fact that these mice produce aggregation-prone arctic Aβ42 in whole brain. In this study, we explored to figure out the potential mechanism preventing Aβ deposition in CB. Methods: (1) In order to compare levels of Aβ between CX and CB, we performed microdialysis experiments on 4-month old APPNL-G-F KI mice. (2) We performed stereotactic injection of HiLyte Fluor555-labeled Aβ42 into brain tissues and examined Aβ diffusion and clearance. (3) We examined the presence of the labeled Aβ42 in deep cervical lymph nodes (DcLNs) as one of drainage routes. Results: (1) We detected no significant difference in ISF Aβ level between CX and CB. (2) Aβ diffusion area in CB was roughly three-times larger than that in CX right after injection. However, we observed 60% decrease in the Aβ diffusion area in CB after 72 h, while that of CX unchanged. (3) Aβ injected into CB was found in DcLNs within 2 h, while that in CX was faint. Conclusions: The level of ISF Aβ in CB is equivalent to that in CX. However, Aβ diffusion and clearance rates in CB is higher than those in CX, which leads to less Aβ deposition in CB.

P012 / #1070


ENDO-LYSOSOMAL ABETA CONCENTRATION AND PH ENABLE FORMATION OF ABETA OLIGOMERS THAT POTENTLY INDUCE TAU MISSORTING

Lecture Title:

W. Hoyer1, M. Schützmann1, F. Hasecke1, S. Bachmann2, M. Zielinski3, S. Hänsch4, G. Schröder3, H. Zempel2 1Heinrich Heine University Düsseldorf, Physical Biology, Düsseldorf, Germany, 2University of Cologne, Institute Of Human Genetics And Center For Molecular Medicine Cologne (cmmc), Cologne, Germany, 3Forschungszentrum Jülich, Institute Of Biological Information Processing (ibi-7) And Justruct: Jülich Center For Structural Biology, Jülich, Germany, 4Heinrich Heine University Düsseldorf, Department Of Biology, Center For Advanced Imaging (cai), Düsseldorf, Germany

Aims: Metastable oligomers of amyloid-β peptide (Aβ) are more effective than Aβ amyloid fibrils at triggering Alzheimer’s disease-related processes such as synaptic dysfunction and Tau pathology. Here we identify physiological conditions that promote Aβ oligomer (AβO) formation. Methods: The pH dependence of AβO formation was determined utilizing Aβ42 as well as the dimeric Aβ construct dimAβ that facilitates analysis of the oligomerization kinetics. AβO formation, clustering, and release were imaged by atomic force microscopy. The structure of the smallest AβOs was investigated by cryo-EM. The capacity of AβOs to bind to dendritic spines, to induce Tau missorting, and to impair neuronal function were studied in primary mouse neuronal cell cultures. Results: The rate of AβO assembly is accelerated 8,000-fold upon pH reduction from extracellular to endo-lysosomal pH, at the expense of amyloid fibril formation. The pH-induced promotion of AβO formation and the high endo-lysosomal Aβ concentration together enable extensive AβO formation of Aβ42 under physiological conditions. Exploiting the enhanced AβO formation of dimAβ we furthermore demonstrate targeting of AβOs to dendritic spines, potent induction of Tau missorting, a key factor in tauopathies, and impaired neuronal activity. Conclusions: The results suggest that the endosomal/lysosomal system is a major site for the assembly of pathomechanistically relevant AβOs.

P013 / #771


THE RELATIONSHIP BETWEEN THE PATH OF AMYLOID-Β FIBRIL FORMATION AND THEIR PROPERTIES.

Lecture Title:

M.T. Islam, K. Brännström, A. Gharibyan, A. Olofsson Umeå University, Medical Biochemistry And Biophysics, Umeå, Sweden

Aims: The aim of this study is to investigate how Aβ1–40 and Aβ1–42 isoforms interact to better understand how pathological fibril formation occurs in vivo. Methods: To learn the cross-incorporation of Aβ1–40 and Aβ1–42 monomeric peptides onto the fibrillar forms of Aβ1–40 and Aβ1–42 respectively, we performed cross-templating experiments using Surface plasmon resonance (SPR)-assay. To learn cross-nucleation between Aβ1–40 and Aβ1–42 in solution and the properties of cross-nucleated fibrils, we performed Thiaflavin-T (ThT-assay) that followed SPR assay. Results: SPR-assay indicates that Aβ1-42 monomers are easily incorporated onto the fibrillar form of Aβ1-

40 and they also acquire the properties of Aβ1-40. On the other hand, Aβ1-40 monomers are inhibited from incorporating onto the fibrillar form Aβ1-42. ThT assay indicates that fibrils derived via SCSN do not acquire the properties of the parental fibrils. Conclusions: In conclusion, there is an intrinsic barrier that prevents a more abundant Aβ1–40 variant from adopting the fibrillar properties of more cytotoxic Aβ1–42. Also, the path of fibril formation controls the transfer of fibrillar properties between amyloid-β fibrils.

P015 / #1142


MELATONIN INHIBIT THE APOPTOSIS BY JNK INHIBTION THROUGH IMPROVED MITOCHONDRIAL FUNCTION IN AD

Lecture Title:

R. Khatoon jamia hamdard, Toxicology, delhi, India

Aims: Alzheimers’s disease is one of the alarming neurodegenerative disease and it is of global concern.The hallmarks of the disease are the amyloid beta (Aβ) aggregation and presence of neurofibrillary tangles (NFTs). The interaction of Aβ with macromolecular targets affects the normal cellular functions. The amyloid peptide interaction with cellular surfaces may trigger the intracellular cascades of signalling. Interaction of Aβ with mitochondria leads to the generation of free radicals.Many studies have suggested the involvement of mitochondrial dysfunction in Alzheimer’s disease. Melatonin prevents mitochondria stress and by means of activating the antioxidant systems it protects the death of neurons. Methods: Two different Elav Gal4 and UAS-Ab42 were used for the study. The F1 offspring of this strain were divided into four Drosophila each: (1) control, (2) 0.43mM melatonin, (3) 0.43mM melatonin + AD (4) AD. Drosophila was concomitantly exposed to a diet containing melatonin for 30 days according to their respective groups. Results: We investigated that 0.43mM of melatonin significantly extended life span and improves the climbing ability of Drosophila owing to the antioxidant properties of melatonin. Melatonin also attenuated the decline of expression of p-jnk protein, mitochondrial membrane potential, mitochondrial ROS level and decreases caspase3 in Drosophila model of AD. Conclusions: These results strongly suggest the neuromodulatory effect of melatonin and that it is probably mediated via jnk inhibition and its potential to attenuate mitochondrial dysfunctions, against AD.

P016 / #1375


PRODUCTION OF AΒ HEXAMERS IN AD PATHOLOGY

Lecture Title:

P. Kienlen-Campard1, D. Vadukul1, C. Vrancx1, F. Perrin2, S. Constantinescu3, N. Papadopoulos3 1UCLouvain, Institute Of Neuroscience, Brussels, Belgium, 2Massachusetts General Hospital, Alzheimer's Disease Research Center, Boston, United States of America, 3UCLouvain, De Duve Institute, Brussels, Belgium

Aims: The formation of Aβ assemblies plays a crucial role in AD pathology. Various Aβ oligomers ranging from dimers to dodecamers have been identified. Our aim was to characterize pathological Aβ oligomers produced by cells and investigate the molecular determinants involved in their formation. Methods: We expressed human APP amyloidogenic fragments (C99) or directly Aβ peptides in cell cultures. Immunoprecipitation combined to mass spectrometry was used for the characterization of Aβ assemblies. Nucleation properties and kinetics of Aβ assemblies were studied by in vitro. Engineered APP constructs and PS-deficient cell lines were used to investigate molecular determinants/conformations driving Aβ assembly and to relate them to specific γ-secretases. Results: Aβ hexamers were readily detectable in cell extracts and culture media of cells expressing C99, Aβ42 or longer isoforms. FAD mutations (central Aβ motif) trigger hexamer formation. Hexameric Aβ42 produced by cells displays nucleating properties which are dependent on the Aβ monomer availability. We assessed the profile of Aβ production in PS-deficient cell lines and found that Aβ hexamers were predominantly linked to PS2-dependent γ secretases. Strikingly, when we investigated APP conformations related to the formation of Aβ assemblies, we observed that a specific conformation of APP dimers strongly favors the release of hexameric-like Aβ. Conclusions: We identified the formation of hexamers in several Aβ enriched cellular contexts and familial AD (FAD) related mutations. Aβ42 hexamers produced by cells have intrinsic nucleation features underlying their pathological properties. The type of γ-secretase (PS1/PS2) and the processing lines controlled by APP conformations are critical in this process.

P017 / #521


EXTRACELLULAR VESICLES ARE ENRICHED IN OLIGOMERIC APP-CTFS IN AD MODELS

Lecture Title:

I. Lauritzen, A. Bécot, A. Bourgeois, R. Pardossi-Piquard, F. Checler Université Cote d'Azur-CNRS UMR7275, Institut De Pharmacologie Moléculaire Et Cellulaire, Valbonne, France

Aims: The β-secretase-derived APP C-terminal fragment (CTF) C99 is known to accumulate in endosomal and lysosomal compartments in Alzheimer disease (AD) mouse brains1,2. We found that γ-secretase inhibitor (D6) treatment led to further increased endolysosomal C99 levels but also to extracellular APP-CTF-associated immunostaining possibly reflecting extracellular vesicle (EV)-associated C99 and other APP-CTFs2. We here characterized these APP-CTFs. Methods: EVs were purified from media or brains from vehicle- or D6-treated C99 or APP expressing cells/mice and analyzed by immunoblot. Combined pharmacological, immunological and genetic approaches (presenilin invalidation and C99 dimerization mutants (GXXXG)) were used to characterize EV APP-CTFs. Subcellular APP-CTF localization was determined by immunocytochemistry. Results: EVs from both AD models contained APP-CTFs and intriguingly D6-treatment led to not only further increased C99 and C99-derived C83 levels but also to the presence of higher molecular weight (HMW) APP-CTFs poorly detected in cell extracts. We established that these HMW APP-CTFs correspond to oligomeric APP-CTFs composed of a mix of C99 and/or C83. Immunocytochemistry showed that oligomers were confined to endosomes and lysosomes, thus explaining the selective recovery of HMW APP-CTFs in EVs. Our data proposed that besides favoring APP-CTF oligomerization by preventing C99 proteolysis, γ-secretase inhibiton also led to a defective SorLA-mediated retrograde transport of HMW APP-CTFs from endosomes to the TGN. Conclusions: We show the presence of oligomeric APP-CTFs in AD models, the levels of which are selectively enriched in endolysosomal compartments including exosomes3. 1. Lauritzen et al, J. Neuroscience 2012 2. Lauritzen et al, Acta Neuropath. 2016 3. Lauritzen et al, Translational neurodegeneration, 2019

P018 / #1394


BRAIN EXPRESSION AND PROCESSING OF THE AMYLOID PRECURSOR PROTEIN IS UNAFFECTED BY APOLIPOPROTEIN E GENOTYPE

Lecture Title:

M. Novy, S. Newbury, J. Morales-Corraliza, M. Alldred, S. Ginsberg, P. Mathews Nathan Kline Institute, Center For Dementia Research, Orangeburg, United States of America

Aims: The apolipoprotein E ε4 allele (APOE4) is the single greatest genetic risk factor for Alzheimer’s disease, and of its many potential deleterious effects APOE4 have been suggested to impact the expression and processing of the amyloid precursor protein (APP). In this study, we examined in the brains of mice the impact of APOE genotype on the expression of the endogenous APP gene as well as in vivo APP and processing. Methods: Endogenous murine APP was examined in the brains of 12-month-old mice humanized and homozygous for the ε2, ε3, or ε4 APOE alleles. APP mRNA levels were determined by quantitative PCR analysis. Soluble Aβ40 and Aβ42 levels were measured by murine Aβ sandwich ELISA. The levels of full length APP, cell-associated APP C-terminal fragments, and soluble APP fragments were determined by Western blot analysis and immunoprecipitation. Results: APP gene expression and the levels of APP protein and APP fragments were not affected by APOE genotype. The levels of both α- and β-cleaved soluble APP fragments were similar across genotypes, as were the levels of the cell-associated α- and β-cleaved C-terminal fragments. Expression of the three APOE alleles did not alter the levels of brain Aβ derived from the endogenous APP in 12-month-old mice. Conclusions: While expression of APOE4 is known to impact Aβ clearance, seeding, and amyloid deposition, in a model without APP overexpression and with endogenous levels of APOE expression, brain expression and processing of APP is unaffected by the expression of the three human APOE alleles.

P019 / #397


TRIFUNCTIONAL NANOPARTICLE DESIGN TO RESCUE NEUROSYSTEM FAILURE CAUSED BY PROTEIN AGGREGATION (TRINADES)

Lecture Title:

V. Redondo, T. Schrader University of Duisburg-Essen, Organic Chemistry, Essen, Germany

Aims: Alzheimer’s disease (AD) is the most prevalent form of dementia, with 50 million people affected around the world. Aβ is one of the proteins involved in AD, which can misfold and form plaques that collect between neurons and disrupt cell function. To this day, an effective treatment against Aβ deposition has yet to be found. With this challenge in mind, a trifunctional nanoparticle was designed to prevent the toxic effect of Aβ aggregation. Each nanoparticle is equipped with three elements: a selector, which binds to small neurotoxic Aβ oligomers; a breaker, which disrupts the existing β-sheet by dissociating its hydrogen bonds; an artificial protease, which cuts the absorbed protein molecules into smaller fragments, regenerating the original trifunctional state and therefore making the cycle catalytic. With this approach, the trifunctional nanoparticle is turned into a disaggregation nanomachine, which may completely abolish the aggregation propensity of Aβ and rescue cell viability.

Methods: A total organic synthesis was carried out to obtain β-sheet selectors, β-sheet breakers and artificial proteases. The effectivity of the artificial proteases to cleave different peptides will be evaluated by proteomic analysis using mass spectrometry. Finally, the synthesized ligands will be supported on gold nanoparticles obtained by Pulsed Laser Ablation in Liquids (PLAL). Results: All target molecules were successfully synthesized. The effectivity of the artificial proteases is currently being evaluated by proteomic analysis.

Conclusions: β-sheet selectors, β-sheet breakers and artificial proteases were successfully synthesized, which will be supported on gold nanoparticles to create a nanomachine against Aβ aggregation.

P020 / #1336


SULFONYLUREA RECEPTOR/SUR DEFICIENCY INCREASED VULNERABILITY TO NEURODEGENERATION IN DROSOPHILA MODELS OF ALZHEIMER’S DISEASE

Lecture Title:

M. Sekiya, X. Quan, Y. Sakakibara, S. Chikamatsu, K. Iijima National Center for Geriatrics and Gerontology, Department Of Alzheimer's Disease Research, Obu, Japan

Aims: Hippocampal sclerosis of aging (HS-aging) is a common senile dementia characterized by sever neuron loss and gliosis in the hippocampus, and clinically mimics Alzheimer’s disease (AD). Polymorphisms in GRN, TMEM106B and ABCC9 have previously shown the association with HS-Aging. However, the mechanisms and a molecular link between HS-aging and AD remain elusive. Sulfonylurea receptors (SURs: ABCC8/SUR1 and ABCC9/SUR2) are the regulatory subunits of ATP-sensitive potassium channels that coordinate energetic status and intracellular Ca2

+ levels in various cells. Mutations in SURs causes diabetes and cardiovascular diseases, both of which are known AD risks. Interestingly, single nucleotide polymorphism that increases the mRNA levels of ABCC9/SUR2 has been associated with a risk for HS-aging, suggesting that ABCC9/SUR2 might influence pathogenesis of AD as well as HS-aging. Methods: Using Drosophila as a model, we asked whether deficiency in Sur, a fly ortholog of SURs, is protective or detrimental to neuronal integrity during aging and under neurodegenerative conditions. Results: We found that Sur deficiency by itself did not affect age-associated changes in neuronal function and integrity. In contrast, Sur deficiency significantly worsened behavioral deficits and neurodegeneration by impairing energy metabolism in a fly model of amyloid-β42 toxicity. We also demonstrated that Sur deficiency significantly exacerbated tau-mediated neurodegeneration and AD-related tau phosphorylation. Conclusions: This study suggests that ABCC9/SUR2 plays neuroprotective roles in the pathogenesis of AD and that increased ABCC9/SUR2 might extend clinical onset of AD.

P021 / #1746


DEPLETION OF CAVEOLIN-1 IN TYPE-2 DIABETES INDUCES ALZHEIMER’S DISEASE PATHOLOGY

Lecture Title:

A. Shetti, J. Bonds, O. Lazarov University of Illinois at Chicago, Anatomy And Cell Biology, Chicago, United States of America

Aims: This study aimed to understand the role of endothelial enriched Caveolin-1 (Cav-1) in the development of the Alzheimer’s disease in type II diabetes. Methods: Brain samples from type II diabetes patients and age- matched healthy subjects, as well as hippocampal samples from type-II diabetic db/db (Leprdb) and wild type mice were examined for Cav-1 levels and AD- related pathology. Levels of Cav-1 were restored by stereotaxic injection of Cav-1 expressing adenovirus into the hippocampus of db/db mice. Learning and memory was assessed in db/db and wild type mice by novel object recognition test. Results: Here we show that levels of Cav-1 and its downstream signal endothelial nitric oxide synthase (eNOS) were reduced in the brains of T2DM patients compared to healthy aging. Cav-1 and eNOS levels inversely correlated with levels of β-amyloid (Aβ). Depletion of Cav-1 was recapitulated in the brains of db/db mice and corresponded with deficits in recognition memory, increased levels of amyloid precursor protein (APP), BACE-1 and hyperphosphorylated tau (p-tau) species. Importantly, we show that restoration of Cav-1 levels in the brains of db/db mice using adenovirus expressing Cav-1 (AAV-Cav-1) rescued learning and memory deficits and reduced pathology. Conclusions: Cav-1 regulates AD- linked proteins. Restoration of Cav-1 in the brains of T2DM mice can attenuate the development of AD- like pathology.

P022 / #512


AMYLOID-B SEEDING IN A SPORADIC MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

L. Trujillo-Estrada1, K. Do Huynh2, M. Minh Thu Nguyen2, A. Cheung2, J. Pham Tran2, C. Nuñez-Diaz1, S. Forner2, A. Martini2, C. Da Cunha2, M. Shahnawaz3, C. Soto3, I. Moreno Gonzalez1, A. Gutierrez1, F. Laferla2, D. Baglietto-Vargas1 1University of Malaga/CIBERNED/IBIMA, Cell Biology, Malaga, Spain, 2University of California, Irvine, Institute For Memory Impairments And Neurological Disorders, Irvine, United States of America, 3University of Texas Health Science Center at Houston, The Mitchell Center For Alzheimer’s Disease And Related Brain Disorders, Department Of Neurology, Houston, United States of America

Aims: Recent evidence indicates that Aβ can misfold and aggregate into seeds that structurally corrupt native proteins, mimicking a prion-like process. Several studies using FAD animal models have demonstrated that intracerebral infusion of brain extracts from APP-transgenic mice or AD patients induce Aβ deposition and cerebral amyloid angiopathy. To carry out most of these Aβ-seeding studies, APP-transgenic animal have been used. Nevertheless, it remains to be elucidated whether Aβ deposition can be induced by Abeta-seeds in a sporadic AD model that does not overexpress APP and produces wild type human Aβ. Methods: We used an innovative model to better understand the amyloidogenic events that occur in sporadic AD. This hAβ-KI model, expresses wild-type human Aβ under the control of the endogenous mouse APP gene. Aβ-seeds from AD patients (stage C) from the AD Research Center (UCI) were administered into 7-8-month-old hAβ-KI and as positive controls 3xTg-AD mice were employed. Results: We demonstrated that amyloid seeds can stimulate Aβ aggregations in 3xTg-AD and hAβ-KI models. We found that Aβ aggregates occur earlier in the 3xTg-AD vs hAβ-KI and that a longer term of treatment is necessary to accelerate diffusible Aβ pathology in the hAβ-KI mice. Conclusions: This hAβ-KI model represents an important step towards the development of next-generation animal models that will provide better predictive outcomes for human patients. Grants: UCI MIND Pilot project (DBV), Ministry of Science PID2019-108911RA-100 (DBV), U54 AG054349 (FML), Institute of Health Carlos III PI18/01557 (AG) co-financed by FEDER funds (European Union), NIH/NIA Grant P50 AG16573 (UCI-ADRC).

P023 / #1244


ER AGGREGATION SITES MODULATE OMEGASOME AND AUTOPHAGOSOME FORMATION IN AN OLIGOMERIC Β-AMYLOID (1-42) MODEL OF EARLY AUTOPHAGY

Lecture Title:

S.S.H. Yeung1, Y.-S. Ho2, R.C.-C. Chang1,3 1The University of Hong Kong, Laboratory Of Neurodegenerative Diseases, Lks Faculty Of Medicine, Pok Fu Lam, Hong Kong, Hong Kong PRC, 2Hong Kong Polytechnic University, School Of Nursing, Hong Kong, Hong Kong PRC, 3The University of Hong Kong, State Key Laboratory Of Brain And Cognitive Sciences, Pok Fu Lam, Hong Kong PRC

Aims: Cellular degradation can be modulated by the Autophagy Lysosome Pathway (ALP), where dysfunctional proteins and organelles are degraded. Many neurological disorders are prominently linked with dysfunction in the ALP – leading to the aggregation of misfolded proteins (e.g. β-Amyloid). However, the underlying cellular pathomechanism of how β-Amyloid modulates autophagy remains elusive. Methods: To investigate, a murine model of primary hippocampal neurons were transfected with different constructs: GFP-KDEL, DsRed-KDEL, DsRed-Atg14L, DsRed-LC3, GFP-p62 and GFP-p62 L343A to allow confocal live-imaging of protein recruitment in ALP following timed Aβ-oligomer(1-42) treatment. Transfected cells were fixed and analyzed through immunohistochemistry, Western blot and the Proximity Ligation Assay to study the colocalization between proteins important in both omegasome and autophagosome recruitment in the ER Results: Here we found that Aβ1-42 oligomer (AβO) treatment increased AMPK-dependent recruitment of ULK-1 to the ER in early autophagy, independent of mTOR activation. Moreover, AβO treatment upregulated recruitment of components of the PI3K Complex III (i.e. Beclin-1, Atg14L and Vps34) to the ER, and subsequently induced increases in LC3 puncta distribution, autophagosome formation and accumulation within the endoplasmic reticulum. In parallel, p62, an autophagosome cargo protein was also found to translocate within ER aggregation sites following AβO treatment in both wildtype p62, and p62-L343A LIR mutant counterpart, suggesting roles of p62 not just in cargo shuttling, but omegasome maturation as well. Conclusions: Collectively, these data indicate that Aβ-induced autophagy begins at aggregation sites within the ER, and that Aβ primarily exerts cellular damage by upregulating autophagic machinery particularly in the ER during early autophagy.

P024 / #1230

Topic: Theme A: β-Amyloid Diseases / A1.b. Disease Mechanisms, Pathophysiology: Cell to cell transmission, spreading of pathology, prion-like

THE SPEED LIMITS OF TAU SPREADING - THE CONTRIBUTION OF EDUCATION AND REGIONAL AMYLOID

Lecture Title:

M. Hönig1,2, G. Bischof2, M. Sitter2, V. Dzialas2, T. Van Eimeren2,3,4, A. Drzezga1,2,4, F.T.A.D.N.I. (Adni)1 1Research Center Jülich, Neuroscience And Medicine Ii, Jülich, Germany, 2University of Cologne, University Hospital of Cologne, Nuclear Medicine, Multimodal Neuroimaging, Köln, Germany, 3University of Cologne, Neurology, Köln, Germany, 4German Center for Neurodegenerative Diseases, Pet Imaging, Bonn, Germany

Aims: Here, we investigated the contribution of education and regional amyloid as potential de- and accelerators in the spread of tau pathology. Methods: 85 amyloid-positive subjects (age=76.80±1.08, education=16.70±.50 years, M/F=42/43) were included, for whom [18F]AV-1451 PET at two timepoints, baseline [18F]AV-45 PET, ApoE-status and clinical information were available at ADNI. [18F]AV-1451 PETs were intensity-standardized (reference: inferior cerebellum) and z-transformed (control sample: 35 amyloid-negative subjects). Baseline and follow-up tau-PET z-maps were converted to volume-maps (z-score threshold > 1.96). Based on these volume-maps, tau-changes over time were assessed in terms of 1) tau speed (i.e. newly affected volume at follow-up), and 2) tau level rise (i.e. tau increase in previously affected volume). These two dependent variables were used in multiple regression analyses including education, global amyloid, baseline tau, ApoE, cognitive function, sex, and age as predictors. Additionally, baseline amyloid-burden derived from 32 Freesurfer-ROIs was regionally correlated with the newly tau-affected volume and mean z-score change in five Meta-ROIs (entorhinal cortex, inferior-temporal, precuneus, middle-occipital, oribito-frontal), correcting for multiple comparisons. Results: Education contributed as decelerator (β=-.322, p=.037), whereas global amyloid (β=.262, p=.029) and baseline tau (β=.283, p=.028) facilitated tau speed. Tau level rise was negatively associated with age (β=-.211, p=.017) and positively related with baseline tau (β=.710, p<.001). Tau spread to the precuneus was associated with greater cingulate amyloid-burden, whereas tau level rise in temporal, parietal and occipital regions was positively correlated with orbito-frontal and cingulate amyloid-burden. Conclusions: A combination of de- and accelerating factors and regional dissociations likely contribute to the temporo-spatial evolution of tau pathology.

P025 / #465


THE ROLE OF THE RETROMER, AUTOPHAGY AND EXOSOMES IN THE CLEARANCE AND PROPAGATION OF AMYLOID BETA OLIGOMERS IN ALZHEIMER’S DISEASE

Lecture Title:

L. Johansson1, N. Albargothy1, C. Sackmann1, M. Hallbeck2 1Linköping University, Biomedical And Clinical Sciences, Division Of Neurobiology, Linköping, Sweden, 2Linkoping University, Department Of Clinical And Experimental Medicine, Division Of Neurobiology And Clinical Pathology, Linkoping, Sweden

Aims: The vacuolar sorting protein 35 (VPS35) is a central component of the retromer complex responsible for intracellular cargo sorting of endosomal molecules. Reduced expression and mutations in VPS35 were detected in the hippocampus of late-onset Alzheimer’s disease patients. Our recent work identified novel mechanisms for this disease’s prion-like propagation via small extracellular vesicles known as exosomes. In this study, we unravel the role of the retromer system, autophagy and exosomes in the clearance and propagation of toxic oligomeric Amyloid beta (oAβ). Methods: We established a stable cell line of differentiated SH-SY5Y cells expressing truncated non-functional VPS35, by utilizing CRISPR/Cas9. We then investigated the effect of retromer dysfunction on the clearance and transfer of oAβ in a 3D co-culture system that allows the quantification of cell-to-cell protein transfer between donor and recipient cells. Results: We show that retromer dysfunction increases oAβ accumulation and decreases clearance regardless whether oAβ originates from extracellular milieu or direct neuronal transfer. Retromer dysfunction also decreases release of exosomes and upregulates autophagy and these effects are amplified following oAβ treatment. Exosomes derived from oAβ treated cells with retromer dysfunction significantly increase cell death in organotypic hippocampal slices. Conclusions: Our results provide evidence that retromer dysfunction and autophagy impairment decrease the ability of neurons to clear oAβ resulting in their accumulation and propagation. Together with current evidence, our data points to the retromer system, autophagy and exosome biogensis as potential therapeutic targets to enhance the proper recycling and clearance of toxic oAβ and reduce their spread to slow down disease progression.

P026 / #1090


SOLUBLE HIGH-MOLECULAR-WEIGHT AΒ OLIGOMERS DERIVED FROM THE AΒ-LADEN BRAINS INDUCE CEREBRAL Β-AMYLOIDOSIS.

Lecture Title:

M. Kashiwagi-Hakozaki1,2, Y. Naka1, P. Dooley3, M. Frosch3, B. Hyman3, M. Fukayama2, T. Ushiku2, T. Hashimoto1,4, T. Iwatsubo1 1Graduate School of Medicine, The University of Tokyo, Department Of Neuropathology, Tokyo, Japan, 2Graduate School of Medicine, The University of Tokyo, Department Of Pathology, Tokyo, Japan, 3Massachusetts General Hospital, Massachusetts Alzheimer's Disease Research Center, Charlestown, United States of America, 4Graduate School of Medicine, The University of Tokyo, Department Of Innovative Dementia Prevention, Tokyo, Japan

Aims: Extracellular deposition of amyloid-β (Aβ) peptides is a key neuropathological feature of Alzheimer’s disease (AD). We aimed to identify the “seed Aβ” strains that induce the initiation and propagation of Aβ pathology in AD brains. Methods: The Tris buffer-soluble fractions from the brains of APP tg mice and autopsied human AD brains were separated by Seperdex-75 size-exclusion chromatography. The Aβ-positive fractions verified by ELISA were stereotaxically injected into the unilateral hippocampus of the 9-month-old APP tg mice, and Aβ-positive area in the hippocampus was assessed after 4 months to evaluate the seeding capacity. Results: Tris-soluble Aβ from the brains of APP tg mice was eluted into three fractions, at ~200-300, ~50-60 and ~10-20 kDa, among which the presence of Aβ oligomers was verified by an Aβ oligomer-specific ELISA in the ~200-300 kDa fraction. Intrahippocampal injection of the ~200-300 kDa fraction induced a unique laminar pattern of Aβ deposition extending into the layers and fiber tracts in the hippocampi. Immunodepletion of Aβ with anti-Aβ antibodies from the ~200-300 kDa fraction abolished the potency to induce Aβ deposition. The Tris-soluble ~200-300 kDa Aβ-positive fraction from AD brains also induced a similar pattern of Aβ deposition in the brains of APP tg mice. Conclusions: Soluble Aβ oligomers with a size of ~200-300 kDa derived from the brains of APP tg mice and AD patients exhibited a potency to induce cerebral β-amyloidosis, which might be involved in the spatiotemporal spreading of β-amyloidosis in AD brains.

P027 / #847


EVALUATION OF ALZHEIMER’S DISEASE HETEROGENEITY THROUGH HUMAN BRAIN INOCULATIONS IN A MOUSE MODEL OF AMYLOID AND TAU LESIONS

Lecture Title:

S. Lam1, L. Stimmer2, S. Boluda2,3, A.-S. Hérard1, F. Petit1, C. Duyckaerts2,3, B. Delatour2, S. Haïk2,3, M. Dhenain1 1Institut F. Jacob, UMR9199 CEA/CNRS, Molecular Imaging Research Center (mircen), Fontenay-aux-Roses, France, 2Institut du cerveau, INSERM U1127, CNRS UMR7225, Sorbonne Universités, Paris, France, 3Laboratoire de neuropathologie Raymond Escourolle, Sorbonne Université, Hôpital de la Pitié-Salpêtrière, Département De Neuropathologie - Pôle Des Maladies Du Système Nerveux, Paris, France

Aims: A major challenge in clinical research is to better understand the heterogeneity of Alzheimer’s disease (AD). While classical sporadic forms of AD (clAD) evolve over approximately 10 years, the "rapidly progressive form of AD" (rpAD) is a non-genetic aggressive form characterized by a rapid clinical decline over 2 years. Several studies have shown that inoculations of brain samples with Aβ or tau pathology into transgenic (tg) mouse models overexpressing mutant human APP or tau promotes either Aβ or tau aggregation. This work aims to study the impact of rpAD versus clAD human brain inoculations in a tg mouse model of β-amyloidosis and explore the mechanisms associated to AD heterogeneity. Methods: Human brain samples were characterized pathologically and biochemically. rpAD, clAD and ctrl (non-AD) samples were inoculated in the CA1 of 2-month-old APP/PS1dE9 mice (n=15-20/group). After 8 months, memory performance was evaluated. Aβ deposits, tau inclusions and synaptic density were histologically assessed. Results: Morphometric differences were observed in amyloid plaques between clAD and rpAD patients (Fig. 1), while Aβ/tau loads were similar. In mice, this same lesionnal phenotype was transmitted (Fig. 1) along with heterogeneous clinical outcomes and synaptic deficits. Indeed, only rpAD-inoculated animals displayed cognitive alterations (Fig. 2) and synaptic loss (Fig. 3).

Conclusions: The inoculation of different human brain samples to mice is a new tool to explore AD heterogeneity as AD-like lesionnal and clinical heterogeneity can be experimentally transmitted. Our results suggest that, independently from the severity of amyloid and tau pathologies, synaptic health variability may contribute to AD heterogeneity.

P028 / #976


ANTI-TAU ANTIBODY SEMORINEMAB REDUCES TAU LEVELS IN BRAIN INTERSTITIAL FLUID IN VIVO

Lecture Title:

S.-H. Lee1, C. Wallace2, K. Stark1, W. Meilandt1, S. Schauer3, P. Chan4, S. Sadekar5, R. Fuji6, A. Easton1, J. Cirrito2, K. Wildsmith3 1Genentech, Neuroscience, South San Francisco, United States of America, 2Washington University in St. Louis, Neurology, St. Louis, United States of America, 3Genentech, Devsci Omni-biomarker, South San Francisco, United States of America, 4Genentech, Biochemical And Cellular Pharmacology, South San Francisco, United States of America, 5Genentech, Devsci Ptpk, South San Francisco, United States of America, 6Genentech, Devsci Sa Pathology, South San Francisco, United States of America

Aims: The burden and distribution of tau pathology correlates with cognitive decline in Alzheimer’s disease (AD), making tau an attractive therapeutic target. Semorinemab is a humanized anti-tau monoclonal IgG4 in development for the treatment of AD. The proposed mechanism of action of semorinemab is to bind and intercept tau in brain interstitial fluid (ISF), thus blocking cell-to-cell spread of tau pathology. In previous in vivopreclinical studies, a peripherally administered mouse surrogate of semorinemab showed efficacy in slowing the spread of tau pathology in a transgenic (Tg) mouse model. However, the pharmacokinetic (PK)/pharmacodynamic (PD) relationship and mechanism of action of semorinemab in ISF requires confirmation. Methods: We examined the effect of a mouse surrogate of semorinemab (semorinemab CDR on a murine IgG1 Fc; muSEMO) on tau levels in brain ISF in wild-type human tau knock-in mice. Control antibody or muSEMO was peripherally administered and ISF was collected via microdialysis every 90 min for 72 hrs to measure tau levels and antibody concentrations. Additional PK/PD assessments were performed on plasma and brain. Results: Significant reductions in ISF tau levels were seen in the muSEMO group relative to the control group. Conclusions: These results support the mechanistic hypothesis that semorinemab engages and facilitates removal of tau in ISF, thereby attenuating the cell-to-cell spread of pathological tau.

P029 / #759


SPREADING OF AMYLOID-Β PATHOLOGY AND MATURATION OF AMYLOID-Β AGGREGATES: A TWO-ACT PLAY IN MAN AND MOUSE

Lecture Title:

X. Li1, T. Robberechts1, S. Ospitalieri1, K. Balakrishnan2,3, L. Hofmann3, C. Schmid3, A. Upadhaya3, A. Ronisz1, M. Koper1, C. Von Arnim4,5, S. Kumar6, M. Willem7, J. Walter6, D. Thal1,3 1KU Leuven, Department Of Imaging And Pathology, Leuven, Belgium, 2Ulm University, Department Of Gene Therapy, Ulm, Germany, 3Ulm University, Institute Of Pathology, Ulm, Germany, 4Ulm University, Department Of Neurology, Ulm, Germany, 5University of Goettingen Medical School, Department Of Geriatrics, Goettingen, Germany, 6University of Bonn, Department Of Neurology, Bonn, Germany, 7Ludwig-Maximilians-University Munich, Biomedical Center, Munich, Germany

Aims: 1. To clarify whether propagation of Aβ pathology follows a similar sequence of Aβ aggregate maturation events as observed in the neocortex. 2. To analyze whether the initiation of mature plaques occurs once a maturation stage has been reached in a given brain or whether it depends on the composition of the seed. Methods: Biochemical and histopathological analysis of Aβ plaque composition with antibodies against non-modified Aβ, AβN3pE and AβpSer8 in the neocortex, cingulate gyrus, basal ganglia and cerebellum of 37 human autopsy cases. Injection of human brain-derived Aβ seeds from brains in early and late maturation stages of Aβ pathology into the hippocampus of 2-month-old APP23 mice and subsequent histopathological investigation at 6 months. Results: All investigated regions in the human brain showed more Aβ plaques detectable with antibodies against non-modified Aβ than with antibodies against AβN3pE, and the number of AβpSer8-positive plaques was the least. On the other hand, injection of seeds into APP23 mouse brain always induced Aβ deposits positive for non-modified Aβ, AβN3pE and AβpSer8 at 6 months of age regardless of the composition of the seeds injected, although non-modified Aβ-positive plaques were more widespread than AβN3pE and/or AβpSer8-positive plaques. Conclusions: Propagation of Aβ plaque pathology in the human brain is followed by Aβ aggregate maturation in each newly affected region, similar as in the neocortex. On the other hand, in APP23 mice, propagation and maturation of Aβ aggregates are induced by Aβ seeds, regardless of their composition.

P030 / #1519

Topic: Theme A: β-Amyloid Diseases / A1.c. Disease Mechanisms, Pathophysiology: Inflammation

QUERCETIN REDUCES MANGANESE NEUROTOXICITY ON THE HIPPOCAMPUS OF ADULT MICE

Lecture Title:

A. Adekeye, O. Ofogba, A. Fafure Afe Babalola University, Anatomy; Neuroscience Unit, Ado-Ekiti, Nigeria

Aims: To check the effects of Quercetin on manganese-neurotoxicity in the hippocampus Methods: In this study, 40 adult mice of average weight of 18 – 29g were randomly distributed into five groups of eight each. The first group served as the control group, the second group received manganese alone, the third group received Quercetin alone, the fourth group served as the group that received manganese first then received Quercetin, the Fifth group received both manganese and Quercetin at the same time. Before and after final administration, neurobehavioral test such Novel recognition (NORt) and Ymaze tests were used to evaluate the level for the working spatial memory. The animals were sacrificed using ketamine followed by intracardiac perfusion fixation (0.9% followed by 4% paraformaldehyde) while the animals set out for biochemical analysis were sacrificed through cervical dislocation. The brain was harvested and the region of the hippocampus was grossed for histological, immunohistochemical and biochemical analysis. Statistical analysis were done using one-way ANOVA with Newman-keuls for post hoc tests. The data statistical analysis were represented in graphs and bar charts with error bar representing the mean ± SEM Results: The results from immunohistochemical study revealed that the group that received manganese showed an increase in the amount of TNFα when compared to the control and Quercetin. an alterations at the granular layer was observed in the hippocampus to affect neuronal integrity Conclusions: Manganese may have possibilities of causing neuronal degeneration of the dentate gyrus and administration of Quercetin may proffer an ameliorative solution.

P031 / #1359


ALTERED EXPRESSION OF GLIAL GAP JUNCTION PROTEINS CX43, CX30 AND CX47 IN THE 5XFAD MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

S. Angeli1, I. Kousiappa1, K. Kleopa2, S. Papacostas3 1The Cyprus Institute of Neurology and Genetics, School of Molecular Medicine, Neurobiology Department, Nicosia, Cyprus, 2The Cyprus Institute of Neurology and Genetics, School of Molecular Medicine, Neuroscience Department, Center For Neuromuscular Disorders, Center For Multiple Sclerosis And Related Disorders, Nicosia, Cyprus, 3Medical School University of Nicosia,The Cyprus Institute of Neurology and Genetics, School of Molecular Medicine, Neurobiology Department, Cognitive Disorders Center, Nicosia, Cyprus

Aims: Connexins form gap junctions in the CNS to allow bidirectional cytosolic exchange of molecules between adjacent cells. Their involvement in inheritable diseases and the use of animal models that closely mimic such diseases revealed the critical role of glial gap junctions in myelination and homeostasis. Connexins are also implicated in acquired demyelinating disorders, such as Multiple Sclerosis and Alzheimer’s disease. The aim of this study was to investigate the astrocytic connexins, Cx43 and Cx30 in relation to oligodendrocytic Cx47 in the cortex and thalamus of the 5XFAD mouse model of AD. Methods: 5XFAD and wild-type (WT) mice were used at the ages of 3, 6 and 9-months of age. Immunostainings were performed for Aβ, Cx43, Cx30, Cx47, Cx32, Olig2, CC1 and PLP. Also, real-time PCR and immunoblot experiments were performed to measure Cx43 and Cx30 mRNA and protein levels. Results: The model was characterized by increased Aβ deposition, gliosis, neuronal loss, and memory impairment. Cx43 and Cx30 showed increased immunoreactivity in older 5XFAD compared to WT mice, reflecting astrogliosis, while Cx47 immunoreactivity was reduced. Moreover, Cx47 GJ plaques showed reduced colocalization with Cx43 plaques. Oligodendrocyte precursor cells and mature oligodendrocyte populations were also depleted, and myelin deficits were observed. Conclusions: Our findings indicate reduced astrocyte-oligodendrocyte gap junction connectivity and possibly a shift in Cx43 expression towards astrocyte-astrocyte gap junctions and/or hemichannels, that could impair oligodendrocyte homeostasis and myelination. Our study provides evidence that connexins might have implications in the progression of AD, although the role of oligodendrocyte connexins in AD requires further investigation.

P032 / #629


APOE4 PRIMES INFLAMMATION IN HUMAN ASTROCYTES THROUGH THE DOWNREGULATION OF A NEWLY DISCOVERED TARGET CANDIDATE

Lecture Title:

L. Arnaud, L. Greetham, D. Stephan, A. Jimenez, N. Jullien, P. Belio-Mairal, P. Benech, K. Baranger, S. Rivera, E. Nivet Institute of NeuroPhysiopathology (INP) (UMR 7051), Faculty Of Medical And Paramedical Sciences, Marseille Cedex , France, France

Aims: The polymorphism on APOE is of outmost interest for studying the mechanisms involved in Alzheimer’s disease (AD), as it is the major genetic risk, and thought to alter key functions that lead to increased susceptibility to the disease. Neuroinflammatory changes are amongst the earliest brain manifestations and are increasingly seen as critical in the pathogenesis. Yet, the role of APOE4 in neuroinflammatory processes remains poorly understood despite being primarily produced by astrocytes, which are major neuroinflammatory regulators. Therefore, studying how APOE4 alters inflammatory functions under the prism of astrocytes represents a new path to better understand the so far elusive contribution of these glial cells to AD pathogenesis. Methods: We leveraged on the use of patient-specific induced pluripotent stem cell (hiPSC)-derived astrocytes, combined with CRISPR-Cas9-based genome editing. Along with hiPSC lines from patients/donors carrying different APOE polymorphism, we generated isogenic APOE4 Knock-In (KI) and Knock-Out (KO) lines from APOE3 hiPSCs to evaluate the specific impact of different APOE variants on the inflammatory status/responses in human astrocytes. Results: We found that astrocytes displayed a pro-inflammatory signature characterized by sustained and exacerbated inflammatory responses specific to APOE4. We identified a new mechanism by which APOE4 alters the NFkB signaling pathway to establish a disease phenotype in human astrocytes. Conclusions: Our results highlight APOE as a key regulator of neuroinflammation in astrocytes through the modulation of a newly discovered mechanism, thus unveiling a new potential therapeutic target (undisclosed yet) for further development aiming at controlling the detrimental inflammatory component in AD.

P033 / #839


MIF ROLE IN NEURODEGENERATION

Lecture Title:

S. Besson-Girard1, L. Liu1, B. Bulut1, H. Ji1, F. Usifo1, J. Bernhagen2,3, O. Gokce1,3 1Institute for Stroke and Dementia Research, Klinikum der Universität München, System Neuroscience Laboratory, Munich, Germany, 2Institute for Stroke and Dementia Research, Klinikum der Universität München, Ludwig-Maximilians-University, Department Of Vascular Biology, Munich, Germany, 3Munich Cluster for Systems Neurology (SyNergy), -, Munich, Germany

Aims: Alzheimer’s disease (AD) pathology is shaped by the interplay between the neuronal and immune compartments. The misfolded and aggregated proteins in the neuronal compartment initiate release of inflammatory mediators such as macrophage migration-inhibitory factor (MIF). MIF has a multitasking potential that allows for an elaborate fine-tuning of AD pathology in the extra- and intracellular space. In extracellular space, MIF modulates acute and chronic inflammatory diseases such as septic shock, acute respiratory distress syndrome, neuroinflammation, or rheumatoid arthritis. Recent studies also highlight MIF’s surprising role as the long-searched missing nuclease for the AIF-dependent cell death pathway, which drives neuronal loss in various nervous system disorders. Methods: To elucidate MIF functions in AD, we characterized the impact of MIF on an AD mouse model using behavior, single-cell RNA sequencing (scRNAseq), and imaging. Results: Our results suggest MIF plays an essential role in AD phenotypes, including neuronal survival, scRNAseq signatures, and behavior. MIF modulates AD pathologies such as neuronal loss, neuroinflammation and behavioral problems. Conclusions: Overall our results suggest that MIF plays an essential role in AD phenotypes, including neuronal survival, scRNAseq signatures, and behavior. Furthermore, targeting MIF functions is a promising treatment candidate for AD.

P034 / #975


P53 ORCHESTRATES THE MICROGLIAL RESPONSE UPON AMYLOID-BETA-INDUCED NEURODEGENERATION IN VIVO

Lecture Title:

S. Gonzalez-Guerrero1,2, R. Lapresa1,2, J. Agulla1,2, A. Almeida1,2 1Institute for Biomedical Research of Salamanca (IBSAL), Molecular Neurobiology, Salamanca, Spain, 2Institute of Functional Biology and Genetics (IBFG), Molecular Neurobiology, Salamanca, Spain

Aims: Amyloid-β (Aβ) oligomers trigger signalling cascades involving alteration of calcium homeostasis, ROS production, inflammatory processes and mitochondrial dysfunction, culminating in neuronal apoptosis. According to a functional scale, microglia can be categorized into the classical pro-inflammatory/neurotoxic phenotype, known as M1, and an alternative anti-inflammatory/neuroprotective M2 phenotype, which is involved in the resolution of inflammation, phagocytosis and tissue repair. Previously, we described a key role of p53 in neurodegeneration induced by Aβ-neurotoxicity in vivo. Recently, p53 has emerged as a key modulator of the immune response in microglia. Here we evaluated whether p53 is modulating the microglial response to Aβ25-35 oligomers leading to neurodegeneration. Methods: Aβ25-35 oligomers (9 nmol) were stereotaxically injected in the cerebral right ventricle of WT and p53KO mice. Microglial phenotype and neurodegeneration were assessed by using ELISA, Western blot and immunofluorescence. Memory status was evaluated by using the AnyMaze™ system. Results: We found that oligomerized Aβ25-35 triggered p53 accumulation leading to an early microglial activation. Besides, p53 dictates the microglial M1/M2 phenotype. Moreover, Aβ injection also induced reactive astrogliosis. Altogether, these events lead to dendrite disruption, neuronal death and memory impairment. Furthermore, these effects were prevented by the genetic (p53KO) and pharmacological (PFT-alpha) inhibition of p53. Conclusions: Our results highlight a key role of p53 in the Aβ-induced inflammatory response, which may contribute to neurodegeneration and cognitive decline. Funded by The Instituto de Salud Carlos III (PI18/00265); European Union’s Horizon 2020 Research and Innovation Programme (Grant Agreement 686009) and Programa Operativo FSE Castilla y León 2014-2020.

P035 / #610


CELLULAR SENESCENCE, INFLAMMATION, AND COGNITION IN AGING AND ALZHEIMER’S DISEASE: WHAT’S THE CONNECTION?

Lecture Title:

K. Hascup1, N. Esperant-Hilaire2, C. Findley3, S. Mcfadden2, Y. Fang2, A. Bartke4, E. Hascup3 1Southern Illinois Univ. School of Medicine, Neurology, Cadrd, Pharmacology, And Mmicb, Springfield, United States of America, 2Southern Illinois Univ. School of Medicine, Neurology, Cadrd, Springfield, United States of America, 3Southern Illinois Univ. School of Medicine, Neurology, Cadrd, And Pharmacology, Springfield, United States of America, 4Southern Illinois Univ. School of Medicine, Internal Medicine And Cadrd, Springfield, United States of America

Aims: Cellular senescence and inflammation may contribute to the pathogenesis of Alzheimer’s disease (AD). We sought to define their inter-relationship in cell culture and animal models of AD and determine if senolytic compounds could alleviate these conditions thereby improving cognition. Methods: Hippocampal tissue from 12 month old male AβPP/PS1 and C57BL/6 mice underwent RNA sequencing (RNAseq) to assess cellular senescence and inflammation. Primary neurons were treated with Aβ42 (10µM) and fisetin (15µM), dasatinib (1µM), quercetin (10µM), dasatinib+quercertin, or vehicle control. Cell viability, senescent cell burden, and cytokine profiles were assessed. 16 month old male and female APPNL-F/NL-F mice and C57BL/6 mice underwent hippocampal infusion of dasatinib+quercetin. Learning and memory, inflammation, and cellular senescence were assessed at 18 months of age. Results: RNAseq results support elevated senescent and inflammatory markers in the hippocampus of AβPP/PS1 mice compared to C57BL/6 mice. Preliminary data revealed that primary neurons treated with Aβ42 and senolytic compounds tended to have decreased senescence and inflammatory markers compared to controls. In vivo studies are ongoing to determine if preventative and/or intervention treatment with senolytic compounds are effective at altering AD progression. Conclusions: Preliminary results support elevated cellular senescence and inflammation in the hippocampus of an AD mice. Senolytic treatments in primary neuronal cultures were able to reduce Aβ42–related cellular senescence. Elimination of senescent cells and the corresponding inflammatory profile may be effective at treating AD. Ongoing studies to evaluate the in vivo effectiveness of senolytic compounds in mouse models of AD will be presented. Supported by NIH R01AG057767 and R01AG061937.

P036 / #806


PATIENT IPSC-DERIVED NEURONS AND MICROGLIA FOR DISEASE MODELING OF ALZHEIMER’S DISEASE

Lecture Title:

H. Haukedal, K. Freude University of Copenhagen, Department Of Veterinary And Animal Sciences, Frederiksberg, Denmark

Aims: Alzheimer’s disease (AD) is the most common cause of dementia with no curative treatments available. Investigating precise disease pathology is therefore crucial, potentially facilitating a new therapeutic approach. Methods: To understand underlying molecular mechanisms induced pluripotent stem cells (iPSC) were derived from patients with familial AD (fAD), caused by Presenilin 1 mutations, and sporadic AD (sAD), with no apparent heritability. Isogenic controls for fAD lines were generated by correcting the mutations using CRISPR/Cas9. Additionally, CRISPR/Cas9 was used to introduce mutations into healthy iPSC to establish new cell lines. iPSC were differentiated into forebrain-specific cortical neurons. Moreover, to understand the non-neuronal aspect of AD, iPSC were differentiated into microglia, suspected to be proinflammatory drivers of AD. Results: fAD neurons confirmed increased amyloid beta production and Tau phosphorylation, accompanied by aberrant mitochondria, metabolic defects, Golgi fragmentation and synaptic deficits. The cellular phenotypes were rescued in isogenic controls, indicating that they are related to the mutation, and can be considered early AD phenotypes. Genetic risk factors for developing sAD have been identified, and iPSC from sAD patients were categorized based on single nucleotide polymorphisms (SNP) to determine if microglia with genetic profiles associated with high risk of developing AD expressed elevated proinflammatory cytokines levels, being more detrimental to surrounding neurons compared to low risk microglia. The inflammatory profile was also assessed in fAD-, control- and knock-in lines creating a full overview of neuroinflammation in various AD conditions. Conclusions: Recapitulation of various disease phenotypes in our in-vitro patient-specific disease models make it attractive for drug target development.

P037 / #1188


CROSSTALK BETWEEN NEURONS AND GLIA IN ALZHEIMER’S DISEASE: A TRIPLE CO-CULTURE MODEL

Lecture Title:

C. Luchena1,2,3, J. Zuazo-Ibarra1,2,3, E. Alberdi1,2,3, C. Matute1,2,3, E. Capetillo-Zarate1,2,3,4 1Achucarro Basque Center for Neuroscience, Laboratory Of Neurobiology, Leioa, Spain, 2CIBERNED, Centro De Investigación Biomédica En Red Enfermedades Neurodegenerativas, Madrid, Spain, 3Faculty of Medicine and Nursery, University of the Basque Country (UPV/EHU), Department Of Neuroscience, Leioa, Spain, 4IKERBASQUE, Basque Foundation For Science, Bilbao, Spain

Aims: Given that neuroinflammation is a key contributor to Alzheimer’s disease (AD) progression, we wanted to establish an in vitro model where we could recapitulate the disease and study the communication between neurons and glial cells. Methods: We established a triple co-culture model where neurons, microglia and astrocytes were present, and immunofluorescence, western blot and ELISA techniques were used to characterize it. Additionally, oligomeric Aβ (oAβ) was added to model AD. Results: We found that microglia increased the expression of anti-inflammatory marker Arginase I and decreased the secretion of pro-inflammatory cytokine IL-1β in the triple co-culture, compared to microglia alone. Astrocytes displayed a more ramified morphology and decreased the expression of pro-inflammatory A1 marker C3. The presence of neurons also induced astrocytes to increase the expression of glutamate receptor GLT-1 by astrocytes. Neurons showed an increase of post-synaptic markers PSD-95 and Homer, and developed more and longer branches when co-cultured with glial cells. Treatment of the triple co-culture with oAβ caused a reduction of pre-synaptic marker Synaptophysin and post-synaptic marker Homer, and increased the expression of CD11b in microglia. Conclusions: Our results suggest that in the triple co-culture model, microglia are less inflammatory, astrocytes are less reactive and neurons display a more mature morphology than in individual primary cultures. Moreover, we were able to recapitulate Aβ-induced synaptic loss and activation of microglia. Thus, this triple co-culture model could be a more appropriate in vitro model for AD and neuroinflammation than individual primary cultures.

P038 / #1288


THE LINK BETWEEN BLOOD-BASED BIOMARKERS AND MILD COGNITIVE IMPAIRMENT AND ALZHEIMER'S DISEASE IN A PANAMANIAN SAMPLE

Lecture Title:

D. Oviedo1,2, A. Villarreal2, C. Posada2, S. Rodriguez2, G. Rangel2, M. Carreira2, A. Perez-Lao2, G. Britton2 1Universidad Santa María La Antigua, Psychology, Panama, Panama, 2INDICASAT AIP, Center For Neuroscience, Panama, Panama

Aims: Research has established that studying brain changes in a prodromal phase in Alzheimer’s disease (AD), is key to improving intervention mechanisms and to delay the expression of symptoms. The aim of this study was to evaluate the link between blood-based biomarkers and mild cognitive impairment (MCI) and AD in a sample of elderly Panamanians. Methods: A descriptive, cross-sectional, observational study was conducted. Participants were 84 Panamanians (controls, n=41; MCI, n=32; and AD, n=11). They were part of an outpatient geriatric sample from a prospective study carried out by the Panama Aging Research Initiative (PARI). Non-fasting blood samples were collected. Participants were assessed with MMSE and Clock Test, and were genotyped for ApoΕ4. Results: CRP levels among AD participants were significantly increased relative to MCI participants (p < 0.05). A2M levels in AD participants were significantly increased compared to normal controls (p < 0.05). SAA levels in AD participants were significantly increased when compared to MCI participants (p < 0.05). Conclusions: These results show that in the AD group CRP, A2M and SAA are significantly increased. The link between MCI and AD and blood-based markers can produce an accurate tool that can aid in early diagnosis of AD.

P039 / #492


SYNTHETIC AMYLOID BETA DOES NOT INDUCE A ROBUST TRANSCRIPTIONAL RESPONSE IN IMMUNE CELL CULTURE SYSTEMS

Lecture Title:

I. Quiroga1, E. Cruikshank2, K. Reed3, B. Evangelista4, J.-H. Tseng4, R. Meeker4, M. Bond1, T. Cohen4, D. Phanstiel5 1University of North Carolina, Thurston Arthritis Research Center, Chapel Hill, United States of America, 2University of North Carolina, Postbaccalaureate Research Education Program, Chapel Hill, United States of America, 3University of North Carolina, Curriculum In Genetics & Molecular Biology, Chapel Hill, United States of America, 4University of North Carolina, Department Of Neurology, Chapel Hill, United States of America, 5University of North Carolina, Thurston Arthritis Research Center, 5department Of Cell Biology And Physiology, Chapel Hill, United States of America

Aims: Alzheimer’s disease (AD) is a progressive neurodegenerative disease that impacts nearly 50 million people worldwide, but no effective treatments are available, in large part because the mechanisms underlying this disease are unclear. Recent evidence suggests that the main factor contributing to AD progression is the development of a chronic neuroinflammatory state in response to the accumulation of Amyloid beta (Ab). As a result, many in vitro systems have been established to study Ab-mediated activation of innate immune cells and the role of AD genetic risk variants in this process. Nevertheless, the resemblance of these models to the AD brain has never been comprehensively studied on a genome-wide scale. Our aim is to quantify the global trascriptional response of cultured innate immunce cells treated with Ab. Methods: We treated several established, primary and iPSC-derived immune cell lines (including macrophages, microglia and astrocytes) with different formulations and concentrations of synthetic Ab, and performed RNA-seq. Results: Surprisingly, our results reveal that in most cases synthetic Ab does not produce a robust transcriptional response in these cells. Conclusions: Our results suggest that alternative in vitro models to mimic the inflammatory estate in the AD brain need to be developed to further understand the molecular mechanisms underlying this disease.

P040 / #618


INVESTIGATING THE ROLE OF THE ADAPTIVE IMMUNE SYSTEM IN ALZHEIMER’S DISEASE PATHOGENESIS

Lecture Title:

J. Sanchez1, H. Davtyan2, M. Blurton-Jones2 1UC Irvine, Neurobiology & Behavior, Irvine, United States of America, 2Institute for Memory Impairments & Neurological Disorders Sue & Bill Gross Stem Cell Research Center, Department Of Neurobiology & Behavior, Irvine, United States of America

Aims: The innate immune system, particularly microglia, have been strongly implicated in Alzheimer’s disease (AD) pathogenesis, however the role of adaptive immunity remains poorly understood still. Our lab has generated two transgenic models of AD. Methods: To determine whether genetic deletion of adaptive immunity alters AD pathogenesis, we backcrossed the well-established 5xfAD model onto a Rag2/il2rg double knockout background (RAG2ɣc-/-5xfAD). Interestingly, genetic deletion of adaptive immune cells in these mice led to a dramatic increase in amyloid beta plaque load, an activated microglial neuroinflammatory state, and a corresponding decrease in the ability of microglia to phagocytose beta-amyloid. Results: To further investigate the specific role of T cells in these findings, I have performed adoptive transfer experiments including bone marrow transplantation and isolated T cell transfers into RAG2ɣc-/-5xfAD and RAG2ɣc-/- mice. Immunohistochemistry and flow cytometry have revealed a significant increase in memory CD8+ T cells within the RAG2ɣc-/-5xfAD brain parenchyma. Interestingly, T cells were sometimes observed adjacent to plaque-associated microglia in the RAG2ɣc-/-5xfAD brain. I have also sought to determine whether a similar effect can be observed within immune intact AD mice. I confirmed that CD8 T cell infiltration is dramatically increased in immune intact AD mice. Conclusions: This data shows that CD8+ T cells can induce both protective and detrimental responses within the AD brain. Therefore, I hypothesize that CD8 cells directly interact with microglia to modulate their activity and decrease beta-amyloid, but conversely also contribute to neuronal death in AD.

P041 / #1655


INTESTINAL DYSBIOSIS PROMOTES IMMUNE DYSREGULATION IN AN ANIMAL MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

B. Santos-Lima1, E. Zenaro1, A. Slanzi1, E. Pietronigro1, N. Lopez1, E. Terrabuio1, N. Vitulo2, V. Della Bianca1, G. Angelini1, A. Bani1, A. Suli1, G. Constantin1 1University of Verona, Department Of Medicine, Verona, Italy, 2University of Verona, Department Of Biotechnology, Verona, Italy

Aims: Current evidence suggests that systemic inflammation promotes Alzheimer’s Disease (AD) and recent reports demonstrated that circulating neutrophils migrate into the AD brain inducing neuroinflammation and cognitive deficits in animal models of AD. However, how peripheral immune dysregulation occurs and promotes neuroinflammation and memory decline in AD is still unknown. Even though descriptive reports suggest the involvement of the microbiome in AD, the mechanisms behind AD pathology are not explored. In this context, we aim to better understand the dysregulation of leukocyte subpopulations in AD by addressing the gut-blood-brain axis. Methods: By taking advantage of the 3xTg-AD mouse model we addressed peripheral inflammation during early disease stages. To this aim, we performed: 1) an intestinal microbiota analysis; 2) characterization of circulating inflammatory mediators; and 3) immunophenotypization of peripheral blood. Results: In our study we show a clear intestinal dysbiosis in the 3xTg-AD mice, characterized by higher abundance of taxa known to cause proinflammatory conditions. In agreement, we also found increased intestinal permeability and accumulation of immature precursors and aged neutrophils in circulating blood, suggesting a strong peripheral dysregulation in the early phases of the disease. Conclusions: Altogether, our data suggest microbiota as a modulator of leukocyte activation and migration into the brain. Therefore, targeting the gut-blood-brain axis in AD could lead to new immunomodulatory strategies for AD treatment.

P042 / #1178


CALCIUM-DEPENDENT CYTOSOLIC PHOSPHOLIPASE A2 ACTIVATION IS IMPLICATED IN NEUROINFLAMMATION AND OXIDATIVE STRESS ASSOCIATED WITH APOE4

Lecture Title:

S. Wang1, B. Li1, V. Solomon1, A. Fonteh2, H. Yassine1 1University of Southern California, Keck School Of Medicine, Los Angeles, United States of America, 2Huntington Medical Research Institutes, Neurosciences, Pasadena, United States of America

Aims: Apolipoprotein E4 (APOE4) is associated with a greater response to neuroinflammation and risk of developing late-onset Alzheimer disease (AD), but the mechanisms for this association are not clear. Activation of calcium-dependent cytosolic phospholipase A2 (cPLA2) is involved in inflammatory signaling and is elevated within plaques of the AD brain. The relation between APOE4genotype and cPLA2 activity is not known. Methods: Mouse primary astrocytes, mouse and human brain samples differing by APOE genotypes were collected for measuring cPLA2 expression, phosphorylation and activity in relation to measures of inflammation and oxidative stress. Results: Greater cPLA2 phosphorylation and activity were identified in ApoE4 compared to ApoE3 in primary astrocytes and brains of ApoE-targeted replacement (ApoE-TR) mice. These differences were also demonstrated in brain homogenates from the inferior frontal cortex from AD patients carrying APOE3/4 carriers compared to APOE3/3 carriers. Higher cPLA2 activation with APOE4 was associated with greater activation of the MAPK p38 pathway, as well as with higher levels of leukotriene B4 (LTB4), reactive oxygen species (ROS) and inducible nitric oxide synthase (iNOS). Inhibition of cPLA2 reduced LTB4, ROS and iNOS levels in ApoE4 primary astrocytes to those of ApoE3 astrocytes. Conclusions: Our findings implicate an induced cPLA2 leukotriene signaling system with APOE4, that could represent a potential target for mitigating the increased neuroinflammation with APOE4 and AD.

P043 / #791

Topic: Theme A: β-Amyloid Diseases / A1.d. Disease Mechanisms, Pathophysiology: Synaptic plasticity & synapse pathology

HIGH-CONTENT SCREENING OF ALZHEIMER'S DISEASE GENETIC RISK FACTORS BASED ON SYNAPTIC CONNECTIVITY ANALYSIS

Lecture Title:

A. Coulon, T. Mendes, D. Kilinc, J.-C. Lambert, J. Dumont, J. Chapuis Institut Pasteur de Lille, U1167, Lille, France

Aims: Synaptic loss is one of the earliest pathological hallmarks and the strongest marker of cognitive decline in Alzheimer’s disease (AD). A strong genetic predisposition is present in AD, and genome-wide association studies have pointed out more than hundreds of genes associated with the risk of developing the disease. With this background, the objective is to develop a cell-based screen to assess the impact of each genetic risk factor on synaptic connectivity. Methods: Immunofluorescences are performed on primary rat hippocampi neurons in 384-wells format, to stain pre- and post-synaptic compartments and neuronal network. Synaptic connectivity is then assessed through High Content analyses using Colombus and Matlab software. Results: We succeed in developing High Content analyses to assess the synaptic connectivity by assigning each post-synaptic structure to the nearest pre-synaptic structure. This approach could then be used to assess the impact of AD genetic risk factors on synaptic connectivity. For that, we plan to perform a screen of a lentiviral shRNA library, and to assess the impact of these genes under-expression on synaptic connectivity. Conclusions: Here, we have developed an high-content screening approach which will allow us to define the genetic risk factors that are involved in the pathophysiological synaptic dysregulation observed in the early stages of AD.

P044 / #1350


LOSS OF LOCUS COERULEUS-NORADRENERGIC AFFERENTS IN THE APP KNOCK-IN MOUSE MODEL OF AΒ AMYLOIDOSIS

Lecture Title:

K. Iijima1, Y. Sakakibara1, K. Ibaraki1, Y. Hirota1, K. Takei1, T. Saito2, T. Saido3, M. Sekiya1 1National Center for Geriatrics and Gerontology, Department Of Alzheimer's Disease Research, Obu,

Japan, 2Nagoya City University, Department Of Neurocognitive Science, Kawasumi−１, Japan, 3RIKEN,

Center For Brain Science, Wako-shi, Japan

Aims: Locus coeruleus (LC) is a nucleus containing the cell bodies of noradrenergic neurons in the brain and is one of the earliest regions affected by Alzheimer’s disease (AD). However, mechanisms underlying neurodegeneration in LC-noradrenergic system remain elusive. We have previously reported that AppNL-G-

F/NL-G-F mice, which harbor three familial AD mutations (Swedish, Beyreuther/Iberian, and Arctic) exhibited cognitive deficits and severe neuroinflammation accompanied by massive amyloid-β (Aβ) pathology. In this study, we asked whether and how LC- noradrenergic system is altered in AppNL-G-F/NL-G-F mice. Methods: To ask whether AppNL-G-F/NL-G-F mice exhibit noradrenergic degeneration, we compared the density of norepinephrine transporter (NET)-positive fibers in the cortex and hippocampus as well as the number of dopamine β-hydroxylase (DBH)-positive neurons in the LC between AppNL-G-F/NL-G-F and wild-type (WT) C57BL/6J mice at 24 months of age. We also examined whether tau pathology occurs in the LC-NE system in these mice. Results: Histochemical analyses revealed that 24-month-old AppNL-G-F/NL-G-F mice exhibited significant decreases in the density of NET-positive fibers in the prefrontal and entorhinal cortices and the hippocampal CA1, the regions associated with learning and memory, compared to WT mice. In contrast, AppNL-G-F/NL-G-F mice did not display loss of DBH-positive neurons in the LC even at 24 months of age. Furthermore, AppNL-G-F/NL-G-F mice did not develop prominent tau pathology either in the nerve terminals or in the cell bodies of LC neurons. Conclusions: This study demonstrates that Aβ pathology is sufficient to reduce the density of noradrenergic fibers independent of neuron loss and tau pathology in the LC.

P045 / #716


SPECIFIC PRESYNAPTIC LOSS IN THE OUTER MOLECULAR LAYER OF THE DENTATE GYRUS IN ALZHEIMER’S DISEASE

Lecture Title:

T. Jorda1, H. Haytural2, B. Winblad3, C. Mulle1, L. Tjernberg2, S. Frykman3, G. Barthet1 1Institut interdisciplinaire de neurosciences, Synapse Physiology, BORDEAUX, France, 2Karolinska Institutet, Division Of Neurobiology, Care Sciences And Society, Stockholm, Sweden, 3Karolinska Institutet, Dept Of Neurobiology, Care Sciences And Society, Stockholm, Sweden

Aims: Synaptic degeneration has been reported as the best pathological correlate of cognitive deficit in Alzheimer’s Disease (AD). However, the location of these synaptic alterations within hippocampal sub-regions, the vulnerability of the presynaptic versus postsynaptic compartments, and finally the causes of these impairments remain unknown. Here, we aimed to study synaptic changes in the OML of the human AD hippocampus Methods: We performed immunofluorescence labelings of different synaptic proteins in human hippocampal sections Results: Here we report reduced levels in AD of several presynaptic proteins of the neurotransmitter release machinery (complexin-1, syntaxin-1A, synaptotagmin-1 and synaptogyrin-1). This deficit was restricted to the outer molecular layer (OML) of the dentate gyrus whereas other hippocampal sub-fields where preserved. Interestingly, standard markers of postsynaptic densities (SHANK2) and dendrites (MAP2) were unaltered, as well as the number of granule cells in the dentate gyrus, indicating that the deficit is specifically presynaptic. Besides, there was no correlation between the reduction in presynaptic proteins in OML and the extent of the amyloid load or of the numbers of neurofibrillary tangles. Notably, the axonal components, myelin basic protein, SMI-312 and Tau, were unaffected whereas there was a remarkable abundance of Tau-immunoreactive neurites arranged in islands specific of the OML. Altogether, this study highlights the specific vulnerability of the OML of dentate gyrus and supports the notion of presynaptic failure in AD Conclusions: Altogether, this study highlights the specific vulnerability of the OML of dentate gyrus and supports the notion of presynaptic failure in AD

P046 / #1431


DECIPHERING THE ROLE OF THE FORMIN MDIA1 IN OAΒ1-42 SYNAPTOTOXICITY

Lecture Title:

A. Kumar1, M.E. Pero1, X. Qu1, J. Parato1, J. Teoh2, H. Zhang3, E. Argyrousi3, S. Choi4, E. Mosharov4, D. Sulzer4, O. Arancio3, F. Bartolini1 1Columbia University, Pathology And Cell Biology, New York, United States of America, 2Columbia University, Institute For Genomic Medicine, New York, United States of America, 3Columbia University, Taub Institute For Research On Alzheimer's Disease And The Aging Brain, New York, United States of America, 4Columbia University Medical Center, Psychiatry, New York, United States of America

Aims: Whether defects in MT dynamics and organization at synapses play a pathogenic role in neurological disease remains unknown. We recently found that activity-evoked de novo nucleation of dynamic MTs at excitatory presynaptic boutons is rate limiting for synaptic vesicle (SV) exocytosis by regulating the intrabouton transport of SVs and SV precursors (Qu et al., 2019). We also reported that the formin mDia1 contributes to oligomeric amyloid beta 1-42 (oAb1-42)synaptotoxicity through the modulation of MT dynamics (Qu et al., 2017) and tau phosphorylation.The mechanism by which mDia1 acts as a permissive factor in oAb1-42 synaptotoxicity through MT dynamics remains, however, not completely understood. Methods: We crossed mDia1 KO mice with J20 mice and compared the four resulting littermate genotypes in hippocampal synaptic plasticity, spatial memory function and spine density. Results: We found that presynaptic MT nucleation and interbouton SV bidirectional transport was perturbed by oAβ1-42 and this activity was abolished by loss of tau or mDia1 inhibition, implicating this MT pathway in the induction of oAb1-42- and tau-mediated synaptic injury. Strikingly, we found that even if mDia1 deficiency alone exhibited abnormal LTP, loss of mDia1 in J20/mDia1KO mice normalized LTP damage, memory deficits, spine density depletion and presynaptic synapsin 1 activation in J20 mice. Conclusions: Our findings reveal a novel role for mDia1 in regulating presynaptic MT nucleation perturbed by oAb1-42 and suggest that loss of its presynaptic MT activity in mDia1 KO mice may underlie the mechanism of synaptic rescue in mice affected by amyloid pathology.

P047 / #1440


CYP46A1 OVEREXPRESSION INDUCES SEX-SPECIFIC CHANGES IN SYNAPTIC FUNCTIONS IN AGED MICE.

Lecture Title:

M. Latorre Leal1, P. Rodriguez1, F. Eroli1, L. Franchini2, P. Merino-Serrais3, I. Björkhem4, A. Cedazo-Minguez1, S. Maioli1 1Karolinska Institutet, Nvs, Division Of Neurogeriatrics, Stockholm, Sweden, 2Università degli Studi di Milano, Pharmacological And Biomolecular Sciences, Milan, Italy, 3Cajal Institute, Functional And Systems Neurobiology, Madrid, Spain, 4karolinska Institutet, Laboratory Medicine, Stockholm, Sweden

Aims: CYP46A1 has been demonstrated to be implicated in neurodegenerative disorders such as Alzheimer’s disease (AD). In our lab, in vivo, and in vitro studies show a beneficial role of CYP461 overexpression and 24 (S)-hydroxycholesterol (24OHC) in cognitive functions. The purpose of this work is to investigate whether CYP46A1 activation and 24OHC can promote neuroprotection in a sex-specific manner by activating estrogen signaling. Methods: We have performed behavioral studies of males and females CYP46A1 overexpressing (Cyp46 Tg) mice, with high levels of 24OHC in the brain and plasma. The morphology of dendritic spines in hippocampal regions of Cyp46Tg mice has been investigated by Golgi Staining. 24OHC-treated hippocampal neurons from rat primary cultures were used for mechanistic studies. Results: Cyp46 Tg old females showed cognitive enhancement and increased levels of synaptic proteins in the hippocampus when compared to control mice (Maioli S, 2013). Noteworthy, these results were not found in old males, where CYP46A1 overexpression led to an impairment of spatial memory and an increase of anxiety like-behavior. These results were further confirmed by changes in the morphology of dendritic spines in the CA1 hippocampal region in these mice. Studies in vitro suggest that 24OHC is able to activate estrogen signaling in neurons. Conclusions: We show sex-specific differences in vivo studies in synaptic functions, where the neuroprotective effects promoted by the upregulation of CYP46A1 seem to be activated only in female mice. These novel findings can have a great clinical impact and help to define new targets for the treatment of AD.

P048 / #1420


ANATOMICAL AND FUNCTIONAL ANALYSES OF PARIETAL CORTEX SYNAPSES IN ALZHEIMER’S DISEASE REVEAL EXCITATORY/INHIBITORY IMBALANCE

Lecture Title:

J. Lauterborn1, P. Scaduto2, G. Lynch1, C. Gall1, A. Limon2 1University of California Irvine, Anatomy And Neurobiology, Irvine, United States of America, 2University of Texas Medical Branch at Galveston, Neurology, Galveston, United States of America

Aims: Disturbances in synaptic excitatory to inhibitory (E/I) balance in forebrain circuits are thought to contribute to the progression of Alzheimer’s disease (AD) and cognitive decline, although direct evidence for such imbalance in humans is lacking. Using anatomical and electrophysiological approaches, we evaluated AD subjects for global synaptic E/I ratios in the inferior parietal cortex (PCx), a region within the default mode network that is overly active in the disorder. Methods: Anatomical and electrophysiological synaptic E/I ratios in post-mortem samples from middle-aged individuals with AD (n = 5) or Down syndrome (DS; n = 6) compared to non-demented controls (n = 5) were assessed by fluorescence deconvolution tomography (FDT) and microtransplantation of synaptic membranes (MSM). Results: FDT analyses of layers 1 and 2 show that total counts of PSD-95 (excitatory) and gephyrin (inhibitory) immunoreactive puncta were not significantly different across groups; however, the ratio of the peaks of the immunolabeling frequency distributions for PSD-95 to gephyrin within the layer 2 was significantly larger in the AD group as compared to controls (Welch ANOVA, p = 0.0316). MSM analysis also showed that the maximal amplitude of ion currents through AMPARs and GABAARs were highly correlated across subjects, and that the AD group exhibited an increase in electrophysiological E/I ratio significantly greater than for controls (F (2,13) = 4.299, p < 0.0369). Conclusions: Our converging evidence suggests that a pro-excitatory synaptic imbalance may underlie the hyperexcitability and reduced resting state deactivation that have been consistently observed in the PCx of AD patients.

P049 / #1214


IL-17 TRIGGERS THE ONSET OF COGNITIVE AND SYNAPTIC DEFICITS IN EARLY STAGES OF ALZHEIMER'S DISEASE

Lecture Title:

L. Lopes1, M. Ribeiro2, H. Brigas2, J. Coelho2, R. Gomes3, V. Gomez-Murcia4, K. Carvalho4, S. Pereira2, A. Antunes De Almeida2, E. Faivre4, L. Buee4, P. Pousinha5, D. Blum4, B. Silva-Santos2, J. Ribot2 1Instituto de Medicina Molecular João Lobo Antunes, Faculdade de Medicina de Lisboa, Universidade De Lisboa, Lisbon, Portugal, 2Universidade de Lisboa, Instituto De Medicina Molecular, Faculdade De Medicina De Lisboa, Lisbon, Portugal, 3Faculdade de Ciencias de Lisboa, Universidade De Lisboa, Lisboa, Portugal, 4Inserm UMR-S 1172 Lille Neuroscience & Cognition, Alzheimer & Tauopathies, Lille, France, 5Centre National de la Recherche Scientifique, Universite de Cote d’Azur, Institut De Pharmacologie Moleculaire Et Cellulaire,, Valbonne, France

Aims: Neuroinflammation in Alzheimer’s disease (AD) patients and related mouse models have been recognized for decades, but the contribution of recently described resident meningeal immune population to AD pathogenesis remains to be addressed. While we have shown that meningeal gd T cell-derived IL-17 promotes synaptic plasticity and short-term memory at steady state (Ribeiro et al, Sci Immunology, 2019), the role of this cytokine in the context of AD neurodegeneration is unclear. Methods: Here, using the 3xTg-AD model, we report an accumulation of IL-17-producing cells, mostly gd T cells, in the brain and the meninges of female, but not male mice, concomitant with the onset of cognitive decline. Results: Critically, IL-17 neutralization into the ventricles was sufficient to prevent short-, but not long-term memory deficits, and synaptic plasticity dysfunction at early stages of disease. These effects preceded blood-brain barrier disruption and amyloid-beta or tau pathology, implying an early involvement of IL-17 in AD pathology. When IL-17 was neutralized at later stages of disease, the onset of short-memory deficits and amyloidosis- related splenomegaly were delayed. Conclusions: Altogether, our data suggests that meningeal accumulation of IL-17 in AD contributes to early synaptic dysfunction, thus supporting the idea that cognition relies on a finely regulated balance of “inflammatory” cytokines derived from the meningeal immune system. Preprint: https://papers.ssrn.com/sol3/papers.cfm?abstract_id=3689053 / Funding: Fundação para a Ciência e Tecnologia PTDC/BIM- MEC/4778/2014 e Santa Casa da Misericórdia MB-7-2018

P050 / #485


SYNAPSE IMBALANCE IN ALZHEIMER’S DISEASE: CONTRIBUTION OF THE SECRETED WNT ANTAGONIST DKK3

Lecture Title:

M. Podpolny, N. Martin Flores, F. Mcleod, P. Salinas UCL, Cell & Developmental Biology, London, United Kingdom

Aims: Alzheimer’s disease (AD) is manifested by synapses loss which best correlates to cognitive impairment. Wnt signalling plays important roles in synapse formation, stability and function. Increasing evidence suggests that deregulation of this pathway contributes to synapse vulnerability in AD. Recent studies showed that the secreted Wnt antagonist Dickkopf-3 (Dkk3) is increased in plasma and CSF of AD patients. Dkk3 also accumulates in Aβ plaques in the human brain. However, the function of Dkk3 in the brain remains unexplored. Here, we investigate the impact of Dkk3 on synapses and in AD. Methods: Dkk3 levels and localisation were analysed in the hippocampus of the J20 transgenic AD mouse model by western blot and immunofluorescence. In vivo loss-of-function of Dkk3 was performed by stereotactic injection of adeno-associated viruses. Changes in synapse number and function were evaluated by confocal microscopy and patch clamp recordings, respectively. Results: Our results demonstrate that Dkk3 secretion is enhanced prior to Aβ plaque formation in the AD mouse brain and accumulates at Aβ plaques, co-localising to dystrophic neurites. Our gain-of-function experiments show that Dkk3 reduces excitatory synapse number but increases inhibitory ones in the adult hippocampus. Consistently, Dkk3 decreases the frequency of mEPSCs whereas increases mIPSC frequency. In contrast, in vivo Dkk3 loss-of-function decreases inhibitory synapse number. Importantly, we found that Dkk3 loss-of-function in J20 mice ameliorates synapse imbalance before and after amyloid plaque deposition. Conclusions: Our studies identify Dkk3 as a new player in AD-mediated synapse imbalance and as a potential target to reduce synapse dysfunction in AD.

P051 / #1333


NEURONAL SUBTYPE VULNERABILITY TO BETA-AMYLOID INDUCED HYPER-EXCITABILITY

Lecture Title:

I. Martinsson1, L. Quintino2, C. Lundberg2, G. Gouras1 1Lund University, Experimental Dementia Research Unit, Experimental Medical Sciences, Lund, Sweden, 2Lund University, Cns Gene Therapy, Department Of Experimental Medical Science, Lund, Sweden

Aims: Multiple lines of evidence implicate the beta-amyloid peptide as a driver of hyper-excitability in Alzheimer’s disease. How beta-amyloid drives hyper-excitability and hyper-activity is currently poorly understood. Methods: In this study we use an in-vitro live-cell calcium imaging approach where we set out to dissect the role of beta-amyloid in network hyper-excitability. We used lentiviral tools to differentiate excitatory and inhibitory cell populations in culture. Results: We show that neurons from APP/PS1 mice have increased spontaneous calcium currents in CaMKII positive cells whereas addition of synthetic beta-amyloid in picomolar levels increased activity in both CaMKII positive and negative neurons. Interestingly, while adding picomolar beta-amyloid increased neuronal activity adding micromolar levels led to a decrease in calcium transients indicating that beta-amyloid could have dose dependent effects on neuronal activity. Further, treating neurons with thiorphan, an inhibitor of the beta-amyloid degrading enzyme neprilysin was able to increase calcium oscillations but had no effect in APP knockout culture. We show that exogenously added beta-amyloid has a strong preference for binding CaMKII positive neurons at synaptic sites Conclusions: Together our results suggest that beta-amyloid has concentration dependent roles in regulating synaptic activity. We hypothesize that a more complete understanding of the role of beta-amyloid and APP in neurons and in the development of AD could be important for development of better therapy for AD.

P052 / #1236


INHIBITION OF PHOSPHOLIPASE D1 AS THERAPEUTICS AGAINST ALZHEIMER’S DISEASE: PRECLINICAL STUDIES

Lecture Title:

B. Krishnan, C. Natarajan, C. Cook, K. Bourne The University of Texas Medical Branch, Neurology, Galveston, United States of America

Aims: We have conducted a preclinical investigation utilizing a small-molecule inhibitor of inducible isoform of Phospholipase D (PLD1) that is aberrantly recruited post-developmentally. We interpret this as a potential pathway amenable to therapeutics that can be potentially used in combination with other clinically validated approaches to prevent the progression of synaptic dysfunction and underlying memory deficits in AD and related dementia. Methods: Western blot analysis of synaptosomes derived from 3xTg AD mice hippocampus (model that has overexpression of presenelin 1gene (PSEN1), human amyloid precursor protein (APP) and microtubule associated protein tau (MAPT) driving neuropathological mechanisms comparable to that of human AD patients) were used to investigate neuronal PLD1 expression. Long -term potentiation was studied on ex vivo slices prepared from PLD1 inhibitor and control treated transgenic mice that were first assessed by Novel object recognition (NOR) and fear conditioning (FC) paradigms. Brain tissues from these animals were subjected to western blot analysis to understand the role of signaling mechanism that is perturbed /altered by overexpression of PLD. Results: Abnormally elevated levels of PLD1 drives synaptic mechanisms causing memory deficits associated with AD and related dementia. Conclusions: In our previous study, we have demonstrated the role of PLD1 overexpression in synaptic dysfunction and neuropathological mechanisms that drive AD and related dementia, using human clinical samples and 3XTg-AD mice. Here we characterize the role of PLD1 signaling mechanism in membrane trafficking, cytoskeletal reorganization, neuroinflammation and autophagy.

P053 / #846


IMPLICATION OF SFRP1 IN ALTERED SYNAPTIC PLASTICITY ASSOCIATED WITH ALZHEIMER'S DISEASE.

Lecture Title:

G. Pereyra Gomez1, M. Mateo1, M. Martin Bermejo1, P. Esteve Pastor1,2, P. Bovolenta1,2 1Centro de Biología Molecular Severo Ochoa - Consejo Superior de Investigaciones Científicas (CBMSO-CSIC), Tissue And Organ Homeostasis, Madrid, Spain, 2Centro de Investigación Biomédica en Red de Enfermedades Raras CIBERER, U709, Madrid, Spain

Aims: We have demonstrated that SFRP1 is a novel, promising therapeutic target for Alzheimer Disease (AD). SFRP1 acts as a secreted endogenous regulator of ADAM10, a brain a-sheddase which controls the activity of several substrates including APP and proteins regulating synaptic plasticity and neuroinflammatory crosstalk. Consistently, astrocyte-derived SFRP1 is upregulated in the brain of AD patients, localizing to amyloid plaques and interacting to Ab peptides. Neutralization of SFRP1 activity in AD-like mouse models decreases the formation of Ab peptides, counteracts brain inflammation and maintains synaptic plasticity. Here we aimed at determining if SFRP1 has a direct effect on synaptic plasticity and neuroinflammation. Methods: We generated a transgenic mouse model overexpressing Sfrp1 in astrocytes (GFAP-tTA;Sfrp1-tetO) and analysed it by immunohistochemistry and RT-qPCR to determine the possible presence of neuroinflammatory and molecular alterations. Mice were further characterized for cognitive abilities with behavioural tests and analysed for possible dendritic and spine modification via viral-mediated visualization of their morphology. Results: We show that GFAP-tTA;Sfrp1-tetO mice present an allele-dependent increase in SFRP1 expression, which is associated with a decrease in dendritic and spine density already at two months of age. These defects are associated with an age-dependent appearance of cognitive decline, the development of a low degree neuroinflammation and reduced expression of a number of genes normally down-regulated in AD patients. Conclusions: These data support the idea that SFRP1 has a direct impact on synaptic plasticity which is not secondary to its effect on APP processing, indicating that SFRP1 might have a pleiotropic effect in AD.

P054 / #1292


PILOT STUDY OF SYNAPTIC MRNA CHANGES IN THE EARLIEST STAGES OF ALZHEIMER'S DISEASE

Lecture Title:

J. Wiley1, D. Mack2, W. Ladiges3, P. Amieux4 1Sage Bionetworks, Neurodegeneration Research, SEATTLE, United States of America, 2University of Washington, Institute Of Stem Cell And Regenerative Medicine, Rehabilitation Medicine, Seattle, United States of America, 3University of Washington, Department Of Comparative Medicine, Seattle, United States of America, 4Bastyr University, Research, Kenmore, United States of America

Aims: Synaptic loss is one of the hallmark events in the early stages of Alzheimer disease (AD) pathogenesis. In this study, we examine changes in mRNA targeting to the synaptic compartment of AD transgenic mice. Our primary objective is to determine whether synaptic mRNA targeting changes in the earliest stages of AD pathology that could contribute to synaptic dysfunction. Methods: C57BL6 APPswe/PSEN1deltaE9 heterozygote transgenic mice were bred, and the transgene carriers and non-carriers were used as AD transgenic and wild-type controls respectively. 3 male AD transgenic and 3 wildtype mice were aged to 3 months (pre-pathology) or 6 months (early pathology). The hippocampus and cortex were removed and synaptosomes were generated and the mRNA was extracted and employed in whole genome microarray experiments to assess changes from pre-pathology to early AD pathology. Results: The synaptosomal mRNAs were enriched in synapse specific genes, with a set of nearly 2000 transcripts identified. The 3-month old animals demonstrated no difference in mRNAs between wild-type and transgenic animals, with the sole exception of up-regulation of APP and PSEN within the APPswe/PSEN1deltaE9 animals. The 6-month old animals demonstrated significant differences between the wild-type and transgenics with 92 up-regulated synaptic transcripts and 338 down-regulated transcripts. The upregulated transcripts were primary RNA processing, mitochondrial gene expression, and immune regulation related. In contrast, the down-regulated transcripts centered upon neuronal projections, ionic transport and synaptic plasticity. Conclusions: Synaptic mRNA localization changes in the earliest stages of AD pathology in APPswe/PSEN1deltaE9 transgenic mice that could impact core neuronal physiology, plasticity and metabolism.

P055 / #226

Topic: Theme A: β-Amyloid Diseases / A1.e. Disease Mechanisms, Pathophysiology: Cellular signaling, kinases, phosphatases, calcium

PKG PHOSPHORYLATES TAU PROTEIN AT DISTINCT SER/THR SITES

Lecture Title:

G. Montalto1, F. Caudano1, L. Sturla1, S. Bruzzone1, A. Salis1, G. Damonte1, J. Prickaerts2, E. Fedele3, R. Ricciarelli1 1University of Genoa, Department Of Experimental Medicine, Genoa, Italy, 2Maastricht University, Department Of Psychiatric And Neuropsychology, Maastricht, Netherlands, 3University of Genoa, Department Of Pharmacy, Genoa, Italy

Aims: Tau protein can be phosphorylated by multiple kinases at several sites. Among such kinases, the cAMP-dependent protein kinase A (PKA) phosphorylates tau at Ser214, an event that exerts a protective effect against the assembly of tau into paired helical filaments. The activation of PKA by cAMP-enhancing drugs also sustains long-term potentiation (LTP) through the stimulation of Aβ production. In a similar manner, cGMP was found to boost Aβ levels and to favor LTP and memory, but an effect of cGMP on tau phosphorylation has never been reported. To investigate this issue, we evaluated the possibility of a PKG-induced phosphorylation of tau. Methods: The cGMP enhancer vardenafil was used in different model systems: neuro N2a cells, rat hippocampal slices and adult male C57BL/6 mice. Tau phosphorylation was analyzed by immunoblotting, gene silencing, in vitro enzymatic assays and nano-HPLC mass spectrometry. Results: Our data show for the first time that the cGMP-activated PKG phosphorylates tau at Ser214. Additionally, other 7 Ser/Thr tau residues appear to be phosphorylated by this kinase, but not Ser202, which is considered one of the earliest markers of tau aggregation. Conclusions: Our results indicate the existence of a PKG-mediated mechanism that might shift tau from a pro-aggregant to an anti-aggregant conformation, which has been reported to exert neuroprotective functions.

P056 / #1346


SLEEP IS BI-DIRECTIONALLY MODIFIED BY AMYLOID BETA OLIGOMERS

Lecture Title:

G. Ozcan, S. Lim, J. Rihel University College London (UCL), Cell And Developmental Biology, London, United Kingdom

Aims: Disrupted sleep is a major feature of Alzheimer’s disease (AD), often arising years before symptoms of cognitive decline. Prolonged wakefulness exacerbates the production of amyloid-beta (Aβ), a major driver of AD progression. Previous studies have thus suggested that reduced sleep during AD may further accelerate Aβ accumulation and neuronal damage, creating a vicious cycle that leads to further neuronal dysregulation and increased sleep-wake cycle abnormalities (Roh et al., 2012). However, the mechanisms by which Aβ affects sleep are unknown. We tested whether Aβ may directly modulate sleep-regulatory pathways independently of neuronal cell death. Methods: We developed an injection assay in which the amount and type of the Aβ oligomers can be controlled. We generated Aβ oligomeric pools with significantly different lengths and assessed how each Aβ pool affected sleep and wake behavior in zebrafish using automated video-monitoring (Prober et al., 2006; Rihel et al., 2010). Results: We demonstrate that Aβ acutely and reversibly enhances or suppresses sleep and brain activity as a function of oligomer length and independently of neuronal cell death. Genetic disruptions revealed that short Aβ oligomers induce acute wakefulness through Adrb2 and Pgrmc1, while longer Aβ forms induce sleep through a pharmacologically tractable Prion protein signalling cascade. Conclusions: Our experiments indicate that Aβ modulates normal sleep/wake behavior due to acute signalling events. We propose that Aβ can trigger a bi-directional sleep/wake switch and alterations to the relative concentrations of different Aβ oligomeric forms during healthy aging and AD will have opposing consequences on sleep and wake behaviour.

P057 / #1521


NEURONAL KV2.1 POTASSIUM CHANNEL AS A POSSIBLE PLAYER IN HIPPOCAMPAL HYPEREXCITABILITY AND ENDOPLASMIC RETICULUM REMODELLING IN TG2576 ALZHEIMER’S DISEASE MICE

Lecture Title:

I. Piccialli1, M.J. Sisalli1, A. Secondo1, F. Boscia1, V. Tedeschi1, A. Pannaccione1, L. Annunziato2 1Federico II University of Naples, Italy, Department Of Neuroscience, Reproductive And Dentistry Sciences, Naples, Italy, 2Fondazione IRCSS, Sdn, Naples, Italy

Aims: Amyloid-β (Aβ)-induced ionic dyshomeostasis and endoplasmic reticulum (ER) Ca2+ dysregulation are crucial pathomechanisms in Alzheimer’s disease (AD), leading to neuronal damage and aberrant network activity. The neuronal KV2.1 voltage-gated channel, acting as a tethering protein inducing and remodelling plasmamembrane (PM)-ER junctions, in addition to its canonical K+ conducting function, has been implicated in AD-related neuronal hyperexcitability, despite its role has been poorly investigated. Hence, we explored the possible modulation of KV2.1 expression and activity in the Tg2576 mouse model mimicking Aβ pathology and the implication of KV2.1 PM clusters in the modulation of PM-ER junctions, which are important sites of Ca2+ signalling between PM and ER, in Tg2576 hippocampal neurons. Methods: We performed 1) electrophysiological recordings to assess KV2.1 activity, 2) Western blot and immunofluorescence analyses to assess KV2.1 protein expression, and 3) immunofluorescence analyses to analyze ER morphology, in both primary hippocampal neurons and in the hippocampus from wild-type and Tg2576 mice. Results: Tg2576 hippocampal neurons displayed upregulated KV2.1 protein expression accompanied by increased cell surface channel clustering in comparison to wild-type neurons. Meanwhile, in accordance with previous studies, increased channel clustering was associated to a significant reduction of KV2.1-mediated currents and a concomitant increase of neuronal excitability. Interestingly, primary Tg2576 hippocampal neurons displayed changes in ER morphology in comparison to wild-type neurons. Conclusions: Our results show that increased KV2.1 clustering may be associated to neuronal hyperexcitability and changes in ER morphology observed in Tg2576 hippocampal neurons, thus bringing out KV2.1 channels as a possible target in AD.

P058 / #1675


BLOCKING H-RAS SIGNALING RESCUES MEMORY DEFICITS IN ALZHEIMER’S MICE

Lecture Title:

W. Qu1, A. Jeong2, L. Li2 1University of Minnesota, Graduate Program In Neuroscience, Minneapolis, United States of America, 2University of Minnesota, Department Of Experimental And Clinical Pharmacology, Minneapolis, United States of America

Aims: Alzheimer’s disease (AD) affects millions of people without effective treatment. The molecular mechanism of AD remains obscure. Emerging evidence suggests that a key post-translational lipid modification of proteins, called farnesylation, plays an important role in the pathogenesis of AD. As an exclusively farnesylated protein, the small GTPase H-Ras governs essential cellular functions including cell-cycle regulation and neuronal synaptic plasticity. Both upstream and downstream signaling of H-Ras is elevated in AD, leading to the hypothesis that blocking H-Ras signaling mitigates the pathogenesis of AD. Methods: To test this hypothesis, H-Ras knockout mice were crossed with transgenic APP/PS1 mice of AD, followed by assessment of cognitive function and neuropathology. A battery of behavioral tests including the open-field, the elevated-plus maze, and the Morris water maze, were conducted to evaluate their cognitive function. Amyloid deposition and neuroinflammation are quantified by immunohistochemical staining. Dendritic spine density/morphology, synaptic markers, and H-Ras signaling molecules are assessed using Golgi staining, immunoblot analysis, and transcriptomic analysis, respectively. Results: Our preliminary data show that H-Ras deficiency rescues memory deficits in APP/PS mice. Consistent with the data in mice, activation of the H-Ras/ERK pathway correlates with cognitive decline and AD neuropathology in human brains. Further analyses are underway to elucidate the molecular mechanisms underlying the role of H-Ras in AD. Conclusions: Our preliminary results demonstrate that blocking H-Ras signaling preserves memory function in AD mice, opening a novel therapeutic avenue for treating AD.

P059 / #1159


DATA DRIVEN NETWORK RECONSTRUCTION OF PATHWAY MODELS IN ALZHEIMER’S DISEASE

Lecture Title:

J. Wiley1, J. Gockley2, J. Young3, B. Logsdon1, A. Greenwood1, L. Mangravite1 1Sage Bionetworks, Neurodegeneration Research, SEATTLE, United States of America, 2Sage Bionetworks, Neurodegeneration Research, Seattle, United States of America, 3University of Washington, Laboratory Medicine And Pathology, Seattle, United States of America

Aims: In order to aid target selection in ongoing AD drug development efforts, we built an application that draws from the Pathway Commons (PC) data resource, dynamically reconstructs pathway models based on seed clusters inputted into the system, and we use it to analyze a kinase dataset identified in a high-throughput genetic knockdown study. Methods: PC is an integrated resource of 22 different pathway databases (e.g., Reactome, KEGG, and Wikipathways) and protein interaction databases (e.g., BioGrid, IntAct and BIND) built upon the Biological Pathway Exchange (BioPax) language. The individual undirected protein-protein interactions and directed nodal binaries are employed as individual construction elements to link gene clusters or modules submitted to the resource. The connection algorithm makes the fewest number of inferences to create an interaction network. Sentinel AD genes are selected to represent disease linked processes and provide anchors to network reconstruction. Results: In order to test the network reconstruction capacity of the application, we employed an identified kinase set that was identified through high-throughput screening (HTS) genetic knockdown studies of APP proteolysis. The seed kinases were employed in the reconstruction with APP processing related sentinel genes. The application revealed numerous non-obvious connections between nodes in the kinase set, with strong centrality around APP and kinase motifs associated with neuroimmune signaling. Conclusions: The data driven network reconstruction of the HTS data gave novel insights into potential signaling mechanisms linking kinases to AD disease process, suggesting this may be a viable approach to nominate candidate hypothetical mechanisms and future drug targets.

P060 / #882

Topic: Theme A: β-Amyloid Diseases / A1.f. Disease Mechanisms, Pathophysiology: Lysosomes, ubiquitin, proteasome, ER stress, chaperones

IMPAIRMENT OF THE ENDOSOME-LYSOSOME PATHWAY BY THE ALZHEIMER DISEASE GENETIC RISK FACTOR BIN1 ISOFORM1

Lecture Title:

P. Dourlen1, E. Lambert1, F. Abdelfettah1, N. Barois2, D. Vandekerkhove3, X. Hermant1, A.-M. Ayral1, L. Davoine1, C. Dupont1, F. Lafont2, P. Amouyel1, J. Chapuis1, P. Verstreken3, B. Dermaut1,4, J.-C. Lambert1 1Institut Pasteur de Lille, Univ. Lille, CHU Lille, Inserm U1167, LILLE, France, 2CNRS UMR 8204 – INSERM U1019, Institut de Biologie de Lille, Institut Pasteur de Lille, Centre D'infection Et D'immunité De Lille, Lille, France, 3KU Leuven, Department of Neurosciences, Laboratory for Neuronal Communication, Vib Center For Brain & Disease Research, Leuven, Belgium, 4Ghent University Hospital, Center For Medical Genetics, Ghent, Belgium

Aims: BIN1 is the most associated risk gene for AD after APOE. A 3bp insertion allele was shown to increase BIN1 expression in human brains and AD risk. The contribution of BIN1 and its isoforms to AD pathogenesis remains unclear. The objective of this work was to assess the neurotoxicity of BIN1 isoforms and understand the underlying mechanisms. Methods: We generated Drosophila transgenic lines expressing brain, muscular and ubiquitous human BIN1 isoforms 1, 8 and 9 respectively. We analyzed BIN1 isoform neurotoxicity using cornea neutralization, immunofluorescence, electrophysiology and electron microscopy in the fly eye photoreceptor neurons. Results: Adult eye-specific expression of brain BIN1 isoform1 only resulted in an age-dependent loss of photoreceptor neurons. This neurodegeneration was characterized by electrophysiological defects starting with altered synaptic transmission. In addition, photoreceptor neuron degeneration was characterized by a strong accumulation of vesicles with endosomal markers suggesting a blockade of the endosome-lysosome pathway. Finally, modulation of Rab5, Rab4 and Rab11 abrogated photoreceptor degeneration, indicating that BIN1 isoform1 neurotoxicity is due to the blockade of the endosome-lysosome pathway likely at the level of early endosomes. Conclusions: Altogether, these results show that an increase of the BIN1 isoform1 impairs the endosome-lysosome pathway, thereby affecting synaptic transmission and inducing neurodegeneration. Interestingly, enlarged endosomes have been shown to be one of the first cytopathological markers of the disease and synaptic transmission is affected early in AD pathogenesis. These results suggest that an increase in BIN1 isoform1 could contribute to early steps of Alzheimer’s disease.

P061 / #795


CHARACTERIZATION OF THE NEUROTOXICITY OF THE ALZHEIMER DISEASE RISK FACTOR BIN1 IN DROSOPHILA

Lecture Title:

E. Lambert1, F. Abdelfettah1, N. Barois2, D. Vandekerkhove3, X. Hermant1, A.-M. Ayral1, L. Davoine1, C. Dupont1, F. Lafont2, P. Amouyel1, J. Chapuis1, P. Verstreken3, B. Dermaut1,4, J.-C. Lambert1, P. Dourlen1 1Institut Pasteur de Lille, Univ. Lille, CHU Lille, Inserm U1167, LILLE, France, 2CNRS UMR 8204 – INSERM U1019, Institut de Biologie de Lille, Institut Pasteur de Lille, Centre D'infection Et D'immunité De Lille, Lille, France, 3KU Leuven, Department of Neurosciences, Laboratory for Neuronal Communication, Vib Center For Brain & Disease Research, Leuven, Belgium, 4Ghent University Hospital, Center For Medical Genetics, Ghent, Belgium

Aims: BIN1 has more than 10 isoforms. Preliminary results have shown that the expression of the brain-specific isoform (BIN1-1) specifically induces photoreceptor neuron degeneration with age in the Drosophila eye. The degeneration is also characterized by endosome-positive large vesicles. The first objective of this work was to identify the domain of BIN1-1 responsible for the degeneration. The second objective was to evaluate the impact of the endosome-lysosome pathway regulation on the BIN1-1 induced toxicity. Methods: Using the drosophila model, we tested truncated forms of BIN1-1 for the exon 7 and the CLAP domain and regulators of the endosome-lysosome pathway (clathrin-mediated endocytosis, early/recycling/late endosomes, lysosome, PIP2 metabolism). We quantified photoreceptor neurodegeneration thanks to the cornea neutralization technique and confocal microscopy in living flies. Results: We showed that truncation of the CLAP domain abolished BIN1-1-induced degeneration, indicating that this domain is necessary for the BIN1-1 induced toxicity. This domain is involved in endocytosis, a process related to the endosome-lysosome pathway. To assess if an alteration of the endosome lysosome pathway could be a cause of the degeneration, we tested the effect of the modulation of this pathway on BIN1-1 associated degeneration. We found that modulators of the early and recycling endosomes, Rab5 and Rab11 respectively, abrogated photoreceptor degeneration. Conclusions: Altogether, these results suggest that BIN1 isoform 1 could impair the endosome-lysosome pathway at the level of early and/or recycling endosomes, leading to neuronal death and thus contribute to Alzheimer’s disease.

P062 / #395


VEGETABLE OIL-DERIVED HYDROXYNONENAL CAUSES DIVERSE CELL DEATH VIA HSP70.1 DEPLETION

Lecture Title:

T. Yamashima Kanazawa University Graduate School of Medical Sciences, Psychiatry & Neurobiology, Cell Metabolism & Nutrition, Kanazawa City, Japan

Aims: Ample evidence implicates reactive oxygen species for Alzheimer neurodegeneration, however, the underlying mechanism remains unelucidated. Recent data advocate for dual roles of Hsp70.1 not only as molecular chaperone for damaged/aged proteins but also as guardian of lysosomal integrity. Thus, in case of Hsp70.1 dysfunction, not only failure of protein degradation but also lysosomal destabilization occurs. In the monkey experimental paradigm, the author’s group previously found that Hsp70.1 can become an in-vivo target of carbonylation by hydroxynonenal. Further, in the in-vitro experiments Hsp70.1 carbonylated by hydroxynonenal was found to be susceptible to cleavage by activated μ-calpain. Here, I focused on hydroxynonenal as a cause of diverse cell death. Methods: Monkeys after consecutive injections of the synthetic hydroxynonenal were histologically studied to determine whether it can induce cell degeneration/death in the brain, liver, and pancreas. Results: In all of the five monkeys injected, neurons, hepatocytes, and β-cells after hydroxynonenal injections revealed similar microcystic degeneration and scattered cell death by light microscopy. By electron microscopy, degenerating cells generally showed lysosomal rupture, dilatation of rough ER, and accumulation of autophagosomes containing cell debris. Immunofluorescence histochemical analysis showed an increased colocalization of activated µ-calpain and Hsp70.1, and extralysosomal release of cathepsin B. Western blotting showed increments of µ-calpain activation and calpain-mediated Hsp70.1 cleavage. Conclusions: Targeting ‘hydroxynonenal’ would help elucidate the pathogenesis of not only Alzheimer’s disease but also related lifestyle diseases. Since ω-6 PUFAs can induce both GPR40/109A activation leading to calpain activation, Hsp70.1 carbonylation, and calpain-mediated cleavage of carbonylated Hsp70.1. Hydroxynonenal was thought to be a real culprit of Alzheimer's disease.

P063 / #384

Topic: Theme A: β-Amyloid Diseases / A1.g. Disease Mechanisms, Pathophysiology: Mitochondrial dysfunction, oxidative damage

MITOCHONDRIAL QUALITY CONTROL ABNORMALITIES IN ALZHEIMER'S DISEASE

Lecture Title:

K. Barnes, S. Bell, L. Ferraiuolo, D. Blackburn, H. Mortiboys University of Sheffield, Sheffield Institute For Translational Neuroscience, Sheffield, United Kingdom

Aims: Mitochondrial dysfunction has long been associated with Alzheimer’s disease (AD), including dysfunction seen in complex IV of the electron transport chain, reduced mitochondrial membrane potential, and reduced levels of ATP. Mitochondrial morphology has also been seen to be altered. The aim of this study is to understand the pathogenic mechanisms leading to mitochondrial abnormalities in AD using patient cells, focusing on mitochondrial dynamic processes, fission and fusion. Methods: Control and AD patient fibroblasts are cultured from skin biopsies. Protein levels of key proteins involved in mitochondrial fission were assessed via western blotting. Results: Our previously published work shows an increased percentage of long mitochondria in AD patient fibroblasts compared to controls, as well as a lower mitochondrial count, suggesting a more fused mitochondrial network. Furthermore, reduced levels of mitochondrial fission protein, Drp1, were seen. Protein levels of the four Drp1 receptors on the outer mitochondrial membrane were also assessed, and a significant decrease was seen in Fis1, Mff, and MiD49, in AD patient cells. However, no significant difference was seen in MiD51. Conclusions: Three of the four Drp1 receptors on the outer mitochondrial membrane are reduced. Current work is investigating Drp1 interactions with each receptor, as well as post-translational modifications of Drp1. Future work will focus on upstream modulators of mitochondrial fission, including pre-constriction sites, as well as studying these processes in neurons. Fibroblasts will be directly converted to neural progenitor cells, which retain the ageing features of the donor, before being differentiated into neurons, which will then be assessed for mitochondrial phenotypes.

P064 / #1102


PRELIMINARY EVIDENCE OF DYSFUNCTIONAL GLP-1 SIGNALING AND OXIDATIVE STRESS IN GASTROINTESTINAL SYSTEM OF A RAT MODEL OF SPORADIC ALZHEIMER'S DISEASE

Lecture Title:

J. Homolak1,2, A. Babic Perhoc1,2, A. Knezovic1,2, J. Osmanovic Barilar1,2, M. Salkovic-Petrisic1,2 1University of Zagreb School of Medicine, Croatian Institute For Brain Research, Zagreb, Croatia, 2University of Zagreb School of Medicine, Department Of Pharmacology, Zagreb, Croatia

Aims: Gastrointestinal (GI) hormone glucagon-like peptide-1 (GLP-1) is being explored as a potential treatment in sporadic Alzheimer's disease (sAD). The aim of our research was to analyze oxidative stress (OS) and GLP-1-related signaling in the GI tract of rats following intracerebroventricular administration of streptozotocin (STZ-icv). Additionally, the effect of central inhibition of GLP-1 receptor (GLP-1R) on the GI system was explored. Methods: Male Wistar rats (n=40) were treated with STZ-icv (3mg/kg) or vehicle and either 85ug/kg Exendin9-39 (Ex-icv) or saline after one month. Duodenum and ileum were extracted. Catalase, superoxide dismutase, lipid peroxidation, oxidation-reduction potential and nitrocellulose redox permanganometry were analyzed spectrophotometrically. GLP-1R expression and distribution were analyzed by quantitative immunofluorescence. Results: Our results suggest for the first time that STZ-icv affects the GI system, disrupting GLP-1 homeostasis and inducing OS. Interestingly, the effect of STZ-icv and Ex-icv, and differences in the response to central GLP-1 inhibition were more pronounced in duodenum than in ileum, both for OS and GLP-1R. Conclusions: Dysfunctional GI GLP-1 signaling and GI OS are present in the STZ-icv rat model of sAD. Pronounced susceptibility of duodenum to the central GLP-1 (ili metabolic) regulation should be further explored in the context of sAD etiopathogenesis.

P065 / #901


NEURONAL MITOCHONDRIAL FUSION PROTECTS C. ELEGANS AGAINST AGE AND AΒ-MEDIATED LOSS OF NEURONAL PLASTICITY

Lecture Title:

C. Escoubas, V. Laversenne, E. Tabakovic, H. Weir, N. Clark Harvard T.H. Chan School of Public Health, Molecular Metabolism, Boston, United States of America

Aims: Our objective is to identify metabolic pathways contributing to age-related cognitive decline and assess whether modulating these pathways have beneficial effects in an Alzheimer's disease (AD) animal model. In this study, we investigate the effect of neuronal mitochondria fusion and fission on associative learning in C. elegans and the potential beneficial effect of AMPK activation in this context. Methods: To assay neuronal plasticity in C. elegans, we used a negative associative learning protocol. This protocol is based on the ability of C. elegans to modulate its behavior (chemotaxis) after forming an association between a neutral element in the media (NaCl) with a deleterious cue (starvation). Results: C. elegans lose their ability to form associative learning with age, and this process is modulated by mitochondrial fragmentation in the neurons. By maintaining mitochondrial homeostasis, AMPK activation reverses this age-dependent loss of plasticity. Based on these data, we investigate the effect of amyloid beta (Aβ) accumulation in the neurons. As expected, animals expressing neuronal Aβ have impaired associative learning even at a young age. Interestingly, AMPK activation was able to rescue this Aβ-mediated loss of neuronal plasticity. Conclusions: AMPK activation both rescues age-mediated and Aβ-meditated loss of neuronal plasticity. As AMPK contributes to neuronal health during aging by maintaining a connected mitochondrial network, we believe that the modulation of mitochondria homeostasis to maintain a healthy mitochondrial network could be protective against neuronal Aβ accumulation.

P066 / #573


AAV-C99 MICE: A NEW AD MODEL WITH LYSOSOMAL AND MITOCHONDRIAL ALTERATIONS

Lecture Title:

R. Pardossi-Piquard1, L. Vaillant-Beuchot2, M. Arnaud1, A. Bourgeois1, I. Lauritzen1, M. Chami1, F. Checler1 1Université Cote d'Azur- CNRS UMR7275, Institut De Pharmacologie Moléculaire Et Cellulaire, Valbonne, France, 2Institut de Pharmacologie Moléculaire et Cellulaire, Cnrs Umr 7275, Valbonne, France

Aims: The 3xTgAD (APPswe, TauP301L, PS1M146V) mice recapitulate both Abeta and Tau pathology of Alzheimer’s disease (AD) with synaptic and memory deficits. In these mice, we have reported an early accumulation of C991 that induces an Abeta-independent lysosomal/autophagic dysfunction2. We have also demonstrated that C99 contributes to synaptic alterations, apathy-like behavior and spatial learning deficits3. To focus on the C99-mediated pathology in wild-type mice devoid of any other AD-related mutations, we have developed a new mouse model based on C99 viral expression. Methods: Newborn mice are injected into the ventricle with an adeno-associated virus AAV expressing C99 (AAV-C99). To discriminate between C99 and Abeta toxicity, we treated mice with the gamma-secretase inhibitor ELND006 (D6) that blocks Abeta production and enhances C99 accumulation. Results: Mice injected with AAV-C99 virus (AAV-C99 mice) display early expression of C99 in hippocampus and cortex as well as Abeta accumulation and plaques formation in old animals. In this model, we also observed an Abeta-independent lysosomal/autophagic dysfunction2 and more recently we reported mitochondrial and mitophagic alterations associated to C99 expression4. Conclusions: Overall, we developed a new AD mouse model for studying specifically the amyloidogenic pathway. The AAV-C99 mice display C99 accumulation, lysosomal/autophagic dysfunction, mitochondrial and mitophagic alterations, Abeta production and plaques formation. Thus C99 could contribute to early stage of AD pathology while Abeta could be involved at later stages of disease progression. 1-Lauritzen et al., J Neurosci. 2012;32:16243-55 2-Lauritzen et al., Acta Neuropathol. 2016;132:257-76 3-Bourgeois et al., Neurobiol Aging. 2018;71:21-31 4-Vaillant-Beuchot and Mary et al., Acta Neuropathol. 2020, under revision

P067 / #1082


ENDOTHELIAL MITOCHONDRIA AS A MEDIATOR OF VASCULAR DYSFUNCTION IN CAA IN THE PRESENCE OF HYPERHOMOCYSTEINEMIA

Lecture Title:

R. Parodi-Rullan1, J. Sone2, S. Fossati1 1Temple University, Alzheimer's Center At Temple (act), Philadelphia, United States of America, 2NYU School of Medicine, Department Of Psychiatry, NY, United States of America

Aims: Cerebral Amyloid Angiopathy (CAA) pathology may be potentiated by cardiovascular (CV) risk factors through vascular dysfunction, contributing to neurodegeneration. The mitochondria, mediators of cell-survival and death, may play a crucial role in disease pathogenesis. The role of brain endothelial (EC) mitochondria in CAA complicated by CV risk factors is unknown. We have previously shown that carbonic anhydrase inhibitors (CAi) reduce amyloid-induced cell death and mitochondrial dysfunction. Here, we aim to understand the role of the mitochondria in the pathogenesis of mixed AD and vascular dementias, and the effects of CAi in preventing the toxic effects of amyloid β (Aβ) and hyperhomocysteinemia (HHcy). Methods: Human ECs were challenged with Aβ and HHcy in the presence or absence of CAi. Mitochondrial and blood-brain barrier (BBB) effects were evaluated. Results: BBB permeability was increased by both Aβ and HHcy, and HHcy worsened amyloid-induced barrier permeability. These effects were prevented by the CAi methazolamide. Aβ impaired mitochondrial respiration and ATP production in ECs after 24h but not after 3-or-6 hours; HHC had no additive effects. CAi prevented Aβ-induced cytochrome-c release, mitochondrial membrane potential loss, and ROS production. Conclusions: Both HHcy and Aβ had detrimental effects on BBB resistance, and HHcy worsened the effects of Aβ. Methazolamide protected the EC barrier in presence of Aβ and HHcy. Mitochondrial dysfunction was observed 24 h post-Aβ treatment, although it was not increased by HHcy. Our studies support the importance of better understanding mitochondrial and EC pathways responsible for cerebrovascular dysfunction in CAA and mixed dementias.

P068 / #1796


MITOCHONDRIAL RESPIRATORY PROTEIN COMPONENTS IN NEURONAL EXTRACELLULAR VESICLES POINT TO NEURONAL MITOCHONDRIAL DYSFUNCTION IN ALZHEIMER’S DISEASE

Lecture Title:

P. Yao, E. Eren, E.J. Goetzl, D. Kapogiannis NIA/NIH, Laboratory Of Clinical Investigation, Baltimore, United States of America

Aims: Neuronal mitochondrial abnormalities are involved in Alzheimer’ disease (AD) pathogenesis by decreasing energy supply and generating reactive oxygen species. Circulating extracellular vesicles (EVs) contain mitochondrial cargo (proteins, mt-RNA, mt-DNA). Here, we sought to examine neuronal-enriched (N)EVs isolated from plasma of AD patients show reduced activity and/or levels of mitochondrial respiratory protein components and superoxide dismutase 1 (SOD1) compared to controls. Methods: We isolated NEVs from 29 patients with early AD and 22 cognitively normal age/sex-matched controls and measured complex I, III, IV, V and SOD1. In a second cohort of 14 patients with early AD and 14 age/sex-matched controls, we measured activity of complex IV and V, and superoxide dismutase 1 in the EVs of AD Results: AD patients compared to controls had reduced levels of complex I (306 ± 39.6 vs 1254 ± 135 pg/ml, p<0.0001), III (399 ± 24.2 vs 1692 ± 153 pg/ml, p<0.0001), IV (3448 ± 488 vs 6209 ± 752 pg/ml, p=0.0061), V (2762 ± 221 vs 4863 ± 213 pg/ml, p<0.0001) and SOD1 (769 ± 96.4 vs 2252 ± 313 pg/ml, p<0.0001). AD patients compared to controls had impaired activity of complex IV (0.14 ± 0.003 vs 1.27 ± 0.43 normalized activity, p = 0.0214) and V (0.26 ± 0.019 vs 0.87 ± 0.096 normalized activity, p<0.0001) Conclusions: NEV biomarkers provide evidence for neuronal mitochondrial dysfunction in AD. Mitochondrial respiratory protein reductions suggest reduced capacity for ATP generation, whereas SOD1 reduction suggests impaired defenses against oxidative stress.

P069 / #345


DISCOVERY AND CHARACTERIZATION OF A SMALL MOLECULE AS MITOCHONDRIA COMPLEX I MODULATOR

Lecture Title:

S. Zhang1, J. Green1, Q. Chen2, Y. Xu1 1Virginia Commonwealth University, Medicinal Chemistry, Richmond, United States of America, 2Virginia Commonwealth University, Internal Medicine, Richmond, United States of America

Aims: The aims of this study are to understand the mechanism of action for a small molecule compound that exhibits promising neuroprotectant activities both in vitro and in vivo. Methods: Chemical biology, biochemical, and cellular/moleular biology assays were employed to characterize the compound for its effects on the OXPHOS and complex I activity. Results: Our studdies demonstarted that the compound inhibited the complex I activity in both NADH oxidase and NFR assays, thus suggesting that it interacts with the FMN binding site of complex I. Photoaffinity labeling studies also demonstrated that a probe derived from the parent compound mainly recognizes the components of the FMN binding site of mitochondrial complex I. The competetion with known complex I inhibitors or substrates suggested a novel model of action for this compound. A cellular thermal shift assay also supported its interaction with the NDUFV1 subunit. Conclusions: The results from the studies strongly suggest that the small molecule compound specifically interacts with the FMN binding site of the mitochondiral complex I and moderately modulates the complex I activity. This represent a novel mechanism of action. Given the unclear roles of mitochondrial dysfunction and oxidative stress in AD and other neurodegenerative disorders, this compound may serve as a chemical provbe to help understanf the pathological roles of complex I and mitochondiral dysfunction in these diseases, thus facilitating development of mitochondria targted therapeutics.

P070 / #1395

Topic: Theme A: β-Amyloid Diseases / A1.h. Disease Mechanisms, Pathophysiology: Lipids, lipoproteins and membrane trafficking

UPREGULATION OF RIN3 INDUCES ENDOSOMAL DYSFUNCTION IN ALZHEIMER’S DISEASE

Lecture Title:

R. Shen1,2, C. Wu1, J. Ding2 1UCSD, Neuroscience Department, san diego, United States of America, 2Ruijin Hospital, Neurology, Shanghai, China

Aims: Ras and Rab Interactor 3(RIN3) is a guanine nucleotide exchange factor (GEF) for the Rab5 small GTPase family and has been implicated to be a risk factor for both late onset AD (LOAD) and sporadic early onset AD (sEOAD). However, how RIN3 is linked to AD pathogenesis is currently undefined, our work will dedicate to show how RIN3 work in AD pathogenesis. Methods: Quantitative PCR and immunoblotting were used to measure the RIN3 expression level in mouse brain tissues and cultured basal forebrain cholinergic neuron (BFCNs). Recombinant flag-tagged RIN3 protein was purified from HEK293T cells and was used to define RIN3-interactomes by mass spectrometry. Live imaging of primary neurons was used to examine axonal transport of amyloid precursor protein (APP) and β-secretase 1 (BACE1). Immunoblotting was used to detect protein expression, processing of APP and phosphorylated forms of Tau. Results: RIN3 mRNA level was significantly increased in the hippocampus and cortex of APP/PS1 mouse brain. Basal forebrain cholinergic neurons (BFCNs) cultured from E18 APP/PS1 mouse embryos also showed increased RIN3 expression accompanied by early endosome enlargement. In addition, via its proline rich domain, RIN3 recruited BIN1 and CD2AP, two other AD risk factors, to early endosomes. Interestingly, overexpression of RIN3 or CD2AP promoted APP cleavage to increase its carboxyl terminal fragments (CTFs) in PC12 cells. Upregulation of RIN3 or the neuronal isoform of BIN1 increased phosphorylated Tau level. Conclusions: RIN3 is significantly upregulated and correlated with endosomal dysfunction in APP/PS1 mouse. Increased RIN3 expression alters axonal trafficking and procession of APP.

P071 / #1096

Topic: Theme A: β-Amyloid Diseases / A1.h. Disease Mechanisms, Pathophysiology: Lipids, lipoproteins and membrane trafficking

SMALLER HDL PARTICLES IN CEREBROSPINAL FLUID ARE ASSOCIATED WITH HIGHER CSF Β-AMYLOID LEVELS AND BETTER COGNITION

Lecture Title:

H. Yassine1, A. Martinez1, G. Weissberger2, X. He1, H. Chui1, M. Harrington3, S. King4, D. Han1, R. Krauss4 1University Southern California, Keck School Of Medicine, Los Angeles, United States of America, 2Bar-Ilan, Medicine, Ramat Gan, Israel, 3Huntington Medical Research Institutes, Neuroscience, Pasadena, United States of America, 4University of California, San Francisco, Pediatrics, Oakland, United States of America

Aims: In animal models of Alzheimer’s disease (AD), enhancing brain ATP binding cassette 1 (ABCA1) expression or activity reduces brain amyloid accumulation. Small HDL particles are activators of ABCA1 mediated cholesterol transport. However, the relationships between cerebrospinal fluid (CSF) HDL particles with ABCA1 activity and brain amyloid pathology are not known. Our aim was to determine the association of CSF HDL particle concentrations with CSF Aβ42, ABCA1 cholesterol activity of CSF, and measures of cognition. Methods: HDL particle concentrations as a function of particle diameter by ion mobility in CSF and plasma were measured in 181 older participants with no cognitive impairment or mild cognitive impairment. Particles spanning the size range of plasma HDL were grouped into two categories in both plasma and CSF based on their diameter: small (7.00 – 10.5 nm), and large HDL particles (10.5 – 14.5 nm). The association of HDL particle concentrations with CSF Aβ42 levels, CSF ABCA1 activity, and neuropsychological performance were assessed. Results: A positive correlation between levels of small HDL, but not large HDL particles, in CSF and plasma was observed. Levels of small CSF HDL particles were positively associated with CSF Aβ42 peptide levels, and better performance in three domains of cognitive function (memory, attention/executive function, and global cognition) independent of age, sex, education years, or APOE4. Greater levels of small HDL particles were also associated with greater ABCA1 mediated cholesterol efflux in CSF. Conclusions: Higher levels of small CSF HDL particles may be protective against AD and could represent novel targets for AD prevention.

P072 / #495

Topic: Theme A: β-Amyloid Diseases / A1.i. Disease Mechanisms, Pathophysiology: Microglia

PLAQUE-ASSOCIATED HUMAN MICROGLIA ACCUMULATE LIPID DROPLETS IN A CHIMERIC MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

C. Claes, J. Hasselmann, E. Danhash, J. Silva Hidalgo, M. Coburn, J.P. Chadarevian, T.E. Lim, S. Kiani Shabestari, H. Davtyan, M. Blurton-Jones Institute for Memory Impairments & Neurological Disorders Sue & Bill Gross Stem Cell Research Center, Department Of Neurobiology & Behavior, Irvine, United States of America

Aims: Disease-associated microglia (DAMs), that surround beta-amyloid plaques, represent a transcriptionally-distinct microglial profile in Alzheimer's disease (AD). Activation of DAMs is dependent on triggering receptor expressed on myeloid cells 2 (TREM2) and the AD TREM2-R47H risk variant reduces microglial activation and plaque association in carriers. Interestingly, TREM2 has also been identified as a microglial lipid-sensor. However, whether lipid droplets (LDs) are present in DAMs in vivo and how the R47H mutation affects this, so far remains unknown. Methods: TREM2-R47H (homozygous) and TREM2-WT human stem cell-derived microglia were transplanted in chimeric AD mice. 7 months after transplantation, mice were sacrificed and human microglia extracted for single-cell RNA sequencing. Half brains were used for quantitative immunohistochemical analysis. Results: Here we report that the transcriptome of human wild-type TREM2 and isogenic TREM2-R47H DAM xenografted microglia (xMGs), isolated from chimeric AD mice, resembles atherosclerotic foam cells. In addition, plaque-bound xMGs are highly enriched in lipid droplets, which is significantly diminished in TREM2-R47H xMGs. Notably, TREM2-R47H xMGs overall show reduced reactivity to plaques, including plaque-proximity, upregulation of CD9 and secretion of APOE. Conclusions: Altogether, these results indicate lipid droplet accumulation occurs in xMGs in AD and is triggered by a TREM2-dependent microglial response to plaques.

P073 / #1291


BETA-ADRENERGIC RECEPTOR ANTAGONISM IS PROINFLAMMATORY AND EXACERBATES NEUROINFLAMMATION IN A MOUSE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

A. Evans, H.H. Park, M. Shamloo Stanford University, Neurosurgery, Palo Alto, United States of America

Aims: These studies were designed to examine proinflammatory consequences of beta-blocker administration in an acute lipopolysaccharide (LPS) model as well as to examine chronic effects of beta-blockers on neuroinflammation and behavior in an amyloid-beta protein precursor (APP) mouse model of AD Methods: C57BL6/J mice were administered LPS with beta-adrenergic pharmacology and plasma and brain were collected. APP mice were administered beta blockers chronically, underwent behavioral testing, and tissue was collected. Gene and protein markers of inflammation were assessed in both models with qrtPCR, Western Blot, ELISA, and multiplex cytokine assays. Primary microglia cultures were treated with beta blockers and underwent live cell analysis of phagocytic function. Results: We show robust potentiation of peripheral inflammation with 4 different beta-blockers in the acute LPS model and potentiation of CNS inflammation in the chronic APP model which was accompanied with impairment in learning and memory. Notably, in the acute LPS model, prior genetic knockdown of either beta1- or beta2-adrenergic receptors in microglia did potentiate CNS inflammation. The beta-blocker, metoprolol also induced markers of phagocytosis and impaired cognitive behavior in both wildtype and APP mice. We examined phagocytosis of synaptosomes in an in vitro primary microglia culture and show that beta-blockers enhanced whereas beta-adrenergic agonists inhibited phagocytosis of synaptosomes. Conclusions: In conclusion, beta-blockers potentiated inflammation peripherally in a systemic model of inflammation and centrally in an amyloidosis model of neuroinflammation. Additionally, beta-blockers impaired learning and memory, and modulated synaptic phagocytosis with implications for synaptic degeneration.

P074 / #1303


UNDERSTANDING THE FUNCTION OF THIOREDOXIN-80 IN THE BRAIN AND ITS IMPLICATION IN ALZHEIMER´S DISEASE

Lecture Title:

J. Goicolea1, P. Rodriguez2, G. Gereñu-Lopetegui3, A. Cedazo-Minguez2, S. Maioli2 1Karolinska Institute, Department Of Neurobiology, Care Sciences And Society (nvs), Solna, Sweden, 2Karolinska Institutet, Neurobiology, Cara Sciences And Society, Stockholm, Sweden, 3Biodonostia, Neuroscience, Vasque Country, Spain

Aims: Thioredoxin-1 is a redox-active protein that can be cleaved into a smaller peptide, Thioredoxin-80 (Trx80). Previous studies in our group reported that Trx80 levels are depleted in Alzheimer´s Disease (AD) postmortem brains. We aim to define the biological function of Trx80 and the mechanisms that regulate endogenous Trx80 production and secretion in the brain. Also, we aim to explore its contribution to AD progression and its potential as an AD biomarker. Methods: This study is carried out in vitro primary brain cell cultures, in vivo mouse model and serum and Cerobrospinal Fluid (CSF) samples from AD patients. We have performed RNA-sequencing analysis on Trx80-treated primary microglia. Trx80 protein levels were analyzed in neuronal primary cultures treated with rotenone and in cortical samples of 3 and 10 months old APP knock-in mice by western-blot and immunohistochemistry. Results: RNA-sequencing analysis of Trx80-treated microglia revealed a significant upregulation of the interferon signaling pathway. In vitro, rotenone treatment on neuronal primary cultures showed a significant increase of Trx1 and Trx80 protein levels. Cortical samples from APPki mice showed that Trx80 is significantly elevated in 3 months old APPki mice in comparison to controls whereas in 10 months old mice, Trx80 levels are significantly decreased. Conclusions: This study points at Trx80 and its regulation in the brain, as a potential new neuronal mechanism to communicate with microglia under stress. The decrease in Trx80 levels that are observed in both, adult APPki mice and AD autopsy brains indicates that Trx80 could potentially be used as a novel AD biomarker.

P075 / #1449


AGE AND REGION SPECIFIC DIFFERENCES IN MICROGLIAL SYNAPTIC PRUNING IN A C5AR1 DEFICIENT MOUSE MODEL OF ALZHEIMER’S DISEASE.

Lecture Title:

A. Gomez Arboledas1, M. Fonseca1, A. Tenner1,2,3 1University of California Irvine, Molecular Biology And Biochemistry, Irvine, United States of America, 2University of California Irvine, Neurobiology And Behavior, Irvine, United States of America, 3University of California Irvine, Pathology And Laboratory Medicine, Irvine, United States of America

Aims: Objectives: The complement system (C’), which can be activated by fibrillar Aβ, can generate C5a that binds microglial C5aR1 and contributes to neuroinflammation. Many reports have shown an excessive synaptic loss in Alzheimer’s disease (AD) mouse models that is dependent on C’ at the synapses. Here, we examined whether C5a-C5aR1 signaling contributes to synaptic engulfment by microglia. Methods: Immunofluorescence with markers for microglia, lysosomes and synapses were assessed in brain sections of WT, C5aR1KO, Arctic (AD) and Arctic-C5aR1KO mouse models at different ages. 3D reconstruction analysis and quantification (of whole brain sections and single microglial cells) were carried out to test differences in microglial activation and phagocytosis. Results: The hippocampus of Arctic mice was characterized by an intense microglial response that increased with age. However, Arctic mice lacking C5aR1 showed lower Iba1 field area relative to the C5aR1 sufficient AD. This reduction was also accompanied by a slight reduction in the lysosomal marker CD68. 3D analysis of microglial cells showed less phagocytosis of synaptic marker VGlut1 in Arctic-C5aR1KO mice, which was dependent on the age and hippocampal region analyzed. In addition, microglia volume showed age and region dependent changes in AD mice lacking C5aR1. Conclusions: Our results suggest that in 10 month-old Arctic mice, C5aR1 influences synaptic pruning that occurs during Alzheimer’s disease. This contribution seems to vary across different ages and hippocampal regions. While the mechanisms underlying this remain to be delineated, modulation of C5a-C5aR1 signaling could be a potential therapeutic target to treat this neurodegenerative disease.

P076 / #400


MICROGLIAL NLRP3 INFLAMMASOME CONTRIBUTES TO AMYLOID PATHOLOGY: ANALYSIS OF THE PROCESSES INVOLVED

Lecture Title:

M.Á. Gutiérrez De Ravé1, P. Botella1, C. Lodder1, S. Vanherle1, N. Cremers2, I.-C. Stancu1, I. Dewachter1 1University of Hasselt, Biomedical Sciences Faculty, Neurosciences, Diepenbeek, Belgium, 2KU Leuven, Rega Institute, Laboratory Of Virology And Chemotherapy, Leuven, Belgium

Aims: Brains of AD patients are characterized by the presence of amyloid plaques and neurofibrillary tangles along with microgliosis. NLRP3 inflammasome, a key player in microglial activation, has been shown to be activated by Aβ contributing to amyloid pathology by different mechanisms. It is increasingly clear that microglia and their responses change during AD. Different NLRP3-dependent mechanisms can contribute or dominate at different stages of the disease progression. Here, we focus on a detailed analysis of NLRP3 inflammasome activation by Aβ, and its contributory effect on amyloid pathology by different microglia related processes in vitro and in vivo. Methods: To investigate this, primary microglia cultures were used to measure NLRP3 activation response to Aβ oligomers (o) and fibrils (f), reflected in IL-1β levels. We furthermore crossed 5xFAD mice with NLRP3ko (5xFAD.NLRP3ko) and compared them to 5xFAD.NLRP3wt at 6.5 months to correlate amyloid pathology and microgliosis by IHC analyses of different brain regions. Results: Aβo and Aβf activate the inflammasome in vitro when microglia are primed with LPS, in an NLRP3 dependent way. In vivo, amyloid pathology was significantly decreased in 5xFAD.NLRP3ko mice at 6,5 mo of age in prefrontal and frontal cortex while no significant effect was demonstrated in subiculum. Conclusions: Our work indicates that NRLP3 is activated by Aβ in vitro and in vivo. Ongoing analysis will further analyze the contribution of NLRP3-dependent microglial processes involved, including phagocytosis, ASC-speck formation, and cytokine/chemokine production in the different brain regions.

P077 / #1455


EFFECT OF TREM2 DELETION IN DEPOSITION OF PYROGLUTAMATE-3 AMYLOID-BETA IN TRANSGENIC MOUSE BRAIN

Lecture Title:

P. Joshi1, K. Satoh2, M. Enomoto3, S. Qamar4, S. Kumar1, C. Haass5,6, P. Fraser2, P. St. George-Hyslop2,4, J. Walter1 1Universitätsklinikum Bonn, Department Of Neurology, Bonn, Germany, 2Tanz Centre for Research in Neurodegenerative Diseases, Departments Of Medical Biophysics And Medicine (neurology), Toronto, Canada, 3Princess Margaret Cancer Centre Research Institute, University Health Network, Toronto, Canada, 4Cambridge Institute for Medical Research, University of Cambridge, Department Of Clinical Neurosciences, School Of Clinical Medicine, Cambridge, United Kingdom, 5German Center for Neurodegenerative Diseases, Dzne, Munich, Germany, 6Ludwig-Maximilians-Universität München, Metabolic Biochemistry, Munich, Germany

Aims: Triggering Receptor Expressed on Myeloid cells 2 (TREM2) with rare mutations in its ectodomain has been implicated in Alzheimer’s disease (AD). Recent studies have shown that Amyloid-β (Aβ) interacts with TREM2. However, the role of TREM2 in its interaction and in the deposition of various post-translationally modified Aβ species is unknown. In recent years, N-truncated pyroglutamate-modified Aβ, AβpE(3-x), has been studied extensively. Therefore, we wanted to investigate the effect of this modification on the interaction with TREM2, and the role of TREM2 in the deposition of AβpE(3-x) in the brains of transgenic mouse models. Methods: Characterization of aggregation states of Aβ variants was performed by negative stain Transmission electron microscopy (TEM) followed by Aβ binding studies using dot blot and Bio-layer interferometry (BLI) techniques. The characteristics and deposition of AβpE(3-x) species in age and sex matched APP/PS1 TREM2 knockout mice were analyzed with confocal microscopy using AβpE(3-x) specific antibody. Results: We observed that the TREM2 has a higher affinity to oligomeric than to monomeric forms of AβpE(3-42). Moreover, the binding of TREM2 to oligomeric AβpE(3-42) was similar to that of oligomeric non-modified Aβ(3-42). Furthermore, we show that TREM2 deletion leads to an increased number of plaques containing AβpE(3-x) species while not having a specific effect on its deposition as extracellular plaques. Conclusions: The N-terminal truncation and pyroglutamation of Aβ did not increase the interaction with TREM2. The AβpE(3-x) deposits in the extracellular plaques have not been altered in the TREM2 knockout transgenic mouse brain.

P078 / #1290


AMELIORATION OF ALZHEIMER’S PATHOLOGY WITH LONG TERM GLATIRAMER ACETATE TREATMENT IS CORRELATED WITH SUBTLE CHANGES IN MICROGLIAL TRANSCRIPTOME

Lecture Title:

B. Karaahmet, D. Dionisio-Santos, L. Owlett, J. Olschowka, M. O'Banion University of Rochester, Neuroscience, Rochester, United States of America

Aims: It is established that immune mechanisms or interventions may alter histopathological hallmarks of Alzheimer’s disease (AD). In this study, we investigated the therapeutic potential of a well-tolerated immunomodulatory Multiple Sclerosis drug, Glatiramer Acetate (GA), on the 3xTg model. We further aimed to determine the impact of GA on the microglial transcriptome. Lastly, we examined whether changes in Th cell phenotypes correlated with our observations. Methods: Female 3xTg mice (>15 month-old) were treated weekly with 0.1 mg of GA for approximately 8 weeks. Hallmarks of AD pathology were quantified through immunohistochemistry, ELISA and Westerns. RNAseq on FACS isolated microglia from a separate cohort was performed. Immunophenotypes of isolated peripheral CD4+ Th cells from this cohort were analyzed by qRT-PCR for key transcriptional markers. Results: We were able to show that GA treatment decreased amyloid plaque load and phosphorylated-Tau. This was associated with a subtle transcriptomic change in hippocampal microglia. In particular, only DCSTAMP was significantly upregulated. From single-cell RNAseq datasets, we noticed that DCSTAMP expression is commonly observed in Disease Associated Microglia (DAM). Furthermore, gene set based analyses revealed that GA treatment is associated with response to anti-inflammatory interleukin-4. We did not observe significant expression changes in transcriptional markers of Th phenotypes. Conclusions: Here, we show that GA treatment may be beneficial in ameliorating hallmarks of AD and its mechanism may partly be associated with microglia. Specifically, we hypothesize that GA may be inducing the DAM phenotype. Interestingly, we could not detect the Th2-shift with GA treatment in our paradigm. Further investigations are underway.

P079 / #1829


IMBALANCE BETWEEN GLYCOLYSIS AND OXIDATIVE PHOSPHORYLATION IN MICROGLIA CONTRIBUTES CRITICALLY TO ALZHEIMER’S DISEASE PATHOLOGY

Lecture Title:

Y. Liu1, H. Yoon2, D.-W. Li3, G. Dos Santos3, J. Ryu1, D. Scharre4, L. Bruschweiler-Li3, R. Bruschweiler3, K. Obrietan2, S. Yoon1 1The Ohio State University, Department Of Biological Chemistry And Pharmacology, Columbus, United States of America, 2The Ohio State University, Department Of Neuroscience, Columbus, United States of America, 3The Ohio State University, Department Of Chemistry And Biochemistry, Columbus, United States of America, 4The Ohio State University, Department Of Neurology, Columbus, United States of America

Aims: To investigate the mechanisms leading to the metabolic disruption in AD and how they contribute to the disease progression. Methods: Analyses of brain samples from AD patients, control individuals and 5XFAD mice. Results: Our metabolomic study demonstrated that serum lactate levels decreased in 6-month-old 5XFAD mice and human AD cases. From subsequent analyses, we discovered that the levels of active tetrameric PKM2 were significantly increased in the brains of AD patients and 6-month-old 5XFAD mice. Of particular interests, PKM2 was primarily expressed among activated microglia associated with Aβ plaques in 5XFAD mice. We hypothesized that aberrant activation of PKM2 in microglia in AD induces glycolytic imbalance, thereby obstructing microglial phagocytic function. Indeed, pharmacological activation of PKM2 in vivo with its agonist, TEPP46, significantly attenuated recruitment of Cx3Cr1-GFP+ microglia toward the lesion epicenter upon 2-photon imaging. These results correlated with a significant drop in glycolysis with coincident reduction in lactate production when TEPP46 was added to primary microglia. Together these results suggest that activation of PKM2 favors oxidative phosphorylation over lactate production, thereby restricting microglial damage responses. Indeed, when oxidative phosphorylation was inhibited by ~50% in primary microglia with IACS010759, a complex I inhibitor of the mitochondrial respiratory chain, phagocytosis of Aβ42 aggregates was increased by greater than 2-fold. In further support, a selective deletion of PKM2 among microglia in 5XFAD mice resulted in a significant reduction in microglial activation and Aβ plaques. Conclusions: These results highlight the importance of homeostatic balance between glycolysis and oxidative phosphorylation for proper microglial function in AD.

P080 / #664


THE R522 ALZHEIMER'S PROTECTIVE PLCG2 HYPERMORPH DEPLETES SUBSTRATE AND ALTERS CELL FUNCTION

Lecture Title:

E. Maguire1, G. Menzies2, T. Phillips1, M. Sasner3, H. Williams3, M. Czubala4, N. Evans1, E. Cope1, R. Sims1, G. Howell2, E. Lloyd-Evans2, J. Williams1, N. Allen1, P. Taylor1 1Dementia Research Institute, Cardiff University, Cardiff, United Kingdom, 2Cardiff University, School Of Biosciences, Cardiff, United Kingdom, 3The Jackson Laboratory, Alzheimer's Disease Center, Maine, United States of America, 4Cardiff University, School Of Medicine, Cardiff, United Kingdom

Aims: Evidence from large scale genetic studies suggests the R522 variant in the enzyme PLCγ2 protects against Alzheimer’s disease (AD). Within cells, PLCγ2 breaks down the phospholipid PI(4,5)P2, releasing IP3 and DAG and activating various signalling pathways. PLCγ2 lies downstream of TREM2, expression of which is known to impact AD development. Within the brain, PLCγ2 is predominantly expressed in microglia; a cell type increasingly recognized as important to AD pathology. We aimed to characterize how the R522 mutation affects both PLCγ2 enzyme activity and subsequent microglial function. Methods: We generated novel human and mouse models to study the effects of the R522 mutation, beginning by performing Ca2+ and PI(4,5)P2 measurements in the presence of a PLCγ2 activator to characterize PLCγ2 enzyme activity in microglia and macrophages. Phagocytosis, cytokine secretion, and TREM2 function were examined to characterize phenotypic changes within the protective variant. Results: The R522 variant generated a hyper-functional PLCγ2-mediated Ca2+ response compared to controls, with this hyper-function appearing to result in a reduced PI(4,5)P2 pool. In addition, microglia and macrophages showed comparable defects in phagocytosis, reduced LPS-mediated cytokine production, and increased TREM2 signalling. Conclusions: Consistent findings in physiologically relevant human and mouse models suggest that increased enzyme activity within R522 microglia and macrophages leads to reduced PLCγ2 substrate. This reduction appears to impact function in various ways, including changes in phagocytic engulfment, cytokine production, and TREM2 signalling. These observations suggest reduced activation of microglia in response to stimuli, and could go part way to explaining the observed protective effects of R522 in patients.

P081 / #631


APOE SHAPES THE RESPONSE OF HUMAN MICROGLIA TO ALZHEIMER'S DISEASE PATHOLOGY IN VIVO

Lecture Title:

R. Mancuso1,2, A. Martinez-Muriana1, N. Fattorelli1, L. Wolfs1, P. Preman1, I. Geric1, S. Poovathingal1, M. Fiers1,3, B. De Strooper1,3 1VIB-KU Leuven, Vib Center For Brain & Disease Research, Leuven, Belgium, 2VIB-UAntwerpen, Department Of Biomedical Sciences, Antwerpen, Belgium, 3UK Dementia Research Institute at UCL, University College London, London, United Kingdom

Aims: Genetics place microglia in the center of the pathogenic cellular network in Alzheimer’s disease (AD). ApoE, the main risk factor for AD, is expressed by microglia and has been shown to be essential for their phenotypical changes in response to disease. However, the ApoE polymorphisms crucial to understand its role in disease are absent in mouse models, and us and others have shown important divergence in the response of human microglia to AD in comparison to lower mammals. Therefore, it is crucial to investigate the role of ApoE in a humanized system. Methods: We assessed how ApoE shapes the response of human microglia in AD in vivo, by transplanting isogenic series of iPSC-derived microglia carrying different APOE2/2, APOE3/3 and APOE4/4 allelic variants, and APOE-KO into the brain of APP-NLGF mice. We profiled microglia transcriptome and analyze their association to plaques by morphological studies. Results: We found a genotype dependent effect in the response of human microglia to amyloid beta plaques, with significant changes in their phenotypic transition from homeostatic to activation states. Morphological analysis also revealed differences in the recruitment of human microglia to amyloid beta plaques depending on the expression of ApoE variants, pointing to important cell autonomous effects of ApoE in microglia function. Conclusions: ApoE is able to shape the response of human microglia to amyloid beta in AD. This confirms the relevant place that microglia have in the pathogenic events leading to disease and situates neuroinflammation as an attractive target for future therapeutic interventions.

P082 / #718


ROLE OF MICROGLIA IN THE RESOLUTION OF INFLAMMATION IN ALZHEIMER’S DISEASE

Lecture Title:

A. Martinez-Muriana1, L. Wolfs1, N. Fattorelli1, N. Thrupp1, M. Fiers1, J. Dalli2, R. Mancuso1, B. De Strooper1 1VIB-KU Leuven, Vib Center For Brain & Disease Research, Leuven, Belgium, 2Queen Mary University of London, William Harvey Research Institute, London, United Kingdom

Aims: Chronic neuroinflammation is a pathological hallmark of Alzheimer’s disease (AD). Lipidome profiling of hippocampal and entorhinal cortex samples from AD patients showed reduced levels of specialized pro-resolving lipid mediators (SPMs), pointing to impaired resolution as a potential player in the pathology. Genome wide association studies (GWAS) also linked microglia and lipid metabolism as main players in AD, suggesting that lipids dysregulation may influence microglial inflammatory responses. Hence, we hypothesize that AD pathology may impair SPMs’ synthesis in microglia resulting in chronic inflammation. Here, the main goal is to determine if human microglia is involved in the synthesis of SPMs and characterize the inflammatory response upon an AD insult. Methods: To stablish the role of human microglia in the resolution of inflammation, we analyzed the transcriptomic and lipidomic profiles in primary and stem cell-derived human microglia and assessed the expression inflammatory mediators and synthesis enzymes. To determine the impact of AD pathology during the resolution of inflammation, we stimulated human microglia with oligomeric Aβ (oAβ) and LPS in vitro and assessed cytokines expression by qPCR. Results: Human microglia express all the necessary enzymes and receptors to resolve inflammation. Besides, lipidome profiling revealed that human microglia present steady-stage levels of pro-inflammatory and pro-resolving lipid mediators. Upon stimulation in vitro, oAβ triggered a higher pro-inflammatory response compared to LPS. Conclusions: Our preliminary data reveals that human microglia are capable of synthetizing SPMs and that oAβ induced an enhanced and longer pro-inflammatory response compared to LPS, suggesting that resolution pathways might be affected in AD.

P083 / #509


UNDERSTANDING MICROGLIAL RESPONSES IN THE FRONTAL CORTEX OF ALZHEIMER´S DISEASE PATIENTS

Lecture Title:

M. Mejias Ortega1, E. Sanchez-Mejias1, V. Navarro2, C. Nuñez-Diaz1, R. Sanchez-Varo1, S. Jimenez2, J.C. Davila1, J. Vitorica2, A. Gutierrez1 1University of Malaga/CIBERNED/IBIMA, Cell Biology Department, Faculty Of Sciences, Malaga, Spain, 2University of Seville/CIBERNED/IBIS, Biochemistry And Molecular Biology, Seville, Spain

Aims: Microglial cells, the immune cells of the brain, and the neuroinflammatory process associated, have been postulated as a critical factor in AD pathogenesis, since the identification of genetic risk factors related to microglial function. However, the microglial role in the development/progression of AD has not been determined yet. In this sense, we have previously reported a limited activation and microglial degeneration in the hippocampus of AD patients in contrast to the proinflammatory view based on findings in amyloidogenic models. Here, we have further analyzed the functional/phenotypic profile displayed by microglial cells in other vulnerable brain region of AD patients, the frontal cortex. Methods: Immunohistochemistry and image analysis approaches were performed in the frontal cortex of post mortem samples from controls (Braak 0-II) and AD patients (Braak V-VI) including familial cases. Results: Microglia of Braak V-VI individuals were observed forming clusters and showed, both plaque (Iba1+/TMEM119+/P2ry12-/CD45high/Trem2+) and inter-plaque (Iba1+/ TMEM119+/P2ry12-/CD45high/Trem2-

) microglial activation, similar that observed in amyloidogenic mice. By contrast, homeostatic and ramified microglial cells of non-demented Braak II cases presented Iba1+/P2ry12+/TMEM119+/CD45low/Trem2-

profile. Furthermore, different microglial responses were observed between sporadic and familial AD cases. Conclusions: These different microglial phenotypes associated with AD pathology show the heterogeneity and complexity of the microglial phenotypes and suggest different functional states of these glial cells in a region-specific manner. These data need to be considered for better understand the immunological mechanisms underlying AD progression. Modulating brain inflammatory responses might be a promising avenue to prevent cognitive dysfunction in AD patients. ISCiii:PI18/01557(AG)-PI18/01556(JV);Junta Andalucia:UMA18-FEDERJA211(AG). All cofinanced by FEDER funds (European-Union).

P085 / #1417


MODELING THE IMPACT OF AD GENETIC RISK ON MICROGLIA STATES AND FUNCTION

Lecture Title:

M. Therrien1, M. Dolan1, S. Jereb1, N. Lojek1, T. Kamath2, S. Marsh3, M. Johnson1, E. Macosko2, K. Eggan4, B. Stevens3,5,6 1Broad Institute of MIT and Harvard, Stanley Center For Psychiatric Research, Cambridge, United States of America, 2Broad Institute of MIT and Harvard, Stanley Center For Psychiatric Research, Cambridge, United States of America, 3Boston Children's hospital, F.m.kirby Neurobiology Center, Boston, United States of America, 4Harvard University, Department Of Stem Cell And Regenerative Biology, Cambridge, United States of America, 5Broad Institute of Harvard and MIT, Stanley Center For Psychiatric Research, Cambridge, United States of America, 6Howard Hughes Medical Institute, Boston Children's Hospital, Boston, United States of America

Aims: Microglia, the brain’s resident immune cells, are highly dynamic and reactive to environment and genetic challenges. More than 40 genomic loci have been linked to late onset Alzheimer’s disease (AD) and many risk genes are highly expressed in microglia. Our goal is to connect insights from genetic association studies to new ways of functionally modeling the cellular and molecular causes of disease to enable predictive tracking and targeting of detrimental immune cell states in patients in the early stages of disease. Methods: Single cell transcriptomic studies reveal diverse microglial states in human and mouse brains, however we currently lack specific approaches to track and manipulate specific populations of microglia in Alzheimer’s and other diseases. To answer this question, we turned toward human iPSC-induced microglia (iMGL) and single cell transcriptomics. Results: Single cell transcriptomics revealed the presence of multiple microglia states (up to 14) when iMGLs are stimulated with different brain-relevant challenges, including apoptotic neurons, synaptosomes, myelin and amyloid. Moreover, alignment of these data using Liger (Welch et al. Cell 2017) shows these states are similar to the ones observed in human and mouse in vivo, revealing several disease associated states, including disease associated microglia (DAM). We also observed changes in microglia states depending on the challenge and genetic background. Conclusions: Together, our data identified key elements causing the formation of DAM and how AD risk genes affect them. This work will open the door to the identification of modulators of DAM and highlight new therapeutics avenues of AD

P086 / #576

Topic: Theme A: β-Amyloid Diseases / A1.j. Disease Mechanisms, Pathophysiology: Astroglia

SUITABILITY OF HUMAN-DERIVED CELLS AS A PLATFORM FOR ALZHEIMER´S DISEASE MODELING

Lecture Title:

L. Cáceres Palomo1, J.A. García León2, J.C. Davila3, J. Vitorica4, A. Gutierrez5 1University of Malaga/CIBERNED/IBIMA, Cell Biology, Genetics And Physiology. *first Authors., Málaga, Spain, 2University of Malaga/CIBERNED/IBIMA, Cell Biology, Genetics And Physiology. *first Authors. #corresponding Authors., Málaga, Spain, 3University of Malaga/CIBERNED/IBIMA, Cell Biology, Genetics And Physiology., Málaga, Spain, 4University of Seville/CIBERNED/IBIS, Biochemistry And Molecular Biology, Seville, Spain, 5University of Malaga/CIBERNED/IBIMA, Cell Biology, Genetics And Physiology. #corresponding Authors., MALAGA, Spain

Aims: Alzheimer's disease (AD) is characterized by presenting a complex pathology, not fully resolved yet. This fact, together with the lack of reliable models, has impeded the development of effective therapies. Recently, several studies have shown that functional glial cell defects have a key role in the pathology of AD. However, this glial dysfunction, currently, cannot be correctly modeled using the available animal models, so we hypothesized that cells derived from Alzheimer's patients can serve as a better platform for studying the disease. In this sense, human pluripotent stem cells (hPSC) allow the generation of different types of neural cells, which can be used for disease modeling, identification of new targets and drugs development. Methods: We have a collection of hiPSCs derived from patients with sporadic forms of AD. We have differentiated these cells towards neural lineage to obtain neurons and astrocytes. For the generation of oligodendrocytes (OLs), we have developed a fast and robust protocol to generate mature OLs in just 22 days. Results: We have generated neural precursors from all the lines tested. In the case of OLs, the cells generated resemble primary OLs and can myelinate neurons in vivo and in vitro using a screening compatible platform. This platform is being transferred for the generation of the other glial cells. Conclusions: This methodology can be used to elucidate the pathogenic pathways associated with neurodegeneration and to identify new therapeutic targets susceptible to modulation, contributing to the development of new effective drugs against AD.

P087 / #1179


AN ASTROCYTIC MECHANISM OF CHOLESTEROL-INDUCED HYPEREXCITABILITY IN THE BRAIN

Lecture Title:

R. Loera-Valencia1, G. Gereñu-Lopetegui2, P. Merino-Serrais1, M. Perez Peiró1, K. Gomez1, L. Alvarez1, F. Spanos1, M. Latorre Leal3, J. Goicolea4, M. Gomez5, H. Balleza-Tapia4, A. Fisahn4, S. Maioli6, A. Cedazo-Minguez6 1Karolinska Institute, Nvs, Solna, Sweden, 2Biodonostia, Neuroscience, Vasque Country, Spain, 3Karolinska Institutet, Nvs, Division Of Neurogeriatrics, Solna, Sweden, 4Karolinska Institute, Department Of Neurobiology, Care Sciences And Society (nvs), Solna, Sweden, 5Karolinska Institute, Physiology And Pharmacology, Solna, Sweden, 6Karolinska Institutet, Neurobiology, Cara Sciences And Society, Stockholm, Sweden

Aims: To unravel the role of 27-hydroxycholesterol in the function of astrocytes and their impact on neuronal synaptogenesis in vitro and in vivo. Hypothesis: Elevated 27-hydroxycholesterol levels in the brain will cause activation in astrocytes and thus, decrease astrocyte synaptic function. Methods: We studied a transgenic in vivo model and mixed neuron-astrocyte cultures in 3D for in vitro studies described below. CYP27 Transgenic Mice (Cyp27Tg): CYP27-overexpression transgenic mice were generated using a human full-length CYP27 coding region controlled by a constitutive promoter. These mice have 3-5 higher levels of circulating 27-hydroxycholesterol during their whole life. To test the hypothesis that 27-OH could be implicated in astrogenesis in vitro, we will study their effect on neuronal development in cultured murine astroglia and neurons in close three-dimensional (3D) interaction. Results: When we evaluated GLT-1 and GLAST in protein extracts from CYP27Tg mice brains and we encounter a decrease in protein levels compared to WT controls. Additionally, we encountered increased GFAP signaling that suggests astrocyte activation. Together these results point towards decreased astrocyte function since both GLT-1 and GLAST are important in maintaining neuronal excitability through recapturing glutamate from excitatory synapses. When we analyzed astrocytes in 3D cultures, we encountered diminished glutamate transporters in samples treated with 27-hydroxycholesterol. The reduced levels of GLT-1 and GLAST of astrocytes would mean a limited capability to recapture glutamate in synapses, thus affecting neurotransmission. Conclusions: We show for the first time that 27-hydroxycholesterol has the capability of activating astrocytes and decrease their glutamate uptake function, affecting synaptic transmission and inducing hyperexcitability on neurons.

P088 / #1273


A SYSTEMATIC REVIEW OF HUMAN POSTMORTEM IMMUNOHISTOCHEMICAL STUDIES UNVEILS THE COMPLEXITY OF ASTROCYTE REACTION IN ALZHEIMER'S DISEASE

Lecture Title:

A. Noori1,2, L. Viejo De Navas2,3, B. Hyman2,4, S. Das2,4, A. Serrano-Pozo2,4 1Harvard College, Molecular And Cellular Biology, Cambridge, United States of America, 2Massachusetts General Hospital, Neurology, Boston, United States of America, 3Universidad Autónoma de Madrid, Farmacología Y Terapéutica, Madrid, Spain, 4Harvard Medical School, Neurology, Boston, United States of America

Aims: Astrocytes undergo morphological and functional changes in CNS diseases, collectively termed "astrocyte reaction" or "reactive astrogliosis". While reactive astrocytes have traditionally been characterized by increased immunoreactivity for the cytoskeletal intermediate filament glial fibrillary acidic protein (GFAP), transcriptomics investigations such as single nuclei RNA-sequencing have begun to decipher their complexity and heterogeneity in various CNS disorders. We hypothesized that a systematic review of previously published human postmortem immunohistochemical studies coupled with bioinformatics analyses would unravel this complexity in Alzheimer's disease (AD). Methods: We conducted a systematic review following PRISMA guidelines (search strategy "Alzheimer's disease" AND "Astrocytes" in PubMed, WoS-SCI, and APA PsycInfo; prespecified eligibility criteria for article selection). We applied bioinformatics tools on the resulting protein set [e.g. pathway enrichment analysis (PEA), transcription factor enrichment analysis (TFEA), protein-protein interaction (PPI) functional network], followed by hypergeometric enrichment tests against published human brain/CSF proteomic and astrocyte transcriptomic datasets. Results: Total 361 included articles rendered 196 proteins. PEA implicated cytokine signaling, extracellular matrix, lipoprotein metabolism, trophic factors, response to ROS, and protein degradation, among other functional changes. TFEA suggested CTCF as a novel driver of these changes. PPI functional network analysis yielded a highly connected network with interleukin-6, tumor necrosis factor-α, and MAP-kinase-1, -3 and -8 as top hub proteins. Cross-validation with published human multiomics datasets demonstrated statistically significant enrichment (p<1E-11). Conclusions: Our systematic review provides clues about the complexity of astrocyte reaction in AD, revealing altered immune response, extracellular matrix, lipid metabolism, trophic factors, oxidative stress, and proteostasis. Our findings could inform ongoing biomarker research efforts.

P089 / #703


ASSOCIATION OF AΒ PLAQUES AND CONNEXINS IN THE SPINAL CORD OF 5XFAD ALZHEIMER’S MOUSE MODEL

Lecture Title:

M. Pechlivanidou1, I. Kousiappa1, S. Angeli1, K. Kleopa2, S. Papacostas3 1Cyprus Institute of Neurology and Genetics, School of Molecular Medicine, Neurobiology Department, Nicosia, Cyprus, 2The Cyprus Institute of Neurology and Genetics, School of Molecular Medicine, Neuroscience Department, Center For Neuromuscular Disorders, Center For Multiple Sclerosis And Related Disorders, Nicosia, Cyprus, 3Medical School University of Nicosia,The Cyprus Institute of Neurology and Genetics, School of Molecular Medicine, Neurobiology Department, Cognitive Disorders Center, Nicosia, Cyprus

Aims: The aim is to explore the amyloid pathology along with alteration in glia in the spinal cord (SC) of the 5xFAD mouse model. Methods: Motor performance tests were performed in 5XFAD and non-transgenic mice groups (3 and 12 months). Fluorescence immunohistochemistry (IHC) in 3rd-4th Cervical and 2nd-3rd Lumbar frozen spinal sections was carried out for 6E10, GFAP, Iba1, CC1, Olig2 and NeuN markers. Results: Twelve-month-old 5XFAD mice showed a significant impairment in all motor performance tests. Also, they showed abundant extracellular Aβ plaque deposition in spinal cord, which appeared at 3 months and exhibited a time dependent increase. Older 5xFAD mice showed increased immunoreactivity for GFAP+ astrocytes and Iba1+ microglia compared to 3-month-old mice. Lastly, 12M 5XFAD mice exhibited a significant neuronal loss and oligodendrocytic loss in the white matter of the abovementioned spinal segments. Conclusions: We detected increased astrogliosis and oligodendrocytic loss in the SC which may contribute to Alzheimer’s neurodegeneration. Future experiments will be conducted to invistigate Cxs contribution to the spinal axonal damage and to the motor deficiencies in 5xFAD and 3xTG AD mice.

P090 / #1805


GABA METABOLISM IS DECREASED IN HUMAN INDUCED PLURIPOTENT STEM CELLS DERIVED ASTROCYTES WITH MUTATION IN THE APP OR PSEN-1 GENE

Lecture Title:

C. Salcedo1, A. Wagner1, H. Waagepetersen1, K. Freude2, B. Aldana1 1University of Copenhagen, Department Of Drug Design And Pharmacology, Copenhagen, Denmark, 2University of Copenhagen, Department Of Veterinary Clinical And Animal Sciences, Faculty Of Health And Medical Sciences, Copenhagen, Denmark

Aims: Alzheimer´s Disease (AD) is a neurodegenerative disorder which impairs cognition predominantly manifested as memory loss. AD principal hallmarks comprise extracellular accumulation of amyloid β plaques and intracellular neurofibrillary tangles. Likewise, metabolic and mitochondrial dysfunction has been strongly implicated in AD pathophysiology, resulting in neurotransmitter disturbances. Neurotransmitter recycling is mainly mediated by glia, and we recently showed that astrocytes, the main glial cell type, take up and metabolize gamma-aminobutyric acid (GABA), released from the neurons. Subsequently, the astrocytes produce and release glutamine to support neuronal transmission. The aim of the study was to assess GABA metabolism in human induced pluripotent stem cells (hIPSC) derived astrocytes with mutation in the APP or PSEN-1 gene as models of AD. Methods: hIPSC-derived astrocytes with or without APP or PSEN-1 mutation were incubated with uniformly 13C labeled GABA ([U-13C]GABA) (50μM or 200μM) for 90min at 37°C. 13C incorporation in selected metabolites was determined by mass spectrometry and amino acids amounts were quantified using HPLC. Results: Reduced 13C incorporation in TCA cycle intermediates was observed after incubation with [U-13C]GABA at both concentrations in APP and PSEN-1 mutated astrocytes compared to control astrocytes. Intracellular labeled amounts of GABA and glutamate, were reduced in AD mutated astrocytes. Interestingly, the labeled amounts of glutamine were increased after incubation with 200μM [U-13C]GABA in AD mutated astrocytes. Conclusions: These results revealed an altered GABA metabolism in hIPSC derived AD astrocytes, further contributing to the synaptic transmission disturbances present in AD.

P091 / #1247


COMPARED OF IGF-1 INDUCED GDNF AND BDNF FROM C6 -GLIOBLASTOMA CELLS IN BETWEEN DEPOSITION OF ROTENONE AND OF BETA-AMYLOID NEUROTOXICITY

Lecture Title:

T. Watanabe Teikyo University of Science, Neuropathology, TOKYO, AGACHI-city, Japan

Aims: Alzheimer’s disease (AD), one of most common neurodegenerative disorders is elderly and oxidative stress, is characterized deposition massive loss of neuronal cells with Amyloid-beta in corticocerebral region. In the previous study, we analyzed secretion of GDNF and BDNF in effect of IGF-1 for C6 glioma cells with/without rotenone- and MPTP+-induced Neurotoxicity of parkinson disease (survival of PD). Methods: 1.The all of these experiments were used C6 glioblastoma cell line. 2.Cell count measurement: Under plated at low density (2x104cells/35mm plate), cells were cultured 1day, 2day or 3day with IGF-1 /normal serum. And then, counted means of triplicate determination. 3.Cell-Viability measurement: MTT –assay for absorbance at 540nm 4.RT-PCR for mRNA: All primer were denatured by 30 cycler at 94℃,

and by annealing at 55℃ and by elongation at 72℃ Results: In present study, to determine factors that would enhance GDNF and BDNF expression, we analyzed the effect of IGF-1 in C6 glioma cells. Treatment of C6 cells with IGF-1 elicited increase in the level of GDNF and BDNF content. In addition, our results indicate that IGF-1 is 14.5% of effective at recovery of beta-Amyloid induced C6 cell. Conclusions: These data indicated that IGF-1 was a potent inducer of GDNF and of BDNF expression and suggested that IGF-1 might contribute to the regulation of GDNF and BDNF in vivo survival situation with Gliosis under not only PD but also AD Neurotoxicity.

P092 / #1831

Topic: Theme A: β-Amyloid Diseases / A1.k. Disease Mechanisms, Pathophysiology: Neurogenesis

ADULT HUMAN HIPPOCAMPAL NEUROGENESIS IN AGING AND ALZHEIMER’S DISEASE.

Lecture Title:

A. Disouky1, M. Tobin1, K. Musaraca1, A. Shetti1, A. Bheri1, W. Honer2, N. Kim3, R. Dawe3, D. Bennett3, K. Arfanakis3, O. Lazarov1 1University of Illinois at Chicago, Anatomy And Cell Biology, Chicago, United States of America, 2University of British Colombia, Psychiatry, British colombia, Canada, 3Rush University, Rush Alzheimer’s Disease Center, Rush University Medical Center, Chicago, United States of America

Aims: The existence and extent of neurogenesis in the adult human hippocampus has been a topic of debate in recent years. Methods: Immunohistochemistery Stereological counting Results: Here, we provide evidence for the persistence of neurogenesis not only in aging individuals with no cognitive impairments, but also in the brains of patients with mild cognitive impairments (MCI) and Alzheimer’s disease (AD). We show that neural progenitor cells, neuroblasts and immature neurons are distributed throughout the ventral-dorsal axis of the hippocampus. The number of neuroblasts and immature neurons is significantly reduced in MCI. In addition, higher number of neuroblasts (DCX+PCNA+) is associated with better cognitive diagnosis. Furthermore, we show that the number of newly generating neurons correlates with the expression of key presynaptic proteins, previously shown to be associated with cognitive reserve. Conclusions: These findings imply that a higher number of neuroblasts is less likely to be associated with cognitive decline. In addition, these observations suggest that neurogenesis is compromised in an early stage of cognitive deterioration. Taken together, these results suggest that hippocampal neurogenesis persists in the brain throughout life and that enhancing neurogenesis may support cognitive function in AD.

P093 / #691

Topic: Theme A: β-Amyloid Diseases / A1.k. Disease Mechanisms, Pathophysiology: Neurogenesis

TRACE AMINE-ASSOCIATED RECEPTORS (TAARS) AND ADULT NEUROGENESIS

Lecture Title:

R. Gainetdinov SPBU, Institute Of Translational Biomedicine, St. Petersburg, Russian Federation

Aims: Trace amine-associated receptors (TAARs) are a class of sensory G protein-coupled receptors that detect amines, products of decarboxylation of amino acids. The majority of TAARs (TAAR2-TAAR9) are found in the olfactory epithelium and are believed to serve as a new class of olfactory receptors sensing innate odors. However, there is evidence that one of the members of this family TAAR5 is expressed also in the limbic "emotional" areas of the brain receiving projection from the olfactory system. Methods: We investigated a mouse line lacking TAAR5 (TAAR5-KO mice) that express beta-galactosidase mapping TAAR5 expression. Results: In TAAR5-KO mice, the number of dopamine neurons, the striatal levels of dopamine and its metabolites, as well as striatal levels of GDNF mRNA, are elevated indicating a potential increase in dopamine neuron proliferation. Analysis of TAAR5 beta-galactosidase expression reveled that TAAR5 is present in the major neurogenic areas of the brain such as the subventricular zone (SVZ), the subgranular zone (SGZ), and in the less characterized potentially neurogenic zone surrounding the 3rd ventricle. Analysis of neurogenesis by using specific markers Doublecortin (DCX) and Proliferating Cell Nuclear Antigen (PCNA) revealed at least a 2-fold increase in the number of proliferating neurons in the SVZ and SGZ of TAAR5-KO mice. Conclusions: TAAR5 is involved in adult neurogenesis and TAAR5 antagonists should be explored as potential treatments of neurodegenerative disorders such as Parkinson’s disease. The “olfactory” TAAR-mediated sensory function of detecting decarboxylated amino acids inside the brain may represent a novel mechanism for regulating neurogenesis in response to the various insults to the brain.

P094 / #854

Topic: Theme A: β-Amyloid Diseases / A1.l. Disease Mechanisms, Pathophysiology: Vasculature, microbleeds, hypertension, angiogenesis

EFFECT OF PERIPHERAL ADMINISTRATION OF RECOMBINANT HUMAN APOJ ON THE OCCURRENCE OF CEREBRAL MICROBLEEDS IN APP23 MICE

Lecture Title:

A. Bonaterra-Pastra, M. Solé, P. Marazuela, S. Fernández De Retana, M. Hernández-Guillamon Vall d'Hebron Research Institute, Neurovascular Diseases, Barcelona, Spain

Aims: Apolipoprotein J (ApoJ) is a multifunctional chaperone related to β-amyloid (Aβ) aggregation and clearance. Our prior study (Fernández-de-Retana, 2019) demonstrated that treatment with recombinant human ApoJ (rhApoJ) in 14-month-old APP23 mice prevented the Aβ accumulation in cerebral arteries and decreased insoluble Aβ brain levels. The present study aims to determine the effects of rhApoJ treatment in older APP23 mice in terms of potential amelioration of vascular damage and reduction of cerebral microbleeds (CMB) associated with cerebral amyloid angiopathy (CAA). Methods: Twenty-month old APP23 mice received 25 intraperitoneal doses of rhApoJ (1mg/kg) (n=9) or saline (n=8) during 13 weeks. Wild-type (WT) animals were treated with saline (n=13) as controls. After treatments, postmortem T2*-weighted magnetic resonance imaging (MRI) was performed immersing brains in Galden®D05 PFPE and using a horizontal magnetic system (7T, BioSpec 70/30USR, Bruker). Hypointense signals on T2* were counted as hemorrhages and classified as CMB (50-300 µm diameter) or larger hemorrhages (> 300 µm). Results: The chronic treatment with rhApoJ was safe in very old mice. APP23 brains presented a higher number of cerebral hemorrhages than WT mice (p<0.001). However, rhApoJ treatment reduced the total number of cerebral hemorrhagic lesions compared to saline-treated mice (p=0.009). In fact, brains from rhApoJ-treated mice presented a lower number of cortical CMB (p=0.012) and fewer larger hemorrhages in the cortex (p=0.002) than saline-treated mice. These differences were not obtained when deep areas were analyzed. Conclusions: Our results suggest that chronic peripheral treatment with rhApoJ in very old APP23 mice ameliorates the neurovascular damage associated with CAA pathology.

P095 / #807


ELEVATED BLOOD PRESSURE AND DECREASED VASCULAR CONTRACTILITY UPON CARDIOVASCULAR PHENOTYPING OF THE APP23+/- OVEREXPRESSING AD MOUSE MODEL

Lecture Title:

J.O. Hendrickx1, S. De Moudt1, D. Van Dam2,3, G. De Meyer1, P. Fransen1 1Laboratory of Physiopharmacology, Faculty Of Pharmaceutical, Biomedical And Veterinary Sciences, University Of Antwerp, Wilrijk, Belgium, 2Laboratory of Neurochemistry and Behaviour, Institute Born-Bunge, Department Of Biomedical Sciences, University Of Antwerp, Wilrijk, Belgium, 3University of Groningen and University Medical Center Groningen, Department Of Neurology And Alzheimer Research Center, Groningen, Netherlands

Aims: Increasing epidemiological and experimental evidence confirms the occurrence of cardiovascular (CV) disease in the progression of AD. Therefore, we aimed for a cardiovascular phenotyping of a mutated hAPP overexpressing mouse model (APP23+/-) of AD. Methods: APP23+/- mice (male, n=10) were compared to C57BL/6 (male, n=24) at the age of 6 months including in vivo analysis of blood pressure (BP, CODA), aortic pulse wave velocity (aPWV, VEVO2100), and echocardiography (high-frequency echocardiography, VEVO2100). Serum corticosterone levels and vascular smooth muscle cell (VSMC) phenotypic markers were ascertained via ELISA and qPCR, respectively. Vascular reactivity studies were performed ex vivo in isometric organ chambers. Data are presented as mean ± SEM. Results: Increased peripheral systolic BP (systolic BP: 110 ± 5 vs. 99 ± 3 mmHg) and aPWV was obtained in APP23+/- mice (aPWV: 4.08 ± 0.3 vs. 2.94 ± 0.1 m/s, p=0.0004) in vivo. No differences were observed in systolic or diastolic heart function upon echocardiographic analysis. Vascular reactivity studies revealed decreased (32%) adrenoreceptor-dependent contractions of the thoracic aorta from APP23+/- mice. Endothelial-dependent (acetylcholine) and -independent (diethylamine nonoate) relaxations were unaffected. Phenotypic markers of VSMC contractility remained unchanged. APP23+/- mice experience a lower survival rate compared to C57BL/6 littermates (39% vs. 100%, p=0.001) and APP23+/- mice showed increased corticosterone levels compared to C57BL/6 mice (13.39 ± 5.15 vs. 2.30 ± 2.52 µg/mL, p=0.0025) at 6 months of age. Conclusions: In conclusion, APP23+/- mice present with increased peripheral BP, aPWV and serum corticosterone levels in vivo and decreased vascular contractility ex vivo.

P096 / #829


CORTICOSTERONE LEVELS AFFECT IN VIVO ARTERIAL STIFFNESS INDIRECTLY WITHOUT AFFECTING EX VIVO ARTERIAL WALL BIOMECHANICS IN AMYLOID-Β OVEREXPRESSING MOUSE MODELS

Lecture Title:

J.O. Hendrickx1, S. De Moudt1, D. Van Dam2,3, G. De Meyer1, P. Fransen1 1Laboratory of Physiopharmacology, Faculty Of Pharmaceutical, Biomedical And Veterinary Sciences, University Of Antwerp, Wilrijk, Belgium, 2Laboratory of Neurochemistry and Behaviour, Institute Born-Bunge, Department Of Biomedical Sciences, University Of Antwerp, Wilrijk, Belgium, 3University of Groningen and University Medical Center Groningen, Department Of Neurology And Alzheimer Research Center, Groningen, Netherlands

Aims: Given the suggested interplay between arterial stiffness (AS) and Alzheimer’s disease (AD), we aimed to investigate arterial stiffness (AS) by a mutated human APP overexpression (APP23+/-) in C57BL/6 mice and an endothelial dysfunction mouse model of AS (eNOS-/-). Methods: C57BL/6 (male; n= 10), APP23+/- (male; n= 9), eNOS-/- (male; n=14) and eNOS-/-/APP23+/-

(male; n= 11) were studied at the age of 6 months. AS was determined by aortic pulse wave velocity (aPWV, VEVO2100) in vivo and by aortic Peterson Moduli (Ep) ex vivo through the in-house developed Rodent Oscillatory Tension set-up for Arterial Compliance (ROTSAC). Corticosterone levels were analysed on blood serum via ELISA. Data are represented as mean ± SEM. Results: Corticosterone levels were elevated by APP overexpression (C57BL/6 = 2.5 ± 1.4; APP23+/- = 10.1 ± 3.0; eNOS-/- = 13.1 ± 4.4; eNOS-/-/APP23+/- = 16.5 ± 2.4 µg/mL). Similarly, increased aPWV values were observed upon APP overexpression (C57BL/6 = 2.9 ± 0.1; APP23+/- = 4.1 ± 0.3; eNOS-/- = 4.6 ± 0.3; eNOS-/-/APP23+/- = 5.0 ± 0.9 m/s), which led to a positive Pearson correlation with aPWV values (p=0.0048, r=0.6335). No effect of APP overexpression was obtained in Ep values in C57BL/6 or eNOS-/- mice at baseline (KREBS), α1-adrenergic receptor-dependent contracted (phenylephrine (PE)) and fully contracted (PE and Nω-Nitro-L-arginine methyl ester hydrochloride) conditions. Conclusions: These observations suggest that overexpression of the APP protein might exert an indirect corticosterone-dependent effect on in vivo arterial stiffness without affecting ex vivo arterial wall biomechanics.

P097 / #12


BLOOD BETA-AMYLOID TO THE SEVERITY OF ALZHEIMER'S DISEASE.

Lecture Title:

Y.-H. Yang Kaohsiung Medical University, Neurology, Kaohsiung, Taiwan

Aims: Cerebral spinal fluid (CSF) beta-amyloid 1-40(Aβ1-40) andbeta-amyloid 1-42(Aβ1-42) peptides are considered the biomarkers to differentiate Alzheimer’s disease (AD) from healthy control (HC) although their roles are not well recognized in blood except Aβ1-40 was hypothesized more prominent in vascular amyloid deposit. Given to the invasiveness of CSF examination and the heterogeneous results in blood to separate AD from HC, whether these two blood biomarkers can clarify disease severity in order to increase their clinical significance are practicable and important. Methods: We have recruited 32 clinically diagnosed AD patientsbased on NINCDS-ADRDA criteria withthe annual comprehensive psychometrics including mini-mental status examination (MMSE) and clinical dementia rating (CDR) from our memory clinic. The venous blood samples were obtained in EDTA tube from all 32 participants in 2 groups: CDR=0.5:11 and CDR>0.5 group, CDR=1:17, and CDR=2: 4, for the Quantification of Aβ1-42 and Aβ1-40 in plasma using a specific ELISA kit :Human Amyloidβ(1-40) Assay Kit – IBL®with code number 27713 and Human Amyloidβ(1-42) Assay Kit – IBL®with code number 27711. All assays were performed according to manufacturer’s protocol. Results: For plasma Aβ1-40 level, patients with CDR=0.5 (n =11) was 201.3±152.7 (mean ±SD) pg/ml, which was significant lower to that, 369.5±168.5, of CDR>0.5 group (n=21) (p=0.010). There is no significant difference for Aβ1-42 in both groups (CDR=0.5: 40.9±28.5; CDR>0.5:47.2±47.1; p=0.691). Conclusions: Aβ1-40 was significantly prominent in advanced stage of AD. It highlighted the possibility of Aβ1-40 might contribute to advanced AD through vascular component. These relationships could be examined in coming study.

P098 / #349

Topic: Theme A: β-Amyloid Diseases / A1.m. Disease Mechanisms, Pathophysiology: Blood-brain barrier

THE PROTECTIVE EFFECT OF HIGH-FAT DIET ON ALZHEIMER'S DISEASE-RELATED PATHOLOGY AND COGNITIVE FUNCTION.

Lecture Title:

A. Amelianchik, J. Merkel, S. Kaneki, P. Palanisamy, E. Hyatt, E. Norris The Rockefeller University, Laboratory Of Neurobiology And Genetics, New York, United States of America

Aims: Alzheimer’s disease (AD) is a complex multifactorial disease that can develop as a result of various genetic and lifestyle factors. It has been reported that obesity may influence AD pathogenesis, yet there is no consistent association between high-fat diet (HFD)-induced obesity and AD pathophysiology in AD mouse models. Conflicting data may be due to experimental variations in previous studies, such as the mouse line and timeline of HFD consumption. Methods: The current study determined the effect of varied onset of HFD on AD-related pathology and cognitive function in transgenic Tg6799 AD mice. Results: HFD consumption starting at or before 3 months-of-age reduced extracellular amyloid-beta (Aβ) deposition and CD11b- positive microglia cell recruitment in the brain and improved cognitive function in AD mice. Additionally, HFD improved blood-brain barrier integrity and decreased fibrinogen extravasation into the brain parenchyma. However, delaying the onset of HFD until 6 months-of-age reduced the protective effect of HFD feeding, since HFD did not affect Aβ pathology or cognitive function in 11-month-old AD mice. Surprisingly, HFD still reduced the extravasation of fibrinogen into the brain despite the delayed onset of HFD consumption. Conclusions: Overall, we demonstrate that the onset of HFD consumption plays an important role in how dietary fats affect AD-related pathology and cognitive function in a transgenic mouse model of AD. These results suggest that dietary interventions may delay or prevent the onset of AD symptoms and underscore the importance of developing early therapeutic interventions to treat AD before patients develop cognitive deficits.

P099 / #451


CARBONIC ANHYDRASE INHIBITION AMELIORATES COGNITIVE DYSFUNCTION, ATTENUATING ABETA-INDUCED NEUROVASCULAR PATHOLOGY IN VIVO.

Lecture Title:

E. Canepa, R. Vazquez-Torres, R. Parodi-Rullan, S. Fossati Temple University, Alzheimer's Center At Temple (act), Philadelphia, United States of America

Aims: Neuropathological studies have established that 90% of Alzheimer's disease (AD) cases present cerebral amyloid angiopathy (CAA), a detrimental condition characterized by abnormal vasculotropic deposition of amyloid beta (Aβ). Our former in vitro work has shown that, in cells composing the neurovascular unit (NVU), including neurons, endothelial, glial and smooth muscle cells, vascular amyloidosis elicits mitochondrial dysregulation and caspase-mediated apoptotic pathways activation. Our group has demonstrated that acetazolamide (ATZ) and methazolamide (MTZ), FDA-approved carbonic anhydrase inhibitors (CAIs) which cross the blood-brain barrier (BBB), can hamper these pathological processes. Carbonic anhydrases (CAs) are metalloenzymes catalyzing the reversible hydration of carbon dioxide, and their inhibition improves cerebral blood flow and vasoreactivity. Methods: We fed Tg-SwDI (APP-Swedish, Dutch, Iowa) mice, model of CAA, a short- (3 months) or long-term (7/8 months) CAI-diet, following which behavioral analysis has been performed, and brains harvested for biochemical and IHC assessments. Results: CAIs ameliorate cognitive dysfunction, decreasing vascular Aβ accumulation and astrogliosis. In Tg mice, Aβ overload occurs in both endothelial and glial cells, triggering cell-specific caspase-3 activation. CAI-diet attenuates endothelial and astrocytic Aβ burden, the resulting caspase activation, and brain Aβ deposition. The increase of cerebral CD68+ cells in CAI-fed animals suggests a boost of the perivascular phagocytic activity that might explain the observed reduction of Aβ deposition. Conclusions: CAs mediate Aβ-induced apoptotic and inflammatory pathways, typically associated with CAA and AD, in endothelia and astrocytes. CAIs improve cerebrovascular health and provide neuroprotection.

P100 / #1128


NEUROVASCULAR SSAO/VAP-1 EXPRESSION ENHANCES AMYLOID-BETA-DEPENDENT NEURONAL SENSITIVITY IN A NEW NEUROVASCULAR UNIT IN VITRO MODEL.

Lecture Title:

C. Fábregas-Ordóñez1, J. Català-Solsona2, D. Siedlecki-Wullich2, M. Unzeta2, J. Rodríguez-Álvarez2, A.J. Miñano-Molina2, M. Solé-Piñol3 1Autonomous University of Barcelona, Biochemistry And Molecular Biology, Bellaterra (Cerdanyola del Vallès), Spain, 2Autonomous University of Barcelona, Biochemistry And Molecular Biology, Bellaterra (Cerdanyola del Vallés), Spain, 3Autonomous University of Barcelona, Institut De Recerca De La Vall D'hebron, Barcelona, Spain

Aims: To study possible alterations of the neurovascular unit (NVU) induced by the expression in cerebral endothelial cells of semicarbazide-sensitive amine oxidase/vascular adhesion protein-1 (SSAO/VAP-1). This enzyme is increased in the brain microvasculature and in plasma from Alzheimer’s disease patients and is involved in a pathologic cross-talk with amyloid-beta (Aβ). Methods: We established an in vitro model where we co-culture mouse cortical cells (neurons and glia) with the human brain microvascular endothelial cell line (hCMEC/D3) expressing or not the SSAO/VAP-1. In this new NVU model we analysed possible disruptions of the blood-brain barrier (BBB) by determining tight-junction proteins levels. Through multiplex immunoassay we studied alterations in the released angioneurins, signaling molecules affecting neurovascular functions. By immunoblotting and immunofluorescence experiments we determined the impact of these released molecules on synaptic proteins from neurons within our NVU model. Results: hCMEC/D3 cells expressing SSAO/VAP-1 displayed alterations in the BBB function and an increase in the release of some inflammatory angioneurins (IL-6, IL-8 and VEGF) along with a decrease in some trophic factors like BDNF, which is involved in glutamate receptors expression. The study of GluA1-staining intensity from neurons co-cultured in our NVU model revealed the presence of different cellular subpopulations. A decrease of the more intense GluA1 subpopulation on NVUs expressing SSAO/VAP-1 was found after treating our system with soluble forms of Aβ. Conclusions: Our co-culture model is a good system to study communicating events within the NVU. We identified neuronal affectations that may be related to an altered angioneurins pattern associated to SSAO/VAP-1 expression.

P101 / #1265


NEUROPATHOLOGICAL AND AMYLOID PEPTIDE DIFFERENCES BETWEEN DOWN SYNDROME AND FAMILIAL ALZHEIMER’S DISEASE WITH DUPLICATIONS AND MISSENSE MUTATIONS IN APP GENE

Lecture Title:

A. Kasri1, L. Durix1, S. Boluda1, L. Stimmer1, E. Gkanatsiou2, G. Brinkmalm3,4, Y. Vermeiren5,6, S. Pape7, P. De Deyn5,6, H. Zetterberg4,8, A. Strydom7, M.-C. Potier1 1Paris Brain Institute, ICM, Cnrs Umr7225 - Inserm U1127 – Upmc, Paris, France, 2. Institute of Neuroscience and Physiology, The Sahlgrenska Academy At The University Of Gothenburg, Gothenburg, Sweden, 3Sahlgrenska University Hospital, Clinical Neurochemistry Laboratory, Mölndal, Sweden, 4The Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Institute Of Neuroscience & Physiology, Mölndal, Sweden, 5University of Groningen, Department Of Neurology And Alzheimer Research Center, Groningen, Netherlands, 6Institute Born-Bunge, University of Antwerp, Department Of Biomedical Sciences,laboratory Of Neurochemistry And Behaviour, Wilrijk, Belgium, 7Institute of Psychology and Neuroscience, King’s College London, London, United Kingdom, 8UCL Institute of Neurology, Department Of Neurodegenerative Disease, London, United Kingdom

Aims: Alzheimer's disease (AD) is characterized by the extracellular amyloid-β peptide (Aβ) deposits and intraneuronal neurofibrillary tangles enclosing hyperphosphorylated tau protein. Additionally, Aβ can be found deposited in blood vessel walls as cerebral amyloid angiopathy (CAA). While amyloid plaques in post-mortem brains are common to all AD cases, including sporadic, familial, and Down syndrome (DS), CAA is a more prominent phenotype in familial cases. The study focuses on rare and poorly studied patient groups but potentially very informative ones, i.e., those with APP mutations or duplications (DUP-APP), and DS, to explain differences that may provide essential treatment clues. Methods: We are investigating the diversity of clinical and neuropathological phenotypes associated with various alterations in the APP gene. We study the alterations in the endo-lysosomal pathway (by immunohistochemistry) and Aβ species (by mass spectrometry) in post-mortem human brain tissues. Results: Our results showed increased Rab5 puncta size in all cases except in AD with APP point mutations compared to healthy controls. Moreover, Aβ peptide profiles showed an increase of Aβ40 but not Aβ42 species in DUP-APP and DS groups. These patients are characterized by a higher grading CAA, suggesting that (i) CAA is due to the specific accumulation of Aβ40 species appearing predominant. Alternatively, (ii) Aβ40 is the principal Aβ species produced in DUP-APP and DS cases that is more prone to aggregate in blood vessels. Conclusions: Analysis of Aβ and CAA grading is ongoing to establish a global comparison between patients and to unravel pathophysiological mechanisms involved in specific Aβ production and deposits.

P102 / #290


INORGANIC CLAY NANOCOMPOSITE SYSTEM FOR IMPROVED CHOLINESTERASE INHIBITION AND BRAIN PHARMACOKINETICS OF DONEPEZIL

Lecture Title:

A. Singh, S. Singh Institute of Medical Sciences, Banaras Hindu University, Center Of Experimental medicine & Surgery, Varanasi, India

Aims: To analyze the pharmacokinetics/pharmacodynamics parameters of laponite (LAP) nanocomposites: an effective approach for neurodegenerative disorder management. Methods: In the present study, laponite (LAP) nanocomposites were exploited for the improved brain delivery of donepezil (DZ) following encapsulation approach due to their nano-size and positive charge at pH < 9. Results: The size of prepared nanocomposites was 53.7 ± 4.0 to 137.7 ± 11.0 nm. The drug was released in a sustained manner till 120 h in phosphate buffer saline (pH 7.4) and acid phthalate buffer (pH 4.0). LAPDZ formulations inhibited acetylcholinesterase approximately by 82%, significantly higher (p< 0.05) than plain DZ (30%). Swiss albino mice exhibited enhanced brain uptake of LAPDZ administered via intravenous route. Promising pharmacokinetic parameters were observed in animals treated with LAPDZ. LAPDZ formulation showed half-life (t1/2), volume of distribution (Vd) and clearance (Cl) as 5.53 ± 0.40 h-1, 0.129 ± 0.02 L, 0.015 ± 0.002 L/h, respectively. While DZ solution showed the same parameters as 1.06 ± 0.12 h-1, 0.168 ± 0.01 L, 0.106 ± 0.013 L/h, respectively. The brain uptake of LAPDZ formulation was improved with quintuplet t1/2. Conclusions: Based on the results of present study, it is proposed that the formulated nanocomposite would result in improved patient compliance with therapeutic effect at lower doses.

P103 / #1628

Topic: Theme A: β-Amyloid Diseases / A1.n. Disease Mechanisms, Pathophysiology: Metabolism, insulin

27-HYDROXYCHOLESTEROL INFLUENCES OLIGODENDROGENESIS AND MYELIN INTEGRITY – IMPLICATIONS FOR ALZHEIMER’S DISEASE

Lecture Title:

V. Alanko1, R. Loera-Valencia1, A. Solomon2, I. Björkhem3, G. Tabacaru1, A. Cedazo-Minguez1, S. Maioli1, T. Quintela-López4, A. Gaminde-Blasco4, C. Matute4, M. Kivipelto1, E. Alberdi4, A. Matton1 1Karolinska Institutet, Neurobiology, Care Sciences And Society, Solna, Sweden, 2University of Eastern Finland, Neurology, Kuopio, Finland, 3Karolinska Institutet, Laboratory Medicine, Stockholm, Sweden, 4Faculty of Medicine and Nursery, University of the Basque Country (UPV/EHU), Department Of Neuroscience, Leioa, Spain

Aims: Midlife hypercholesterolemia is an established risk factor for developing Alzheimer’s disease (AD), still peripheral cholesterol does not pass the blood-brain-barrier. A proposed factor linking elevated systemic cholesterol levels and AD is the oxidized cholesterol metabolite 27-hydroxycholesterol (27-OH). It is known that high levels of the 27-OH has negative impact on the brain, yet the effect of high 27-OH levels on oligodendrocyte function remains unexplored. Abnormal myelin structure results in the disconnection of neural networks that is an early phenomenon in AD. This project aims to determine whether 27-OH affects myelination in the brain. Methods: Effects of 27-OH treatment was investigated both in a cell line and primary 3D cell cultures from mice. Brain sections from wild type (WT) and transgenic mice with increased systemic 27-OH levels (CYP27A1 tg) were immunohistochemically stained. CSF samples from subjective controls, mild cognitive impairment (MCI) and AD were analyzed for associations between AD biomarkers, 27-OH, and myelin proteins. Results: High concentrations of 27-OH induce cell death in human oligodenroglioma cells. 27-OH treatment stimulated oligodendrogenesis in mixed 3D cultures. Further, levels of Myelin Basic Protein (MBP) were increased in brain sections from CYP27A1 tg mice compared to WT. Patient CSF levels of MBP and the oligodendrogenesis regulating enzyme CNPase was associated to levels of 27-OH and AD biomarkers. CSF levels of MBP and CNPase were increased in MCI. Conclusions: The hypercholesterolemia associated 27-OH alters the rate of differentiation from oligodendrocyte progenitor cells into mature oligodendrocytes, suggesting that 27-OH reduces oligodendrocytic ability for appropriate remodeling in the aging brain.

P104 / #1084


REGULATION OF PERIPHERAL METABOLISM MEDIATED THROUGH CNS AMYLIN RECEPTORS ON ALZHEIMER’S DISEASE PATHOLOGY IN APP/PS1 MICE.

Lecture Title:

R. Corrigan1, G. Casadesus Smith2 1Kent State University, Biomedical Sciences, Kent, United States of America, 2Kent State University, Biology, Kent, United States of America

Aims: Amylin, a pancreatic hormone, known to regulate glucose homeostasis through its ability to sensitize insulin also acts as a satiety signal in conjunction with leptin. Interestingly, amylin is also an amyloid protein like amyloid-beta (Aβ); and, has been shown to be toxic when aggregated. Furthermore, amylin has also been shown by us and others to be neuroprotective in a non-aggregated state in Alzheimer’s disease (AD)-modeled mice. The amylin receptor (AMYR) is expressed throughout the brain, including areas important in both cognition and AD pathology. We have previously shown that an analogue of amylin, Pramlintide (PRAM), reduces Aβ plaque burden and recuses hippocampal-dependent cognitive decline in AD-mouse models. However, while beneficial in therapy, how PRAM signaling via AMYR to reduce Aβ burden remains largely unknown. Methods: Here we begin to address whether peripheral regulation of metabolism via the hypothalamus or other mechanisms in regions like the hippocampus are responsible for pramlintide neuroprotection. To address this question, male and female APP/PS1 mice were treated chronically with PRAM or saline peripherally in the presence or absence of AC187, an AMYR antagonist, delivered centrally. Results: Preliminary data show that PRAM reduces soluble AB1-42 in the cortex and increases alpha secretase in the hippocampus showing that the two mechanisms of PRAM may be independent of each other. Conclusions: Our data thus far suggests differential effects of peripheral amylin and even AMYR activation or antagonism on cognitive behavior, APP processing enzymes, and soluble amyloid-beta levels when in the presence or absence of receptor antagonism.

P105 / #789


CHANGES IN PERIPHERAL IMMUNE LANDSCAPE AND CIRCULATING METABOLITES INDUCED BY HIGH-FAT DIET IMPACT NEURONAL RESILIENCE IN AN ALZHEIMER’S DISEASE MODEL

Lecture Title:

S. Suzzi1, T. Croese1, A. Ravid2, O. Gold2, S. Medina1, S. Colaiuta1, S. Malitsky3, M. Itkin3, K. Vernon4,5, A. Clark4,5, A. Greka4,5, N. Habib2, M. Schwartz1 1Weizmann Institute of Science, Neurobiology, Rehovot, Israel, 2The Hebrew University of Jerusalem, Edmond & Lily Safra Center For Brain Sciences, Jerusalem, Israel, 3Weizmann Institute of Science, Life Sciences Core Facilities, Rehovot, Israel, 4Brigham and Women's Hospital and Harvard Medical School, Medicine, Boston, United States of America, 5MIT and Harvard, Broad Institute, Cambridge, United States of America

Aims: Mid-life obesity is an established risk factor for Alzheimer’s disease (AD), and the immune system an important common player in the two morbidities. We investigated whether obesity exacerbates AD, and if so, whether it is linked to immune system dysfunction and diet-associated metabolites. Methods: We fed 2-month-old 5xFAD mice, a model of AD-like amyloidosis, with a high-fat diet (HFD, 60% fat by caloric intake) for 28 weeks. Cognitive state was longitudinally assessed via novel object recognition (NOR) and radial arm water maze tasks. We used flow cytometry and cytometry by time-of-flight to analyze the peripheral immune landscape, and histo-/immunohistochemistry and ELISA to examine brain pathology. To identify the molecular signatures specific to the comorbid state as compared to AD or obesity alone, we performed untargeted metabolomic analysis on plasma, and single-nucleus RNA sequencing on whole hippocampus. Results: 5xFAD HFD mice exhibited early loss of working memory (NOR) relative to mice under normal diet. Furthermore, we found a significant increase in neuronal loss in the brain cortex in the 5xFAD HFD group relative to normal diet mice. HFD led to elevation of circulating Tregs, which positively correlated with weight gain, and to elevation of specific metabolites, some of which present at the highest level in the mice with both morbidities. Conclusions: Long-term HFD accelerates neuronal demise and cognitive impairment in 5xFAD mice. We suggest HFD-induced systemic immune suppression as the functional linkage between the two morbidities, which could be due, at least in part, to diet-induced metabolites, together affecting neuronal resilience.

P106 / #1766


AGE-DEPENDENT EVALUATION OF INSULIN PATHWAY RELATED GENES IN THE APPNL-F KNOCK-IN MOUSE MODEL OF ALZHEIMER’S DISEASE.

Lecture Title:

E. Trojan, M. Szuster-Głuszczak, A. Basta-Kaim Institute of Pharmacology Polish Academy of Sciences, Department Of Experimental Neuroendocrinology, Kraków, Poland

Aims: Data demonstrated that Alzheimer’s disease (AD) and type 2 diabetes (T2D) are among the most prevalent chronic diseases affecting the aging population. Data indicated that T2D is a well—established risk for AD, however the molecular mechanism underlying this association is still unclear. Similarly, open case remains a question how T2D may contribute to the progression of AD. The recent development of knock-in mouse model of AD provides distinct advantages over traditional transgenic mouse models that rely on over-expression of amyloid precursor protein. The aim of this study was to further biochemically characterize this model, in particular evaluate the age-dependent expression of brain insulin pathway genes expression in the AppNL-F knock-in mice. Methods: To address this issue, at 3- and 6-months of age, female mice were killed by rapid decapitation Hippocampi and frontal cortices were dissected and stored at -80°C until biochemical analysis. Using qRT-PCR we measured age-dependent mRNA expression of insulin, insulin receptor, and phospho-insulin receptor in both brain areas. Results: Biochemical study demonstrated that mRNA expression of all analyzed genes was significantly up-regulated in young 3 months of age APPNL-F females in hippocampus as well frontal cortex in comparison with control C57BL mice. At the age of 6 months we observed the down-regulation of all analyzed genes expression. Conclusions: Our study demonstrated that the expression of genes related with brain insulin pathway is age-dependent and shows signs of malfunction already at the age of 6 months in APPNL-F mice. Supported by the grant no. JPND/13/2019 National Centre for Research and Development

P107 / #178

Topic: Theme A: β-Amyloid Diseases / A1.o. Disease Mechanisms, Pathophysiology: Neural networks, plasticity

ALZHEIMER'S DISEASE, NEURAL RECEPTORS, AND VITAMIN D DEFICIENCY

Lecture Title:

A. Câmara Federal University of Rio Grande do Norte, Biophisics And Pharmacology, Natal, Brazil

Aims: This study aimed to review the relationship between VD deficiency, neural receptors, and Alzheimer disease (AD). Methods: We describe the proteins involved in AD pathogenesis and how those proteins can be influenced by VD deficiency. We investigated the receptors family most involved with AD as well as the tissue that most of these receptors are expressed. We also investigated a relationship between AD death rate and solar radiation worldwide using NASA data. The statistical analyses were performed using SPSS and R language. Results: We found an increased AD death rate in countries with low sunlight. It was also observed that amyloid precursor protein, ryanodine receptor, mammalian target of rapamycin complex 1, and receptor for advanced glycation end products are associated with a worse prognosis in AD. While the Klotho protein, phosphatase and tensin homologue, and VD receptor are associated with a better prognosis in the disease. In addition, nuclear transcription receptors and G-protein-coupled receptors were the most linked to AD. Most of these receptors are expressed in the cerebral cortex and hippocampus. Additionally, a great number of studies evaluated the receptors related to beneficial effects in the disease. The depletion of amyloid protein or the blockade of pathways related to its synthesis were the main functions performed by these receptors. Conclusions: Decline in VD concentrations may be involved in the establishment and progression of AD. According to sunlight data, we can conclude that countries with low average sunlight have high AD death rate.

P108 / #1307


A NOVEL IN VITRO PLATFORM TO STUDY ENTORHINAL NETWORKS IN ALZHEIMERS DISEASE

Lecture Title:

K.S. Hanssen1, A. Kobro-Flatmoen1, A. Sandvig2, I. Sandvig2, M. Witter1 1Kavli Institute for Systems Neuroscience, Faculty Of Health And Medicine, Trondheim, Norway, 2The Norwegian University of Science and Technology, Department Of Neuromedicine, Trondheim, Norway

Aims: Alzheimer’s disease (AD) is a progressive neurodegenerative disease where layer II of the entorhinal cortex (EC) is one of the first brain structures affected, suffering extensive early tangle pathology and neuron loss. We recently showed that the vulnerable layer II-neurons express the glycoprotein reelin, and that the layer II projection to the hippocampal dentate gyrus and CA3 arises exclusively from this reelin-expressing population. The reelin expressing EC layer II-neurons located close to the rhinal sulcus not only express higher levels of reelin but are also increasingly subject to early accumulation of intracellular amyloid-β. Here we aim to develop a platform to study these neurons at their adult stage in vitro. Methods: Our method involves surgically extracting and then culturing EC layer II-neurons from young adult APP/PS1 model mice on microelectrode arrays, enabling electrophysiological recordings. Our aim is to investigate early cell- and network-level dysfunctions hypothesized to arise due to early build-up of amyloid-β. Results: First results show survival of EC layer-II neuron cultures, establishing structural connections after 3 days in vitro. These neuronal networks start displaying spiking activity and local field potentials after 14 days in vitro and the network activity gradually increases up to 30 days in vitro. Alongside obtaining electrophysiological recordings on microelectrode arrays, ongoing work includes assessing the neuronal population in vitro by labelling for reelin expression in AD-model neurons and age-matched healthy controls. Conclusions: The current study offers an opportunity to explore EC layer II-neurons in vitro to further understand early cell- and network-level dysfunctions caused by early AD-pathology.

P109 / #851


ENHANCING THE POTASSIUM CHLORIDE CO-TRANSPORTER KCC2 REVERSES FUNCTIONAL DEFICITS ASSOCIATED WITH ALZHEIMER’S DISEASE-RELATED MUTATIONS IN MICE

Lecture Title:

I. Keramidis1, B. Mcallister2, J. Mehla3, P. Degagne2, A. Godin1, M. Mohajerani2, Y. De Koninck1 1CERVO Brain Research Center, Cellular And Molecular Neuroscience, Quebec, Canada, 2University of Lethbridge, Canadian Centre For Behavioural Neuroscience, Lethbridge, Canada, 3Washington University School of Medicine, Neurosurgery, St. Louis, United States of America

Aims: Growing evidence indicates that during early stages of Alzheimer’s disease (AD) an abnormal brain activity appears due to disruption of GABAA-mediated transmission. While disrupted GABAA signaling may result from several mechanisms, recent evidence points to deficits in the potassium-chloride cotransporter KCC2, responsible for maintaining low intracellular chloride in neurons to maintain robust inhibition. In this study, we validate whether KCC2 is downregulated in two transgenic lines that accumulate AD-like symptoms and whether by restoring KCC2 function we can alleviate deficits associated with AD Methods: The 5xFAD transgenic and APPNL-G-F knock-in lines are used in this study. Immunofluorescence analysis is conducted in layer II/III of the medial prefrontal cortex (mPFC) to assess the total and membrane KCC2 protein levels. The network activity of the CA1 pyramidal layer is recorded with a chronic electrode bundle. The memory and cognitive deficits are measured with the Morris Water Maze, Fear Conditioning and Elevated Plus Maze tests Results: We found a decrease in total and membrane protein levels of KCC2 in 4-month-old 5xFAD mPFC. In addition, the hippocampal gamma oscillation power was decreased in 9-month-old APPNL-G-F mice, consistent with predictions from deficits in KCC2. Treatment with CLP290, a KCC2 activity enhancer, restored the gamma oscillation power. Acute administration of CLP290 in APPNL-G-F mice also improved memory performance in the Morris Water Maze test the Contextual Fear Conditioning test Conclusions: These results indicate that KCC2 may be a viable target for reversing deficits in GABAA-mediated inhibition in AD and attenuating several symptoms associated with AD pathology

P110 / #1633


RECONFIGURATION OF THE CORTICAL-HIPPOCAMPAL INTERACTION IN APP/PS1 MICE MAY SUPPORT SPATIAL MEMORY FORMATION

Lecture Title:

N. Macrez1, B. Jura2, D. Młoźniak2, H. Goszczyńska2, K. Blinowska2, N. Biendon3, P. Meyrand4, T. Bem2 1Institut des Maladies Neurodégénératives, UMR 5293, CNRS Université de Bordeaux & INSERM, Neurocentre Magendie, U1215, Neurosciences, Bordeaux, France, 2Nałęcz Institute of Biocybernetics and Biomedical Engineering,, Polish Academy Of Sciences, Warsaw, Poland, 3Institut des Maladies Neurodégénératives, UMR 5293, CNRS Université de Bordeaux, Neurosciences, Bordeaux, France, 4INSERM, Neurocentre Magendie, U1215, Neurosciences, Bordeaux, France

Aims: Hippocampal-cortical dialogue is necessary for long-term consolidation of spatial memories. We recently demonstrated that freely moving APP/PS1 mice, a model of Alzheimer's Disease (AD), are able to learn a spatial reference memory task despite a major deficit in Sharp-Wave Ripples (SWRs). Here, we tested whether a reconfiguration of hippocampal-cortical dialogue occurs in these APP/PS1 mice by studying causal relations between hippocampal and cortical circuits during hippocampal SWRs. Methods: We analyzed the data set obtained from multielectrode intracranial recording of transgenic and wild-type mice undergoing consolidation of spatial memory and we applied Directed Transfer Function in order to determine the effective coupling between distributed circuits in some brain regions in which Aβ-amyloid plaques were quantified. Results: Hippocampal-cortical coupling in epochs containing SWRs display stronger coupling in the slow gamma range and the strength of effective coupling from the cortex to hippocampus (CA1) in the ripple band involves cortical areas that were different in the two groups of animals. In the wild-type group, retrosplenial cortex and posterior cingulate cortex interacted with the hippocampus most strongly, whereas in the APP/PS1 group more anterior structures (anterior cingulate cortex and prefrontal cortex) interacted with the hippocampus. Immunostaining of amyloid Aβ-plaques showed that the anterior cingulate and prefrontal cortex were less invaded by amyloid pathology than the posterior cortices. Conclusions: The reconfiguration of cortical-hippocampal interaction patterns may be an adaptive mechanism bypassing Aβ-plaques impact and allowing a less efficient though operational spatial memory consolidation in AD mice model.

P111 / #1074


EARLY VULNERABILITY OF CONTEXTUAL CIRCUITS TO AΒ DEPOSITION REVEALED WITH A NOVEL BEHAVIORAL MANIPULATION AND TWO-PHOTON IMAGING IN MICE

Lecture Title:

S. Moore1, A. Wang1, J. Lawlor1, K. Fogelson2, S. Ostojic3, K. Kuchibhotla1 1Johns Hopkins University, Department Of Psychological And Brain Sciences, Baltimore, United States of America, 2University of California San Diego, Biomedical Sciences Graduate Program, La Jolla, United States of America, 3École Normale Supérieure, Départment D’Études Cognitives, Paris, France

Aims: AD impacts the functional integrity of neural circuits before large-scale neurodegeneration, suggesting that targeting vulnerable circuits could be a powerful intervention strategy. An early AD symptom is context-dependent memory loss. Does this loss arise from deficits in neural encoding of ‘contexts’ or ‘memories’? Methods: We recently demonstrated that sensorimotor learning is context-dependent. We trained mice to lick after a tone for a water reward and to withhold from licking after another tone. We interleaved reinforced trials with trials without reinforcement (‘probe-context’, no licktube). Surprisingly, early in learning, animals performed better in the probe-context versus the reinforced-context suggesting that sensorimotor memories formation can be dissociated from its contextual access. Eventually, all animals performed similarly in the reinforced and probe-context, demonstrating that control mice have full contextual access to their memories. Here, we test whether mice with Aβ accumulation (APP/PS1+) exhibit memory (assessed in the ‘probe-context’) and/or contextual (assessed in the ‘reinforced-context’) deficits at plateau. Results: In 6-8mo APP/PS1+ mice, contextual-performance is significantly impaired compared to controls. Surprisingly, these same animals show only minor memory impairments. However, older APP/PS1+ mice (10-12mo) show both contextual and memory deficits. Two-photon calcium imaging in the auditory cortex of behaving animals revealed a reduction in stimulus selectivity in APP/PS1+ mice in reinforced trials. Conclusions: These results suggest that contextual circuits are selectively vulnerable to Aβ deposition early in disease progression, even before circuits involved in maintaining memories. We propose a role for aberrant inhibitory network integration of contextual signals, a conclusion further supported by computational modeling.

P112 / #929

Topic: Theme A: β-Amyloid Diseases / A1.p. Disease Mechanisms, Pathophysiology: Transcriptional & translational regulation, micro RNAs

THE TMEM106B FTD-PROTECTIVE VARIANT, RS1990621, IS ALSO ASSOCIATED WITH INCREASED NEURONAL PROPORTION

Lecture Title:

C. Cruchaga, F. Farias, B. Benitez, O. Harari Washington University School of Medicine, Psychiatry, St Louis, United States of America

Aims: Apart from amyloid β deposition and tau neurofibrillary tangles, Alzheimer disease (AD) is a neurodegenerative disorder characterized by neuronal loss and astrocytosis in the cerebral cortex. The goal of this study is to investigate genetic factors associated with the neuronal proportion in health and disease. Methods: To identify cell-autonomous genetic variants associated with neuronal proportion in cortical tissues, we inferred cellular population structure from bulk RNA-seq derived from 1,536 individuals Results: We identified that the variant rs1990621 located in the TMEM106B gene region was significantly associated with neuronal proportion (p=6.40×10-07) and replicated in an independent dataset (p=7.41×10-

04) surpassing the genome-wide threshold in the meta-analyses (p=9.42×10-09). We stratified the samples by disease status, and identified that this variant modulates neuronal proportion not only in AD cases but also in elderly neuropathology-free controls. This demonstrates that the association with neuronal proportion is not driven by individuals with FTD. Furthermore, we did not find a significant association in younger controls or schizophrenia patients, suggesting that this variant might increases neuronal survival or confer resilience to the neurodegenerative process. Our Mendelian Randomization approaches also indicate that lower neuronal proportions lead to increased AD and FTD risk Conclusions: We identified a protective variant in the TMEM106B gene that has neuronal protection effect in general aging, independent of disease status, which could help understand the relationship between aging and neuronal survival in the presence or absence of neurodegenerative disorders.Our findings suggest that TMEM106B could be a potential target for neuronal protection therapies to ameliorate cognitive and functional deficits.

P113 / #494


IMPLICATIONS OF NEURONAL MIRNA-124 MODULATION ON APP PROCESSING, TAU PHOSPHORYLATION AND CELL SECRETOME CONTENT USING HUMAN AD PLATFORMS

Lecture Title:

G. Garcia1, S. Pinto1, A. Fernandes1,2, T. Malm3, J. Koistinaho3, D. Brites1,2 1iMED.ULisboa - Research Institute for Medicines, Neuron-glia Biology In Health And Disease, Lisbon, Portugal, 2Faculty of Pharmacy of Universidade de Lisboa, Department Of Biochemistry And Human Biology, Lisbon, Portugal, 3University of Eastern Finland, A. I. Virtanen Institute, Kuopio, Finland

Aims: Alzheimer’s disease (AD) is the most common neurodegenerative disorder, with Amyloid-β (Aβ) and Tau as prime drivers. Recently, microRNAs (miRNAs) emerged as promising biomarkers/targets due to their multitarget/pleiotropic properties. For instance, miR-124 deregulation has been reported in neurodegeneration. Since in AD there is still no consensus on its role, we investigated the consequences of miR-124 modulation in SH‐SY5YAPP695 neuroblastoma (SH-SWE) and PSEN1DE9 mutant iPSCs-derived neurons (iNEU-PSEN) as human AD models. Methods: Cells were differentiated before transfection with miR-124-3p inhibitors/mimics. Tau/Aβ levels were quantified by Western Blot, Aβ release by ELISA, and APP expression and miRNAs by qRT-PCR. Exosomes were isolated by sequential ultracentrifugation and characterized. Results: Neuronal miR-124 was overexpressed in SH-SWE and iNEU-PSEN AD models, together with APP, comparing to wild-type controls. Interestingly, miR-124 mimic normalized the APP overexpression in both models, counteracted the elevation of low-molecular-weight oligomers (LMW) Aβ oligomers in SH-SWE and inhibited TAU phosphorylation in iNEU-PSEN. Although no alterations were found in Aβ1-40/42 release, we noticed that exosomes from both mutated cells were enriched in inflammation-associated miRNAs. Interestingly, miR-124 mimic depleted such exosomes in miR-146a and miR-155. Conclusions: This study involved two different models that match in endogenous miR-124 overexpression, despite each one represents a different AD-associated mutation. Further miR-124 overexpression with mimic revealed multiple cellular and paracrine therapeutic benefits. Despite the need for further validation studies, miR-124 hijack might be a promising strategy whenever patients show suppressed levels. Additional studies are essential to explore whether miR-124 neuroprotective effects are extended to glia, and how it revamps microglia-induced neuroinflammation.

P114 / #1355


BRAIN CELL-TYPE REGULATORY LANDSCAPES AND ASSOCIATIONS WITH ALZHEIMER'S DISEASE

Lecture Title:

A. Nott1, I. Holtman2, N. Coufal3, C. Glass3 1Imperial College London, Brain Sciences, London, United Kingdom, 2University Medical Center Groningen, Biomedical Sciences Of Cells & Systems, Groningen, Netherlands, 3University of California, San Diego, Cellular And Molecular Medicine, La Jolla, United States of America

Aims: There is no clear genetic cause for most cases of Alzheimer’s disease (AD), however, genome-wide association studies have identified genetic variants associated with increased risk of AD. These AD-risk variants are often located in non-coding DNA regulatory regions called enhancers, which upon transcription factor binding, modulate the expression of nearby genes. A major challenge for the AD field is to assign cell-type specificity and functionality to these genetic variants. Methods: Nuclei of microglia, neurons and glia were isolated from human cortical tissue and processed using ChIP-seq to define cell-type promoters and enhancers and ATAC-seq to define open chromatin regions. PLAC-seq, a chromatin conformation assay, was used to link distal enhancers to target genes. Enhancers were validated by CRISPR-mediated deletion of putative regions in induce-pluripotent stem cells (iPSCs) derived into brain cell-types followed by gene expression analysis. Results: Genetic variants associated with increased risk of AD were largely confined to microglia enhancers, suggesting that brain immune cells are an important cell-type for AD risk. Enhancer-promoter interactome maps connected disease-risk variants located in enhancers to promoters, providing putative microglia target genes. Lastly, the BIN1 AD-risk locus has a microglia-specific enhancer that harbors AD-risk variants. CRISPR-deletion of the microglia enhancer in iPSCs ablated BIN1 gene expression in iPSC-derived microglia. Conclusions: There has been intense interest in elucidating mechanisms that link human non-coding genetic variation to AD risk. Integrative analysis of promoter-enhancer interactome maps has revised and expanded the gene repertoire influenced by AD-risk variants and revealed the probable cell types impacted.

P115 / #239


APOE4 AFFECTS BASAL AND NMDAR MEDIATED PROTEIN SYNTHESIS RESPONSE IN NEURONS BY PERTURBING CALCIUM HOMEOSTASIS.

Lecture Title:

S. Ramakrishna1,2, S. Konings3, B. Schmid4, B. Holst4, K. Freude5, G. Gouras3, R. Muddashetty1 1Institute for Stem Cell Science and Regenerative Medicine, Institute For Stem Cell Science And Regenerative Medicine, Bangalore, India, 2The University of Trans-Disciplinary Health Sciences and Technology, The University Of Trans-disciplinary Health Sciences And Technology, Bangalore, India, 3Lund University, Experimental Dementia Research Unit, Department Of Experimental Medical Science, Lund, Sweden, 4Department of Bioneer A/S, Department Of Bioneer A/s, Hosrholm, Denmark, 5University of Copenhagen, Department Of Veterinary Clinical And Animal Sciences, Faculty Of Health And Medical Sciences, Copenhagen, Denmark

Aims: To study the effect of APOE4 on basal and synaptic activity mediated protein synthesis in neurons. Methods: Image for methods-

Results: Treatment of neurons with APOE4 (but not APOE3) for 20minutes causes an inhibition of global protein synthesis.This is shown through an increase in eEF2 phosphorylation, FUNCAT and polysome profiling (shift of ribosomes from heavier polysomes to lighter fractions).Further, the temporal profile of the translation response showed that both APOE3 and APOE4 lead to translation inhibition with 1minute treatment, but they have different recovery kinetics.The translation remains low in APOE4 treated neurons till 20minutes whereas APOE3 treated neurons recover. Further, we show that APOE4 mediated translation dysregulation is mediated by disruption of calcium homeostasis. While both APOE3 and APOE4 lead to calcium influx in neurons, the amount of calcium and the sources of calcium differ significantly. APOE3 causes a small peak of calcium through NMDARs only; whereas APOE4 leads to a sustained increase in calcium involving both NMDARs and L-VGCCs. As a consequence, along with the basal translation, NMDAR mediated translation response is also perturbed in the neurons which are treated with APOE4 for 20minutes. Interestingly, the physiological stimulation of NMDA receptors seems to evoke a stress response in APOE4 treated neurons. Hence,we hypothesize that the APOE4 mediated synaptic defects are caused due to perturbation of calcium homeostasis resulting in the inhibition of both basal and cue mediated synaptic translation. Conclusions: APOE4 causes sustained calcium influx through NMDARs and L-VGCCs leading to inhibition of global and NMDAR mediated protein synthesis response.

P116 / #1484


HOMER1 RNA MOLECULES CHANGES IN ALZHEIMER´S DISEASE HUMAN BRAIN

Lecture Title:

A. Urdanoz-Casado1,2, M. Robles1,2, M. Roldan1,2, I. Blanco-Luquin1,2, M. Mendioroz1,2,3 1IDISNA, Neuroepigenetica, Pamplona, Spain, 2Navarrabiomed Biomedical Research Center, Neuroepigenetica, Pamplona, Spain, 3Complejo Hospitalario de Navarra, Neurology Department, Pamplona, Spain

Aims: HOMER1 gene is closely implicated in synaptic plasticity and learning and memory. Recent studies show that circular RNA originated from HOMER1 (circHOMER1) expression is altered in some Alzheimer´s disease (AD) brain regions. Also, HOMER1 messenger (mRNA) levels are associated with Beta amyloid deposits in cortical regions. Our aim was to study the expression of circHOMER1 and its linear form in AD human entorhinal cortex, a brain region most vulnerable to AD pathology. Methods: We isolated RNA from human entorhinal cortex samples from 29 AD patients and 16 controls with RNeasy Lipid Tissue Mini kit (Qiagen). Genomic DNA was removed with recombinant DNase (TURBO DNA-free™ Kit, Ambion). Next, we perfomed Complementary DNA (cDNA) conversion 500 ng total RNA with SuperScript® III First-Strand Synthesis Reverse Transcriptase (Invitrogen) after priming with random primers. RT-qPCR reactions were performed in triplicate with Power SYBR Green PCR Master Mix (Invitrogen) in a QuantStudio 12K Flex Real-Time PCR System (Applied Biosystems) Results: A 2.32-fold decrease in circHOMER1 levels was observed in entorhinal cortex of AD cases compared to controls (p-value < 0.01). In addition, HOMER1 mRNA expression was downregulated in AD entorhinal cortex region compared to controls (Fold-change=2.37; p-values <0.01). Conclusions: We show that HOMER1 is deregulated in the entorhinal cortex of AD patients. These results add to the notion that HOMER1 and a its circular form could be playing a role in the pathogenesis of AD disease

P117 / #1429

Topic: Theme A: β-Amyloid Diseases / A1.q. Disease Mechanisms, Pathophysiology: Autophagy, apoptosis, cell death

AUTOPHAGY DISORDER IN RPE AS A POSSIBLE PATHOGENIC WAY OF Β-AMYLOIDOPATHY DEVELOPMENT IN AMD IN CORRELATION WITH AD

Lecture Title:

V. Ermilov1, A. Nesterova2 1Volgograd State Medical University, Forensic Medicine And Pathology, Volgograd, Russian Federation, 2Pyatigorsk Medical Pharmaceutical Institute of Volgograd Medical State University, Morphology, Pyatigorsk, Russian Federation

Aims: Autophagy plays a significant role in homeostasis of retinal pigment epithelium (RPE). Injury of RPE is a pathogenic base of the age-related macular degeneration (AMD) development. AMD associated with Alzheimer’s disease (AD) is morphologically characterized by the formation of β-amyloid deposits in both brain and eye. Our aim is to evaluate morphological features of β-amyloidopathy in the fundus of AMD eyes in correlation with AD. Methods: We examined 111 AMD eyes and 56 brains (both species from 56 Alzheimer’s donors) with electron mycroscopy and histochemical methods of amyloid detection. Morphometry was performed. Results: Microscopically, we identified the amyloid deposits in 39% (39 eyes of 100 AD eyes) with dry AMD and 80% (8 eyes of 11 AD eyes) with wet AMD. B-amyloid aggregations were detected in drusen, Bruch's membrane (BM), retina, and choroid vessels as well as in senile plaques and vessels of brain. The number of RPE nuclei was the lowest in the wet AMD in AD eyes with identified amyloidosis, in the age group 81-100 years. Conclusions: Based on the data obtained, we demonstrate that β-amyloid deposits correlate with retinal neurodegeneration, and it plays a key role in the pathogenesis of AMD. We hypotized that Aβ accumulation is associated with the impairment of autophagy in the RPE cells. This view creates the preconditions for new therapeutic strategies aimed at preventing autophagy dysfunction which leads to the formation of Aβ and its pathological aggregation.

P118 / #1539


NEUROPROTECTIVE EFFECT OF ARTEMETHER ON ALZHEIMER'S DISEASE MODEL BY ACTIVATING AMPK /GSK3Β(SER9)/ NRF2 SIGNALING PATHWAY

Lecture Title:

S. Li, W. Zheng University of Macau, Faculty Of Health Sciences, Macau, China

Aims: Alzheimer's disease is a progressive neurodegenerative disease and the most common cause of senile dementia. Inflammation, mitochondrial dysfunction and oxidative stress play significant roles in the progression of Alzheimer's disease. It has been suggested that the multifactorial nature of Alzheimer's disease pathogenesis requires the design of antioxidant drugs with a broad spectrum of neuroprotective activities. For this reason, the use of natural products, characterized by multiple pharmacological properties is advantageous over the single-targeted chemicals as AD-modifying drugs. Methods: We tested the protective effect of artemether on AD.The protective effect of artemether in neuronal cells confers Aβ1-42 induced cell injury was measured by MTT and LDH assay. Oxidative stress was followed up by measurements of intracellular ROS and mitochondrial membrane potential. In AD mice model, Mice memory was evaluated by Morris water maze test. Cell apoptosis was performed by TUNEL assay. Oxidative stress was determined by MDA and SOD activity measurements. Aβ expression and tau proteins phosphorylation was determined by Immunohistochemistry. The phosphorylation of signaling proteins was measured by western blotting. Results: Artemether attenuated neurotoxicity induced by Aβ1-42 in cell cultures. A temporal correlation was found between Artemether neuroprotection towards Aβ-induced neurotoxicity. In 3xTg-AD mice, Artemether therapy attenuated learning and memory, inhibited neuronal apoptosis and glial activation, inhibited oxidative stress and reduced Aβ deposition and tau phosphorylation and activated AMPK/GSK3β and Nrf2 signaling in vitro and in vivo. Conclusions: These findings suggest that artemether produced neuroprotective effects in neuronal cells and AD mice model which might be responsible for the neurotherapeutic effects of AD.

P119 / #1427


RNA SEQUENCING ANALYSIS OF ALZHEIMER APP KNOCK-IN MODEL MICE REVEALS MITOCHONDRIAL IMPAIRMENT, NEUROINFLAMMATION AND AUTOPHAGY IMPAIRMENT

Lecture Title:

M. Shimozawa1, L. Naia2, E. Bereczki1, R. Jiang1, X. Li3, J. Liu3, M. Ankarcrona2, P. Nilsson1 1Karolinska Institutet, Department Of Neurobiology, Care Science And Society, SOLNA, Sweden, 2Karolinska Institutet, Neurobiology, Care Science And Society, Stockholm, Sweden, 3Karolinska Institutet, Department Of Medicine, Huddinge, Sweden

Aims: Alzheimer Disease (AD) is a neurodegenerative disorder characterized by progressive impairment of memory and cognitive functions caused by the pathological accumulation of amyloid β (Aβ) peptides into Aβ plaques. Two novel App knock-in mouse models AppNL-F and AppNL-G-F recapitulate the Aβ pathology of AD, but the pathological role downstream of Aβ pathology of these mice still needs to be elucidated. Methods: We performed RNA-sequencing analysis on hippocampus from these two mouse models at three different ages, 1) before onset of pathology, 2) at onset of pathology and 3) at late stage in the pathology progression. Furthermore, we performed several validation studies of altered genes. Autophagy flux was measured in synaptosomal preparations of hippocampus. Results: We sequenced approximately 45,000 transcripts. Pathway analysis revealed that several inflammatory pathways such as Toll-like receptor signaling and TNF signaling were up-regulated. Mitochondrial pathways such as oxidative phosphorylation were altered in both AppNL-F and AppNL-G-F mice. We also found that pathways regulating autophagy were slightly up-regulated before onset of pathology, and it gradually decreased at late stage mimicking the processes present in AD brain. Several autophagy related genes were altered especially in AppNL-G-F mice. We also found the accumulation of autophagosomes in the isolated crude synaptosomal fraction from 12-month-old AppNL-G-F mice. Conclusions: We revealed neuroinflammation, mitochondrial and autophagy impairment in App knock-in mouse modes. These data suggest that novel App knock-in mouse models exhibit several phenotypes similar to AD brain, and these models are important tools for AD research.

P120 / #383


TAU EXPRESSION IS REQUIRED FOR CELLULAR SENESCENCE

Lecture Title:

A.C. Zaia Rodrigues1, V. Garbarino2, M. Orr1 1Wake Forest University, Gerontology-internal Medicine, Winston-Salem, United States of America, 2UT Health San Antonio, Gerontology, San Antonio, United States of America

Aims: Our previous work demonstrated that tau protein accumulation induced cellular senescence in transgenic mice and was associated with neuronal senescence in postmortem human brain. The objective of this project was to begin investigating the role of endogenous tau protein expression on cellular senescence. Methods: Male and female 20-month-old wild type and tau knockout (Mapt0/0) mice were used. We assessed physiological and behavioral measures associated with senescence including body weight, insulin resistance, anxiety, and cognitive impairment. Postmortem analyses on brain included tissue mass, histology, gene, protein and lipid analyses. Results: As previously reported Mapt0/0 mice were obese and insulin resistant and displayed anxiety like behavior, all previously attributed to cellular senescence in the brain. However, postmortem analyses revealed that Mapt0/0 mice expressed lower levels of senescence-associated genes (i.e., Cdkn2a, Cdkn1a, Tnfα, Il1β, Cxcl1), displayed lower senescence-associated β-galactosidase activity, and contained lower levels of lipids associated with senescence (i.e., acyl carnitine). Conclusions: Our data indicate that endogenous mouse tau expression is required for obesity and insulin resistance-induced cellular senescence in the brain. Our results suggest that physiological levels of tau are required to activate the cellular senescence response by upregulating senescence associated gene expression, lysosomal activity and lipid accumulation. Further studies are underway to elucidate how tau mechanistically regulates the senescence stress response.

P121 / #1514

Topic: Theme A: β-Amyloid Diseases / A1.r. Disease Mechanisms, Pathophysiology: Aging

A NOVEL COMPLEMENT C3 INDUCIBLE CONDITIONAL GLOBAL KNOCKOUT MOUSE MODEL

Lecture Title:

A. Batista1,2, M. Schroeder2, K. Khan2, J. Presumey3,4, E. Yalcin3,4, M. Carroll3,4, C. Lemere1,2 1Harvard Medical School, Department Of Neurology, Boston, United States of America, 2Brigham and Women's Hospital, Ann Romney Center For Neurologic Diseases, Boston, United States of America, 3Harvard Medical School, Department Of Pediatrics, Boston, United States of America, 4Boston Children’s Hospital, Program In Cellular And Molecular Medicine, Boston, United States of America

Aims: Complement component C3 is an innate immune protein involved in synapse elimination. Cerebral C3 is elevated with aging and Alzheimer’s Disease (AD). We previously reported that germline C3-deficiency protected aged wildtype and APPswe/PS1dE9 mice against hippocampal synapse loss and cognitive decline. Here, we generated a novel C3 inducible conditional knockout mouse line (C3iKO) to allow global C3 lowering at any age. Methods: We generated C3 floxed (C3fl/fl) mice and crossed them to an inducible, global Cre line (Rosa26-Cre-ERT2+/-; Jackson Laboratories) to produce C3fl/fl; Rosa26-Cre-ERT2+/- mice (C3iKO). Two-to-three month-old C3iKO mice were injected intraperitoneally with tamoxifen (TAM, 75 mg/kg) or corn oil daily for 5 days. We analyzed serum C3 levels at various intervals 7-180 days post-treatment, C3 levels in the brain at 60- and 150-days post-treatment, and C3, C1qa, C1qb, C1qc, C4b, CD11b and CD18 mRNA expression in the brain and liver 60- and 150-days post treatment. Results: Serum C3 levels were consistently reduced 85-90% in C3iKO+TAM mice compared to controls at all timepoints. Brain C3 levels were reduced ~60% at days 60 and 150. C3 mRNA expression was reduced ~90% in brain and ~82% in liver. None of the other complement component/receptor mRNAs were altered except C1q mRNA, which was elevated in brain and liver at day 60 but not day 150. Conclusions: Our novel C3iKO mouse model allows for global C3 lowering at any age and will be crossed with AD-like models to evaluate C3 lowering in early-stage AD pathogenesis.

P122 / #1322


MS-BASED ANALYSIS OF PROTEIN COMPLEXES FOR UNRAVELING INTERACTING PROTEIN MYSTERIES IN ALZHEIMER’S DISEASE PATHOLOGY.

Lecture Title:

B. Hasan1, A. Khan1, A. Asif2, C. Lenz2,3, N. Ahmed4 1University of Karachi, Department Of Biochemistry, karachi, Pakistan, 2Institute of Clinical Chemistry, University Medical Center Göttingen, Institute Of Clinical Chemistry, Göttingen, Germany, 3Max Planck Institute for Biophysical Chemistry, Bioanalytical Mass Spectrometry Unit, Göttingen, Germany, 4University of Karachi, Department Of Biochemistry, Karachi, Pakistan

Aims: Alzheimer’s disease (AD) is a complex neurodegenerative disorder with impaired protein activities. An in-depth understanding of protein complexes and their components is crucial not only for gaining insight of brain physiology but also towards developing drug therapies The aim of this study is to identify novel disease associated protein complexes and their potential role in the progression of AD pathology. Methods: Protein complexes were obtained from AD brain prefrontal cortex and age matched controls by Blue Native-Polyacrylamide Gel Electrophoresis (BN-PAGE) , a method of choice due to its simplicity and suitability for isolation and separation of interacting proteins working in a complex. A quantitative proteomic analysis was performed using second dimension SDS-PAGE followed by nano LC-MS/MS. Differentially expressed proteins were mapped to existing biological networks by Ingenuity Pathway Analysis (IPA). Results: A total of 13 protein complexes with their interacting proteins were resolved on SDS-PAGE. We identified 34 protein spots and found significant abundance difference between the two experimental samples. IPA analysis revealed degeneration of neurons and cell death as a major consequence of protein dysregulation. Furthermore, focused network analysis suggested an integrated regulation of the identified proteins through APP and MAPT dependent mechanisms. Conclusions: The interacting differentially expressed proteins in AD were found to be part of concomitant signaling cascades terminating in neuronal cell death. The identified protein networks and pathways warrant further research to study their actual contribution to AD pathology.

P123 / #1202


BIOCHEMICAL VARIANTS OF BETA-AMYLOID AND THEIR ASSOCIATION WITH HYPERPHOSPHORYLATED TAU IN BRAIN BIOPSIES FROM SUBJECTS WITH IDIOPATHIC NORMAL PRESSURE HYDROCEPHALUS

Lecture Title:

S. Libard1, I. Alafuzoff2 1Uppsala University Hospital, Surgical Pathology Department, Uppsala, Sweden, 2Akademiska sjukhuset, Klinisk Patologi, Uppsala, Sweden

Aims: To study different variants of beta-amyloid, and their association with hyperphosphorylated tau, in brain biopsies, from patients with idiopathic Normal Pressure Hydrocephalus (iNPH) and in a tissue micro array (TMA) including post mortem brain tissue samples from subjects with different stages of Alzheimer’s disease neuropathological change (ADNC). Methods: 127 brain biopsies from iNPH subjects with notable beta-amyloid pathology, with or without hyperphosphorylated tau pathology, and a TMA with core samples from 19 subjects with different stages of ADNC, were assessed and immunochistochemically (IHC) stained with antibodies (Ab) towards different beta-amyloid variants and an Ab towards hyperphosphorylated tau. The outcome of the IHC was assessed semi-quanititaively and in a subset of samples morphometrically. Results: All iNPH biopsies displayed beta-amyloid and 91% hyperphosphorylated tau. Significant increase of beta-amyloid -6F/3D, -4G8 and -phosphorylated 7H3D6 was seen with age. All beta-amyloid markers correlated with each other and with hyperphosphorylated tau in the whole cohort. In subjects 75 years old and older, the hyperphosphorylated tau correlated only with the modified pyroglutaminated- and phosphorylated beta-amyloid markers. In the TMA all the markers increased with the level of ADNC. All the markers correlated with each other except beta-amyloid 4G8 that did not correlate with the phosphorylated beta-amyloid species or with hyperphosphorylated tau. Conclusions: In the setting of iNPH the ADNC, the extent of pyroglutaminated- and phosphorylated species of beta-amyloid increase with age. In older population the pyroglutaminated- and phosphorylated species of beta-amyloid associates with the extent of hyperphosphorylated tau pathology in iNPH as previously reported in the setting of AD.

P124 / #443


UNDERSTANDING ROLES OF GWAS ALZHEIMER’S RISK GENES ON AGEING GLIA BIOLOGY, USING THE POWER OF DROSOPHILA MELANOGASTER.

Lecture Title:

E. Mackinnon, J. Williams, O. Peters Cardiff University, Dementia Research Institute, Cardiff, United Kingdom

Aims: Genome Wide Association Studies have identified several genes associated with increased risk of developing Alzheimer’s disease (AD), yet our knowledge of their biological functions and contribution to Alzheimer’s pathology is incomplete. Our primary goal is to ascertain the functional importance of these genes on glia biology in the ageing brain and explore their influence on amyloid beta neuropathology. Methods: To study glia autonomous effects of AD risk gene activity, we exploited the UAS/Gal4 binary system in Drosophila to conduct a RNAi screen of conserved Alzheimer’s associated genes. Using a well-established behavioural assay that correlates locomotion with CNS function, we detected modifiers of age-related locomotion behaviour as well as glia specific survival phenotypes. We secondly explored glia roles of the protective PLCG2 gene on amyloid beta pathology, using both the fly ortholog and cloned human PLCG2 transgenes. Results: We identified AD genes where loss of function, exclusively in glia, appears to shorten longevity. Preliminary findings also suggest reduced glial expression of small wing (sl)/PLCG2 modifies amyloid beta dependent neurodegenerative phenotypes. Conclusions: Our data suggests that glial expression of GWAS AD risk genes are required for the longevity and health of adult flies and that glial sl/PLCG2 expression modifies amyloid beta phenotypes. Future experiments aim to further study screen candidates, characterising mechanisms that underlie glia dysfunction throughout age and AD pathophysiology.

P125 / #445


LACTADHERIN (MFG-E8) ACCUMULATES IN AΒ-AFFECTED CORTICAL VESSELS IN CEREBRAL AMYLOID ANGIOPATHY

Lecture Title:

P. Marazuela1, M. Solé1, A. Bonaterra1, J. Camacho2, E. Martínez-Sáez2, O. Pancorbo3, C. Vert4, A. Rovira4, D. Rodríguez-Luna4, M. Hernández-Guillamon1 1Vall d'Hebron Research Institute, Universitat Autonoma de Barcelona, Neurovascular Diseases, BARCELONA, Spain, 2Vall d'Hebron University Hospital, Universitat Autonoma de Barcelona, Pathology Department, BARCELONA, Spain, 3Vall d'Hebron University Hospital, Stroke Unit, Neurology Department, Barcelona, Spain, 4Vall d'Hebron University Hospital, Radiology Department, Barcelona, Spain

Aims: Brain accumulation of amyloid-beta (Aβ) is a crucial feature in Alzheimer´s Disease (AD) and Cerebral Amyloid Angiopathy (CAA), although the pathophysiological relationship between them remains unknown. Numerous proteins are associated with Aβ in parenchymal plaques and/or brain vessels. We hypothesize that the study of these proteins will allow understanding specific biological differences between these two pathologies. Methods: Laser capture microdissection approach combined with proteomics was used to detect vascular and parenchymal Aβ-associated proteins in the APP23 transgenic mouse model of cerebral-β-amyloidosis. Validation of results was performed in brain and plasma from mouse and human samples and using cultured human brain vascular smooth muscle cells by western blot, immunochemistry and ELISA techniques. Results: We focused our study on one of the principal proteins identified in the Aβ-affected cerebrovasculature: Lactadherin (MFG-E8). We found that MFG-E8 brain levels were higher in APP23 compared to WT mice. Furthermore, MFG-E8 was strongly detected in Aβ-positive vessels in AD/CAA human brains, whereas Aβ-plaques did not present MFG-E8 immunodetection. Circulating levels of MFG-E8 were also analyzed in a cohort of patients clinically diagnosed with CAA; an inverse association was found between plasma MFG-E8 levels and the presence of MRI visible enlarged perivascular spaces in the centrum-semiovale. Finally, we found that the expression of MFG-E8 increased after the incubation of smooth muscle cells with the toxic Aβ40-Dutch peptide in vitro. Conclusions: Our results demonstrate a high presence of MFG-E8 in brain Aβ-positive vessels. Further functional studies are ongoing to elucidate its implication in Aβ accumulation and drainage trough the cerebrovasculature.

P126 / #1071


INVOLVEMENT OF CNS LUTEINIZING HORMONE RECEPTOR IN ALZHEIMER’S DISEASE PATHOGENESIS

Lecture Title:

M. Mey, G. Casadesus Smith Kent State University, Biology, Kent, United States of America

Aims: CNS Luteinizing hormone and its receptor have been shown to modulate neuronal plasticity and cognition. Importantly, levels of brain LH are inversely related to levels in the periphery, thus low upon menopause-related upregulation of peripheral LH. Previously we have shown that downregulation of peripheral LH improves function and plasticity in AD mouse models, improvements that are positively correlated with brain LH. However, whether these improvements are driven directly through the activation of central LHR signaling is not known. Therefore, the current study addresses whether central LHR activation was able to rescue cognitive and neuronal plasticity changes previously observed by peripheral LH downregulation. Methods: Female APP/PS1 transgenic and wild-type mice underwent ovariectomy or sham surgery and received ICV hCG or aCSF treatment for 8 weeks. Upon completion cognitive testing, tissue was collected to determine changes in structural plasticity and Abeta pathology. Results: Preliminary data demonstrates the ability of hCG treatment to differentially modulate cognition, pathology, and plasticity markers. Conclusions: This work supports the involvement of LHR in CNS function and begets its further study as a contributor linking menopause and increased AD risk.

P127 / #368


SINGLE-CELL RNA EXPRESSION CHANGES IN ALZHEIMER'S DISEASE

Lecture Title:

L. Soreq, C. Frigerio UCL, Molecular Neuroscience, Ramat Aviv, United Kingdom

Aims: Alzheimer's disease (AD) is a devastative late-life disease for which the age is a major risk factor. The underlying molecular mechanisms are mainly unknown. We propose to conduct a single-cell RNA sequencing experiments on 30 post mortem AD brain samples (including 3 brain regions and several BRAAK stages). We will analyze the produced data using Matlab to produce classification figures and statistical measures. We will use a Chromium machine. We will search for cell type-specific marker genes (microglia, neurons, oligodendrocytes) and will compare the outcomes to our recent aging brain study. We will complement the experiment by staining of 10 samples brain sections for 3 different cell types (neurons, microglia, oligodendrocyte) followed by machine learning computational quantification. The project findings may provide hope for patients and clues for potential genome-based therapeutic approaches. questions: 1. Which cell types contribute to disease progression? 2. Which splicing factors change in the disease? 3. What are the quantitative changes in microglia in AD? 4. Comparison to aging brains. Keywords: Alzheimer's disease, aging, brain, RNA, neurons, microglia, machine learning, bioinformatics Methods: We will use a Chromium machine for the single-cell sequencing experiment and will stain the samples using microscopy. 2. 10X Genomics kits (including ATAC+GEX kits - 16 reactions, on 30 samples) and Matlab for analysis. Results: RNA expression changes, cell type marker genes, alternative splicing detection. ,Cell quantification, Classification. Conclusions: We expect to gain insights on disease progression and underlying mechanisms and compare the changes to aging brains. We will discover changes in alternative splicing and inflammation.

P128 / #1425


THE ROLE OF SENESCENCE AND TELOMERE ATTRITION IN THE PATHOPHYSIOLOGY OF ALZHEIMER’S DISEASE

Lecture Title:

N. Suelves1, T. Ibrahim1, L. Quinton2, A. Decottignies1, P. Kienlen-Campard1 1Université Catholique de Louvain - UCLouvain, Institute Of Neuroscience, Brussels, Belgium, 2Université de Liège - ULiège, Biological Chemistry, Liege, Belgium

Aims: Aging is the main risk factor for most neurodegenerative diseases, including Alzheimer's disease (AD). Recent studies suggest that AD pathology correlates with the accumulation of cells that become senescent due to the progressive build-up of cellular insults, but the identification of senescent cells in the brain and their implication in brain dysfunction and neurodegenerative pathologies still needs to be explored. The progressive shortening of telomeres with each cellular division plays a central role in the senescence of dividing cells. We explored how brain senescence interconnects with brain dysfunction and AD. Methods: We used a mouse model of accelerated aging that is genetically deficient for the RNA component of the telomerase (Terc-/-), showing significant telomere attrition after 3 generations. We analyzed the brain proteome of Terc-/- mice and we validated classical and novel markers of senescence by cell-type specific immunostainings in primary cultures of major brain cell types (neurons and astrocytes). We finally investigated how pathological aging affects the onset and progression of AD by crossing our Terc deficient mice with the 5xFAD AD mouse model and evaluating different Aβ-induced neuropathological features. Results: Proteomic analysis uncovered key cellular pathways related to autophagy and mitochondrial function and novel protein candidates potentially involved in the transition towards a senescent state, which were validated in primary cultures. We also demonstrated that telomerase attrition induces an array of senescence markers and modifies the progression of amyloid-induced pathology in AD. Conclusions: Our results provide crucial knowledge that might led to the identification of potential targets for therapeutic development in AD.

P129 / #1668

Topic: Theme A: β-Amyloid Diseases / A1.s. Disease Mechanisms, Pathophysiology: Microbiome

INTESTINAL INFLAMMATION AND COMPROMISED BARRIER FUNCTION IN IDIOPATHIC PARKINSONISM: A SYSTEMIC REVIEW

Lecture Title:

C. Umamahesan, A. Augustin, D. Taylor, C. Weller, A. Charlett, R. Dobbs, S. Dobbs King's College London, Institute Of Pharmaceutical Sciences, London, United Kingdom

Aims: To address: how common are intestinal inflammation and compromised barrier function in idiopathic parkinsonism (IP); any potential treatment benefits; the outcome of not treating; whether screening is worthwhile. This may provide the missing link between systemic/brain inflammation in IP and implicated gastrointestinal microbiota/specific pathogens. Methods: Search strategy was based on PRISMA guidelines. Fifteen of 1395 articles (1995-2020) identified met inclusion criteria. Seven gave results on more than one intestinal modality: inflammation, permeability to passive diffusion, integrity, bacterial translocation. Results: The inter-relationship of IP with intestinal inflammation and bacterial translocation is firmly established, lacking only random sample surveys to meet Level-1 OCEBM evidence. Evidence for reduced integrity is limited to 2 small studies of tight-junction proteins in colonic biopsies. No overall conclusion can be drawn regarding faecal and circulating markers of integrity: evidence based on an assay that recognises the whole zonulin family, not the specific peptide, was censored. Evidence for increased permeability is insubstantial: further work is needed in IP, with and without small-intestinal-bacterial-overgrowth, and including a non-fermentable sugar-absorption test. Concentrations of markers of intestinal inflammation (faecal calprotectin) and bacterial translocation (circulating lipopolysaccharide-binding protein) appear not to change with IP duration or severity. This is compatible with a pre-presentation insult. There are no longitudinal studies on inflammation or translocation to guide design of interventional studies. Neither are cut-points discriminant for IP-facets, or gradients prognostic for its evolution, defined. Conclusions: Intestinal inflammation and barrier function is a strategic junctional point in the hypothesis for the aetipathogenesis of IP.

P130 / #1749

Topic: Theme A: β-Amyloid Diseases / A1.t. Disease Mechanisms, Pathophysiology: Other

UNDERSTANDING THE ROLE OF PLATELETS IN ALZHEIMER'S DISEASE

Lecture Title:

D. Bessa De Sousa1,2, M. Unger1,2, H. Mrowetz1,2, R. Poupardin2,3, T. Fröhlich4, L. Aigner1,2, K. Kniewallner1,2 1Paracelsus Medical University, Institute Of Molecular Regenerative Medicine, Salzburg, Austria, 2Paracelsus Medical University, Spinal Cord Injury And Tissue Regeneration Center Salzburg (sci-trecs), Salzburg, Austria, 3Paracelsus Medical University, Cell Therapy Institute, Salzburg, Austria, 4Ludwig Maximilian University of Munich, Laboratory Of Functional Genome Analysis (lafuga), Gene Center, Munich, Germany

Aims: In Alzheimer’s disease (AD) platelets become dysfunctional contributing to vascular injury and neuroinflammation. However, the molecular changes of platelets in AD remain largely unexplored. Aiming to characterise platelets at the molecular and cellular level, we analysed platelet’s contribution to AD pathology in a transgenic mouse model of AD (APP Swedish PS1 dE9, APP-PS1). Methods: We assessed the activation status (CD62P expression) and proteome of platelets in 14 months old APP-PS1 mice and wild type age-matched controls (WT). Further, we induced short-term (up to 9 days) immune-mediated platelet depletion in APP-PS1 mice (6-7months and 12-13 months old) by intraperitoneal injections of an anti-CD42b antibody. Results: APP-PS1 mice showed significantly higher percentages of activated platelets in the brain, but only a slight, non-significant, higher platelet activation in the bloodstream. Nevertheless, preliminary proteomics data revealed the existence of 77 differentially expressed proteins in APP-PS1 blood isolated platelets compared to WT mice. Interestingly, in the APP-PS1 mouse brain, about 20% of the platelets located extravascularly, where they associate with astrocytes. Antibody-mediated depletion successfully induced thrombocytopenia (>99%) in APP-PS1 mice for 5 to 9 days. Withdrawal of the antibody treatment had a rebound effect on platelet production, resulting in thrombocytosis. Conclusions: Platelets might present an altered cellular and molecular profile in AD. Short-term platelet depletion will help to enlighten the involvement of platelets in AD pathogenesis, particularly in amyloid plaque formation and microglial function, and potentially to identify novel targets of platelet biology, which might be developed as new drugs or drug targets.

P131 / #1544


ULTRASTRUCTURAL LOCALIZATION OF PORPHYROMONAS GINGIVALIS RGPB VIRULENCE FACTOR IN THE MIDDLE TEMPORAL GYRUS (MTG) OF THE ALZHEIMER’S DISEASE HUMAN BRAIN

Lecture Title:

S. Dominy1, F. Ermini1, U. Haditsch1, D. Raha1, M. Nguyen1, S. Broce1, L. Rodriguez1, S. Arastu-Kapur1, L. Holsinger1, C. Lynch1, V. Low2, J. Song2, M. Dragunow2, M. Curtis2 1Cortexyme, Research, South San Francisco, United States of America, 2The University of Auckland, Neurovalida, Auckland, New Zealand

Aims: Using light microscopy and immunohistochemistry, we recently identified gingipain virulence factors from the periodontal pathogen Porphyromonas gingivalis (Pg) in the MTG of postmortem Alzheimer’s disease (AD) brains (Dominy et al. 2019). Here, we aimed to validate techniques to determine the ultrastructural localization of the gingipain-R (RgpB) virulence factor in the MTG of the human brain. Transmission electron microscopy (EM) was employed to detect the subcellular immunoreactivity of the antibody CAB101 (anti-RgpB antibody). Methods: We processed fixed frozen MTG tissue sections at 50 μm thickness to detect the antigenicity of CAB101. In addition to rabbit anti-CAB101, we used an antibody against GFAP as a positive control for the experiments. For immunogold detection of the target antigen, we incubated sections with goat anti-rabbit 6 nm gold (anti-RgpB) and goat anti-mouse 15 nm gold (anti-GFAP) overnight. To ensure there was no non-specific binding of gold particles to the tissue sections, we included no primary antibody controls. Results: Using immunogold-EM, the RgpB antigen from Pg was detected in the MTG of AD brain within and in close proximity to cell nuclei and within blood vessel endothelial cells. When the primary CAB101 antibody was omitted, we were unable to detect any non-specific gold particle binding to tissue sections. Conclusions: We demonstrate that immunogold is a suitable and specific technique for the detection of the ultrastructural location of the RgpB-antigen from Pg in the human AD MTG using EM techniques.

P132 / #1483


EXAMINING METAL BIOCHEMISTRY IN ALZHEIMER’S DISEASE BRAINS USING SYNCHROTRON X-RAY SPECTROMICROSCOPY

Lecture Title:

J. Everett1, F. Lermyte2, J. Brooks2, V. Tjendana-Tjhin2, G. Plascencia-Villa3, I. Hands-Portman4, J. Donnelly2, G. Perry3, X. Zhu5, P. Sadler6, P. O'Connor6, J. Collingwood2, N. Telling1 1Keele University, School Of Pharmacy And Bioengineering, Stoke-on-Trent, United Kingdom, 2University of Warwick, School Of Engineering, Coventry, United Kingdom, 3UTSA, Department Of Biology And Neurosciences Institute, San Antonio, United States of America, 4University of Warwick, School Of Life Sciences, Coventry, United Kingdom, 5Case Western Reserve University, School of Medicine, Department Of Pathology, Division Of Experimental Pathology, Cleveland, United States of America, 6University of Warwick, Department Of Chemistry, Coventry, United Kingdom

Aims: Disrupted metal homeostasis is linked to the development of Alzheimer’s disease and the formation of amyloid plaques. However, our knowledge of metal (bio)chemistry in Alzheimer’s lacks the chemical and spatial sensitivity required to develop effective metal-targeting technologies for disease diagnosis/treatment. Here, advanced synchrotron x-ray spectromicroscopy was used to examine the nanoscale chemistry of metal inclusions within amyloid plaques isolated from Alzheimer’s disease brains. Methods: Amyloid plaques from two Alzheimer’s brains were examined using Scanning Transmission X-ray Microscopy (STXM) at Diamond Light Source beamline I08 and the Advanced Light Source beamline 11.0.2. STXM was performed over the calcium, copper and iron L-edges at nanoscale (< 100 nm) spatial resolution to determine the distribution and chemical state of these metals within the plaques. Results: Using x-ray absorption techniques coupled with magnetically-sensitive X-ray Magnetic Circular Dichroism (XMCD), we achieved a detailed chemical characterisation of metals harboured within Alzheimer’s amyloid plaques. STXM showed amyloid plaques to be calcified, and contain nanodeposits of copper and iron. Furthermore, these approaches allowed the identification of nanoscale variations in copper and iron oxidation state within the same amyloid plaques [1]. Conclusions: These results demonstrate the power of synchrotron x-ray spectromicroscopy to examine the biochemistry of Alzheimer’s disease. The identification of metal types uniquely associated with Alzheimer’s pathology will provide a critical step-change in our understanding of how metals contribute to the disease, thus facilitating the development of new viable therapies designed to restore metal balance in Alzheimer’s brains. References: 1. J. Everett et al. Nanoscale. 10, 11782-11796 (2018).

P133 / #550


HUMAN IPSC-DERIVED BRAIN TISSUE MODELS TO INVESTIGATE ALZHEIMER’S DISEASE

Lecture Title:

J. Klimmt1,2, C. Cardoso Goncalves1,2, S. Robinson1,2, G. Kleinberger3, C. Haass4,5,6, D. Paquet1,2,3,6 1University Hospital, LMU Munich, Institute For Stroke And Dementia Research (isd), Munich, Germany, 2Ludwig Maximilians Universität München, Graduate School Of Systemic Neurosciences, Planegg-Martinsried, Germany, 3ISAR Bioscience GmbH, Neurodegeneration Program, Planegg, Germany, 4Ludwig-Maximilians-Universität München, Metabolic Biochemistry, Munich, Germany, 5German Center for Neurodegenerative Diseases, Dzne, Munich, Germany, 6Ludwig-Maximilians-Universität München, Munich Cluster For Systems Neurology (synergy), Munich, Germany

Aims: Human induced pluripotent stem cell (hiPSC-) based models of Alzheimer’s disease (AD) enable investigation of pathomechanisms in disease-relevant, human brain cell types. They therefore offer great potential for mechanistic and translational studies, possibly complementing disadvantages of mouse models such as species differences or incompleteness of pathologies. However, current hiPSC-AD models enable mostly investigation of early pathologies including Aβ accumulation and Tau hyperphosphorylation, but fail to recapitulate later hallmarks such as widespread protein aggregation. Therefore, we aim to develop novel, advanced hiPSC-AD models that show late-stage phenotypes including Aβ plaques and tau tangles, without the use of gene overexpression or pathogenic tau mutations. Methods: To create these models, we generated human iPSC lines with AD mutations using genome editing to prime the cells for pathogenesis, and optimized protocols to differentiate these hiPSCs into highly pure cultures of disease-relevant cell types, including cortical neurons, astrocytes and microglia. Currently, we establish controllable models of human brain tissue, by 3D co-culturing all cell types to elicit AD pathogenesis in a physiologically relevant system. Results: We already observed typical early AD phenotypes such as increased Aβ secretion and phospho-Tau levels, Aβ accumulation in the extracellular space, and microglial activation. Currently, we are optimizing the model to elicit later-stage pathology such as Aβ plaques and Tau tangles. Conclusions: Our model will form the basis for studies elucidating novel, potentially human-specific pathomechanisms and provide a framework for translational and screening approaches to develop disease-modifying therapies.

P135 / #761


SIGNIFICANCE OF N-GLYCOSYLATION FOR PATHOGENIC PROCESSING AND TRAFFICKING IN ALZHEIMER DISEASE

Lecture Title:

T. Lin1, E. Berlin2, L. Tjernberg1, S. Schedin Weiss1 1Karolinska Institutet, Division Of Neurobiology, Care Sciences And Society, Stockholm, Sweden, 2Linköping University, Department Of Clinical And Experimental Medicine (ike), Linköping, Sweden

Aims: N-glycosylation is essential for cellular function of membrane and secretory proteins. It is disturbed in Alzheimer disease (AD) and inhibition of N-glycosylation leads to altered processing of the amyloid precursor protein (APP) including reduced production of the amyloid β-peptide (Aβ). Here, we study the effects of N-glycosylation on APP processing, sorting and trafficking in cultured neurons via introduced mutations at N-glycosylation sites on APP. Methods: • Plasmid vectors encoding human APP695 (wild-type or with mutations introduced at N-glycosylation sites) and a C-terminal SNAP-tag that can be fluorescently labeled in the live cells were used. • The APP-SNAP vectors were delivered into HEK293T cells and cultured primary mouse cortical neurons with transfection reagent Lipofectamine 3000. • Transfected cells were labeled with organelle markers and SNAP-tag reagents. Samples were visualized by using live cell imaging and confocal microscopy. Results: APP-SNAP was localized in a variety of vesicles and at the plasma membrane. The mutations that prevent N-glycosylation in APP alter the processing and trafficking of APP. Conclusions: N-glycosylation is one of the most essential post-translational modifications and contributes to correct APP processing and trafficking. Lack of individual N-glycans due to mutations in N-glycosylation sites may affect the subcellular localization of APP and, thus, result in alterations in APP processing pathways. These mechanisms could be a contributing factor in the development of AD.

P136 / #594


LINKING AMYLOID TO COGNITION IN THE PATHOGENESIS AND TREATMENT OF ALZHEIMER’S DISEASE: TOWARD THE DEVELOPMENT OF A “QUANTITATIVE A/T/N MODEL"

Lecture Title:

N. Mazer1, C. Hofmann1, D. Lott1, R. Gieschke1, G. Klein2, M. Traber3, G. Kerchner4 1Roche Innovation Center Basel, Pharmaceutical Sciences - Clinical Pharmacology, Basel, Switzerland, 2F. Hoffmann-La Roche Ltd, Biomarkers And Translational Technology, Neuroscience And Rare Diseases, Basel, Switzerland, 3F. Hoffmann-La Roche Ltd, Product Development Medical Affairs, Basel, Switzerland, 4F. Hoffmann-La Roche Ltd, Product Development Neuroscience, Basel, Switzerland

Aims: Jack and colleagues proposed a research framework for Alzheimer’s Disease, based on biomarkers of beta-amyloid, tau, and neurodegeneration (“A/T/N”) (Jack 2016; 2018). To elucidate the mechanisms underlying the canonical sequence from A to T to N to clinical outcomes, and to simulate the potential downstream effects of anti-amyloid therapy, we developed a mathematical representation of this framework—the “Quantitative A/T/N Model” (Q-ATN). Methods: Published data and biologically plausible mechanisms were used to construct, calibrate and validate the Q-ATN. Its five modules (anti-amyloid effect, amyloid PET, tau PET, cortical thickness [CT], and cognitive impairment [CDR-SB]) and four linkages (L1-L4) are shown in Figure 1. Treatment with a hypothetical anti-amyloid antibody was simulated with the Q-ATN model over 5 years.

Results: Linkage 1 (inset A) quantifies amyloid dynamics with a first-order rate constant (kDE) for antibody-mediated plaque removal and a parabolic growth function. Linkage 2 (inset B) quantifies the effect of amyloid on tau PET based on data from the HABS study (Johnson 2020) and hypothesized mechanisms of tau production and clearance. Linkage 3 (inset C) quantifies the effect of tau on the rate of

change of CT (dCT/dt). Linkage 4 (inset D) quantifies the relationship between medial temporal CT and CDR-SB based on data from Dickerson 2009. The 5-year simulation (Figure 2) suggests that antibody-mediated amyloid removal (A) will reduce tau PET (B), slow cortical thinning (C), and decrease the rate of cognitive decline (D).

Conclusions: The Q-ATN model provides a novel quantitative framework that connects anti-amyloid therapy to the amelioration of cognitive decline.

P137 / #686


INVESTIGATING ABETA-INDUCED TOXICITY IN TAU-DEFICIENT HUMAN NEURONS AND ALZHEIMER’S DISEASE PATIENT-DERIVED NEURONS

Lecture Title:

B. Ng1, J. Vowles2, D. Beccano-Kelly1, T. Caffrey1, Z. Cader3, S. Cowley2, N. Connor-Robson4, R. Wade-Martins4 1University of Oxford, Physiology, Anatomy And Genetics, Oxford, United Kingdom, 2University of Oxford, Dunn School Of Pathology, Oxford, United Kingdom, 3University of Oxford, Weatherall Institute Of Molecular Medicine, Oxford, United Kingdom, 4University of Oxford, Department Of Physiology, Anatomy And Genetics, Oxford, United Kingdom

Aims: Amyloid-beta (Abeta) and tau aggregates are two major pathological hallmarks and known to be dysregulated in Alzheimer’s disease (AD), leading to widespread aggregation. We now know from tau-deficient animal models that tau does not merely facilitate Abeta-induced toxicity but is also essential in that particular toxic cascade. However, a tau-deficient human in vitro model has not been available to corroborate these studies. We set out to understand the effects of tau depletion in human induced pluripotent stem cell (iPSC)-derived neurons and further extended the study to AD patient-derived neurons in a clinical cohort. Methods:

Here we generated the first isogenic MAPT-/- human induced pluripotent stem cell (iPSC) lines in order to study the effects of tau deficiency in human biological context, together with iPSC lines from 14 sporadic AD patients. We then differentiated the lines into cortical neurons in co-culture with rat astrocytes, and utilised AD brain homogenate to serve as the source of extrinsic Abeta in addition to synthetic Abeta oligomers. Results: iPSC-derived MAPT-/- neurons exhibited functional deficits compared to MAPT+/+ neurons, while expressing similar number of synapses. On the other hand, MAPT-/- neurons demonstrated impaired axonal outgrowth over 5 days. Upon extrinsic Abeta insults, MAPT-/- neurons appeared to be protected from cytotoxicity and functional hyperactivation as compared to MAPT+/+ neurons. However, the MAPT-/-

background was unable to prevent Abeta-induced loss of synapses.

Conclusions: Taken together, the absence of tau in human neurons resulted in phenotypic changes at baseline and could result in neuroprotection in MAPT-/- neurons from Abeta-induced toxicity.

P138 / #1180


THE AD KNOWLEDGE PORTAL: DATA, ANALYSES, AND TOOLS FROM THE NATIONAL INSTITUTE ON AGING'S ALZHEIMER'S DISEASE TRANSLATIONAL RESEARCH PROGRAM

Lecture Title:

M. Peters1, A. Greenwood2, W. Poehlman2, Z. Leanza1, J. Gockley3, L. Heath2, J. Wiley2, T. Thyer2, N. Kauer1, J. Schneider1, K. Woo1, C. Suver4, K. Montgomery1, S. Sieberts1, B. Logsdon2, L. Omberg1, L. Mangravite2 1Sage Bionetworks, Systems Biology, Seattle, United States of America, 2Sage Bionetworks, Neurodegeneration Research, SEATTLE, United States of America, 3Sage Bionetworks, Neurodegeneration Research, Seattle, United States of America, 4Sage Bionetworks, Research Governance & Ethics, Seattle, United States of America

Aims: There is a need to understand the molecular complexity of AD in order to develop therapeutics that target the full range of disease presentation. Three major challenges are associated with this: 1) identifying new molecular hypotheses and mechanisms, 2) vetting hypotheses via independent experimental assessments, 3) prioritizing new molecular mechanisms for therapeutic development. We posit that resources and technologies which empower open community-driven solutions will be key to the overall success of creating new AD therapeutics. Methods: We have developed the AD Knowledge Portal to tackle these challenges. The Portal publicly shares data from several NIA funded research consortia, including the Accelerating Medicines Partnership in AD. Data release is based on a rapid model without embargoes where data is available for general research use. To enable reproducible analysis of data on the portal, open-source computational workflows for processing common data types such as raw RNA sequencing reads are available. In addition, an Analytical Workspace allows users to easily provision cloud computing resources such as RStudio notebooks in a secure AWS environment. Results: Portal data has been used by investigators studying AD and aging, basic neuroscience, and cell biology. Data reuse has led to new insights into disease molecular mechanisms, including the role of herpes simplex virus in AD pathophysiology, the identification of sex-specific transcriptomic differences in AD, and the development of tools for the prediction of disease pathology. Conclusions: Our framework is designed to accelerate AD target discovery by expanding community-driven efforts to tackle each component of the AD therapeutic development process.

P139 / #1780


ACUTE RESPONSES OF THE GUT AND THE ENTERIC NERVOUS SYSTEM TO Β-AMYLOID EXPOSURE

Lecture Title:

A. Christmann, M. Gries, S. Schulte, K.-H. Schäfer University of Applied Sciences, Imst, Zweibrücken, Germany

Aims: The number of patients suffering from Alzheimer's disease rises with increasing life expectancy. Unfortunately, the disease becomes only clinically apparent when a significant neural damage already occurred. Usually, this is too late for a curative therapy. Since there is growing evidence that onset and course of neurodegenerative disorders might be influenced by or even start in the gastrointestinal tract, its intrinsic nervous system, the enteric nervous system (ENS) might be an ideal target for both, early diagnosis and treatment. We therefore investigated acute effects of β-amyloid (Aβ42) on the ENS. Methods: To evaluate the impact of Aβ42 on gastrointestinal function, a mesenterial perfusion of rodent gut was performed with nicotine, nicotinic blockers and the pathological peptide. To analyze the role of the intrinsic enteric nervous system, myenteric plexus was isolated, cultured and treated with Aβ42. Neuronal activity was measured by multi-electrode-arrays (MEA) and Ca-Imaging. Results: Mesenterial perfusions with nicotine or Aβ42 lead to significantly increased contractions, which could both be blocked by nicotinic inhibitors. MEA recordings revealed a dose-dependent activity change in cultured enteric neurons, which were acutely treated with Aβ42. In addition, Ca-Imaging measurements showed an increase of synaptic activity in cultured enteric neurons. Conclusions: In this study we showed functional changes of the gut in the presence of amyloid. Moreover, we could deliver evidences for an acute response of the ENS to the application of the neuropathological peptide Aβ42 in electrophysiological in vitro studies.

P140 / #900


DISRUPTED CIRCADIAN RHYTHMS ARE ASSOCIATED WITH INCREASED COGNITIVE DECLINE IN MILD-MODERATE ALZHEIMER’S DISEASE PATIENTS

Lecture Title:

A. Targa1, I. Benitez1, F. Dakterzada2, J. Fontenele-Araújo3, A. Carnes2, M. Pujol4, L. Elkhayat2, O. Minguez4, M. Dalmases1, M. Sanchez-De-La-Torre1, F. Barbe1, G. Piñol-Ripoll2 1Hospital Universitari Arnau de Vilanova-Santa Maria, IRBLleida; Centro de Investigación Biomédica en Red de Enfermedades Respiratorias (CIBERES), Translational Research In Respiratory Medicine, LLeida, Spain, 2Hospital Universitari Santa Maria-IRBLleida, Cognitive Disorders Unit, LLeida, Spain, 3Federal University of Rio Grande do Norte, Physiology And Behavior, Natal, Brazil, 4Hospital Universitari Arnau de Vilanova-Santa Maria, IRBLleida, Translational Research In Respiratory Medicine, Lleida, Spain

Aims: To investigate the association between circadian rhythms and the cognitive evolution of mild-moderate Alzheimer’s disease patients. Methods: We performed an observational, prospective, single-center study including patients with mild-moderate Alzheimer’s disease (NCT02814045). The rest-activity rhythms of the individuals were monitored through the use of an actigraph for 14 days. Based on this, we evaluated different aspects of circadian rhythms such as the synchronization with zeitgebers (interdaily stability), the fragmentation of rest-activity rhythms (intradaily variability), and the amplitude (relative amplitude). This was followed by neuropsychological evaluations, which were also performed after 12 months of follow-up. Results: Our cohort included 101 subjects with a mean (SD) age of 75.6 (5.07) years. We observed an estimated decrease of 3.276 (95% CI: 0.527 to 6.025; p = 0.020) points in Mini-Mental State Examination (MMSE) for a one-unit increase in intradaily variability. This was maintained after adjustment for age, sex, educational level, and Apolipoprotein E status. Similarly, there was an estimated decrease of 8.087 (95% CI: 0.432 to 15.743; p = 0.039) points in MMSE for a one-unit increase in the relative amplitude after adjusting for conditional factors. There was no association between interdaily stability and cognitive evolution. Conclusions: This preliminary analysis suggests that increased fragmentation of rest-activity rhythms as well as increased amplitude are associated with a worse cognitive evolution in patients with mild-moderate Alzheimer’s disease.

P141 / #678


EFFECT OF TIMES TO BLOOD PROCESSING ON THE STABILITY OF BLOOD PROTEINS ASSOCIATED WITH DEMENTIA

Lecture Title:

J.-F. Chang1, C.S.Y. Yang1, P.-N. Wang2, H.-C. Liu1, H.-H. Chen1 1MagQu Co., Ltd., President Office, New Taipei City, Taiwan, 2Taipei Veterans General Hospital, Department Of Neurology, Taipei, Taiwan

Aims: The stability of proteins in collecting tubes after blood draw is critical to the measured concentrations of proteins. Although the guidelines issued by Clinical & Laboratory Standards Institute (CLSI) suggest the centrifugation of blood within two hours after blood draw, it is very difficult to follow these guidelines in hospitals or clinics. It is necessary to study the effect of times to blood processing on the stability of proteins of interest. Methods: In this work, plasma proteins of interest are those relevant to dementia, such as amyloid β 1-40 (Aβ1-40), Aβ1-42, Tau protein (Tau) and α-synuclein. The times to blood processing after blood draw ranged from 0.5 to 8 hours. The storage temperatures of blood were room temperature (~25 °C) and 30 °C. After storage, blood samples were centrifuged at room temperature to obtain plasma samples. Ultrasensitive immunomagnetic reduction was applied to assay these proteins in plasma. Results: The levels of plasma Aβ1-40, Tau, and α-synuclein did not significantly change until 8 hours after blood draw when stored at room temperature. Plasma Aβ1-42 levels did not change significantly after 8 hours of storage at room temperature before blood processing. Higher storage temperatures, such as 30 °C, for the blood samples accelerated the significant variations in the measured concentrations of Aβ1-40, Tau and α-synuclein in plasma. Conclusions: According to these results, it is suggested to store blood samples at room temperature for no longer than 4.5 hours for processing centrifugation after blood draw in clinical practice for the assay of dementia biomarkers in plasma.

P142 / #487

Topic: Theme A: β-Amyloid Diseases / A2.a. Therapeutic Targets, Mechanisms for Treatment: Abeta, truncated & pGlu-Abeta

ELEVATED LEVELS OF SOLUBLE AMYLOID BETA PROTOFIBRILS IN DOWN’S SYNDROME AND ALZHEIMER’S DISEASE

Lecture Title:

T. Odergren1, M. Johannesson1, C. Sahlin1, L. Söderberg1, H. Basun1, J. Fälting1, C. Möller1, O. Zachrisson1, D. Sunnemark2, A. Svensson2, L. Lannfelt1 1BioArctic AB, R&d, Stockholm, Sweden, 2Offspring Biosciences, R&d, Södertälje, Sweden

Aims: The hallmarks of Alzheimer’s disease (AD) are the presence of senile plaques in the brain containing amyloid β (Aβ) and neurofibrillary tangles with hyperphosphorylated tau and neuronal loss. Aβ is produced by cleavage of the amyloid precursor protein, which gene is located on chromosome 21. Down’s syndrome (DS), trisomy of chromosome 21, leads to the propensity to develop Aβ brain pathology followed by cognitive and behavioral deterioration. The purpose of this study was to provide characterization of amyloid species in brains from subjects with DS and AD, and to compare with non-demented controls (NDC). Since similarities in the Aβ species pattern in AD and DS could be anticipated, binding to BAN2401 (an antibody in phase 3 development for AD) was included in the study. BAN2401 has high selectivity for soluble toxic Aβ aggregates, such as oligomers or protofibrils. Methods: Brain samples from DS and AD subjects as well as NDC were analyzed with respect to Aβ species. Immunohistochemical staining using antibodies towards Aβ was also performed. Results: Elevated levels of soluble Aβ protofibrils and insoluble Aβx-40 and Aβx-42 in brain extracts, and elevated immunohistochemical staining of Aβ deposits were demonstrated with the antibody BAN2401 in DS and AD compared with NDC. Conclusions: These data and the recent promising data with BAN2401 in a large phase 2 AD clinical trial, suggest that BAN2401 may warrant clinical study in subjects with DS with signs of functional or cognitive deterioration.

P143 / #1647

Topic: Theme A: β-Amyloid Diseases / A2.b. Therapeutic Targets, Mechanisms for Treatment: Immunotherapy

TAPAS-PART-1: HUMANIZED ANTIBODIES AGAINST N-TRUNCATED AMYLOID-PEPTIDES AMINO ACIDS (TAPAS) AND DEVELOPMENT OF NOVEL VACCINE WITH COMPARABLE FEATURES

Lecture Title:

P. Bakrania1, S. Davies1, H. Payne1, J. Price1, C. Mpamhanga1, E. Love1, M. Carr2, G. Hall2, R. Cowan2, D. Matthews1, T. Bayer3 1LifeArc, Centre For Therapeutics Discovery, Stevenage, United Kingdom, 2University of Leicester, Molecular And Cell Biology, Leicester, United Kingdom, 3University Medicine Goettingen, Psychiatry, Goettingen, Germany

Aims: We have previously identified a mouse antibody (TAP01), which bound to non-plaque forms of Aβ and showed therapeutic potential in mouse models of Alzheimer’s disease (AD). Here, we report the development of novel humanized antibodies, the crystal structure of the binding pocket and the discovery of a novel epitope representing a vaccine for active immunization. The therapeutic effects of these new therapeutic approaches were evaluated using two transgenic Alzheimer mouse models. Methods: Transgenic mice (5XFAD, Tg4-42), ELISA, peptide generation, protein expression and purification, Biacore analyses, site-directed mutagenesis, antibody humanisation Results: Crystal structures of N-truncated Abeta bound to TAP01 revealed that Aβ adopted a novel structure, not related to any fibrillar or amyloid associated conformations reported previously and accounts for the non-plaque specificity of TAP01. The crystal structures enabled the design of stable and novel Abeta peptide used as a vaccine. Binding ELISA data of sera against the novel epitope was analyzed in wildtype, 5XFAD and Tg4-42 mice. The sera showed comparable binding to the novel epitope and N-truncated Abeta peptides and no binding to full-length Abeta, as has been demonstrated for humanized TAP01 antibodies. Furthermore, we compared the binding properties of the humanized TAP01 antibodies and the sera against the novel Abeta epitope with other possible therapeutic antibodies. We clearly demonstrate unique binding features not seen with any other comparator antibodies. Conclusions: We have identified a new conformational state of N-terminal region of Aβ, which clearly plays a key role in AD progression and reveals at least two attractive routes for therapeutic development.

P144 / #1354


LONGITUDINAL COGNITIVE STIMULATION AND AΒ IMMUNOTHERAPY AMELIORATE COGNITIVE IMPAIRMENT IN THE 3XTG-AD MOUSE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

A.R. Roda Universitat Autònoma de Barcelona, Bioquímica I Biologia Molecular, Cerdanyola del Vallès, Spain

Aims: Background. AD is characterized by accumulation of extracellular A-beta peptide in the form of senile plaques and intracellular hyperhosphorylated tau protein in neurofilament tangles, which causes neuronal degeneration and eventually neuronal death leading to dementia. The 3xTg-AD mouse model replicates both histopathological markers and behavioral and cognitive deficits. Behavioral alterations are first detected when mice are 2.5 month-old and long-term memory deficits between 3- and 5-month old. Objectives. To test the effect of monthly administration of the Aβ-directed antibody fragment scFv-h3D6 and repeated cognitive stimulation on behavioral and cognitive abilities. Methods: . Once a month NTg and 3xTg-AD mice were intraperitoneally administered with PBS or 100 micrograms of scFv-h3D6. Twenty-four hours after treatments at 5, 7, and 9 months of age, behavioral (CT and OFT) and cognitive (MWM) testing were performed. Results: . Neophobia and anxious-like behavior were evident in 5-month old 3xTg-AD female mice, supporting an early onset of behavioral and psychological symptoms of dementia (BPSD). Both NTg and 3xTg-AD mice exhibited a decrease in exploratory activity over the longitudinal study because of normal aging process. Although long-term memory was clearly impaired in 5-month old 3xTg-AD mice, learning abilities were not seriously affected. ScFv-h3D6-treatment was able to ameliorate early cognitive impairment detected at 5 month-old but not BPSD-like symptoms. Interestingly, cognitive stimulation, by task repetition in the Morris Water Maze over time, was also effective in reducing cognitive deficits. Conclusions: Conclusion. Combination of both, A-beta immunotherapy and cognitive stimulation could constitute a good therapeutic approach for the early treatment of AD.

P145 / #542


PROTEOLYTIC DEGRADATION OF HIPPOCAMPAL AΒ42 WITH A BRAIN-PENETRATING NEUROPEPTIDE

Lecture Title:

F. Rofo1, C. Yilmaz Ugur1, N. Metzendorf1, T. Gustavsson2, C. Beretta2, F. Sandbaumhüter1, E. Jansson1, A. Erlandsson2, D. Sehlin2, S. Syvänen2, P. Nilsson3, G. Hultqvist1 1Uppsala University, Department Of Pharmaceutical Biosciences, Uppsala, Sweden, 2Uppsala University, Department Of Public Health And Caring Sciences, Uppsala, Sweden, 3Karolinska Institutet, Dept Of Neurobiology, Care Sciences And Society, Stockholm, Sweden

Aims: Aggregation of the amyloid-beta peptide (Aβ) is one of the main pathological events in Alzheimer’s disease. A potential therapeutic intervention in Alzheimer’s is to enhance the degradation of Aβ prior to aggregates formation. This can be achieved by enhancing the activity of the major Aβ degrading enzymes, including neprilysin. Immunotherapy using neuropeptides has previously been suggested as a promising therapeutic strategy. However, poor delivery over the blood-brain barrier (BBB) and short plasma half-life of neuropeptides limit their therapeutic significance. Methods: We developed a fusion protein based on a neuropeptide recombinantly linked to a BBB transporter binding to the transferrin receptor. The ability of the new protein drug to pass BBB, activate neprilysin and degrade Aβ was tested using mice overexpressing the Aβ-precursor protein with the Swedish mutation (APPswe). Using quantitative LC/MS, the effects of the formed construct on the proteome in the brain was evaluated. Results: The new construct exhibited long plasma half-life and high brain uptakes when injected intravenously. A significant increase in the concentration of neprilysin and degradation of hippocampal Aβ42 could be detected in the brain of APPswe mice after only three injections. The fusion protein could also demonstrate some effects in regulating mitochondrial and synaptic proteins involved in Alzheimer’s disease pathology. Conclusions: Since the hippocampus is the central area in the initiation of Alzheimer’s disease, our new protein-based drug can serve as a potential treatment option in the early stages of the disease with a promising safety profile targeting only the disease affected areas.

P146 / #456


EVALUATION OF CLASS C CPG ODN EFFICACY AND SAFETY PROFILE IN A NON-HUMAN PRIMATE MODEL OF SPORADIC CAA

Lecture Title:

H. Scholtzova1, A. Patel1, P. Nehete2, B. Nehete2, S. Karimi1, K. Leigh Dowling-Campbell1, T. Genovese1, D. Guttman1, B. Drittel1, L. Williams2, T. Wisniewski1 1NYU School of Medicine, Neurology, New York, United States of America, 2UT/MD Anderson Cancer Center, Michale E. Keeling Center For Comparative Medicine And Research, Bastrop, United States of America

Aims: Many immunotherapeutic clinical trials for Alzheimer’s disease (AD) have failed due to poor translatability of promising transgenic mouse data to humans. Our recent data from a non-human primate (NHP) model of sporadic cerebral amyloid angiopathy (CAA), squirrel monkey (SQM), indicate that treatment with TLR9 agonist, class B CpG ODN, results in CAA reduction without toxicity. Here, we advanced our study using class C CpG ODN, which shows favorable safety profiles in a variety of human diseases. This is the first study assessing the effectiveness of immunostimulatory class C CpG in an aged population whose immune system might be senescent. Methods: Elderly female SQMs received either CpG ODN or saline every 4 weeks as part of an ongoing chronic-treatment study. Plasma was analyzed to identify autoantibody responses toward Abeta 40/42 and cytokine/chemokine profiles using Luminex technology. Changes in mRNA expression levels were periodically evaluated in isolated PBMCs prior to and 24hrs post-injection using Nanostring nCounter System. Results: Administration of CpG ODN was effective in inducing plasma cytokine/chemokine levels without toxicity. Nanostring analysis demonstrated significant up-regulation of IFN-inducible genes (ISG-54K/IFIT2, Mx2/MxB, GBP1) and specific cytokine-chemokine genes following CpG ODN injection. IgG/IgM autoantibodies against Abeta did not increase when assessed periodically. Furthermore, longitudinal MRI monitoring for Amyloid-Related Imaging Abnormalities (ARIA) enhanced evaluation of CpG ODN safety, and post-treatment behavioral assessments are currently underway. Conclusions: The present study provides essential information prior to any clinical use of CpG ODN in AD. Use of this advantageous NHP model is important to reach efficient translation for human benefit.

P147 / #797

Topic: Theme A: β-Amyloid Diseases / A2.c. Therapeutic Targets, Mechanisms for Treatment: Secretases, proteases

FRAGMENTS GENERATED FROM APP CLEAVAGE BY ETA-SECRETASE IN ALZHEIMER'S DISEASE: PROTEOLYSIS AND FUNCTIONS

Lecture Title:

E. Afram, F. Checler, R. Pardossi-Piquard Institut de pharmacologie moléculaire et cellulaire, Alpes-maritimes, VALBONNE, France

Aims: Amyloid precursor protein (APP) is a transmembrane protein undergoing canonical cleavages by α, β and γ-secretases. Recently, the matrix metalloprotease MT5-MMP, referred to as η-secretase, has been identified as a novel APP cleaving enzyme (Baranger et al., 2016). This enzyme produces a transmembrane ηCTF that undergoes subsequent cleavages by α and β-secretases generating Aηα and Aηβ (Willem et al., 2015). Methods: The fate and functions of ηCTF and its related fragments are poorly understood. In our study, we transiently overexpressed ηCTF, Aηα and Aηβ in various cell models (SH-SY5Y, HEK293, SHAPPwt, SHAPPswe) and analyzed their degradation by pharmacological interference with classical degradation pathways. Thus, we treated cells either with inhibitors of the proteasomal machinery (lactacystine and MG132) or with compounds affecting authophagic process (Bafilomycine A1 and Smer28). Furthermore, we more selectively evaluated the effect of secretases on the expression of ηCTF by treating the cells with various secretases inhibitors. Results: Our results demonstrate that overexpressed ηCTF undergoes degradation in the lysosomal pathway, and that it is cleaved by α, β and γ-secretases as expected. Conclusions: Given that the ηCTF protein that we are overexpressing is cleaved and processed as shown for endogenous ηCTF, we have undertaken biochemical, immunohistochemical, electrophysiological and behavioral approaches to assess the respective contribution of the various catabolites generated by η-secretase in Alzheimer’s Disease.

P148 / #1246


PYRUVATE KINASE M2 REGULATES GAMMA-SECRETASE ACTIVITY AND AGGRAVATES MEMORY IMPAIRMENT IN ALZHEIMER’S DISEASE MICE

Lecture Title:

J. Hyun1, J. Han2, J.-S. Park3, S.-M. Jung3, E.-I. Jeong3, S.-H. Kim3, Y. Oh3, D.-G. Jo4, Y.-K. Jung3 1Seoul National University, Life Science, Seoul, Korea, Republic of, 2University of Texas Texas Southwestern Medical Center, Department Of Neuroscience, Texas, United States of America, 3Seoul National University, School Of Biological Sciences, Seoul, Korea, Republic of, 4Sungkyunkwan, School Of Pharmacy, Suwon, Korea, Republic of

Aims: Gamma (γ)-secretase is a key enzyme that catalyzes the generation of amyloid beta peptides (Aβ). In Alzheimer’s disease (AD), Aβ is thought to cause synaptic dysfunction and memory deficits. Hypoxia is a major risk factor for AD and enhances Aβ generation. Yet, the relationship between hypoxia and γ-secretase regulation remains unclear. Here, we report that pyruvate kinase M2 (PKM2), an enzyme involved in glycolysis, acts as a positive regulator of γ-secretase under hypoxia. Methods: We identified PKM2 from a genome-wide functional screen using a cell-based assay and cDNA expression library. PKM2 expression significantly increased in the brains of patients with AD and a murine model of AD (PDAPP AD model mice). Using cDNA overexpression and shRNA knockdown, We measured Abeta40 and Abeta42 production. Gamma-secretase holo-complex level were checked by Blue Native- PAGE. Real-Time qPCR were performed to examine transcriptional regulation of APH1a Results: PKM2 expression level in AD patient and model mice were increased. Overexpression of PKM2 increased the production of Aβ1-42 and Aβ1-40, while shRNA-mediated knockdown of PKM2 reduced Aβ1-42 and Aβ1-40 generation. PKM2 dimer can act as transcirption co-activator. PKM2 affect APH1a mRNA transcriptional regulation. Through APH1a expression by transcription, APH1a-NCT subcomplex were enhanced by PKM2 effect. Hypoxia condition induced PKM2 expression via HIF-1α and thus promoted γ-secretase activity and APP intracellular domain (AICD) generation in mouse primary neuron. Conclusions: These results suggest that dimeric PKM2 is an important γ-secretase regulator that promotes Aβ production and memory impairment under hypoxia condition

P149 / #574


MT5-MMP AT THE CROSSROADS OF NEUROINFLAMMATION AND AMYLOIDOGENESIS: STUDY IN MURINE AND HUMAN NEURAL CELLS

Lecture Title:

D. Pilat1, J.-M. Paumier2, L. Garcia Gonzalez1, L. Arnaud1, L. Greetham1, L. Louis1, D. Stephan1, A. Bernard1, E. Di Pasquale1, M. Khrestchatisky1, E. Nivet1, K. Baranger1, S. Rivera1 1Aix-Marseille University, Institute Of Neurophysiopathology, Cnrs, Marseille, France, 2Northwestern University, Feinberg School Of Medicine, Chicago, United States of America

Aims: We have shown that MT5-MMP deficiency in the 5xFAD mouse model of AD strongly reduces accumulation of Abeta, C99 and neuroinflammation, all concomitant with the prevention of deficits in synaptic activity and learning. However, the impact of MT5-MMP on those processes at pre-symtomatic stages of the pathology is still elusive. Methods: To address this question, we used MT5-MMP deficient mouse primary neurons and iPS-derived neurons from non-AD and AD patients where MT5-MMP was suppressed using Crispr/Cas9. We investigated the role of MT5-MMP in these settings using molecular and cell biology, biochemistry, immunocytochemistry and whole cell patch clamp. Results: We demonstrated that the lack of MT5- MMP prevents the accumulation of toxic APP metabolites and reduces inflammatory response to IL-1beta at early stages of the development in primary neuronal cultures from 5xFAD mice, but also in iPS-derived human neural cells. Moreover, we showed in 5xFAD cells that MT5-MMP deficiency prevented the loss of dendritic spines and neuronal hyperexcitability. Interestingly, MT5-MMP deficiency also negatively impacted synaptic integrity in the non-AD background. Overall, MT5-MMP seems to play a physiological role in synaptogenesis and also contribute to early pathogenic APP metabolism, neuroinflammation and synaptic disruption. Conclusions: Overall, these data confirm that MT5-MMP is a new multifaceted player in AD pathogenesis and a potential target that could pave the way for the study of new therapeutic strategies.

P150 / #341


BETA-SECRETASE MODULATORS WITH A DUAL ACTIVITY AGAINST BOTH AMYLOID AND TAU PATHOLOGY

Lecture Title:

M. Tautou1, S. Eddarkaoui1, F. Descamps2, P.-E. Larchanché2, C. Lamarre1, D. Blum1, L. Buee1, P. Melnyk2, N. Sergeant1 1Inserm UMR-S 1172 Lille Neuroscience & Cognition, Alzheimer & Tauopathies, Lille, France, 2Inserm UMR-S 1172 Lille Neuroscience & Cognition, Brain Biology & Chemistry, Lille, France

Aims: Small compounds have been synthesized and share different strengths of lysomotropic and beta-secretase modulating activities. The structure activity relationship has identified two molecules MAGS and PEL differing from each other by a single azote that is necessary for the lysomotropic activity (Gay et al, Neurobiol Dis 2019). Since several compounds of this family are efficient toward both amyloid and tau pathologies, our objective is to determine which of the lysomotropic or beta-secretase modulating activities is necessary for the curative effect and cognitive improvement in a mouse model of neurofibrillary degeneration. Methods: In vitro experiments were performed using SY5Y-APPwt cells. Thy-Tau22 transgenic mice were treated for 1 month in a curative paradigm. Short term memory was evaluated using the Y-maze, followed by biochemical experiments including Western-blot and Immunohistochemistry to investigate the effect on the neurofibrillary degenerating process. Results: MAGS, with both lysomotropic and b-secretase activity, had no effect on the behavioral activity of Thy-tau22 treated mice while PEL, with the b-secretase modulatory activity, was shown to restore the short term memory and to reduce the neurofibrillary degenerating process. This effect was associated with a reduced phosphorylation of Tau and a decrease of inflammatory markers. Conclusions: PEL is a new molecule acting on APP proteolysis by modulating the β-secretase activity and is also effective on Tau pathology and cognitive functions. Although the mechanism of action is not yet fully understood, we already know that the lysomotropic activity is dispensable for the effect on both Aβ and Tau pathologies.

P151 / #343


MULTI-EFFECT MOLECULES WITH A NOVEL PHARMACOLOGICAL TARGET: AN INDIRECT ΒETA-SECRETASE INHIBITOR FOR THE TREATMENT OF AΒ LESIONS IN ALZHEIMER’S DISEASE

Lecture Title:

M. Tautou1, S. Eddarkaoui1, F. Descamps2, P.-E. Larchanché2, C. Lamarre1, L. Buee1, D. Blum1, P. Melnyk2, N. Sergeant1 1Inserm UMR-S 1172 Lille Neuroscience & Cognition, Alzheimer & Tauopathies, Lille, France, 2Inserm UMR-S 1172 Lille Neuroscience & Cognition, Brain Biology & Chemistry, Lille, France

Aims: Alzheimer's disease is the most common neurodegenerative disorder in the elderly and has still no effective treatment. Targeting the two main pathological processes, Tau pathology and Aβ plaques, with molecules acting on both lesions is a interesting strategy since the two processes are considered as synergistic. Derived from our pharmacophore, we have selected two molecules MAGS and PEL differing from each other by a single azote. PEL has an inhibitory activity on Aβ production but no longer the characteristic lysomotropic activity. We aim in this study to determine which of the lysomotropic or b-secretase modulating activity is efficient against amyloidogenesis. The effect on Tau pathology is the aim of a complementary study. Methods: 8 months-old APP/PS1 mice (double-transgenic model of amyloidogenesis) were treated for four months. Cognition benefits were measured using Y-Maze and Barnes tests. Then the amyloid pathology was analysed in the hippocampus and the cortex using immunohistochemistry, ELISA and also qRT-PCR to investigate several inflammatory markers. Results: In the present study we showed that PEL with an indirect β-secretase activity but no lysomotropic activity had a positive effect on the Aβ pathology and cognitive functions in vivo, as short –term and long-term memory were restored. Conclusions: PEL is a new molecule acting on APP proteolysis by modulating the β-secretase activity with a promising efficiency on Aβ pathology and cognitive functions. Although the mechanism of action remains ill-defined, we already know the b-secretase modulatory activity is essential to reduce both the Aβ and Tau pathology.

P152 / #295

Topic: Theme A: β-Amyloid Diseases / A2.d. Therapeutic Targets, Mechanisms for Treatment: Kinases, other enzymes

THE CUTTING EDGE OF DYRK1A IN AXONAL TRANSPORT REGULATION AND ITS IMPLICATIONS IN NEURODEGENERATIVE DISEASES.

Lecture Title:

I. Fernandez Bessone, J. Navarro, J.E. Martinez, M. Holubiec, T. Saez, T. Falzone University of Buenos Aires, Institute Of Cell Biology And Neuroscience "prof. E. De Robertis", Buenos Aires, Argentina

Aims: In Alzheimer Disease (AD) the abnormal intracellular distribution of the amyloid precursor protein (APP) affects its processing and, consequently, the generation of Aβ. Axonal transport play key roles in the neuronal distribution of APP. The dual-specificity-tyrosine phosphorylation-regulated-kinase-1A (DYRK1A) has been associated with AD onset since over-expression was found in Down syndrome and sporadic AD patients. Moreover, specific APP and tau phosphorylations are mediated by DYRK1A. Therefore, we tested whether DYRK1A has a role in APP axonal transport regulation. Methods: We developed highly-polarized human-derived neurons in 2D cultures. At day 15 we inhibited DYRK1A for 48 hs with harmine (7.5 μM). We then generated a plasmid and a lentiviral vector to over-express DYRK1A for 48hs (short-term) and 21 days (long-term). Using live-cell imaging we recorded APP-loaded vesicles in axons and analyzed transport dynamics using custom-made MATLAB routines. Results: Short-term harmine treatment reduced axonal APP vesicles density, due to a reduction in non-stationary particles. Contrarily, DYRK1A over-expression enhanced axonal APP density, due to an increase in stationary and retrograde particles. Moreover, long-term DYRK1A over-expression, revealed differential changes in APP dynamics compared with short-term over-expression. Conclusions: Taken together our results suggest that DYRK1A has a relevant role in the regulation of axonal transport and sub-cellular positioning of APP vesicles. Therefore, our work shed light on the role of DYRK1A on axonal transport regulation, and the putative role of harmine restoring axonal transport impairments.

P153 / #535


EFFECT OF E2027, A NOVEL PHOSPHODIESTERASE-9 INHIBITOR, ON COGNITIVE FUNCTION AND HIPPOCAMPAL CYCLIC GMP IN TG2576 MOUSE MODEL OF ALZHEIMER’S DISEASE.

Lecture Title:

Y. Goto, S. Kotani, N. Watanabe, T. Fukushima Eisai Co., Ltd., Neurology Business Group, Tsukuba, Japan

Aims: Phosphodiesterase (PDE) 9 is a cyclic guanosine monophosphate (cGMP) degrading enzyme and highly expressed in the brain. Thus, PDE9 inhibitor increases cGMP, which is involved in synaptic plasticity and cognitive function, and it is expected to be a target for memory improvement. The purpose of this study was to examine the effects of E2027 on cognitive impairment in contextual fear conditioning and cGMP level in Tg2576 mouse model of AD. Methods: E2027 (3 and 10 mg/kg) or vehicle (10 mL/kg) was orally administrated to the aged Tg2576 mice and vehicle was orally administrated to the wild-type mice. Then, cognitive function was evaluated by contextual fear conditioning task. After 8-day washout period, E2027 (3 and 10 mg/kg) or vehicle (10 mL/kg) was orally administrated to the same individuals to the same dosing design as contextual fear conditioning. After that, the heads of the mice were focally irradiated by microwave applicator under anesthesia and the hippocampus was isolated to measure cGMP level. Results: E2027 treatment at 10 mg/kg showed significantly longer freezing behavior in contextual fear conditioning test and significantly improved the reduced cGMP level in the hippocampus in aged Tg2576 mice. Conclusions: Amyloid deposition in aged Tg2576 mice is associated with reduced cGMP and impaired contextual fear conditioning, which are improved by E2027.

P154 / #891


TREATMENT OF APPSL TRANSGENIC MICE WITH AN ALDH2 ACTIVATOR AS A PROMISING TREATMENT OPTION FOR ALZHEIMER’S DISEASE

Lecture Title:

B. Hinteregger1,2, S. Flunkert2, T. Madl1, J. Neddens2, W. Yang3, B. Hutter-Paier2 1Gottfried Schatz Research Center, Medical University of Graz, Division Of Molecular Biology And Biochemistry, Graz, Austria, 2QPS Austria GmbH, Neuropharmacology, Grambach, Austria, 3Foresee Pharmaceuticals Co, Ltd., Foresee Pharmaceuticals, Taipei, Taiwan

Aims: It is known that pathological alternations in AD brains can cause oxidative imbalance which further lead to the generation of toxic aldehydes, resulting in an accumulation of highly reactive, toxic intermediates in the brain and blood. Therefore, the detoxification by enzymes such as aldehyde dehydrogenase 2 (ALDH2) which are able to enhance the catalytic activity of ALDH2, lead to an increased detoxification and are crucial to protect cells and tissues from damage. Thus, the aim of this study was to evaluate the effects of an ALDH2 agonist on AD disease progression in APPSL mice. Methods: In this study, male transgenic APPSL mice and non-transgenic littermates received either an ALDH2 activator or vehicle via the drinking water for the duration of four months. Behavioral tests, immunohistochemistry, biochemical analyses as well as untargeted NMR-based metabolic phenotyping of all mice were performed. Results: In vivo results revealed a highly significant improvement in spatial learning in APPSL + AD-9308 compared to APPSL animals. Furthermore, histological analyses of the brains were performed and metabolic changes in serum and brain regions like hippocampus, cerebellum and cortex analyzed by biochemical assays or NMR-based metabolic phenotyping will be shown. The results provide a first insight in the value of this ALDH2 activator for the treatment of AD. Conclusions: By performing behavioral studies, immunofluorescent labeling and NMR-based metabolic phenotyping, we show that the ALDH2 activator can ameliorate AD symptoms in the APPSL transgenic mouse model and thus could be an effective drug against AD.

P155 / #1674


NEURONAL FARNESYLTRANSFERASE KNOCKOUT RESTORES COGNITIVE FUNCTION AND AMELIORATES ALZHEIMER’S NEUROPATHOLOGY

Lecture Title:

A. Jeong1, S. Cheng1, R. Zhong1, W. Qu2, L. Li1 1University of Minnesota, Experimental And Clinical Pharmacology, Minneapolis, United States of America, 2University of Minnesota, Graduate Program In Neuroscience, Minneapolis, United States of America

Aims: Alzheimer’s disease (AD) is the leading cause of age-related dementia, yet its pathogenesis is not fully understood. Compelling evidence from studies of human postmortem brain tissue as well as cellular and animal models suggests that protein prenylation, a critical posttranslational lipid modification of proteins including farnesylation and geranylgeranylation, contributes to AD pathogenesis. Specifically, systemic haplodeficiency of farnesyltransferase (FT), responsible for protein farnesylation, and pharmacological inhibition of FT have been shown to mitigate amyloid and tau pathology and behavioral impairment in transgenic mouse models. However, little is known about the specific cellular role of protein farnesylation in AD. This study aims to elucidate the impact of neuronal FT on cognitive function and AD neuropathology. Methods: Transgenic APP/PS1 mice were crossed with FT-floxed and CamK2α-Cre mice to generate forebrain neuron-specific FT knockouts (APP/PS1/nFTKO). At 9-10 months of age, APP/PS1/nFTKO and APP/PS1 littermates were subjected to behavioral tests including the Morris water maze, followed by neuropathology assessments using immunohistochemical staining, ELISA, and immunoblot and RNA-seq analyses for relevant signaling pathways. Results: Neuronal FT knockout attenuated memory deficits and amyloid deposition in APP/PS1/nFTKO compared with APP/PS1 mice. It regulated APP proteolytic processing without affecting its protein expression. The reduction of neuropathology was associated with downregulation of farnesylation-dependent small GTPase signaling. RNA-seq analysis is underway to identify molecular networks underlying the impact of neuronal FT. Conclusions: Our data demonstrate that neuronal protein farnesylation regulates the pathogenic process of AD and identify neuronal FT as a potential therapeutic target for AD.

P156 / #1098


ROLE OF POLO-LIKE KINASE 2 IN THE PATHOGENESIS AND TREATMENT OF ALZHEIMER'S DISEASE

Lecture Title:

L. Martinez-Drudis1, R. Sheta1, R. Pellegrinato1, M. Bérard1, S. Loukili1, F. Calon2, S. Rivest1, A. Oueslati1 1Université Laval, Department Of Molecular Medicine & Chu De Quebec Research Center, Québec, Canada, 2Université Laval, Faculty Of Pharmacy & Chu De Quebec Research Center, Québec, Canada

Aims: Increasing evidence suggest that phosphorylation plays an important role in the aggregation and toxicity of amyloid beta (Aβ) resulting from amyloid precursor protein (APP) cleavage and tau, the major neuropathological hallmarks of Alzheimer’s disease (AD). Our laboratory has reported a dramatic accumulation of Polo-like kinase 2 (PLK2) in the brains of AD patients. This observation, in association with the recent notion of a direct interaction between PLK2 and APP, suggests that the aberrant accumulation and activity of PLK2 may contribute to AD. Our goal is focused on examining the effect of PLK2 pharmacological inhibition on APP and tau accumulation and toxicity in cells and transgenic mouse models of AD. Methods: HEK293T cells were used to examine the effect of PLK2 and its inhibition on APP and Tau protein levels by immunoblotting. In vivo, behavioral analysis incorporated evaluation of different learning and memory task. Biochemical and histological analysis of AD neuropathology (APP, tau, their phosphorylated forms, and synaptic dysfunction) were performed using immunoblotting and immunohistochemistry. Results: We observed that PLK2 overexpression decreases APP and tau levels in a PLK2-concentration dependent manner, counteracted by PLK2 pharmacological inhibition in cells. In vivo, our results showed cognitive decline and AD hallmarks in symptomatic mice, as well as a decrease in some aspects of APP and tau pathology upon PLK2 inhibition both at the behavioral and molecular levels. Conclusions: Overall, this project will shed light onto novel mechanisms by which phosphorylation regulates tau and APP aggregation and toxicity, providing a novel therapeutic target for AD and related dementia.

P157 / #1160


KINOME SCREEN IDENTIFIES APP PROTEOLYTIC MODIFIERS AS POTENTIAL ALZHEIMER'S DISEASE DRUG TARGETS

Lecture Title:

J. Wiley1, J. Annis2, J. Gockley3, A. Greenwood1, B. Logsdon1, L. Mangravite1, H. Frankowski2, M. Bothwell4, J. Young5 1Sage Bionetworks, Neurodegeneration Research, SEATTLE, United States of America, 2University of Washington, Institute For Stem Cell And Regenerative Medicine, Seattle, United States of America, 3Sage Bionetworks, Neurodegeneration Research, Seattle, United States of America, 4University of Washington, Physiology And Biophysics, Seattle, United States of America, 5University of Washington, Laboratory Medicine And Pathology, Seattle, United States of America

Aims: We screened the entire kinome to identify kinases that regulate APP proteolytic processing as potential new AD therapeutic targets. We selected kinases that increase overall levels of APP proteolysis to combat 'loss of function' aspects of APP processing. Pharmacological inhibitors for the lead candidates were employed in human AD iPSC derived neurons to assess their impact on AD pathology. Methods: Gamma-secretase reporter cells generated from the N2A neuroblastoma line were interrogated in an siRNA kinome-wide knockdown high-throughput assay. The reporter line employs an APP-Gal4VP16 driver construct and a Gal4-luciferase reporter to monitor gamma-secretase mediated cleavage of APP. The reporter is activated once APP is cleaved by gamma-secretase, allowing the APP AICD-Gal4VP16 moiety to migrate to the nucleus and transactivate the Gal4luciferase reporter. Pharmaceutical inhibitors for Axl and Cerk, two of the primary hits identified in the screen, where tested in human PS1delta9 and APPswe iPSC derived neurons to assess their impact on amyloid and tau production. Results: We identified 45 kinases that regulate APP proteolysis in an upward (n=21) or downward (n=24) direction at statistically significant levels. Our goal was to identify kinases whose targeting up-regulates APP proteolysis in a non-pathogenic direction, so we selected two of the most robust negative regulators--Axl and Cerk--for exploration in human iPSC derived AD neurons. Treatment of human AD neurons with Axl or Cerk inhibitors decreased pathological amyloid and tau production. Conclusions: Identification of APP proteolytic modifiers may enable amelioration of amyloid and tau pathology and elucidate potential AD drug targets.

P158 / #950

Topic: Theme A: β-Amyloid Diseases / A2.e. Therapeutic Targets, Mechanisms for Treatment: Neurotransmitters & receptor-based

RILUZOLE ATTENTUATES SPATIAL LEARNING AND MEMORY IMPAIREMENTS IN FEMALE TRANSGENIC ALZHEIMER’S DISEASE MICE

Lecture Title:

C. Findley1, S. Mcfadden2, L. Sime2, K. Hascup3, E. Hascup1 1Southern Illinois Univ. School of Medicine, Neurology, Cadrd, And Pharmacology, Springfield, United States of America, 2Southern Illinois Univ. School of Medicine, Neurology, Cadrd, Springfield, United States of America, 3Southern Illinois Univ. School of Medicine, Neurology, Cadrd, Pharmacology, And Mmicb, Springfield, United States of America

Aims: Women are almost twice as likely to develop Alzheimer’s disease (AD) compared to men (≥65 years of age). Yet, previous studies on AD interventions have primarily focused on males. This study aimed to determine the long-term impact of a glutamatergic modulator (riluzole) treatment on peripheral glucose metabolism and cognition. Methods: Female C57BL/6 (normal aging; n= 15-16), AβPP/PS1 (transgenic; n= 2-8), and APPNL-F/NL-F

(knock-in; n= 18-19) mice received either vehicle (1% sucrose) or Riluzole (356 µM) treated drinking water (voluntary oral administration) between 2-6 months of age. At 12 months, all mice underwent insulin (ITT) and glucose tolerance testing (GTT) to examine metabolic function and Morris water maze (MWM) to determine cognitive ability. Results: Preliminary results indicated no differences in insulin sensitivity and glucose metabolism between genotypes or treatment. Additionally, no differences were observed in spatial learning (MWM). Riluzole-treated AβPP/PS1 mice showed trending improvement of impaired long-term spatial memory compared to vehicle-treated AβPP/PS1 mice (MWM: annulus 40 cumulative distance, percent time, and entries per distance swam). Vehicle-treated APPNL-F/NL-F mice did not display cognitive decline at this time point compared to treatment-matched C57BL/6 mice, nor responsiveness to riluzole treatment. Conclusions: Prodromal glutamatergic modulation may be an effective preventative strategy for cognitive decline in females for familial AD models. Studies are ongoing to evaluate sex- and age-related differences in response to riluzole treatment.

P159 / #1115

Topic: Theme A: β-Amyloid Diseases / A2.e. Therapeutic Targets, Mechanisms for Treatment: Neurotransmitters & receptor-based

POSSIBLE ROLE OF GHRELIN IN NEURONAL CONDUCTION IN IMPROVED MEMORY COGNITION

Lecture Title:

R. Kumar shoolini university, Pharmacology, solan, India

Aims: The study aims to investigate the role of Ghrelin signaling in memory cognition and retention using intracerebroventricular streptozotocin (ICV-STZ) model in Wistar rats. Methods: Male Wistar rats (250-280 g) were employed into the study with sample size n=6. Streptozotocin (STZ) 3 mg/kg was administered on day one through intra-cerebroventricular route. n-Octanoic acid, a Ghrelin activator was given in two doses 50 and 100uL for 21 daysby dissolving in polyethylene glycol 600whereas Nifedipine, Ca++ channel blocker was given at 10 mg/kg, i.p. to one group underwent to administration of n-octanoic acid high dose.Impairments in cognition, memory consolidation and retention were assessed using Morris water maze, Y maze, balance beam, open field test and photoactometer test. The biochemical estimations for oxidative stress i.e. lipid peroxidation, glutathione, and acetylcholinesterase activity were done in rat brain homogenate. Statistical analysis was carried out using graph pad prism 5. Results: ICV-STZ treated animals exhibited memory deficits in Morris water maze, Y maze, balance beam test. Administration of low and high doses of n-Octanoic acid produced significant restoration of memory retention. However, nifedipine abolished the memory improvement produced by n-octanoic acid. The level of oxidative stress and AChE activity observed in rat brain was also reversed. Conclusions: The finding may reveal that ghrelin plays pivotal role in improving cognition, retentionand working memory possible through Ca signaling.

P160 / #734

Topic: Theme A: β-Amyloid Diseases / A2.f. Therapeutic Targets, Mechanisms for Treatment: ApoE & lipoprotein-based

CONNECTION BETWEEN APOLIPOPROTEIN E AND THE PROGRESSION OF ALZHEIMER'S DISEASE AND CANNABINOIDS AS A THERAPEUTIC METHOD, A SYSTEMATIC REVIEW.

Lecture Title:

A. Gavioli1, A.C. Ortiz2, S.R. Ortiz3 1Universidade São Judas Tadeu, Pós Graduação, São Paulo, Brazil, 2Faculdade de Medicina do ABC, Centro Universitário Saúde ABC, Medicine, Santo André, Brazil, 3São Judas Tadeu University, Postgraduate Program In Aging Sciences, São Paulo, Brazil

Aims: Alzheimer's disease (AD) is the most common cause of dementia and is characterized by progressive loss of memory and cognition, especially in the elderly population. The most common genetic risk factor for late-onset AD is related to an ε4 variant of the Apolipoprotein gene E (ApoE). ApoE4 can lead to neural hyperactivity and increase pathology, leading to neurodegeneration due to the accumulation of beta amyloid and TAU clearance. As a therapeutic method, cannabinoids (CB) have been successfully inhibiting the formation of beta amyloid plaques, on account of its anti-inflammatory and antioxidant role. The present systematic review aims to analyze the studies linking the therapeutic function of CBs with the deposit of beta amyloid related to the ApoE gene. Methods: The papers for the review were selected from PubMed. Articles containing the therapeutic effects of CBs related to AD and ApoE were taken into consideration. Results: Until now, 57 articles have been selected describing the therapeutic effects of CBs in AD. However, there is still no consensus on the dose that should be administered in order to provide better quality of life without adverse effects. Conclusions: The literature is still scarce and therefore inconclusive; however, studies present great prospect for the treatment of AD with the use of CBs due to its neuroprotective role, preventing the accumulation of beta amyloid plaques, even with its accelerated production because of the presence of the ApoE gene.

P161 / #1042


GENERATION OF BLOOD–BRAIN BARRIER FUNCTION FROM IPSCS HARBORING MUTATIONS ASSOCIATED WITH NEURODEGENERATIVE DISORDERS

Lecture Title:

D. Rajesh1, M. Ohshima1, M. Goedland2, C. Munn3, E. Arndt3 1FujiFilm Cellular Dynamics International, R&d Life Science, Madison, United States of America, 2Fujifilm Cellular Dynamics International, R &d Life Science, Madison, United States of America, 3Fujifilm Cellular Dynamics Inc., R&d Life Science, Madison, United States of America

Aims: Objective: Recent GWAS studies have shown that genetically inherited variants apolipoprotein E4 (APOE4) is a genetic risk factor for Alzheimer’s disease (AD) and is also associated with other forms of neuropathology. APOE4 is thought to compromise the blood-brain barrier (BBB), but the underlying mechanisms is unclear. In vitro BBB generated by human induced pluripotent stem cells (iPSCs) derived brain microvascular endothelial cells (BMECs), pericytes and astrocytes can be used to develop a BBB model to understand the causative relationship between APOE4/4 and impairment of BBB function. Methods: Episomally-derived iPSCs from two apparently healthy normal (AHN) donors and two donors harboring the APOE4/4 genotype were successfully differentiated to end-stage BMECs and pericytes using a novel defined differentiation protocol. These cells were characterized by flow cytometry, immunohistochemistry and qPCR analysis. End-stage BMECs and pericytes retained a high purity and are cryopreservable. Results: End-stage BMECs expressed high levels CD31, Glut-1, P-glycoprotein,, Z0-1, Claudin-5 and displayed transendothelial electrical resistance (TEER) over multiple days. End-stage pericytes expressed >90% PGDFR-β+/NG2+/CD13+/CD146+ along with the co-expression of Desmin+/DLK1+/αSMA+ and absence of CD274- and VCAM1, which are-associated with a contractile arteriolar subtype of pericytes. Pericytes exhibited phagocytosis of pHrodo-labeled S. aureus bioparticles and displayed enhancement of barrier function when co-cultured in the presence of isogenic BMECs. Conclusions: Conclusion: The successful generation of a panel of cryopreserved normal and disease associated BMECs and pericytes can be used to generate BBB in vitro to investigate the role of AOPE4/4 containing BMECs and pericytes and BBB dysfunction

P162 / #557


APOLIPOPROTEIN E2 AFFECTS THE BRAIN ENDOSOME AND EXOSOME SYSTEM DURING AGING

Lecture Title:

K. Peng1,2, B. Liemisa2, S. Penikalapati2, E. Levy1,2,3,4, P. Mathews1,2,4 1New York University School of Medicine, Psychiatry, New York, United States of America, 2Nathan S. Kline Institute, Center For Dementia Research, Orangeburg, United States of America, 3New York University School of Medicine, Biochemistry & Molecular Pharmacology, New York, United States of America, 4New York University School of Medicine, Neuroscience Institute, New York, United States of America

Aims: The polymorphic apolipoprotein E (APOE) gene is the greatest genetic determinant of sporadic Alzheimer’s disease (AD) risk: the APOE4 allele increases risk while the APOE2 allele is neuroprotective compared with the common risk-neutral APOE3 allele. We investigated APOE2’s impact on brain exosomal and endosomal systems. Methods: We performed pathway analyses following RNA sequencing of 12-month-old targeted-replacement APOE2 and APOE3 mice brains. Extracellular vesicles (EVs) were isolated from APOE2 and APOE3 mice brains at 6, 12, and 18 months of age. Exosome levels within EV-containing fractions were quantified using nanoparticle tracking and Western blot analyses. Following subcellular fractionation of 18-month-old APOE2 and APOE3 mice brains, early endosome levels were measured using Western blot analysis. Results: Pathway analyses revealed differential regulation of brain endosomal pathways in 12-month-old APOE2 vs. APOE3 mice. Brain exosome levels were higher in 18-month-old APOE2 vs. APOE3 mice while remaining unchanged at younger ages. Early endosome levels were lower in endosome-containing subcellular fractions prepared from 18-month-old APOE2 vs. APOE3 mice brains. Conclusions: Our findings revealed an aging-dependent APOE2-driven increase in brain exosome number and decrease in early endosome levels compared with APOE3 mice. We previously showed that impaired exosome production leads to detrimental accumulation of early endosomes, contributing to age-related APOE4-driven neuronal vulnerability. In contrast, our findings revealed that APOE2 supports clearance of endocytic pathways through robust exosome production. Given that functional interdependent endosomal and exosomal pathways are essential for catabolic cellular processes, we propose that APOE2 exerts its neuroprotective effects in part through the endosomal-exosomal system.

P163 / #793


HUMAN RECOMBINANT APOA-I-MILANO TREATMENT MOBILIZES CEREBRAL ABETA AND IMPROVES COGNITION IN OLD APP23 MICE

Lecture Title:

M. Solé, P. Marazuela, A. Bonaterra, M. Hernández-Guillamon VALL D'HEBRON RESEARCH INSTITUTE, Neurovascular Diseases, BARCELONA, Spain

Aims: To evaluate the therapeutic effect of recombinant Apolipoprotein A-I carrying the genetic Milano variant (rApoA-I-M) on an experimental in vivo model with established AD pathology. Methods: Twenty-one months old APP23 male mice were treated for 8 weeks with human rApoA-I-M or saline, and compared with wild-type (wt) mice. Working memory was evaluated through the T-Maze test. Aβ presence was analyzed in brain, plasma and CSF. NeuN immunohistochemistry was used to evaluate neuronal loss. Markers of vascular function and inflammation were studied in vascular-enriched brain fractions and brain lysates. Results: Saline-treated APP23 mice displayed increased exploration time in the T-maze compared to wt mice. However, rApoA-I-M treatment restored this alteration, indicating a therapeutic impact in this working memory paradigm. The chronic treatment with rApoA-I-M also recovered the neuronal loss observed in saline-treated APP23 mice in the polymorphic layer of the Dentate Gyrus. Although no differences were observed in number or size of parenchymal Aβ-positive deposits, the soluble brain Aβ40 was significantly decreased in rApoA-I-M-treated mice. Furthermore, rApoA-I-M treatment induced an increase of Aβ levels in CSF, confirming its ability to mobilize brain Aβ from the periphery. Vascular levels of occludin and ICAM-1 were significantly increased in rApoA-I-M-treated brains, while P-selectin levels were decreased. In addition, plasma soluble RAGE was remarkably elevated in rApoA-I-M-treated mice, which drastically decreased the vascular damage marker AGEs/sRAGE ratio. Conclusions: The treatment with human rApoA-I-M is safe and may have beneficial effects to prevent the cognitive decline associated with AD through mechanisms involving brain Aβ mobilization and restoring cerebrovascular function.

P164 / #1676


HDL-MIMETIC PEPTIDE 4F ATTENUATES NEUROPATHOLOGY AND COGNITIVE DEFICIT IN A TRANSGENIC MOUSE MODEL OF CEREBRAL AMYLOID ANGIOPATHY AND AD

Lecture Title:

R. Zhong1, D. Chernick2, D. Hottman1, L. Li1,2 1University of Minnesota, Department Of Experimental And Clinical Pharmacology, Minneapolis, United States of America, 2University of Minnesota, Department Of Pharmacology, Minneapolis, United States of America

Aims: Cerebral amyloid angiopathy (CAA), one of the pathological hallmarks of Alzheimer’s disease (AD), features amyloid-β (Aβ) deposition in the cerebral vasculature. Substantial evidence from clinical and animal studies has shown that elevated levels of high-density lipoprotein (HDL), and its main protein component, apoA-I, are associated with reduced CAA and superior cognitive function. The present study aims to investigate whether acute or chronic treatment with the HDL mimetic peptide 4F modulates cerebral amyloid angiopathy (CAA) and associated cognitive deficits and neuropathologies in the transgenic APPSwDI mouse model of CAA/AD. Methods: APPSwDI mice (C57BL/6-Tg(Thy1-APPSwDutIowa)BWevn/Mmjax), an established model of CAA/AD, were treated with intraperitoneal injections of D-4F, the D-isomer of 4F that exhibits a longer half-life. Two cohorts of age- and sex-matched APPSwDI mice received either a 1-week (acute) or 12-week (chronic) daily treatments of D-4F or vehicle (PBS), respectively. Behavioral function was assessed by tests in the open field, the elevated plus maze, and the Morris water maze, followed by (immuno)histochemistry and immunoblotting analysis to assess CAA/AD pathology. Student’s t-test was used to compare the differences between groups. Results: In the acute treatment study, soluble Aβ levels were significantly reduced in the brain of D-4F treated APPSwDI mice; in the chronic study, D-4F treatment attenuated parenchymal amyloid deposition and CAA, neuroinflammation, and memory deficits. Additional analyses are underway to unravel the molecular mechanisms underlying D-4F treatment effects in APPSwDI mice. Conclusions: Our findings suggest that HDL-memetic peptides could be a potential therapeutic agent to mitigate CAA/AD.

P165 / #1721

Topic: Theme A: β-Amyloid Diseases / A2.g. Therapeutic Targets, Mechanisms for Treatment: Anti-inflammatory

SPATIAL TRANSCRIPTOMICS AND IN SITU SEQUENCING TO STUDY ALZHEIMER’S DISEASE

Lecture Title:

W.-T. Chen1, A. Lu1, K. Craessaerts1, B. Pavie1, C. Frigerio1, N. Corthout1, X. Qian2, J. Lalakova2, M. Kühnemund2, I. Voytyuk1, L. Wolfs1, R. Mancuso1, E. Salta1, S. Balusu1, A. Snellinx1, S. Munck1, A. Jurek3, J. Navarro3, T. Saido4, I. Huitinga5, J. Lundeberg6, M. Fiers1, B. De Strooper1 1VIB-KU Leuven, vib-ku Leuven Center For Brain & Disease Research, Leuven, Belgium, 2Science for Life Laboratory, Department Of Biochemistry And Biophysics, Stockholm, Sweden, 3Science for Life Laboratory, School Of Biotechnology, Stockholm, Sweden, 4RIKEN, Center For Brain Science, Wako-shi, Japan, 5Netherlands Institute for Neuroscience, Netherlands Brain Bank, Amsterdam, Netherlands, 6KTH Royal Institute of Technology, Dept Gene Technology, Stockholm, Sweden

Aims: Although complex inflammatory-like alterations are observed around the amyloid plaques of Alzheimer’s disease (AD), little is known about the molecular changes and cellular interactions that characterize this response. We speculate that Aβ accumulation might trigger a multicellular gene regulatory network, which propagates the pathology and drives the disease progression. We aim to identify the molecules of plaque-induced netoworks at cellular levels. Methods: We investigate the transcriptional changes occurring in tissue domains of mice brains in a 100 μm diameter around amyloid plaques via spatial transcriptomics. We identify 12 gene regulatory networks co-expressed across 10327 tissue domains at 3-, 6-, 12-, and 18-months of age in wild-type and AD mouse models. However, the spatial transcriptomics does not provide single cell resolution. We apply an orthogonal technology, in situ sequencing, to confirm the observed alterations at the cellular level on mouse and human brain sections. Results: We demonstrate early alterations in a gene co-expression network enriched for myelin and oligodendrocyte genes (OLIGs), whereas a multicellular gene co-expression network of plaque-induced genes (PIGs) involving the complement system, oxidative stress, lysosomes, and inflammation is prominent in the later phase of the disease. Conclusions: Genome-wide spatial transcriptomics analysis provides an unprecedented approach to untangle the dysregulated cellular network in the vicinity of pathogenic hallmarks of AD and other brain diseases.

P166 / #905


NEUROPROTECTIVE AGENTS FROM FUNGI: A FUTURE OPPORTUNITY FOR TREATMENT OF ALZHEIMER’S DISEASE?

Lecture Title:

V.T.T. Nguyen1, G. Erkel2, K. Endres1 1University Medical Center of the Johannes Gutenberg University, Department Of Psychiatry And Psychotherapy, Mainz, Germany, 2University of Kaiserslautern, Molecular Biotechnology & Systems Biology, Kaiserslautern, Germany

Aims: Currently only drugs for the treatment of the symptoms of Alzheimer’s dementia are clinically available. This neurodegenerative disease is characterized by damage of the nerve cells and thus the loss of cognitive, motor or sensory abilities. In the search for agents with neuroprotective properties which counterbalance the molecular and biochemical events following impairment, exploring natural resources like fungi are promising. Methods: In a screening, 186 fungal extracts were tested for different properties: toxicity, anti-inflammatory potential, anti-oxidative function and protection against Amyloid-beta (A-beta) challenge. For this, initially the monocytic cell line THP-1 and the neuroblastoma cell line SH-SY5Y, both of human origin, were used. Subsequently, promising candidates were tested in different reporter strains of C. elegans and ex vivo in organotypic brain slice cultures. Results: We identified a small number of promising crude extracts with anti-inflammatory and neuroprotective activity. In addition, single extracts were also able to inhibit Amyloid-beta aggregation and to counterbalance the effect of this neurotoxic peptide. It was investigated whether the in vitro results can be transferred to more complex biological systems. Conclusions: In long term these experiments might lead to finding active substances which could serve as a drug for neural cell survival and re-establishment of normal function of the brain, which could be applied for the treatment of patients with Alzheimer’s disease.

P167 / #447


STIMULATION OF TNF RECEPTOR 2 AS POTENTIAL THERAPY FOR ALZHEIMER'S DISEASE

Lecture Title:

N. Orti-Casañ1, H. Wajant2, I. Zuhorn3, P. De Deyn4, P. Naudé4, U. Eisel1 1University of Groningen, Molecular Neurobiology, Groningen, Netherlands, 2University Hospital Würzburg, Molecular Internal Medicine, Würzburg, Germany, 3University of Groningen, Biomedical Ingeneering, Groningen, Netherlands, 4University of Groningen, Neurology And Alzheimer Centre, Groningen, Netherlands

Aims: Our approach consists on treating an APP overexpressing mouse model of Alzheimer’s disease with a TNF Receptor 2 agonist to investigate its effectiveness in AD-related pathology. Methods: Mice were treated for a period of 6 weeks by using two different ways of administration: infusion of the drug into the brain via implantation of osmotic pumps or systemic administration by intraperitoneal injection. Treated and control animals were evaluated in behavioral tests and pathological changes were investigated. Three behavioural tests were performed: elevated-plus maze to measure anxiety-like behaviour, Y-maze to assess working memory and Morris water maze to evaluate spatial memory. Regarding the pathology, changes in Aβ plaques and different neuroinflammation markers were measured. Results: Our results showed significant differences when comparing the treated vs. untreated mice that received a systemic administration of the drug. We observed in the treated mice a significant improvement in both working and spatial memory with no changes in anxiety levels. Nonetheless, mice that received the drug centrally via implantation of osmotic pumps showed no differences in anxiety or working memory. However, both treated and untreated mice in this group showed comparable results to the systemically-injected mice in terms of spatial memory. Conclusions: Treatment with a TNF Receptor 2 agonist is able to improve working and spatial memory in an Alzheimer’s disease context. Moreover, we observed a differential effect depending on the way of administration. These results may provide valuable information to understand and design future potential treatments for Alzheimer’s disease.

P168 / #711


ETHANOLIC EXTRACT OF ZANTHOXYLUM SP. REDUCED TAU BURDEN, NEUROINFLAMMATION, AND INCREASED APOE EXPRESSION IN THE TRIPLE TRANSGENIC MICE MODEL OF ALZHEIMER´S DISEASE

Lecture Title:

J.C. Ruiz-González1, A. Sandoval-Hernández1, G. Arboleda-Bustos2 1Universidad Nacional de Colombia, Facultad De Ciencias - Departamento De Química, Bogotá, Colombia, 2Universidad Nacional de Colombia, Facultad De Medicina, Bogotá, Colombia

Aims: To evaluate the potential beneficial effect of an ethanolic extract from Zanthoxylum sp. in the 3xTg-AD mouse model of AD. Methods: Twelve months old 3xTg-AD mice were treated for 18 weeks with ethanolic extract of Zanthoxylum sp., (25 mg/kg/day). Immunohistochemical changes (AT8, Iba-1, ApoE staining) were analyzed by confocal microscopy. Results were compared with control animals and 3xTg-AD mice treated with vehicle (water). Results: Treated animals showed a reduction in immunoreactivity AT8 antibody, as well as decreased neuroinflammation across all hippocampal regions. These changes were accompanied by increased expression of ApoE, as compared to untreated animals. Conclusions: Zanthoxylum sp. ethanolic extract reverses cognitive impairment in 3xTg-AD mice associated with reduction in neuroinflammation, pathological Tau burden and increased ApoE expression. Acknowledgements: Funded by Colciencias (110177758004 and 202010013184) and DIEB-Universidad Nacional de Colombia.

P169 / #1391


NEAR-INFRARED LIGHT REDUCES GLIAL ACTIVATION IN HIPPOCAMPUS AND CORTEX OF DIET-INDUCED OBESE MICE

Lecture Title:

S. Saieva, A. Fracassi, M. Marcatti, W.-R. Zhang, G. Taglialatela University of Texas Medical Branch, Neurology, Galveston, United States of America

Aims: Obesity is a major risk factor for Alzheimer’s Disease (AD), characterized by systemic insulin resistance and neuroinflammation, this latter prodromal event to neurodegeneration. Hippocampal neuroinflammation, in rodents, correlates with poor memory performance, while in humans, growing evidence shows that obesity increases three times the risk to develop AD. Therefore, any treatment aimed at reducing the impact of obesity on neuroinflammation may delay the onset of dementia, the overarching objective of this work. Our previous works showed that a transcranial delivery of near-infrared (NIR) light reduced Aβ and Tau oligomers in hippocampal synaptosomes and improved memory function in mouse models of AD. Here, we tested whether NIR light prevents obesity-induced glial activation, the triggering event of neuroinflammation. Methods: 5-weeks old wild-type mice were fed a high-fat diet (HFD) for 12 weeks to induce obesity prior the beginning of treatment, which consisted of a transcranial delivery of NIR light for 4 weeks throughout daily sessions of 90 seconds each. After sacrifice, we performed free-floating immunofluorescence of brain slices stained for microglia (Iba1 and CD68) and astrocytes (GFAP) markers to evaluate glial activation. Results: In both hippocampus and cortex, HFD caused increased expression of CD68 and GFAP, which were reversed by NIR light treatment. However, Iba1 was not changed by HFD or NIR. Conclusions: Our results show a reduction of glial activation in both hippocampus and cortex. This evidence poses NIR light as a potential preventive therapeutic approach against obesity-induced CNS deficits that are known to concur to AD neuropathology. Supported by R01AG069433, R01AG060718, R56063405 to GT

P170 / #282


DEVELOPMENT OF NLRP3 INHIBITORS FOR ALZHEIMER'S DISEASE

Lecture Title:

S. Zhang, H. Blevins, J. Green, Y. Xu Virginia Commonwealth University, Medicinal Chemistry, Richmond, United States of America

Aims: NLRP3 inflammasome is an essential component of innate immunity and recent studies have indicated a critical role for the NLRP3 inflammasome in the pathogenesis of Alzheimer’s disease (AD), where neuroinflammation has been recognized as an essential player. Thus, novel NLRP3 inhibitors represent a novel approach to develop AD therapeutics. The aim of this study is to ellucidate the mode of action for the newly developed small molecule NLRP3 inhibitors in our laboratory and develop new inhibitors with CNSA drug-like properties. Methods: Photoaffinity labeling (PAL) probes based on a novel lead NLRP3 inhibitor that exhibits promising neuroprotective activities in AD models were synthesized to understand mode of action. A new chemical scaffold was also designed to develop NLRP3 inhibitors with CNS drug-like properties. The new analogs were charatcerizaed for their inhibitory potency and selectivity both in vitro and in vivo. Results: PAL studies confirmed the selective interaction of the PAL probe with the NLRP3 protein. Binding studies also confirmed the direct interaction of our inhibitor with the NLRP3 protein. We also confirmed that our NLRP3 inhibitor does not inhibit the ATPase activity of NLRP3. The newly design NLRP3 inhibtors exhibited potent and selective inhibittion on the NLRP3 inflammasome both in vitro and in vivo. Conclusions: Chemical biology studies established that our small molecule NLRP3 inhibitors directly bind to the NLRP3 protein without interfering its ATPase activity. The newly designed NLRP3 inhibitors exhibited improved CNS drug-like properties and strongly encourage further development as potential AD therapeutics.

P171 / #517


CELLULAR SIGNALING AND ANTI-APOPTOTIC EFFECTS OF PROLACTIN-RELEASING PEPTIDE AND ITS ANALOG ON SH-SY5Y CELLS

Lecture Title:

A. Zmeškalová, A. Popelová, A. Exnerová, B. Železná, J. Kuneš, L. Maletínská AS, Institute Of Organic Chemistry And Biochemistry, Prague, Czech Republic

Aims: Prolactin-releasing peptide (PrRP), is a neuropeptide with anorexigenic and antidiabetic properties. Because of the suggested link between obesity and/or type 2 diabetes and Alzheimer’s disease (AD) development, anorexigenic and/or antidiabetic compounds were repurposed as potential neuroprotective compounds. Methods: In this study, we focused on the molecular mechanisms of action of natural PrRP31 and its lipidized analog palm11-PrRP31 in the human neuroblastoma cell line SH-SY5Y to describe their cellular signaling and possible anti-apoptotic properties. Furthermore, the potential neuroprotective properties through activation of anti-apoptotic pathways of PrRP31 and palm11-PrRP31 were studied using the SH-SY5Y cell line and rat primary neuronal culture stressed with toxic methylglyoxal (MG). MG is a side-product or glycolysis which promotes the pro-apoptic pathways. The cell viability was measured by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test. Results: Using in vitro models of neurodegeneration, we tested potential beneficial effects of PrRP analog palm11-PrRP31. PrRP31 significantly upregulated the phosphoinositide-3 kinase-protein kinase B/Akt (PI3K-PKB/Akt) and extracellular signal-regulated kinase/cAMP response element-binding protein (ERK-CREB) signaling pathways that promote metabolic cell survival and growth. In addition, we proved via protein kinase inhibitors that activation of signaling pathways is mediated specifically by PrRP31 and its palmitoylated analog. Pre-treatment with natural PrRP, as well as with palm11-PrRP, protected the sh-SY5Y from MG-induced cytotoxic effect using MTT. Conclusions: In summary, lipidized analogs of PrRP seem to be potential neuroprotective agents but the exact mechanism of action must be further studied.

P172 / #488

Topic: Theme A: β-Amyloid Diseases / A2.h. Therapeutic Targets, Mechanisms for Treatment: Anti-oxidants

AMYLOID-ΒETA-INDUCED OXIDATIVE STRESS BOOSTS CERIUM OXIDE NANOPARTICLES UPTAKE BY CHANGING BRAIN ENDOTHELIUM MICROVILLI PATTERN

Lecture Title:

R. Dal Magro1, A. Vitali2, S. Fagioli1, A. Casu3, A. Falqui3, B. Formicola1, L. Taiarol1, U. Anselmi-Tamburini2, P. Sommi4, F. Re1 1University of Milano-Bicocca, Medicine And Surgery, Vedano al Lambro (MB), Italy, 2University of Pavia, Department Of Chemistry, Pavia, Italy, 3King Abdullah University of Science and Technology (KAUST), Biological And Environmental Sciences And Engineering Division, Thuwal, Saudi Arabia, 4University of Pavia, Department Of Molecular Medicine, Pavia, Italy

Aims: Cerebrovascular dysfunctions are a common feature of several neurodegenerative disorders, including Alzheimer’s disease (AD). Amyloid-β peptides contribute to the impairment of the brain vasculature toward the generation of reactive oxygen species (ROS) in endothelial cells, leading to a cascade of events that promotes neurodegeneration. In this scenario, the use of cerium oxide nanoparticles (CNP) as ROS scavenging agents has gained increasing interest. Here we aimed to investigate the ability of CNP to hinder Aβ-induced oxidative stress on human cerebral microvascular endothelial cells (hCMEC/D3) and to elucidate the mechanisms promoting CNP uptake under oxidative stress conditions. Methods: The free radical scavenging activity of CNP after hCMEC/D3 exposure to Aβ oligomers was assessed by measuring ROS/RNS levels in the presence and absence of CNP. The modification of endothelial microvilli pattern after cell exposure to Aβ species was evaluated by SEM analysis and WB. Results: Treatment with CNP restored basal ROS levels in hCMEC/D3 cells both after acute or prolonged exposure to Aβ oligomers. Interestingly, the uptake of CNP increased after cell incubation with Aβ. To gain insight into this result, cell surface modifications were investigated under Aβ treatment. We demonstrated that vascular pro-oxidant stimuli, i.e. Aβ and hydrogen peroxide, induced microvilli-like protrusions on the surface of endothelial cells, which enhance CNP binding to the cell surface. Conclusions: Endothelial microvilli formation under oxidative stress conditions could be exploited to boost the uptake of anti-oxidant nanoparticles at the vascular level as potential therapy for ROS-mediated cerebrovascular dysfunction in brain disorders.

P173 / #845


A MULTIFUNCTIONAL THERANOSTIC PLATFORM COUNTERACTING IRON INDUCED OXYDATIVE STRESSES IN ALZHEIMER’S DISEASE BRAINS

Lecture Title:

E. Ficiarà1, S. Ansari1, F. D'Agata1, R. Cavalli2, C. Guiot1 1University of Torino, Department Of Neuroscience, Torino, Italy, 2University of Torino, Department Of Drug Science And Technology, Torino, Italy

Aims: The etiology of Alzheimer’s disease (AD) is not still completely clear, making current treatments almost ineffective. Since many evidences support a crucial role of iron in neurodegeneration [1], iron is one the main target for innovative therapeutic approaches. Although systemic iron chelator drugs are already under clinical assessment in AD as off label treatment, there is room for optimization at different levels, i.e. by improving personalization (based on accurate measurement of iron concentration in cerebrospinal fluid [2] and serum), drug formulation(safer and nanostructured), and drug-delivery strategies. This work aims at investigating a multifunctional theranostic nanostructured platform to improve therapeutic strategies based on iron chelation. Methods: Physico-chemical characterization of nanobubble structures with an oxygenated perfluorocarb core and a double-decker shell encapsulating superparamagnetic iron oxide nanoparticles (SPIONs) covered by glycol-chitosan combined with chelating agents (EDTA, or Deferoxamine DFO) was performed. Biocompatibility, chelating and magnetic properties were tested. Results: Following characterization(average diameter = 330.2 +/- 2.4 nm; Polydispersity Index =0.27 +/- 0.17; Z potential = -27.5 +/- 4.2 mV) the chelating properties were evaluated (Table).

Formulation Chelated Fe (II) Chelated Fe (III)

MCNBs+EDTA 97.2% 52.3%

MCNBs+DFO 46.2% 89.4%

Conclusions: Due to their biocompatibility, such Magnetic Chelating Nanobubbles (MCNBs) may be delivered either systemically or by intrathecal administration to CSF and monitored via MRI as well as by sonography. Magnetic driving may force the MCNBs across the brain membranes enhancing the effectiveness of the iron chelation process. [1] Ndayisaba et al., Front Neurosci. 2019; 13: 180. [2] Ficiarà E. et al., Abstract 459, 2nd AAT-AD/PD Meeting 2020.

P174 / #1223


MYRICETIN PREVENTS HIGH MOLECULAR WEIGHT AΒ1-42 OLIGOMER-INDUCED NEUROTOXICITY ON MITOCHONDRIA

Lecture Title:

A. Kimura1, Y. Kiuchi2, M. Tsuji3, K. Ono1 1Showa University, Department Of Medicine, Division Of Neurology, Shinagawaku, Japan, 2Showa University, Department Of Pharmacology, Shinagawaku, Japan, 3Showa University, Pharmacology Research Center, Shinagawaku, Japan

Aims: Excessive accumulation of β-amyloid peptide (Aβ) is one of the primary mechanisms that cause neuronal death in Alzheimer’s disease (AD). We’ve previously shown high molecular weight Aβ1-42 oligomer could play a major role. Though many materials have been thought to be protective against Aβ oligomer, the mechanism remains unclear. Myricetin is one of the natural flavonols from fruit, vegetable, tea, and medicinal plants that have been reported to protect against several neurodegenerative diseases such as Parkinson’s and Alzheimer’s. The purpose of this study was to clarify the mechanism of the protective effect of myricetin against the neurotoxicity of Aβ oligomer in SH-SY5Y cells. Methods: We examined myricetin's protective effect on cellular mitochondrial damage by 5 μM Aβ oligomer in vitro, including reactive oxygen species (ROS) generation, MPT(mitochondrial membrane permeability transition), Mitochondrial ROS, MnSOD, and intracellular ATP. Results: Aβ oligomer disturbed mitochondria integrity with an increase in ROS generation, mitochondrial ROS generation, and with decrease in MnSOD, ATP, and MTP. Meanwhile, the treatment with myricetin significantly prevented neuronal damage induced by Aβ oligomer as follows. Myricetin significantly inhibited Aβ oligomer-induced oxidative stress and disruption of mitochondrial membrane integrity. Furthermore, it prevented the Aβ oligomer-induced decreasing MnSOD and ATP and suppressed the decrease of MPT. Conclusions: These results suggest that myricetin has multiple functions to counter the progress of AD by the reduction of the detoxification of Aβ oligomer, and it may be developed as neuroprotective agents against AD.

P175 / #945

Topic: Theme A: β-Amyloid Diseases / A2.i. Therapeutic Targets, Mechanisms for Treatment: Neurotrophic, synaptic plasticity, repair, regenerative medicine

HRG80 : A NEW PREVENTIVE DIET SUPPLEMENT WITH NEUROPROTECTIVE EFFECTS.

Lecture Title:

A. Henriques1, N. Callizot2, P. Poindron2, L. Rouvière2, C. Farrugia2, S. Defrère3, P.-A. Mariage3 1Neuro-Sys, Pharmacology, GARDANNE, France, 2Neuro-Sys, Pharmacology, Gardanne, France, 3Botalys, R&d, Ath, Belgium

Aims: Alzheimer’s disease (AD) cognitive decline is primary due to weaker neuronal plasticity and progressive neuronal loss. The hippocampus plays a major role in neuroplasticity and is a key structure for memory processing and consolidation. In animal models, increase adult neurogenesis is associated to improvement in acquisition, formation and maintenance of memory. AD is characterized by excitotoxic levels of extracellular glutamate, accumulation of soluble Amyloid peptide (Ab) and hyperphosphorylated tau leading to neurodegeneration. Soluble Ab oligomers (AbO) results in synaptic failure, through modifications to the glutamatergic system. Panax ginseng is a plant belonging to the Panax genus that originates from Eastern Asia. Extensive evidence suggests that the roots of Panax ginseng could promote neuroprotection and neuroplasticity. A number of neuroactive compounds have been identified in ginseng extracts and include ginsenosides, ginsan or gintonin. Here, we present the neuroprotective effects of a ginseng extract called HRG80. Methods: Using primary hippocampal/cortical neurons injured with glutamate or AbO, we showed that HRG80 was able to prevent both injuries in a dose dependent manner. Results: A protective effect on neurons and their neurite network was observed but also on the number of synapses. In addition, the chronically treatment of hippocampal neurons significantly increased the maturation and the number of synapses. HRG80 reduced the Tau accumulation into the neurites Conclusions: Altogether, these results are in favor of a preventive effect of ginseng extract against neuronal injuries and a potential effect on the neuronal plasticity promoting the synaptogenesis.

P176 / #1458


BENEFICIAL EFFECT OF POSITIVE MODULATORS OF TRPC6 CHANNELS IN ANIMAL MODELS OF ALZHEIMER’S DISEASE

Lecture Title:

E. Popugaeva1, D. Chernyuk1, N. Zernov1, I. Bezprozvanny1,2 1Peter the Great St. Petersburg Polytechnic University, Medical Physics, St Petersburg, Russian Federation, 2UT Southwestern Medical Center, Physiology, Dallas, United States of America

Aims: Alzheimer’s disease (AD) is the dominant form of elderly dementia. Most AD clinical trials target β-amyloid pathway, but success has been very limited so far. Therefore, there is a need to identify alternative targets for treatment of AD patients. Neuronal store-operated calcium entry (nSOCE) is a fine tuning mechanism that regulates intracellular Ca2+ content. We previously reported that activation of TRPC6-mediated nSOCE in neurons prevented synaptic loss and rescued synaptic plasticity defects in animal models of AD (Zhang at al, 2016, J Neurosci. v 36, pp 11837-11850). The aim: to identify novel positive modulators of TRPC6 channels and evaluate their biological activity in animal models of AD. Methods: bioinformatical screening of existing drugs database, calcium imaging in soma and spines, cell culture model of amyloid synaptotoxicity, confocal imaging of dendritic spines in fixed culture and brain slices, long-term potentiation (LTP) experiments in 5xFAD mouse model of AD. Results: we have defined several potential positive modulators of TRPC6 channels. Some of these compounds demonstrated neuroprotective effects in in vitro and ex vivo models of AD. Conclusions: identified compounds provide a lead for development of potential AD therapeutic that target TRPC6-dependnent nSOCE and rescue synaptic loss. This work was funded by Russian Science Foundation grant No 20-75-10026.

P177 / #1332


THE INFLUENCE OF FINGOLIMOD ON THE GENE EXPRESSION OF SELECTED PRESYNAPTIC PROTEINS IN A MURINE MODEL OF ALZHEIMER'S DISEASE.

Lecture Title:

I. Wieczorek1, P. Wencel1, H. Jesko2, R. Strosznajder1 1Mossakowski Medical Research Centre Polish Academy of Sciences, Laboratory Of Preclinical Research And Environmental Agents, Warsaw, Poland, 2Mossakowski Medical Research Centre Polish Academy of Sciences, Department Of Cellular Signalling, Warsaw, Poland

Aims: The aim of the study was to investigate the changes in gene expression of selected presynaptic proteins in the hippocampus of AD mice and to determine the influence of sphingosine-1-phosphate receptor modulator (fingolimod) on these processes. Methods: The experiments were conducted on the hippocampus of 3- and 12-month-old FVB mice: with overexpression of the human mutant gene of β-amyloid precursor protein (FVB-APP+) and without the transgene (FVB-APP-), which received fingolimod (1 mg/kg) or vehicle (0.9% NaCl) for 2 weeks. The expression of genes encoding the following presynaptic proteins: synaptobrevin-1 (VAMP1), syntaxin-1a (STX1a), synaptosomal-associated protein 25 kDa (SNAP-25), complexin-1 (CPLX1), synaptotagmin-1 (SYT1), synaptophysin-1 (SYP1), and neurexin-1 (NRXN1) was investigated using the qRT-PCR method. Results: A significant reduction in expression of Vamp1 and Syt1 was found in the hippocampus of 3-month-old transgenic mice. Administration of fingolimod had no influence on any gene expression in this age group. In 12-month-old transgenic animals, a significant decrease in Cplx1, Nrxn1, Snap25, Syt1, and Stx1a expression was observed. In contrast to younger mice, the administration of fingolimod to 12-month-old FVB-APP+ animals increased the expression of all genes except Syt1. Conclusions: Changes in gene expression in the hippocampus were observed in both groups, however, they were more common in 12-month-old animals. Administration of fingolimod to FVB-APP+ mice had a beneficial effect, restoring reduced expression of most studied genes to control values (those of FVB-APP-), but it only occurred in older animals. Supported by grant no. NCN 2014/15/B/NZ3/01049 and statutory budget of MMRC, theme no. 7.

P178 / #755

Topic: Theme A: β-Amyloid Diseases / A2.j. Therapeutic Targets, Mechanisms for Treatment: Protein aggregation, misfolding, chaperones

NATURAL PRODUCT BASED PEPTIDES: BRIDGING THE GAP BETWEEN INTRINSICALLY DISORDERED PROTEINS OF DIABETES AND ALZHEIMER’S

Lecture Title:

S. Das1,2, S. Dutta1, R. Paidi3, A. Bej1, S. Biswas3, U. Halder2, D. Bhattacharyya1 1CSIR-INDIAN INSTITUTE OF CHEMICAL BIOLOGY, Structural Biology And Bioinformatics, KOLKATA, India, 2JADAVPUR UNIVERSITY, Chemistry, KOLKATA, India, 3CSIR-INDIAN INSTITUTE OF CHEMICAL BIOLOGY, Cell Biology And Physiology Division, West Bengal, India

Aims: Targeting amyloidal fibrils of Aβ and insulin by natural product based peptides of bromelain and development of therapeutic targets that bridge the gap between diabetes and Alzheimer's Methods: Fruit bromelain peptide pool having Mw<500Da was separated following human digestive protease digestion. Aggregation kinetics of both insulin and Aβ in presence and absence of bromelain peptides was followed using Thioflavin-T assay, transmission electron and atomic force microscopy. Sequence homology was performed and corresponding synthetic peptide using bromelain template showed interaction specificity. FT-IR, circular dichroism spectra and extraneous fluorophore(8-ANS) helped analyse secondary structure and hydrophobic interaction. Ex vivo analysis in SHSY5Y and HepG2 cells was performed using MTT assay and microscopic imaging respectively. Bilateral intracerebroventricular administration of Aβ to cause AD-type neurodegeneration in rats was developed both in control and diabetic models. Results: Bromelain peptides prevent toxic oligomer formation and dissociate preformed amyloid fibrils, thereby demonstrating a changed configuration of dissociated and/or preventive states of insulin and Aβ from their native forms. Current hypothesis indicate possible interaction of peptides at a common hydrophobic region that highlights a close spatial correlation between brain expression implicated in IR and beta-amyloid pathologies. Cytotoxic effect of Aβ was relieved by peptides. Peptide infusion caused plaque dissociation in vivo and showed subsequent physiological and behavioral changes in rats. Conclusions: Fruit bromelain peptides irreversibly prevent Aβ and insulin amyloidogenesis as well as dissociate preformed aggregates in vitro, ex vivo and in vivo. Prevention of insulin aggregation and targeting diabetic patients, thereby act as an early marker in the pathogenesis of AD.

P179 / #253

Topic: Theme A: β-Amyloid Diseases / A2.k. Therapeutic Targets, Mechanisms for Treatment: TREM2

CXCR4-DEPENDENT MIGRATION IS DEFECTIVE IN HUMAN TREM2 KNOCKOUT MICROGLIA

Lecture Title:

A. Mcquade1, Y.J. Kang2, A. Jairaman3, J. Hasselmann1, M. Coburn1, S. Kiani Shabestari4, J.P. Chadarevian1, M. Blurton-Jones1 1University of California, Irvine, Neurobiology And Behavior, Irvine, United States of America, 2University of North Carolina Charlotte, Department Of Mechanical Engineering And Engineering Science, Charlotte, United States of America, 3UCI, Department Of Physiology And Biophysics, Irvine, United States of America, 4Institute for Memory Impairments & Neurological Disorders Sue & Bill Gross Stem Cell Research Center, Department Of Neurobiology & Behavior, Irvine, United States of America

Aims: The discovery of TREM2 as a myeloid-specific Alzheimer’s disease (AD) risk gene has accelerated research into the role of microglia in AD. Recent studies have consistently found that mutations or deletion of TREM2 results in failure to exit homeostatic states despite exposure to disease pathology. While this phenotype has been profiled extensively, the mechanisms that drive this impaired response remain unclear. Understanding these molecular targets will be crucial to the development of therapies which could beneficially re-activate microglia against disease pathology. Methods: To mimic TREM2 loss of function mutations which increase disease-risk, we developed CRISPR-modified TREM2 knockout induced pluripotent stem cell (iPSC) lines. Using a novel chimeric model of AD we examined human microglial activation in vivo via single-cell sequencing and histological analysis. Using these unbiased experiments as a guide, we next assayed migration in vitro to narrow down receptor-ligand pairs which may be responsible for activation deficits seen in vivo. Results: Loss of TREM2 leaves human microglia unable to properly activate towards beta-amyloid plaques in vivo. RNA-sequencing of xenotransplanted microglia highlighted several receptor-ligand pairs as likely candidates for these migration defects, including CXCR4/CXCL12. Using in vitro approaches, we further confirmed that CXCR4 is reduced in TREM2 knockout microglia and necessary for migration towards beta-amyloid producing neural cultures. Conclusions: Taken together, these studies highlight CXCR4 as a potential target to restore migration and activation of TREM2 mutant microglia. Ongoing studies will determine whether in vivo migration and the ‘DMA phenotype’ can be restored through CXCR4 manipulation.

P180 / #1164


TREM2 RESTRAINS THE ENHANCEMENT OF TAU ACCUMULATION AND NEURODEGENERATION BY BETA-AMYLOID PATHOLOGY

Lecture Title:

W. Meilandt1, S.-H. Lee1, L. Xie2, V. Gandham2, H. Ngu3, K. Barck2, J. Imperio1, G. Lalehzadeh1, K. Stark1, O. Foreman3, R. Carano2, M. Sheng4, A. Easton1, C. Bohlen1, D. Hansen5 1GENENTECH, INC., Neuroscience, SOUTH SAN FRANCISCO, United States of America, 2GENENTECH, INC., Biomedical Imaging, SOUTH SAN FRANCISCO, United States of America, 3GENENTECH, INC., Pathology, SOUTH SAN FRANCISCO, United States of America, 4Stanley Center for Psychiatric Research, Broad Institute Of Mit And Harvard, Cambridge, United States of America, 5Brigham Young University, Chemistry, Provo, United States of America

Aims: Loss-of-function TREM2 (triggering receptor expressed on myeloid cells 2) mutations strongly increase Alzheimer’s disease (AD) risk. Deletion of Trem2 has revealed a protective role of Trem2 function in preclinical models of β-amyloid accumulation. How TREM2 influences later AD stages characterized by tau-mediated neurodegeneration is less clear. Here, we aimed to determine the effects of TREM2 deletion on the accumulation and spread of tau pathology in the presence or absence of β-amyloid pathology Methods: To understand Trem2 function in the context of both β-amyloid and tau pathologies, we examined Trem2-deficient mice expressing mutant tau alone (P301L, pR5-183 model) or in the TauPS2APP model, in which β-amyloid pathology exacerbates tau pathology and neurodegeneration. Changes in disease-associated pathology and neurodegeneration were measured by Western blot and Immunohistochemistry in TauP301L mice (6, 10, and 18 months) and in TauPS2APP mice (9, and 17 months) with wild-type, heterozygous and homozygous deletions of Trem2. Changes is brain atrophy were measured longitudinally by (v)olumetric MRI. Results: In the presence of β-amyloid pathology, Trem2 deletion further exacerbated tau accumulation within neuronal cell bodies and processes, including dystrophic neurites. We also found increased tau spreading and hyperphosphorylation of endogenous mouse tau especially in the hippocampus. Brain atrophy was also significantly elevated in Trem2-deficent TauPS2APP mice measured by degenerative staining and vMRI. Without β-amyloid pathology, Trem2 deletion did not affect these processes. Conclusions: TREM2 may slow AD progression and reduce tau-driven neurodegeneration by restricting the degree to which β-amyloid facilitates the spreading of pathogenic tau.

P181 / #643


ANTI-HUMAN TREM2 INDUCES MICROGLIA PROLIFERATION AND REDUCES PATHOLOGY IN AN ALZHEIMER’S DISEASE MODEL

Lecture Title:

S. Wang1, M. Mustafa2, C. Yuede3, S. Salazar2, P. Kong2, H. Long2, M. Ward2, O. Siddiqui2, R. Paul2, S. Gilfillan1, A. Ibrahim2, H. Rhinn4, I. Tassi2, A. Rosenthal2, T. Schwabe2, M. Colonna1 1Washington University School of Medicine in St Louis, Pathology And Immunology, saint louis, United States of America, 2Alector LLC, Alector Llc, South San Francisco, United States of America, 3Washington University School of Medicine in St Louis, Psychiatry And Neurology, saint louis, United States of America, 4Alector, Inc, Bioinformatics, South San Francisco, United States of America

Aims: TREM2 is a receptor for lipids expressed in microglia. The R47H variant of human TREM2 impairs ligand binding and increases Alzheimer’ s disease (AD) risk. In mouse models of amyloid β (Aβ ) accumulation, defective TREM2 function affects microglial response to Aβ plaques, exacerbating tissue damage, whereas TREM2 overexpression attenuates pathology. Thus, AD may benefit from TREM2 activation. Methods: Here, we examined the impact of an anti-human TREM2 agonistic mAb, AL002c, in a mouse ADmodel expressing either the common variant (CV) or the R47H variant of TREM2. Results: Single-cell RNA-seq of microglia after acute systemic administration of AL002c showed induction of proliferation in both CV- and R47H-transgenic mice. Prolonged administration of AL002c reduced filamentous plaques and neurite dystrophy, impacted behavior, and tempered microglial inflammatory response. Conclusions: We conclude that AL002 is a promising candidate for AD therapy.

P182 / #1243


PK/PD MODEL OF THE EFFECTS OF AL002, AN ANTI-TREM2 ANTIBODY, FROM A FIRST-IN-HUMAN PHASE 1 STUDY

Lecture Title:

M. Ward1, R. Paul1, F. Yeh1, H. Long1, M. Spencer1, T. Schwabe1, S. Kathman2, C. Hines2 1Alector LLC, Alector Llc, South San Francisco, United States of America, 2PPD, Pk/pd, Wilmington, United States of America

Aims: AL002 is a monoclonal IgG1 antibody that activates the triggering receptor expressed on myeloid cells -2 (TREM2) pathway on microglia and is being developed by Alector for the treatment of Alzheimer’s disease. A semi-mechanistic exposure-response model was developed using data from the AL002 first-in-human Phase 1 study AL002-1 (INVOKE). Methods: INVOKE is a multi-site, first-in-human, Phase 1 study in which healthy volunteers (HV) received single-dose (SD) IV AL002. HV data from this study was used to build an exposure-response model combining a population pharmacokinetic (PK) model for AL002 in serum and CSF with a population pharmacodynamic (PD) model for soluble TREM2 (sTREM2) in CSF. The population PK model for AL002 in serum was a two-compartment model. A third compartment was added for AL002 concentration in the CSF. A turnover model was utilized for CSF sTREM2. Results: Based on simulation results from the model, dosing every 4 weeks with low, medium and high IV doses of AL002 is expected to lead to decreased levels of CSF sTREM2. Reductions in CSF sTREM2 are similar with the medium and high doses, suggesting that a maximum effect in HVs is being reached at these levels. Conclusions: A semi-mechanistic exposure-response model of AL002 on sTREM2 was developed using INVOKE study data. Diagnostic plots, including visual predictive checks, show the model was predictive of the data. The model was used to identify an intravenous (IV) regimen for a Phase 2 study that is predicted to activate microglia TREM2 signaling for sustained treatment periods.

P183 / #541

Topic: Theme A: β-Amyloid Diseases / A2.m. Therapeutic Targets, Mechanisms for Treatment: Microglia

TARGETING THE MOESIN-CD44 PATHWAY FOR THERAPEUTIC INTERVENTION IN ALZHEIMER’S DISEASE

Lecture Title:

V. Katis1, W. Bradshaw1, R. Betarbet2, Y. Du3, K. Qian3, C. Jimenez Antunez1, C. Mulhern4, B. Slakman4, Y. Zhang5, J. Guilinger5, A. Keefe5, C. Huguet5, M.-A. Guie5, N. Seyfried2, H. Fu3, A. Levey2, L. Mangravite6, P. Brennan7, O. Gileadi1 1University of Oxford, Structural Genomics Consortium, Oxford, United Kingdom, 2Emory University School of Medicine, Department Of Neurology, Atlanta, United States of America, 3Emory University School of Medicine, Dept. Of Pharmacology And Chemical Biology, Atlanta, United States of America, 4ZebiAI, Zebiai, Waltham, United States of America, 5X-Chem Pharmaceuticals, X-chem, Waltham, United States of America, 6Sage Bionetworks, Neurodegeneration Research, SEATTLE, United States of America, 7University of Oxford, Alzheimer’s Research Uk Oxford Drug Discovery Institute, Oxford, United Kingdom

Aims: Investigation of the Alzheimer’s Disease (AD) brain proteome, along with weighted co-expression network analysis, revealed a module enriched with proteins involved in inflammation (PMID: 27989508). Moesin (MSN) and CD44 emerged as key inflammation module hub proteins. MSN is a cytoskeletal protein with roles in focal adhesion-mediated cell motility. CD44 is a hyaluronic acid (HA) receptor that binds MSN. This project aims to develop chemical tools to disrupt this pathway. Methods: To investigate the AD-relevant distribution of MSN, tissue sections from AD patients were stained for MSN, amyloid beta, and Iba1 (microglial marker). Recombinant FERM domains of MSN and EPB41L3 were co-crystallized with ligands. CD44 ectodomain was expressed in bacteria and HEK293 cells. DNA-encoded library (DEL) screens were performed as described (http://www.x-chemrx.com/our-science/). Results: MSN localised to endothelial cells and microglia in human brain sections. In AD brains, MSN-positive microglia were observed surrounding amyloid plaques. Co-crystal structures of MSN and EPB41L3 revealed the binding mode of CD44 cytoplasmic tail to MSN. An HTRF-based assay was developed for high-throughput screening of molecules disrupting this interaction. In parallel, we performed an EPB41L3 crystal-based fragment screen. Primary hits are currently being validated. DEL screening of CD44 ectodomain-HA interaction revealed micromolar hits, which are being further developed to obtain potent brain-penetrant inhibitors. Conclusions: • MSN expression is associated with AD pathology • The CD44-MSN signalling axis was targeted for potential points of intervention • We have developed the assays, structural biology and chemical starting points to develop small-molecule inhibitors of the CD44 ectodomain and the CD44-MSN interaction.

P184 / #1206

Topic: Theme A: β-Amyloid Diseases / A2.m. Therapeutic Targets, Mechanisms for Treatment: Microglia

MICROGLIAL ACTIVATION BY MCSF PREVENTS AΒ-INDUCED SYNAPTIC DYSFUNCTION AND REDUCES EXTRACELLULAR AΒ IN ALZHEIMER’S DISEASE

Lecture Title:

J. Zuazo Ibarra1,2, C. Luchena2,3,4, E. Alberdi2, C. Matute2, E. Capetillo-Zarate2,3,5,6 1University of Basque country, Neurosciences, Leioa, Spain, 2Achucarro Basque Center for Neuroscience, Laboratory Of Neurobiology, Leioa, Spain, 3CIBERNED, Centro De Investigación Biomédica En Red Enfermedades Neurodegenerativas, Madrid, Spain, 4Faculty of Medicine and Nursery, University of the Basque Country (UPV/EHU), Department Of Neuroscience, Leioa, Spain, 5IKERBASQUE, Basque Foundation For Science, Bilbao, Spain, 6University of the Basque Country (UPV/EHU), Faculty of Medicine and Nursery, Department Of Neuroscience, Leioa (Vizcaya), Spain

Aims: Accumulation of β-amyloid (Aβ) peptide and the synaptic dysfunction are the main hallmarks of Alzheimer´s disease (AD) neuropathology. Loss of synapses occurs early in AD and is considered the best pathological correlate of cognitive decline. The role of microglia in AD remains controversial. In one hand, microglia mediates early synapse loss in AD models. In contrast, activation of microglia by immunotherapy or the cytokine macrophage colony stimulating factor (MCSF) results in a more efficient Aβ degradation. Methods: To study the role of microglia in Aβ-related synapse pathology we performed immunofluorescence to measure levels of synaptic markers and Aβ in primary cultures treated with Aβ oligomers. We also carried out immunoprecipitation for extracellular Aβ detection. Additionally, microglia behavior was study in an ex vivo AD model. Results: We found that microglia decreased extracellular Aβ in microglia-neuron co-cultures in presence of extracellular Aβ oligomers, although this was not enough to restore Aβ-induced synapse loss. Synapse pruning was not observed in or co-culture model. Notably, microglial activation by MCSF was not only able to reduce extracellular Aβ load more consistently but also to prevent synapse damage. Moreover, MCSF treatment induced an increase in Aβ internalization by microglia and an increase in microglial population in vitro. Also, MCSF treatment reverted Aβ-induced microglial area increase, measured by Iba1 immunofluorescence, in cortex organotypic cultures. Conclusions: These results strongly suggest that Aβ oligomers are deleterious to synaptic function by interfering with neurons, and that microglial activation by MCSF might constitute an important therapeutic target for the prevention of synapse toxicity.

P185 / #647

Topic: Theme A: β-Amyloid Diseases / A2.o. Therapeutic Targets, Mechanisms for Treatment: Gene therapy and gene editing

MULTIFACTORIAL THERAPEUTIC CAPACITY OF A MUTANT FORM OF E2F4 IN A MURINE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

J.M. Frade1, M. Ramón-Landreau1,2, C. Trujillo1, A. Garrido-García1,2, N. López-Sánchez1,2 1Cajal Institute, Molecular, Cellular And Developmental Neurobiology, Madrid, Spain, 2Tetraneuron, -, Valencia, Spain

Aims: Alzheimer’s disease (AD) has a multifactorial etiology, which requires a single multi-target approach for an efficient treatment. We have studied the putative role of E2F4 in AD, as this transcription factor regulates cell quiescence and tissue homeostasis, controls gene networks affected in AD, and is upregulated in the brain of Alzheimer’s patients and of APP/PS1 and 5xFAD transgenic mice. Methods: E2F4 contains an evolutionarily-conserved Thr-motif that, when phosphorylated, modulates its activity, thus constituting a potential target for intervention. We have generated transgenic mice with neuronal expression of a dominant negative form of E2F4 lacking this Thr-motif (E2F4DN), which was crossed with 5xFAD mice. Then, their descendants and control EGFP/5xFAD mice were subjected to transcriptomic analysis as well as biochemical and functional studies. Results: We have found that neuronal expression of E2F4DN in 5xFAD mice potentiates a transcriptional program consistent with global brain homeostasis. The latter correlates with attenuation of both microglial immune response and astrogliosis, modulation of Aβ proteostasis, and blockade of neuronal tetraploidization. Moreover, E2F4DN prevents cognitive impairment and body weight loss, a known somatic alteration associated with AD. Conclusions: We propose E2F4DN-based gene therapy as a promising multifactorial approach against AD.

P186 / #1485

Topic: Theme A: β-Amyloid Diseases / A2.p. Therapeutic Targets, Mechanisms for Treatment: ASO and RNAi

SEX-DEPENDENT EFFECTS OF ALPHA-SYNUCLEIN SILENCING IN A MOUSE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

J. Brown1,2,3, G. Boyle2,3, D. Hart2,3, T. Brown2,3,4, M. Lacroix3,4, M. Kim2,3, M. Sherman2,3, M. Sung2,3, T. Cole5, J. Schneider6, M. Lee2,3, D. Bennett6, H. Kordasiewicz5, S. Lesne1,2,3 1University of Minnesota, Graduate Program In Neuroscience, Minneapolis, United States of America, 2University of Minnesota, Department Of Neuroscience, Minneapolis, United States of America, 3University of Minnesota, Institute Of Translational Neuroscience, Minneapolis, United States of America, 4University of Minnesota, Medical Scientist Training Program, Minneapolis, United States of America, 5Ionis Pharmaceuticals Inc., Neurodegeneration Unit, Carlsbad, United States of America, 6Rush University, Rush Alzheimer’s Disease Center, Rush University Medical Center, Chicago, United States of America

Aims: Evidence is accumulating that Alzheimer’s disease (AD) and Parkinson’s disease, two of the most common neurodegenerative disorders, might share signaling pathways that could account for common pathology and symptomology. We previously described how genetically ablating SNCA encoding alpha-synuclein (aSyn) in a mouse model of AD improved survival and spatial memory. Antisense oligonucleotides (ASOs) are a clinically tested method of transiently lowering target gene transcripts. We investigated whether treatment with ASO targeting murine SNCA mRNA improved cognition in mice overexpressing human amyloid precursor protein. Methods: Male and female mice were injected with ASOSNCA and learning and memory was tested using the Barnes circular maze. Results: The murine ASOSNCA was successful at lowering both SNCA mRNA and protein, and improving cognition in male mice. Female mice, however, displayed impaired memory retention. In addition, constitutive ablation of SNCA in female mice also worsened spatial memory. Since mediators of SNCA and its abundance might impact responses to ASOSNCA treatment, we used human samples from the Religious Orders Study to investigate potential sex differences in aSyn protein and its modulators. While confirming that soluble aSyn was increased in male and female AD brains, we also observed sex-specific differences in beta-synuclein (bSyn), an endogenous regulator of aSyn. bSyn was decreased in female AD brains but not in male brains, and cognitive scores for women in the AD group were inversely correlated with bSyn abundance. Conclusions: These results highlight a novel sex difference in our understanding of AD pathogenesis and may have profound implications for future translational studies.

P187 / #231

Topic: Theme A: β-Amyloid Diseases / A2.r. Therapeutic Targets, Mechanisms for Treatment: Other

TRANSCRANIAL PULSE STIMULATION (TPS) – A SEMINAL NEW BRAIN THERAPY

Lecture Title:

R. Beisteiner Medical University of Vienna, Neurology, Vienna, Austria

Aims: Transcranial Pulse Stimulation (TPS) may become a powerful new technique to modulate the human brain in a focal and targeted manner. For this new brain therapy, long term benefits for patients with Alzheimer’s have already been published. Due to its unprecedented clinical capabilities for precise network targeting and deep brain stimulation, particular TPS advantages are also expected for therapy of movement disorders. Methods: The presentation will introduce the new clinical sonication technique TPS that is based on single ultrashort acoustic pulses, which markedly differ from existing focused ultrasound techniques. Results: from the comprehensive preclinical and clinical studies will be presented. The preclinical TPS results show large safety margins and dose dependent neuromodulation. In 35 patients with Alzheimer’s disease, clinical long term effects after TPS were observed for the first time (Beisteiner et al. 2020, Abbasi 2020). The clinical results reveal high treatment tolerability and no major side effects. Neuropsychological scores improved significantly after TPS and improvements lasted for three months and correlated with an upregulation of the memory network (fMRI data). Conclusions: The talk will conclude with an outlook on potential benefits for movement disorders and other network specific diseases (Beisteiner & Lozano 2020). References: Beisteiner R, Matt E, Fan C, et al. Transcranial Pulse Stimulation with Ultrasound in Alzheimer’s Disease—A New Navigated Focal Brain Therapy. Advanced Science 2020, 7, 1902583. Abbasi J. Ultrasound Brain Stimulation Piloted in Alzheimer Study. JAMA 2020;323(6):499. Beisteiner R, Lozano A. Treating the Brain at the Speed of Sound. Brain Stimul. 2020 May 5. pii: S1935-861X(20)30099-1.

P188 / #1231


ALZHEIMER’S DISEASE TARGET PRIORITIZATION VIA GENETIC, MULTI-OMIC, NEUROPATHOLOGICAL, LITERATURE, AND PATHWAY EVIDENCE INTEGRATION

Lecture Title:

G. Carter1, G. Cary1, J. Gockley2, B. Lehallier3, K. Carter4, K. Leal5, L. Grinberg6, N. Seyfried4, J. Young7, S. Jayadev8, R. Betarbet4, L. Omberg9, A. Greenwood5, J. Shulman10, F. Longo3, A. Levey11, L. Mangravite5, B. Logsdon5 1The Jackson Laboratory, Research, Bar Harbor, United States of America, 2Sage Bionetworks, Neurodegeneration Research, Seattle, United States of America, 3Stanford University, Neurology, Palo Alto, United States of America, 4Emory University, Neurology, Atlanta, United States of America, 5Sage Bionetworks, Neurodegeneration Research, SEATTLE, United States of America, 6University of California at San Francisco, Neurology, San Francisco, United States of America, 7University of Washington, Laboratory Medicine And Pathology, Seattle, United States of America, 8University of Washington, Neurology, Seattle, United States of America, 9Sage Bionetworks, Systems Biology, Seattle, United States of America, 10Baylor College of Medicine, Neurology, Houston, United States of America, 11Emory University, Department Of Neurology, Atlanta, United States of America

Aims: Genome-scale studies are generating thousands of potential molecular targets for treating late-onset Alzheimer’s disease (AD). Here we combine evidence across mutually supporting but independent studies to prioritize specific genes, proteins, and pathways for further resource and reagent generation in the Target Enablement to Accelerate Therapy Development for Alzheimer’s Disease (TREAT-AD) consortium. Methods: We propose a gene level scoring method that scores evidence across genome-wide association studies of large AD cohorts, genetic evidence from model systems, transcriptomic and proteomic evidence from human post-mortem brain tissue in the Accelerating Medicine Partnership in Alzheimer’s Disease (AMP-AD) cohorts, neuropathological evidence from high throughput functional screens in D. melanogaster, and literature evidence of disease relevance. Subscores from each category of evidence are combined in a final target score for therapeutic prioritization. Results: We scored a pilot set of 1742 genes implicated in AD from recent work in the AMP-AD consortium. From this list, we identified 128 with target scores in the high tail of the distribution. These 128 targets are highly networked across relevant pathways in AD including endolysosomal trafficking, synaptic function, structural stabilization, immune response, and regulation of apoptosis. Ten targets were further prioritized for immediate development in TREAT-AD. Conclusions: Our method provides an integrative, evidence-based strategy for prioritizing candidate molecules for further assay development, structural biology, and medicinal chemistry studies in the TREAT-AD consortium. This de-risking of understudied molecular candidates will enable the rational development of critical resources that will be openly shared with the research community.

P189 / #998


COMBINATION OF MICRODOSE LITHIUM NP03 AND S-ADENOSYLMETHIONINE ALLEVIATES COGNITIVE DEFICITS AND AMYLOID-DRIVEN PATHOLOGY IN MCGILL-R-THY1-APP RATS

Lecture Title:

S. Do Carmo1, J. Emmerson1, M. Foret1, L. Welikovitch1, A.C. Cuello1,2,3,4 1McGill University, Pharmacology & Therapeutics, Montreal, Canada, 2McGill University, Anatomy And Cell Biology, MONTREAL, Canada, 3McGill University, Integrated Program In Neuroscience, MONTREAL, Canada, 4McGill University, Neurology And Neurosurgery, MONTREAL, Canada

Aims: We have previously shown that treatment with the microemulsion formulation of lithium NP03 and S-adenosylmethionine (SAM), a global methyl donor, separately, slows the progression of amyloid-driven pathology and alleviates cognitive impairment. The present study was conducted to assess the potential therapeutic value of a combination of NP03 and SAM when applied at early disease-stages, prior to plaque deposition, to a rat model of Alzheimer-like amyloid pathology. Methods: Three-month-old (pre-plaque) McGill-R-Thy1-APP transgenic rats were treated with NP03 (rectal administration; 40 ug Li/Kg; 5 days per week), SAM (orally; 20 mg/Kg; 3 days per week), a combination of NP03-SAM, or vehicle for 12 weeks followed by a wash-out period of 3 weeks. Rats underwent behavioral testing before tissue harvest followed by immunohistochemical and biochemical analyses. Results: McGill-APP rats treated with a combination of NP03-SAM presented less cognitive deficits in the auditive fear conditioning task compared with either monotherapy. These improvements in cognition were accompanied by a lower abundance of amyloid-β peptides and reduced BACE 1 expression and activity. Beneficial effects of NP03-SAM treatment also included a reduction in microglia recruitment, increased neurogenesis, and global DNA remethylation. Conclusions: Complex conditions such as Alzheimer’s disease call for preventive and treatment interventions that can engage several targets simultaneously. Our findings provide preclinical evidence suggesting that combined treatment with NP03 and SAM could be a promising, non-amyloid targeted, therapeutic approach in the early stages of Alzheimer’s disease.

P190 / #365


ELUCIDATING ALZHEIMER’S DISEASE SUBGROUPS TO IMPROVE PATIENT OUTCOME

Lecture Title:

E. Hascup1, K. Hascup2 1Southern Illinois Univ. School of Medicine, Neurology, Cadrd, And Pharmacology, Springfield, United States of America, 2Southern Illinois Univ. School of Medicine, Neurology, Cadrd, Pharmacology, And Mmicb, Springfield, United States of America

Aims: We will discuss refining AD into overlapping subgroups based upon known genetic risk factors (APP, PSEN1, ApoE ε4, TREM2, and TOMM40) and environmental contributors including blood-brain barrier disruption, metabolic syndrome, mitochondrial dysfunction, traumatic brain injury, hyperactive neuronal networks, cellular senescence, sleep disturbances, and microbial infiltration. Methods: Alzheimer’s disease (AD) is an age-related neurodegenerative disorder characterized by progressive anterograde amnesia, cerebral atrophy, and eventual death. The prevailing dogma implicates amyloid or tau aggregation causes AD progression, but available treatments clearing these proteinopathies have been ineffective. Failure to find disease modifying treatments for AD may be due to its broad diagnosis. Refining AD into overlapping subgroups allows for biomarker identification across a narrower patient population for improved pharmacotherapeutic opportunities. Results: Multiple, interrelated causalities contribute to the subsequent cognitive and functional decline that are funneled into a single diagnosis – AD. Furthermore, genetic risk factors for early and late-onset AD may accelerate or aggravate particular environmental subgroups based on their mechanisms of action. Conclusions: Identifying biomarkers across a narrower patient population can aide in designing appropriate preclinical models that more accurately recapitulate subgroup-specific disease pathologies for testing novel therapeutics. The interrelatedness of many of these proposed subgroups indicates no one single factor may initiate the cognitive and functional decline. As such, combination therapeutic approaches based upon subgroup identification could be warranted. This work was supported by the National Institutes of Health [NIA R01AG057767 and NIA R01AG061937], CADRD, and the Kenneth Stark Endowment at SIU SoM.

P191 / #581


TREATMENT OPTIONS FOR SLEEP-RELATED EPILEPTIC SPIKING IN EARLY ALZHEIMER’S DISEASE

Lecture Title:

H. Tanila, N. Jin, I. Gureviciene University of Eastern Finland, A. I. Virtanen Institute, Kuopio, Finland

Aims: Epileptic spiking without motor manifestations can be found in more than 40% patients with early AD [1]. This number may still be an underestimate since this spiking occurs almost exclusively during sleep. Further, AD patients with subclinical epileptiform activity had more than twice faster cognitive decline during a 5-year follow-up [1]. Treating epileptic spiking during sleep may offer a way to slow down AD progression, but so far there is no published clinical study on the topic, calling for preclinical testing to lead the choice of treatment and further drug development. We recently identified medial temporal giant spikes occurring in APP/PS1 mice but not in their wild-type littermates and only during sleep [2]. Using this model we aim to test the efficacy of different categories of drugs against these giant spikes. [1] Vossel KA et al. Ann Neurol. 2016;80(6):858-870. [2] Gurevicience I et al., Front Neurol. 2019;10:1151. Methods: Ten male APP/PS1 mice (3-5 months) were implanted bilaterally with cortical screw electrodes and hippocampal staggered triple wire electrodes to record local field potentials and neck EMG wire electrodes. Sleep is monitored by overhead video-recording and EMG. We will test the standard AD-drugs donepezil and memantine, anti-epileptic drugs levetiracetam and lamotrigine, gamma-secretase inhibitor semagacestat and an anti-inflammatory tetracycline derivative minocycline at doses with no overt motor manifestations. Results: All mice displayed giant spikes during sleep and preliminary data suggest their suppression by some of the treatments. Conclusions: Sleep-related epileptic spiking in AD is treatable with clinically approved drugs.

P192 / #1799


ACTIVELY TARGETED POLY(ETHYLENE GLYCOL)-POLY(Ε-CAPROLACTONE) NANOPARTICLE OF ROSIGLITAZONE ON REVERSAL OF COGNITIVE DEFICITS IN ANIMAL MODEL OF ALZHEIMER’S TYPE OF DEMENTIA

Lecture Title:

S.L. K C Birla Institute of Technology and Science - Pilani (BITS-Pilani), Pharmacy, Pilani, India

Aims: To evaluate the efficacy of brain targeted poly(ethylene glycol)-poly(ε-caprolactone) (PEG-PCL) nanoparticle of rosiglitazone in neuro-protection along with its effect on various genes implicated in memory and cognition. Methods: Mice model of Alzheimer’s type of dementia was developed with intra-hippocampal injection of beta-amyloid1-42. The PEG-PCL based nanoparticle of rosiglitazone was prepared by emulsion-diffusion-solvent evaporation method further the nanoparticles were linked with TAT peptide. The rosiglitazone (5mg/kg, 10mg/kg in free form and 5mg/kg dose in nanofomulation) was administered for 28 days. Neuroprotective effect of rosiglitazone was assessed by evaluating the expression of genes which are implicated in cognitive function, such as CREB, BDNF, GDNF and NGF with respect to internal control gene by RT-PCR method, the behavioral parameters was assessed by Morris water maze test and neuronal degeneration was estimated by histo-chemical analysis. Results: We found that free form of rosiglitazone up-regulated the mRNA expression of CREB, BDNF, GDNF, NGF with respect to internal control in a dose dependent manner, However the nanoformulation markedly up regulated the mRNA expression than that of its free form. Further, the behavioral study and the histo-chemical analysis revealed significant improvement on cognitive deficits and neuronal degeneration after rosiglitazone nanoformulation treatment. Conclusions: In view of our study outcomes it is recommended that PPARγ agonist rosiglitazone in its nanoformulation have efficiently attenuated the cognitive deficits and neuronal degeneration in beta-amyloid induced mice model of Alzheimer type of dementia along with up-regulation of genes implicated for memory.

P193 / #439


SCO-SPONDIN DERIVED PEPTIDES, INNOVATIVE DRUG CANDIDATES FOR THE TREATMENT OF ALZHEIMER’S DISEASE

Lecture Title:

J. Le Douce1, N. Delétage1, S. Lemarchant1, Y. Godfrin1,2 1Axoltis Pharma, Preclinical R&d, Lyon, France, 2Godfrin Life Sciences, -, Caluire-et-Cuire, France

Aims: SCO-spondin is a brain-specific glycoprotein whose secretion starts during embryogenesis and stops after birth in humans. A small peptide derived from this protein, named NX, displays neuroprotective and neuroregenerative properties. Because drugs reducing cognitive symptoms in Alzheimer’s disease (AD) do not altered its progression, NX effect on cognition was evaluated as a: (1) Single treatment, (2) Combinatory treatment with donepezil (DPZ), (3) Rescue treatment following DPZ resistance. Methods: Mice were injected with Aβ25-35 intracerebroventricularly to mimic AD and treated as follows: (1) NX alone, (2) A combination of subactive doses of NX + DPZ, (3) DPZ until resistance and then NX. Several behavioral tests (Y-maze, passive avoidance, Morris water maze) and biochemical analyses of AD biomarkers were performed. Results: (1) NX-treated mice displayed a full restoration of all memory-types evaluated. Cognitive recovery by NX was accompanied by a significant beneficial modulation of cerebral levels of AD pathological biomarkers such as hyperphosphorylated Tau, Aβ1–42, neuroinflammation, oxidative stress and synaptic markers. (2) NX/DPZ-treated mice were protected from Aβ25-35-induced cognitive deficits for 17 weeks without losing efficacy. (3) Four weeks after Aβ25-35 administration, mice were resistant to DPZ. Interestingly, NX administration completely rescued memory loss in DPZ-resistant mice for up to 17 weeks. Conclusions: These results support the proof of concept of NX for efficacy either alone, in combination or as a rescue treatment to DPZ. NX represents a promising drug-candidate to adjust therapeutic protocols intended for AD patients depending on the stage, tolerance, and treatment duration.

P194 / #1432


THE EMORY-SAGE-SGC TREAT-AD CENTER FOR NEW MEDICINES IN ALZHEIMER’S DISEASE

Lecture Title:

L. Mangravite1, G. Carter2, S. Frye3, H. Fu4, O. Gileadi5, A. Greenwood6, K. Leal6, R. Betarbet7, F. Longo8, L. Omberg9, K. Pearce, Jr.3, A. Edwards10, A. Levey11 1Sage Bionetworks, Neurodegeneration Research, SEATTLE, United States of America, 2The Jackson Laboratory, Research, Bar Harbor, United States of America, 3University of North Carolina at Chapel Hill, Eschelman School Of Pharmacy, Chapel Hill, United States of America, 4Emory University School of Medicine, Dept. Of Pharmacology And Chemical Biology, Atlanta, United States of America, 5University of Oxford, Structural Genomics Consortium, Oxford, United Kingdom, 6Sage Bionetworks, Neurodegeneration Research, Seattle, United States of America, 7Emory University School of Medicine, Department Of Neurology, Atlanta, United States of America, 8Stanford University, Neurology, Palo Alto, United States of America, 9Sage Bionetworks, Systems Biology, Seattle, United States of America, 10University of Toronto, Department Of Molecular Genetics, Toronto, Canada, 11Emory University, Department Of Neurology, Atlanta, United States of America

Aims: The Emory-Sage-SGC TREAT-AD Center was established with the goal of developing and openly distributing experimental tools for use in the evaluation of a diverse set of novel Alzheimer’s Disease (AD) targets. Methods: Targets are derived from systems biology studies within the Accelerating Medicines Partnership in AD (AMP-AD) consortium and further evaluated using data from multiple AD consortia and existing literature. All TREAT-AD investigators – and institutions – will place all data, knowledge, reagents, and tools including chemical and biological probes into the open domain with no intellectual property claims. Results: Targets nominated through the AMP-AD consortium were mapped to mechanistic hypotheses and prioritized based on unbiased bioinformatic assessments across multiple lines of evidence. Hypotheses prioritized in the initial evaluation included those relating to immune function, endocytosis, and retromer-mediated endosomal protein sorting. Target Enabling Packages, including expression constructs, knockout cell lines, assays, antibodies and crystal structures, are being generated for 20 proteins identified within these mechanisms. Hit characterization and chemical probe development has started for CD44. All data and reagents will be publicly shared through standard repositories and cataloged at the Agora site (https://agora.adknowledgeportal.org). Conclusions: The open drug discovery approach of TREAT-AD is aimed to de-risk potential AD therapeutics to support industry investment by catalyzing robust and independent evaluation of a diverse portfolio of promising yet untested AD therapeutic hypotheses.

P195 / #1361


EFFECT OF PLASMA EXCHANGE WITH ALBUMIN REPLACEMENT ON ALBUMIN LEVELS AND BLOOD BRAIN BARRIER INTEGRITY OF ALZHEIMER’S DISEASE PATIENTS

Lecture Title:

A.M. Ortiz1, C. Minguet1, L. Núñez1, A. Ruiz2, O. López3, M. Boada2, A. Páez1, M. Costa1 1Grifols, Alzheimer's Research Group, Sant Cugat del Vallès, Barcelona, Spain, 2Fundació ACE, Institut Català de Neurociències Aplicades - Universitat Internacional de Catalunya, Research Center And Memory Clinic, Barcelona, Spain, 3University of Pittsburgh School of Medicine, Neurology And Psychiatry, Pittsburg, United States of America

Aims: To assess the effects of plasma exchange with albumin replacement on brain blood barrier (BBB) integrity using cross-sectional and longitudinal changes over time of albumin in CSF and serum in Alzheimer’s disease patients participating of the AMBAR study. Methods: Albumin levels were measured in serum (n=319) and CSF (n=109) at baseline, intermediate visit (after 6-week period of conventional therapeutic PE-A [TPE]; 1TPE/week), and final visit (after 12-month period of low volume PE-A [LVPE]; 1LVPE/month). The integrity of the BBB function was measured with the CSF/serum albumin ratio (QAlb) (abnormal values: >8; normal reference values: 32-56g/L serum albumin and 0.08-0.42g/L CSF albumin). Results: Baseline serum and CSF albumin levels were within normal range (44.0g/L [33.8-56.3] and 0.199g/L [0.105-0.542] respectively), with 89% of patients (97/109) presenting no BBB disruption. Men (p<0.05) and patients aged <75 (p<0.01) showed the highest baseline albumin levels and QAlb, as previously described. PE-A-treated patients presented higher change from baseline in both serum and CSF albumin levels after TPE period compared to non-treated (2.10g/L vs 0.7g/L [p<0.0001] and 0.025 g/L vs 0.002g/L [p<0.0001], respectively), with no BBB integrity worsening. Albumin levels of both compartments remained within normal range after 14-month treatment (42.9g/L [40.7-44.9] and 0.197g/L [0.165-0.275], respectively) with 81% of patients (35/43) exhibiting no BBB disruption. Conclusions: This study showed that in Alzheimer’s disease patients treated with plasma exchange, the levels of albumin in CSF and serum remained stable, as well as the QAlb, suggesting that PE-A treatment does not affect BBB integrity.

P196 / #511


MICROTUBULE STABILIZATION REDUCES AMYLOID PATHOLOGY AND IMPROVES SYNAPTIC/MEMORY DEFICITS IN APP/PS1 MICE

Lecture Title:

R. Sanchez-Varo1, J.J. Fernandez-Valenzuela1, C. Muñoz-Castro2, V. De Castro1, E. Sanchez-Mejias1, V. Navarro2, S. Jimenez2, C. Nuñez-Diaz1, A. Gomez-Arboledas1, I. Moreno Gonzalez1, F. Moyano2, M.L. Vizuete2, J.C. Davila1, J. Vitorica2, A. Gutierrez1 1University of Malaga/CIBERNED/IBIMA, Cell Biology, Genetic And Physiology, Malaga, Spain, 2University of Seville/CIBERNED/IBIS, Biochemistry And Molecular Biology, Seville, Spain

Aims: Cognitive decline in Alzheimer's disease (AD) is highly related to synaptic/neuronal loss. Tau hyperphosphorylation destabilizes microtubules leading to axonal transport failure and generation of dystrophic neurites, thus contributing to synaptic dysfunction. The effect of microtubule stabilization on amyloid-β (Aβ) pathology has not been assessed in vivo yet. This study evaluated the effect of the microtubule-stabilizing agent, Epothilone D (EpoD) in the pathology of an amyloidogenic mouse model. Methods: APP751SL/PS1M146L mice (3-month-old) were treated weekly with intraperitoneal injections of EpoD (2 mg/kg) or vehicle for 3 months. For memory performance, animals were tested on the object-recognition, Y-maze and Morris water maze. Hippocampal proteinopathies were quantified by image analysis after immunostaining. Somatostatin (SOM)-numerical density was calculated by stereology. APPswe-N2a cells were treated with EpoD 100nM for 12/24 hours. Protein levels were analysed by Western/dot-blot. Results: EpoD-treated mice improved their performance of cognitive tests, while hippocampal phospho-tau and Aβ (especially oligomers) accumulation decreased, together with synaptic/neuritic pathology. Remarkably, EpoD exerted a neuroprotective effect on SOM-interneurons, a highly AD-vulnerable GABAergic subpopulation. Conclusions: EpoD improved microtubule dynamics and axonal transport in an AD-like context, reducing tau and Aβ accumulation and promoting neuronal and cognitive protection, underlining the cross talk between cytoskeleton pathology and proteinopathy. Therefore, microtubule-stabilizing drugs could be candidates for slowing AD at both tau and Aβ pathologies. Supported by PI18/01557 (to AG) and PI18/01556 (to JV) grants from ISCiii of Spain, co-financed by FEDER funds (European Union), CIBERNED collaborative grant (to AG and JV), and by PPIT.UMA.B1.2017/26 grant (to RSV).

P197 / #1537


NEUROPROTECTIVE EFFECTS OF 17Β-ESTRADIOL METABOLITES ON SH-SY5Y CELLS

Lecture Title:

L. Zacharia1, C. Eleftheriou1, V. Gkretsi2 1University of Nicosia School of Sciences and Engineering, Life And Health Sciences, Nicosia, Cyprus, 2European University Cyprus, Life Sciences, Nicosia, Cyprus

Aims: It is well known that 17β-estradiol offers protection in Alzheimer’s disease (AD) as evidenced from a number of clinical and non clinical studies through diverse mechanisms. In this study we hypothesize that the protective effects of 17β-estradiol are partly afforded by its biologically active metabolites and specifically 2-methoxyestradiol (2ME). Methods: To test the hypothesis, SH-SY5Y neuronal cells were subjected to H2O2 induced cell death, in the presence and absence of 17β-estradiol and its biologically active metabolite 2-methoxyestradiol. Cell death was assessed by the MTT assay as well as methylene blue viability test. Measurement of free radicals/oxidative stress was conducted by the DCFDA method, and RT-PCR was conducted on selected pro and anti apoptotic genes. Results: Our results indicate that 2ME protects SHSY-5Y cells from H2O2 induced neuronal cell death as evidenced both by the MTT assay and the methylene blue viability test. Measurement of oxidative stress with the DCFDA method indicated that 2ME was able to reduce the oxidative stress induced by H2O2. Full reversal of H2O2 induced cell death was not possible indicating that the protective effect of 2ME is partial, and can be partly attributed to its antioxidative effect. Furthermore, RT-PCR results indicate that 2ME increases anti apoptotic genes and decreases pro apoptotic genes. Conclusions: In this work we demonstrate that the protective effects of 17β-estradiol are partly attributed to its metabolite 2-methoxyestradiol. Our findings may explain partly why hormone replacement therapy not containing 17β estradiol does not protect from AD.

P198 / #1441

Topic: Theme A: β-Amyloid Diseases / A3.c. Drug Development, Clinical Trials: Amyloid clearance

DESIGNED MACROCYCLIC AMYLOIDOGENESIS INHIBITORY PEPTIDE MITIGATES ALZHEIMER’S DISEASE PATHOLOGY IN A MURINE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

H. Ji1, C. Zan2, B. Dalla-Volta3, L. Liu1, A. Spanopoulou3, F. Usifo1, S. Besson-Girard4, J. Bernhagen2,5, A. Kapurniotu3, O. Gokce4,5 1Institute for Stroke and Dementia Research (ISD) University Hospital, LMU Munich, System Neuroscience Laboratory, Munich, Germany, 2Institute for Stroke and Dementia Research (ISD) University Hospital, LMU Munich, Department Of Vascular Biology, Munich, Germany, 3Technische Universität München, Division Of Peptide Biochemistry, Freising, Germany, 4Institute for Stroke and Dementia Research, Klinikum der Universität München, System Neuroscience Laboratory, Munich, Germany, 5Munich Cluster for Systems Neurology (SyNergy), -, Munich, Germany

Aims: Alzheimer’s disease (AD) pathology involves the production and self-assembly of beta-amyloid peptide, but no anti-amyloid treatment is currently available. We previously showed that designed islet amyloid polypeptide (IAPP)-derived macrocyclic peptides (MCIPs) can act as nanomolar inhibitors against amyloidogenesis in -vitro. Here, our main objective was to evaluate the amyloidogenesis inhibitory effect of one such MCIP, the IAPP analog 2E in an AD mouse model. Methods: To elucidate the impact of 2E peptide treatment on AD pathology, we performed chronic intraperitoneal injections in parallel to disease development in the 5xFAD model. Next, we characterized the 2E effect on the AD mouse model using behavioral tests, imaging, and serological tests. Results: 2E peptide treatment significantly reduces b-amyloid levels and neuronal damage. The treatment also substantially lowered spatial learning and memory deficits in the AD mouse model. These improvements in AD pathology are accompanied by increased plaque-associated activated astrocytes numbers and microglia phagocytosis. Conclusions: Treatment of 5x FAD mice with the 2E peptide mitigates hallmarks of AD pathology such as amyloid burden, behavior deficits, and neuronal damage. These results suggest 2E and related MCIPs are promising treatment candidates for AD.

P199 / #1294


PHARMACOLOGICAL EVALUATION OF NEUROPROTECTION PROPERTIES OF NATURALLY INSPIRED NOVEL MULTIFUNCTIONAL MOLECULE F-24 (EJMC-7A) THROUGH IN-VITRO, IN-VIVO, AND IN-SILICO APPROACHES IN ALZHEIMER’S DISEASE MODELS

Lecture Title:

Y.P. Singh1, N. Kumar2, K. Priya3, B. Chauhan4, G. Singh1, A. Kashyap1, S. Srikrishna4, G. Singh5, P. Garg2, G. Rai3, G. Modi1 1Indian Institute of Technology (BHU), Department Of Pharmaceutical Engineering & Technology, varanasi, India, 2National Institute of Pharmaceutical Education and Research (NIPER), Department Of Pharmacoinformatics, SAS Nagar, India, 3Banaras Hindu University, Department Of Molecular And Human Genetics, varanasi, India, 4Banaras Hindu University, Department Of Biochemistry, varanasi, India, 5Central University of South Gaya, Bihar, Department Of Pharmacy, gaya, India

Aims: In our continued efforts to discover naturally inspired multifunctional molecules for AD, we have carried out a systematic extensive SAR and identified in-vivo active multifunctional molecule F-24 (7a) in our earlier publication. Here, we report the detailed pharmacological evaluation of F-24 as a novel potent cholinestrases inhibitor as a symptomatic and neuroprotective agent for AD. Methods: A. In-vitro neuroprotection studies against H2O2 induced neurotoxicity were carried out with the SHSY5Y cell line. B. Evaluation of in-vivo and ex-vivo anti-AD property in water maze model. C.The efficacy of F-24 against Aβ induced neurotoxicity in Drosophila melanogaster was evaluated. D. Molecular docking and molecular dynamics simulation studies of F24 on Aβ1-42 protofibrils (PDB ID 2BEG). Results: The significant neuroprotection was conferred by pretreatment of F-24 in wide concentration range (1-20 µM) against H2O2 induced neurotoxicity in SHSY5Y cell line. F24 exhibited promising in-vivo and ex-vivo activities upon administration through the i.p. route in the scopolamine-induced water maze AD model. This supports our earlier reported in-vivo activity upon oral administration of F-24 through oral route in the Y maze AD model. The AD flies treated with F24 exhibited dose-dependent increase in rescue percentages of eye phenotypes such as 43% (0.2mg/ml), 53% (0.4 mg/ml), 60% (0.6mg/ml) and 68% (0.8mg/ml), respectively. F24 also forms two hydrogen-bonding interactions with Leu17C and Leu17E of pentameric Aβ42 fibrils. The computed parameters highlight the destabilization of Aβ42 protofibrils in presence of ligand F24 Conclusions: Our studies strongly support that F24 can act as a promising lead molecule for the management of AD.

P200 / #1046


IN VITRO CHARACTERIZATION OF THE BRAIN-SPECIFIC SIGNAL PEPTIDE PEPTIDASE LIKE 2 B (SPPL2B): A NOVEL THERAPEUTIC APPROACH TO TREAT ALZHEIMER'S DISEASE

Lecture Title:

S. Tambaro1, R. Maccioni2, S. Zerial3, A. Wagener4, Y. Andrade-Talavera1, A. Fisahn1, T. Mentrup5, B. Schröder5, P. Nilsson6 1Karolinska Instituet, Department Of Neurobiology, Care Sciences And Society, Solna, Sweden, 2University of Cagliari, Department Of Biomedical Sciences, Neuroscience And Clinical Pharmacology, Cagliari, Italy, 3University of Trieste, Department Of Life Science, Trieste, Italy, 4Heidelberg University, Interdisciplinary Center For Neuroscience, Heidelberg, Germany, 5Technische Universität Dresden, Institute Of Physiological Chemistry, Dresden, Germany, 6Karolinska Institutet, Dept Of Neurobiology, Care Sciences And Society, Stockholm, Sweden

Aims: Alzheimer's disease (AD) is a multifactorial disorder in which the abnormal brain production of amyloid β-peptide (Aβ) plays a crucial role in the disease onset. Identifying new proteins and new pathways involved in the Aβ cascade is an essential step to provide novel effective therapeutic targets. The intramembrane enzyme SPPL2b is a new potential target since it is involved in the proteolysis of two AD-related proteins: TNF-alpha and BRI2, involved in the inflammatory response and Aβ production, respectively. Here we have investigated for the first time the expression levels and the pathogenic role of SPPL2b in AD. Methods: The pathophysiological role of SPPL2b in Aβ metabolism was evaluated in vitro by using human neuroblastoma cell line SH-SY5Y, mouse primary neuronal cell cultures, acute brain slices from WT mice, and brains of a new App knock-in AD mouse model (AppNL-G-F) exhibiting high levels of Aβ42. Results: APP overexpression in SH-SY5Y cells increased the levels of SPPL2b. A reduction in Aβ40 production was observed in SH-SY5Y cells upon treatment with a selective SPPL2b inhibitor. In brain slices-maintained ex vivo, Aβ42 exposure induced a strong up-regulation of SPPL2b. Increased levels of SPPL2b were also observed in the cortex of in 3 months old AppNL-G-F mice, however the levels were significantly decreased in aged mice. Furthermore, immunofluorescence staining showed that SPPL2b is expressed in neurons and glia deposited in the Aβ plaques. Conclusions: These results strongly support the involvement of SPPL2b in AD pathology. Most importantly, Aβ42 induces SPPL2b expression in neurons and Aβ plaque-associated microglia.

P201 / #1837


SEARCHING FOR THE THERAPEUTIC DRUGS FOR AD USING DROSOPHILA AND MOUSE

Lecture Title:

L. Tsuda, R. Minami, Y. Lim National Center for Geriatrics and Gerontology, Animal Models Of Aging, -, Japan

Aims: Alzheimer's disease (AD) is the most common cause of dementia among the elderly, however, no effective drug is available for the disease, so far. This therapeutic gap reflects the lack of an ideal model system for evaluating small compounds for AD. To identified new therapeutic drugs for AD, we have established animal model systems using Drosophila and mouse. Methods: Using Drosophila system, we have searched for chemical compounds, which can suppress toxic effect by Aβ. To check the binding between chemical compounds and Aβ we performed surface plasmon resonance method. To search for compounds that bind to Aβ in a manner similar to E3, we performed docking simulation. Results: We identified E3 from chemical library derived from natural products (consisted with 800 compounds). Our analysis suggest that E3 can bind directly to Aβ42, and that administration of E3 causes the degradation of amyloid-β (Aβ) protein in the brain. Since E3 was expected to have less transfer to the blood brain barrier (BBB), we searched for compounds that bind to Aβ in a manner similar to E3. As a result, eight compounds (N1 to N8) were identified out of more than 4 million chemical compounds and expected to have high penetrance to the BBB. We found that among three of them (N5, N6, and N8) suppressed the behavioral abnormalities of flies caused by Aβ. Conclusions: Our data suggests that N5, N6, and N8 may act on Aβ clearance in the mouse brain. We are currently using a mouse model of AD to analyze the efficacy of these compounds.

P202 / #1362


GINSENOSIDE COMPOUND K REDUCES NEURONAL OXIDATIVE DAMAGE AND IMPROVES NEURONAL SYNAPTIC DYSFUNCTION BY TARGETING AΒ

Lecture Title:

Q. Yang1, H. Li2,3, X. Sui1, J. Lin1, M. Kan1, X. Shi1, J. Li1, J. Wang1, X. Liu1, S. Ming1, Z. Zhang1, X. Qu1, N. Li1 1Changchun University of Chinese Medicine, Jilin Ginseng Academy, Changchun, China, 2Qian Wei Hospital of Jilin Province, Endocrinology Department, Changchun, China, 3Qian Wei Hospital of Jilin Province, Department Of Endocrinology, Changchn, China

Aims: Alzheimer’s disease (AD) is the most common neurodegenerative condition worldwide, amyloid β (Aβ) oligomers are the main pathological features of AD. Therefore, this study investigated whether Ginsenoside Compound K (CK) can target Aβ, and through what ways to reduce synaptic damage and cognitive impairment. Methods: Cellular AD models were established by Aβ42 insult in HT22 cells, mouse AD models were established by intraperitoneal injection of 2mg/kg scopolamine hydrobromide (SCOP). Detecting the ability of CK to regulate Aβ oligomers were performed in vitro and in vivo using transmission electron microscopy (TEM), immunohistochemistry, and Western Blotting. Molecular functional analyses of brains in mouse AD model were performed by the microarray analyses. The effect of CK on the expression of Nrf2/Keap1 signaling pathway and neuronal synaptic function was detected in HT22 cells. Results: CK reduced the aggregation of Aβ by Transmission electron microscopy. CK regulates the balance between Aβ production and clearance. CK can inhibit the accumulation of Aβ in neuronal extracellular and produce in vitro and in vivo. Based on microarray showing that CK affects the production of oxidative free radicals in neurons. Further studies indicate that CK improves neuronal synaptic function and cognitive function by regulating the expression of Nrf2/Keap1 signaling pathway. Conclusions: CK can target Aβ inhibition, regulate Aβ production and degradation balance, reduce neuronal oxidative damage, improve neuronal synaptic dysfunction, improve cognitive function, and protect neurons.

P203 / #675

Topic: Theme A: β-Amyloid Diseases / A3.f. Drug Development, Clinical Trials: Neuroprotective & mitochondrial compounds

TOPLINE RESULTS OF PHASE 2 STUDY OF AR1001 IN MILD TO MODERATE ALZHEIMER'S DISEASE PATIENTS

Lecture Title:

P. Scheltens1, D. Greeley2, M. Choung3, J. Rock4, V. Hingorani4, F. Kim4 1Amsterdam University Medical Center, Alzheimer Center, AMSTERDAM, Netherlands, 2University of Washington School of Medicine, Neurology, Seattle, United States of America, 3AriBio Co., Ltd., Headquarters, Seongnam, Korea, Republic of, 4AriBio Co., Ltd., Clinical Operations, Seongnam, Korea, Republic of

Aims: AR1001, a small molecule, is being investigated as a polypharmacological therapeutic agent for mild to moderate Alzheimer's disease. This Phase 2 study aims to evaluate the safety and efficacy of AR1001 in patients with mild to moderate Alzheimer's disease. Methods: This double-blind, randomized, placebo-controlled, parallel-group comparison study of AR1001 investigated its efficacy and safety in mild to moderate Alzheimer's disease patients. A total of 210 eligible patients were randomized into placebo, AR1001 10 mg, or AR1001 30 mg groups. AR1001 was administered as a daily oral dose for 26 weeks. Results: The Phase 2 study will be completed by the end of 2020, and the topline results will be available by January or February of 2021. We will update the results section as soon as the topline analysis is completed. Conclusions: The Phase 2 study will be completed by the end of 2020, and the topline results will be available by January or February of 2021. We will update the conclusion section as soon as the topline analysis is completed.

P204 / #593

Topic: Theme A: β-Amyloid Diseases / A3.l. Drug Development, Clinical Trials: Medicinal chemistry approaches, drug repurposing

SMALL-MOLECULE CHAPERONES OF THE INTERACTION OF TRANSTHYRETIN AND ABETA PEPTIDES AS DISEASE-MODIFYING DRUG CANDIDATES FOR ALZHEIMER’S DISEASE

Lecture Title:

G. Arsequell1, E. Cotrina1, J. Llop2, I. Cardoso3, J.R. Quintana4 1IQAC-CSIC, Biological Chemistry, Barcelona, Spain, 2CIC biomaGUNE, Basque Research and Technology Alliance (BRTA), Radiochemistry And Nuclear Imaging Lab, San Sebastian, Guipúzcoa, Spain, 3i3S Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Molecular Neurobiology, Porto, Portugal, 4Universitat Pompeu Fabra (UPF), Research Programme On Biomedical Informatics, Barcelona, Spain

Aims: Our transdisciplinary drug discovery program is focused on the transthyretin (TTR) and Amyloid-beta (Aβ) interaction. We aim to characterize a set of 10 selected compounds that behave as small-molecule chaperones (SMCs) enhancing the TTR/Aβ interaction, among them three registered drugs that can be repurposed and our small-molecule compound iododiflunisal. We aim to progress these SMCs as real disease-modifying AD therapies. Methods: By a combination of computational drug discovery approaches and in vitro biological assays we have selected a list of compounds that bind TTR and stabilize its tetrameric conformation. Using our HTS assay combined with Isothermal calorimetric studies (ITC) with ternary complexes TTR / Aβ(12-28) / compound, we prioritized a list of SMCs. Here, we used different biophysical methods (e.g. Thioflavin T fluorescence assays, Isothermal Titration Calorimetry, Dynamic light scattering) to characterize the ternary complexes [TTR/SMC/Abeta] of the 10 best SMCs. Results: A complete thermodynamic profile of the ternary complexes TTR/SMC/Abeta has been obtained by ITC confirming the chaperoning effect of the SMCs at enhancing the TTR/Aβ interaction. All the SMCs efficiently enhanced the TTR ability to prevent Aβ42 fibril formation, as determined by ThT and DLS analyses. An analogous chaperoning effect could not be observed with other TTR stabilizer drugs like diflunisal and Tafamidis. Conclusions: 10 SMCs of the TTR/Abeta interaction have been discovered and characterized, constituting good candidates for preclinical safety / clinical studies for Alzheimer's disease. The three SMCs drugs, olsalazine, sulindac and flufenamic acid, can be repurposed for AD therapeutics.

P205 / #676

Topic: Theme A: β-Amyloid Diseases / A3.m. Drug Development, Clinical Trials: Personalized medicines

HUMAN ALZHEIMER’S CAUSAL TRANSGENES MEDIATE MEMORY DEFICIT, CIRCADIAN AND SLEEP DYSFUNCTION IN DROSOPHILA MELANOGASTER

Lecture Title:

S. Ahmed University of Bristol, Pharmacology, TL, United Kingdom

Aims: Alzheimer’s disease (AD) is a progressive neurodegenerative disorder, impairing cognition and behavior. Pathological hallmarks include Amyloid Beta (Aβ) plaques and hyperphosphorylated Tau tangles. Pathogenesis implicates cortical impairment as a result of neurodegeneration, although aetiology remains unclear. This study aimed to understand the underlying mechanisms, using Drosophila melanogaster as a model of AD. Methods: Overexpression of human AD casual transgenes (secAβ, Tau and Tandem) via the GAL4-UAS system and orthologue knockdown of the top 20 hits from an epigenetic wide association study (EWAS) using RNA interface (RNAi) allowed study in neurodegeneration and behavioral assays. Results: Tau had the most severe significant (p<0.01) reduction in photoreceptor neurodegeneration (24%), memory deficits (-0.53), locomotor dysfunction (36%), circadian and sleep irregularities, compared to control. Tandem displayed further detrimental effects on photoreceptor degeneration, sleep and circadian, compared to secAβ42, which had more effect on memory and locomotor function. ANK1, human orthologue ANK3 involved in neuronal membrane attachment, showed photoreceptor degeneration. Conclusions: Overall, there appears to be many mechanistic pathways at interplay, and further investigation is to be done exploring the effects of ANK1,before it can be considered a potential therapeutic target.

P206 / #1649

Topic: Theme A: β-Amyloid Diseases / A3.m. Drug Development, Clinical Trials: Personalized medicines

FORECAST PARKINSON DISEASE PROGRESSION TO BETTER SELECT PATIENTS INTO TRIALS

Lecture Title:

J. Ortholand1, I. Koval2, E. Maheux2, A. Valladier2, S. Durrleman2,3 1Paris Brain Institute, Aramis, Paris, France, 2ICM, Institut Du Cerveau, Paris, France, 3INRIA Paris, Aramis, Paris, France

Aims: Subject recruitement is a burden that hampers clinical trials, especially in neurodegenerative diseases inducing long-term and subtle worsening of abilities. Better selecting them allows to reduce their required number - or conversely to improve the proven effect size. Methods: We extracted one-year separated visits in PPMI to model the placebo arm. For each patient, we derive a treated counterpart by applying an individual treatment effect calibrated so to get an effect size that correspond to a 4 point MDS-UPDRS II + III improvement and from which we derive a required sample size. Then, thanks to individual outcome predictions [Schiratti et al, 201] from which we derive a individual ratio of MDS-UPDRS II + III change, we select a subset of patients that minimizes the sample size for the previous effect size. The optimal ratio value is estimated on 80% of the PPMI patients and tested on the last 20%. We then test this ratio on DIGPD. The operations are bootstrapped 100 times on 50% of the data. Results: By selecting patients whose one-year MDS-UPDRS II+III ratio of predicted change is of 0.078 and for 0.4 of effect size, the number of patients needed to show a 0.4 effect size is reduced by 45±(11.4)SD% (resp. 55±(7.6)SD%) on the test set (resp. DIGPD evaluation). Conclusions: The use of our prediction model can concretly improve clinical trials on Parkinson’s disease by helping to better select patients and thus reduce the cohort size for a fixed treatment effect size.

P207 / #1558

Topic: Theme A: β-Amyloid Diseases / A3.o. Drug Development, Clinical Trials: Non-pharmacological interventions

IMPACT OF TRANSCRANIAL DIRECT CURRENT STIMULATION ON COGNITIVE FUNCTION, BRAIN FUNCTIONAL ACTIVITY IN MCI PATIENTS ACCORDING TO AMYLOID-BETA RETENTION AND APOE Ε4 ALLELE

Lecture Title:

D. Kang Seoul Saint Mary's hospital, Geriatric Psychiatry, Seoul, Korea, Republic of

Aims: Previous studies have demonstrated that anodal transcranial direct current stimulation (anodal-tDCS) improves cognition and normalizes abnormal network configuration during resting-state fMRI in mild cognitive impairment (MCI) patients. This study aimed to evaluate the impact of anodal-tDCS on memory performances, intra-regional activity, and intrinsic connectivity of default mode network in MCI patients depending on amyloid-beta (Aβ) deposition and APOE ε4 allele. Methods: In 32 patients with MCI ([18F] flutemetamol -: n=10, [18F] flutemetamol +: n=22; APOE ε4-: n=13, APOE ε4+: n=17, missing values of APOE ε4 allele: n=2), we delivered anodal tDCS (2mA/day, five times per week for 2 weeks) over the left dorsolateral prefrontal cortex and assessed neuropsychological test battery (CERAD-K) and resting-state fMRI measurements before and after 2 weeks of stimulation. Results: We observed a statistical trend toward an impact of an anodal tDCS-by-Aβ retention interaction on change in MMSE score, and toward a negative association between baseline [18F] flutemetamol SUVRPONS and change in score of word list recognition. We found a significant effect of tDCS-by-APOE ε4 allele interaction on change in amplitude of low-frequency fluctuation of temporal pole and also observed a negative association between baseline [18F] flutemetamol SUVRPONS and change in regional integrity of hippocampal formation. Conclusions: These findings suggest a possibility that anodal tDCS could have a therapeutic effect of improving cognitive function and enhancing compensatory intrinsic functional change in MCI patients, being modulated by the presence of Aβ retention and APOE ε4 allele

P208 / #1133


MODULATION OF WORKING MEMORY AND RESTING STATE FMRI BY TDCS OF THE RIGHT FRONTO-PARIETAL NETWORK

Lecture Title:

M. Pupíková1,2, P. Šimko1,2, M. Gajdoš1, I. Rektorova1,2,3 1CEITEC MU, Applied Neuroscience, Brno, Czech Republic, 2Masaryk University, Faculty Of Medicine, Brno, Czech Republic, 3School of Medicine, St. Anne's University Hospital, First Department Of Neurology, Brno, Czech Republic

Aims: We targeted the right fronto-parietal network by one session of tDCS to induce behavioral effects upon online/offline working memory tasks (WMT; Gazzaley et al., 2005, Elfmarková et al., 2017) in a double-blind cross-over trial. The online WMT was used to enhance effects of tDCS on the offline WMT. We further explored the neural underpinnings of induced changes by resting-state fMRI (rs-fMRI). Methods: Twenty-seven volunteers (age: 27 ± 4.1 years) participated in the behavioral testing and a subgroup (n=22) underwent rs-fMRI prior to and after each tDCS condition (active or sham; tDCS: 2 mA, 20 minutes). The anode was positioned over the right middle frontal gyrus (rMFG). Paired t-tests compared differences in reaction times (RT) in the offline WMT performance between stimulation conditions. Resting-state functional connectivity between the rMFG seed (engaged in the fronto-parietal network) and the default mode network (DMN) was analyzed. Results: We observed a significant difference in RT changes between stimulation conditions (p=0.049) for higher difficulty level of the offline WMT and increased connectivity between the stimulated rMFG and the DMN (p = 0.042) after the sham stimulation. We found significant positive correlation of the rMFG -DMN connectivity with the tDCS-induced behavioral changes (r=0.459; p=0.032) for the active tDCS, i.e.faster responses were associated with lower rMFG-DMN connectivity. Conclusions: Targeting the right fronto-parietal network by tDCS leads to positive cognitive aftereffects upon WMT already in young healthy subjects. Specific tDCS-induced cognitive changes are associated with changes in resting state functional connectivity between the task-positive (fronto-parietal) and the task-negative (DMN) networks.

P209 / #1145


ACUTE BEHAVIORAL AND NEURAL EFFECTS OF BI-FRONTAL OR THE RIGHT FRONTO-PARIETAL TDCS ON VISUAL WORKING MEMORY OF HEALTHY SENIORS

Lecture Title:

P. Šimko1,2, M. Pupíková1,2, M. Gajdoš1, I. Rektorova1,3 1CEITEC MU, Applied Neuroscience, Brno, Czech Republic, 2Masaryk University, Faculty Of Medicine, Brno, Czech Republic, 3School of Medicine, St. Anne's University Hospital, First Department Of Neurology, Brno, Czech Republic

Aims: The combined tDCS-cognitive training intervention may enhance working memory in healthy aging. In this pilot study, we assessed acute effects of transcranial direct current stimulation (tDCS) combined with an online visual working memory (VWM) task performance in healthy seniors. Our main objective was to identify an optimal tDCS montage with a beneficial VWM after-effect. Using resting state fMRI, we also investigated neural correlates of the tDCS-induced behavioral changes. Methods: In this double-blind crossover study, tDCS was applied to 25 older adults (68.8 ± 4.6 years) as a bi-frontal montage targeting the left DLPFC or as a right fronto-parietal setup with anode placed over the ventrolateral prefrontal cortex. A visual object matching task (VOMT) was used as the main behavioral outcome. Futher, we paired tDCS with an online VWM training task. Linear Mixed Models (LMM) were used for the behavioral and seed-based functional connectivity analysis. Results: Based on our behavioral analysis, the right frontoparietal montage had no effect while the bilateral frontal montage had a significant time × stimulation effect on the overall accuracy of the responses (F(92)=4.72, p=0.03) favoring the active stimulation condition compared to the sham. In addition, we observed a significant time × stimulation effect of the bi-frontal montage on the (enhanced) functional connectivity between the anode seed and seeds within the fronto-parietal network. Conclusions: Preliminary results indicate that bi-frontal tDCS may enhance visual working memory in healthy aging which is coupled by the enhanced intrinsic connectivity within the frontoparietal network.

P210 / #630

Topic: Theme A: β-Amyloid Diseases / A3.p. Drug Development, Clinical Trials: Other

TOPLINE ETHERAL PHASE II TRIAL DATA

Lecture Title:

R. Bullock1, M. Ropacki1, M. Boada2, S. Ratcliffe3, D. Watson4, S. Gutierrez1, J. Xaus1, C. Buesa1 1Oryzon Genomics, Clinical Research, Cornella de llobregat, Spain, 2Fundacio ACE. Institut Catala de Neurociencies Aplicades, Research Center And Memory Clinic, Barcelona, Spain, 3Sant Pancras Clinical Research, Clinical Research, London, United Kingdom, 4Alzheimer’s Research and Treatment Center (ARTC), Clinical Research, Wellington, United States of America

Aims: ETHERAL is a randomized, double-blind, parallel-group, placebo-controlled, 12-month phase II trial to evaluate safety and tolerability of two doses of vafidemstat (0.6 mg & 1.2 mg) in a mild-to-moderate Alzheimer’s disease (AD) population. Interim 6-month analyses from ETHERAL-EU, presented at AD/PD 2020, confirmed vafidemstat was safe and well-tolerated. Topline 12-month results from ETHERAL and ETHERAL-US cohorts, including efficacy and biomarkers data will be presented. Methods: ETHERAL-EU and ETHERAL-US trials were conducted under nearly identical protocols, including a 6-month double-blind placebo-controlled treatment period followed by a double-blind non-placebo controlled 6-month open extension period. Subjects were randomized to 0.6 mg or 1.2 mg of vafidemstat or placebo for the first 6-months. In the second 6-month period, placebo subjects were randomized to 0.6 mg or 1.2 mg, while previously vafidemstat treated subjects maintained their original dosing. Although ETHERAL’s primary endpoint is safety and tolerability, other endpoints including CSF biomarkers and proteomics, as well as measures of cognition, behaviour and function will be explored. Results: 117 randomized subjects were recruited into ETHERAL-EU. Despite the COVID-19 pandemic, in April 2020 ETHERAL-US closed recruitment with 24 randomized subjects, for a combined N = 141. Unblinding and final data analyses are planned after last patient last visit in November 2020 and topline data will be presented. Conclusions: ETHERAL is the first epigenetic Phase II trial in AD which aims to demonstrate that 12-month vafidemstat treatment is safe and well-tolerated in a mild-to-moderate late onset AD population.

P211 / #1387

Topic: Theme A: β-Amyloid Diseases / A3.p. Drug Development, Clinical Trials: Other

9-MER CATLYTIDES, JAL-TA9 IMPROVED THE COGNITIVE IMPAIRMENT OF ALZHEIMER’S MODEL MICE THROUGH THE SINGLE DIRECTION DOSE INTO HIPPOCAMPUS

Lecture Title:

R. Nakamura1,2, Y. Higashi2, T. Aratake2, M. Konishi3, M. Saito2, T. Akizawa1,2 1O-Force Co., Ltd., Delopment, Kochi, Japan, 2Kochi University, Laboratory Of Pharmacology, School Of Medicine, Nankoku, Japan, 3Setsunan University, Department Of Integrative Pharmaceutical Science, Faculty Of Pharmaceutical Sciences, Osaka, Japan

Aims: The synthetic 9-mer peptide, JAL-TA9 (YKGSGFRMI) is identified the first Catalytide termed as the general name of the hydrolase peptide [1, 2]. JAL-TA9 cleaved not only authentic soluble form amyloid-β protein (Aβ42) but also solid type Aβ42 on the central region with a serine protease activity [3]. These data suggest that JAL-TA9 may be effective against Alzheimer’s disease with a new strategy. In this study, we evaluated the effects of JAL-TA9 to APP knock-in mice through the single direction dose into the both side CA1 of the hippocampus by Y-maze test. Methods: A dose of 2 uL JAL-TA9 (2.5 ug/uL) per side CA1 of the hippocampus of APP knock in-mice (C57BL/6-APP) was infused for 4 minieuts with a syringe. Y-maze test was conducted at before injection and after injection and alternation rate was calculated. Results: As a result, only after a week the alternation % of JAL-TA9 administration group was significantly higher than non-treat group and improved almost same level with normal mice. So that the spatial working memory was improved by JAL-TA9. Conclusions: We concluded that JAL-TA9 is the most attractive candidate for the fundamental drug for Alzheimer’s disease with new strategy. [1] Rina Nakamura, et. al., Peptide, 116: 71-77 (2019) [2] Rina Nakamura, et. al., Integra. Mol. Med., 6: 1-4 (2019) [3] Rina Nakamura, et. al., Journal of Royal Science., 1 (2) 30-35 (2019)

P212 / #1657

Topic: Theme A: β-Amyloid Diseases / A4.a. Imaging, Biomarkers, Diagnostics: Structural MRI, MR spectroscopy

ASSOCIATION BETWEEN MICROSTRUCTURAL WHITE MATTER ABNORMALITIES AND COGNITIVE FUNCTIONING IN PATIENTS WITH TYPE 2 DIABETES MELLITUS: A DIFFUSION TENSOR IMAGING STUDY

Lecture Title:

R. Abdelrazek1, A. Osama1, T. Hassan2, M. Negm1, H. Abou Elhamd1 1faculty of medicine, Suez Canal University, Neurology, Ismailia, Egypt, 2Suez canal university, Diagnostic Radiology, Ismailia, Egypt

Aims: To predict early cognitive dysfunction and detection of microstructural white matter changes in patients with type 2 DM by diffusion tensor imaging. Methods: A case control study included thirty patients aged ≥ 18 years old of both sexes with type 2 DM and thirty controls. All subjects underwent to Montreal Cognitive Assessment (MoCA) "Arabic version": to detect Mild Cognitive Impairment (MCI) and Diffusion Tensor Imaging study (DTI). Results: Mild Cognitive Impairment is related to type 2DM (56.7% of diabetic group), Reduced Fractional anisotropy (FA) values and elevated mean diffusivity (MD) values were related to cognitive impairment evaluated through Montreal Cognitive Assessment (MoCA) in patients with type 2 DM. Conclusions: The integrity of the white matter measured using DTI vary in MCI diabetics compared with non-MCI diabetics. Such changes have major implications on the cognitive function

P213 / #1785


COMPARISON OF THREE DIFFERENT METHODS FOR THE POST-HOC ANALYSIS OF LOCUS COERULEUS MAGNETIC RESONANCE IMAGING

Lecture Title:

A. Galgani1, N. Martini2, D. Della Latta2, A. Vatti2, H. Hlavata3, F. Baldacci1, D. Chiappino3, F. Fornai4,5, G. Siciliano1, F. Lombardo3, F. Giorgi1,4 1Pisa University Hospital, Neurology Unit, Pisa, Italy, 2G. Monasterio Foundation/National Research Council - Tuscany Region, Deep Health Unit, Pisa, Italy, 3G. Monasterio Foundation/National Research Council - Tuscany Region, Magnetic Resonance Imaging And Neuroradiology Unit, Pisa, Italy, 4University of Pisa, Department Of Translational Research And Of New Surgical And Medical Technologies, Pisa, Italy, 5IRCCS Neuromed, Neurobiology Of Movement Disorders, Pozzilli, Italy

Aims: To compare three post-processing approaches to analyze Locus Coeruleus (LC) magnetic resonance imaging features. Methods: 45 subjects (27M; 60-80yy) underwent LC-MRI scan, using a 2D-Fast Spin Echo (FSE) T1-weighted sequence (TE/TR 14/600 ms; flip angle 90°; voxel size 0.39x0.39x2.2 mm; NEX 5). Post-processing analysis was performed by a Semi-Automatic (SA), a Deep-Learning (DL), and a Template-Based (TB) approach. With SA approach, the most intense voxels automatically highlighted by the software in the LC area were selected by a trained operator. In the DL method, fully automatic segmentation of the two LC was performed through a supervised approach in the native space In the TB approach, a common template space was obtained performing a nonlinear coregistration of individual T1-w scans, where reference and LC regions were manually drawn. Then, LC voxels were automatically extracted from individual template-warped T1-w scans. Three different LC metrics were calculated: the normalized maximum intensity (LCmax), the mean intensity (LCmean), and the volume (LCvol) of the LC. Results: SA and DL methods analyzed LC signal in the native space while the TB method offered the advantage of a high-resolution template (0.5mm isotropic). We observed good agreement (r=0,70) between the three different post-processing approaches concerning LCmax. The moderate correlation (r=0.44) we observed for LCvol may be partly explained by the different resolution of native and template spaces. Conclusions: There is global agreement among LC-analysis in template or native space; the template-based approach shows the advantage of loco-regional analysis. Founding: Italian Ministry of Health, ricerca finalizzata #PE-2013-02359574 (P.I.: FSG).

P214 / #1564


DIFFERENCES IN CORTICAL STRUCTURE BETWEEN COGNITIVELY NORMAL EAST ASIAN AND CAUCASIAN OLDER ADULTS: A SURFACE-BASED MORPHOMETRY STUDY

Lecture Title:


Aims: There is a growing literature on the impact of ethnicity on brain structure and function. Despite the regional heterogeneity in age-related changes and non-uniformity across brain morphometry measurements in the aging process, paucity of studies investigated the difference in cortical anatomy between the East Asian and Caucasian older adults. The present study aimed to compare cortical anatomy measurements, including cortical thickness, volume and surface area, between cognitively normal East Asian (n = 171) and Caucasian (n = 178) older adults. Methods: The current study used semi-automated brain morphometry tools for evaluating cortical thickness, volume, and surface area. Vertex-wise statistical test was performed to compare the differences of these cortical anatomy measurements between East Asian and Caucasian groups, adjusting for age, gender, education, and MMSE score. Results: The East Asian group showed greater cortical thickness and larger cortical volume in the right superior temporal gyrus, postcentral gyrus, bilateral inferior temporal gyrus, and inferior parietal cortex. The Caucasian group showed thicker and larger cortex in the left transverse temporal cortex, lingual gyrus, right lateral occipital cortex, and precentral gyrus. Additionally, the difference in surface area was discordant with that in cortical thickness. Differences in brain structure between the East Asian and Caucasian might reflect differences in language and information processing, but further studies using standardized methods for assessing racial characteristics are needed. Conclusions: The research results represent a further step towards developing a comprehensive understanding of differences in brain structure between ethnicities of older adults, and this would enrich clinical research on aging and neurodegenerative diseases.

P215 / #1793


DEVELOPING NOVEL BIOLOGICAL MEASURES OF COGNITIVE RESERVE USING DEEP LEARNING

Lecture Title:

A. Marseglia, G. Mårtensson, J.-S. Muehlboeck, D. Ferreira, E. Westman Karolinska Institutet, Division Of Clinical Geriatrics, Center For Alzheimer Research, Department Of Neurobiology, Care Sciences And Society, Stockholm, Sweden

Aims: Although research showed that lifetime stimulating activities reduce the risk of dementia, measuring the underlying compensatory mechanisms has been challenging. We aimed to develop biological measures or reserve-related mechanisms (resilience/resistance) based on differences between predicted brain age (PBA) and the person’s chronological age (CA) using deep learning. Methods: The sample included 16,845 T1-weighted MRI from multiple time-points of healthy individuals from four cohorts: 1) UK Biobank (n=13882, aged 44-81 yrs.); ADNI (n=1489, aged 55-90 yrs.); AIBL (n=973, aged 60-93 yrs.); GENIC (n=501, aged 29-85 yrs.). Healthy status was defined as absence of dementia/cognitive impairment, neuro-psychiatric disorders, and self-reported good health. Cognitive impairment was also identified with a Mini-Mental State Examination score≤26. Medical diagnoses were collected through physician-/self-reports or medical records following the International Classification of Diseases criteria. MRIs were first labelled with the person’s chronological age. A deep learning model (DLM), based on convolutional neural networks, was trained to predict participants’ age from MRIs. The training-set included 80% of the UK Biobank/ADNI/AIBL/GENIC participants, whereas the validation-set included 10% of the remaining participants. Results: We trained the DLM and applied it to MRIs of people in the validation set. Our preliminary results show that the mean absolute error between predicted and chronological ages was 3.7 years, indicting accurate prediction of the brain age. Conclusions: Preliminary results show the potential of DLM to obtain a measure of brain resilience/resistance from MRI. In the next steps, we will test our DLM on MRIs from another longitudinal aging dataset to define resilience (PBA>CA) and resistance (PBA<CA) mechanisms.

P216 / #252


DEVELOPMENT OF AUTOMATED BRAIN VOLUMETRIC PROGRAM TO ASSIST THE DIAGNOSIS OF MILD COGNITIVE IMPAIRMENT AND ALZHEIMER’S DISEASE DEMENTIA

Lecture Title:

D.-W. Ryu, D.W. Yang College of Medicine, The Catholic University of Korea, Department Of Neurology, Seoul, Korea, Republic of

Aims: A quantitative analysis of brain volume can help to assess alzheimer's disease (AD) and other types of dementia. To validate the automated analysis program developed for volumetric measurements and to evaluate its performance in discriminating AD dementia (ADD) and mild cognitive impairment (MCI) patients from normal controls (NC). Methods: Based on T1-weighted MRI, regional volume measurements were conducted with the automated program named Quick Brain Volumetry (QBraVo). The validation of QBraVo was evaluated by intra-rater reliability and inter-rater reliability with a manual method. The discriminating performances to distinguish among ADD, MCI and NC were analyzed with receiver operating characteristic curves. The diagnostic performances were compared between MMSE, QBraVo, and the combination of both tools. Results: In total, 30 ADD patients, 30 MCI patients and 36 NCs were enrolled in this study. The average running time for single data analysis was 5 minutes and 36 seconds. Intra-rater reliability was 0.999. Inter-rater reliability as compared with manual measurements were high for total brain volume, total intracranial volume, and cerebrospinal fluid volume (R=0.97, 0.93 and 0.89, respectively) and superior to other volumetric methods. The medial temporal ratio showed the highest diagnostic performance for discriminating ADD from NC (area under the curve=0.901, sensitivity=80.0%, specificity=83.3%). Using combined MMSE and QBraVo could improve the diagnostic performance for ADD and MCI. Conclusions: Our results demonstrated that QBraVo could be an easy, fast, and accurate method to measure brain volume. The brain volumes measured by QBraVo could help to diagnosis ADD and MCI and assist in increasing diagnostic accuracy.

P217 / #830

Topic: Theme A: β-Amyloid Diseases / A4.b. Imaging, Biomarkers, Diagnostics: Functional MRI

STEPWISE CONNECTIVITY PAVES THE WAY TO REVEAL FUNCTIONAL NETWORK VULNERABILITY IN AGE-RELATED NEURODEGENERATIVE DISORDERS

Lecture Title:

S. Basaia1, C. Cividini2, E.G. Spinelli2, V. Castelnovo2, M. Leocadi2, D. Calderaro1, E. Canu1, F. Agosta2, M. Filippi3 1IRCCS San Raffaele Scientific Institute, Division Of Neuroscience, Milano, Italy, 2IRCCS San Raffaele Scientific Institute and Vita-Salute San Raffaele University, Division Of Neuroscience, Milano, Italy, 3IRCCS San Raffaele Scientific Institute; Vita-Salute San Raffaele University, Division Of Neuroscience; Unit Of Neurology, Milano, Italy

Aims: Ageing is the main risk factor for most neurodegenerative diseases and results in complex transformations of the human brain function. The aim of this study was to investigate how topological organization of the brain connectome changes with age using resting-state functional MRI and stepwise functional connectivity (SFC) analyses. Methods: 138 controls were recruited and divided into two groups according to age: 55 young (20-30 years [YC]) and 83 old (41-84 years [OC]). SFC analysis aims to characterize regions that connect to specific seed brain areas at different levels of link-step distances. Eight well-known hubs of the human connectome were selected as seeds: middle frontal gyrus, rostral anterior and posterior cingulate cortex, precuneus, inferior parietal, middle temporal and lingual gyri and pericalcarine cortex. Whole-brain two-sample t-test comparisons between groups were performed. Results: At one-link step distance, in OC, all the seed regions displayed decreased regional–local functional connectivity with superior frontal and medial orbital frontal gyri, rostral anterior and isthmus cingulate cortex, precuneus and middle and inferior temporal gyri relative to YC; across intermediate link-steps, a reduced connectivity was observed between all seed regions and frontal-parietal lobes. By contrast, at the first link-step distance, YC showed lower connectivity only between few seed regions and precentral, paracentral and lateral occipital gyri compared to OC. At intermediate link-step distances, increased connectivity with sensorimotor regions was found in OC relative to YC. Conclusions: SFC approach might have important implication providing a starting point for evaluating network disruptions in age-related neurodegenerative disorders. Supported by: European Research Council (StG-2016_714388_NeuroTRACK).

P218 / #1677

Topic: Theme A: β-Amyloid Diseases / A4.c. Imaging, Biomarkers, Diagnostics: PET – amyloid

SEX-RELATED DIFFERENCES IN AMYLOID-PET POSITIVITY AMONG SUBJECTIVE COGNITIVE DECLINE PATIENTS

Lecture Title:

M.Y. Chun1, G.H. Kim1, H.K. Park1, D.W. Yang2, Y.J. Hong3, S. Kim4, M.J. Wang5, K.H. Park6, S.H. Choi7, J.H. Jeong8 1Ewha Womans University Mokdong Hospital, Department Of Neurology, Seoul, Korea, Republic of, 2St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Department Of Neurology, Seoul, Korea, Republic of, 3Uijeongbu St. Mary's Hospital, Department Of Neurology, Gyeonggi-do, Korea, Republic of, 4Seoul National University Bundang Hospital, Department Of Neurology, Seongnam, Korea, Republic of, 5Roa Neurology Clinic, Department Of Neurology, Gyeonggi-do, Korea, Republic of, 6Gil Medical Center, Gachon University College of Medicine, Department Of Neurology, Incheon, Korea, Republic of, 7Inha University School of Medicine, Department Of Neurology, Incheon, Korea, Republic of, 8Ewha Womans University Seoul Hospital, Department Of Neurology, Seoul, Korea, Republic of

Aims: Subjective cognitive decline(SCD) is considered as preclinical Alzheimer’s disease(AD). Sex-related differences in cognitive decline and brain atrophy suggest risk factors such as age, cardiovascular factors and cortical thickness can have different effects according to sex. The aim of study is to demonstrate associations risk factors and amyloid PET results according to sex among amnestic SCD. Methods: We selected 120 amnestic SCD subjects aged 60 years or older in the range of 7-50% of the memory domain through neuropsychological tests. They also underwent body composite analysis, apolipoprotein E(APOE) genotype, MRI, and amyloid PET. We performed regression analyses in men and women separately to evaluate the associations variables and amyloid positivity. Results: 120 subjects consisted of 68 women and 52 men. Men had higher level of education, high-density lipoprotein cholesterol(HDLC), Framingham score, less visceral fat than women. Men exhibited higher visual memory and lower verbal memory scores than women. There was no difference in APOE4 between sex. In both men and women, APOE4 was related to amyloid positivity. Among men, age, HDLC, APOE4 and lower temporal anterior volume were associated with amyloid positivity. Among women, body mass index, weight circumference, visceral fat, APOE4 and visual memory were associated with amyloid positivity. Conclusions: This study analyzed risk factors of AD according to sex among amnestic SCD. Increased age, HDLC and APOE4 in amnestic SCD men are associated with amyloid positivity. Lower weight circumference and APOE4 are related in amnestic SCD women. Further studies need to prevent AD progression from the preclinical stage with sex-specific intervention.

P219 / #1767


PREDICTIVE SCALE FOR AMYLOID POSITIVITY BASED ON CLINICAL AND MRI VARIABLES IN PATIENTS WITH AMNESTIC MILD COGNITIVE IMPAIRMENT

Lecture Title:

M.Y. Chun1, G.H. Kim1, H.K. Park1, D.W. Yang2, S. Kim3, S.H. Choi4, J.H. Jeong5 1Ewha Womans University Mokdong Hospital, Department Of Neurology, Seoul, Korea, Republic of, 2St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Department Of Neurology, Seoul, Korea, Republic of, 3Seoul National University Bundang Hospital and Seoul National University College of Medicine, Department Of Neurology, Gyeonggi-do, Korea, Republic of, 4Inha University School of Medicine, Department Of Neurology, Incheon, Korea, Republic of, 5Ewha Womans University Seoul Hospital, Department Of Neurology, Seoul, Korea, Republic of

Aims: Early detection of Alzheimer's disease (AD) in mild cognitive impairment (MCI) stage is important because early intervention can prevent MCI from progression to AD dementia. The confirmatory diagnostic tools for AD such as amyloid positron emission tomography (PET) and cerebrospinal fluid β-amyloid (Aβ) studies have limitation for routine clinical usage. The study aims to develop a predictive scale for Aβ positivity using easily accessible tools such as neuropsychological tests, brain magnetic resonance imaging (MRI) and apolipoprotein E (APOE) genotype tests for routine clinical practice. Methods: We recruited 147 subjects who had memory impairment in either verbal or visual memory tests. All subjects underwent neuropsychological tests, brain MRI, APOE genotype test and amyloid PET. Medial temporal atrophy (MTA) was evaluated with axial visual rating scale (aVRS). We assessed he association between clinical variables and Aβ positivity with univariate and multivariate logistic regression analysis. We performed receiver operating characteristic (ROC) curve to determine the optimal model and developed a predictive scale with integers. With the ROC curve, the optimal cutoff score of the scale could be selected. Results: Seoul Verbal Learning Test-delayed recall (SVLT-DR), Rey Complex Figure Test-delayed recall (RCFT-DR), MTA aVRS and APOE genotype are associated with Aβ positivity in univariate associations. We developed a predictive scale with simple integers with 0.847 AUC value (95% CI 0.782-0.912). The optimal cutoff score was ≥3 with a sensitivity of 71.4% and a specificity of 87.0%. Conclusions: Amnestic MCI subjects with Aβ positivity can be identified using a predictive scale consisting of SVLT-DR, RCFT-DR, MTA aVRS and APOE genotype.

P220 / #1487


PERFORMANCE OF CSF AD BIOMARKERS IN PREDICTING AMYLOID PET POSITIVITY IN EARLY AD: DATA FROM EISAI’S MISSIONAD PROGRAM

Lecture Title:

J. Kaplow1, M. Kanekiyo1, D. Verbel1, C. Roberts2, M. Gee2 1Eisai Inc, Nbg, Woodcliff Lake, United States of America, 2Eisai Inc, Nbg, Hatfield, United Kingdom

Aims: Eisai’s MissionAD program comprised two global Phase 3 studies investigating elenbecestat (BACE inhibitor) in early AD. Eligibility required amyloid burden confirmation by CSF and/or amyloid PET. In this analysis data from subjects with both modalities were evaluated to determine the best performing CSF measure(s) for concordance with amyloid PET positivity. Methods: Baseline amyloid PET scans were read visually (per approved tracer labels). Aβ(1-42), Aβ(1-40), total tau and ptau181 were measured in subjects with baseline CSF (Fujirebio’s Lumipulse). Additional covariate baseline analysis included neurogranin and neurofilament light, performed using ELISA and Simoa technology, respectively. Cutoffs were evaluated by receiver operator curves using Youden’s index for individual analytes and ratios. Results: 526 subjects provided both baseline amyloid PET (florbetaben 84%, flutemetamol 11%, florbetapir 5%) and CSF assessments. The AUC of CSF Aβ(1-42), total tau, and ptau181 were 0.81, 0.83, 0.86; the overall percent agreement (OPA) of each analyte was 71%, 78%, 82%, respectively. CSF ptau181/Aβ(1-42), Aβ(1-42)/Aβ(1-40), and total tau/Aβ(1-42) ratios resulted in AUCs of 0.90, 0.89, 0.90 and OPAs of 90%, 85%, 89%, respectively demonstrating better performance. Flutemetamol performance was lower than the other tracers for Aβ(1-42)/Aβ(1-40) ratio. Inclusion of covariates did not improve performance of any of the CSF biomarker ratios. Conclusions: CSF biomarker ratios have greater concordance with visually-read amyloid PET positivity than single biomarkers. Both ptau181/Aβ(1-42) and total tau/Aβ(1-42) demonstrated the highest overall performance.

P221 / #1646


FLUTAMETAMOL PET SCANS IN COGNITVELY NORMAL APOE4 HETEROZYGOTES- RESULTS FROM GENERATION 2 ALZHEIMER’S DISEASE PREVENTION TRIAL AT GLASGOW MEMORY CLINIC

Lecture Title:

J. Lynch1, F. Inglis1, V. Mclean1, L. Wallace1, A. Cranmer1, E. Van Beek2, D. Martin2, D. Brian2, A. Fletcher2, C. Lucatelli2 1Glasgow memory clinic, Research, Motherwell, United Kingdom, 2Edinburgh Imaging facility QMRI, Radiology, Edinburgh, United Kingdom

Aims: The Generation 2 programme co -sponsored by Novartis, Amgen, and the Banner Institute, investigated the effects of experimental medication on AD related biomarkers including amyloid deposition measured by PET- CT. Here we focus on the PET- CT scans we conducted at Edinburgh QMRI, prior to the early cessation of this clinical trial. Methods: Cognitively normal subjects who were previously identified as having at least one copy of the APOE e4 genotype were invited to participate in the Generation programme. To be eligible for randomisation subjects also had to demonstrate elevated amyloid level as determined by PET scan or Lumbar puncture. Results: 424 subjects were screened .190 reached the biomarker testing stage and had a 18F-flutemetatmol PET -CT study, which is a marker for amyloid deposition. 78 subjects were screen failed due to early cessation of the programme before reaching biomarker assessment. Age range 61-76 years, 250 female, average age 68yrs, 174 male, average age 69yrs. The PET – CT scanning results showed 65 (34%) were deemed amyloid positive and 125 (66%) were deemed negative, as per protocol. Conclusions: As anticipated the majority of screen fails for this programme were at amyloid testing stage. A third of cognitively normal APOE4 heterozygotes were positive for amyloid by PET assessment. The use of 18F-flutematamol PET-CT imaging can facilitate identification of patients most likely to benefit from novel treatments.

P222 / #399


UNIQUE REGIONAL PATTERNS OF AMYLOID BURDEN PREDICT PROGRESSION TO PRODROMAL AND CLINICAL STAGES OF ALZHEIMER’S DISEASE.

Lecture Title:

J. Pfeil1, M. Hönig1,2, E. Doering1,3, T. Van Eimeren1,3,4, A. Drzezga1,2,3, G. Bischof1, For The Alzheimer'S Disease Neuroimaging Initiative1 1University of Cologne, University Hospital of Cologne, Nuclear Medicine, Multimodal Neuroimaging Group, Köln, Germany, 2Research Center Jülich, Neuroscience And Medicine Ii, Jülich, Germany, 3German Center for Neurodegenerative Diseases, Positron Emission Tomography (pet), Bonn, Germany, 4University of Cologne, Neurology, Köln, Germany

Aims: Given the low predictive power of global amyloid-beta status for conversion from cognitively-normal (CN) to mild cognitive impairment (MCI) or Alzheimer’s disease (AD), we examined the prognostic potential of regional amyloid-beta burden compared to global amyloid-beta, CSF, genetic-, and cognitive measures. Methods: Data from ADNI (http://adni.loni.usc.edu/) of four age-matched amyloid-beta-positive groups were included: cognitively stables (CN-CN, n=38), CN to MCI/AD converters (CN-MCI/AD, n=38), MCI stables (MCI-MCI, n=104), and MCI to AD converters (MCI-AD, n=104). For all, a baseline 18F-florbetapir scan, neuropsychological assessments, CSF, and clinical follow-up information were available. 18F-florbetapir scans were pre-processed using SPM12 including normalization and intensity-standardization (reference: cerebellum). To assess regional differences in amyloid-beta burden, voxel-wise group-wise comparisons (FWE-corrected) were performed between CN-CN vs. CN-MCI/AD and MCI-MCI vs. MCI-AD, respectively. Mean amyloid-beta burden of resulting significant clusters was subsequently used in logistic regression analyses together with CSF, genetic-, and cognitive measures to determine the best predictive biomarker for conversion. Results: CN to MCI/AD conversion was predicted by amyloid-beta burden in the precuneus, lingual- and angular gyrus, and middle- and superior temporal gyrus. MCI to AD conversion was predicted by amyloid-beta in the anterior cingulate gyrus, middle- and superior temporal gyrus, precuneus, medial frontal cortex, posterior- and anterior insula, and central operculum. APOE4 carriership was a significant risk factor for conversion from CN to MCI/AD, while a composed ADNI-memory score predicted conversion from MCI to AD. Conclusions: Regional amyloid-beta burden is a more sensitive predictor of conversion to MCI or AD than global amyloid-beta, pointing towards its usefulness as prognostic biomarker.

P223 / #1239


CLINICALLY BASED PREDICTION OF AMYLOID PET SCAN OUTCOMES IN COGNITIVELY NORMAL ELDERLY

Lecture Title:

L. Schneider1, A. Yau1, G. Cutter2, T. Goldberg3, R. Kennedy2, W. Mack1 1University of Southern California, Keck School Of Medicine, Los Angeles, United States of America, 2University of Alabama, Birmingham, School Of Public Health, Birmingham, United States of America, 3Columbia University Medical Center, Psychiatry And Anesthesiology, New York, United States of America

Aims: Although most elderly people with dementia have positive Aβ PET scans over 30% of cognitively unimpaired also do. Demographic and clinical predictors of a positive Aβ PET scan in this population are uncertain. We describe a multivariable prediction model that uses readily available demographic and mainly self-assessed clinical characteristics to predict Aβ PET outcomes, and perhaps consequently greater or lesser risk for MCI or dementia. Methods: 4465 cognitively unimpaired people, ages 65 to 85 years, with similar demographic and clinical characteristics seeking entry into a randomized clinical trial of solanezumab for preclinical AD (the A4 trial) received Aβ PET scans. A total of 1223 had a positive scan (SUVR ≥ 1.15) and therefore fulfilled criteria for ‘preclinical Alzheimer disease.’ Potential predictors including demographics, APOE genotype, family history of dementia, clinical and cognitive ratings; and self-assessed cognitive function (CFI), ADLs, concerns about AD, and their perspective on their future, were modeled using multivariable logistic regression. Results: Increasing age and APOE4 genotype together predicted Aβ-PET positivity (PPV = 45.41%, NPV = 85.39%, AUC=0.7395 (95% CI: 0.7231, 0.7559)). Model discrimination was improved about 2% after race (white), family history, delayed recall, CFI, and concerns about AD were added. Sensitivity analyses and validation results will be detailed as well. Conclusions: The above predictors are easily obtained and, together, are substantially predictive of Aβ PET outcomes; and may inform the clinical need and screening yield for an Aβ PET scan. Funding: NIH P30 AG066530, R01 AG057684, R01 AG051346, P50 AG05142; California DHS grant 15-10291

P224 / #1360


SLEEP DYSFUNCTION, MEASURED OBJECTIVELY, CORRELATES WITH CEREBRAL AMYLOID DEPOSITION.

Lecture Title:

C. Tissot1, H. Blais2, C. Thompson2, A. Benedet1, T. Pascoal1, J. Therriault1, M. Chamoun1, F. Lussier1, S. Gauthier1, N. Gosselin2, P. Rosa-Neto1 1The McGill University Research Centre for Studies in Aging, Translational Neuroimaging Laboratory, Montreal, Canada, 2Université de Montréal, Center For Advanced Research In Sleep Medicine, Montreal, Canada

Aims: Sleep problems are common along the Alzheimer’s disease (AD) spectrum. CSF Aβ correlates with poor sleep quality through self-reported questionnaires in cognitively unimpaired (CU) individuals, while AD patients show tau-PET increase. Accumulating evidence proposes that sleep disturbances are an early marker for AD pathology and of risk of cognitive impairment. Our objective is study the relationship between sleep efficiency as assessed with polysomnography and AD biomarkers: amyloid, tau and neuroinflammation. Methods: 28 non-demented individuals underwent [18F]AZD4694 amyloid-PET, [18F]MK6240 tau-PET, [11C]PBR28 neuroinflammation-PET scans, MRI, neuropsychological evaluation and a polysomnography. [18F]AZD4694, [18F]MK6240 and [11C]PBR28 standardized-uptake-value-ratios used the cerebellar grey as the reference region and were calculated 40-70 min, 90-110 min and 0-90 min post-injection respectively. Voxel based regression models evaluated the relationship between AD pathophysiologies, amyloid, tau and neuroinflammation, and sleep efficiency. Model’s covariates included age and diagnosis. Results: We found a negative correlation between sleep efficiency and [18F]AZD4694. The impacted regions were the temporal poles, inferior parietal cortex and subcortical regions as the thalamus. However, no correlation was found with [18F]MK6240 nor [11C]PBR28. When corrected for apnea-hypopnea-index, sex and education years, results were similar.

Conclusions: These preliminary results show poor sleep quality correlates with amyloid deposition in CU and MCI. The regions impacted are related to sleep regulation, such as the thalamus. Others are known to be vulnerable to AD pathophysiology, as temporal and inferior parietal cortices. Our study adds to previous research by identifying anatomical correlates. It uses objective assessment of sleep quality, and corroborates the idea that sleep deprivation promotes Aβ deposition.

P225 / #684


PLASMA PYROGLUTAMATE-MODIFIED AMYLOID BETA DIFFERENTIATES AMYLOID PATHOLOGY

Lecture Title:

C.S.Y. Yang1, P.-N. Wang2, K.-J. Lin3, U. Andreasson4, K. Blennow5, H. Zetterberg6 1MagQu Co., Ltd., President Office, New Taipei City, Taiwan, 2Taipei Veterans General Hospital, Department Of Neurology, Taipei, Taiwan, 3Linkou Chang Gung Memorial Hospital, Department Of Radiology, New Taipei City, Taiwan, 4Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden, 5Univ. of Gothenburg, Inst Of Neuroscience And Phsyiology, Mölndal, Sweden, 6UCL Institute of Neurology, Department Of Neurodegenerative Disease, London, United Kingdom

Aims: AβpE3 could be a biomarker specific for Aβ plaque pathology in the brain. The concentration of AβpE3 in human body fluid is extremely low. An ultra-high-sensitive assay technology is needed for detecting AβpE3-40 in human body fluids. The measured concentrations of plasma AβpE3-40 of these subjects were compared with amyloid PET. Methods: Immunomagnetic reduction was utilized for quantification of pyroglutamate-modified amyloid β 3-40 (AβpE3-40) in human plasma. Forty-six subjects aged more than 55 years were enrolled. Magnetic resonance (MR) imaging was used to exclude subjects with major neuropathologies. Patients with AD and aMCI patients follows criteria of National Institute on Aging and the Alzheimer's Association (NIA-AA). The 18F-florbetapir PET (Aβ PET) images were acquired for each enrolled subject. Results: The lower limit of detection of assaying AβpE3-40 using IMR was 5 fg/ml, while the upper limit of detection was 100 pg/ml. No significant interference by Aβ3-40 (≦ 20 pg/ml), Aβ1-40 (≦ 80 pg/ml), or Aβ1-42

(≦ 20 pg/ml) was observed. AβpE3-40 concentration in human plasma was 44.1 ± 28.2 fg/ml in PET- (n = 28) and 91.6 ± 54.6 fg/ml in PET+ (n = 18) (p < 0.05). The cut-off value of plasma AβpE3-40 for discriminating PET- from PET+ was 55.5 fg/ml, resulting in a sensitivity of 83.3%, a specificity of 71.4% and an AUC of 0.808. The concentration of plasma AβpE3-40 showed a moderate correlation (r = 0.437) with PET SUVR. Conclusions: These results reveal the feasibility of detecting Aβ pathology using quantification of a plaque-derived Aβ molecule in plasma.

P226 / #1319

Topic: Theme A: β-Amyloid Diseases / A4.d. Imaging, Biomarkers, Diagnostics: PET – glucose

ESTABLISH THE CORRELATION BETWEEN THE FUNCTIONAL, ANATOMIC AND CSF-BIOMARKERS, INTHE NEURODEGENERATIVE DETERIORATION OF REFERRED PATIENTS TO THE PET/UNIT

Lecture Title:

J.M. Nogueiras Alonso1, C. Martnez Ramos1, V. Rodríguez Morales2, G. Castillo Simón1 1Hospital do Meixoeiro, Nuclear Medicince, Vigo, Spain, 2Hospital do Meixoeiro, Nuclear Medicince, Tui, Spain

Aims: Asses the correlation between the results of 18F-FDG-PET/CT, SPECT, brain-MR and CSF in thediagnosis of neurodegenerative dementia Methods: Retrospective review (March-November 2018) of cerebral 18F-FDG-PET/CT scans by two double-blindnuclear physicians, conducted in 54 patients(p) studied for possible neurodegenerative deterioration and itscorrelation by groups with the results of anatomical (NMR), functional-SPECT- and biochemical tests (TAU-t/TAU-p &β-amyloid). 18F-FDG-PET/CT acquired at 30-45' injection intravenous of 180MBq (18F-FDG), by brain PET locatedof 15 'duration Results: 54p with 18F-FDG-PET/CT, 57%♂, 43%♀, age: 63a [29-83]. Reason for consultation: mild cognitiveimpairment-MCI-(7p), FTD(11p), AD(8p), subjective memory complaints-SMC-(4p). In 51(94%)patients 18F-FDG-scans was pathological result: 68.5% anterior pattern and 22% posterior pattern -CSF-biomarkers and 18-FDG-PET/TC MATCH in 85.2% (3p normal pattern,37p-FTD, 6-AD) and 8p (14.8%) with notcoincidence. -Only 44p(81%) with MRI, 11% (6p) with hippocampal atrophy concordant with hipometabolism at 18F-FDG-PET/CT. -36p(67%) had SPECT 75% (27p) was pathological and coincident with 18F-FDG-PET/CT 55%(20p). In no case was there a triple coincidence 18F-FDG-PET/TC-NMR and CSF, possibly due to the poor findings of theMRI, influenced by acquisition with a 1.5T device Conclusions: Anterior pattern was the most frequent, it´s porbably related to the bias generated from the Neurology consultation.We found a good correlation between our results and the CSF biomarker, like the tendency to rely more on theclinical, neuropsicological and PET tests, reserving the CSF for difficult diagnosis. Better correlation with MRI, can getwith better hippocampal volumetry (how we are observing with MR-3T).

P227 / #614

Topic: Theme A: β-Amyloid Diseases / A4.g. Imaging, Biomarkers, Diagnostics: Multimodal imaging

MULTI-COMPARTMENT DIFFUSION MRI MEASURES CAPTURE MICROSTRUCTURAL ASSOCIATIONS WITH AMYLOID AND TAU PET MEASURES IN THE CORTEX

Lecture Title:

T. Nir1, J.E. Villalon-Reina1, A. Zhu1, S. Thomopoulos1, P. Maiti1, L. Salminen1, R. Reid2, M. Bernstein2, B. Borowski3, C. Jack3, M. Weiner4, P. Thompson1, N. Jahanshad1 1University of Southern California, Imaging Genetics Center, Mark & Mary Stevens Neuroimaging & Informatics Institute, Keck School Of Medicine, Marina del Rey, United States of America, 2Mayo Clinic and Foundation, Department Of Information Technology, Rochester, United States of America, 3Mayo Clinic and Foundation, Department Of Radiology, Rochester, United States of America, 4University of California San Francisco School of Medicine, Department Of Radiology, San Francisco, United States of America

Aims: Multi-shell diffusion MRI (dMRI) biophysical models can estimate intracellular (ICVF), extracellular (ECVF) and free water (ISOVF) volume fractions (VFs) in brain microstructure. These microstructural measures can be calculated in the cortex using a gray matter specific three compartment model that may be sensitive to hydrophobic aggregates of intracellular tau and extracellular amyloid, the pathological hallmarks of Alzheimer’s disease. Here, we evaluated relationships between regional PET tau and amyloid accumulation and cortical microstructural VFs. Methods: Multi-shell dMRI and FAV1451, FBB, or FAV45 PET data were available for 98 ADNI-3 participants (age: 72.1±6.4 yrs; 43 M). Cortical ICVF, ECVF and ISOVF were estimated with multi-tissue SMT using a gray matter specific response function. Mean cortical dMRI and PET measures were extracted from 4 lobes and the cingulate cortex. Random-effects linear regressions tested associations between regional dMRI measures and tau or amyloid burden, adjusting for age, sex, education, and grouping by scanning site. Analyses were also conducted within cognitively impaired or normal (CI/CN) and amyloid positive or negative (Aβ+/Aβ-) subgroups.

Results: are reported in Figures 1-2.

Conclusions: These results are preliminary and, as many factors influence dMRI measures, should be interpreted with caution. Greater amyloid and tau with higher ISOVF and lower ICVF suggest neuronal loss, particularly in CI and Aβ+ participants. Moreover, in CI individuals, lower ECVF with greater amyloid may reflect accumulation of large amyloid plaques in the extracellular space. In Aβ- individuals, higher ICVF with greater tau may reflect, for example, accumulation of phosphorylated tau along microtubules before axonal instability and degeneration.

P228 / #1597


MULTIMODAL NEUROIMAGING OF THE MOUSE CHOLINERGIC PROJECTION SYSTEM

Lecture Title:

K. Onuska1, H. Shanks1, M. Fox1, J. Hicks1, V. Prado2, J. Thiessen1,3, M. Prado1,2, T. Schmitz1 1University of Western Ontario, Schulich Medicine And Dentistry, London, Canada, 2The University of Western Ontario, Robarts Research Institute, London, Canada, 3Lawson Health Research Institute, Imaging, London, Canada

Aims: Cholinergic neurons become damaged early on in Alzheimer’s disease (AD). However, degeneration of cholinergic neurons is invisible to structural magnetic resonance imaging (sMRI). Positron emission tomography (PET) with the [18F] FEOBV radiotracer overcomes this problem. [18F] FEOBV binds to the vesicular acetylcholine transporter (VAChT), a protein found in cholinergic nerve terminals, thereby providing a cell-type specific measure of the cholinergic projectome. However, the sensitivity and specificity of in vivo [18F] FEOBV–PET to the endogenous expression of VAChT is unknown. This has created a major obstacle for assessing the utility of [18F] FEOBV-PET as a preclinical biomarker for AD. The goal of this study is therefore to validate [18F] FEOBV-PET as a cell-type specific tool for measuring cholinergic integrity by evaluating the binding intensity patterns and uptake profiles of [18F] FEOBV in wild type mice (WT) and a knockdown (KD) mouse model with 60% global brain reductions in VAChT expression (VAChTHOMKD). Methods:

We acquired high-resolution sMRI and [18F] FEOBV-PET images in 4-month-old WT and VAChTHOMKD mice. These images were then preprocessed using MATLAB’s SPM toolbox. Results:

First, we demonstrated that [18F] FEOBV-PET binding patterns reflect the underlying anatomy of the mouse cholinergic projectome in both WT and VAChTHOMKD mice.

Next, we established that [18F] FEOBV uptake is decreased within regions that receive cholinergic input in the VAChTHOMKD mouse when compared to the WT mouse. Conclusions: Overall, these preliminary results support [18F] FEOBV-PET as a sensitive in vivo cell-type specific tool for measuring cholinergic integrity that could be used as a preclinical biomarker for AD.

P229 / #1419


DOPAMINERGIC AND SEROTONERGIC PATHWAYS IN ALZHEIMER’S DISEASE: A 123I-FP-CIT STUDY

Lecture Title:

A. Pilotto1, S. Caminiti2, C. Liguori3, L. Presotto2, V. Garibotto4, A. Sala2, R. Turrone1, S. Caratozzolo1, A. Scalvini1, L. Rozzini1, M. D'Amelio3, N. Mercuri3, B. Paghera5, E. Premi1, M. Rizzetti6, A. Stefani3, A. Padovani1, D. Perani2 1university of Brescia, Department Of Clinical And Experimental Sciences, Brescia, Italy, 2Vita-Salute San Raffaele University, Nuclear Medicine, Milano, Italy, 3University of Rome 'Tor Vergata", Sleep Center, roma, Italy, 4University of Geneve, Nuclear Medicine, geneve, Switzerland, 5university of brescia, Nuclear Medicine, brescia, Italy, 6FERB ONLUS, Department Of Parkinson's Disease Rehabilitation, Trescore Balneario, Italy

Aims: to evaluate extrastriatal dopaminergic and serotonergic pathways in AD patients by using a 123I-FP-CIT SPECT imaging. Methods: Alzheimer’s disease patients were included in a multicenter study and underwent a standardized neurological examination, structural imaging and CSF, amyloid and metabolism imaging in order to reach a biomarker diagnosis of AD (i.e. A+T+N+ classification). Each patient underwent 123I-FP-CIT SPECT imaging and the bindings of extrastriatal regions of interests were calculated from spatially normalized images. The occipital-adjusted specific to non-displaceable binding (SBR) in the different regions was compared between AD and controls adjusting for the effect of age, sex, disease duration and serotonergic/dopaminergic treatment. Results: 52 AD patients A+T+N+ and 75 controls entered in the study. AD patients showed lower 123I-FP-CIT SPECT SBR in cingulate (p=0.001) and temporal lobe (p=0.007) as well as in insula (p=0.01) and thalamus (p=0.025) compared to controls. When dividing AD according to severity, MCI due to AD (n=17) showed significantly lower parietal SBR compared to controls (p=0.002) and significantly higher SBR in insula (p=0.01), thalamus (p=<0.001) and temporal lobe (p=0.008) compared to AD dementia (n=35). Conclusions: We demonstrated extrastriatal dopaminergic and serotonoergic impairment in Alzheimer’s disease. In the parietal cortex the alterations was present already at MCI stage, while in the temporal regions, thalamus and insula alterations appeared later in the AD dementia stage. Longitudinal studies will be necessary in order to evaluate the clinical value of extrastriatal 123I-FP-CIT SPECT assessment for possible different pattern of progression and response to treatment in AD patients.

P230 / #702


THE VIRTUAL BRAIN CLOUD – TOWARDS PERSONALIZED MULTI-SCALE BRAIN SIMULATION USING NEUROIMAGING COHORTS FOR ALZHEIMER’S DISEASE RESEARCH

Lecture Title:

L. Stefanovski1, R. Pai1, K. Bülau1, L. Martin1, M.-A. Diaz-Cortes1, A. Kapsis1, P. Triebkorn1, D. Perdikis1, T. Lett1, J. Meier1, A. Solodkin2, L. Alberi Auber3, M. Mohajerani4, V. Jirsa5, A. Mcintosh6, A. Kodamullil7, M. Hofmann-Apitius7, P. Ritter1 1Charité - Universitätsmedizin Berlin, Brain Simulation Section, Department Of Neurology, Berlin, Germany, 2University of Texas in Dallas, Behavioral And Brain Sciences, Dallas, United States of America, 3Swiss Integrative Center for Human Health, Sichh, Fribourg, Switzerland, 4University of Lethbridge, Canadian Center For Behavioural Neuroscience, Lethbridge, Canada, 5Aix Marseille Université, Institut De Neurosciences Des Systèmes, Marseille, France, 6Baycrest Health Sciences, Rotman Research Institute, Toronto, Canada, 7Fraunhofer Institute for Algorithms and Scientific Computing SCAI, Department Of Bioinformatics, Sankt Augustin, Germany

Aims: We aim to present a conceptual framework of the large-scale EU consortium VirtualBrainCloud, that combines big data science, computational linguistics, and personalized multi-scale brain network simulation for mechanistic predictive disease modeling to improve prevention, diagnostics and treatment of Alzheimer’s Disease (AD). Methods: To achieve interoperability and standardization, we first transfer brain imaging and selected associated data of the Alzheimer’s Disease Neuroimaging Initiative (ADNI) into Brain Imaging Data Structure (BIDS) format. This allows for automated image processing with BIDS Apps developed by our team. Second, we apply computational linguistics to characterize relationships between knowledge available in existing knowledge bases (as molecular signaling cascades) and ADNI data. Third, we annotate available mathematical dynamic models of The Virtual Brain simulation platform (TVB, www.thevirtualbrain.org) with explanatory information regarding the biological processes they represent and build an ontology of the available model variables and parameters. Results: We subsequently propose quantitative relationships (coupling functions) between relevant endpoints of biological signaling cascades taken from knowledge bases and the TVB model parameters. We identify candidate quantitative “Cause-and-effect” models how alterations on the subcellular level modify parameter values of the local neural mass model and thus the resulting neural dynamics. Brain simulation outputs are mapped back to empirical ADNI data for empirical falsification. We describe how this approach can be utilized in AD-relevant clinical studies. Conclusions: Our framework allows the identification of overlapping entities between TVB dynamics and existing knowledge inventories about potential disease mechanisms. This enables multi-modal and multi-scale AD-linked brain simulations on the basis of large cohort databases like ADNI.

P231 / #274

Topic: Theme A: β-Amyloid Diseases / A4.h. Imaging, Biomarkers, Diagnostics: CSF, blood, body fluid biomarkers

A PROMISING DIAGNOSTIC TOOL: DIFFERENTIALLY METHYLATED POSITIONS OF PERIPHERAL BLOOD LEUCOCYTES BETWEEN ALZHEIMER'S DISEASE PATIENTS AND CONTROLS.

Lecture Title:

B. Acha Santamaría1,2, I. Blanco-Luquin1,2, J. Corroza3, M. San Miguel3, S. Zueco4, A. Urdanoz-Casado1,2, M. Roldan1,2, C. Cabello3, I. Elizalde Beiras5,6,7, J. Sánchez Ruiz De Gordoa3, M. Mendioroz1,2,3 1IDISNA, Neuroepigenetica, Pamplona, Spain, 2Navarrabiomed Biomedical Research Center, Neuroepigenetica, Pamplona, Spain, 3Complejo Hospitalario de Navarra, Neurology Department, Pamplona, Spain, 4Complejo Hospitalario de Navarra, Medicina Interna, Pamplona, Spain, 5Osasunbidea, Gerencia De Atención Primaria, Pamplona, Spain, 6Universidad Pública de Navarra, Idisna, Pamplona, Spain, 7IDISNA, Primary Care,servicio Navarro De Salud - Osasunbidea, Pamplona, Spain

Aims: A great effort is currently being made to find non-invasive and cost-effective diagnostic tools. So we chose blood as a source of harmless biomarkers for diagnosis of AD to explore epigenetics biomarkers. As a first approach, our aim was to find differentially methylated genes in the DNA of leukocytes in the peripheral blood of patients with AD compared with controls. Methods: This study is a case-control study in which blood was collected in EDTA tubes from 45 patients with probable AD dementia based on the NIA-AA criteria and 45 age- and sex- matched healthy controls. DNA was isolated from the buffy coat using FlexiGen DNA Kit (Qiagen) and bisulfite treated. We analyzed differentially methylation positions (DMP) from BIN1, HAND2, HOXB6, SIX3 and HOXA3 genes, selected after a comprehensive bibliographic search and our previous results in brain tissue. The percentage of methylation of each DMP was measured by pyrosequencing. Mann–Whitney U Test was used to compare the methylation level between both groups. Results: Differences were found between AD patients group and controls group in two DMPs, corresponding to SIX3 and HOXA3 genes, [median (IQR) 64.91 (8.03) in AD vs. 66.67 (6.50) in controls, p-value<0.05] and [92.21 (1.34) in AD vs. 90.37 (1.05) in controls; p-value< 0.05], respectively. No significant differences in DMPs from BIN1, HAND2 and HOXB6 were detected between AD patients and controls. Conclusions: These results suggest that these genes may be useful in the diagnosis of AD as blood-based biomarkers and are worth exploring in larger AD cohorts.

P232 / #566


PLASMA BIOMARKERS IN THE AT(N) CATEGORIES FOR THE DETECTION OF ALZHEIMER’S DISEASE

Lecture Title:

D. Alcolea1,2, C. Delaby2,3, L. Muñoz1,2, S. Torres1,2, T. Estellés1,2, N. Zhu1,2, I. Barroeta1,2, M. Carmona-Iragui1,2, I. Illán-Gala1,2, M. Santos1,2, M. Altuna1,2, I. Sala1,2, M.B. Sánchez-Saudinós1,2, L. Videla1,2, S. Valldeneu1,2, A. Subirana1,2, C. Hirtz3, J. Vialaret3, S. Lehmann3, K. Blennow4,5, H. Zetterberg4,5,6,7, O. Belbin1,2, R. Blesa1,2, J. Clarimon1,2, J. Fortea1,2, A. Lleó1,2 1Centro de Investigación Biomédica en Red en Enfermedades Neurodegenerativas, Ciberned, Madrid, Spain, 2Institut d’Investigacions Biomèdiques Sant Pau - Hospital de Sant Pau, Universitat Autònoma de Barcelona, Sant Pau Memory Unit, Department Of Neurology, Barcelona, Spain, 3IRMB, INM, Université de Montpellier, INSERM, CHU de Montpellier, Laboratoire De Biochimie-protéomique Clinique, Montpellier, France, 4Sahlgrenska University Hospital, Clinical Neurochemistry Laboratory, Mölndal, Sweden, 5Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden, 6UCL Institute of Neurology, Department Of Neurodegenerative Disease, London, United Kingdom, 7UCL, Uk Dementia Research Institute, London, United Kingdom

Aims: To compare levels of plasma Aβ42/Aβ40, pTau181 and neurofilament light chain (NfL) between AT(N) categories in neurodegenerative dementias. We evaluated their correlation with levels in CSF and assessed their single and combined accuracy to detect Alzheimer’s disease (AD). Methods: We measured Aβ42/Aβ40, pTau181 and NfL in plasma from 150 participants of the Sant Pau Initiative for Neurodegeneration (SPIN). Participants had a diagnosis of AD, Lewy body dementia, frontotemporal lobar degeneration or were cognitively normal and were classified as A+T+, A+T-, A-T+ or A-T- according to CSF Aβ42/Aβ40 and CSF pTau181 validated cutoffs. Amyloid species in plasma were measured by immunoprecipitation coupled with mass spectrometry. Levels of plasma pTau181 and NfL were quantified using Single Molecule Array. Results: After adjusting by age and multiple comparisons, the A+T+ group had lower Aβ42/Aβ40 (p<0.001) and higher levels of pTau181 (p<0.001) compared to the A-T-. There were no significant differences in NfL between groups (p=0.92, Figure 1). Aβ42/Aβ40, pTau181 and NfL in plasma correlated with their measures in CSF across categories (Rho=0.36, Rho=0.51 and Rho=0.78, respectively, all p<0.001). In the ROC analysis, pTau181 and its combination with Aβ42/Aβ40 yielded the greatest areas under the curve, 0.82 (95%CI 0.75-0.90) and 0.85 (95%CI 0.78-0.92), respectively, to discriminate participants in the A+T+ category. The accuracy increased to 0.90 (95%CI 0.85-0.96) when age and APOE status were included (Figure 2).

Conclusions: Plasma markers of the AT categories are useful to detect participants with CSF pathophysiological evidence of AD, while N markers lack sufficient accuracy.

P233 / #1264


CEREBROSPINAL FLUID AΒ42 AND AΒ40 AND THEIR RELATION TO SOLUBLE AND INSOLUBLE AΒ IN THE BRAIN IN ALZHEIMER’S DISEASE

Lecture Title:

E. Andersson1, K. Blennow2,3, H. Zetterberg2,3,4,5, O. Hansson1,6 1Lund University, Clinical Memory Research, Lund, Sweden, 2Sahlgrenska University Hospital, Clinical Neurochemistry Laboratory, Mölndal, Sweden, 3Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden, 4UCL Institute of Neurology, Department Of Neurodegenerative Disease, London, United Kingdom, 5UCL, Uk Dementia Research Institute, London, United Kingdom, 6Skåne University Hospital, Memory Clinic, Malmö, Sweden

Aims: In patients with Alzheimer’s disease (AD), the concentration of Aβ42 in cerebrospinal fluid (CSF) is reduced while Aβ40 remains unchanged. It has been proposed that the reduced Aβ42 concentration is due to aggregation of this peptide into insoluble plaques, resulting in less soluble Aβ42 available for transport to the CSF. However, studies investigating how changes in soluble and insoluble Aβ42 and Aβ40 in the brain relate to the concentrations of these Aβ peptides in CSF are limited. Methods: CSF and brain tissue was collected from the 5xFAD mouse model of AD at 2, 4, 6, and 12 months (n=45). The concentrations of CSF Aβ42 and Aβ40 were measured using the Single Molecule Array (Simoa) technology. Soluble brain fractions were prepared and the concentrations of Aβ42 and Aβ40 were determined using the Meso Scale Discovery platform. Furthermore, immunohistochemistry was performed on brain sections using antibodies specific for Aβ40 and Aβ42. Results: The concentration of CSF Aβ42 was reduced over time whereas no changes of CSF Aβ40 were found (Fig1). Both Aβ42 and Aβ40 were deposited in insoluble plaques from 2 months of age (Fig2), however the concentrations of soluble Aβ42 and Aβ40 in the brain were increased over time (Fig3). Moreover, there was a negative correlation between CSF Aβ42 and soluble Aβ42 in the brain, whereas such a correlation for Aβ40 was lacking (Fig4).

Conclusions: Our results suggest that mechanisms other than aggregation of Aβ42 into insoluble plaques may contribute to decreased CSF concentrations of this Aβ peptide during plaque development in 5xFAD mice.

P234 / #1497


BIOSENSOR FOR THE SIMULTANEOUS DETECTION OF ALZHEIMER’S DISEASE BIOMARKERS

Lecture Title:

P. Carneiro1, J. Loureiro1, C. Delerue-Matos2, S. Morais2, M.D.C. Pereira1 1Faculty of Engineering of the University of Porto, Chemical Engineering, Porto, Portugal, 2School of Engineering of the Polytechnic of Porto, Chemical Engineering, Porto, Portugal

Aims: Early diagnosis of Alzheimer’s disease is still very challenging as a result of the clinical overlap with other diseases and also due to the low accuracy and high cost of the existing technology . This way, the goal of this work is to perform the simultaneous detection of Aβ42 and Aβ40 in biological fluids and determination of their ratio by electrochemical sensing. The proposed electrochemical immunosensor is based on a screen printed carbon electrode (SPCE) modified in a layer-by-layer approach with carbon nanotubes (CNTs) and gold nanoparticles (AuNPs). The formation of a stable layer of biological recognition elements on the nano-modified SPCE is achieved through chemical modification (thiolation) of monoclonal antibodies specific to amino acids 1-17 of human Aβ, promoting the formation of a strong gold-thiol bond between the antibodies and AuNPs. Moreover, to overcome the problem with non-specific binding albumin from bovine serum is also immobilized on the SPCE surface. Finally, a sandwich assay is carried out through the use of specific secondary antibodies. Methods: Square-wave voltammetry, cyclic voltammetry, electrochemical impedance spectroscopy and scanning electron microscopy were used to characterize the development of the biosensor. Results: Under the optimal conditions, an analytical curve was attained using the peak current observed in square-wave voltammetry after exposure of the immunosensor to different concentrations of Aβ42 and Aβ40. Conclusions: The developed immunosensor enables a successful, simple and highly sensitive detection of Aβ42/Aβ40.

P235 / #1349


DEREGULATED MICRORNA PLASMA LEVELS AS CANDIDATE BIOMARKERS OF SENESCENCE AND ALZHEIMER´S DISEASE

Lecture Title:

N. Csicsátková, M. Čente, P. Filipčík, K. Mátyásová Institute of Neuroimmunology, Slovak Academy of Science, Molecular And Cellular Neurobiology, Bratislava, Slovak Republic

Aims: Alzheimer´s disease (AD) as the most widespread type of neurodegeneration is typical by pathological changes presented years before the onset of AD symptomatic stage. Group of small non-coding RNAs microRNAs execute a role as post-transcriptional negative regulators which lead to translational inhibition or mRNA degradation. The biological function and stability of miRNAs in body fluids point out their potential as diagnostic biomarkers for AD neurodegeneration and progression. Our study aimed to identify miRNAs as potential biomarkers linked to the neurodegeneration of AD. Methods: Plasma samples were collected from 24 subjects (Alzheimer's disease patients n= 8, healthy age-matched individuals n= 8, healthy young individuals n= 8) in the equivalent representation of women and men. Transcriptomic analysis was conducted to compare AD patients to healthy age-matched controls for analysis of changes in expression profiles of miRNAs. Results: Transcriptomic analysis of plasma samples defined panel of 15 deregulated microRNA profiles distinguishing the AD patients from healthy age-matched individuals. Concurrently, downregulation of one miRNA identified as potential senescence marker differentiating healthy age-matched controls from healthy young controls. Conclusions: Our results show the diagnostic potential of identified deregulated microRNA panel to bring higher resolution for differentiation of AD individuals from healthy controls and related tauopathies. This project was supported by research grant: VEGA 2/0118/19, 2/0154/19, APVV-17-0668, APVV-19-0568 and ERA-NET NEURON Neu-Vasc.

P236 / #564


INCREASED NEUROLEUKIN LEVELS IN CEREBROSPINAL FLUID OF ALZHEIMER’S PATIENTS

Lecture Title:

A. De Kort1, B. Kuiperij1, D. Alcolea2, I. Kersten1, A. Versleijen3, F. Schreuder1, A. Lleó2, J. Claassen4, M. Verbeek1,3 1Radboud University Medical Center, Department Of Neurology, Nijmegen, Netherlands, 2Memory Unit, Sant Pau Biomedical Research Institute, Hospital de la Santa Creu i Sant Pau, Neurology, Barcelona, Spain, 3Radboud University Medical Center, Department Of Laboratory Medicine, Nijmegen, Netherlands, 4Radboud University Medical Center, Geriatrics, Nijmegen, Netherlands

Aims: Neuroleukin (NLK) is a recently identified cerebral cytokine, associated with senile plaques and cerebral amyloid angiopathy (CAA). The aim of our study was to investigate the discriminative value of CSF NLK in patients with CAA and in patients at various clinical stages of Alzheimer’s disease (AD). Methods: CSF NLK levels were quantified by ELISA in three cohorts: (1) a CAA-control cohort (n=26 CAA patients and n=27 controls); (2) an AD-control study cohort from Nijmegen consisting of 19 patients with mild cognitive impairment (MCI; clinical dementia rating (CDR) = 0.5), 28 patients with AD CDR 1.0 and 12 patients with AD CDR 2.0 and 27 controls, and (3) an AD-control validation cohort from Barcelona consisting of 33 MCI patients, 17 AD patients and 50 controls. Results: We found that CSF NLK levels were similar in CAA patients and controls (p=0.97). However, in the Nijmegen cohort we found a significant elevation of CSF NLK in both MCI (p<0.0001) and AD CDR 1 (p<0.0001) compared to controls. We could confirm this elevation in the Barcelona cohort, both in MCI (p<0.0001) and AD patients (p<0.0001) compared to controls. In addition, we found a correlation of NLK with YKL-40 (rsp=0.51, p<0.0001). Conclusions: Our findings indicate that CSF NLK levels are elevated in MCI and AD patients compared to controls and support an increased neuroinflammatory state in both MCI and AD.

P237 / #713


BIOCHEMICAL MARKERS OF ALZHEIMER'S DISEASE: FABP3 AS A POTENTIAL CANDIDATE.

Lecture Title:

M. Dulewicz1, A. Kulczynska-Przybik1, A. Klimkowicz-Mrowiec2, J. Pera2, A. Słowik2, B. Mroczko1,3 1Medical University of Bialystok, Neurodegeneration Diagnostics, Białystok, Poland, 2Jagiellonian University, Neurology, Krakow, Poland, 3Medical University of Bialystok, Biochemical Diagnostics, Białystok, Poland

Aims: Fatty acid binding protein 3 (FABP-3) is gaining importance in relation with the dopaminergic system as one of the crucial factor of AD pathology. FAB proteins, including FABP-3, regulates lipid transport, which is crucial for neuronal survival, synaptic activity and immune responses of glial cells in the brain. FABP-3 may also influence on isoform E4 of apolipoprotein, which is known as a risk factor of Alzheimer’s disease (AD). Moreover, FABP-3 binds and modulates the dopaminergic D2 receptors, and overexpression of FABP-3 may indirectly affect the formation of neurofibrillary tangles. Therefore, the aim of the study was a measurement of the FABP-3 level in the cerebrospinal fluid of patients with AD and non-demented controls. Methods: The cerebrospinal fluid levels of FABP-3 and classical AD biomarkers, such as Aβ-42, Aβ-42/Aβ-40, hTau and pTau181 were assessed by Luminex 200 Magnetic Bead platform and immunoenzyme assays. The study included 18 patients with AD and 15 non-demented controls. Results: In our research, we showed that the concentration of FABP3 was significantly higher in cerebrospinal fluid of AD patients compared to non-demented controls. The increased CSF levels of FABP3 were significantly correlated with Aβ-42/Aβ-40 ratio and pTau181 in AD subjects. Conclusions: Ours preliminary study has revealed that FABP3 seems to be related with pathogenesis of AD. However, the changes of FABP3 levels in CSF appeared in full-blown AD; therefore it may be an early indicator for later development of AD,although further studies on larger cohort are needed.

P238 / #1744


NEURON DERIVED EXTRACELLULAR VESICLES MIRROR BRAIN PATHOLOGY

Lecture Title:

E. Eren1, J. Leoutsakos2,3,4, C. Lyketsos2,3,4, E. Oh2,3,4, D. Kapogiannis1,4 1National Institute on Aging (NIA/NIH), Laboratory Of Clinical Investigation, Baltimore, United States of America, 2Johns Hopkins School of Medicine, Richman Family Precision Medicine Center Of Excellence In Alzheimer's Disease, Baltimore, United States of America, 3Johns Hopkins School of Medicine, Alzheimer's Disease Research Center, Baltimore, United States of America, 4Johns Hopkins School of Medicine, Memory And Alzheimer’s Treatment Center, Baltimore, United States of America

Aims: Neuron-derived Extracellular Vesicles EVs (NDEVs) can be isolated from peripheral blood and provide biomarkers for Alzheimer’s disease (AD). To further establish NDEV biomarkers as “liquid biopsy” for AD, we sought to demonstrate their relationship with ante-mortem cognition and brain pathology at autopsy Methods: We immunoprecipitated NDEVs by targeting neuronal marker L1CAM from ante-mortem plasma samples from 65 patients with definitive AD (Age, sex, interval from autopsy) and measured Aβ42, p181-Tau, total Tau, synaptic proteins and three EV markers, CD63, CD81 and CD9. Results: Higher levels of NDEV Aβ42 (and at trend p181-Tau) were associated with lower Braak stages (p=0.014, p=0.079 respectively). Higher levels of NDEV Aβ42 levels were consistently associated with better cognitive status, memory, fluency, working memory and executive function (all p<0.01). Higher levels of NDEV synaptophysin and synaptodopin were associated with better performance on working memory and executive function tasks (all p<0.05). Interestingly, higher levels of EV marker CD81 were associated with worse cognitive status, memory, and working memory (all p<0.05). Conclusions: Ante-mortem NDEV Aβ42 levels are positively associated with ante-mortem cognition and negatively associated with AD Braak stage at autopsy. Releasing Aβ42-laden NDEVs may be an adaptive mechanism in clinical AD.

P239 / #328


ULTRA-DEEP PROTEOMIC PROFILING OF CEREBROSPINAL FLUID FOR UNBIASED AD BIOMARKER DISCOVERY AND SUBJECT STRATIFICATION

Lecture Title:

Y. Feng1, R. Bruderer2, D. Heinzmann2, L. Reiter2 1Biognosys AG, Bd, SCHLIEREN - ZURICH, Switzerland, 2Biognosys AG, R&d, SCHLIEREN - ZURICH, Switzerland

Aims: We seek to address the unmet need in currently available CSF biomarkers by applying a novel mass spectrometry(MS)-based discovery workflow which enabled the unbiased proteomic characterization of CSF from AD subjects in an unprecedented depth. Methods: CSF samples were obtained from subjects with late onset Alzheimer’s disease (LOAD; n = 16) and age-matched normal controls (CO; n = 8). The samples were analyzed using high field asymmetric waveform ion mobility spectrometry (FAIMS) coupled to data-independent acquisition mass spectrometry (DIA-MS). Results: On average 2’606 proteins were quantified in 24 CSF samples. Across all samples 3’473 proteins were identified and quantified. The pool of quantified proteins comprises well characterized species associated with AD and other neurological disorders such as BACE1, APP, Tau, SNCA and NFL. Moreover, the depth and breadth of protein quantification covers numerous pathological mechanisms such as AB and Tau pathology, synaptic dysfunction (e.g. SPTX2 and SNAP25), neuronal injury, iron toxicity and inflammation. Differential expression analysis identified 431 proteins being significantly dysregulated between LOAD and CO groups (Q-value < 0.05 and Log2 FC > 0.58) from which 85% are upregulated in LOAD. Among these 369 upregulated proteins, we observed significant enrichments in immune-mediated processes, vesicle trafficking and secretion pathways. Conclusions: The FAIMS-DIA-MS workflow is scalable to the profiling of 1000s of samples and can thus be applied for subject stratification and biomarker discovery in clinical cohorts. We envision the unbiased quantification of >3’000 proteins in CSF to aid the discovery of new AD biomarkers as well as the elucidation of complex disease mechanisms.

P240 / #893


NEUROPATHOLOGIC CORRELATES OF ELECSYS-DERIVED CEREBROSPINAL FLUID BIOMARKERS OF ALZHEIMER’S DISEASE IN COMPARISON TO A NOVEL PLASMA-BASED P-TAU181 BIOMARKER

Lecture Title:

M. Grothe1,2,3, N. Ashton2,3, A. Moscoso2,3, H. Zetterberg2,4, K. Blennow2,4, M. Schöll2,3 1Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, Unidad De Trastornos Del Movimiento, Sevilla, Spain, 2Institute of Neuroscience and Physiology, The Sahlgrenska Academy, University of Gothenburg, Department Of Psychiatry And Neurochemistry, Gothenburg, Sweden, 3University of Gothenburg, Wallenberg Centre For Molecular And Translational Medicine, Gothenburg, Sweden, 4Sahlgrenska University Hospital, Clinical Neurochemistry Laboratory, Mölndal, Sweden

Aims: To analyze neuropathological correlates of the fully automated Elecsys cerebrospinal fluid (CSF) biomarkers of Alzheimer’s disease (AD) in comparison to a novel blood-derived biomarker of phosphorylated tau (p-tau181). Methods: We analysed ante-mortem Elecsys CSF biomarkers in comparison to standardized post-mortem assessments of neuropathologic changes in 45 participants from the ADNI autopsy cohort. In a subset of 26 participants we also analyzed ante-mortem plasma p-tau181 levels. We studied the diagnostic accuracy of these biomarkers to distinguish neuropathologically-confirmed AD dementia cases from Aβ-PET-negative healthy controls (N=146) and non-AD dementia cases at autopsy, and further assessed pathology-specific associations of the different biomarkers. Results: All CSF biomarkers clearly distinguished pathologically-confirmed AD dementia patients from Aβ-PET-negative controls, reaching perfect group separation with the p-tau181/Aβ1-42 ratio (AUC=1.00). Total-tau (t-tau) and p-tau181 levels, as well as their ratios with Aβ1-42, also clearly distinguished pathologically-confirmed AD from non-AD dementia (AUCs=0.94-0.97). Thal amyloid phase showed highest correlation with Aβ1-42 (rho=-0.62), and CERAD neuritic plaques and Braak tau stage showed highest correlation with p-tau181 (rho=0.61/0.64, all p<0.001). Optimal pathology-specific biomarker cut-offs were estimated as 1097 pg/ml for Aβ1-42, 229 pg/ml for t-tau, and 19 pg/ml for p-tau181. Compared to CSF, plasma p-tau181 demonstrated comparably high diagnostic accuracies for distinguishing pathologically-confirmed AD from Aβ-PET-negative controls (AUC=0.91) and non-AD dementia cases (AUC=0.96), and showed similar, though weaker, pathology-specific associations with neuritic plaques and Braak stage (rho=0.47/0.43, p<0.028). Conclusions: Elecsys CSF biomarkers detected AD neuropathologic changes with very high discriminative accuracy in-vivo. Preliminary findings also indicate pathology-specific associations for plasma-based p-tau181 measurements.

P241 / #1788


ASSOCIATION OF NEURONAL PENTRAXIN 2 IN CEREBROSPINAL FLUID WITH COGNITION IN THE BIOFINDER-2 STUDY

Lecture Title:

D. Jacobs1, S. Janelidze2, E. Stomrud2, O. Hansson2, E. Vanmechelen1 1ADx NeuroSciences, Technologiepark 94, Ghent, Belgium, 2Lund University, Clinical Memory Research Unit, Department Of Clinical Sciences Malmö, Malmö, Sweden

Aims: Alzheimer’s disease is a slowly progressing neurodegenerative disease affecting several specific neuronal circuits in a stereotypical, sequential and hierarchical way. The synaptic organizer protein, Neuronal pentraxin 2 (NPTX2), which defines an interneuronal circuit activity is currently the most robust biochemical marker in cerebrospinal fluid (CSF) of cognitive decline among the several studied synaptic fluid biomarkers. Methods: We used a monoclonal antibody based ELISA (ADx NeuroSciences) to detect CSF levels of NPTX2 in the Biofinder-2 cohort (NCT03174938). In this cohort CSF total tau (tTau), CSF phospho-tau181 (Ptau181) were determined using the Innotest, while CSF phospho-tau231 (Ptau231) was determined using a research ELISA (ADx NeuroSciences). Results: We confirmed that NPTX2/tau (Ptau) ratios were the most performant diagnostic biomarkers discriminating AD and AD-MCI (n=342) from cognitively unimpaired individuals (n=473) (Table). In contrast to NPTX2 as a single marker the NPTX2/tTau ratio showed a robust association with MMSE in AD (r=0.24; p=0.003), with MMSE ranging from 8-28 and additionally a statistically significant correlation was also demonstrated in the AD-MCI cohort (r=0.15; p=0.036) with MMSE scores ranging from 22-30. Table: Area under Curve (AUC) of Receiver Operating Curves (ROC) for discriminating AD, MCI due to AD (n=342) from cognitively unimpaired individuals (CU, n=473).

Variable AUC (95% CI)

CSF-NPTX2 0.645 (0.611-0,678)

CSF-tTau 0.792 (0.763-0.820)

CSF-Ptau181 0,765 (0,734-0,794)

CSF-Ptau231 0,803 (0,774-0,830)

CSF-NPTX2/tTau 0,869 (0,844-0,891)

CSF-NPTX2/Ptau181 0,857 (0,831-0,880)

CSF-NPTX2/Ptau231 0,856 (0,830-0,879)

Conclusions: The ratio NPTX2/tau equals the NPTX2/Ptau ratio and is superior to single analyte diagnosis of clinical AD in CSF.

P242 / #1341


AMYLOID-ΒETA PEPTIDES IN CSF DISCRIMINATE CEREBRAL AMYLOID ANGIOPATHY FROM CONTROLS

Lecture Title:

B. Kuiperij1, T. Marques1, A. De Kort1, F. Schreuder1, K. Klijn1, E. Stoops2, G. Brinkmalm3, E. Portelius3, E. Gkanatsiou3, M. Verbeek1 1Radboud University Medical Center, Department Of Neurology, Nijmegen, Netherlands, 2ADx NeuroSciences, , Ghent, Belgium, 3Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden

Aims: Cerebral amyloid angiopathy (CAA) is characterized by the accumulation of amyloid-beta (Abeta) peptides in the cerebral vasculature and has been associated with dementia and intracerebral haemorrhages. Currently, the diagnosis of CAA is based on neuroimaging and includes the identification of lobar (micro-)bleeds and cortical superficial siderosis. However, these MRI markers reflect only late-stage manifestations and are therefore not suitable for an early diagnosis of CAA. We aimed to evaluate the diagnostic potential of the Abeta peptides AB1-38, AB1-40, AB1-42, and AB1-43 in cerebrospinal fluid (CSF) to discriminate CAA from controls. Methods: Our cohort included patients diagnosed with probable or possible CAA (n=28), and non-neurological controls (n=40). We used immunoassays to quantify the concentrations of AB1-38, AB1-40, AB1-42 and AB1-43 in CSF. Results: In addition to the well-described lower levels of AB1-40 in CAA, we found decreased levels of AB1-38 (p=0.03), AB1-42 (p<0.0001) and AB1-43 (p<0.0001) in CSF of CAA patients compared to controls. The areas under the curve (AUC), which reflect the diagnostic accuracy, were 0.64 (AB1-38), 0.90 (AB1-42), and 0.92 (AB1-43). The concentrations of all Abeta peptides strongly correlated with each other (Spearman r>0.6, p<0.0001 for each comparison). Conclusions: We confirm previous findings that Abeta peptides have strong biomarker potential for the diagnosis of CAA. Interestingly, our results indicate that AB1-43, which has shown to be relatively low-abundant in vascular amyloid deposits, has the highest diagnostic accuracy to discriminate CAA from controls. In addition, we show for the first time that CSF AB1-38 levels are decreased in CAA compared to controls.

P243 / #938


THE RETICULON4A AS A CANDIDATE BIOMARKER OF ALZHEIMER’S DISEASE

Lecture Title:

A. Kulczynska-Przybik1, A. Klimkowicz-Mrowiec2, M. Dulewicz1, R. Borawska1, A. Litman-Zawadzka1, A. Słowik2, B. Mroczko1,3 1Medical University of Bialystok, Department Of Neurodegeneration Diagnostics, Białystok, Poland, 2Jagiellonian University, Neurology, Krakow, Poland, 3Medical University of Bialystok, Department Of Biochemical Diagnostics, Białystok, Poland

Aims: Alzheimer’s disease (AD) is a heterogeneous and complex disease. Research in AD biology has made substantial progress but the precise mechanism of the disease is not well understood. Some evidence suggests that RTN4A protein is engaged in the pathology of Alzheimer’s disease by regulation of BACE1 function or APP processing. Moreover, it was revealed that RTN4A protein may influence act as a negative regulator of synaptic plasticity, as well as impairment of cognitive functions. Therefore, the objective of the present study was the evaluation of RTN4A levels in the cerebrospinal fluid (CSF) of AD patients and non-demented controls, and the assessment of a relationship between this protein and classical AD biomarkers. Methods: The concentration of RTN4A and classical biomarkers (Aβ-40, Aβ-42, tau, and pTau181) were measured in the CSF of 18 patients with AD and 18 non-demented controls by means of quantitative sandwich enzyme immunoassay technique. Results: The CSF concentrations of RTN4A were significantly higher in the AD group in comparison with cognitively normal individuals. Moreover, the level of this protein was observed significantly correlated with tau and pTau181 proteins in AD group. Conclusions: Presented findings indicate on a potential role of RTN4A proteins in the pathology of AD, and as a candidate biomarker of AD supporting classical biomarkers and improve clinical outcomes, but these investigations need to be further clarified. Acknowledgment: The study was conducted with the use of equipment purchased by Medical University of Białystok as part of the RPOWP 2007-2013 funding, Priority I, Axis 1.1, contract No. UDA-RPPD.01.01.00-20-001/15-00 dated 26.06.2015.

P244 / #1534


ALTERED MICRORNAS RELATED TO SYNAPTIC FUNCTION AS POTENTIAL PLASMA BIOMARKERS FOR EARLY STAGES OF ALZHEIMER’S DISEASE

Lecture Title:

A. Miñano-Molina1,2, D. Siedlecki-Wullich1,2, J. Català-Solsona1,2, C. Fábregas-Ordóñez1,2, I. Hernandez3, J. Clarimon1,4, A. Lleó1,4, M. Boada1,3, C. Saura1,2, J. Rodríguez-Álvarez1,2,5 1Instituto de Salud Carlos III, Centro De Investigacion Biomedica En Red Enfermedades Neurodegenerativas - Ciberned, Madrid, Spain, 2Institut de Neurociències, Universitat Autònoma de Barcelona, Departament De Bioquímica I Biologia Molecular, Barcelona, Spain, 3Fundació ACE, Institut Català de Neurociències Aplicades - Universitat Internacional de Catalunya, Research Center And Memory Clinic, Barcelona, Spain, 4Institut d’Investigacions Biomèdiques Sant Pau - Hospital de Sant Pau, Universitat Autònoma de Barcelona, Sant Pau Memory Unit, Department Of Neurology, Barcelona, Spain, 5Albert Einstein College of Medicine, Dominick P. Purpura Department Of Neuroscience, New York, United States of America

Aims: Failure of synaptic function occurs at preclinical stages of Alzheimer’s disease (AD) preceding neuronal loss and AD pathological hallmarks. Nowadays, there is an urgent need to identify reliable biomarkers that could be obtained with non-invasive methods to improve AD diagnosis at early stages. Our aim was to perform a miRNA massive analysis in AD brain samples and examine plasma levels of a group of miRNAs related to synaptic proteins in a cohort composed of cognitive healthy controls (HC), mild cognitive impairment (MCI) and AD subjects potentially useful as biomarker. Methods: Affymetrix miRNA 4.1 microarray was used to analyze enthorrinal cortex and hippocampal brain tissue at different stages of the disease. Plasma levels of 8 miRNAs related to synaptic function, selected by bioinformatics approaches, were analyzed in two different cohorts including 38 HC, 26 MCI, 56 AD dementia patients and 27 Frontotemporal dementia (FTD) patients. D’Agostino-&-Pearson and Shapiro-Wilk tests were used to evaluate data normality. miRNA levels between groups were compared using a two-sided nonparametric Mann-Whitney test and sensitivity and specificity was determined by receiver operating characteristic curve analysis. Results: Significant up-regulation of miR-92a-3p, miR-181c-5p and miR-210-3p was found in the plasma of both MCI and AD subjects. MCI patients that progress to AD showed higher plasma levels of these miRNAs. By contrast, no changes in miR-92a-3p, miR-181c-5p or miR-210-3p levels were observed in plasma obtained from a cohort of FTD. Conclusions: Our study shows that plasma miR-92a-3p, miR-181c-5p and miR-210-3p constitute a specific molecular signature potentially useful as a potential biomarker for AD.

P245 / #1259


ASSOCIATION OF BLOOD PHOSPHORYLATED TAU181 AND NEUROFILAMENT LIGHT WITH PROGRESSIVE NEURODEGENERATION IN ALZHEIMER DISEASE

Lecture Title:

A. Moscoso1, M. Grothe1,2, N. Ashton3, H. Zetterberg4, K. Blennow5, M. Schöll6 1Wallenber Center for Molecular Imaging, Department Of Psychiatry And Neurochemistry, Institute Of Neuroscience And Physiology, The Sahlgrenska Academy, University Of Gothenburg, Sweden, Göteborg, Sweden, 2Unidad de Trastornos del Movimiento, Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del RocíoCSIC/Universidad de Sevilla, Sevilla, Spain, Unidad De Trastornos Del Movimiento, Instituto De Biomedicina De Sevilla (ibis), Hospital Universitario Virgen Del Rocío/csic/universidad De Sevilla, Sevilla, Spain, Sevilla, Spain, 3University of Gothenburg, Inst Of Neuroscience & Physiology, Dept Of Psychiatry & Neurochemistry, Mölndal, Sweden, 4Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden, 5Univ. of Gothenburg, Inst Of Neuroscience And Phsyiology, Mölndal, Sweden, 6University of Gothenburg, Wallenberg Centre For Molecular And Translational Medicine And The Department Of Psychiatry And Neurochemistry, Gothenburg, Sweden

Aims: To study the potential of plasma p-tau181 for assessing neurodegeneration progression and cognitive decline in AD in comparison to plasma neurofilament light chain (NfL), a non-disease specific neuronal injury marker. Methods: We included 1113 subjects (374 cognitively unimpaired (CU) and 734 impaired (CI)) from the Alzheimer’s Disease Neuroimaging Initiative (ADNI) with available plasma p-tau181 and neurofilament light chain (NfL) measurements, as well as at least one [18F]fluorodeoxyglucose (FDG) PET or structural MRI scan. We explored associations of plasma p-tau181 and NfL levels with longitudinal cognitive decline and prospective neurodegeneration (hypometabolism and atrophy) using brain-wide voxel-wise and region-of-interest analyses in predefined regions typically affected in AD. Results: Baseline levels of plasma p-tau181 correlated with cognitive decline (CU: β=-0.11, p=0.04; CI: β=0.34, p<0.001) and prospective glucose hypometabolism (CU: r=-0.06, p=0.40; CI: r=-0.28, p<0.001) and grey matter atrophy (CU: r=-0.11, p=0.03; CI: r=-0.28, p<0.001) in a highly AD-characteristic regional pattern. Similar and stronger correlations were found for longitudinal changes of plasma p-tau181. These associations were limited to amyloid-β-positive individuals. Both plasma p-tau181 and NfL were independently associated with cognition and neurodegeneration in AD-vulnerable regions. However, NfL also correlated with neurodegeneration in brain regions exceeding this AD-typical spatial pattern in amyloid-β-negative subjects. Mediation analyses revealed that ~30-50% of plasma p-tau181 effects on cognitive decline were mediated by imaging neurodegeneration markers, suggesting links between plasma p-tau181 and cognition independent of these measures. Conclusions: Plasma p-tau181 is an accessible marker for predicting and monitoring AD-related neurodegeneration and cognitive decline and, unlike plasma NfL, is AD-specific.

P246 / #953


POTENTIAL CLINICAL USEFULNESS OF MICROGLIA MARKERS: TREM2 AND YKL-40 IN ALZHEIMER’S DISEASE

Lecture Title:

B. Mroczko1,2, A. Kulczynska-Przybik1, A. Klimkowicz-Mrowiec3, J. Pera3, R. Borawska1, M. Dulewicz1, A. Litman-Zawadzka1, A. Słowik3 1Medical University of Bialystok, Department Of Neurodegeneration Diagnostics, Białystok, Poland, 2Medical University of Bialystok, Department Of Biochemical Diagnostics, Białystok, Poland, 3Jagiellonian University, Neurology, Krakow, Poland

Aims: Alzheimer’s disease (AD) is a multifactorial disease characterized by a robust inflammatory response of microglia. Mounting evidence has supported the crucial role of TREM2 and YKL-40 expressed by microglia in the pathology of Alzheimer’s disease. The overexpression of TREM2 has been found in patients with amyloid and tau pathology and seems to be associated with the recruitment of microglia to amyloid plaques. The purpose of the present study was to assess the cerebrospinal fluid (CSF) concentrations of TREM2 and YKL-40 to test their potential usefulness in diagnosing Alzheimer’s disease. Methods: The concentrations of TREM2, YKL-40 and classical biomarkers (Aβ-42, Aβ-42/Aβ-40, tau and pTau181) were measured in the CSF of 18 AD patients and 15 elderly subjects without cognitive deficits using ELISA and Multiplexing methods. Results: The CSF concentration of TREM2 and YKL-40 was significantly higher in the group of AD patients in comparison to non-demented controls. The levels of TREM2 and YKL-40 significantly correlated with age, Aβ-42/Aβ-40 ratio, tau proteins in the whole study group. Moreover, in the AD group was revealed the association between TREM2 and age, as well as between YKL40 and Aβ-42/Aβ-40 ratio. Conclusions: Presented findings indicate a potential role of TREM2 and YKL-40 in the pathology of AD and the possible usefulness of these proteins in the early diagnosis of patients with AD. Acknowledgment: The study was conducted with the use of equipment purchased by the Medical University of Białystok as part of the RPOWP 2007-2013 funding, Priority I, Axis 1.1, contract No. UDA-RPPD.01.01.00-20-001/15-00 dated 26.

P247 / #766


BETA-SYNUCLEIN IS A NOVEL SYNAPTIC MARKER FOR ALZHEIMER´S DISEASE WITH CONSISTENT CHANGES IN CSF AND BLOOD

Lecture Title:

P. Oeckl1, S. Halbgebauer1, S. Anderl-Straub1, C. Von Arnim2, J. Diehl-Schmid3, L. Froelich4, T. Grimmer3, L. Hausner4, J. Denk5, H. Jahn5, P. Steinacker1, J. Weishaupt1, A. Ludolph1, M. Otto1 1Ulm University Hospital, Department Of Neurology, Ulm, Germany, 2Ulm University, Department Of Neurology, Ulm, Germany, 3Technical University of Munich, School of Medicine, Klinikum rechts der Isar, Center For Cognitive Disorders, Department Of Psychiatry And Psychotherapy, Munich, Germany, 4Central Institute of Mental Health Mannheim, Gerontopsychiatry, Mannheim, Germany, 5University Hospital Hamburg, Department Of Psychiatry, Hamburg, Germany

Aims: Synaptic degeneration is a major hallmark of Alzheimer´s disease (AD) and the best pathological correlate of cognitive dysfunction. Synaptic markers are therefore a highly desired read-out for clinical trials in the mainly unsuccessful AD drug development. Several synaptic markers for AD are described in cerebrospinal fluid (CSF) but studies in blood were not successful so far. Methods: We used quantitative mass spectrometry (IP-MS, MRM) to measure the presynaptic protein beta-synuclein in CSF and blood from 363 patients: AD (n=64), Creutzfeldt-Jakob disease (CJD, n=25), behavioural variant frontotemporal dementia (bvFTD, n=16), dementia with Lewy bodies/Parkinson´s disease dementia, (DLB/PDD, n=13), Parkinson´s disease (PD, n=25), amyotrophic lateral sclerosis (ALS, n=30) and non-neurodegenerative controls (Con, n=45). Changes in AD were validated in two independent validation cohorts from other clinical centers (AD: n=40 and n=49, Con: n=44 and n=25). Results: We observed increased concentrations of beta-synuclein in CSF and blood of AD patients (CSF: median 979pg/mL, IQR: 738-1223pg/mL vs. 659pg/mL, IQR: 521-860pg/mL in controls; Serum: 12.9pg/mL, IQR: 10.6-16.4pg/mL vs. 8.9pg/mL, IQR: 7.1-10.4pg/mL in controls, p<0.01) and confirmed this finding in the two validation cohorts. Changes in CSF were additionally confirmed with an in-house ELISA as a second, independent method (r=0.92 for ELISA and antibody-free MRM). Beta-synuclein was already increased in patients with mild cognitive impairment due to AD (p<0.01). Conclusions: These findings suggest beta-synuclein as a candidate blood marker for synaptic degeneration which might be used in clinical AD trials and patient follow-up as part of the recently suggested ATN biomarker panel.

P248 / #995


PERIPHERAL INFLAMMATORY BIOMARKERS AND APOE GENOTYPE AS RISK FACTORS OF COGNITIVE IMPAIRMENT IN THE CHILEAN GERO COHORT: PRELIMINARY STUDY

Lecture Title:

P. Orellana1,2, C. Duran-Aniotz1,3, F. Henríquez1,2,4, V. Cabello1,2,4, P. Lillo1, R. Villagra1, D. Thumala1, T. Parrao1, C. Gonzalez-Billault1,5,6, A. Slachevsky1,2,4 1Geroscience Center for Brain Health and Metabolism (GERO), University Of Chile, Santiago, Chile, 2Neuropsychology and Clinical Neuroscience Laboratory (LANNEC), Physiopathology, University Of Chile, Santiago, Chile, 3University Adolfo Ibáñez, Center For Social And Cognitive Neuroscience (cscn), School Of Psychology, Santiago, Chile, 4Neurology Department, Hospital del Salvador and Faculty of Medicine, University of Chile, Memory And Neuropsychiatric Clinic (cmyn), Santiago, Chile, 5University of Chile, Aboratory Of Cell And Neuronal Dynamics, Faculty Of Sciences, Santiago, Chile, 6The Buck Institute for Research on Aging, The Buck Institute For Research On Aging, Novato, United States of America

Aims: We aim to study and correlate the inflammatory biomarkers, ApoE genotype with different cognitive domains in the GERO cohort (49 subjective complaint (SC) and 62 mild cognitive impairment (MCI), >70 years) and 33 healthy controls (HC). Methods: Plasma inflammatory proteins IL-2, IL-6, IL-10, TNF-α, CRP, SAP and ApoE genotype were analyzed and correlated with neuropsychological assessment. Results: CRP showed a decreased expression in the SC compared to MCI and HC. SAP showed a decreased expression in SC as well as MCI compared to HC. A correlation of CRP with memory and social cognition in the SC and with memory in the MCI was found. SAP correlations were observed with nomination and repetition in the SC and with repetition in the MCI. Additionally, ApoE analysis was performed in 3 genotype groups: e2 (ε2/ε2-ε2/ε3, n=10), e3 (ε3/ε3, n=83) and e4 (ε2/ε4-ε3/ε4-ε4/ε4, n=20). Differences between SC and MCI were found in the e2 allele in social cognition. In the e3 allele in attention, executive function, memory, verbal fluency, language, social cognition and visuoconstructive skills. In the e4 allele in executive function, language, emotional recognition and functionality. Conclusions: SAP and CRP showed correlations with memory, language and social cognition, suggesting that these proteins may be associated with pathological aging. The ApoE e3 allele, the MCI showed an increased cognitive impairment compared with the e2 and e4 alleles, which can not be explained by the ApoE effect. The e4 allele, the MCI was observed to have an increased cognitive and functionality impairment.

P249 / #777


EARLY AD DIAGNOSIS BY MIRNAS PLASMA BIOMARKERS

Lecture Title:

C. Peña-Bautista, A. Tarazona, L. Álvarez, A. Cuevas, M. Baquero, C. Cháfer-Pericás Health Research Institute La Fe, Neurology, Valencia, Spain

Aims: There is a general consensus on the need to identify the Alzheimer’s disease (AD) cases at an early stage, so that existing treatments and those available in the near future could stop its evolution, avoiding the serious and costly consequences of dementia produced by the AD. Several genes implied in AD pathogenesis altered their expression patterns trough epigenetics by means of microRNAs. This miRNAs could be detected in plasma samples and are postulated as good AD biomarkers. The main objective of this work is the identification of miRNA biomarkers in early stages of AD in a minimally invasive and economically acceptable manner, allowing the development of a personalized, specific and transferable diagnostic model. Methods: MiRNAs were analysed in RNA samples purified from plasma samples from MCI and healthy control participants’ groups y next generation sequencing (NGS). MiRNAs sequences were identify by using miRExpress tool. MiRNAs with a differential expression between both groups were included in a multivariate diagnosis tool. Results: Several identified miRNAs showed different expression levels between healthy control and MCI groups. MiRNAs with a significant different expression between both groups were included in a multivariate diagnosis model that showed promising diagnosis indices. Conclusions: MiRNAs seem to be useful in early AD detection in plasma samples. However, target quantification of miRNAs is necessary in order to verify these results and bring these biomarkers closer to clinical practice.

P250 / #786


ALZHEIMER DISEASE EARLY AND MINIMALLY INVASIVE DIAGNOSIS BASED ON LIPID BIOMARKERS

Lecture Title:

C. Peña-Bautista, M. Roca, L. Álvarez, I. Ferrer, L. García, M. Baquero, C. Cháfer-Pericás Health Research Institute La Fe, Neurology, Valencia, Spain

Aims: Alzheimer Disease diagnosis is nowadays too late in many cases to achieve a beneficial effect from conventional and new treatments. In last few years, lipid have been postulated as potential biomarkers for the pathology due to the high lipid composition in brain. The aim of this work is to show the utility of lipid compounds as early and minimally invasive biomarkers for AD and to develop a diagnosis tool able to be applied in clinical practise. Methods: A method based on HPLC-MS/MS was developed for the quantification of 4 lipid biomarkers (LysoPC18:1, Lyso PE18:0, L-alfa-fosfatisilcholine, DHA) in plasma samples. These compounds were analysed in 58 participants divided in early Alzheimer Disease (AD) (n=29) and Healthy control (HC) (n=28) groups. Results: highlighted the correlation between LysoPC 18:1 and tau and phosphorylated tau levels from cerebrospinal fluid biomarkers, as well as with RBANSL neuropsychological scale score. Individually, these compound were no able to achieve an accurate AD diagnosis. However, a multivariate model including the 4 biomarkers with age and gender is able to make a diagnosis with an AUC-ROC close to 0.8. In addition, this model shows a specificity above 90%. Although it does not have a good sensitivity, it can be useful as a first screening test Conclusions: Lipid biomarkers could be useful in AD diagnosis. However, a clinical validation with an external cohort is need in order to apply these findings to clinical practise.

P251 / #356


ASSESSMENT OF THE CONCORDANCE AND DIAGNOSTIC ACCURACY BETWEEN ELECSYS AND LUMIPULSE FULLY AUTOMATED PLATFORMS AND INNOTEST

Lecture Title:

G. Piñol-Ripoll1, F. Dakterzada2, R. López-Ortega1, A. Arias1, I. Riba-Llena1, M. Ruiz-Julian1, R. Huerto1, N. Tahan1 1Hospital Universitari Santa Maria Lleida-IRBLleida, Cognitive Impairment Unit, Lleida, Spain, 2Hospital Universitari Santa Maria-IRBLleida, Cognitive Disorders Unit, LLeida, Spain

Aims: Manual ELISA assays are the most commonly used methods for quantification of CSF AD biomarkers; however, they show inter- and intra-laboratory variability. We compared the Innotest ELISA method with two fully automated platforms (Lumipulse and Elecsys) to determine whether these new methods can provide effective substitutes for ELISA assays. Methods: We included 149 males and females with AD (n = 34), MCI (n = 94) and non-AD dementias (n = 21). Aβ42, T-tau, and P-tau were quantified using the ELISA method (Innotest, Fujirebio Europe), CLEIA method on a Lumipulse G600II, and ECLIA method on a Cobas e 601 instrument. Results: We found a high correlation between the three methods. Both Lumipulse and Elecsys methods were highly concordant with clinical diagnoses, and the combination of Lumipulse Aβ42 and P-tau had the highest discriminating power (AUC 0.915, 95% CI 0.822–1.000). The use of Aβ42/Aβ40 ratio instead of Aβ42 alone in AT(N) classification enhanced the diagnostic accuracy (AUC 0.798, 95% CI 0.649–0.947 vs AUC 0.778, 95% CI 0.617–0.939). Finally, the biomarker ratios (P-tau/Aβ42 and T-tau/Aβ42) were more consistent with the Aβ42/Aβ40 ratio for both Lumipulse and Elecsys methods, and Elecsys P-tau/Aβ42 had the highest consistency with amyloid pathology (AUC 0.994, 95% CI 0.986–1.000 and OPA 96.4%). Conclusions: The Lumipulse and Elecsys CSF AD assays showed high analytical and clinical performances so their use is recommended for the measurement of CSF AD biomarkers compared with unstandardized manual methods.

P252 / #358


IDENTIFICATION AND VALIDATION OF ENDOGENOUS CONTROL MIRNAS IN PLASMA SAMPLES FOR NORMALIZATION OF QPCR DATA FOR ALZHEIMER’S DISEASE

Lecture Title:

F. Dakterzada1, G. Piñol-Ripoll2, A. Targa3, I. Benitez3, G. Torres3, L. Romero1, D. De Gonzalo-Calvo3, A. Moncusi4, R. Huerto2, M. Sanchez-De-La-Torre3, F. Barbe3 1Hospital Universitari Santa Maria-IRBLleida, Cognitive Disorders Unit, LLeida, Spain, 2Hospital Universitari Santa Maria Lleida-IRBLleida, Cognitive Impairment Unit, Lleida, Spain, 3Translational Research in Respiratory Medicine, Hospital Universitari Arnau De Vilanova-santa Maria, Irblleida, LLeida, Spain, 4Hospital Universitari Arnau de Vilanova-Santa Maria, IRBLleida, Translational Research In Respiratory Medicine Hospital Universitari Arnau De Vilanova-santa Maria, LLeida, Spain

Aims: Several studies had explored the role of miRNAs in Alzheimer’s disease (AD). The identification of endogenous controls (ECs) would contribute to the standardization of these biomarkers in AD but they have not yet been established. The objective of the study was to identify miRNAs that can be used as ECs in AD. Methods: We evaluated 145 patients divided into two different cohorts. One was a discovery cohort of 19 women diagnosed with mild to moderate AD (Mini-Mental State Examination (MMSE) score ≥ 20) and with confirmed pathologic levels of Aβ42 in CSF. The stability assessment cohort consisted of 126 individuals: 24 control subjects, 25 subjects with MCI and negative for CSF biomarkers, 22 subjects with MCI and positive for CSF biomarkers and 55 subjects with AD and positive for CSF biomarkers. In the discovery cohort, a profile of 384 miRNAs was determined in plasma by TaqMan Low Density Array. The best EC candidates were identified by mean-centering and concordance correlation restricted normalization methods. The stability of the EC candidates was assessed using the GeNorm, BestKeeper and NormFinder algorithms. Results: Nine miRNAs (hsa-miR-324-5p, hsa-miR-22-5p, hsa-miR-103a-2-5p, hsa-miR-362-5p, hsa-miR-425-3p, hsa-miR-423-5p, hsa-let-7i-3p, hsa-miR-532-5p, and hsa-miR-1301-3p) were identified as EC candidates in the discovery cohort. The validation results indicated that hsa-miR-103a-2-5p was the best EC, followed by hsa-miR-22-5p, hsa-miR-1301-3p, and hsa-miR-425-3p, which had similar stability values in all three algorithms. Conclusions: We identified a profile of four miRNAs as potential plasma ECs to be used for normalization of miRNA expression data in studies of subjects with cognitive impairment.

P253 / #359


CIRCULATORY MIRNAS AS BIOMARKERS FOR COGNITIVE EVOLUTION IN PATIENTS WITH ALZHEIMER’S DISEASE

Lecture Title:

G. Piñol-Ripoll1, F. Dakterzada2, A. Targa3, I. Benitez3, A. Lladó4, G. Torres3, L. Romero2, D. De Gonzalo-Calvo3, A. Tort-Merino5, A. Moncusi6, R. Huerto1, M. Sanchez-De-La-Torre3, F. Barbe3 1Hospital Universitari Santa Maria Lleida-IRBLleida, Cognitive Impairment Unit, Lleida, Spain, 2Hospital Universitari Santa Maria-IRBLleida, Cognitive Disorders Unit, LLeida, Spain, 3Translational Research in Respiratory Medicine, Hospital Universitari Arnau De Vilanova-santa Maria, Irblleida, LLeida, Spain, 4Neurology Service, Hospital Clínic of Barcelona, Barcelona, Spain and Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Alzheimer’s Disease And Other Cognitive Disorders Unit, Barcelona, Spain, 5Hospital Clínic of Barcelona / Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Neurology, Barcelona, Spain, 6Hospital Universitari Arnau de Vilanova-Santa Maria, IRBLleida, Translational Research In Respiratory Medicine Hospital Universitari Arnau De Vilanova-santa Maria, LLeida, Spain

Aims: To detect and validate circulatory miRNAs that can be associated with the speed of cognitive decline in patients with AD after two years of follow-up. Methods: We evaluated 77 patients divided into two different cohorts. One was a discovery cohort of 19 women diagnosed with mild to moderate AD (Mini-Mental State Examination (MMSE) score ≥ 20) at baseline and with confirmed pathologic levels of Aβ42 in CSF. The validation cohort consisted of 58 male and female with the same inclusion criteria than discovery cohort and with a 2 years of cognitive follow-up. In the discovery cohort, a profile of 384 miRNAs was determined in plasma by TaqMan Low Density Array. The miRNA candidates were studied in the validation cohort by RT-qPCR. Results: The patients were grouped according to the rate of decline in the MMSE score along the two years. We identified 17 miRNAs that were differentially expressed between two groups those who lost less than 4 points (N = 11) and those who lost four or more points (N = 8) in the Discovery cohort. The validation results indicated that low expression of miRNA-342-5p allows to differentiate those patients more than 4 points in MMSE after 2 years of follow-up. Conclusions: miRNA-342-5p could be useful to predict, at the time of diagnosis, those AD patients who will have a faster cognitive decline.

P254 / #1287


CROSS-PLATFORM EVALUATION OF HIGHLY SENSITIVE IMMUNOASSAY TECHNOLOGIES FOR SERUM MEASURES OF PTAU181: PERFORMANCE IN ALZHEIMER’S DISEASE

Lecture Title:

C. Rubel1, J. Czerkowicz1, K. Ferber2, E. Milliman3, L. Bekris4, J. Leverenz5, D. Graham1 1Biogen, Biomarkers, Cambridge, United States of America, 2Biogen, Biostatistics, Cambridge, United States of America, 3Biogen, Data Sciences, Cambridge, United States of America, 4Cleveland Clinic, Lerner Research Institute, Genomic Medicine Institute, Cleveland, United States of America, 5Cleveland Clinic, Neurological Institute, Department Of Neurology, Lou Ruvo Center For Brain Health, Cleveland, United States of America

Aims: Recent studies suggest phosphorylated-tau-181 (pTau181) in blood is significantly elevated in Alzheimer’s disease (AD) relative to healthy controls (HC) and may be associated with development of AD in cognitively normal (CN) and Mild Cognitive Impairment (MCI) subjects (Janelidze et al.; Rodriguez et al.). These observations suggest significant potential for blood pTau181 as a non-invasive prognostic biomarker as both a patient enrichment tool and biomarker of disease progression in AD. Leveraging CN controls (n=89), MCI (n=33) and AD (n=121) patient samples from the Australian Imaging, Biomarker and Lifestyle Flagship Study of Aging (AIBL) and the Cleveland Clinic Center for Brain Health Biobank we sought to evaluate the performance of two pTau181 assays. Methods: pTau181 levels were determined in 243 serum samples utilizing a Meso-Scale-Discovery S-PLEX immunoassay and Quanterix Simoa pTau-181 V2 Advantage Kit. For assay comparison, we estimated the Pearson correlation coefficient and 95% confidence interval and generated a Bland-Altman plot. To assess the assays’ ability to discriminate clinical groups, we compared the mean concentrations of pTau181. Results: Serum pTau181 concentrations calculated by the two assays were highly correlated (r= 0.84; 95% CI (0.80, 0.88)), though S-PLEX concentrations were approximately 30% higher. 97% of samples were detectable by both assays and mean pTau181 concentrations distinguished AD + MCI patients from CN. Conclusions: Results from both assays suggest strong potential as suitable methods for quantitating pTau181 in blood for use as a biomarker in AD. Additional data is required to validate these assays and demonstrate the value of this measure in interventional studies.

P255 / #1284


ALTERNATIVE PROTOCOLS TO SUPPORT BLOOD COLLECTION IN CHALLENGING PRE-ANALYTICAL CONDITIONS

Lecture Title:

J. Simrén1,2, K. Blennow1,2, H. Zetterberg1,2,3,4, N. Ashton1,5,6 1University of Gothenburg, Department Of Neurochemistry And Psychiatry, Institute Of Neuroscience And Physiology, Sahlgrenska Academy, Gothenburg, Sweden, 2Clinical Neurochemistry Laboratory, Sahlgrenska University Hospital, Mölndal, Sweden, 3UCL Institute of Neurology, Department Of Neurodegenerative Disease, London, United Kingdom, 4UCL, Uk Dementia Research Institute, London, United Kingdom, 5University of Gothenburg, Wallenberg Centre For Molecular And Translational Medicine, Gothenburg, Sweden, 6King's College London, Institute Of Psychiatry, Psychology & Neuroscience, Maurice Wohl Clinical Neuroscience Institute, London, United Kingdom

Aims: To investigate alternative preanalytical handling to aid the analysis of plasma Neurofilament Light (NfL) for AD and other neurological diseases in remote settings. Methods: As reference, plasma was obtained using standard protocols from n=10 individuals being investigated at the Sahlgrenska University Hospital, Mölndal. Whole blood was collected from each individual and; i) spotted onto Noviplex™ dry plasma spot (DPS) cards; ii) Whatman 903 protein saver dry blood spot (DBS) cards and; iii) left at room temperature for 48 hours as whole blood. In another data set (n=21), whole blood was left for 48 hours; either incubated at 37°C or shaken for 48 hours, aiming to simulate remote conditions or transportation of whole blood. This was compared to Noviplex™ DBS and standard NfL. Single molecule array (Simoa) HD-X analyzer measured NfL (NF-light®, Quanterix, Bilerica, MA). Spearman correlations were employed to investigate the associations between novel methods and the reference. Results: Plasma stored at room temperature (Rho=1.0, P<0.0001) and Noviplex™ DPS (Rho=0.933, P<0.001) strongly correlated with plasma, whereas Whatman DBS (Rho=0.782, P<0.01) was slightly less accurate. Results from the pilot study were then in large replicated (DPS: Rho=0.842, P<0.0001). High temperature exposure (Rho=0.989, P<0.0001) and shaking (Rho=0.983, P<0.0001) of the samples did not affect NfL concentrations. Conclusions: This study suggests that Noviplex™ DPS or delayed processing of whole blood are feasible alternative sample handling methods for plasma NfL. This can have advantages of, i) no processing required at collection and ii) measurement of NfL in remote regions where immediate access to advanced laboratory equipment is limited.

P256 / #1408


PLASMA AMYLOID-BETA STABILITY AND PRE-ANALYTICAL PROCEDURES

Lecture Title:

J. Vogelgsang1,2, J. Wiltfang2,3, M. Walter2 1McLean Hospital, Harvard Medical School, Dpartment Of Basic Neuroscience, Translational Neuroscience Laboratory, Belmont, United States of America, 2University Medical Center Goettingen (UMG), Department Of Psychiatry And Psychotherapy, Goettingen, Germany, 3German Center for Neurodegenerative Diseases (DZNE), -, Goettingen, Germany

Aims: Pre-analytical sample handling and storage conditions seem to play an important role on Amyloid-β (Aβ) stability, impacting the reliability and reproducibility of the AD-specific biomarkers. While recent studies on blood-based Alzheimer’s disease (AD) biomarker suggest a high association with cerebral Aβ pathology, a less invasive blood test might be a valuable screening tool for AD pathology. Methods: To elucidate the impact of pre-analytical sampling conditions on plasma Aβ peptides, we measured Aβ levels in venous and capillary blood under different storage conditions. Results: Important pre-analytical parameters affecting the stability of Aβ40 and Aβ42 besides the processing time is the effect of storage temperature showing a high stability of Aβ peptides at lower temperatures. Aβ40, Aβ42 and Aβ42/40 levels of whole blood or plasma stored at room temperature significantly start to decrease 6 hours after sampling. While Aβ42 was most prone to the time of storage, Aβ42/40 was stable up to 8 hours. At 4°C, Aβ peptide concentrations were stable up 72 hours. In addition, capillary blood Aβ42/40 could be measured and remained stable up to 72 hours at 4°C. Conclusions: Improving and establishing a standardized procedure in pre-analytical handling and storage condition is essential for reliable Aβ measurements in blood. These findings provide information that need to be respected in pre-analytical standard operating procedures for clinical and research practice to generate most reliable plasma Aβ measurements. Furthe,r we showed that capillary blood-based analysis of Aβ peptides might be a future tool in AD diagnostics.

P257 / #1801


MEASUREMENT OF PLASMA AND SERUM P-TAU181 IN ALZHEIMER’S DISEASE USING THE FULLY-AUTOMATED LUMIPULSE G PLATFORM: A FEASIBILITY STUDY

Lecture Title:

E. Wilson1, M. Swarovski1, M. Vandijck2, B. Lind3, D. Channappa4, J. Hall1, N. Le Bastard2, V. Henderson1, F. Longo5, J. Quinn3, K. Andreasson1 1Stanford University, Neurology, Stanford, United States of America, 2Fujirebio Europe N.V., Alzheimer's Disease, Ghent, Belgium, 3Portland VA Medical Center, Neurology, Seattle, United States of America, 4Stanford University, Pathology, Stanford, United States of America, 5Stanford University, Neurology, Palo Alto, United States of America

Aims: Promising therapeutic agents addressing the underlying biology of Alzheimer’s disease (AD) have renewed the call for fast, reliable blood-based biomarkers for wide scale population screening. Plasma P-Tau181 has high accuracy in distinguishing early AD from other neurodegenerative diseases. The Lumipulse G platform from Fujirebio is fully-automated and would facilitate high-throughput testing at a low turn-around time. We sought to evaluate the diagnostic performance of a modified version of the Lumipulse G P-Tau181 CSF test when applied to plasma and serum. Methods: Participants were enrolled in the Alzheimer’s Disease Research Centers at Stanford University and Oregon Health & Science University. The plasma cohort consisted of 37 control participants, 12 MCI, and 8 AD participants. The serum cohort was comprised of 11 Controls, 4 MCI, and 20 AD. ROC curve analysis compared P-Tau181 levels from plasma and serum in distinguishing AD from Controls. Results: Plasma and serum P-Tau181 were significantly elevated in AD participants compared to MCI and Controls. Analyte concentrations were lower in serum compared to plasma. P-Tau181 distinguished AD from control participants in plasma (AUC: 0.934) and serum (AUC: 0.907). Plasma and serum P-Tau181 were positively associated with CSF P-Tau181 in amyloid-positive participants and were positively associated with impaired cognition using the Montreal Cognitive Assessment (MoCA) and Mini-Mental State Exam (MMSE). Conclusions: These results support further investigation into the potential application of this modified Lumipulse G P-Tau181 CSF protocol for wide scale AD screening.

P258 / #498

Topic: Theme A: β-Amyloid Diseases / A4.i. Imaging, Biomarkers, Diagnostics: EEG, brain mapping, MEG

THE DELETERIOUS IMPACT OF AMYLOID-BETA OLIGOMERS ON SLEEP IN ALZHEIMER’S DISEASE

Lecture Title:

A. Hector1,2, C. Provost2, V. Mongrain2,3, J. Brouillette1,2 1Université de Montréal, Pharmacology And Physiology, Montréal, Canada, 2Centre de Recherche du CIUSSS du Nord de l'île de Montréal, Neurophysiology, Montréal, Canada, 3Université de Montréal, Neurosciences, Montréal, Canada

Aims: Synapse loss and ensuing neuronal death are the best predictors of memory deficits in Alzheimer’s disease (AD). There is mounting evidence from recent studies that soluble low-molecular-weight amyloid-beta oligomers (Aβo), especially oligomers derived from Aβ1-42 peptides, are the most neurotoxic species and correlate extensively with memory deficits in AD patients and animal models. It is also well-established that sleep loss impairs the function of the hippocampus, and that sleep alterations are among the first clinical symptoms observed in AD. The main objective of this project is to determine the impact of soluble Aβo-induced neurodegeneration on sleep architecture in rats. We also propose to identify molecular mechanisms underlying Aβo-driven hippocampal neurodegeneration. Methods: We performed chronic hippocampal injections of soluble Aβ1-42 oligomers in rats and electroencephalographic (EEG) measurements were performed to assess sleep alterations. The effects of Aβo on signaling pathways were analyzed by Western blot and immunofluorescence. Results: Although we observed no change in the time animals spent in REM sleep, NREM sleep or awake , our preliminary results suggest that the alpha waves are decreased after six days of Aβ injection. Sleep architecture and power spectral analyses will be investigated across the six days of injection to determine how the progression of Aβ pathology affects sleep, and if it is associated with a progressive increase of neuroinflammation within the hippocampus. Conclusions: Identifying the specific signature of hippocampal neurodegeneration on sleep features might serve as a non-invasive marker of early AD.

P259 / #1827

Topic: Theme A: β-Amyloid Diseases / A4.i. Imaging, Biomarkers, Diagnostics: EEG, brain mapping, MEG

GENETIC ASSOCIATION OF PICALM GENOTYPE WITH COMPLEXITY AND FUNCTIONAL CONNECTIVITY OF EEG IN NON-DEMENTED ADULTS

Lecture Title:

N. Ponomareva1, T. Andreeva2, M. Protasova3, A. Mitrofanov4, V. Fokin1, S. Illarioshkin1, E. Rogaev5 1Research Center of Neurology, Brain Research, Moscow, Russian Federation, 2Vavilov Institute of General Genetics of Russian Academy of Science, Department Of Human Genetics And Genomics, Moscow, Russian Federation, 3Vavilov Institute of General Genetics of Russian Academy of Science, Moscow, Russia., Department Of Human Genetics And Genomics,, Moscow, Russian Federation, 4Research Center of Mental Health, Russia, Fgbnu, Moscow, Russian Federation, 5University of Massachusetts Medical School,, Department Of Psychiatry, Worcester, MA, United States of America

Aims: PICALM genotype is a validated genetic risk factor for Alzheimer’s disease (AD). In particular, the PICALM rs3851179 A allele has a protective role, while the G allele serves as a strong genetic risk factor. Recent studies have revealed that genetic predisposition for AD can modify cortical functional connectivity and complexity many years before clinical onset of AD . The aim of this study was to analyze the association of PICALM rs3851179 genotype with complexity estimated by Shannon spectral entropy (SSE) and lagged linear connectivity (LLC) of spontaneous EEG in non-demented adults Methods: We examined the association of the PICALM rs3851179 genotype with the characteristics of LLC and SSE of spontaneus EEG in 130 non-demented adults (age range: 20-79 years). The LLC analysis was performed using exact low-resolution brain electromagnetic tomography (eLORETA) freeware (Pascual-Marqui et al., 2011). Results: We found that the carriers of the PICALM GG genotype have lower interhemispheric LLC of alpha sources compared to the carriers of protective PICALM A allele (PICALM AA and AG genotypes). Brain activity in the carriers of PICALM GG genotype was also characterized by lower SSE values in left temporal and frontal regions than in the carriers of PICALM A allele. Conclusions: Genetic risk for AD in the non-demented carriers of PICALM rs3851179 GG genotype is associated with lower cortical functional connectivity and complexity of brain networks. Cost-effective non-invasive identification of predisposition for AD using EEG biomarkers may provide the possibility for prophylactic before the appearance of any clinical symptoms.

P260 / #1407

Topic: Theme A: β-Amyloid Diseases / A4.j. Imaging, Biomarkers, Diagnostics: Cognitive, psychometric & behavioral tests

PENTAGON COPYING TASK AS A KEY INDICATOR FOR DIFFERENTIATING ALZHEIMER’S DISEASE FROM DEMENTIA WITH LEWY BODIES

Lecture Title:

D. Bakir1, S. Goel1, T. Ala2 1Southern Illinois University School of Medicine, Neuroscience Institute: Center For Alzheimer's Disease And Related Disorders, Springfield, United States of America, 2Southern Illinois University School of Medicine, Cenetr For Alzheimer's Disease And Related Disorders, Springfield, United States of America

Aims: It is often difficult to differentiate dementia with Lewy bodies (DLB) from Alzheimer’s disease (AD). There is no biomarker that reliably diagnoses DLB, and there are many overlapping features with AD. Previous work has shown that DLB patients often have marked visuospatial and constructional difficulty in contrast to AD patients. Therefore, the purpose of this study was to determine if the pentagon copying task of the Mini-Mental State Exam (MMSE) is a helpful parameter in distinguishing DLB from AD. Methods: MMSEs are routinely administered in our memory clinic. The MMSEs were reviewed for 136 consecutive AD patients and 24 consecutive DLB patients (AD: MMSE average 20.0 ± 6.0, range 1-30; DLB: MMSE average 21.8 ± 5.1, range 10-28). The pentagon copying scores of the two groups were compared using the original MMSE binary ("correct" or "incorrect") scoring criteria (two figures, each with five sides, with correct intersection). Results: The ability of the DLB patients to copy the pentagons was more impaired than the DLB patients (p=0.0248). An incorrect copy of the pentagons predicted DLB with sensitivity 67%, specificity 60%, positive predictive value 23%, and negative predictive value 91%. Conclusions: The pentagon copying task of the MMSE is useful to distinguish DLB from AD. Whereas the task had marginal sensitivity, marginal specificity, and poor positive predictive value, it had good negative predictive value for the diagnosis of DLB. Considering our population of LBD and AD patients, those who could copy the pentagons correctly were 91% more likely to have AD.

P261 / #993


AGE-RELATED DIFFERENCES IN THE COGNITIVE CONNECTOME USING GRAPH THEORY

Lecture Title:

E. Garcia Cabello1, L. González Burgos1, J.B. Pereira2, J.A. Hernandez Cabrera1, E. Westman2, G. Volpe2, D. Ferreira2, J. Barroso1 1University of La Laguna, Department Of Clinical Psychology, Psychobiology And Methodology, San Cristóbal de La Laguna, Spain, 2Karolinska Institutet, Division Of Clinical Geriatrics, Center For Alzheimer Research, Department Of Neurobiology, Care Sciences And Society, Stockholm, Sweden

Aims: Objectives: Cognitive aging has been extensively investigated using univariate analyses. However, more sophisticated approaches are required in order to capture the complex interactions between multiple cognitive variables. Our objective was to study the interrelationship between multiple cognitive variables during aging with graph theory analysis applied to cognitive data. Methods: A total of 334 cognitively unimpaired individuals were divided into early middle-age (37-50 years), late middle-age (51-64 years) and elderly (65-78 years) groups. We analysed 47 cognitive variables using BRAPH (BRain Analysis using graPH theory, http://braph.org/) to build and characterize cognitive networks. Results: We identified a modular organization of 5 communities: verbal memory, visual memory/visuospatial, procedural memory, executive functions and processing speed. A pattern of segregation was observed in the elderly group, characterized by reduced transitivity and strength, and increased efficiency compared with the early middle-age group. The late middle-age group showed lower efficiency and modularity than the early middle-age group. Nodal analyses showed the important role of executive and processing speed variables in explaining the differences between age groups. Conclusions: We conclude that graph theory is useful to capture complex interactions between multiple cognitive variables and showed age-related differences in several features of the cognitive networks. Our results suggest potential compensatory mechanisms to maintain cognitive performance during late middle-age, which seem to be less effective in elderly subjects.

P262 / #1445


DETECTION OF EPISODIC MEMORY IMPAIRMENT IN MCI USING THE TABLET-BASED BREIF ASSESSMENT OF COGNTION (BAC)

Lecture Title:

S. Karas1, A. Atkins2, H. Stevens3, K. Welsh-Bohmer1,4 1VeraSci, Neurodegenerative Disorders, Durham, United States of America, 2VeraSci, Scientific Development, Durham, United States of America, 3VeraSci, Data Integration And Product Solutions, Durham, United States of America, 4Duke University Medical Center, Psychiatry, Durham, United States of America

Aims: The aim of this work is to assess the utility of the tablet-based Brief Assessment of Cognition (BAC) to identify and quantify episodic memory deficits in MCI. Methods: Participants include 239 Healthy Older Controls (HC), and 36 participants with clinical diagnoses of MCI. All participants completed an expanded version of the BAC to assess 6 cognitive domains: episodic verbal memory, processing speed, working memory, verbal fluency, motor function and executive function. Data were analyzed using independent t-tests. Z-scores were generated to describe demographically-corrected performance of the MCI group relative to a robust normative data set of 600 + healthy controls. Results: Participants with MCI performed significantly worse on BAC App measures of episodic memory, visuospatial memory, and processing speed (p<.007 for all). No group differences occurred on verbal fluency, digit sequencing or Tower of London. On episodic memory (delayed free recall), the MCI group (M = 4.94, SD = 4.05) performed significantly worse compared to the HC (M = 9.46, SD = 3.18; t(272) = 7.55, p < .001). The MCI group Z-scores for episodic memory measures fell 1 to 1.5 standard deviations below the mean, consistent with recent criteria for clinically diagnosed MCI (Alberts et al 2011). Conclusions: Findings confirm the utility of the tablet-based BAC to assess objective cognitive impairments in MCI and preclinical AD. Further, the observed MCI episodic memory performance of 1 to 1.5 standard deviations below the normative mean support use of the BAC in screening and evaluation of established diagnostic criteria.

P263 / #642


THE EFFECT OF APOE AND BDNF VAL66MET POLYMORPHISMS ON SPATIAL NAVIGATION IN SUBJECTIVE COGNITIVE DECLINE

Lecture Title:

J. Laczó1,2, K. Cechova1,2, M. Parizkova1,2, O. Lerch1,2, V. Matuskova1,2, Z. Nedelska1,2, M. Vyhnalek1,2, J. Hort1,2 1Department of Neurology, Charles University, 2nd Faculty Of Medicine And Motol University Hospital, Prague, Czech Republic, 2International Clinical Research Center, St. Anne’s University Hospital Brno, Brno, Czech Republic

Aims: The apolipoprotein E (APOE) ɛ4 allele is associated with episodic memory deficits and less accurate spatial navigation in nondemented older adults. The brain-derived neurotrophic factor (BDNF) Met allele may further worsen memory impairment in APOE ɛ4 carriers but its role in APOE ɛ4-related spatial navigation deficits has not been established. We examined the influence of combination of the APOE and BDNF Val66Met polymorphisms on spatial navigation, cognitive functions and volumes of selected brain regions associated with spatial navigation including the hippocampus, entorhinal cortex, precuneus, inferior parietal and posterior cingulate cortex in individuals with subjective cognitive decline (SCD). Methods: In total, 71 older adults with SCD from the Czech Brain Aging Study were stratified based on the APOE and BDNF Val66Met polymorphisms into four groups: ɛ4–/BDNFVal/Val (n =27), ɛ4–/BDNFMet (n = 15), ɛ4+/BDNFVal/Val (n =18), and ɛ4+/BDNFMet (n=11). The participants underwent comprehensive neuropsychological examination, brain MRI and spatial navigation testing of egocentric, allocentric, and allocentric delayed navigation in a real-space human analogue of the Morris water maze. Results: The ɛ4+/BDNFMet group, although similar to other polymorphism groups in demographic characteristics and global cognitive function, had less accurate allocentric and allocentric delayed navigation performance than the ɛ4–/BDNFVal/Val group (p<0.05). There were no significant differences in egocentric navigation performance, cognitive performance in any cognitive domain and volumes of selected brain regions associated with spatial navigation between the polymorphism groups (p>0.05). Conclusions: The combination of the APOE ɛ4 and BDNF Met polymorphisms is associated with less accurate allocentric spatial navigation in individuals with SCD.

P264 / #1379


COGNITIVE PATTERNS OF FAST AND SLOW PROGRESSION IN ALZHEIMER’S DEMENTIA: A FEASIBILITY STUDY USING ADNI DATA

Lecture Title:

T. Matosevic Institute of Neurology, Institute Of Neurology, London, United Kingdom

Aims: The aims of this study are to identify which cognitive tests are good at predicting a diagnostic status at 48 months follow-up and to assess whether the selected cognitive measures can be used to differentiate between slow and fast cognitive decliners. Methods: The study uses a longitudinal data from the Alzheimer’s Disease Neuroimaging Initiative. A sample of 97 cognitively normal (CN), 77 mildly cognitively impaired (MCI) and 46 individuals with AD is included in the analyses. Results: The sensitivity analysis indicates that overall cognitive and functional tests are more sensitive to detecting cognitive changes in MCI than AD individuals as compared to CN. The findings show that the predictive diagnostic accuracy for MCI, the Clinical Dementia Rating Sum of Boxes (CDRSB), the Mini-Mental State Examination and the Delayed recall tests are good predictors of diagnostic status at 48 months. The results for AD indicate that, at 48 months, only a CDRSB test has a significant diagnostic accuracy. The progression analysis reveals that CDRSB scores at baseline, Functional Activities Questionnaire and Everyday Cognition study partner measures are best at differentiating between the fast and slow cognitive decliners. Conclusions: The findings represent an important contribution in identifying which cognitive measures are best at diagnosing preclinical AD and in monitoring cognitive decline in AD.

P265 / #715


PERSPECTIVE TAKING AND ITS STRUCTURAL CORRELATES IN EARLY ALZHEIMER´S DISEASE

Lecture Title:

M. Parizkova1,2, J. Kalinova1, Z. Nedelska1,2, M. Vyhnalek1,2, J. Hort1,2, J. Laczó1,2 1Department of Neurology, Charles University, 2nd Faculty Of Medicine And Motol University Hospital, Prague, Czech Republic, 2International Clinical Research Center, St. Anne’s University Hospital Brno, Brno, Czech Republic

Aims: Spatial orientation assessment could be used for early diagnosis of Alzheimer´s disease (AD). We aimed to evaluate the potential of perspective taking tests to identify individuals with early AD and to determine associations between perspective taking and volumes of the selected brain regions affected in early AD. Methods: 66 participants with amnestic mild cognitive impairment (aMCI) and positive AD-biomarkers (aMCI due to AD, n=22), aMCI and negative biomarkers (aMCI AD-negative, n=15), mild AD dementia (n=7) and cognitively normal (CN) older adults (n=22) underwent clinical and neuropsychological evaluation, MRI brain scan with automated volumetry, AD-biomarker assessment and two perspective taking tasks: Standardized Road-Map test of Direction Sense (RMTDS) and Perspective Taking/Spatial Orientation Test (PTSOT). In the RMTDS, the participants followed a pathway on a city map indicating a direction of turning at each intersection. The PTSOT included pictures of arrays of objects and participants indicated direction between selected objects. Results: The aMCI due to AD and mild AD dementia groups had lower scores than the CN group (p<0.05) in the PTSOT. The aMCI due to AD group had a lower score compared to the CN group (p<0.05) in the RMTDS. There were no differences between the CN and aMCI AD-negative groups in both tests. PTSOT scores correlated with volumes of the right inferior temporal gyrus and right and left precuneus. Conclusions: The perspective taking tests reliably detect spatial orientation impairment typical for early stages of AD. Worse performance in the PTSOT was associated with smaller volumes of the brain regions affected early in AD.

P267 / #429


DIFFERENTIAL DIAGNOSIS OF CORTICAL AND SUBCORTICAL DEMANTIAL SYNDROMES BY PRAXIS TESTS

Lecture Title:

Z. Sanli1, A. Evlice2 1Adana Sehir Training and Research Hospital, Neurology, Adana, Turkey, 2ÇUKUROVA UNİVERSİTY, Neurology, adana, Turkey

Aims: It is known that cortical dementia ,such as Alzheimer's disease, frequently has high cerebral dysfunction such as apraxia. In this study, it was aimed to determine whether practical bedside praxia tests can be useful in differential diagnosis of cortical and subcortical dementia syndromes. Methods: 34 Alzheimer 's disease,31 Parkinson's disease dementia,29 mild cognitive impairment and28 healthy subjects followed at Outpatient Clinic at Department of Neurology Cukurova University Faculty, were included in the study.Complete blood count, liver/kidney /thyroid function tests,vitamin b12,folic acid levels and cerebral imaging were evaluated. In addition,minimental sititation test(MMSE), clock drawing test(CDT) , and DEKODa and AST tests which are the tests of experimental status and apraxia screening tests were performed.All data were comparedamong the groups.SPSS version 21.0 was used for statistical analysis of the data,p <0.05 was considered statistically significant. Results: There was a significant difference between all groups in terms of MMSE, CDT, DEKODa and AST scores(p <0.001).DEKODa and AST test scores were significantly lower in Alzheimer's disease compared to the other groups.The cut-off value was 12 for DEKODa and 10.5 for AST.Sensitivity and specificity for DEKODa were 76.5% and 100%, respectively; For AST, the sensitivity was 79.4% and the specificity was 100%. Conclusions: This work has shown that DEKODa and AST tests can be useful differentiate Alzheimer's disease from subcortical dementia. In addition, the DEKODa test, which is used to evaluate apraxia, was observed to be close to the AST test

P268 / #353


USING MACHINE LEARNING TO DEVELOP A CLINICAL MODEL THAT PREDICTS DEMENTIA SEVERITY

Lecture Title:

J. Pakanati1, K. Kolev1, E. Schonfeld2, R. Hunter1, K. Hussain1, M. Meredith3, E. Schonfeld4, R. Senno1 1Rosalind Franklin University, Chicago Medical School, North Chicago, United States of America, 2Stanford University, School Of Humanities And Sciences, Stanford, United States of America, 3University of Illinois at Urbana-Champaign, College Of Liberal Arts And Sciences, Urbana, United States of America, 4Glenbrook North, High School, Northbrook, United States of America

Aims: Clinical applications of machine learning in dementia include predicting both diagnosis and severity. Many current methods are MRI/PET scan based; however, often in the long term care setting, scans are either not accessible or outdated. Clinical testing, such as the Mini Mental State Examination, is biased towards aphasia, poor vision, deafness, educational level, stroke, and psychosis among others. Using machine learning techniques, we develop a non-linear model to predict cognitive impairment in dementia only using clinical variables from medical charts. Methods: With IRB approval, 153 patients with dementia in three long term memory care assisted living communities were administered the MMSE to assess level of cognition; medical and social variables were gathered from medical charts. To limit overfitting, clinical variables were mapped to a feature vector composed of most relevant variables. Linear regression with L2 loss was performed to determine the model’s parameters. Results: The model predicted MMSE score; the score was classified as mild, moderate, or severe. The confusion matrix for validation and test sets shows high accuracy (≤ 65%), demonstrating proof of concept for a clinical model. The algorithm performed especially well for those with severe dementia. Conclusions: The model can be used in research and has potential clinical applications for non-participatory patients. The model can be used for triage. By combining imaging with clinical data, more advanced models can be developed for predictive Alzheimer's care. These may include connecting our feature vector, with its associated parameters, to dense layers of deep convolutional models based on MRI/PET to measure dementia progression.

P269 / #1737


DYNAMIC COGNITIVE RESERVE AS A PREDICTOR OF DAILY COGNITIVE AND BEHAVIORAL DEFICITS IN MCI

Lecture Title:

J. Točík1,2, I. Rektorova1,2 1Faculty of Medicine, Masaryk university, First Department Of Neurology, Brno, Czech Republic, 2Central European Institute of Technology, Masaryk University, Applied Neuroscience Research Group, Brno, Czech Republic

Aims: The objective of the study was to evaluate whether residual episodic memory variance is useful in the prediction of the cognitive and behavioral deficits in patients with amnestic MCI. Methods: We used neurocognitive and neuroimaging data from 686 patients with aMCI from the Alzheimer's Disease Neuroimaging Initiative (ADNI) database. To estimate the episodic memory reserve, we regressed Rey's Verbal Learning Test results at baseline on the available volumetric data. We tested whether this dynamic cognitive reserve indicator does predict deterioration of daily cognitive and behavioral problems assessed by the clinician (Clinical Dementia Rating Scale), by the patient (ECog scale) and by the patient's significant other (also ECog scale). Results: The level of memory reserve predicted the rate of cognitive and behavioral decline as measured by CDR-SB. Over four years period, patients with a lower reserve showed a significantly higher rate of decline than those with a higher reserve level. Similar results were also seen in the rate of deterioration reported by a significant other. We found no moderation effect of dynamic cognitive reserve on the rate of self-reported behavioral and cognitive decline. Conclusions: Dynamic cognitive reserve estimated as residual episodic memory variance is an important predictor of the rate of cognitive and behavioral decline in patients with aMCI. Patients with higher levels of reserve showed a slower rate of decline assessed by the clinician and reported by the significant other. This recently developed indicator of cognitive reserve calls for further research on its potential clinical relevance of predicting everyday difficulties in aMCI.

P270 / #303


ACCELERATED LONG-TERM FORGETTING IN INDIVIDUALS WITH SUBJECTIVE COGNITIVE DECLINE AND AMYLOID-BETA POSITIVITY

Lecture Title:

A. Tort-Merino1, N. Valech1, M. Laine2, J. Olives1, M. León1, M. Ecay-Torres3, A. Estanga3, P. Martínez-Lage3, J. Fortea4, J.L. Molinuevo1,5, R. Sánchez-Valle1, A. Rodriguez-Fornells6,7,8, L. Rami1 1Alzheimer’s Disease and Other Cognitive Disorders Unit, Neurology Service, Hospital Clínic of Barcelona, Institut D'investigacions Biomèdiques August Pi I Sunyer (idibaps), Barcelona, Spain, 2Department of Psychology, Åbo Akademi University, Turku, Finland, 3Fundación CITA-Alzhéimer Fundazioa, Centro De Investigación Y Terapias Avanzadas, Donostia, Spain, 4Memory Unit, Sant Pau Biomedical Research Institute, Hospital de la Santa Creu i Sant Pau, Neurology, Barcelona, Spain, 5Barcelonaβeta Brain Research Center, Pasqual Maragall Foundation, Barcelona, Spain, 6Cognition and Brain Plasticity Group, Bellvitge Biomedical Research Institute-idibell, L’Hospitalet de Llobregat, Spain, 7Catalan Institution for Research and Advanced Studies, Icrea, Barcelona, Spain, 8Department of Cognition, Development and Education Psychology, University Of Barcelona, L'Hospitalet de Llobregat, Spain

Aims: Accelerated long-term forgetting (ALF) has been defined as a rapid loss of information over days or weeks despite normal acquisition/encoding. We investigated ALF in cognitively unimpaired individuals with and without subjective cognitive decline (SCD). We also explored the relationships between the performance on a new highly demanding learning and memory test (the Ancient Farming Equipment Test; AFE-T), subjective cognitive ratings measured by the Subjective Cognitive Decline Questionnaire (SCD-Q), and cerebrospinal fluid (CSF) Alzheimer’s disease (AD) biomarkers. Methods: Fifty-two individuals were included, and SCD was quantified through the SCD-Q using its validated cutoff to classify subjects as non-SCD (n=21) or SCD (n=31). SCD subjects were further subdivided according to the presence or absence of abnormal Aβ42 levels (SCD/Aβ+ or SCD/Aβ-). The AFE-T requires acquisition, maintenance and recall of novel object/name pairs. Its comprehensive design encompasses two learning sessions and a long-term maintenance/recall phase with free and cued recall at one week, three months, and six months after the initial learning. Results: The SCD group showed a significantly higher free forgetting rate at 3 months compared to the non-SCD (F(1,44)=4.72; p<0.05; Fig. 1). When stratifying by amyloid status, SCD/Aβ+ showed a significantly lower performance than SCD/Aβ– on the learning scores and on the 1-week cued forgetting rate and 3-month recall of the AFE-T (all p<0.05; Fig. 2). Higher SCD-Q scores predicted higher forgetting rates on the AFE-T.

Conclusions: By using a highly demanding learning and memory task, it is possible to detect ALF in subjects with SCD, especially in those with abnormal CSF Aβ42 levels.

P271 / #1623

Topic: Theme A: β-Amyloid Diseases / A4.k. Imaging, Biomarkers, Diagnostics: Other

MORPHOLOGICAL TYPING OF DIVERSE ABETA DEPOSITS IN AD USING FLUORESCENT PROBES

Lecture Title:

R. Al-Lahham, A. Mukherjee, C. Soto University of Texas Health Science Center at Houston, The Mitchell Center For Alzheimer’s Disease And Related Brain Disorders, Department Of Neurology, Houston, United States of America

Aims: Accumulation of Aβ into amyloid plaques (APs) and tau into neurofibrillary tangles (NFTs) is a pathological hallmark of Alzheimer’s disease (AD). Aβ can deposit in a spectrum of morphology from diffuse to fibrillar, and dense-cored plaques. The pathological relevance of these varied morphologies in AD remains unclear. Inter- and intra-individual diversity in Aβ aggregates may be associated with substantial clinical heterogeneity in AD. Our goal is to develop an array of fluorescent dyes to probe the conformational and morphological diversity of Aβ accumulates in AD Methods: We screened a small library of fluorescent compounds with in vitro generated Aβ aggregates. Subsequently, we validated the hit by staining paraffin-embedded sections of frontal and temporal cortex as well as midbrain from AD, LBD patients and healthy control. Mirror image sections were used for immunostaining with 4G8, AT8, phospho-α-synuclein and α-synuclein Results: We identified a novel compound PMDAM006 that selectively stained Aβ accumulates, both parenchymal and vascular, in AD brain sections and displayed no preference for NFTs in AD and Lewy bodies in LBD. No apparent staining is observed in healthy control brain. Interestingly, derivatives of this novel scaffold, PMDAM022 and PMDAM015, demonstrated selective preference for different types of Aβ accumulates. PMDAM0022 also stained NFTs and LBD, while PMDAM015 did not Conclusions: We believe that these novel fluorescent probes will help us understand the physiological relevance of Aβ plaque diversity and help address the heterogeneity observed within AD. Consequently, these probes will facilitate the development of novel and refined diagnostic tools for early detection of AD

P272 / #1279


RETINAL VASCULAR PATHOLOGY IN PRECLINICAL ALZHEIMERʼS DISEASE: EARLY OCULOVASCULAR BIOMARKERS USING SWEPT SOURCE-OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY IN VIVO

Lecture Title:

S. Asanad1,2, M. Fantini3, W. Sultan2, M. Nassisi4, C. Felix5, R. Karanjia2,6, M. Harrington7, A. Sadun2 1University of Maryland School of Medicine, Ophthalmology & Visual Sciences, Baltimore, United States of America, 2Doheny Eye Institute - UCLA, Neuro-ophthalmology, Los Angeles, United States of America, 3University of Udine, Ophthalmology, Udine, Italy, 4Doheny Eye Institute - UCLA, Ophthalmology, Los Angeles, United States of America, 5David Geffen School of Medicine, UCLA, Ophthalmology, Los Angeles, United States of America, 6University of Ottawa, Ophthalmology, Ottawa, Canada, 7Huntington Medical Research Institutes, Neuroscience, Pasadena, United States of America

Aims: We evaluated the retinal vasculature using swept-source optical coherence tomography angiography (SS-OCTA) in preclinical Alzheimerʼs Disease (AD) classified by cerebral spinal fluid (CSF) β-amyloid/Tau ratio. Methods: Cognitively healthy (CH) participants were classified into two sub-groups, normal (CH-NAT) or pathological (CH-PAT), using a logistic regression model from the CSF β-amyloid/Tau ratio that identified >85% of patients with clinically probable AD. SS-OCTA was performed in 16 CH-PATs (16 eyes) and 16 CH-NATs (16 eyes). Macular (6x6 mm) scans were acquired for the superficial (SCP) and deep capillary plexus (DCP), as well as the choriocapillaris (CC). Vessel density (VD) was compared for the fovea and parafoveal sectors. Mann-Whitney U test was applied for statistical analysis. Results: Mean age (+/- standard deviation) in the CH-PAT group (75.2 +/- 8.4 years) was similar to the CH-NAT group (75.4 +/- 8.1 years; p=0.48). Total SCP was significantly attenuated in preclinical AD relative to controls (3% VD reduction; p < 0.03), most severe in the fovea (8.5%; p < 0.02) and the nasal parafovea (4%; p < 0.005). Total CC was also reduced in preclinical AD relative to controls, specifically within the nasal sector (3% reduction; p < 0.05). Total DCP was overall preserved in preclinical AD (p =0.3). Conclusions: SS-OCTA revealed vascular pathology in asymptomatic, preclinical AD. Perfusion defects corresponded with the vascular supply of the RNFL, which was structurally thinner in this same cohort. These early retinal vascular findings may potentiate the diagnostic value of in vivo ocular biomarkers for AD, prior to the development of symptoms.

P273 / #179


NONINVASIVE MAPPING OF CMRO2 IN THE APPSWE/PS1DE9 MOUSE BRAIN BY 17O-MRI

Lecture Title:

C. Baligand, J.-B. Perot, D. Thenadey, F. Petit, G. Liot, M.-C. Gaillard, O. Barret, J. Flament, M. Dhenain, J. Valette Université Paris-Saclay, CEA, CNRS, Laboratoire des Maladies Neurodégénératives : mécanismes, thérapies, imagerie, Molecular Imaging Research Center, Fontenay-aux-Roses, France

Aims: Accumulation of amyloid-beta has been associated with mitochondrial dysfunction.1 Therapies targeting mitochondrial impairments provide treatment opportunities for Alzheimer’s disease.2 In-vivo follow-up of disease and therapies requires new methods to monitor mitochondrial health. We implemented an MRI method to quantify the cerebral metabolic rate of oxygen consumption (CMRO2) in mouse models of amyloidosis and showed alterations of CMRO2 in APPswe/PS1dE9 mice. Methods: 17O is a stable isotope detectable by MRI. It is not MR-visible as a gas, nor while bound to hemoglobin, but becomes MR-visible once incorporated into water (H2

17O) as a waste of mitochondrial respiration, allowing direct non-invasive measure of O2 utilization. Thus, high-resolution 17O-MRI during inhalation of 17O-enriched gas provides a measure of CMRO2. Experiments were conducted at 11.7T in ~12-month-old APPswe/PS1dE9 (n=4) and wild-type (n=4) mice. A series of 3D 17O-MRI (Fig1b) was acquired (spatial resolution: 1.5x1.5x1.5mm3, time resolution: 17s) before, during (3.5min) and after inhalation of 70%-17O-enriched oxygen (Fig1d) to generate CMRO2 maps (Fig1c). Post-mortem analysis included RT-qPCR and histology. Results:

CMRO2 was significantly lower in APPswe/PS1dE9 than in WT mice (1.52±0.07 versus 1.27±0.08 μmol/g/min, Fig1e, **p=0.0028). Brain and ventricles volumes were not different between groups. Post-mortem analysis confirmed amyloid-beta plaques and astrocytes reactivity in APPswe/PS1dE9 . Mitochondrial biogenesis and mitochondrial complexes mRNA levels were overall down in APPswe/PS1dE9. Conclusions: 17O-MRI can detect decreased CMRO2, a marker of mitochondrial activity, in 12mo APPswe/PS1dE9 mice. This provides a new biomarker to monitor mitochondrial impairments in amyloidosis models and experimental therapies. References: 1)Yao et al. PNAS,2009,106,34,14670–14675; 2)Bonda et al., Drugs and Aging,2010,27,3,181-192.

P274 / #1310


CO-LOCALIZATION OF AMYLOID BETA PLAQUES AND EXOSOME MARKERS IN THE APP/PS1 MOUSE BRAIN.

Lecture Title:

R. De La Flor1, M. Alavijeh2, J. Inal3 1Pharmidex, Cns, Hatfield, United Kingdom, 2Pharmidex, Managing Director, Hatfield, United Kingdom, 3University of Hartfordshire, Biomedical Science, Hatfield, United Kingdom

Aims: Currently, improvement of cognitive deficits and/or reduction in amyloid plaques in transgenic models of Alzheimer’s disease (AD-Tg) is used as preclinical evidence of efficacy of novel chemical entities. Despite the validity of AD-Tg to study the pathology of AD, the predictive power of current cognitive/histopathology parameters in the models is still low. Taking advantage of current advances in isolation of neural derived exosomes from blood, we aim to identify disease progression and drug efficacy biomarkers in blood to improve the translational potential of well-established AD-Tg models. Methods: To validate the approach, we investigated the correlation between exosomes and amyloid beta plaques by performing a histological examination of an APP/PS1 overexpressing AD-Tg mouse brain using the exosome marker CD9 and an antibody for amyloid beta 1-14 region. Results: Our data showed a marked co-localisation of CD9 with plaques of amyloid beta in the overexpressing APP/PS1 mouse brain. Conclusions: This data suggests that brain derived exosome cargo is a valid source of AD pathophysiological biomarkers.

P275 / #1293


DEVELOPMENT OF NOVEL NEAR INFRARED FLUORESCENT (NIRF) IMAGING PROBE FOR EARLIER DIAGNOSIS OF ALZHEIMER’S DISEASE

Lecture Title:

H. Rai1, G. Shankar1, R. Renu2, S. Wadhwa1, S. Gupta3, S. Kumar2, G. Modi1 1Indian instittue of technology (BHU), Department Of Pharmaceutical Engineering & Technology, varanasi, India, 2All India Instittue of Medical Sciences, Department Of Biophysics, New Delhi, India, 3National Institute of Immunology, National Institute Of Immunology, New Delhi, India

Aims: Our objective is to develop a novel NIRF probe able to selectivity detect the soluble and/or insoluble Aβ aggregates at earlier stage and in long term to monitor the therapeutic effect of Beta-secretase 1 (BACE1) inhibitor. Methods: A) Design and synthesis of novel probes for detection of Aβ aggregates: In the present study, we have carried out the structural modifications in the compounds developed in our laboratory to impart donor-π- acceptor, the replacement of phenolic OH with N,N-dimethylaniline, etc. The developed probes were fully characterized with help of 1H & 13C NMR, and high-resolution mass spectroscopy. B) Preparation and characterization of Aβ1-42 aggregates was performed as per protocol followed by (Singh, Tej et al. 2020). C) The In silico ADMET predictions tool was used to predict chemical properties, and uv-spectroscopy was utilized an Autodock to theoretically investigated affinity and binding mechanism of probes with on Aβ1-42 protofibrils (PDB: 2LMO). Results: The calculation of physiochemical properties indicated the drug-like properties of the developed molecules. The UV absorption measurement in PBS implicated the potential of the developed probes as imaging agents with optimal absorption maxima (λexcitaion) ranging from 408 nm to 560 nm at 100 Conclusions: The findings clearly indicate that these derivatives are worthy of further investigation. The detailed study be presented.

P276 / #772


DEVELOPING A SALIVA-BASED POLYGENIC RISK SCORING TEST FOR EFFECTIVE STRATIFICATION OF POTENTIAL ALZHEIMER’S DISEASE RELATED CLINICAL TRIAL PARTICIPANTS

Lecture Title:

A. Gibson1, R. Pither1, P. Daunt2, C. Ballard3, B. Creese3, G. Davidson4, T. Hill1, O. Oshota2 1Cytox, R&d, Oxford, United Kingdom, 2Cytox, R&d, Manchester, United Kingdom, 3University of Exeter, Medicine, Exeter, United Kingdom, 4Ledcourt Associates, R&d, Cambridge, United Kingdom

Aims: To develop a polygenic risk scoring (PRS) assessment for use in clinical trial recruitment strategies. Specifically, to identify those most likely to decline cognitively towards Alzheimer’s disease. Methods: Participants who entered the Alzheimer Disease Neuroimaging Initiative (ADNI) longitudinal study with a diagnosis of mild cognitive impairment (MCI) that have available genetic data and clinical follow up for at least 4 years were selected for this analysis. For each participant a PRS (0-1) was calculated and the population separated into higher and lower risk groups using a threshold of 0.6. The performance of the algorithm was tested for its ability to enrich a clinical trial population for those individuals most likely to decline cognitively over 4 years. Results: 285 participants with an MCI diagnosis at baseline were selected for this study, of which 61 individuals, declined by at least 10 ADAS-Cog13 points over 4 years. Using a threshold of 0.6, the PRS identified 54 of those cognitively declining individuals, whilst excluding 90 participants whose cognitive performance remained relatively stable over that period. Enrichment for individuals most likely to decline cognitively improved by 33% by using the PRS algorithm. Conclusions: Using DNA easily accessible from saliva, can provide a more efficient manner for identifying participants most likely to decline cognitively and therefore enriching clinical trials with more suitable patients. Using PRS algorithms prior to more invasive, expensive or burdensome procedures provides a strategy to screen out unsuitable patients very early in the recruitment process.

P277 / #483


IS THERE A GENERAL DYSREGULATION OF THE ENDOLYSOSOMAL COMPARTMENT IN FIBROBLASTS OF AD PATIENTS?

Lecture Title:

L. Xicota1, S. Landron2, A. Botté1, C. Gautier3, J. Lagarde4, M. Sarazin4, M.-C. Potier1 1Paris Brain Institute, Alzheimer’s Disease And Prion Disease Group, Paris, France, 2Servier Research Institute, Pôle Sciences Cellulaires - Pex Biotechnologie, Croissy-sur-Seine, France, 3Servier Research Institute, External Research And Innovation, Croissy-sur-Seine, France, 4Université Paris Descartes, Sorbonne Paris Cité, INSERM UMR S894, Centre Hospitalier Sainte Anne, Neurologie De La Mémoire Et Du Langage, Service De Neurologie,, Paris, France

Aims: The objective of this study was i) to analyze the dysregulation of the endolysosomal pathway in peripheral cells of Alzheimer’s disease patients; ii) to identify a specific profile that would allow the use of such cells as biomarkers for disease risk assessment. Methods: Thirty fibroblast lines from Control, MCI, and AD from the IMABIO3 and SHATAU studies were analyzed (10/group). Cells were cultured in 96-well plates, fixed and immunofluorescence was performed against Rab5 and EEA1 (early endosomes), LBPA (late endosomes), and LAMP1 (late endosomes/lysosomes). Images were obtained through an automatic imager Opera Phenix. Statistical analyses were performed through either nested-ANCOVA with correction for age or Kruskal-Wallis depending on the number of samples and the variance, followed by a posthoc analysis. Results: Preliminary analyses (12 lines, 4 per group) shows a trend towards an increased mean area of EEA1 puncta in cells of MCI subjects as compared to controls (p=0.08), as well as a significant reduction of the number of EEA1 positive puncta in the AD group as compared to controls (p<0.01). ForRab5, LBPA and LAMP1, although there were several trends suggesting a dysregulation of the endolysosomal pathway, no significant results could be observed in this pilot study. We will present the final analysis of the 30 fibroblast lines. Conclusions: Our pilot (12 fibroblast lines) shows a general dysregulation of the endolysosomal pathway. Completion of the study (30 lines) should allow us to ascertain the potential use of peripheral fibroblasts as a biomarker of AD disease.

P278 / #1076

Topic: Theme A: β-Amyloid Diseases / A4.l. Digital and AI-Based biomarkers

PRECISION FUNCTIONAL ASSESSMENT FOR ALZHEIMER’S DISEASE (PFA-AD) USING DIGITAL TECHNOLOGY: A PILOT STUDY FRAMEWORK

Lecture Title:

G. Hernandez1, C. Lopez1, Y. Wang1, D. Wershiner2, T. Gisler2, V. Seyfert-Margolis2, R. Brinton1 1University of Arizona, Center For Innovation In Brain Science, Tucson, United States of America, 2My Own Med, My Own Med, Bethesda, United States of America

Aims: To develop the Precision Functional Assessment for Alzheimer’s Disease (PFA-AD) mobile application to enable: 1) identification of key behavioral challenges that are relevant to the person with AD and their caregivers; 2) assessment of therapeutic effect in real time across multiple domains of behavioral function; and 3) innovate clinical trial assessment of cognitive function. Methods: PFA-AD proof-of-concept development was embedded as part of a clinical trial (Clinicaltrials.gov:NCT03748303) to further develop allopregnanolone, a first-in-class regenerative therapeutic for AD. This study is enrolling 10 persons with AD to assess the intramuscular formulation of allopregnanolone. Participants consent to use and interact with the “Allo IM app” daily for the duration of the 12-week trial period. The digital platform, created by MyOwnMed (MOM), captures first person patient experience, clinical treatment efficacy and outcomes from real world patient experiences. Allo IM app content was personalized for each patient/caregiver based on their input. Wearable technologies were integrated to capture physical activity and sleep, using available interfaces that seamlessly integrate into the app. Results: Outcomes of PFA-AD analysis will include: 1) App usability; 2) Fidelity of personalized measures; 3) Correlation of personalized measures with standardized assessments; 4) Feasibility for inclusion in clinical trials. 4) Correlation of personalized real-life functions with clinical trial MRI-based brain analytics and 5) Feasibility of scale up for inclusion in the Allo phase 2 clinical trial. Conclusions: Enabling the assessment of patient’s real-life function in the real-world can advance the development of precision patient-centered drug development in Alzheimer’s disease.

P279 / #1185

Topic: Theme A: β-Amyloid Diseases / A4.l. Digital and AI-Based biomarkers

WEB-BASED AMYLOID RISK PREDICTION TO IDENTIFY PRECLINICAL ALZHEIMER’S DISEASE: APPLYING MACHINE-LEARNING TO JAPANESE TRIAL-READY COHORT WEBSTUDY POPULATION

Lecture Title:

K. Sato1, R. Ihara2, K. Suzuki3, Y. Niimi4, A. Iwata2, T. Iwatsubo5 1University of Tokyo Hospital, Neurology, Tokyo, Japan, 2Tokyo Metropolitan Geriatric Medical Center Hospital, Neurology, Tokyo, Japan, 3National Defense Medical College, Neurology, Saitama, Japan, 4The University of Tokyo Hospital, Unit For Early And Exploratory Clinical Development, Tokyo, Japan, 5Graduate School of Medicine, The University of Tokyo, Department Of Neuropathology, Tokyo, Japan

Aims: Selection of cognitively normal elderly individuals with higher risk of amyloid deposition in brain is critical to the success of prevention trials for Alzheimer’s disease (AD), whereas effective algorithms that can predict amyloid risks from basic clinical and cognitive data are yet to be established. Here we aimed to predict the standard uptake value ratio (SUVr) of amyloid PET in our ongoing Japanese Trial-Ready Cohort (J-TRC) webstudy, which is web-based feeder registry, using the clinical and cognitive variables available in the online settings. Methods: We built machine learning models consisting of age, sex, education years, family history of dementia, and online cognitive scores (Cognitive Function Instrument (CFI) and CogState), by using the screening data of cognitive normal participants (n = 4,277) from the A4 study as a reference to train. We then applied the models to the J-TRC webstudy participants registered within the initial 9 months of launch to obtain predicted SUVr. Results: Age, family history, CFI-study participant, and CogState score were the top important variables in the derived models. In a small subgroup of J-TRC webstudy participants who know their amyloid status (n = 37), the predicted SUVr corresponded well with the self-reported amyloid test results (AUC = 0.81). Conclusions: Our web-based prediction algorithms of brain amyloid status based on the A4 data may be usable for automatic prioritization of candidate participants with higher amyloid risks to be preferentially recruited from the J-TRC webstudy population to the in-person, J-TRC onsite study, and maximize the efficiency for the identification of preclinical AD participants.

P280 / #1266

Topic: Theme A: β-Amyloid Diseases / A5.a. Genetics, Epidemiology: Whole genome sequencing

USING LINKAGE ANALYSIS TO IDENTIFY NOVEL GENE-GENE INTERACTIONS IN ALZHEIMER’S DISEASE

Lecture Title:

M. Grunin1,2, N. Wheeler1, W. Bush1,2, J. Haines1,2, A.D.S.P. For The Alzheimer'S Disease Sequencing Project1 1Case Western Reserve University, Population And Quantitative Health Sciences, Cleveland, United States of America, 2Case Western Reserve University, Cleveland Institute For Computational Biology, Cleveland, United States of America

Aims: Linkage analysis was a workhorse of disease gene mapping until genome-wide association studies(GWAS). However, GWAS’ ability to detect gene-gene interactions needs huge sample sizes. Whole Genome Sequencing(WGS) captures nearly all allelic variation enabling direct measurement of disease co-segregation for each allele/family through linkage analysis without considering recombination fractions. Our approach integrates co-segregation data with biological interaction to identify interacting genes involved in disease using WGS of Alzheimer’s Disease(AD) families. Methods: Two point linkage(MERLIN) was performed on WGS of 46 AD Sequencing Project(ADSP) families. Per family, an empirical maximum LOD score(MaxLOD) was calculated. Interactions between proteins were identified by querying multiple functional/pathway databases. Variants reaching family-specific MaxLOD were filtered by impact(missense+stop/loss) and functional interaction. Extensive analysis of further ADSP families are being performed. Results: Genes of interest were defined as genes of known interaction where >2 biologically interacting loci/family reached family-specific MaxLOD and were found in >1 family, creating single shared transchromosomal locus. Transchromosomal MaxLODs were combined across multiple families sharing the same biologically interacting loci. Multiple families had biologically interacting genes with individual variants attaining MaxLOD>1.2. One interaction of shared gene pairs/family had transchromosomal MaxLOD across 3 families of 4.05, linking variants in SLC10A6, DHCR7, SOAT1, ABCA1 and SREBF2 in the cholesterol metabolism/lipid processing pathways. Additional 5 families had MaxLOD variants in at least one gene. 4 families had interacting pairs with transchromosomal MaxLOD score>2.4 unique to the family. Conclusions: This approach utilizes biological interactions to identify potential disease loci despite genomic distance, identifying possible epistatic effects not easily observable in case-control datasets.

P281 / #1087

Topic: Theme A: β-Amyloid Diseases / A5.a. Genetics, Epidemiology: Whole genome sequencing

THE ALZHEIMER’S DISEASE SEQUENCING PROJECT – FOLLOW UP STUDY (ADSP-FUS): INCREASING ANCESTRAL DIVERSITY IN ALZHEIMER’S GENETICS

Lecture Title:

B. Kunkle1, P. Mena1, K. Faber2, L. Adams1, T. Foroud2, D. Reyes-Dumeyer3, A. Kuzma4, A. Naj4, E. Martin1, C. Dalgard5, G. Schellenberg6, L.-S. Wang6, B. Vardarajan3, R. Mayeux7,8, J. Vance1, M. Cuccaro1, M. Pericak-Vance1 1University of Miami, John P. Hussman Institute For Human Genomics, Miami, United States of America, 2Indiana University, School Of Medicine, Indianapolis, United States of America, 3Columbia University, Taub Institute, New York, United States of America, 4University of Pennsylvania, School Of Medicine, Philadelphia, United States of America, 5Uniformed Services University of the Health Sciences, Uniformed Services University Of The Health Sciences, Bethesda, United States of America, 6University of Pennsylvania, Pathology And Laboratory Medicine, Penn Neurodegeneration Genomics Center, Philadelphia, United States of America, 7Columbia University, Neurology, New York, United States of America, 8Columbia University, Taub Institute For Research On Alzheimer's Disease And The Aging Brain, New York, United States of America

Aims: AD genetic studies lack racial-ethnic diversity. The ADSP-FUS is a National Institute on Aging (NIA) initiative focused on identifying genetic risk and protective variants for late-onset Alzheimer Disease (LOAD) through collection and sequencing of existing cohorts from diverse ancestries. Additional cohorts are presently being recruited. Methods: The cohorts consist of participants from studies of AD, dementia, and aging-related conditions. Existing biospecimens are processed primarily through the National Centralized Repository for Alzheimer’s Disease (NCRAD), and subsequently delivered to Uniformed Services University of the Health Sciences (USUHS) for whole genome sequencing. The resulting raw data is delivered to the Genome Center for Alzheimer’s Disease (GCAD) for processing followed by quality control analysis at University of Pennsylvania and University of Miami. Clinical, genotype and sequence data are stored at NIA Genetics of Alzheimer Disease Data Storage Site (NIAGADS), the ASDP-FUS data management and sharing center. Results: Over 30,000 samples have been ascertained: 7,896 with African ancestry; 9,475 with Hispanic ancestry; 13,531 with European ancestry, and 89 with Amerindian ancestry. Power studies show ~16,100 cases and ~16,100 controls are needed per ancestry for discovery of risk/protective variants with minor allele frequency (MAF) of 0.5% and odds ratios (OR) of 2.0 at genome-wide significance (P=5 x 10-8). Using gene-based testing ~10,000 cases and ~10,000 controls are needed for finding variants with MAF=0.005% and ORs=1.4. Conclusions: The ADSP-FUS is designed to enhance ongoing efforts for the identification of shared and novel genetic risk factors for LOAD among different populations, with the hope of developing effective treatments for all populations.

P282 / #599

Topic: Theme A: β-Amyloid Diseases / A5.c. Genetics, Epidemiology: GWAS, genetic associations, susceptibility & protective genes

KL-VS HETEROZYGOSITY ASSOCIATES WITH REDUCED BRAIN AMYLOID IN PRE-SYMPTOMATIC APOE4 CARRIERS.

Lecture Title:

M. Belloy1, S. Eger1, Y. Le Guen1, V. Napolioni1, K. Deters1, H.-S. Yang2, M. Scelsi3, T. Porter4, S.-N. James5, A. Wong5, J. Schott6, S. Laws4, E. Mormino1, Z. He1, S. Han1, A. Altmann3, M. Greicius1 1Stanford University, Neurology And Neurological Sciences, Stanford, United States of America, 2Brigham and Women's Hospital, Neurology, Boston, United States of America, 3University College London, Centre For Medical Image Computing (cmic), London, United Kingdom, 4Edith Cowan University, Collaborative Genomics And Translation Group, Joondalup, Australia, 5University College London, Medical Research Council Unit For Lifelong Health And Ageing, London, United Kingdom, 6University College London, Dementia Research Centre, London, United Kingdom

Aims: Perform a large-scale study to validate whether Klotho-VS heterozygosity (KL-VSHET+) is associated with reduced amyloid burden in cognitively normal older Apolipoprotein E4 (APOE4) carriers. Methods: We performed a large-scale meta-analysis across five independent studies (ADNI, A4, AIBL, Insight-46, HABS) comprising 3581 white non-Hispanic (Northwestern European ancestry) pre-clinical participants ages 60-80, to investigate whether KL-VSHET+ reduces the risk of having an amyloid-positive positron emission tomography scan (using study-specific thresholds). Analyses were stratified by APOE4 status and outcomes meta-analyzed (fixed effects inverse variance) across studies. The formal interaction between KL-VSHET+ and APOE4 was also evaluated. Outcomes are reported as odds ratios (OR) with 95% confidence intervals (CI). A priori power analyses were conducted for a range of expected effect sizes. Results: KL-VSHET+ reduced the risk of amyloid positivity in APOE4 carriers (OR=0.67 [0.52,0.88]; P=3.5x10-3), but not in non-carriers (OR=0.94 [0.73,1.21]; P=0.63) (Table1). The APOE4-by-KL-VSHET+ interaction analysis indicated that KL-VSHET+ displayed a stronger protective effect against amyloid positivity in APOE4 carriers compared to non-carriers, with borderline significance (OR=0.70 [0.48,1.02]; P=0.062). APOE4-stratified analyses were well-powered at observed effect sizes (Power>0.85).

Table1. Association between KL-VSHET+ and amyloid status by APOE4

status

Stratum Amy- (KL-

VSHET+/total) Amy+ (KL-

VSHET+/total) OR [95% CI] P-value

APOE4+ 182/621 (29.3%) 145/631 (23.0%) 0.67 [0.52-0.88]

0.0035

APOE4- 504/1926 (26.2%) 101/403 (25.1%) 0.94 [0.73-1.21]

0.63

Conclusions: The combination of APOE4 and KL-VS genotypes may help enrich AD clinical trials for pre-symptomatic subjects at increased risk of cognitive decline. Klotho-related pathways may help elucidate protective mechanisms against amyloid accumulation, meriting exploration for novel Alzheimer’s disease drug targets.

P283 / #1072


A GENOME-WIDE ASSOCIATION STUDY FOR CSF AΒ42 AND TAU WITH 6,000 INDIVIDUALS

Lecture Title:

F. Farias1, K. Black1, V. Fernandez2, J. Budde1, A. Fagan3,4, D. Holtzman5, J. Morris5, S. Kim6, A. Saykin7, P. De Jager8,9,10, M. Albert11, R. O'Brien12, M. Riemenschneider13, R. Petersen14,15, K. Blennow16,17,18, H. Zetterberg17,18,19, L. Minthon20, V. Van Deerlin21, V. Lee21, L. Shaw21, J. Trojanowski21, G. Schellenberg21, J. Haines22, R. Mayeux23,24, M. Pericak-Vance25, L. Farrer26,27,28, E. Peskind29, G. Li30,31, A. Ruiz32, L. Dumitresscu33, F.T.A.D.N.I. (Adni)34, On Behalf Of Adgc, Charge And Eadb Consortia35, C. Cruchaga2,36 1Washington University in St. Louis, Psychiatry, St. Louis, United States of America, 2Washington University School of Medicine, Psychiatry, St Louis, United States of America, 3Washington University in Saint Louis, Knight Alzheimer Disease Research Center, SAINT LOUIS, United States of America, 4Washington University in Saint Louis, Neurology, SAINT LOUIS, United States of America, 5Washington University School of Medicine, Hope Center For Neurological Disorders, Saint Louis, United States of America, 6State University of New York, Department Of Electrical And Computer Engineering, Oswego, United States of America, 7Indiana University School of Medicine, Department Of Radiology And Imaging Sciences, Indianapolis, United States of America, 8Columbia University, Taub Institute, New York, United States of America, 9Broad Institute of MIT and Harvard, Cell Circuits And Epigenomics, CAMBRIDGE, United States of America, 10Columbia University Medical Center, Department Of Neurology, New York, United States of America, 11Johns Hopkins University School of Medicine, Department Of Neurology, Baltimore, United States of America, 12Duke Medical Center, Department Of Neurology, Durham, United States of America, 13Saarland University, Clinic Of Psychiatry And Psychotherapy, Saar, Germany, 14Mayo Clinic, Neurology, Rochester, United States of America, 15Mayo Clinic, Department Of Neurology, Rochester, United States of America, 16Univ. of Gothenburg, Inst Of Neuroscience And Phsyiology, Mölndal, Sweden, 17Sahlgrenska University Hospital, Clinical Neurochemistry Laboratory, Mölndal, Sweden, 18Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden, 19UCL Institute of Neurology, Department Of Neurodegenerative Disease, London, United Kingdom, 20Lund University, Clinical Memory Research Unit, Department Of Clinical Sciences, Lund, Sweden, 21University of Pennsylvania, Department Of Pathology And Laboratory Medicine, Philadelphia, United States of America, 22Vanderbilt University, Department Of Molecular Physiology And Biophysics, Vanderbilt Center For Human Genetics Research, Nashville, United States of America, 23Columbia University, Neurology, New York, United States of America, 24Columbia University, Taub Institute For Research On Alzheimer's Disease And The Aging Brain, New York, United States of America, 25University of Miami, The John P. Hussman Institute For Human Genomics, And Dr. John T. Macdonald Foundation Department Of Human Genetics, Miami, United States of America, 26Boston University School of Medicine, Department Of Neurology And Alzheimer’s Disease Center, Boston, United States of America, 27Boston University School of Public Health, Department Of Medicine (biomedical Genetics), Boston, United States of America, 28Boston University School of Public Health, Department Of Biostatistics, Boston, United States of America, 29VA Puget Sound Health Care System, Visn-20 Mental Illness Research, Education, And Clinical Center, Seattle, United States of America, 30University of Washington, Psychiatry And Behavioral Sciences, Seattle, United States of America, 31VA Puget Sound, Grecc, Seattle, United States of America, 32Fundació ACE, Institut Català de Neurociències Aplicades - Universitat Internacional de Catalunya, Research Center And Memory Clinic, Barcelona, Spain, 33Vanderbilt University Medical Center, Vanderbilt Memory And Alzheimer’s Center, Nashville, United States of America, 34University of Cologne, Nuclear Medicine, Köln, Germany, 35University of Pennsylvania School of Medicine, Adgc, Philadelphia, United States of America, 36Washington University School of Medicine, Hope Center For Neurological Disorders, St. Louis, United States of America

Aims: Genome-wide association studies (GWAS) of endophenotypes have been successful in revealing new genes associated with disease risk. Cerebrospinal fluid (CSF) levels of amyloid beta (Aβ42) and tau are well known endophenotypes for Alzheimer’s disease (AD), they have been used in several GWAs studies. We describe here a large GWAS study with about 6,000 participants across twenty cohorts for CSF Aβ42, tau, and phosphorylated tau (ptau181) in order to confirm and identify new loci associated with AD. Methods: CSF Aβ42, tau, and ptau181 levels for each cohort was first log10-transformed to approximate a normal distribution and the mean from each data set was standardized to zero to account for variability in data collection. PLINK v1.9 was used for additive linear regression analysis and METAL was used for meta-analyze all cohorts. Results: CSF Aβ42 analysis showed previously identified APOE loci 19q13.32 (p= 6.68 x 10-155). Three suggestive loci for CSF Aβ42 were observed in chromosomes 3,4,and 6. CSF tau and ptau181 analysis confirmed previously identified APOE loci 19q13.32 (p= 1.02 x 10-42; p= 9.77 x 10-44, respectively) and the loci 3q28 (p= 3.28 x 10-22; p= 3.95 x 10-24, respectively) that was previously identified only with ptau181. Conclusions: Our results confirm previously reported APOE loci for CSF Aβ42, tau, and ptau181 levels. The previously identified loci in 3q28 that was only identified in CSF ptau181 was also identified with CSF tau levels. We are currently performing rare variant analysis and re-imputing the data with TopMed reference panel to repeat the analysis.

P284 / #228


FUNCTIONAL EXPLORATION OF AGFG2, A NOVEL PLAYER IN THE PATHOLOGY OF ALZHEIMER DISEASE

Lecture Title:

V. Fernandez1, J. Budde1, E. Dhungel1, A. Eteleeb1, F. Wang2, R. Martinez3, J. Marsh4, J. Norton2, J. Gentsch1, J. Morris5, R. Bateman6, L. Swisher6, E. Mcdade6, R. Perrin6, O. Harari4, B. Benitez4, C. Karch1, C. Cruchaga1 1Washington University School of Medicine, Psychiatry, St Louis, United States of America, 2Washington University in St. Louis, Psychiatry, St. Louis, United States of America, 3Washington University School if Medicine, Psychiatry, St. Louis, United States of America, 4Washington University School of Medicine, Psychiatry, St. Louis, United States of America, 5Washington University School of Medicine, Neurology, St Louis, United States of America, 6Washington University School of Medicine, Neurology, St. Louis, United States of America

Aims: There are three main clinical presentations in Alzheimer disease (AD) with diversity in phenotype, onset and progression of clinical symptoms: autosomal dominant (ADAD), early onset (EOAD) and late onset (LOAD). Ultimately, AD is characterized by the deposition of Aβ and ptau protein aggregates in the brain. This work aims to identify and characterize common disrupted pathways across AD etiologies. Methods: We examined bulk transcriptomic data from brain donors to the DIAN (ADAD, N=19) and Knight-ADRC (EOAD, N=13; LOAD, N=55; controls, N=16). We performed differential gene expression (DGE) analyses using DSeq2 and cross-checked significant signals with known GWAs loci. We replicated our findings in three AMP-AD independent studies. Results: DGE analysis identified 57 significantly differentiated genes across ADAD, EOAD or LOAD vs controls. Among those, AGFG2 (p=7×10-4) had a higher expression in cases and falls under the AD GWAs signal for NYAP1 (rs1476679). This effect replicated in all independent datasets: Mount Sinai (p=8.63×10-3), Mayo (p=5.88×10-12), ROSMAP (p=3.96×10-05). AGFG2 is expressed primarily in astrocytes and is a member of the HIV-1 Rev binding protein family that mediates the nucleocytoplasmic transfer of proteins and RNAs. AGFG2 has been implicated in APP metabolism, leading to the hypothesis that higher expression of AGFG2 could promote more release of APP to the media. Conclusions: Our results suggest that AGFG2 may be a novel player in the etiology of AD. We are currently modifying AGFG2 expression on iPSC-derived astrocytes using CRIPR-Cas9 technology to evaluate AGFG2 role in APP and Tau metabolism. We will present these results at the conference.

P285 / #1457


GENOTYPE-PHENOTYPE DATA OF RARE ABCA7 MISSENSE MUTATIONS IN FLANDERS BELGIAN ALZHEIMER’S DISEASE PATIENTS

Lecture Title:

E. Hens1,2,3,4,5,6, L. Bossaerts2,3,6, A. Sieben3,6,7, S. Engelborghs3,5,6, Y. Vermeiren3,6, B. Hanseeuw8, R. Vandenberghe9, P. De Deyn1,3,6, P. Cras3,4,6, J.-J. Martin3,6, C. Van Broeckhoven2,3,6 1Hospital Network Antwerp, Department Of Neurology And Memory Clinics, Antwerp, Belgium, 2VIB Center for Molecular Neurology, Neurodegenerative Brain Diseases Group, Antwerp, Belgium, 3University of Antwerp, Department Of Biomedical Sciences, Antwerp, Belgium, 4University Hospital Antwerp, Department Of Neurology, Edegem, Belgium, 5University Hospital Brussels and University Center for Neurosciences VUB, Department Of Neurology, Brussels, Belgium, 6Antwerp, Institute Born-bunge, Antwerp, Belgium, 7Gent University Hospital, Neurology Department, Gent, Belgium, 8University Hospital Saint-Luc and University Institute of Neuroscience UC Louvain, Department Of Neurology, Louvain-la-Neuve, Belgium, 9University Hospitals Leuven and University Department of Neurosciences KU Leuven, Catholic University of Leuven, Department Of Neurology, Leuven, Belgium

Aims: ABCA7 was associated with risk of Alzheimer’s disease (AD) in GWAS. In AD patients, rare premature termination codon (PTC) variants and missense variants are enriched. In a cohort of Flanders-Belgian AD patients, we identified 99 missense mutation carriers and 69 PTC mutation carriers. We aimed to delineate the clinicopathological characteristics of the missense carriers and to compare genotype-phenotype data with AD patients carrying PTC variants. Methods: Available demographic, clinicopathological data of both missense and PTC mutation carriers were reviewed. Results: ABCA7 missense carriers were most frequently diagnosed with probable AD (77%). However, in 16% important vascular involvement was observed. Mean onset age was 68.1±9.9 years (37-92), with mean disease duration of 9.6±4.4 years. Positive first-degree familial history was present in 62.5%, and we observed apparent autosomal dominant co-segregation of AD in a family (p.G1820S). Clinical neurological examination and neuroimaging showed typical signs of AD. Cerebrospinal fluid (CSF) AD biomarkers displayed a typical AD profile in 80%. Brain autopsy (n=7) revealed classical AD changes plus cerebral amyloid angiopathy (CAA) in all carriers. High levels of CAA were observed in the meningeal arteries and capillaries. Compared to missense carriers, total tau levels were significantly higher in PTC carriers. Further, no significant differences in CSF AD-biomarker levels or CAA were observed in PTC carriers and missense carriers. Conclusions: ABCA7 missense mutation carriers presented with a classical AD phenotype and neuropathological AD hallmarks with moderate-to-high levels of CAA. Besides higher total tau levels in PTC carriers, we did not observe clinicopathological differences between ABCA7 PTC and missense carriers.

P286 / #1480


HERITABILITY ANALYSES SHOW ONLY PARTIAL GENETIC OVERLAP BETWEEN EARLY AND LATE ONSET ALZHEIMER DISEASE.

Lecture Title:

E. Lucio Da Fonseca1, M. Jean Francois1, J. Thulaseedhara Kurup2, S. Hackleman Slifer1, E. Martin1, B. Kunkle1, M. Pericak-Vance1, G. Schellenberg3, V. Fernandez4, C. Cruchaga4, C. Reitz2, G. Beecham1 1University of Miami, Miller School of Medicine, John P. Hussman Institute For Human Genomics, Miami, United States of America, 2Columbia University, Depts. Of Neurology And Epidemiology, New York, United States of America, 3University of Pennsylvania, Path & Lab Med, Stellar Chance, School Of Medicine, Philadelphia, United States of America, 4Washington University, School Of Medicine, St. Louis, United States of America

Aims: Alzheimer disease (AD) is the most common cause of progressive dementia. It is highly heritable and has a wide range of clinical symptoms and onset. It is often described as either early onset (EOAD, age at onset, [AAO] <= 65) or late onset (LOAD, [AAO]>65). Few genetic studies have focused on EOAD and it is unclear how much genetic etiology is shared by the two forms. To understand this shared etiology, a genome-wide association study (GWAS) and heritability analyses of non-Mendelian EOAD and LOAD were performed. Methods: Genetic data on 21,622 individuals from the Alzheimer Disease Genetics Consortium (ADGC) were used: (1,476 EOAD, 9,695 LOAD and 10,451 control). Association analyses were performed using logistic regression under two models: ancestry plus SNP, and ancestry, sex, APOE dosage, and SNP. EOAD and LOAD were considered separately. LD score regression was used to estimate the SNP heritability (h2) and genetic correlation (rg). Results: In EOAD models 7 SNPs across 3 novel regions were associated (P<5x10-8). These include chr5, 104.8Mb (intergenic); chr6, 90.6Mb (intronic to BACH2); and chr14, 61.9Mb. Heritability analyses showed higher h2 values for EOAD (h2=0.24, 0.23 for models 1 and 2) than LOAD (h2=0.18 ,0.14). Genetic correlation showed moderate genetic overlap between EOAD and LOAD (rg=0.3515, p=0.0283). Conclusions: Together GWAS and heritability analysis suggest that the genetic etiology of EOAD has some — but not complete — overlap with LOAD. The results also suggest a stronger polygenic effect in EOAD than LOAD, confirming the need for additional genomics efforts in non-Mendelian EOAD.

P287 / #1474


GENETICS OF COGNITIVE RESILIENCY IN THE MIDWESTERN AMISH

Lecture Title:

L. Main1, Y. Song1, R. Laux1, K. Miskimen1, M. Cuccaro2, P. Ogrocki3, J. Haines1, A. Lerner1,3, J. Vance2, M. Fuzzell1, S. Fuzzell1, J. Sewell1, L. Caywood2, M. Prough4, W. Scott4, L. Adams2, M. Pericak-Vance5, J. Clouse4, S. Herington2 1Case Western Reserve University, Population And Quantitative Health Sciences, Cleveland, United States of America, 2University of Miami Miller School of Medicine, John P Hussman Institute For Human Genomics, Miami, United States of America, 3University Hospitals, Cleveland Medical Center, Cleveland, United States of America, 4University of Miami, John P. Hussman Institute For Human Genomics, Miami, United States of America, 5University of Miami, The John P. Hussman Institute For Human Genomics, And Dr. John T. Macdonald Foundation Department Of Human Genetics, Miami, United States of America

Aims: Alzheimer’s disease (AD) is a progressive disease characterized by a buildup of amyloid beta plaques and tau tangles in the brain, leading to extreme atrophy and cognitive decline. Our study focuses on the Midwestern Amish, leveraging their homogeneous genetics and environment. Our goal is to identify genetic variants that delay the onset or protect against the development of AD. Methods: Studying genetics in this isolated population increases our ability to find rare protective variants for AD compared to the general population. We have been ascertaining individuals who are at high risk for developing AD (i.e. have an affected sibling and are of 76 years of age or older), but are still cognitively normal (CN). Each of these individuals is retested every two years. Results: As of August 2020, we have a total of 2,217 individuals genotyped in the Midwestern Amish. The frequency of the APOE-e2 allele in this dataset is 5%, while the APOE-e4 allele frequency is 14%, slightly lower than the general European population. We are currently using KING and GENESIS software to perform analyses since these programs take into account relatedness in a population structure. Association and linkage analyses, taking into account the complex pedigree relationships, is ongoing. As of the most recent analysis, 5 SNPs have p ≤ 0.05. Conclusions: We are using a founder population to have an increased chance of finding protective genetic variants in AD. The closest genes to the SNPs that passed the significance threshold from the association analysis are: YIPF4, SHROOM3, NUMB, ACSBG1, and LINC00654.

P288 / #876


STUDY OF THE LATE-ONSET ALZHEIMER’S DISEASE RISK FACTOR BIN1 IN HIPSC-DERIVED NEURONS

Lecture Title:

A.R. Melo De Farias, O. Saha, J.-C. Lambert, M. Costa Institut Pasteur de Lille, Unit 1167, Lille, France

Aims: The BIN1 (Bridging Integrator-1) gene has been identified to be second on the list of risk genes with a major susceptibility loci linked to late-onset Alzheimer’s disease (LOAD). In this study, we aimed at studying the possible effects of BIN1 deletion in endocytosis using human induced pluripotent stem cell (hiPSC)-derived neurons. Methods: We are utilizing hiPSCs expressing allelic variants of BIN (BIN1+/+ and BIN1-/-) to this end. Human induced neural progenitor cells (hiNPCs) are generated from these hiPSCs, which are further differentiated into mature neurons and glia. We evaluated the expression of different BIN1 isoforms, cell type composition of cultures and endocytic pathway using western blot and immunocytochemistry. Results: Our panel of preliminary data suggests that BIN1 knockout impairs the proliferation and differentiation potential of hiNPCs. Also, analyses of early endosome vesicles in 6-week neuronal cultures using antibodies against the early endosome antigen 1 (EEA1) and RAB5A-GFP fused proteins reveal a significant decrease in the mean volume of endosomes in BIN1-/- compared to BIN1+/+ hiNs. Conclusions: Taken together, our preliminary data seems to suggest a role of BIN1 in the proliferation and differentiation of hiNPCs and also in the endocytic pathway in hiNs.

P289 / #1747


COMBINING THE TOHOKU MEDICAL MEGABANK ORGANIZATION COHORT STUDY’S OPTICAL COHERENCE TOMOGRAPHY AND GENETIC DATA FOR ALZHEIMER’S DISEASE PREDICTION RESEARCH

Lecture Title:

M. Taira1, K. Kojima1, S. Tadaka1, R. Kawasaki2, A. Miki2, A. Saykin3,4, K. Kinoshita1, N. Fuse1, M. Yamamoto1 1Tohoku Medical Megabank Organization, Integrative Genomics, Sendai, Japan, 2Osaka University, Graduate School Of Medicine/faculty Of Medicine, Osaka, Japan, 3Indiana University School of Medicine, Radiology And Imaging Sciences, Indianapolis, United States of America, 4Indiana University School of Medicine, Department Of Radiology And Imaging Sciences, Indianapolis, United States of America

Aims: The Tohoku Medical Megabank Organization (ToMMo) is a prospective cohort-scale biobank established in Northern Japan’s Miyagi Prefecture following the Great East Japan Earthquake and Tsunami in 2011. Under the national mission to promote residents’ health, ToMMo enrolled approximately 120,000 adults. One of our aims is to accelerate the development of specific biomarkers for the early detection of common diseases, including Alzheimer’s disease (AD). This communication reports initial progress. Methods: We propose a new framework based on a modified convolution neural network for the correlations between SD-OCT image and previously reported AD genes. Spectral Domain Optical Coherence Tomography (SD-OCT): With encompassed image data preprocessing, training-validation-testing dataset, and evaluation metrics, we capture retinal nerve fiber layer thickness changes and fundus imaging features. The raw imaging data were digitized and archived. Genetics: whole genome sequencing and/or SNP array data were obtained from participants’ blood samples. Results: Usable SD-OCT data exists for 24,664 participants aged over 20 and 13,696 participants aged over 50. The age distribution of participants who had OCT at Baseline is shown in the Table.

Conclusions: ToMMo’s SD-OCT data is a powerful resource for AD research. Specifically, the combination of SD-OCT and genetic data will support unique disease progression analyses and investigations to elucidate AD pathophysiology. Additional information is being gained by correlating SD-OCT image data and APOE haplotypes in combination with participants’ demographic and relevant health data. Future work will entail identifying biomarkers suggested by analyses of Polygenic Risk Score and Mendelian Randomization.

P290 / #736

Topic: Theme A: β-Amyloid Diseases / A5.e. Genetics, Epidemiology: Aging

ESTROGEN MODULATING BREAST CANCER THERAPIES AND THE INCIDENCE OF ALZHEIMER’S DISEASE

Lecture Title:

G. Branigan, M. Soto, L. Neumayer, K. Rodgers, R. Brinton University of Arizona, Center For Innovation In Brain Science, Tucson, United States of America

Aims: To determine whether estrogen modulating therapy (EMT) exposure is associated with risk of Alzheimer’s (AD) in women with breast cancer in the Humana claims data set. The Humana data set contains claims from private-payer and Medicare insurance data sets from across the United States with a population primarily residing in the Southeast. Methods: Patients receiving EMT for breast cancer treatment were identified and survival analysis, stratified by age and by individual therapeutics, was used to determine the association between EMT exposure and diagnosis of AD. A propensity score approach was used to minimize measured and unmeasured selection bias. Results: In this cohort study of propensity score matched perimenopausal to postmenopausal aged women with breast cancer, EMT exposure was associated with decrease in diagnosis of neurodegenerative disease, most specifically AD (relative risk, 0.82; 95% CI, 0.75-0.90; P < 0.001). Conclusions: Among female patients with breast cancer, exposure to estrogen modulating therapies was associated with a decrease in diagnosis of AD and Dementia. Risk reduction varied by individual therapeutic exposure and by age of treatment where greater age at exposure was associated with greatest risk reduction. As a result, translational studies investigating the mechanisms by which these therapeutics impact AD risk are currently underway.

P291 / #507


FRAILTY AS A RISK FACTOR FOR INCIDENT DEMENTIA AND MEMORY DECLINE: DATA FROM THE “INVECE.AB” LONGITUDINAL STUDY.

Lecture Title:

A. Davin1, M.C. Mimmi2, D. Ardemagni1, R. Vaccaro1, M. Colombo1, T.E. Poloni3,4, V. Medici3, C. Cereda2, A. Guaita1 1Golgi-Cenci Foundation, Laboratory Of Neurobiology And Neurogenetic, Abbiategrasso, Italy, 2IRCCS Mondino Foundation, Genomic And Post-genomic Center, Pavia, Italy, 3Golgi-Cenci Foundation, Department Of Neurology And Neuropathology, Abbiategrasso, Italy, 4ASP Golgi-Redaelli Geriatric Hospital, Department Of Rehabilitation, Abbiategrasso, Italy

Aims: Evaluation of the influence of Frailty syndrome on incident dementia and memory functions decline in the Italian longitudinal cohort study InveCe.Ab (Invecchiamento Cerebrale in Abbiategrasso, clinical trials.gov NCT01345110), during a 4 year of follow up. Methods: All InveCe.Ab study participants (baseline and 4-years follow up wave) free from dementia, and psychiatric/neurological disorders were included (N=1171). Frailty was measured by Frailty Index (FI= 0,0 – 1,0), and participants grouped into Frail (FI>0,25), pre-Frail (FI = 0,08-0,25) and non-Frail (FI<0,08). Dementia was defined using the DSM-IV-TR. Global cognition, memory, executive and visuo-spatial cognitive domains were evaluated by a comprehensive neuropsychological test battery. Results: After 4-years follow-up, 4,2% developed Dementia and baseline Frailty was associated with cumulative incidence of dementia (non-Frail 2,3%; pre-Frail 4,3%; Frail 16,9%; p<0,001). The logistic regression model adjusted for age, gender, level of education, loneliness and APOEɛ4 carrier status shows that frailty categories is significantly associated with incident dementia (OR=8,8, p<0,001), with higher risk of dementia for frail men versus frail women (OR=14,11, p<0,001; OR= 5,80, p=0,017, respectively). The three-way repeated-measures ANOVA analysis showed a statistically significant interaction effect between frailty and gender during 4-years follow-up, F(2,841)=3,680, p=0,026 and a substantial effect of time on memory performance, F(1,841)=14.39, p<0,001. Conclusions: Frailty is associated with cognitive decline over a 4-year period, especially for the male gender, and could be considered as a red flag for early detection of memory decline.

P292 / #1226


THE EXTERNAL VALIDITY OF CONTEMPORARY DEMENTIA RESEARCH ACROSS THE GLOBE: A SYSTEMATIC REVIEW

Lecture Title:

S. Mooldijk, S. Licher, F. Wolters Erasmus MC University Medical Center, Department Of Epidemiology, Rotterdam, Netherlands

Aims: Failure to translate dementia research into efficacious therapies is often attributed to targeting the wrong population in clinical trials, but could also be explained by limited external validity of informative studies. We therefore aimed to describe participants with or at risk for dementia included in contemporary dementia research. Methods: We systematically reviewed PubMed/Medline for studies of persons with dementia published between September 2018 and August 2019 in the top 100 journals in the fields of neurology/neuroscience, geriatrics, psychiatry or general medicine. Study details were extracted by 2 reviewers. Results: We identified 513 studies of individuals with dementia. Of these, 211 (41%) studied <50 individuals with dementia. The remaining 302 studies included a median of 214 participants with dementia. Only 22% of all studies reported participants’ ethnicity. Among those, virtually all participants were of Caucasian descent [median 90%, IQR 79-97] mainly residing in North-American and Western-European populations (Figure). The mean age at diagnosis was 71.8 years (SD 6.4) in clinical studies, compared to 80.6 years (SD 4.7) in population-based studies (p-value for difference <0.001). Use of MRI, PET and CSF, but not blood biomarkers, was mostly limited to clinical studies (87%), and consequently applied to young patients (mean age at diagnosis: 71.6).

Conclusions: Dementia research is limited in terms of ethnic and geographic diversity. Clinic-based studies cover an approximately 10-year younger group of patients. This hampers the transportability of research findings, including the generalizability of treatment interventions and diagnostic criteria for

patients with dementia in the global community.

P293 / #1753


IMMUNE RESPONSE AND ENDOCYTOSIS PATHWAYS ARE ASSOCIATED WITH THE RESILIENCE AGAINST ALZHEIMER’S DISEASE

Lecture Title:

N. Tesi1, S. Van Der Lee1, M. Hulsman1, I. Jansen1, N. Stringa2, N. Van Schoor2, P. Scheltens3, W. Van Der Flier4, M. Huisman2, M. Reinders5, H. Holstege6 1Amsterdam UMC, Department Of Neurology, AMmsterdam, Netherlands, 2Amsterdam UMC, Department Of Epidemiology And Biostatistics, Amsterdam, Netherlands, 3Amsterdam University Medical Center, Alzheimer Center, AMSTERDAM, Netherlands, 4Amsterdam UMC, Alzheimer Center Amsterdam, Department Of Neurology, Amsterdam Neuroscience, Vrije Universiteit Amsterdam, Amsterdam, Netherlands, 5TU Delft, Delft Bioinformatics Lab, Delft, Netherlands, 6Amsterdam University medical Center, Alzheimer Center, AMSTERDAM, Netherlands

Aims: Developing Alzheimer’s disease (AD) is influenced by genetic variants that are involved in five major AD-pathways. These pathways may differentially contribute to the modification of the AD-risk. The pathways involved in the resilience-against-AD have thus far been poorly addressed. We investigated how each molecular mechanism associates with the increased AD-risk and the resilience-against-AD until extreme old age, by comparing pathway-specific polygenic-risk-scores (pathway-PRS). Methods: Using 29 AD-associated single-nucleotide-variants, we developed pathway-PRS for five major pathways involved in AD. We developed an integrative framework that allows multiple genes to associate with a variant, and multiple pathways to associate with a gene. We studied pathway-PRS in AD patients (N=1,895), healthy population controls (N=1,654) and our unique cohort of cognitively healthy centenarians who avoided AD (N=293). Last, we estimated the contribution of each pathway to the genetic risk of AD in the general population. Results: All pathway-PRS significantly associated with AD-risk and with resilience-against-AD (except for angiogenesis). The pathway that contributed most to the modulation of AD-risk was β-amyloid metabolism (29.6%), driven mainly by APOE-variants. After excluding APOE-variants, all pathway-PRS associated with increased AD-risk (except for angiogenesis), while specifically immune-response and endocytosis associated with resilience-against-AD. The variants in these two pathways became the main contributors to the overall genetic risk of AD (45.5% and 19.2%, respectively). Conclusions: The single-genetic-variants associated with the resilience-against-AD indicate which pathways are involved with maintaining cognitive functioning until extreme ages. A favorable immune-response and endocytosis pathways might be involved in neuro-protection, highlighting the need to investigate these pathways, next to β-amyloid metabolism.

P294 / #1006


STATIN THERAPY AND RISK OF ALZHEIMER’S, PARKINSON’S, AND OTHER AGE-RELATED NEURODEGENERATIVE DISEASES

Lecture Title:

G. Torrandell Haro1, G. Branigan1,2, F. Vitali1, R. Brinton1 1University of Arizona, Center For Innovation In Brain Science, Tucson, United States of America, 2University of Arizona, Pharmacology, Tucson, United States of America

Aims: To establish the efficacy of and molecular pathways for statins to assess the impact on incidence of Alzheimer’s, Parkinson’s, and other neurodegenerative diseases (NDD). Methods: This retrospective cohort study surveyed U.S.-based Humana claims, which includes prescription and patient records from private-payer and Medicare insurance. Claims from 288,602 patients, aged 45 years and older without prior history of NDD or neurological surgery, were surveyed for a diagnosis of NDD starting 1 year following statin exposure. Computational system biology analysis was conducted to determine unique target engagement for each statin. Results: Of the 288,602 participants included in the study, 144,214 patients (mean [SD] age, 67.22 [3.8] years) exposed to statin therapies, and 144,301 patients (65.97 [3.2] years) were not treated with statins. The mean (SD) follow-up time was 5.1 (2.3) years. Exposure to statins was associated with a lower incidence of Alzheimer’s (1.10% vs 2.37%; relative risk, 0.4643; 95%CI, 0.44-0.49; P < .001), dementia 3.03% vs 5.39%; RR, 0.56; 95%CI, 0.54-0.58; P < .001), Multiple-Sclerosis (0.08% vs 0.15 %; RR, 0.52; 95%CI, 0.41-0.66; P < .001), Parkinson’s (0.48% vs 0.92%; RR, 0.53; 95%CI, 0.48-0.58; P < .001), ALS (0.02% vs 0.05 %; RR, 0.46; 95%CI, 0.30-0.69; P < .001). All NDD incidence for all statins, except for fluvastatin (RR, 0.91; 95% CI, 0.65-1.30; P = 0.71), was reduced with variances in individual risk profiles. Pathway analysis indicated unique and common profiles associated with risk reduction efficacy. Conclusions: Statins were associated with lower risk of NDD which paralleled activation of neurological targets.

P295 / #1565

Topic: Theme A: β-Amyloid Diseases / A5.f. Genetics, Epidemiology: Environmental risk factors

DIFFERENTIAL RISK OF INCIDENT FRACTURES DEPENDING ON INTENSITY AND FREQUENCY OF PHYSICAL ACTIVITY ACCORDING TO COGNITIVE STATUS: A NATIONWIDE LONGITUDINAL STUDY

Lecture Title:


Aims: There has been growing recognition of the vital effect of physical activity (PA) on delay and prevention of fractures in the trajectory of dementia. This study aimed to evaluate the optimal intensity and frequency of PA needed to prevent fractures in cognitively preserved older adults (CP), participants with subjective cognitive decline (SCD), and dementia patients using a large-scale nationwide cohort study Methods: Data from a nationwide health screening program for individuals at the transitional age of 66 years were used in this study. A total of 968,240 subjects was enrolled, followed from 2007 to 2014, and classified as CP (n=759874), SCD (n=195365), or dementia group (n=13001). Adjusted hazard ratios (aHRs) by demographic and known risk factors for fractures were evaluated to identify the impact of various frequency and intensity PA on the occurrence of hip, vertebral, and limb fractures. Results: In CP participants, the most noticeable reduction of hip and vertebral fracture risk was shown in those performing vigorous-intensity PA at least three times per week. In the SCD group, the risk decrement in hip and vertebral fractures was most prominent in subjects who performed multiple-intensity PAs at least three times a week regardless of intensity. In the dementia group, only high-frequency walking and high-frequency & multiple-intensity PA decreased the risk of hip fractures compared with absence of PA. Conclusions: These findings suggest a role for various PA intensity and frequency levels to prevent hip and vertebral fractures according to cognitive status.

P297 / #421

Topic: Theme A: β-Amyloid Diseases / A5.i. Genetics, Epidemiology: Other

A METHOD OF COMBINING TWO SINGLE COHORTS FOR DEVELOPING DISEASE PROGRESSION MODEL

Lecture Title:

S. Kim1, S.-Y. Woo1, Y. Lim2, J. Lee2, S.W. Seo3 1Samsung Medical Center, Statistics And Data Center, Seoul, Korea, Republic of, 2Chung-Ang University, Department Of Statistics, Seoul, Korea, Republic of, 3Samsung Medical Center, Department Of Neurology, Seoul, Korea, Republic of

Aims: To develop a disease progression model by combining data from cohorts with different disease states but the starting time point of all cohorts is zero regardless of disease state, we present the method to estimate the starting time point of the more advanced cohort that follows the less advanced cohort. Methods: Our approach is to estimate the mixed effect model for each cohort, and to search the time S to start to overlap between the two consecutive cohorts using the predicted outcome and its 95% confidence interval for each subject at each follow-up time based on the estimated model. The method was validated using the simulated data with a size of 100 per cohort under various scenarios, fixed or random effects of intercept and slope, correlation structure among follow-up times, missing pattern, independence between random effects, variability of random effect, and residual of the mixed effect model. Bias and precision of the estimates of S (21 months, 42 months) were evaluated for each scenario. Results: Our method provided good performance. For all scenarios, biases were within 0.5 months and not affected by scenario factors. Precisions were only affected by the variability of the intercept, and standard deviations of the estimates of S increased from 0.5 month to 1.2 months when variability of intercept increased. Conclusions: With this approach, a disease progression course can be modeled using the separate cohorts with different consecutive disease states without following up whole period of the disease. This approach can also be applied to nonlinear progression model.

P298 / #249

Topic: Theme A: β-Amyloid Diseases / A5.i. Genetics, Epidemiology: Other

ASSOCIATION OF BLOOD-BASED TRANSCRIPTIONAL RISK SCORES WITH BIOMARKERS FOR ALZHEIMER'S DISEASE

Lecture Title:

Y.H. Park1, A. Hodges2, M. Weiner3, S. Kim1, A. Saykin4, K. Nho4 1Seoul National University Bundang Hospital, Department Of Neurology, Seongnam, Korea, Republic of, 2King’s College London, Institute Of Psychiatry, Psychology & Neuroscience, London, United Kingdom, 3University of California, San Francisco, Department Of Veterans Affairs Medical Center, San Francisco, United States of America, 4Indiana University School of Medicine, Department Of Radiology And Imaging Sciences, Indianapolis, United States of America

Aims: To determine whether transcriptional risk scores (TRS), a summation of polarized expression levels of functional genes, reflect the risk of Alzheimer's disease (AD). Methods: Blood transcriptome data was from Caucasian participants, which included AD, mild cognitive impairment and cognitively normal controls (CN) in the Alzheimer’s Disease Neuroimaging Initiative (ADNI, n=661) and AddNeuroMed (n=674) cohorts. To calculate TRS, we selected functional genes that were expressed under the control of the AD risk loci and were identified as being responsible for AD by using Bayesian colocalization and Mendelian randomization methods. Regression was used to investigate the association of TRS with diagnosis (AD vs. CN) and MRI biomarkers (entorhinal thickness and hippocampal volume). Regression was also used to evaluate whether expression of each functional gene was associated with AD diagnosis. Results: TRS was significantly associated with AD diagnosis, hippocampal volume, and entorhinal cortical thickness in ADNI. The association of TRS with AD diagnosis and entorhinal cortical thickness was also replicated in AddNeuroMed. Among functional genes identified to calculate TRS, CD33 and PILRA were significantly up-regulated, and TRAPPC6A was significantly down-regulated in AD patients compared to CN, all of which were identified in ADNI and replicated in AddNeuroMed. Conclusions: Blood-based TRS is significantly associated with AD diagnosis and neuroimaging biomarkers. In blood, CD33 and PILRA were known to be associated with uptake of b-amyloid and herpes simplex virus-1 infection, respectively, both of which may play a role in the pathogenesis of AD.

P299 / #318

Topic: Theme A: β-Amyloid Diseases / A6.a. Cell, Molecular and Systems Biology: APP, APLP, Abeta

ABETA42 AND TAU BOTH DISRUPT NUCLEOCYTOPLASMIC TRANSPORT

Lecture Title:

S. Akerman1, G. Multhaup2 1McGill University, Pharmacology And Therapeutics, Montreal, Canada, 2McGill University, Pharmacology & Therapeutics, Montreal, Canada

Aims: There is growing evidence that nucleocytoplasmic transport (NCT) defects and nucleoskeleton deformities are linked to Alzheimer Disease (AD). Ras-related nuclear protein (Ran) is a highly conserved protein, plays a crucial role in NCT and can be an indicator of impaired nucleocytoplasmic transport. Previously, Ran levels were found to be decreased in AD; however, the exact mechanism remains unknown. Here we established a molecular link between Ran function, NCT defects and nucleoskeleton deformities. Methods: We treated SH-SY5Y neuroblastoma cells with Abeta species 40, 42 and 42G33A and analyzed SH-SY5Y cells stably expressing tau P301L for effects on NCT proteins, i.e. Ran, Ran-GAP, RCC1 and importins. Effects due to increased phosphorylation of tau were tested under hypothermic conditions. Results: We found that Abeta42 specifically caused a reduction in Ran levels and led to a mislocalization of an engineered soluble nuclear marker. Tau overexpression resulted in Ran delocalization from the nucleus to the cytoplasm but did not alter Ran levels. Tau specifically caused nucleocytoskeleton deformities which were enhanced under hypothermic conditions. Conclusions: Both, Abeta42 and tau cause defects on NCT. Abeta42 leads to reduced levels of Ran while tau causes a mislocalization of Ran from the nucleus to the cytoplasm. When the NCT is impaired the signaling pathways in neurons could be affected, potentially resulting in inefficient transcription as well as aberrant DNA/RNA maintenance which in turn may lead to neuronal death.

P300 / #837


APP GLYCOSILATION DETERMINES ITS PROTEOLYTIC PROCESSING, AND THIS IS ALTERED IN ALZHEIMER´S DISEASE

Lecture Title:

I. Cuchillo-Ibáñez, C. Boix, I. López-Font, J. Sáez-Valero Instituto de Neurociencias, UMH-CSCI, Molecular Neurobiology And Neuropathology, Sant Joan d'Alacant, Spain

Aims: The amyloid precursor protein (APP) is a transmembrane glycoprotein that undergoes post-translational glycosylation and alternative proteolytic processing. The amyloidogenic processing generates a soluble ectodomain fragment (sAPPβ) and the β-amyloid peptide (Aβ), while the non-amyloidogenic processing generates sAPPα. We aim to examine the levels and glycosylation patterns of sAPPα and sAPPβ in Alzheimer's disease (AD) brain, discriminating between those generated from neuronal-APP695 or glial/KPI isoforms. Methods: In frontal cortex samples from AD patients (n = 7) and non-demented controls (NDC; n = 7) the expression of total APP, APP695 and APP-KPI were analyzed by qRT-PCR. Levels of sAPPα, sAPPβ and citosolic fragments (CTFα and CTFβ) were estimated by western blots. Glycosylation patterns were determined by lectins. CHO-PS70 cells stably over-expressing wild-type human APP were treated with 5mM Aβ42. Results: We detected a higher expression of APP695 and APP-KPI variants in AD cortex relative to NDC. However, sAPPα and sAPPβ protein levels remained unchanged, as did those of CTFα and CTFβ. When glycosilation was examined, sAPPα from APP695 and APP-KPI showed a common glycosylation pattern, but this was different in AD cortex respect to NDC. sAPPβ glycosylation from APP695 was different respect to those from APP-KPI isoforms, but remained the same in AD and NDC brains. Aβ peptide altered glycosylation pattern of sAPPα and sAPPβ in cultured cells. Conclusions: Specific glycosylation may dictate the APP processing and therefore, the generation of different sAPP in neurons and glial cells. sAPPα glycosylation pattern is affected in AD, but not sAPPβ.

P301 / #1638


EXPRESSION OF HUMANIZED AMYLOID-BETA IN A KNOCK-IN STRAIN DRIVES ACCUMULATION OF PERIODIC ACID-SCHIFF GRANULES IN THE HIPPOCAMPUS OF MICE

Lecture Title:

D. Javonillo1, K. Tran1, D. Baglietto-Vargas1,2, J. Phan1, C. Da Cunha1, S. Forner1, S. Kawauchi3, A. Tenner1,2,4, F. Laferla1,2, G. Macgregor3,5, K. Green1,2, F.T.M.-A.C. (Model-Ad)1 1University of California, Irvine, Institute For Memory Impairments And Neurological Disorders, Irvine, United States of America, 2University of California Irvine, Neurobiology And Behavior, Irvine, United States of America, 3University of California, Irvine, Transgenic Mouse Facility, Irvine, United States of America, 4University of California Irvine, Molecular Biology And Biochemistry, Irvine, United States of America, 5University of California, Irvine, Department Of Developmental And Cell Biology, Irvine, United States of America

Aims: Most Alzheimer’s disease (AD) cases are sporadic and late-onset, yet nearly all existing mouse AD models harbor pathogenic mutations, rendering them better representations of familial autosomal-dominant forms of the disease. As a foundational step to model late-onset AD (LOAD), we generated and analyzed knock-in mice that express wildtype human amyloid beta (hAβ-KI) under control of the mouse App locus in exon 14 flanked by loxP sites (FL). Methods: Using confocal microscopy, we performed immunofluorescence stains on coronal brain slices and quantified the appearance of astrocyte-associated clusters of granules, which stained positive via Periodic Acid-Schiff (PAS) stain. To correlate the appearance of PAS granules with hAβ expression, we crossed hAβ-KI-FL mice with UBC-CreERT2 mice to enable Tamoxifen-inducible cre recombinase-mediated inactivation of hAβ expression. Finally, we generated hAβ-KI; PS1M146V mice to investigate whether co-expression of familial-linked AD mutations affect the hAβ-induced presence of PAS granules. Results: Changing 3 amino acids in the mouse Aβ sequence to its wild-type human counterpart produced age-dependent accumulation of PAS granules in the hippocampus of hAβ-KI mice by 10 months of age. Additionally, we observed impairments in cognition and synaptic plasticity in hAβ-KI mice. Remarkably, ablating hAβ expression using inducible cre reduced the formation of PAS granules and rescues cognition. Co-expression of familial-linked AD mutations with hAβ exacerbated the accumulation of PAS granules compared to homozygous hAβ-KI and WT mice. Conclusions: Substituting mouse Aβ with the wild-type human isoform produces significant accumulation of PAS granules and impairments in cognition and synaptic plasticity, highlighting its usefulness in investigating LOAD risk factors.

P302 / #1436


DIFFERENTIAL PLASMA BIOMARKER PROFILE IN FAMILIAL ALZHEIMER DISEASE

Lecture Title:

C. Johansson1, C. Graff1, K. Blennow2, H. Zetterberg3, S. Thordardottir1, J. Pannee3 1Karolinska Institutet, Center For Alzheimer Research, Department Nvs, Division Of Neurogeriatrics, Stockolm, Sweden, 2University of Gothenburg, Institute Of Neuroscience And Phsyiology, Mölndal, Sweden, 3Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden

Aims: We aim to explore if biomarkers in CSF and peripheral blood are congruent as well as indicative of different stages of disease in a familial Alzheimer disease cohort from Sweden. We will also explore the fluid biomarker profiles with respect to mutation type. Methods: Relatives with 50% risk of inheriting one of the disease-causing mutations are invited to participate in a longitudinal prospective study of Swedish kindreds with familial Alzheimer disease. A full study protocol includes examination by physician, neuropsychological assessment, magnetic resonance imaging (MRI), electroencephalography (EEG) and sampling of blood, cerebrospinal fluid (CSF) and skin biopsies. The participants are in preclinical or prodromal phases, but may also convert during follow-up. One hundred and eighty four plasma samples from 89 individuals have been included. Neurofilament light chain (NFL) and total tau (T-tau) will be assessed by using Simoa and amyloid-beta (Aβ) species (Aβ1-38, Aβ1-40, Aβ1-42 and APP669-711) by using an immunoprecipitation-mass spectrometry assay. The results will be investigated regarding correlation to previously published CSF results, years to expected onset and mutation type, applying a linear mixed effects model. Results: Preliminary findings of Aβ species in a subgroup from the APPswe kindred show that MCs (n=3) have higher concentrations of Aβ1-40 (866 ±111pg/mL) and Aβ1-42 (99 ±12pg/mL) compared to NCs (n=8) (Aβ1-40 253 ±38pg/mL and Aβ1-42 28 ±6pg/mL). Conclusions: An in depth understanding of mutation-specific variability is required and might influence the feasibility of these biomarkers being used in a mixed Alzheimer cohort.

P303 / #425


TRIPLE KNOCKOUT OF APP-FAMILY ALTERS BRAIN MORPHOLOGY AND DISRUPTS BASAL SYNAPTIC TRANSMISSION AND SYNAPTIC PLASTICITY

Lecture Title:

S. Klein1, M. Back2, D. Fäßler1, L. Slomianka3, S. Ludewig4, M. Korte4,5, J. Von Engelhardt2, U. Müller1 1Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Department Of Bioinformatics And Functional Genomics, Heidelberg, Germany, 2Institute for Pathophysiology, Universitätsmedizin Mainz, Mainz, Germany, 3Institute of Anatomy and Zurich Center for Integrative Human Physiology, University Of Zurich, Zurich, Switzerland, 4Zoological Institute, TU Braunschweig, Division Of Cellular Neurobiology, Braunschweig, Germany, 5Helmholtz Centre for Infection Research, Neuroinflammation And Neurodegeneration Group, Braunschweig, Germany

Aims: The key role of APP in the pathogenesis of Alzheimer disease is well established. However, postnatal lethality of triple knockout mice has so far precluded the analysis of the physiological functions of APP and the APLPs in the developing and adult brain for the formation of synapses, synaptic function and plasticity. Methods: Here, we generated conditional APP/APLP1/APLP2 triple KO (cTKO) mice lacking the APP family in excitatory forebrain neurons from E12 onwards using NexCre mice. Results: Conditional cTKO mice proved fully viable but showed altered brain morphology with agenesis of the corpus callosum and impaired lamination of the hippocampus, characterized by splitting of the CA1 band into deep and superficial sublayers. Due to this delamination we studied the morphology of superficial (sCA1) and deep (dCA1) pyramidal cells separately. Interestingly, CA1 neurons of cTKO mice showed reduced spine density as well as impairments in dendritic architecture, in a distinct pattern along the radial axis of the CA1 band. Further, electrophysiological recordings in the hippocampus of adult cTKO mice indicated a strong synaptic phenotype with pronounced deficits in the induction and maintenance of hippocampal LTP and impairments in paired pulse facilitation, indicating a possible presynaptic deficit. Conclusions: Collectively, our analysis reveals an essential role of APP family proteins in excitatory principal neurons for mediating normal dendritic architecture, spine density, morphology and synaptic plasticity.

P304 / #325


LRP3, ANOTHER LDLR FAMILY MEMBER AFFECTED BY ALZHEIMER’S DISEASE

Lecture Title:

M. Lennol, I. Cuchillo-Ibáñez, S. Escamilla, I. López-Font, J. Sáez-Valero Universidad Miguel Hernández, Instituto De Neurociencias, Sant Joan d'Alacant, Spain

Aims: The LDL receptor (LDLR) family includes cell surface receptors with key roles in synaptic plasticity, such as LRP8/ApoER2. LRP3 is the most unknown member of this family. Here, we studied the relation between LRP3 and ApoER2 signalling cascade, as well as between LRP3 and APP, as many members of this family have recently been related to APP processing. We have also analysed LRP3 expression in Alzheimer’s disease (AD). Methods: LRP3 expression was analysed by microarray, qRT-PCR, immunoprecipitation and western-blots in human frontal cortex homogenates from AD patients (n= 30) and non-demented controls (NDC, n= 8), and in extracts from cell cultures. Results: Overexpression of full-length ApoER2 enhanced expression of LRP3 mRNA and protein levels in SH-SY5Y cells, being more evident after ApoER2 signalling stimulation by reelin. Overexpression of ICD-ApoER2, an ApoER2 intracellular domain with transcriptional activity, also induced LRP3 expression, while β-amyloid protein (Aβ42) treatment decreased LRP3 expression. LRP3 mRNA and protein levels were found lower in AD patients compared to control subjects. Finally, LRP3 expression affected APP expression. Conclusions: We have demonstrated a novel cross-talk between the ApoER2/reelin signaling and LRP3 expression, and also that LRP3 levels are affected in AD. This receptor, as others of the same family, seems to be related to APP expression or processing.

P305 / #1249


AMYLOID PRECURSOR PROTEIN REGULATION OF GLUTAMATERGIC SYNAPSES IN EARLY DEVELOPMENT

Lecture Title:

J. Rajão-Saraiva1, M. Temido-Ferreira1, S.F. Almeida1, H. Marie2, L. Lopes1, P.A. Pousinha2 1Instituto de Medicina Molecular João Lobo Antunes, Faculdade De Medicina, Universidade De Lisboa, Lisbon, Portugal, 2Université Côte d'Azur (UCA), CNRS UMR7275, Institute Of Molecular And Cellular Pharmacology (ipmc), Valbonne, France

Aims: Recent studies have described amyloid precursor protein (APP) and its derived fragments as regulators of glutamatergic synaptic function. According to the emerging model, APP intracellular domain modulates the composition of synaptic NMDA receptors, promoting an immature-like GluN2B-rich profile (Pousinha et al., 2017). Therefore, we intend to unravel the cellular mechanisms underlying the APP physiological regulation of glutamatergic synapses in early development. Methods: We have conducted experiments in wild-type mice, comparing infant (P7-P10) and adult animals. The distribution of both proteins in the hippocampus/cortex was assessed by immunohistochemistry. Additionally, the protein levels of APP and NMDAR subunits were determined by western blot in hippocampal samples. In parallel, NMDAR synaptic contribution was assessed by Patch-Clamp electrophysiology. Results: We have observed a wide distribution of APP/GluN2B in the hippocampus/cortex of infant mice. When comparing to adult animals, there is a peak in APP protein levels and GluN2B-NMDAR synaptic contribution in early postnatal stages. These observations point towards a crucial role for APP in immature synapses, where it might control synaptic NMDARs subunit composition. Finally, when interfering with APP intracellular domain accessibility, we have induced an alteration in GluN2B-NMDAR mediated currents. Conclusions: Our data suggests that APP might play an important role in synaptogenesis by controlling synaptic NMDA receptors composition at immature synapses.

P306 / #426


TRIPLE KNOCKOUT OF APP-FAMILY LEADS TO SEVERE DEFICITS IN COGNITIVE TESTS AND AUTISM-LIKE BEHAVIOR

Lecture Title:

V. Steubler1, I. Amrein2, D. Wolfer2,3, U. Müller1 1Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Department Of Bioinformatics And Functional Genomics, Heidelberg, Germany, 2Institute of Anatomy and Zurich Center for Integrative Human Physiology, University Of Zurich, Zurich, Switzerland, 3Institute of Human Movement Sciences, Eth Zurich, Zurich, Switzerland

Aims: The key role of the amyloid precursor protein APP in the pathogenesis of AD is well established. Despite this, its physiological role and that of the related APLPs is still poorly understood. The postnatal lethality of constitutive triple knockout mice lacking all APP family proteins has so far precluded the analysis of APP family functions in the adult brain. Methods: Here, we generated conditional APP/APLP1/APLP2 triple KO (cTKO) mice lacking the APP family in excitatory forebrain neurons from E12 onwards using NexCre mice. Results: Conditional TKO mice showed no alterations in grip strength and rotarod performance indicating normal muscle strength and motor coordination. In the open field, that assesses locomotor activity in a novel environment, they failed to habituate and showed hyperactivity. In line with this, we observed overshooting dark phase activity when tested in a familiar homecage. Subsequently, cTKO mice underwent a panel of tests to assess spatial learning and memory (T-Maze, 8-arm radial maze, Barnes and Morris Water maze). In all of these tests the performance of cTKO mice was strongly impaired with no evidence of learning. Further, we observed a high incidence of stereotypic and preserverative behaviors reminiscent of an ASD-like phenotype. Indeed, we could confirm autism-like behavior in cTKO mice including stereotypic repetitive rearing and climbing, abnormal patterns of ultrasonic vocalization in pups and impaired social interaction in the three chambers test. Conclusions: In conclusion, our study reveals an essential role of the APP family for brain circuits important for cognition and social behavior.

P307 / #983

Topic: Theme A: β-Amyloid Diseases / A6.b. Cell, Molecular and Systems Biology: ApoE

CHARACTERIZATION OF NEW ANTI-LRP8 (APOER2) ANTIBODIES TO INVESTIGATE ITS PROCESSING AND ITS ROLE IN AD PATHOGENESIS

Lecture Title:

E. Foderà1, A. Medoro1, D. Mignogna1, C. Porcile1, D. Passarella1, S. Ciampi1, S. Bartollino1, E. Imperlini2, R.M.A. Di Marco1, G. Guerra1, G. Raimo1, M. Intrieri1, A. Pagano3, T. Florio4, C. Russo1 1University of Molise, Department Of Medicine And Health Sciences, Campobasso, Italy, 2IRCCS SDN, -, Naples, Italy, 3University of Genova, Department Of Experimental Medicine, Genova, Italy, 4University of Genova, Department Of Internal Medicine And Centre Of Excellence For Biomedical Research, Genova, Italy

Aims: LRP8 is the main neuronal ApoE receptor and is proteolytically cleaved by γ-secretase, like AβPP, with which interacts through adaptor proteins. Given its role in neuronal migration, formation of cortical layers, memory and learning, a potential aberrant LRP8 processing could be implicated in Alzheimer’s Disease (AD) onset and/or progression. In this scenario, we designed and developed polyclonal and monoclonal antibodies to further investigate LRP8 C-terminal processing and its role in AD pathology. Methods: Monoclonal and polyclonal antibodies were developed by using GenScript service. Polyclonal antibodies against two sequences localized in the C-terminal region of LRP8 were produced after rabbit immunization followed by affinity purification of the antibodies from serum. Monoclonal antibodies production involved the BALB/c mouse immunization following the MonoExpress protocol. The characterization of the antibodies was performed by Western Blot (WB) and immunochemistry. Results: Our polyclonal and monoclonal antibodies detect both the Full-Length and the C-terminal fragments of endogenous and overexpressed LRP8 in N2a wild-type and N2a LRP8-DDK-Myc cells. LRP8-KO mouse brain lysate doesn’t show any LRP8 signal by WB. By immunochemistry, our antibodies localize LRP8 in neurons and cerebral parenchyma of human frontal cortices of healthy brains. Moreover, in N2a LRP8-DDK-Myc cells LRP8 is detected in the cell membrane, axons and dendritic spines. Conclusions: Both polyclonal and monoclonal antibodies show specificity for LRP8 by WB, identifying both the Full-Length receptor and its C-terminal fragments, in particular the ones ranging from 8 to 12 kDa which are not detectable by any of the currently available commercial antibodies.

P308 / #1308


NEUROBIOLOGICAL ROLE OF APOE ON EARLY CELLULAR AND SYNAPTIC CHANGES IN MODELS OF ALZHEIMER’S DISEASE

Lecture Title:

S. Konings, I. Martinsson, L. Torres-Garcia, G. Gouras Lund University, Department Of Experimental Medical Science, Lund, Sweden

Aims: Apolipoprotein E4 (ApoE4), the major genetic risk factor for Alzheimer’s disease (AD), transports lipids from astrocytes to neurons, where it binds receptors and is internalized. It was reported that ApoE4 impairs normal receptor recycling, modulates synaptic receptors and is associated with endosome enlargement. ApoE4 target replacement (TR) mice show neuronal hyperexcitability. The mechanisms, cell types and targets of ApoE relating to AD remain poorly understood. Here we elucidate cellular mechanisms of different ApoE isoforms on early neurobiological changes in AD, including intraneuronal Aβ, endosomes and synapses. Methods: Cortical and hippocampal mouse neurons and astrocytes are derived from wild-type, ApoE KO, ApoE3-TR and ApoE4-TR mice. Astrocyte-conditioned medium from ApoE-TR and recombinant ApoE are used as a source for human ApoE. Primary neurons are analyzed by immunofluorescence, Western blot and live cell calcium imaging. Results: Although ApoE is mainly produced by astrocytes, it can also be made by neurons and microglia. In neurons ApoE appears to localize at neurites and synapses. Both endogenous and added ApoE and Aβ are also in the endosome-lysosome system, a site in neurons where early AD changes occur. Ca2+ live cell imaging shows differences in neuronal excitability with different ApoE isoforms. Conclusions: ApoE appears to localize to sites important in early cellular changes in AD, such as endosomes, neurites and synapses and thereby could influence neuronal excitability. We hypothesize that determining the biology of ApoE within neurons can contribute to the better understanding of the role of ApoE in Alzheimer’s pathogenesis.

P309 / #1365


BRAIN APOLIPOPROTEIN E LEVELS ARE ALLELE DEPENDENT AND MODULATED BY DIETARY FAT

Lecture Title:

B. Liemisa1, S. Newbury1, M. Novy1, E. Levy1,2,3,4, P. Mathews1,2,3 1Nathan Kline Institute, Center For Dementia Research, Orangeburg, United States of America, 2New York University School of Medicine, Psychiatry, New York, United States of America, 3New York University School of Medicine, Neuroscience Institute, New York, United States of America, 4New York University School of Medicine, Biochemistry & Molecular Pharmacology, New York, United States of America

Aims: The apolipoprotein E (APOE) ε4 allele is the single greatest genetic risk factor for Alzheimer’s disease (AD), while ε2 is protective and ε3 risk-neutral. In this study, we examined APOE levels in the blood plasma and brains of mice expressing the three human APOE alleles when challenged by a diet high in fat. Methods: Baseline APOE protein levels were examined by Western blot analysis of mice humanized and homozygous for the ε2, ε3, or ε4 APOE alleles and maintained on a normal chow diet (14% of calories from fat) at three ages: 6, 12, and 18 months of age. Additional mice were placed on a Western diet (39.1% of calories from fat) at 6 months of age until examined in parallel to control diet mice at 12 months of age. APOE protein levels were additionally quantified by a human-specific APOE sandwich ELISA that we developed. Results: At all three ages, brain APOE levels were found to be highest in APOE2 mice, followed by APOE3 and APOE4, similar to the relative levels seen in blood plasma (APOE2 > APOE3 > APOE4). In all three genotypes, Western diet for 6 months increased both brain and plasma APOE levels, with the increase more pronounced in plasma. Conclusions: The three human APOE alleles impact the levels of APOE protein in the brain and in plasma, with the protective APOE2 protein being the most abundant. Additionally, a diet high in fat increases the levels of the three APOE proteins in the brain and plasma.

P310 / #994


DIFFERENTIAL LOCALIZATION AND PROCESSING OF LRP8 (APOER2) IN THE CEREBRAL CORTEX OF SPORADIC AND FAMILIAL ALZHEIMER’S DISEASE PATIENTS

Lecture Title:

A. Medoro, D. Mignogna, D. Leccese, C. Porcile, E. Foderà, D. Passarella, S. Ciampi, M. Intrieri, C. Russo University of Molise, Department Of Medicine And Health Sciences, Campobasso, Italy

Aims: LRP8 is a neuronal receptor of the major risk factor for Alzheimer’s Disease (AD): ApoE. LRP8 is involved in the regulation of neuronal migration, formation of cortical layers, memory and learning processes and is a substrate of γ-secretase, like Amyloid-β Precursor Protein (AβPP), with which interacts through adaptor proteins in the intracellular C-terminus. The aim of this project is to characterize the LRP8 localization and its proteolytic processing in human brain of sporadic (SAD) and familial AD (FAD) patients compared to controls. Methods: Human frontal cortices are analyzed by Western Blot (WB) and immunohistochemistry (IHC). Results: IHC analysis reveals a LRP8 localization in neurons and cerebral parenchyma of all cortical layers, especially in layer I. In SAD and FAD brains LRP8 signal is generally reduced and totally absent in neurons with neurofibrillary tangles. In line with IHC results, AD brains show decreased levels of LRP8 Full-Length variants (105 kDa) compared to control brains by WB. Moreover, we can appreciate a different processing pattern with increased levels of LRP8 C-terminal fragments (between 8 and 12 kDa) in AD cases and a correspondent decrease in controls. Conclusions: Overall, these results show a differential LRP8 localization and processing between control and AD brains characterized by a decreased LRP8 signal and a correspondent increase of its processing in AD brains. Interestingly, both IHC and WB observations show a strict correlation with the severity of the disease, with FAD cases having a more prominent pathological LRP8 phenotype compared to SAD patients.

P311 / #873

Topic: Theme A: β-Amyloid Diseases / A6.c. Cell, Molecular and Systems Biology: Secretases

PRESENILIN 1 FAMILIAL ALZHEIMER’S DISEASE ONSET DEPENDS ON THE EXTENT OF AΒ43 CROSS-TALKING GENERATION

Lecture Title:

N. Kakuda, M. Takami Doshisha University, Neuropathology, Kyotanabe-shi, Kyoto, Japan

Aims: Objective Familial Alzheimer’s disease (FAD) mutations carrying presenilin (PS) has been known as a cause of generating absolute β-amyloid protein (Aβ) Aβ43. In FAD, Aβ42 and Aβ43 make senile plaque earlier than sporadic AD. This study investigated why Aβ43 generation increased in PS1 mutants, based on the γ-secretase stepwise processing mechanism. Methods: The Aβ concentrations in the cerebrospinal fluid (CSF) of all subjects were measured with specific ELISAs. To assess the γ-secretase activity, a large amount of γ-secretase containing lipid-raft fractions collected from wild type human PS1 or mutant PS1 stably expressing cells. To generated Aβs and amyloid precursor protein (APP) intracellular domain (AICD) in vitro assay did and detected with western blotting. Here we also measured released tri and tetra peptides with LC-MS/MS, which generated from βCTF during stepwise processing. Results: The level of Aβ38 and Aβ40 were low concentrations in FAD patients compared with control, mild cognitive impairment, and sporadic AD. In the in vitro γ-secretase assay, FAD mutants decreased Aβ38 and Aβ40 generation from those of precursors, Aβ42 and Aβ43 rather than wild type (WT) PS1. Thus, both ratios of Aβ40/Aβ43 and Aβ38/Aβ42 (product/precursor set) of mutants decreased compared with WT PS1. Importantly, the ratio of Aβ43/AICD increased onset age-dependent, but other Aβs/AICD ratios decreased or unchanged. Further assessment of Aβ43 generation with LC-MS/MS, Aβ43 was generated both Aβ46 and Aβ48. Conclusions: Here, PS1 mutants occurred cross-taking generation from Aβ48 to Aβ43. These data suggest that FAD γ-secretase activity is different from sporadic AD.

P312 / #335

Topic: Theme A: β-Amyloid Diseases / A6.c. Cell, Molecular and Systems Biology: Secretases

MECHANISM UNDERLYING THE SEQUENTIAL GSEC-MEDIATED CLEAVAGE OF TRANSMEMBRANE DOMAINS

Lecture Title:

M. Koch1, T. Enzlein1,2, S. Lismont1, L. Chávez-Gutiérrez1 1VIB/KU Leuven, Vib-ku Leuven Center For Brain & Disease Research, Leuven, Belgium, 2Mannheim University of Applied Sciences, Center Of Applied Research In Biomedical Mass Spectrometry, Mannheim, Germany

Aims: γ-Secretases (GSECs) are intramembrane proteases successively cleaving a variety of type 1 transmembrane proteins, such as the Amyloid-Precursor-Protein (APP). Cleavage of APP by GSECs leads to the production of Aβ peptides of different lengths. Mutations in GSECs and APP cause familiar Alzheimer’s disease (AD). The pathogenic mutations consistently enhance the production of longer Aβ species by destabilizing GSEC-APP interactions. These findings place the enzyme-substrate stability as a central mechanism in AD-pathogenesis. We aim at characterizing the molecular interactions securing the GSEC-substrate complexes during sequential catalysis. We are interested in the development of strategies that target GSEC-APP interactions, and shift the Aβ production towards shorter peptides, without affecting crucial GSEC-mediated signaling cascades. Methods: We performed function-structure analyses of GSEC-mediated cleavage using wild type/mutant enzyme and a selection of GSEC substrates in cell-free and cell-based assays. ELISA, immunoprecipitation and MALDI-TOF mass spectrometry analyses of GSEC-generated products were performed for in-depth analysis of the GSEC-mediated cleavage mechanism. Results: Here, we identified an extracellular GSEC-APP anchoring mechanism that dictates Aβ-length. Conclusions: The here characterized anchor highlights the powerful role of the extracellular GSEC-substrate interface in the regulation of the GSEC processivity. These findings shed light on the molecular foundations of GSEC-mediated cleavage and may guide future strategies safely targeting GSECs in therapeutic settings.

P313 / #749

Topic: Theme A: β-Amyloid Diseases / A6.d. Cell, Molecular and Systems Biology: Growth factors, synaptic plasticity

THE LONG-TERM EFFECT OF SELECTIVE CHOLINERGIC DEPLETION ON COGNITION AND NEUROTROPHIN STATUS IN WILD-TYPE RATS.

Lecture Title:

C. Orciani1, H. Hall2, R. Pentz1, M. Foret2, A.C. Cuello1,2,3 1McGill University, Neurology And Neurosurgery, MONTREAL, Canada, 2McGill University, Pharmacology And Therapeutics, Montreal, Canada, 3McGill University, Anatomy And Cell Biology, MONTREAL, Canada

Aims: Addressing whether a reciprocal interaction between Basal forebrain cholinergic neurons (BFCNs) and the status of the neurotrophins exists. BFCNs represent the main source of cholinergic innervation to the cortex and hippocampus and degenerate early in the progression of Alzheimer’s Disease (AD). BFCNs phenotypic maintenance is dependent on the availability of mature Nerve Growth Factor (mNGF) and mature Brain-Derived Neurotrophic Factor (mBDNF), produced by target neurons and retrogradely transported to the cell body. Methods: We lesioned the nucleus basalis (nb, a basal forebrain cholinergic nuclei that projects mainly to the cortex) by bilateral stereotaxic injection of 192-IgG-Saporin (the cytotoxin Saporin binds the p75ntr receptors expressed exclusively by BFCNs) in 2.5-month-old Wistar rats. At six months post-lesion, we performed a modified version of the 5-Choice Serial Reaction Time Task (5-CSRTT). After behaviour, the rats were perfused, and their brains were collected. Results: A reduction of Choline Acetyltransferase-immunoreactive neurons confirmed the partial lesion of the nb in Saporin-injected rats (SAP). The global score, given by a combination of the overall 5-CSRTT performance, was reduced in SAP rats as compared to controls and correlated with a diminished number of cortical Vesicular acetylcholine transporter-immunoreactive boutons. BDNF transcription levels and NGF precursor (proNGF) protein levels were reduced in the SAP compared to the control. Furthermore, inflammatory markers were elevated in the SAP-treated rats. Conclusions: Long term loss of nb impacted cognitive abilities, cortical levels of proNGF, BDNF and inflammation. These results could have implications in neurodegenerative conditions such as AD.

P314 / #827

Topic: Theme A: β-Amyloid Diseases / A6.f. Cell, Molecular and Systems Biology: Network biology, connectome, protein-protein interations

LINKING MOLECULAR BIOMARKERS AND BEHAVIOR IN PRECLINICAL MODELS AND PATIENTS WITH ALZHEIMER’S DISEASE

Lecture Title:

C. Bjørkli1, M. Hemler1, N. Ebbesen1, M. Witter2, R. Nair2, A. Kobro-Flatmoen2, A. Sandvig1, I. Sandvig1 1The Norwegian University of Science and Technology, Department Of Neuromedicine, Trondheim, Norway, 2Kavli Institute for Systems Neuroscience, Faculty Of Health And Medicine, Trondheim, Norway

Aims: Alzheimer’s disease (AD) is a debilitating neurodegenerative disorder that causes severe deterioration of memory, cognition, behavior, and the ability to perform daily activities. The disease is characterized by the accumulation of two proteins in fibrillar form; Amyloid-β forms fibrils that accumulate as extracellular plaques while tau fibrils form intracellular tangles. Here we aim to translate findings from a commonly used AD mouse model to AD patients. Methods: Here we initiate and chronically inhibit neuropathology in lateral entorhinal cortex (LEC) layer two neurons in an AD mouse model. This is achieved by over-expressing P301L tau virally and chronically activating hM4Di DREADDs intracranially using the ligand dechloroclozapine. Biomarkers in cerebrospinal fluid (CSF) is measured longitudinally in the model using microdialysis, and we use this same system to intracranially administer drugs aimed at halting AD-related neuropathology. The models are additionally tested in a novel contextual memory task. Results: Preliminary findings indicate that viral injections of P301L tau into LEC layer two reveal direct projections between this region and the outer molecular layer of dentate gyrus and the rest of hippocampus. Additionally, phosphorylated tau co-localize with 'starter cells' and appear to spread from the injection site. Preliminary microdialysis results suggest that the concentrations of CSF amyloid-β and tau proteins mirror changes observed along the disease cascade in patients. The disease-modifying drugs appear to halt neuropathological development in this preclincial model. Conclusions: These findings will lead to a novel platform for translational AD research, linking the extensive research done in rodents to clinical applications.

P315 / #694

Topic: Theme A: β-Amyloid Diseases / A6.f. Cell, Molecular and Systems Biology: Network biology, connectome, protein-protein interations

EXPLORING THE AMYLOID ‘CORONA’ AS A MODEL FOR SENILE PLAQUE PATHOLOGY

Lecture Title:

C. Lendel1, M. Rahman1, H. Chaudhary1, J. Löfblom2, H. Zetterberg3 1KTH Royal Institute of Technology, Chemistry, Stockholm, Sweden, 2KTH Royal Institute of Technology, Protein Science, Stockholm, Sweden, 3Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden

Aims: Alzheimer’s disease (AD) pathology involves deposition of amyloid-β (Aβ) in senile plaques. However, Aβ is not the only biomolecule that accumulates in the plaques. Thousands of plaque-associated proteins have been identified from ex vivo tissue. Improved understanding of how these proteins are sequestered and what the functional consequences are could generate new knowledge about AD pathology. We employ a bottom-up approach that relies on the observation that amyloid fibrils, like other nanoparticles, attract a ‘corona’ of proteins when exposed to a biological environment, which may give a first glimpse of the multiprotein aggregates that develops into plaques. Methods: Biochemical methods are used to isolate and identify the human proteins that co-aggregate with Aβ in human cerebrospinal fluid (CSF). In particular, we are developing a method based on flow-cytometry sorting of protein aggregates followed by quantitative mass spectrometry-based proteomics. Results: The flow-cytometry method was demonstrated by a quantitative comparison of the protein ‘coronae’ of Aβ(1-40) and Aβ(1-42) aggregates in CSF from AD patients and controls. The quantitative data allowed speculations about the architecture of the aggregates and we found that all proteins do not interact directly with Aβ. A bioinformatics meta-analysis suggests enrichment of certain functional signatures while the structural properties of the proteins do not correlate with their occurrence in the amyloid ‘corona’. Conclusions: We are establishing an in vitro model for the assembly of Aβ-related aggregates in a biologically relevant environment. Such model can bridge the gap between biochemistry and cell biology, provide new pathological insights and open avenues to the development of novel diagnostics and therapy.

P316 / #434

Topic: Theme A: β-Amyloid Diseases / A6.g. Cell, Molecular and Systems Biology: Metabolomics, transcriptomics, lipidomics, proteomics

PROTEOMIC ANALYSIS OF THE HUMAN ENTORHINAL CORTEX IN ALZHEIMER'S DISEASE

Lecture Title:

V. Astillero-Lopez, S. Villar-Conde, M. Gonzalez-Rodriguez, P. Villanueva-Anguita, A. Flores-Cuadrado, D. Saiz-Sanchez, A. Martinez-Marcos, I. Ubeda-Banon Ciudad Real Medical School (University of Castilla-La Mancha), Medical Sciences, Ciudad Real, Spain

Aims: Alzheimer´s disease (AD), the most prevalent neurodegenerative disorder worldwide, is characterized by cognitive deficits. The olfactory system is prematurely involved, and olfactory deficits are among earliest preclinical symptoms. The entorhinal cortex (EC) is implicated in both olfactory and associative processing information since connections with the hippocampus are essential in learning and memory processes. Neuropathologically, AD brains accumulate deposits of amyloid-β and tau proteins. These proteins can propagate cell-to-cell in a prion-like manner inducing native proteins to become pathological. Therefore, the aim of this work was to analyze the proteomic profile of the human EC to elucidate differences between AD and non-AD patients. Methods: Experimental procedures were approved by the Ethical Committee of Clinical Research at Ciudad Real University Hospital (SAF2016-75768-R and PID2019-108659RB-I00). Post-mortem human brain samples were provided by the Spanish Biobank Network including 6 AD and 6 non-AD age-matched cases. Proteomic analysis was performed through LC/MS, and western blot and immunoassays were performed to validate and evaluate the selected proteins. Results: SWATH analysis quantified the relative expression levels of 1635 proteins, reflecting 170 proteins significantly downregulated and 76 upregulated. Data analysis using enrichment and pathway analyses reflected that main pathways affected were related with neuronal and immune system including strong alterations in axon guidance, vesicle cycle processes and synapse function. Conclusions: Sponsored by the UCLM/ERDF (2020-GRIN-29145 to NPND), the Spanish Ministries of Economy and Competitiveness/ERDF (grant no. SAF2016-75768-R) and Science and Innovation (grant no. PID2019-108659RB-I00) to AMM and the Autonomous Government of Castilla-La Mancha/ERDF (grant no. SBPLY/17/180501/000430) to AMM and DSS.

P317 / #1372


UNRAVELLING DIFFERENT GRADE OF RNA METABOLISM INVOLVEMENT IN ALZHEIMER’S DISEASE, PARKINSON’S DISEASE AND AMYOTROPHIC LATERAL SCLEROSIS

Lecture Title:

M. Garofalo1,2, C. Pandini1,2, M. Bordoni2,3, O. Pansarasa2, S. Gagliardi2, C. Cereda2 1University of Pavia, Biology And Biotechnology "l. Spallanzani", Pavia, Italy, 2IRCCS Mondino Foundation, Genomic And Post-genomic Unit, Pavia, Italy, 3Università degli Studi di Milano, Dipartimento Di Scienze Farmacologiche E Biomolecolari (disfeb), Centro Di Eccellenza Sulle Malattie Neurodegenerative, Milan, Italy

Aims: Alzheimer’s disease (AD), Parkinson’s disease (PD) and Amyotrophic Lateral Sclerosis (ALS) are neurodegenerative disorders characterized by progressive degeneration of central or peripheral nervous system. A central role of RNA metabolism has emerged in these diseases, concerning mRNAs processing and non-coding RNAs biogenesis. We aimed to identify possible crossroads or deviations in the dysregulated pathways of AD, PD and ALS. Methods: We performed RNA-seq analysis to investigate the regulation of both coding and long non-coding RNAs (lncRNAs) in ALS, AD and PD patients and controls (CTRL) in Peripheral Blood Mononuclear Cells (PBMCs). Results: A total of 293 differentially expressed (DE) lncRNAs and 87 mRNAs was found in ALS patients. In AD patients a total of 23 DE genes have emerged, 19 protein coding genes and 4 lncRNAs. By performing KEGG and GO analysis we found common affected pathways and biological processes in ALS and AD. In PD patients only 5 genes were found DE. Conclusions: Our data brought the light on the different importance of lncRNAs and mRNAs regulation in the principal neurodegenerative disorders, offering starting points for new investigations about pathogenic mechanism involved in them.

P318 / #432


PROTEOMIC ASSESSMENT OF HUMAN HIPPOCAMPUS IN ALZHEIMER’S DISEASE.

Lecture Title:

M. Gonzalez-Rodriguez, V. Astillero-Lopez, S. Villar-Conde, P. Villanueva-Anguita, I. Ubeda-Banon, A. Flores-Cuadrado, A. Martinez-Marcos, D. Saiz-Sanchez Ciudad Real Medical School (University of Castilla-La Mancha), Medical Sciences, Ciudad Real, Spain

Aims: Alzheimer’s disease is characterized by executive dysfunction and memory impairment, underlying accumulation of extracellular amyloid-β and intracellular hyperphosphorylated tau. Recently, prion-like hypothesis states that both proteinopathies can spread out from cell-to-cell throughout different brain regions. Hippocampus is a key region in memory formation and it is involved in early stages of disease. State-of-the-art -omic approaches would allow multilevel analysis but are currently underdeveloped. Proteomic studies on human tissue would be especially suitable to identify peptide fingerprint changes during pathology among hippocampal fields. Methods: Post-mortem tissue was provided by IDIBAPS, BTCIEN, BIOBANC-MUR and BPA Spanish National Biobanks. Experimental procedures were approved by Ethical Committee of Clinical Research at Ciudad Real University Hospital (SAF2016-75768-R). A total of 12 cases were used for SWATH analysis. Proteomic analyses consisted in PCA, heatmap, volcano plot and detection of activated/deactivated pathways. Immunofluorescence and western blot analysis were carried out to evaluate selected proteins. Results: A total of 1635 proteins were identified in SWATH analysis. FC > 1.5 and p-value <0.01 were established for proteomic analysis, which reflected 47 upregulated proteins and 102 downregulated. Main pathways altered were related to immune system, neuronal system and cell cycle. Therefore, those proteins implicated in synapses and microtubule organization and their interaction with pathological proteins were evaluated by immunofluorescence and western blot assessments. Conclusions: Sponsored by UCLM/ERDF (2020-GRIN-29145 to NPND), Spanish Ministries of Economy and Competitiveness/ERDF (SAF2016-75768-R) and Science and Innovation (PID2019-108659RB-I00) to AMM and Autonomous Government of Castilla-La Mancha/ERDF (SBPLY/17/180501/000430) to AMM and DSS. MGR received a UCLM/ERDF predoctoral fellowship.

P319 / #1454


SINGLE NUCLEI RNA-SEQ DATA ANALYSIS IDENTIFIES CELL-TYPE SPECIFIC LINEAGES ASSOCIATED WITH ALZHEIMER’S DISEASE SEVERITY

Lecture Title:

L. Heath1, K. Prater2, K. Green2, R. Elyanow3, E. Melief4, P. Valdmanis5, E. Blue5, C. Keene4, P. Crane6, E. Larson7, S. Willis8, J. Young4,9, G. Garden2, S. Jayadev2, A. Greenwood1, B. Logsdon1, L. Mangravite1 1Sage Bionetworks, Neurodegeneration Research, SEATTLE, United States of America, 2University of Washington, Neurology, Seattle, United States of America, 3Adaptive Biotechnologies, Antigen Map Team, Seattle, United States of America, 4University of Washington, Pathology, Seattle, United States of America, 5University of Washington, Division Of Medical Genetics, Seattle, United States of America, 6University of Washington, Division Of Internal Medicine, Seattle, United States of America, 7Kaiser Permanente Washington Health Research Institute, Aging & Geriatrics, Seattle, United States of America, 8University of Washington, Psychiatry And Behavioral Sciences, Seattle, United States of America, 9University of Washington, Laboratory Medicine And Pathology, Seattle, United States of America

Aims: Single nucleus RNA sequencing (snRNA-seq) has the potential to improve our understanding of the cellular-specific drivers of late-onset Alzheimer’s disease (AD). We employ a lineage-based approach to infer brain-tissue-derived celltype-specific gene expression across the continuum of AD-related disease states. Methods: Primary analyses used Mathys et al. 2019 snRNA-seq derived from post-mortem dorsolateral prefrontal cortex tissue of 48 participants (24 cases selected based on AD-related pathology plus 24 age-matched controls) in the ROS/MAP cohort. Data were normalized using scran, with normalized counts pre-processed in Monocle 3 via UMAP-based dimension reduction techniques. Clustering and calculation of pseudotime lineages were learned separately by sex for each cell subtype with Monocle3. We additionally utilized snRNA-seq data from prefrontal cortex tissue from 9 participants across the Adult Changes in Thought (ACT), University of Washington Alzheimer’s Disease Research Center (UW-ADRC), and Seattle Longitudinal Study (SLS) cohorts as a replication data set. Results: We identified cell-type specific lineages that were significantly associated with disease state. Gene expression was significantly associated with pseudotime for multiple genes, at different points across the disease-associated lineages. Results differed by sex. Conclusions: AD is a continuum, not a discrete state; however, brain tissue samples are only collected at autopsy, so we are unable to observe transitions through time directly. By inferring time-ordered lineages across samples of heterogeneous neuropathology, we uncovered celltype-specific gene expression changes that may signal important transitions at different stages in disease progression.

P320 / #1378


TRANSCRIPTIONAL CHARACTERIZATION OF MOLECULAR AND CELLULAR ALTERATIONS ASSOCIATED WITH AMYLOID BETA AND HYPERPHOSPHORYLATED TAU IN FRONTAL CORTEX BRAIN BIOPSIES

Lecture Title:

T. Kamath1,2,3, C. Vanderburg4, V. Gazestani4, J. Li4, A. Abdulraouf4, N. Nadaf4, T. Rauramaa5, S. Marsh4,6, B. Stevens4,6,7, V. Leinonen5, E. Macosko1,8 1Broad Institute, Stanley Center For Psychiatric Research, Cambridge, United States of America, 2Harvard University, Graduate Program In Biophysics, Boston, United States of America, 3Harvard Medical School, Md-phd Program, Boston, United States of America, 4Broad Institute of Harvard and MIT, Stanley Center For Psychiatric Research, Cambridge, United States of America, 5University of Eastern Finland and Kuopio University Hospital, Neurosurgery, Kuopio, Finland, 6Boston Children's hospital, F.m.kirby Neurobiology Center, Boston, United States of America, 7Howard Hughes Medical Institute, Boston Children's Hospital, Chevy Chase, United States of America, 8Massachusetts General Hospital, Psychiatry, Boston, United States of America

Aims: Recent genetic studies implicate multiple neuroimmune populations in the brain as central players in the pathogenesis of Alzheimer’s disease (AD), yet the precise molecular roles of these and other cell types remain largely unclear. Advances in single-cell technology enable the robust characterization of cell states in the human brain. However, cell states change dramatically upon death, and ex vivo brain tissue samples from living donors that harbor Alzheimer’s histopathology are exceedingly rare. Methods: To unbiasedly assess the molecular changes associated with amyloid-beta and hyperphosphorylated tau across all cell types in the neocortex, we performed single-nuclei RNA-sequencing on 58 high-quality frontal cortex biopsies from individuals with suspected idiopathic normal pressure hydrocephalus who have variable degrees of local amyloid and tau histopathology. Using our optimized protocol for nuclei extraction, we sampled 1.5 million nuclei from the human brain identifying 91 cell types/states, unperturbed by death or agonal state. Results: We identified multiple glial subpopulations associated with amyloid-beta and/or amyloid-beta and hyperphosphorylated tau. We further developed a suite of novel benchmarks for differential expression methods tailored for single-cell expression data and used these insights to identify molecular pathways associated with amyloid and amyloid/tau pathologies. Finally, using a curated set of postmortem tissues, we validated a subset of our cellular and molecular shifts, thereby extrapolating our findings outside of our initial cohort. Conclusions: Single-nuclei RNA-sequencing of frontal cortex biopsies ascertain the cellular and molecular changes associated with amyloid-beta and hyperphosphorylated tau unperturbed by postmortem or agonal state.

P321 / #623


METABOLOMICS PROFILING REVEALS DISTINCT SIGNATURES IN THE SERUM AND BRAIN METABOLOMES IN MOUSE MODELS OF ALZHEIMER’S DISEASE

Lecture Title:

R. Pandey1, M. Arnold2, R.F. Kaddurah-Daouk3, G. Carter4, F.T.M.-A.C. (Model-Ad)4, T.A.D.M.C. Admc3 1THE JACKSON LABAROTARY FOR GENOMIC MEDICINE, Carter Lab, FARMINGTON, United States of America, 2Institute of Bioinformatics and Systems Biology, Helmholtz Zentrum München German Research Center For Environmental Health (gmbh), Neuherberg, Germany, 3Duke University, School Of Medicine, Psychiatry And Behavioral Sciences, Durham, United States of America, 4The Jackson Laboratory, Model-ad, Bar Harbor, United States of America

Aims: Progress in development of efficacious therapies for Alzheimer’s disease (AD) is halted limited understanding of underlying pathological mechanisms. Increasing evidence suggests that metabolic impairments prior to symptom development can contribute to disease mechanisms and subsequent dementia. Signals in conserved metabolomic pathways could provide a method to translate experimental findings in preclinical mouse models to humans. Methods: In this study, we systematically investigated the sex-stratified associations of serum and brain metabolites with the same APOE4.Trem2R47Hand the 5xFAD mouse models at six months of age. Metabolites were measured withthe targeted metabolomics platform AbsoluteIDQ®-p180 (Biocrates AG, Innsbruck, Austria). Results: We identified a sex-specific increase of glycerophospholipids levels in male mice, while levels of sphingolipids were more abundant in females. Further, we identified that serum levels of glycerophospholipidswere reduced in APOE4.Trem2R47Hmice compared to C57BL/6J controls, while levels of these metabolites in the same animals were greater in both male and female brains of APOE4.Trem2R47Hand 5XFAD mice. Several of these findings were consistent with recent results from the ADNI cohort, which suggested a similar decrease in the same metabolites in serum of APOE4 carriers compared to non-carriers and indicates a serum-based effect of APOE genotype. Conclusions: Metabolomic signatures were notably different between brain and serum in mouse models. The 5xFAD mice exhibited stronger effects in brain, whereas the APOE4.Trem2R47Hmouse showed more pronounced effects in serum. These findings are consistent with high levels of amyloid pathology in 5xFAD mouse brains and the modifications of serum biomarkers in APOE4.Trem2R47Hmice.

P322 / #302


METABOLOMICS BIOMARKER DISCOVERY IN CEREBROSPINAL FLUID FOR CEREBRAL AMYLOID ANGIOPATHY

Lecture Title:

E. Van Den Berg1, W. Van Nostrand2, T. Peters1, S. De Boer3, K. Coene3, U. Engelke3, B. Kuiperij1, M. Verbeek1,3 1Radboud University Medical Center, Department Of Neurology, Nijmegen, Netherlands, 2University of Rhode Island, George & Anne Ryan Institute For Neuroscience And Department Of Biomedical And Pharmaceutical Sciences, Kingston, United States of America, 3Radboud University Medical Center, Department Of Laboratory Medicine, Nijmegen, Netherlands

Aims: Metabolite levels may be altered due to vascular amyloid-ß deposits and neuroinflammation in cerebral amyloid angiopathy (CAA). In the Cerebral Amyloid Angiopathy Fluid Biomarker Evaluation (CAFE) study, we aim to discover and validate novel CAA-specific cerebrospinal fluid metabolite biomarkers. Methods: A small-vessel CAA transgenic rat model (n=10, 6 months), the rTg-DI model, was used to study a genetically homogenous group, housed in equal environments to age and sex-matched wild-type rats (n=8). Next-generation metabolic screening is our newly developed methodology to study the metabolome, using high-resolution liquid chromatography quadrupole time-of-flight mass spectrometry. Untargeted analysis using univariate and multivariate statistical approaches was performed. Results: Over >20,000 different features were detected in one sample, defined by their unique combination of accurate mass, specific retention time and intensity. Univariate analysis resulted in 48 features differing significantly between groups, irrespective of sex. Multivariate analysis yielded a clear group separation. Human Metabolome Database enabled annotation of interesting features. Probable markers related to oxidative damage, and glutamatergic and cholinergic neurotransmission.

Conclusions: Our powerful innovative technology for untargeted and simultaneous analysis of a large number of metabolites in body fluids shows promise in discovering novel potential biomarkers for CAA. Using a robust animal model allows us to study biomarkers along the trajectory of disease progression. Insight in underlying processes and metabolic pathways involved in the pathophysiology of CAA will be gained. Validation of candidate markers and discovery of additional ones in a novel cohort of 3/6/12 months old rat samples (n=59) is currently ongoing; results will be presented during the conference.

P323 / #474


BRAIN METABOLOME-WIDE ASSOCIATION STUDY WITH ALZHEIMER DISEASE COHORT

Lecture Title:

C. Wang Washington University in St. Louis, Psychiatry, St Louis, United States of America

Aims: Although metabolome-wide association study (MWAS) has been performed in a wide range of tissue types, such as serum, plasma, urine, saliva, and cerebrospinal fluid (CSF), it has not been conducted on any brain tissue. Here we seek to expand our knowledge on the genetic influence of brain metabolism and on the contribution of metabolism abnormality to AD with the metabolomics approach. We aim to identify metabolite quantitative trait loci (metabQTLs) with the largest AD brain cohort available. Methods: We performed the first brain MWAS with 460 parietal cortex brains from the Knight-ADRC. 880 metabolites were measured by the powerful non-targeted Metabolon platform (HD4). European individuals were selected, and the analyses were adjusted for age at death, sex, genotype array methods, and genetic components. Results: We identified 68 locus-metabolite associations in 58 metabolites with genome-wide significance. Six associations were significant after study-wide Bonferroni correction. We are performing functional characterization and replication leveraging datasets from the ROSMAP brain cohort, WADRC CSF cohort, and other tissues’ cohorts. Amongst all, nine nonsynonymous coding variants are found. The strongest locus-metabolite association, found in N6-methyllysine in the discovery phase, was first identified in CSF MWAS (Panyard et al., biorxiv, 2020). This interesting locus includes brain eQTL for the PYROXD2 gene. Conclusions: This first brain metabolome-wide association study discovered the importance of genetic influence on brain metabolites levels. Following expectation, brain metabQTLs can be replicated in the CSF study, implicating that brain and CSF share genetic features in modulating metabolism.

P324 / #1673

Topic: Theme A: β-Amyloid Diseases / A6.h. Cell, Molecular and Systems Biology: Epigenetics, histone modification, DNA methylation

TIP60 HAT AND HDAC2 CROSSTALK IN ALZHEIMER'S DISEASE

Lecture Title:

E. Armour, J. Perhacs, F. Khoa, H. Zhang, F. Elefant Drexel University, Biology, Philadelphia, United States of America

Aims: Histone acetylation mediated synaptic gene activation is implicated in neuroprotection of AD linked cognitive deficits, yet contributing epigenetic mechanisms remain unclear. We previously identified an imbalance of Tip60 histone acetyltransferase (HAT) and histone deacetylase 2 (HDAC2) in the Drosophila AD brain that causes epigenetic repression of synaptic genes and cognitive deficits. We showed that Tip60 HAT action mediates activity dependent synaptic gene activation via its shuttling into the nucleus in response to extracellular cues and that Tip60 is excluded from hippocampal nuclei in AD patients. These results suggest that disruption of Tip60 nuclear shuttling is a contributing factor in AD neuropathology. Here, we aim to characterize Tip60 and HDAC2 subcellular localization in response to external cues in the Drosophila AD brain to elucidate epigenetic modulator nucleocytoplasmic shuttling in cognition and neuroprotection. Methods: Immunohistochemistry and confocal microscopy were performed to visualize Tip60 and HDAC2 localization in the mushroom body (MB) center for learning and memory and in the optic lobe (OL) regions of the Drosophila brain. Results: Tip60 was found predominantly in the cytoplasm of both the MB and OL regions in the adult Drosophila brain, while HDAC2 showed preferential nuclear localization. This pattern of subcellular location was identical during early brain development, suggesting that Tip60 and HDAC2 subcellular localization is tightly controlled throughout brain development. Conclusions: We are the first to explore Tip60 HAT/HDAC2 shuttling mechanisms in the AD brain. Our results can contribute to the development of histone acetylation mediated therapeutic targets for neurodegenerative diseases.

P325 / #1672


TIP60 HAT’S NOVEL RNA BINDING FUNCTION GIVES CUES FOR NEUROPROTECTION IN ALZHEIMER’S DISEASE

Lecture Title:

A. Bhatnagar, B.C. Karisetty, F. Elefant Drexel University, Biology, Philadelphia, United States of America

Aims: Our lab has previously implicated loss of Tip60 histone acetyltransferase (HAT) in early stages of Alzheimer’s disease (AD) that results in reduced histone acetylation, repression of critical synaptic plasticity genes and cognitive decline. Remarkably, Tip60 overexpression prevents these alterations, supporting a neuroprotective role for Tip60 in AD-linked neurodegeneration. Although the epigenetic roles of Tip60 are well studied, Tip60’s alternative cellular roles remain to be elucidated. Here, we aim to uncover a potential RNA binding function for Tip60 that may aid in Tip60-mediated neuroprotection. Methods: RNA-Immunoprecipitation (RIP) assay with anti-Tip60 antibody was used to immunoprecipitate RNA directly bound with Tip60 protein in the wild type Drosophila larval brains. Immunoprecipitated RNA and total RNA were sequenced via a genome-wide approach (RIP-Sequencing). Bioinformatic analyses were performed using DESeq2 software and FlyEnrichr enrichment analysis tool. RNA specifically enriched in the immunoprecipitate were identified as Tip60 targets. Results: We identified an array of Tip60 RNA targets that are enriched for critical neuronal processes and neurodegenerative diseases, including AD. The vast majority of these Tip60 RNA targets were found to be protein-coding RNAs. Tip60’s interaction with RNA is highly specific and reproducible between different biological samples. Remarkably, 77% of these Tip60 RNA targets overlap with Tip60’s chromatin gene targets, suggesting that Tip60 regulates identical sets of genes at both the chromatin and RNA level. Conclusions: We are first to uncover an RNA binding function for Tip60 HAT and propose a novel bi-level gene regulation that may underly Tip60’s neuroprotection in AD-linked neurodegeneration.

P326 / #1374


CHARACTERIZATION OF TWO RELATED CASES AFFECTED BY SLOW AND FAST DEMENTIA REVEALED EPIGENETICS IMPAIRMENTS

Lecture Title:

I. Palmieri1,2, S. Gagliardi1, M. Valente3, S. Zucca4, A. Davin5, V. Medici6, T.E. Poloni6,7, A. Guaita5,6, C. Cereda1 1IRCCS Mondino Foundation, Genomic And Post-genomic Unit, Pavia, Italy, 2University of Pavia, Department Of Molecular Medicine, MILANO, Italy, 3ASL Taranto, P.o.c Pathology Laboratory “s. Annunziata”, t, Italy, 4University of Pavia, Engenome, Pavia, Italy, 5Golgi-Cenci Foundation, Laboratory Of Neurobiology And Neurogenetic, Abbiategrasso, Italy, 6Golgi-Cenci Foundation, Department Of Neurology And Neuropathology, Abbiategrasso, Italy, 7ASP Golgi-Redaelli Geriatric Hospital, Department Of Rehabilitation, Abbiategrasso, Italy

Aims: Clinical diagnosis of patients affected by neurodegenerative diseases is often complex and requires neuropathological examinations. For this reason, it is necessary to strengthen molecular efforts to sustain clinical diagnosis. Here, the molecular characterization of a mother and a son diagnosed as slow and fast-dementia. Methods: NGS was done using a panel of over 6000 genes (Agilent Technologies). Total RNA from the hippocampus, parietal lobe, substantia nigra and basal ganglia was extracted and RNA-Seq analysis was run (Lexogen). Differentially Expressed Genes (DEGs) were identified via R_package_DESeq2. AlphaLisa was performed to investigate the dimethylation status of Histone 3 Lysine 9 (H3-K9) and the acetylation status of H3-K9. Results: No dementia-causative variants were found in the two patients, however, the missense mutation c.2990T>C (p.Leu997Pro) was found in the HDAC4 gene only in the son. DEGs analysis revealed a dysregulation in the son’s substantia nigra, with a 10-fold higher number of DEGs. Moreover, the substantia nigra of the son resulted hypomethylated and hyperacetylated respectively in the H3-K9 residue, while the mother resulted globally hypermethylated in all the brain areas analyzed at the H3-K9 and H3-K4 residues. Conclusions: Our investigation highlighted that beyond the common genetic background shared by the two individuals which may confer genetic susceptibility, the two different forms of dementia may have been influenced by a different epigenetic landscape, opening to an intriguing investigation on the role of the epigenetics in the modulation of neurodegenerative disorders.

P327 / #1172


SINGLE-CELL EPIGENOMIC LANDSCAPE IN ALZHEIMER’S DISEASE

Lecture Title:

S. Morabito1, E. Miyoshi2, N. Michael2, S. Shahin2, V. Swarup2,3 1University of California Irvine, Mathematical, Computational And Systems Biology (mcsb) Program, Irvine, United States of America, 2University of California Irvine, Department Of Neurobiology And Behavior, Irvine, United States of America, 3Biological Sciences 3, Room 3224, Irvine, United States of America

Aims: The gene-regulatory landscape of the brain is dynamic throughout life in health and disease. Dysregulation of the epigenome underpins many of the molecular phenotypes of neurodegenerative disorders such as Alzheimer’s Disease (AD). In this study, we used single-nucleus ATAC-seq (snATAC-seq) to profile the epigenome in all major cell types of the human brain in AD, and we single-nucleus RNA-seq (snRNA-seq) in the same biological samples, together painting a more complete portrait of the changes brought on by disease Methods: We performed snATAC-seq and snRNA-seq using the 10X Genomics Chromium platform, profiling the same biological samples of post-mortem human tissue in AD patients (n=12) and age-matched cognitively normal controls (n=8) for both sequencing assays. Results: Using our multi-omics approach, we profiled a total of 191,897 nuclei in late-stage AD using snATAC-seq and snRNA-seq (n=130,425 for snATAC-seq, n=61,472 for snRNA-seq). We linked genes to cis-regulatory elements by constructing cis co-accessibility networks (CCANs), identifying cell-type and disease-specific regulatory elements. Using LD score regression analysis, we found that accessible chromatin regions in two of the five microglia populations were specifically enriched for AD risk SNPs, indicating that certain sub-populations of microglia are more susceptible to the effects of inherited AD risk SNPs. Conclusions: A major contribution of this study are these cell-type specific regulatory maps between genes and CREs, highlighting regulatory systems in specific cell types. By identifying specific subpopulations of microglia that are enriched for AD risk SNPs. Linking these AD risk SNPs to our cis-regulatory maps, we have constructed detailed regulatory maps of these loci.

P328 / #889

Topic: Theme A: β-Amyloid Diseases / A6.i. Cell, Molecular and Systems Biology: Other

ACUTE GALACTOSE REMODELS ENERGY METABOLISM IN RATS HYPOTHALAMUS AFTER ORAL ADMINISTRATION

Lecture Title:

A. Knezovic, A. Vidovic, D. Zima, J. Homolak, A. Babic Perhoc, J. Osmanovic Barilar, M. Salkovic-Petrisic School of Medicine University of Zagreb, Pharmacology, Zagreb, Croatia

Aims: In our previous research, we showed that long-term oral galactose improves cognitive deficit in rat model of sporadic Alzheimer's disease. We aimed to explore the effect of single acute oral galactose administration on peripheral oxidative stress and hypothalamic metabolism in healthy rats. Methods: Three-month old male Wistar rats were treated orally with galactose (200mg/kg) and sacrificed 30, 60 or 120 minutes after. Glucose and galactose concentration, superoxide dismutase activity, low molecular weight thiols and lipid peroxidation were measured in plasma. Hypothalamic pyruvate dehydrogenase (PHD), cytochrome c oxidase (COXIV), cytochrome c (CytC) and caspase-3 (CASP3) levels were measured by Western blot. Results: With the exception of reduced lipid peroxidation end products 30 and 120 min after oral galactose treatment in comparison to controls, none of the other measured parameters in plasma were affected by galactose treatment. In hypothalamus both PHD and CASP3 expression was found decreased after oral galactose administration, while COXIV expression was found increased 60 and 120 minutes after oral galactose. There was no change in hypothalamic CytC expression. Conclusions: Acute oral galactose decreases peripheral lipid peroxidation and changes metabolic pattern in hypothalamus by promoting increased COXIV expression and preventing cell apoptosis that can have a substantial role in brain function. Supported by Croatian Science Foundation (IP-2018-01-8938). Research was co-financed by the Scientific Centre of Excellence for Basic, Clinical and Translational Neuroscience (project “Experimental and clinical research of hypoxic-ischemic damage in perinatal and adult brain”; GA KK01.1.1.01.0007 funded by the European Union through the European Regional Development Fund).

P329 / #1136

Topic: Theme A: β-Amyloid Diseases / A7.a. Animal Models: Transgenic rodents

NEUROPROTECTIVE EFFECTS OF NEURAL AND MESENQUIMAL STEM CELL HIPPOCAMPAL TRANSPLANTATION ON AMYLOID PLAQUES, NEUROGENESIS AND GLIAL CELLS OF APP/PS1 MICE

Lecture Title:

H. Campos1, D. Hashiguchi1, C. Gimenes1, D. Ribeiro2, S. Romariz1, B. Longo1 1Universidade Federal de São Paulo, Physiology, Laboratory Of Neurophysiology, São Paulo, Brazil, 2Universidade de São Paulo, Lab Of Neuroscience Chemistry Institute, São Paulo, Brazil

Aims: To investigate the effects of neuroprogenitor (NSC) and mesenchymal (MSC) stem cells on the clearance of amyloid plaques, microglial expression and neurogenesis when transplanted in the hippocampus of double-transgenic mice model (APP/PS1) for Alzheimer’s Disease. Methods: Adult male APP/PS1 and wild type (WT) mice were divided in the following groups: WT, AD, AD+NSC, AD+MSC (n=6 for each group). NSC from the telencephalon of eGFP/C57 embryos and bone marrow MSC from eGFP/C57 adult mice were transplanted in the hippocampus of AD+NSC and AD+MSC groups. Thirty days after transplantation, brains were removed, immunohistochemistry slides were prepared, incubated with GFP, NeuN, 6E10, DCX or Iba-1 antibodies, and hippocampus staining was quantified. Results: Transplanted NSC and MSC remained in the hippocampus and differentiated into neurons and glial cells. One-way ANOVA indicated significant differences for the number of amyloid plaques (6E10) showing a decrease in AD+NSC and AD+MSC transplanted groups in the hippocampus when compared with AD and WT groups (F (20) = 48,255, p=0.0001); the number of hippocampal microglial cells (IBA) increased in AD+NSC and AD+MSC groups compared with WT and AD groups (IBA) (F (20) = 21,721, p=0.0001); and hippocampal DCX cells increased in AD+NSC group compared with the AD group (t test (7) =2,663, p=0.003). Conclusions: Both cell types, NSC and MSC, showed neuroprotective effects when transplanted in the hippocampus of AD animals. NSC cells increased hippocampal neurogenesis (DCX), and NSC and MSC decreased the amyloid plaques and increased microglial cell numbers in the hippocampus, suggesting neuroprotective and anti-inflammatory effects.

P330 / #1369


INVESTIGATION OF ADVERSE EVENTS RELATED TO LONG-TERM POLYPHARMACY EXPOSURE IN APP NL-G-F KNOCK-IN MICE

Lecture Title:

F. Eroli1, K. Johnell2, M. Latorre Leal1, S. Hilmer3, P. Nilsson1, J. Wastesson4, A. Cedazo-Minguez1, S. Maioli1 1Karolinska Institutet, Nvs, Division Of Neurogeriatrics, Solna, Sweden, 2Karolinska Institutet, Department Of Medical Epidemiology And Biostatistics, Solna, Sweden, 3Kolling Institute, Royal North Shore Hosptial and University of Sydney, Clinical Pharmacology And Aged Care, Sidney, Australia, 4Karolinska Institutet, Aging Research Center, Solna, Sweden

Aims: Older people represent a growing portion of the population and this evolves in a strong increase of pharmaceutical drug use. Polypharmacy (concomitant use of ≥ 5 medications) is very common in people aged ≥65 years, with a prevalence of inappropriate drug use among persons with dementia, due to pathological brain changes and comorbidities. We recently performed a first polypharmacy mouse study (Eroli et al., Aging, 2020) reporting that polypharmacy treatment impairs exploration and synaptic function already in young wild-type mice. On that basis we decided to investigate the adverse outcomes of multiple-drug exposure in the APP NL-G-F knock-in (APP KI) mouse model of Alzheimer’s disease and whether this could speed up the disease progression at early stages. Methods: Four-month-old C57BL/6J and APP KI mice were fed for 8 weeks with a control or polypharmacy diet containing: metoprolol, paracetamol, simvastatin, citalopram, aspirin. After 4 weeks of treatment we evaluated anxiety-like behavior, motor and cognitive functions through a complete battery of behavioral tests. Results: The treatment was well-tolerated by the mice. Polypharmacy APP KI animals revealed a decrease in locomotor activity compared to WTs. Noteworthy, treated APP KI mice showed a significant reduction of freezing behavior during context test compared to APP KI control and WT groups. Conclusions: Our outcomes suggest that polypharmacy treatment worsens the performance of APP KI mice affecting explorative behavior and impairing contextual memory and learning. Thus, these results indicate that the multiple-medication regimen affects and might accelerate the early progression of cognitive impairment in Alzheimer’s disease.

P331 / #1171


RXR ACTIVATION PROMOTES REMYELINATION IN VERY OLD TRIPLE TRANSGENIC MOUSE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

D. Santos-Gil1, A. Sandoval-Hernández2, G. Arboleda-Bustos3 1Universidad Nacional de Colombia, Instituto De Genética, Bogotá, Colombia, 2Universidad Nacional de Colombia, Facultad De Ciencias - Departamento De Química, Bogotá, Colombia, 3Universidad Nacional de Colombia, Instituto De Genética, Bogota, Colombia

Aims: To evaluate the effect of bexarotene on protein expression changes associated with oligodendrocyte maturation in cortex and hippocampus in 3xTg-AD animals Methods: Triple transgenic animals for Alzheimer Disease (3xTg-AD) were treated with bexarotene (100mg / kg / day for 30 days). Subsequently, immunofluorescence analysis was performed by confocal microscopy of sections of brains treated with vehicle and bexarotene; and vehicle-treated controls (WT) for markers of oligodendrocyte maturation in the cortex and hippocampus regions. Results: Bexarotene increased oligodendrocyte precursor cells in very old 3xTg-AD mice. In addition, there is a greater number of immature oligodendrocytes in the hippocampus and cortex, which co-localized with mitotic markers and the recovery of myelinating mature cells Conclusions: Bexarotene, an RXR receptor agonist, is a pharmacological candidate for the treatment of Alzheimer's disease as it is involved in the proliferation and maturation of oligodendrocyte precursor cells until myelinating oligodendrocytes that achieve the remyelination process.

P332 / #1043


CHRONIC VERUBECESTAT TREATMENT SUPPRESSES AMYLOID ACCUMULATION IN AGED 5XFAD MICE BUT FAILS TO IMPROVE COGNITIVE OUTCOMES.

Lecture Title:

S. Sukoff Rizzo1, S. Quinney2, A. Masters3, C. Biesdorf2, K. Onos4, L. Haynes4, K. Keezer4, Z. Cope1, G. Little1, S.-P. Williams1, J. Meyer5, J. Peters5, L. Figueiredo5, S. Persohn5, A. Bedwell5, K. Eldridge5, R. Speedy5, M. Sasner4, G. Howell4, G. Carter4, A. Oblak6, B. Lamb6, P. Territo6 1University of Pittsburgh, Aging Institute, Pittsburgh, United States of America, 2Indiana University School of Medicine, Obstetrics & Gynecology, Indianapolis, United States of America, 3Indiana University School of Medicine, Clinical Pharmacology Analytical Core, Indianapolis, United States of America, 4The Jackson Laboratory, Research, Bar Harbor, United States of America, 5Indiana University School of Medicine, Radiology, Indianapolis, United States of America, 6Indiana University School of Medicine, Stark Neuroscience Research Institute, Indianapolis, United States of America

Aims: With a major goal for improving preclinical to clinical translation in predicting therapeutic potential of novel compounds for Alzheimer’s disease (AD), the Preclinical Testing Core (PTC) of MODEL-AD established a drug testing pipeline for unbiased assessments of therapeutic agents. This strategy includes: pharmacokinetics (PK); pharmacodynamics (PD) readouts of target engagement, as well as symptom- and disease-modifying activity. To validate the pipeline, verubecestat (VER), was tested in 5XFAD mice. Methods: PK analysis in 6 month aged male and female 5XFAD mice (10-100 mg/kg) was analyzed via LC/MS. PK/PD modeling determined dosing regimen for chronic treatments beginning at 3 months. PD endpoints (n=10-15/sex/treatment) were measured at 6-7mo, and included: 18-FDG PET/MR, 18F-AV45 PET/MR, CSF and plasma Aβ, and behavior. Results: PK analysis revealed rapid clearance of VER, thus all PD experiments were performed via chronic exposure to drug formulated in diet (10-100 mg/kg/day). Prophylactic treatment with VER produced dose-dependent reductions in amyloid deposition via 18F-AV45 PET and in CSF samples, as well as coat color changes similar to hair color changes observed in the clinic. Interestingly, VER did not improve deficits in glucose uptake in 5XFAD mice as measured by 18F-FDG PET nor improved cognitive function. Conclusions: Assessment of VER using the PTC strategy are consistent with clinical findings for improving amyloid deposition in the absence of improvement in cognition, as was observed in patients. Together these data support validation of the MODEL-AD PTC pipeline and highlight improvements in preclinical screening strategies that are sensitive to prediction of cognitive outcome measures.

P333 / #1119

Topic: Theme A: β-Amyloid Diseases / A7.e. Animal Models: Other

UNRAVELLING EARLY SIGNS OF NAVIGATIONAL IMPAIRMENT IN APPSWE/PS1DE9 MICE USING MORRIS WATER MAZE

Lecture Title:

S. Karunakaran Centre for Brain Research, Neuroscience, Bengaluru, India

Aims: Mild behavioral deficits, which are part of normal ageing, can potentially be early indicators of impending Alzheimer’s disease (AD). Using APPswe/PS1dE9 (APP/PS1) mouse model of AD, we systematically evaluated the mild behavioral deficits during the early stages of disease pathogenesis, using the Morris water maze (MWM) spatial learning paradigm. Methods: MWM experiments were based on established protocols using APP/PS1 and WT mice (n=18). We used an alternative method of analysis by focusing on the unsuccessful trials during water maze learning rather than the successful ones, in order to understand the nature of cognitive deficits more accurately. The data was further analysed based on the hierarchical recruitment of strategies used by mice during MWM learning. Results: APP/PS1 mice displayed higher number of unsuccessful trials during initial days of training unlike their wild type counterparts. However, with repeated trial and error learning, they reached levels comparable to WT during later days of training. Individual APP/PS1 mice also showed an enhanced preference for a non-cognitive strategy called circling, that led to abrupt learning transitions, and increased number of unsuccessful trials. Conclusions: Detailed behavioral analysis revealed that WT mice utilized qualitatively different search strategies as they became more proficient at the MWM task. However, APP/PS1 mice exhibited an overt dependence on non-spatial strategies, especially circling, which led to distorted learning transitions.

P334 / #1338


EFFECTS OF WESTERN DIET IN MODELING LATE-ONSET ALZHEIMER’S DISEASE IN LABORATORY MICE

Lecture Title:

K. Kotredes1, A. Oblak2, R. Pandey1, G. Carter1, M. Sasner1, B. Lamb2, G. Howell1 1The Jackson Laboratory, Model-ad, Bar Harbor, United States of America, 2Indiana University School of Medicine, Stark Neuroscience Research Institute, Indianapolis, United States of America

Aims: The Model Organism Development and Evaluation for Late-onset AD (MODEL-AD) Center aims to develop, validate, and distribute novel mouse models of late-onset Alzheimer’s disease (LOAD). In addition to genetic risk factors, the etiology of LOAD is strongly influenced by environmental risk factors. Therefore, we investigated the effects of western diet in mice expressing humanized risk alleles to better align the biology of mouse models with the features observed in human disease. Methods: Metabolic stress is common in LOAD patients due to sedentary lifestyle, obesity, and/or diabetes. Therefore, mice expressing risk alleles (MTHFR, PLCG2) or humanized APP on an APOEε4 and Trem2*R47H background were fed western diet (45% high fat/high sugar) and assessed over time for symptoms of disease. Regular frailty index measurements and biometric samples were collected. Upon endpoints, additional blood and brain tissue was sampled for transcriptomic, neuropathological, and histological analyses. Results: Mice fed western diet displayed significant increases in frailty, irrespective of age and genotype, and had increased glucose and cholesterol measures by eight months of age as well as increased liver enzymes by ten months. We observed that mouse models expressing combinations of LOAD risk variants and fed western diet exhibited strong human AD-relevant signatures with age. Further transcriptomic, neuropathological, and histological analyses are currently underway. Conclusions: Initial characterization of aged mice expressing genetic risk factors of LOAD indicates that a chronic, western diet increases susceptibility to disease. Inclusion of environmental risk factors should be a strong consideration in the modeling of LOAD and related dementias.

P335 / #909


BEHAVIORAL EFFECTS OF INTRACEREBROVENTRICULAR STREPTOZOTOCIN IN FEMALE RATS

Lecture Title:

M.F. Zappa Villar, J. López Hanotte, F. Peralta, P. Reggiani National Council of Scientific and Technical Research (CONICET), Biochemistry Research Institute Of La Plata "prof. Dr. Rodolfo R. Brenner" (inibiolp), National University Of La Plata (unlp), La Plata, Argentina

Aims: Background: Sporadic Alzheimer’s disease (SAD) is the main cause of dementia worldwide with no effective therapy. This multifactorial pathology is characterized by behavioral and cognitive impairment. Despite the fact that women show an increased risk of SAD, female animals are often excluded from basic research. In this line, an intracerebroventricular infusion of streptozotocin (icv-STZ) is commonly employed to generate SAD model in male animals. However, whether sex differences exist in the icv-STZ model remains unknown. Objective: Our objective was to study the female brain changes in rats with experimental SAD. Methods: Animals were submitted into 4 experimental groups (female Sprague Dawley rats, N=8): Sham, STZ, OVX and OVX+STZ. OVX and OVX+STZ groups were ovariectomized and, 14 days later, STZ and OVX+STZ groups were injected with 3 mg/kg icv-STZ. Body weight of all rats was registered along the experiment every week. During the last two weeks until the end of the study (day 30 post icv-STZ), we assessed several behavioural tests: species-typical behaviour (Marble Burying), exploratory and anxiety-related behaviour (Open Field), object recognition memory (Novel Object Recognition), spatial learning and memory (Barnes Maze), and depressive-like behaviour (Forced Swim Test). Results: Our preliminary results show that STZ affected behavioral performance differently in the SAD model depending on the ovarian status of female rats. Conclusions: Conclusion: Overall, we conclude that the potential impact of sex differences on brain function is a relevant issue and it should be considered in future SAD research.

P336 / #1806

Topic: Theme B: Taupathies / B1.a. Disease Mechanisms, Pathophysiology: Tau aggregation, phophorylation, acetylation & modifications

LIPID MEMBRANE MEDIATED MISFOLDING, SELF-ASSEMBLY, AND PHASE SEPARATION OF TAU

Lecture Title:

E. Chi1, J. Majewski2, E. Jones1, C. Vander Zanden3, J. Biernat4, E. Mandelkow4,5 1University of New Mexico, Department Of Chemical And Biological Engineering, ALBUQUERQUE, United States of America, 2National Science Foundation, Division Of Molecular And Cellular Biology, Alexandria, United States of America, 3University of Colorado, Colorado Springs, Department Of Chemistry And Biochemistry, Colorado Springs, United States of America, 4CAESAR, Research Center, Bonn, Germany, 5Deutsches Zentrum für Neurodegenerative Erkrankungen, (dzne) Bonn, Bonn, Germany

Aims: The aggregation of the intrisnically disrodered tau into highly ordered beta-sheet-rich fibrils is implicated in the pathogenesis of many neurodegenerative disorders. The mechanism of tau fibrillogenesis remains unresolved, particularly early events that trigger the misfolding and assembly of the otherwise soluble and stable tau. We investigated the role membrane plays in modulating the aggregation of three tau variants, the largest isoform hTau40, the truncated construct K18, and a hyperphosphorylation mutant hTau40/3Epi. Methods: A suite of biophysical techniques was used to characterize tau interactions with model lipid membranes. Notably, complementary synchrotron X-ray scattering techniques, grazing incidence X-ray diffraction (GIXD) and X-ray reflectivity (XR) were used to characteize in situ Angstrom-level tau and membrane structures. Results: Despite being charged and soluble, the tau proteins were also highly surface active and favorably interacted with anionic, but not zwitterionic, lipid monolayers at the air/water interface. Membrane binding induced tau structural compaction and macroscopic tau phase separation to produce a gelatinous protein-rich layer at the membrane surface. At the molecular level, tau assembled into oligomers composed of ~40 proteins misfolded in a beta-sheet conformation at the membrane surface, as detected by GIXD and XR. Concomitantly, membrane morphology and lipid packing became disrupted. Conclusions: Our findings support a general tau aggregation mechanism wherein tau’s inherent surface activity drives tau-membrane interactions, inducing the misfolding and self-assembly of the disordered protein into beta-sheet enriched oligomers that could subsequently seed tau fibril growth and deposition into diseased tissues.

P337 / #1492


A SINGLE MUTATION MAPT-P301L GENERATES MULTIPLE EVOLVING CONFORMERS INDUCING VARIOUS PHENOTYPES FRONTOTEMPORAL LOBAR DEGENERATION

Lecture Title:

N. Daude1, C. Kim2, S.-G. Kang1, E. Gomez Cardona3, O. Julien3, H. Wille1, J. Safar2, D. Westaway1 1Centre for Prions and Protein Folding Diseases, University of Alberta, Department Of Medicine, Edmonton, Canada, 2Case Western Reserve University, Department Of Neurology And Pathology, Cleveland, United States of America, 3University of Alberta, Biochemistry, Edmonton, Canada

Aims: MAPT mutations cause some types of frontotemporal dementia. Surprisingly, patients with the same and unique mutation can exhibit divergent clinical phenotypes; we sought to understand the origins of this heterogeneity. Methods: We analyzed brain samples from an Iberian kindred of FTLD-MAPT-P301L cases and also from inbred congenic lines of low-expresser 2N, 4R-P301l tau Tg mice. Fixed mouse brain samples were typed for i) the location of tau deposition, while sample homogenates were interrogated by ii) conformational stability assays, iii) a tau seeding assay using reporter cells and iv) mapping of protease resistant cores by mass spectrometry. MAPT expression patterns in the Tg mice was assessed by in situ hybridization with other transcripts profiled by nanostring analysis. Results: Despite uniform and widespread expression of transgene-encoded human tau mRNA and protein in young Tg mice, aged animals diverged to give three distinct signatures of misfolded tau and with a fourth signature in middle-aged mice indicating a mixed, prodromal state. Two signatures in aged mice were shared by P301L patients assigned clinical diagnoses of bvFTD and svPPA. Transcript profiling indicated an neuroinflammatory responses that varied between mice with rostral or caudal deposition of AT8-positive tau species. Conclusions: Heterogeneous yet overlapping signatures in inbred mice and a P301L kindred with a founder effect lead us to conclude that production of divergent conformers of tau is intrinsic to the pathogenesis of this simple, uni-allelic tauopathy. Thus, treatment regimens will need to consider heterogeneous target species in order to be effective.

P338 / #1127


MOLECULAR INTERACTIONS OF TAU WITH THE NUCLEAR PORE COMPLEX (NPC)

Lecture Title:

L. Diez1, T. Kara1, L. Kapinos-Schneider2, S. Hübschmann1, C. Wang3, F. Liu3, R. Lim2, S. Wegmann1 1Deutsches Zentrum für Neurodegenerative Erkrankungen (DZNE), Ag Wegmann, Berlin, Germany, 2Biozentrum, Cellular Nanomechanics And Nucleocytoplasmic Transport Control, Basel, Switzerland, 3Leibniz‐Forschungsinstitut for Molecular Pharmacology (FMP), Structural Interactomics, Berlin, Germany

Aims: In Alzheimer’s disease, Tau accumulating in the neuronal cytosol can interact with components of the nuclear pore complex (NPC) and thereby impair the nucleocytoplasmic transport (NCT) of proteins and nucleic acids. The aim of this study is to decipher the molecular mechanisms of Tau interactions with nucleoporins (Nups), and the human Tau:Nup interactome in Alzheimer’s disease and tauopathies. Methods: We are using proximity-dependent biotin identification (BioID) followed by Mass Spectrometry (MS) to identify interaction partners of Tau in the nuclear envelope of cell models. We are using co-immunoprecipitation and immunostaining for hit validation, and Surface Plasmon Resonance (SPR) and Bilayer interference (BLI) to determine the molecular mechanism of Tau binding to Nups. Results: After establishing the workflow for the BioID analysis of nuclear Tau interactions partners, we are currently analyzing samples obtained from neuronal cells by MS. Using SPR and BLI, we find that Tau binding to the FG-Nups Nup98 and Nup62 is Tau domain- and phosphorylation dependent. Unexpected, the binding of Tau to mutant Nup98 lacking phenylalanine (F), is comparable to wildtype Nup98, suggesting that Tau’s binding to FG-Nups seems somewhat independent of FG-repeats, crucial motifs for NPC diffusion barrier function. Conclusions: We present the first systematic investigation of Tau:NPC interactions aiming to understand the molecular mechanisms behind Tau-induced NCT impairment, which will be elementary for the development of potential therapeutic interventions. Dysfunctional NCT has also been reported in multiple other neurodegenerative diseases and our findings for Tau:Nup interactions will add to the overall goal to therapeutically target NCT deficits in neurodegeneration.

P339 / #1468


UBIQUITINATION OF TAU: IMPLICATIONS IN ALZHEIMER’S DISEASE

Lecture Title:

M. D'Onofrio1, F. Munari1, C. Barracchia1, F. Parolini1, R. Tira1, C. Franchin2, S. Capaldi1, G. Arrigoni2, M. Assfalg1 1University of Verona, Department Of Biotechnology, Verona, Italy, 2University of Padova, Department Of Biomedical Sciences, Padova, Italy

Aims: Accumulation of protein Tau in neurofibrillary tangles in the brain is a hallmark of Alzheimer’s disease (AD). In the pathological fibrillar state Tau is decorated with ubiquitin, suggesting that ubiquitination is involved in AD pathogenesis. Ubiquitination is a post-translational modification participating in many cellular pathways. It is a reversible modification, deubiquitinating enzymes (DUBs) can hydrolyze ubiquitin from the target protein, thus remodeling the ubiquitin signal. We aim to elucidate the effect of ubiquitination on the structure of Tau and on its aggregation pathway to fibrils. We also investigate the role of a DUB in remodeling the ubiquitination signal of Tau with possible effects on its clearance pathway. Methods: We propose an interdisciplinary methodology, combining molecular biology, enzymatic and semisynthetic reactions, biophysical and structural characterizations to obtain Tau ubiquitinated to describe the impact of the modification on aberrant protein aggregation. Results: Our results suggest that enzyme-derived ubiquitinated Tau is unable to convert into fibrils, while the semisynthetic conjugates exhibit diverse capability to form amyloid filaments depending on the modification site. We also obtained evidences of the interaction of a DUB with Tau which contribute novel insights into the structural aspects governing the molecular recognition. Conclusions: We obtained significant results necessary to decipher the effect of ubiquitin modification of Tau on its transition to toxic species. Moreover, the evidences of the interaction of Tau with a DUB could pave the way to gain crucial knowledge on the activity of these enzymes in retailoring the ubiquitination code therefore driving to the formation of pathological Tau forms.

P340 / #510


EVALUATION OF THE TAU PROTEIN AGGREGATION IN DRUG DEVELOPMENT AGAINST ALZHEIMER'S DISEASE

Lecture Title:

J. Janockova1, M. Bartolini2, O. Soukup3 1Faculty of Medicine, P. J. Safarik University in Kosice, Associated Tissue Bank, Košice, Slovak Republic, 2Alma Mater Studiorum, University of Bologna, Department Of Pharmacy And Biotechnology, Bologna, Italy, 3University Hospital Hradec Kralove, Biomedical Research Centre, Hradec Kralove, Czech Republic

Aims: The tau-hypothesis of Alzheimer´s disease process suggests that the disease cascade initiates tau microtubule associated protein abnormalities. It is a soluble protein, but its insoluble aggregates are produced during the formation of neurofibrillary tangles (inside nerve cell bodies) which disrupt the structure and function of neuron. Neurofibrillary tangles are aggregates most commonly known as a marker of AD and tau-cascade is one of the promising target for development of drug candidates for AD treatment. This work was focused on the study of tau protein (τ306-336 peptide) self-aggregation process in vitro followed by a detection of the influence of potential newly synthetized tacrine-phenothiazine inhibitors 1-5 in comparison with standard inhibitor doxycycline. We have also predicted their blood-brain barrier permeability using PAMPA assay and cytotoxicity profile. Methods: tau aggregation - ThT assay BBB permeability - PAMPA assay cytotoxicity - MTT assay Results:

Compound # Inhibition of τ (306-336) peptide aggregation (%) ± SEM *

Tacrine ~ 5

Phenothiazine < 5

Doxycycline 61.5 ± 0.8

Conclusions: - in the case of THA-PHE derivatives 1-5, which are different in the length of alkyl chain, we observed % of inhibition in range from 50 to 60%. The highest inhibitory potencies showed 2 and 3 with 3 and 4 carbons and resulting in being the same potent than standard doxycycline - the search for direct inhibitors of the τ aggregation process acquire deep attention as key mechanism and promising goal in the development of potential drugs against AD

P341 / #1009


COMBINATORIAL NATIVE MS AND LC-MS/MS APPROACH REVEALS HIGH INTRINSIC PHOSPHORYLATION OF HUMAN TAU BUT MINIMAL LEVELS OF OTHER KEY MODIFICATIONS.

Lecture Title:

F. Drepper1,2, J. Biernat3, S. Kaniyappan3,4, H. Meyer5,6, E.-M. Mandelkow7,8, B. Warscheid1,2, E. Mandelkow4,7,8 1Institute of Biology II, University of Freiburg, Biochemistry And Functional Proteomics, Freiburg, Germany, 2University of Freiburg, Signalling Research Centres Bioss And Cibss, Freiburg, Germany, 3Deutsches Zentrum für Neurodegenerative Erkrankungen, (dzne), Bonn, Germany, 4University of Bonn, Department Of Neurodegenerative Diseases And Geriatric Psychiatry, Bonn, Germany, 5Ruhr-University Bochum, Medical Proteome -center, Bochum, Germany, 6Leibniz-Institute für Analytical Sciences (ISAS),, Biomedical Research, Dortmund, Germany, 7CAESAR, Research Center, Bonn, Germany, 8Deutsches Zentrum für Neurodegenerative Erkrankungen, (dzne) Bonn, Bonn, Germany

Aims: Tau aggregation is considered one of the hallmarks of Alzheimer disease. Hyperphosphorylation of tau is thought to take place before aggregation, and it is often assumed that phosphorylation predisposes tau towards aggregation. However, the nature and extent of phosphorylation has remained ill-defined. Tau protein contains up to 85 potential phosphorylation sites, many of which can be phosphorylated by various kinases because the unfolded structure of tau makes them accessible. However, limitations in methods have led to conflicting results regarding the overall degree of phosphorylation of tau in cells. Methods: We employed a combination of native mass spectrometry and LC-MS to define the phosphorylation level of intact full-length tau purified from Sf9 cells. Results: Native mass spectrometry analysis of intact tau reveals tau in different phosphorylation states. The extent of phosphorylation is remarkably heterogeneous with up to ~20 phosphates per molecule and distributed over 51 sites (including all P-sites published so far and additional 18 P-sites). The medium phosphorylated fraction (Pm) showed overall occupancies centered at 8 Pi (± 5 Pi) with a bell-shaped distribution, the highly phosphorylated fraction (Ph) had 14 Pi (± 6 Pi). The distribution of sites was remarkably asymmetric (with 71% of all P-sites located in the C-terminal half of tau). Other known posttranslational modifications of tau were near or below our detection limit (e.g. acetylation, ubiquitination). Conclusions: Tau expressed in eukaryotic cells has high intrinsic extent of phosphorylation. Therefore, hyperphosphorylation of tau may not be specific for a disease state.

P342 / #1460


RETINAL TAUOPATHY IN MILD COGNITIVE IMPAIRMENT AND ALZHEIMER’S DISEASE PATIENTS

Lecture Title:

N. Mirzaei1, J. Sheyn1, D.-T. Fuchs1, A. Rentsendorj1, D. Lee2, M.-L. Selenica3, K. Black1, C. Miller4, Y. Koronyo1, M. Koronyo-Hamaoui5 1Cedars-Sinai Medical Center, Department Of Neurosurgery, Maxine Dunitz Neurosurgical Research Institute, Los Angeles, United States of America, 2University of Kentucky, Department Of Neuroscience, Sanders-brown Center On Aging/alzheimer’s Disease Research Center, Lexington, United States of America, 3University of Kentucky, Department Of Molecular & Cellular Biochemistry, Sanders-brown Center On Aging/alzheimer’s Disease Research Center, Lexington, United States of America, 4University of Southern California, Department Of Pathology Program In Neuroscience, Keck School Of Medicine, Los Angeles, United States of America, 5Cedars-Sinai Medical Center, Departments Of Neurosurgery & Biomedical Sciences, Los Angeles, United States of America

Aims: The field of Alzheimer’s disease (AD) retinopathy is gaining considerable momentum, with cumulative evidence supporting the existence of neuropathological hallmarks, traditionally associated with the brain disease, in the neurosensory retina. These include the proteinaceous accumulation of amyloid-β (Aβ) and hyperphosphorylated tau (pTau) assemblies, chronic gliosis, vascular deficits, pericyte loss, neurodegeneration, and functional abnormalities driven by retinal and visuo-cortical pathway irregularities. Relative to numerous findings of Aβ deposits in post-mortem retinae from mild cognitive impairment (MCI) and AD patients, retinal tauopathy has thus far remained understudied. Despite a small number of semi-quantitative mapping studies on the expression of total tau and pTau species in the human retina, it is still unclear whether paired helical filaments (PHFs) and neurofibrillary tangles (NFTs) exist in and/or affect the integrity of the AD retina. Here, we sought to address these unknowns. Methods: Immunohistochemistry was used to visualize and quantify the spatial burden of pTau species, PHFs, and citrullinated tau in post-mortem retinae from MCI and AD patients, relative to cognitively normal controls. Levels of pre- and post-synaptic markers as well as the thickness of retinal layers were also measured. Results: Elevated levels of various pTau species were found in a geometric-dependent manner in the retinae of MCI and AD patients. Increased burden of PHFs and citrullinated tau were also identified in retinal layers with higher retinal synaptic loss and neuronal atrophy in the same patients. Conclusions: Our study provides further evidence for the presence of AD-specific pathological changes in the retina, similar to those observed in the brain.

P343 / #794


INTRON RETENTION CAUSES ALTERED PROTEIN BEHAVIOUR IN CELLULAR MODEL OF TAUOPATHY

Lecture Title:

C.-T. Ong, W.-Q. Lin, Z.-K. Ngian Temasek Life Sciences Lab, Developmental Biology, Singapore, Singapore

Aims: Aging is the major risk factor for many neurodegenerative diseases such as Alzheimer's disease (AD). Accumulation of amyloid plaques and formation of Tau tangles are two hallmarks of AD pathology. While epigenetic alterations and genetic variations at several AD-susceptibility loci have been attributed to late-onset AD, it is not fully clear how these changes may mould the proteome, and cause the transition of healthy neurons into pathological state. We aim to better understand these mechanisms. Methods: Through analyzing transcriptome from large human cohorts, we (Adusumalli et al 2019) and others have demonstrated that aberrant intron retention is one of the signatures associated with aging and diverse types of tauopathies (including AD). To follow-up on these genome-wide studies, we validated specific intron retention events identified from RNA-sequencing datasets in neural stem cells derived from control, familial AD and sporadic AD patients. We then cloned the specific retained introns into to the cDNA of candidate gene to test how they may affect the biochemical properties of the translated protein in different cell-types. Results: Interestingly, the retained introns, which encode 10-20 amino acids followed by prematured termination codon, change the behavior of the candidate protein drastically in vivo. Translated candidate protein containing the retained intron sequence dimerizes to form high molecular weight species. The protein is also enriched in the Sarkosyl insoluble fraction and have reduced association with microtubules in mature human neurons. Conclusions: Our results indicate that intron retention might cause the protein to form insoluble aggregates, which in turn contributes to tauopathy in human neurons.

P344 / #458


ISOFORM-SPECIFIC KNOCKOUT OF FYNT TYROSINE KINASE ATTENUATED TAUOPATHY, NEUROINFLAMMATION AND SYNAPTIC DEGENERATION IN A TRANSGENIC MOUSE MODEL OF TAUOPATHY

Lecture Title:

M. Tan1, C. Lee1, C. Low1, F. Lim1, M. Lai2 1Singapore General Hospital, Department Of Clinical Translational Research, Singapore, Singapore, 2National University of Singapore, Yong Loo Lin School of Medicine, Department Of Pharmacology, Singapore, Singapore

Aims: Fyn tyrosine kinase has been proposed to be a potential therapeutic target for AD and tauopathies. We reported that FynT but not FynB isoform was selectively upregulated in Alzheimer's disease (AD) and closely associated with neurofibrillary tangle formation and reactive astrogliosis. Furthermore, significant upregulation of FynT was also detected in aged P301S tau transgenic mice (PS19) and positively correlated with tau pathology and neuroinflammation. In this study, we aim to study the critical role of FynT in tau pathology and neuroinflammation using PS19 crossing with FynT knockout (FynT-KO) mice. Methods: WT, FynT-KO, PS19 and double transgenic (DTg) mice (all males) were subjected to contextual and cued fear conditioning test, followed by brain harvesting to assess tau pathology, neuroinflammation and synaptic markers using Cisbio HTRF assay kits, real time RT-PCR, immunoblots and immunostaining. Results: Tau phosphorylation and aggregation, along with glial activation were significantly increased in PS19 when compared with WT littermate controls. Furthermore, synaptic loss and concomitant fear memory impairment were also detected in PS19. Most importantly, isoform-specific knockout of FynT in DTg mice demonstrated significant attenuation of tau pathology and neuroinflammation, as well as restoration of synaptic loss and fear memory deficit when compared with PS19, suggesting that FynT can modulate tau pathology and neuroinflammation. Conclusions: Our findings provide evidence for isoform-specific role of FynT being a regulator of tau pathology and neuroinflammation during the progression of tauopathies. Rational pharmacotherapeutic interventions based on isoform-specific drug targets may be developed to potentially improve drug efficacy and reduce off-target effects.

P345 / #408


TAU MULTIMERISATION AS ONE OF THE EARLIEST EVENTS IN THE DEVELOPMENT OF ALZHEIMER’S PATHOLOGY PRECEDING TAU PHOSPHORYLATION AND MISFOLDING

Lecture Title:

E. Velentza Almpani1, N. Bengoa-Vergniory2, A.M. Silva3, C. Scott4, M. Sastre1, R. Wade-Martins2, J. Alegre-Abarrategui1 1Imperial College London, Department Of Brain Sciences, London, United Kingdom, 2University of Oxford, Department Of Physiology, Anatomy And Genetics, Oxford, United Kingdom, 3Imperial College London, Department Of Physiology, Anatomy And Genetics, London, United Kingdom, 4University of Oxford, Nuffield Department Of Clinical Neurosciences, Oxford, United Kingdom

Aims: Our main aim is to understand the pathophysiological role of the early protein multimers in AD using a novel method for the direct detection of tau-tau interaction in animal and post-mortem human tissue. Our main objectives are: 1) Proof the in vitro specificity of tau-PLA for tau multimers but no monomers. 2) Demonstrate the in situ specificity of the tau-PLA in animal and post-mortem human brain. 3) Comparison of tau-PLA with standard histopathological markers for the detection of early tau pathology. Methods: Recombinant protein assembly, Immunohistochemistry, Immunofluorescence, Proximity ligation assay, Imaging Results: We developed a novel tau-proximity ligation assay (tau-PLA) that allows the direct visualization of tau-tau interactions in situ, and exclusively recognized tau multimers but not monomers. No tau-PLA signal was detected in MAPT K/O mouse brains, while it appeared to be significant strong in P301S transgenic mice and AD brain. Two groups of structures were detected, a previously unreported widespread small-sized diffuse pathology and large, neurofibrillary-like lesions. Tau-PLA-labelled diffuse pathology appeared from the earliest Braak stages, mostly unaccompanied by other tau markers. Conclusions: Tau-PLA is a novel method that allows for the first time the direct visualization of tau-tau interaction both in vitro and in situ with high specificity. We found that tau multimerization precedes tau hyperphosphorylation and conformational changes, being the earliest detectable molecular event of AD tau pathology. Our findings allow the study of early tau pathology, with potential implications in early AD diagnosis and the design of therapeutic strategies.

P346 / #1802


DIFFERENCES OF TAU LIQUID-DENSE CONDENSATES

Lecture Title:

S. Wegmann1, J. Hochmair1, C. Exner2, E. Mandelkow3,4, C. Betzel2 1German Center for Neurodegenerative Diseases, Dzne Berlin, Berlin, Germany, 2University Hamburg / DESY, Structural Biology Of Infection And Inflammation, Hamburg, Germany, 3University of Bonn, Department Of Neurodegenerative Diseases And Geriatric Psychiatry, Bonn, Germany, 4Deutsches Zentrum für Neurodegenerative Erkrankungen, (dzne) Bonn, Bonn, Germany

Aims: Tau can condensate into liquid condensed protein phases in vitro. Liquid-liquid phase separation of Tau at physiological concentrations can be triggered by coacervation with polyanionic molecules like RNA and heparin, or through addition of molecular crowding agents such as polyethylene glycol or dextran. We determined commonalities and differences between crowding induced condensates and coacervates of Tau in vitro. We compared formation and size of Tau condensates formed with RNA or PEG and described their physicochemical properties and the molecular interactions governing the interactions inside Tau condensates. We also described their potency to form oligomeric Tau species able to initiate Tau agrgegation in HEK sensor cells. Methods: We use time-resolved dynamic light scattering and turbidity measurements and microscopy and FRAP to determine size, growth and material properties of Tau condensates. Oligomerization states of Tau in condensates are determined by immunoblotting (SDS-PAGE, Native PAGE, SDD-AGE, dot blot), and the potential to seed cellular Tau aggregtion is determined in HEK cells. Results: The formation kinetics of Tau condensates differs between poyanion and PEG induced LLPS but final "droplet" sizes are similar. Differences in the interior of Tau condensates give rise to different physicochemical behavior and transition of condensates into oligomeric Tau assemblies. Both polyanion and crowding induced Tau condensates produce oligomeric seeding competent Tau species in rather short time (hours). Conclusions: Liquid tau condensates show different hardening and molecular interactions, depending on the mechanism of formation (coacervation vs condensation with crowders). Both kinds of Tau LLPS may be involved in physiological and pathological liquid Tau phases.

P347 / #1758


ROLE OF HYPERPHOSPHORYLATION IN THE SPREAD OF TAU PATHOLOGY

Lecture Title:

S. Zafar, K. Waqar SMME, National University of Sciences and Technology (NUST), Islamabad, Pakistan, Biomedical Engineering And Sciences Department, Islamabad, Pakistan

Aims: Hyper-phosphorylated tau results in neurofibrillary tangles which is a hallmark of Tau pathology in Alzheimer’s disease and other related tauopathies. in the current study we aim to induce and propagated Tau linked post translation modification, experimentally to understand the elusive nature of the prion like transmission character of Tau. Methods: We investigated the tau phosphorylation sites by using LC- MS/MS analysis and identified prion-like activity in vitro and in cultured cells, using the wild-type human 2N4R tau and mutated tau constructs developed by site directed mutagenesis. The level and the sites of hyperphosphorylation was studied by using 32 P-incorporation parallel reaction and Western blots developed with phosphorylation-dependent and site-specific anti-tau antibodies. For investigating the in vivo seeding activity of hyperphosphorylated recombinant tau we used the representative hyperphosphorylated taus and stereotaxically injected each of them into the hippocampus of transgenic mice overexpressing human tau and characterize the induced tau pathology by western blot analysis and immunohistological/histological staining. Results: In line with the findings from the human brain, our in vitro study showed that the tau phosphorylation sites identified by LC-MS/MS were modified with altered prion interactome. The detailed overlay of globe wide proteome identified from hippocampus of transgenic mice/overexpressing human tau showed diferential proteome signature with altered seeding activity of oligomeric hyperphosphorylated taus in cultured cells. Conclusions: In conclusions, despite these differences, the level and the sites of hyperphosphorylation modified and showed altered seeding activity of oligomeric hyperphosphorylated taus in cultured cells.

P348 / #832

Topic: Theme B: Taupathies / B1.b. Disease Mechanisms, Pathophysiology: Cell to cell transmission, spreading of pathology, prion-like

TAU SEEDING ASSAY BASED ON TAURD-GFP FRET PAIRS DOES NOT REPRESENT TEMPLATED ASSEMBLY OF PHFS

Lecture Title:

S. Kaniyappan1, K. Tepper2, J. Biernat2, R.R. Chandupatla2, S. Hübschmann2, S. Irsen3, S. Bicher3, C. Klatt3, E.-M. Mandelkow2, E. Mandelkow4 1University of Bonn, Department Of Neurodegenerative Diseases And Geriatric Psychiatry, Bonn, Germany, 2Deutsches Zentrum für Neurodegenerative Erkrankungen (DZNE), Deutsches Zentrum Für Neurodegenerative Erkrankungen (dzne), Bonn, Germany, 3Center of Advanced European Studies and Research, Center Of Advanced European Studies And Research, Bonn, Germany, 4Deutsches Zentrum für Neurodegenerative Erkrankungen (DZNE), Deutsches Zentrum Für Neurodegenerative Erkrankungen (dzne) Bonne, Bonn, Germany

Aims: The progressive appearance of aggregated tau is a hallmark of Alzheimer disease. It has been proposed that spreading of pathology is caused by transcellular spreading of misfolded tau and templated aggregation of tau fibers (also known as prion-like spreading). In cell culture, the process is monitored by a FRET assay where the cells expressing tau repeat domain (TauRD, with pro-aggregant mutation, e.g. ΔK280 or P301L, ~13.5 kDa) fused to GFP-based FRET pairs (YFP or CFP, ~27kDa). Considering that TauRD and its spacing in Alzheimer-like fibrils (β-sheets, 0.47nm) is much smaller than the size of GFP tag (3-4nm), this raises the question of steric compatibility. Methods: We investigated the influence of GFP linked N- or C-terminally to TauRDΔK or TauFL (full-length tau) on aggregation in-vitro. Using biophysical (light-scattering, fluorescence-anisotropy), atomic force microscopy (AFM) and scanning-transmission electron microscopy (STEM), we characterized assembly states of tau in presence and absence of GFP tags. Results: The aggregation of TauRDΔK-GFP fibers was severely inhibited by GFP tags. Some rare short fiber-like particles were observed but had a very different subunit packing, as judged by AFM and STEM, indicating that the subunit packing is distinct from that of PHF-like fibers. By contrast, aggregation of full-length TauFL-GFP was compatible with PHF-like subunit packing. Conclusions: Both kinetic and structural observations indicate that the tau repeat domain labeled with GFP cannot form fibers with Alzheimer-like packing. This is incompatible with a model of spreading tau pathology by cell-to-cell spreading of tau protein and templated fiber assembly.

P349 / #1323


DEVELOPMENT OF AN AAV-BASED MODEL OF TAUOPATHY IN THE MOUSE VISUAL PATHWAY TO STUDY THE ROLE OF MICROGLIA IN TAU PROTEIN PROPAGATION

Lecture Title:

C. Duwat, A. Vautheny, G. Auregan, C. Josephine, M.-C. Gaillard, C. Jan, A.-S. Hérard, E. Brouillet, P. Hantraye, G. Bovento, K. Cambon, A. Bemelmans Université Paris-Saclay, CEA, CNRS, Laboratoire des Maladies Neurodégénératives : mécanismes, thérapies, imagerie, Molecular Imaging Research Center, Fontenay-aux-Roses, France

Aims: During AD, tauopathy follows a precise spatio-temporal progression, in correlation with symptoms severity. This progression may be due to the spread of pathological forms of Tau between connected neurons and in which microglia and neuroinflammation could play a role. Our main objective is therefore to develop relevant in vivo models to study the molecular and cellular determinants of Tau propagation. Methods: We developed AAV vectors overexpressing human Tau (hTau) in order to trigger a focal tauopathy in transduced neurons after in vivo administration. We selected the visual system, a neural circuit with strictly separated afferent and efferent neurons. To do so, we transduced retinal ganglion cells (RGCs) following intravitreal injection of serotype 2 AAV. We first characterized tauopathy, neuroinflammation and neurodegeneration in wild-type mice. We then performed similar analyses in Trem2-/- mice (an innate immune receptor expressed by microglia and whose polymorphisms are associated to AD) to challenge the role of microglial activation in our experimental paradigm. Results: One month after injection, we found hTau in RGCs, optic nerve and superior colliculi but not in efferent neurons projecting to the primary visual cortex. Morphological analysis highlighted a moderate but significant toxicity induced by hTau in RGCs from wild-type mice. Preliminary results indicated that TREM2 deficiency reduced microglial activation and protected RGCs from hTau-induced toxicity. Conclusions: Results obtained in Trem2-/- animals suggest that tauopathy-triggered neuronal cell death does not rely solely on cell-intrinsic mechanisms. In this context, reducing TREM2 activity appears as a relevant therapeutic target in AD.

P350 / #256


HEPARAN SULFATES 3-O SULFATION MEDIATES CELLULAR INTERNALIZATION OF TAU SEEDS IN HCT-116 CELLS

Lecture Title:

A. Ferreira1,2, D. Moechars2, N. Callewaert1, L. De Muynck2 1VIB-UGent, Center For Medical Biotechnology, Gent, Belgium, 2Janssen Pharmaceutica, Neuroscience Ta, Beerse, Belgium

Aims: Heparan sulfate proteoglycans (HSPGs) have been proposed to mediate the propagation of Tau pathology by internalizing Tau seeds, being the sulfation of HSPGs critical for the uptake. The heparan sulfate (HS) sulfation pattern is highly complex and determines the HSPG function. To date, little is known about the HS structural determinants required for Tau seeds uptake. Here we studied the role of the 3-O-sulfation in the uptake of Tau seeds. Methods: An HS3ST1 KO cell line was generated and 3-O-sulfation at the cell surface was measured using a fluorescence-based method. The uptake of labeled Tau seeds was measured using live-cell confocal microscopy, followed by a rescue assay where HS3ST1 was overexpressed in the KO cell line. Lastly, synthetic (3S-)HS was used to outcompete the HS binding sites of the Tau seeds, and antithrombin III (ATIII) was used to mask cellular 3S-HS prior to the uptake. Results: An HS3ST1 KO cell line that doesn’t express any HS3ST isoforms was successfully generated and shown to be devoided of 3-O-sulfation. This cell line showed reduced uptake of Tau seeds, compared to the WT cell line, and this phenotype could be rescued by overexpressing HS3ST1 in the KO cells. Furthermore, either targeting the Tau seeds with 3S-HS structures or the cell surface 3S-HS with ATIII led to the reduction of Tau seeds uptake by the WT cells. Conclusions: These results suggest that 3S-HS mediate the cellular uptake of Tau seeds, highlighting a potential novel protein class to target Tau pathogenesis.

P351 / #648


THE ROLE OF ECTODERMAL-NEURAL CORTEX 1 IN EARLY ALZHEIMER’S DISEASE

Lecture Title:

H. Fu1, L. Li1, S. Chen1, J. Liang1, L. Venkataraman1, Y.-K. Jung2, M. Diamond3 1The Ohio State University, Neuroscience, Columbus, United States of America, 2Seoul National University, School Of Biological Sciences, Seoul, Korea, Republic of, 3University of Texas Southwestern Medical Center, Center For Alzheimer's And Neurodegenerative Diseases, Dallas, United States of America

Aims: We recently have identified novel subproteome gene signatures in excitatory neurons that may serve as master regulators of selective neuronal and regional vulnerability to tau pathology in early Alzheimer’s disease (AD). The ectodermal-neural cortex 1 (ENC1) is one of such master regulators. This study aims to further investigate the role of ENC1 in early AD, focusing on tau pathology. Methods: The immunostaining and Western blotting were used to measure the localization and protein expression level of ENC1, total tau and p-tau in AD-like tau mouse models and human AD cases at different Braak stages. The DuoLink proximity ligation assay was used to measure the interaction between ENC1 and total tau or p-tau in human non-AD and AD cases. The live imaging was used to measure the DS9 tau seeding in the presence of ENC1 overexpression or knockdown. Results: The nuclear ENC1 levels greatly decreased, while cytoplasmic ENC1 levels actually increased within neurons that accumulated pathological tau species in human entorhinal cortex. Furthermore, the protein interaction between cytoplasmic ENC1 and tau correlates directly with levels of pathological tau. We also found that overexpression of ENC1 significantly increased DS9 tau seeding activity in SH-SY5Y cells harboring P301S mutant tau. In contrast, knockdown of ENC1 decreased tau seeding activity in the SH-SY5Y seeding model. Conclusions: These results suggest that the interaction between ENC1 and pathological tau may aggravate the propagation of tau pathology, thereby contributing to the selective neuronal vulnerability to tau pathology in early AD.

P352 / #992


WIDESPREAD TAU SEED NETWORKS IN ALZHEIMER AND DIFFUSE LEWY BODY DISEASE BRAINS

Lecture Title:

M. Manca1, M. Huntley1, X. Zhu1, M. Cohen2, A. Kraus1 1Case Western Reserve University, School of Medicine, Department Of Pathology, Division Of Experimental Pathology, Cleveland, United States of America, 2University Hospitals Cleveland Medical Center, Departments Of Pathology And Neurology, Cleveland, United States of America

Aims: Prion-like propagation of misfolded tau is implicated as a causal mechanism of Alzheimer disease (AD). Synucleinopathies (with αSynuclein (αSyn) as the primary pathological protein) such as Diffuse Lewy body disease (DLBD), may have comorbid tau pathologies. Currently unknown is how the co-occurrence and distributions of tau and αSyn seeds correlate with disease manifestations. This study evaluates how the distribution of tau seeds, i.e. self-propagating misfolded tau conformers, in AD and DLBD correlate with clinical and pathological outcomes. Methods: We use ultrasensitive seed amplification assays [real-time quaking-induced conversion (RT-QuIC)] to quantitate tau seeds in brain regions involved at different disease stages in AD, DLBD, and AD/DLBD co-pathologies (e.g. primary sensory cortex, association cortex, limbic cortex, entorhinal cortex, amygdala, and hippocampus). Structural/biophysical properties of tau seeds were evaluated using gel and immunoblot analysis. Results: Tau seeds are detectable in all regions assessed in AD cases with multi-log quantitative distinctions among regions. Comparatively low tau seed levels were observed even in regions without histologically visible tau deposits (e.g. cerebellum). Widespread tau seed networks also occurred in DLBD with AD-related pathologies, however, distributions differed among DLBD variants and when compared to AD cases without αSyn pathology. Tau seeds in multiple regions are protease resistant, and can be immunoprecipitated with antibodies targeting phosphorylated tau. Conclusions: Tau seeds are widespread in AD and DLBD brains, providing evidence that tau seeds occur in primary synucleinopathies. RT-QuIC assays can elucidate co-pathologies and help define regional seed distributions with a billion-fold dynamic working range, providing a quantitative methodology for clinicopathological correlative studies.

P354 / #1626


TAU ASSEMBLIES DO NOT BEHAVE LIKE INDEPENDENTLY ACTING PRION-LIKE PARTICLES IN MOUSE NEURAL TISSUE

Lecture Title:

L. Miller1, A. Mukadam1, C. Durrant2, M. Vaysburd3, T. Katsinelos1, B. Tuck1, S. Sanford1, O. Sheppard4, C. Knox3, L. James3, M. Coleman4, W. Mcewan1 1University of Cambridge, Clinical Neurosciences, Cambridge, United Kingdom, 2University of Edinburgh, Centre For Discovery Brain Sciences, Edinburgh, United Kingdom, 3University of Cambridge, Mrc Laboratory Of Molecular Biology, Cambridge, United Kingdom, 4University of Cambridge, Brain Repair Centre, Cambridge, United Kingdom

Aims: Assemblies of the protein tau characterise multiple neurodegenerative diseases, however the mechanism by which tau aggregates appear throughout the brain remains debated. Specifically, the contributions of cell autonomous vs seeded aggregation are not fully understood. Intracranial injection of mice, typically with micromolar concentrations of tau, have shown that seeded aggregation can be induced. However, the dose-response of seeding, and thus the relevance of injection experiments using supraphysiological levels of tau, remains unknown. Methods: Here we develop a tau seeding assay using organotypic hippocampal slice cultures (OHSCs) to enable control of tau assembly concentration, and exposure time, not possible in intracranial injection experiments. Results: Whilst untreated slices displayed no overt signs of pathology, exposure to tau assemblies could result in the formation of intraneuronal, hyperphosphorylated tau structures. Importantly, seeding ability of tau assemblies in OHSCs did not titrate in a one-hit manner that would be expected of independently acting particles (Figure 1). Rather, we only observed seeded aggregation when supraphysiological concentrations of tau assemblies were supplied, at around 100 nM.

Conclusions: Our results suggest that high-dose intracranial challenge experiments cannot be extrapolated down to infer the seeding behaviour of tau assemblies at physiological concentrations. Human CSF concentrations of tau lie in the picomolar range. Our data suggests that neural tissue is able to resist seeding at these concentrations, or that other factors not captured here act to lower the threshold for seeded aggregation in the degenerating brain.

P355 / #1625


ENDOSOMAL ESCAPE IS ESSENTIAL AND RATE-LIMITING IN THE SEEDED AGGREGATION OF TAU

Lecture Title:

B. Tuck, T. Katsinelos, A. Mukadam, W. Mcewan University of Cambridge, Department Of Clinical Neurosciences Irb 406, Cambridge, United Kingdom

Aims: Seeded aggregation potentially underlies the pathological accumulation of tau in neurodegenerative disease. Whilst the uptake of tau species to endosomes is well-documented, the subsequent escape to the cytoplasm is poorly understood. We therefore aimed to identify the molecular mechanism of tau entry and determine its relationship to seeded aggregation. Methods: We established sensitive complementation-based assays to analyse the cytoplasmic entry of tau assemblies in immortalised cells and primary neurons. By manipulating the endo-lysosomal network at various stages, we interrogated the cellular machinery that controls cytosolic entry. Results: We observed that extracellular tau monomers and assemblies were rapidly able to access the cytoplasm, saturating at around 6 h (Fig 1A). We found that tau escapes primarily from RAB7-positive endosomal compartments, and entry can be reduced by inhibition of tau uptake or inhibition of clathrin and dynamin (Fig 1B). In contrast, macropinocytosis was not found to be involved. Importantly, the amount of entry determines the level of seeded aggregation observed.

Figure 1: Quantification of tau entry in live cells. A) 50nM Tau entry over an 8 hour time course. B) Quantification of 50nM tau entry after 1 hour after inhibition of macropinocytosis (amiloride, LY294002), inhibition of tau uptake with heparin or inhibition of clathrin (PitStop) or dynamin (Dyngo). Normalised to no drug. Conclusions: Our study identifies the penetration of endosomal membranes as a critical upstream step in the seeded aggregation of tau. The work demonstrates that contact between tau assemblies and native tau pools is rate-limiting and essential to seeded aggregation.

P356 / #1366

Topic: Theme B: Taupathies / B1.c. Disease Mechanisms, Pathophysiology: Inflammation

IL-1B AS A KEY CYTOKINE TO REGULATE INFILTRATION OF MONOCYTES/MACROPHAGES IN THE BRAIN IN SYSTEMIC INFLAMMATION

Lecture Title:

R.C.-C. Chang1,2, X.K. Chen2, A.C.-H. Ma3 1The University of Hong Kong, State Key Laboratory Of Brain And Cognitive Sciences, Pok Fu Lam, Hong Kong PRC, 2The University of Hong Kong, Laboratory Of Neurodegenerative Diseases, Lks Faculty Of Medicine, Pok Fu Lam, Hong Kong, Hong Kong PRC, 3Hong Kong Polytechnic University, Department Of Health Technology And Informatics, Kowloon, Hong Kong PRC

Aims: Systemic inflammation can be a risk factor for developing cognitive dysfunctions and even Alzheimer’s disease. It is still unclear how systemic monocytes/macrophages respond after sterile inflammation. We aim to use zebrafish and tail amputation for investigating which factor can affect the influence of systemic inflammation in the brain. Methods: Zebrafish was used as an experimental model. Tg(coro1a:DsRed) transgenic fish was used to monitor microglia and macrophages. Tg(mpx:mCherry) was used for monitoring neutrophils, Tg(mepg1:GFP) was used for monitoring macrophages. We have used TALEN method to make knockout or mutation of IL-1β in zebrafish. Quantitative PCR and immunohistochemical staining were used for examination. Acridine orange and TUNEL were employed for labeling apoptotic cells. Tail amputation was performed at day 3 of embryos under anesthetization with tricaine. Results: Tail amputation recruited accumulation of monocytes/macrophages into the wound site in zebrafish tail. Surprisingly, an increased number of macrophages and microglia were also found in the brain, which was tissue-specific as no other organs showed a similar phenomenon. The expression of mRNA for different cytokines was also examined. Among which, IL-1β was significantly increased. Therefore, we prepared IL-1β knockout and mutated zebrafish. After amputation, the basal number of macrophages in the brain did not show any significant increase. We found an increased number of apoptotic cells in the brain after wound injury. IL-1β had a key role in regulating the migration of monocytes/macrophages. Conclusions: Our study demonstrated that IL-1β is a key player to regulate migration of monocytes/macrophages in the brain.

P357 / #1260


EXPERIMENTAL PERIODONTITIS INDUCED BY P. GINGIVALIS EXACERBATED COGNITIVE IMPAIRMENT AND TAU PATHOLOGY IN 3XTG AD MICE

Lecture Title:

R. Wang1, W.-K. Leung2, T. Goto3, Y.-S. Ho4, R.C.-C. Chang1,5 1The University of Hong Kong, Laboratory Of Neurodegenerative Diseases, Lks Faculty Of Medicine, Pok Fu Lam, Hong Kong, Hong Kong PRC, 2The University of Hong Kong, Faculty Of Medicine, Hong Kong, Hong Kong PRC, 3Kagoshima University, Division Of Oral Anatomy And Histology, Graduate School Of Medical And Dental Sciences, Kagoshima, Japan, 4Hong Kong Polytechnic University, School Of Nursing, Hong Kong, Hong Kong PRC, 5The University of Hong Kong, State Key Laboratory Of Brain And Cognitive Sciences, Pok Fu Lam, Hong Kong PRC

Aims: Periodontitis is a common health problem in elderly, which can induce chronic systemic inflammation. Our research team has been studying how systemic inflammation affects the brain immune responses, which can then affect progression of Alzheimer’s disease (AD). Therefore, our aim is to apply experimental model of periodontitis in AD mice and examine cognitive dysfunctions and neuropathology. Methods: Female 3xTg mice at 6 months were injected with heat-killed P. gingivalis bacteria into their buccal mucosa three times per week every other week for a total of 5 weeks. Open field, spontaneous Y maze and puzzle box test were used to assess the sickness behavior and cognitive functions. Following behavioral testing, the mandibular jaws were harvested and subjected to micro-CT scan. Hippocampal regions were harvested for biochemical analysis. Results: Bacterial-induced periodontitis led to significant loss of periodontal bone level and exacerbated impairment of short- and long-term memory in 3xTg mice. When examining the hippocampus regions for phosphorylated tau protein levels, bacterial-induced periodontitis led to significant increase in Tau396 expression in both the total lysate and the sarkosyl-insoluble fractions. Conclusions: As bone loss induced by bacterial infection led to increased tau pathology and worsened hippocampal memory functions. Future work will aim to elucidate the mechanism of how periodontal bone loss lead to the development of AD pathology. Acknowledgement: The study is supported by Health and Medical Research Fund (HMRF 04151216) to RCCC. RPHW is awarded by Hong Kong PhD Fellowship.

P358 / #1653


AN INTEGRIN RECEPTOR COMPLEX MEDIATES FILAMENTOUS TAU-INDUCED ACTIVATION OF PRIMARY ASTROCYTES

Lecture Title:

P. Wang1, Y. Ye2 1National Institutes of Health, National Institute Of Diabetes And Digestive And Kidney Diseases, Bethesda, United States of America, 2National Institutes of Health, National Institute Of Diabetes And Digestive And Kidney Diseases, BETHESDA, United States of America

Aims: Microtubule-associated protein Tau can form protein aggregates transmissible within the brain, correlating with the progression of tauopathies in humans. The transmission of aggregates requires neuron-released Tau to interact with surface receptors on target cells. However, the underlying molecular mechanisms in astrocytes and downstream effects are unclear. Given that astrocytes are the most abundant cell type in the brain and that they perform an critical immune surveillance function to maintain brain homeostasis, it is important to determine whether neuron-derived Tau can have an impact on astrocytes, and if so, how would the interaction of Tau with astrocytes feed back to control neurodegeneration? Methods: We employed a spatially resolved proteomic mapping strategy using recombinant APEX2-tagged Tau as the bait. APEX2 is an engineered peroxidase that can covalently tag proximal lysine side chains with biotin molecules in the presence of biotin-phenoxyl radicals. Results: Here we show that integrin αV/β1 receptor binds recombinant human Tau, mediating the entry of Tau fibrils in astrocytes. Distinct recombinant human Tau species binds the astrocytic αV/β1 receptor and differentially activate integrin signalling. Furthermore, Tau-mediated activation of integrin signaling results in NFκB activation, causing upregulation of several pro-inflammatory cytokines and chemokines, and it induces the expression of a sub-group of neurotoxic astrocytic markers, causing astrocytes to release neurotoxic factors. Conclusions: Our findings suggest that filamentous recombinant human Tau-mediated activation of integrin signaling induces astrocytes conversion towards a neurotoxic state, providing a mechanistic insight into tauopathies.

P359 / #871

Topic: Theme B: Taupathies / B1.d. Disease Mechanisms, Pathophysiology: Synapse pathology

PATHOGENIC FORMS OF TAU ACCUMULATE DIVERSELY IN PRESYNAPSES

Lecture Title:

V. Uytterhoeven, L. Deaulmerie, N. Schoovaerts, P. Verstreken VIB-KU Leuven, center For Brain & Disease Research, Leuven, Belgium

Aims: Synapses are the first to deteriorate in neurodegenerative diseases. Our lab has shown that pathogenic Tau accumulates at presynapses and reduces neurotransmission by sequestering synaptic vesicles. However, little is known about the molecular mechanisms that result in the accumulation of Tau at presynapses. Pathogenic forms of Tau are shown to deregulate autophagic systems, including endosomal microautophagy (eMI). eMI is the selective turnover process in which Hsc70-4 recognizes and stimulates the turnover of cytosolic KFERQ-motif containing proteins in late endosomes. We tested if eMI plays a role in Tau accumulation at presynapses. Methods: We used fruit flies expressing human pathogenic Tau and eMI monitoring and modulation tools that we have generated in flies. Results: Despite the reduced eMI levels at mutant Tau presynapses that we measure, we were still able to increase eMI by the expression of hsc70-4 resulting in the reduction of presynaptic TauP301L and rescue of presynaptic function. However, pathogenic TauV337M is not reduced at presynapses when we increase eMI. TauV337M harbors a mutation in the second KFERQ-motif and we hypothesized that the lack of Tau pathology rescue is because Hsc70-4 fails to recognize TauV337M for degradation. Indeed, using a Tau fluorescent timer, we show that more aged Tau protein is present at TauV337M compared to TauP301L mutant presynapses, indicating that TauV337M turnover is hampered at presynapses. Conclusions: Together, dependent on the pathogenic form of Tau, TauP301L mediated presynaptic dysfunction is restored by increasing eMI via Hsc70-4 expression and defects in TauV337M turnover at presynapses contribute to the accumulation of presynaptic TauV337M.

P360 / #374

Topic: Theme B: Taupathies / B1.e. Disease Mechanisms, Pathophysiology: Cellular signaling, kinases, phosphatases, calcium

DEATH-ASSOCIATED PROTEIN KINASE 1 AS A NOVEL TARGET IN TAU AND ALZHEIMER’S DISEASE

Lecture Title:

D. Chen, Y. Mei, T. Zhang, L. Wang, T.H. Lee Fujian Medical University, Institute For Translational Medicine, Fuzhou, China

Aims: We showed that death-associated protein kinase 1 (DAPK1) increases tau expression and its phosphorylation levels related to Alzheimer’s disease (AD). Moreover, DAPK1 expression is dramatically increased in 75% of patients with AD. However, whether and how DAPK1 is regulated in AD is poorly understood. Methods: We tested DAPK1 protein stability, and tau phosphorylation and functions using comprehensive methods in cells, ex vivo mouse models, and human samples to investigate the effects of melatonin on DAPK1 regulation. We conducted pulldown assay using biotin-melatonin to demonstrate direct binding of melatonin and DAPK1. Moreover, we examined the correlation of melatonin, DAPK1 expression, and Pin1 phosphorylation because suppressing Pin1 activity by DAPK1 leads to tau hyperphosphorylation. Results: We found that melatonin plays an important role in regulating tau hyperphosphorylation and function by directly affecting DAPK1 in AD. Firstly, melatonin decreases DAPK1 protein levels in a post-transcriptional manner. Moreover, melatonin directly binds to DAPK1 and induces its ubiquitination in a proteasome-dependent pathway. Furthermore, inhibition of DAPK1 by melatonin and a DAPK1 inhibitor synergistically reduces tau phosphorylation at multiple sites associated with AD by inhibiting Pin1 activity, a prolyl isomerase known to suppress tau hyperphosphorylation, and promotes neurite outgrowth and microtubule assembly. Finally, DAPK1 and melatonin levels are inversely correlated in human AD brains. Conclusions: We demonstrate for the first time that melatonin directly inhibits DAPK1 protein stability, thereby decreasing tau hyperphosphorylation, and may offer a potential therapeutic option for synergistic targeting of DAPK1 by melatonin and DAPK1 inhibitors in AD.

P361 / #1053

Topic: Theme B: Taupathies / B1.e. Disease Mechanisms, Pathophysiology: Cellular signaling, kinases, phosphatases, calcium

DIFFERENTIAL EFFECTS OF APOLIPOPROTEIN E GENOTYPE ON THE MOLECULAR AND CELLULAR PHENOTYPES ASSOCIATED WITH ALZHEIMER’S DISEASE IN AN ISOGENIC HUMAN IPSC-DERIVED NEURON/ASTROCYTE CO-CULTURE SYSTEM

Lecture Title:

Y. You1, W. Poon2, G. Jun3,4, L. Farrer3,4,5, T. Ikezu1,5,6 1Boston University School of Medicine, Department Of Pharmacology & Experimental Therapeutics, Boston, United States of America, 2University of California, Institute For Memory Impairments And Neurological Disorders, Irvine, United States of America, 3Boston University School of Public Health, Department Of Medicine (biomedical Genetics), Boston, United States of America, 4Boston University School of Public Health, Department Of Biostatistics, Boston, United States of America, 5Boston University School of Medicine, Department Of Neurology And Alzheimer’s Disease Center, Boston, United States of America, 6Boston University, Center For Systems Neuroscience, Boston, United States of America

Aims: We aimed to establish a human iPSC (induced pluripotent stem cell) based co-culture system by deriving isogenic APOE iNeurons and iAstrocytes to explore the differential effects of APOE isoforms on AD-related pathology. Methods: Isogenic human APOE2/2, APOE3/3, APOE4/4 and APOE knock-out (APOE-/-) iPSC lines were induced into functional neurons by overexpressing neural fate-driven gene neurogenin2, and iAstrocytes by generating embryoid bodies. Isogenic human neurons were co-cultured with the same APOE genotype of iAstrocytes to test downstream assays. Results: APOE2 iAstrocytes exhibited the highest intracellular APOE level. Expression of PPP2CB was significantly increased in APOE2 compared to APOE3 and APOE4 iNeurons, while there were no significant differences in C4A/B expression among the APOE-defined groups of iNeurons. No significant differences of Aβ40 and Aβ42 levels among different APOE iNeurons were found. However, we observed the highest level of pTau231/tTau ratio from APOE2 while the pTau181/tTau ratio was the lowest from APOE4 iNeurons, compared that from APOE-/- or other APOE carrying iNeurons. Correlation analysis suggested the positive correlation of PPP2CB expression with pTau231/tTau ratio (P=0.013) but not with pTau181/tTau ratio (P=0.74). C4A/B expression was not correlated with levels of pTau231 or pTau181, while C4A/B expression was inversely correlated with Aβ42 level. Furthermore, we found the significant positive correlation between PPP2CB and C4A/B expression (P=2.0x10-4) in iNeurons co-cultured with isogenic iAstrocytes regardless of the APOE genotype of the iNeurons. Conclusions: Our findings suggest the role of APOE isoforms on pTau accumulation, PPP2CB and C4A/B expression as a cross-talk between human neurons and astrocytes in vitro and potentially in AD brains.

P362 / #1373

Topic: Theme B: Taupathies / B1.i. Disease Mechanisms, Pathophysiology: Microglia

MICROGLIAL REMODELING OF ACTIN NETWORK FOR G PROTEIN-COUPLED PURINERGIC RECEPTOR, P2RY12-DRIVEN CHEMOTAXIS BY TAU OLIGOMERS

Lecture Title:

R. Das1, S. Chinnathambi2 1CSIR-National Chemical Laboratory, Neurobiology Group, Biochemical Sciences Division, Pune, India, 2National Chemical Laboratory, Neurobiology Group, Biochemical Sciences Division, Pune, India

Aims: Purinergic signaling specially P2Y12 found to be important in the field of platelet aggregation, but recently got interest in neurodegeneration as it expressed only in microglia among all brain cells. Microglia P2Y12 plays an important role in sensing ATP leakage from damaged neurons, hence mediates chemotaxis. Methods: To understand the mechanism of microglial migration upon extracellular Tau oligomers, the P2RY12-mediated actin remodeling, reorientation of tubulin network and rate of migration were studied in the presence of ATP. The extracellular Tau species directly interacted with microglia P2RY12 and induced cellular expression. Microglial P2RY12 has colocalized with remodeled membrane-associated actin network as a component of migration to phagocytose Tau oligomers. Results: Here, the extracellular Tau species have directly interacted with P2RY12 and also induced this purinoceptor expression in microglia. Microglial P2RY12 has colocalized with remodeled membrane-associated actin network as a component of migration to phagocytose Tau oligomers. As an inducer of P2RY12, ATP has facilitated the increased incorporation of P2RY12 in lamellipodia and the number of filopodia during accelerated microglial migration. The direct interaction of extracellular Tau oligomers with microglial P2RY12 would facilitate the signal transduction in both way, directional chemotaxis and receptor-mediated phagocytosis. Conclusions: First time it has been observed that P2Y12 is directly interacts with Tau oligomers and induces cellular expression. Moreover, external Tau oligomers have induced actin remodeling by more orchestrated lamellipodia and filopodia, which is related to P2Y12 accumulation. This would insight more potential of P2Y12 as a key regulator of microglia function and clearance of pathological Tau in AD.

P363 / #753


AMYLOID PLAQUE DEPOSITION ACCELERATES TAU PROPAGATION VIA ACTIVATION OF MICROGLIA IN A HUMANIZED APP MOUSE MODEL

Lecture Title:

K. Clayton1, J.C. Delpech1, S. Herron1, N. Iwahara1, T. Saido2, T. Saito3, S. Ikezu1, T. Ikezu1 1Boston University School of Medicine, Pharmacology And Experimental Therapeutics, Boston, United States of America, 2RIKEN, Center For Brain Science, Wako-shi, Japan, 3Nagoya City University,

Department Of Neurocognitive Science, Kawasumi−１, Japan

Aims: Aim 1: Determine the effect of microglia depletion on amyloid plaque deposition. Aim 2: Determine the effect of amyloid plaque deposition on pTau propagation. Aim 3: Determine the effect of microglia depletion on pTau propagation. Methods: In order to examine the effect of microglia depletion on plaque pathology and tau propagation, APPNL-G-F (homozygote for the App loci with APPNL-G-F) and WT mice aged 4 months were fed chow containing the CSF1R inhibitor (PLX5622) or placebo for 4 weeks prior to AAV-Tau P301L injection and for an additional month following AAV injection until euthanasia. Results: We show that propagation of phosphorylated tau to the granular cell layer (GCL) of the dentate gyrus is highly exacerbated in humanized APPNL-G-F knock-in mice compared to wild type. Microglia depletion increased plaque burden and ablated the prevalence of plaque-associated microglia (MGnD/DAM), which colocalize strongly with the exosome marker, Tumor susceptibility gene 101. MGnD in APPNL-G-F knock-in mice produce significantly more GFP+ extracellular vesicles (EVs) compared to homeostatic microglia when injected with mEmerald-CD9 lentivirus in the MEC, suggesting MGnD hypersecrete EVs. Conclusions: Our findings indicate that plaque-mediated activation of microglia may supply seeding factors through secreted EVs, exacerbating tau propagation along the perforant pathway.

P364 / #1040


USING CHIMERIC MICE TO EXAMINE HUMAN MICROGLIAL RESPONSES TO AMYLOID AND TAU IN VIVO

Lecture Title:

M. Coburn, J. Hasselmann, A. Mcquade, H. Davtyan, M. Blurton-Jones Institute for Memory Impairments & Neurological Disorders Sue & Bill Gross Stem Cell Research Center, Department Of Neurobiology & Behavior, Irvine, United States of America

Aims: Microglia are strongly implicated in the pathogenesis of Alzheimer’s disease (AD) and several microglial-enriched genes have been associated with altered risk of developing AD. Although both in vitro iPS-derived microglia (iMGL) and in vivo rodent models have provided important tools for studying microglial biology, there remain two critical limitations with these models, namely the rapid transcriptional changes that occur with culturing of microglia and the inability to study the influence of complex human genetic backgrounds on microglia function in mice. Thus we sought to develop a new approach to examine the unique responses of human microglia to AD pathologies in vivo. Methods: iPS-derived hematopoietic progenitor cells (HPCs) were transplanted directly into the brains of xenotransplantation-compatible P1 hCSF-PS5x pups, where they readily differentiated into human microglia, exhibiting typical markers and morphology. 6 months later, human xenotransplanted microglia (XMGs) were isolated from the brain with negative magnetic bead sorting for single-cell RNA sequencing and half brains were fixed and examined by confocal microscopy. Results: We find that XMGs respond and migrate towards amyloid beta plaques, upregulating key disease associated microglia (DAM) markers. Single-cell RNA-seq data from isolated human XMGs is currently undergoing analysis and will also be reported. Conclusions: We anticipate that these studies will elucidate important differences in how human and mouse microglia respond to AD pathology in vivo, greatly expanding our understanding of the transition of homeostatic microglial to pathological and disease associated microglia, helping to fill an important gap in our understanding of human microglia biology in AD.

P365 / #982


NOVEL TREM2 SPLICING ISOFORM THAT LACKS THE V-SET IMMUNOGLOBULIN DOMAIN IS ABUNDANT IN THE HUMAN BRAIN.

Lecture Title:

O. Korvatska1, K. Kiianitsa2, W. Raskind1,3 1University of Washington, Department Of Psychiatry, Seattle, United States of America, 2University of Washington, Department Of Immunology, Seattle, United States of America, 3University of Washington, Division Of Medical Genetics, Seattle, United States of America

Aims: TREM2 is an immune receptor expressed by certain myeloid cells, such as macrophages, dendritic cells, osteoclasts and microglia. Certain missense variants in the TREM2 are associated with an increased risk of Alzheimer’s disease. Human TREM2 gene is a subject to alternative splicing, so we set out to analyze its splicing in the human brain. Methods: Total RNA was isolated from human brain, TREM2 cDNA cloned and sequenced. Transcript levels were quantified with isoform-specific qRT-PCR assays, TREM2 isoforms were visualized at protein level in THP-1 cells with antibodies specific to the C-terminus. Phagocytosis of amyloid beta peptide was measured by flow cytometry. Results: We discovered a novel TREM2 splice isoform that lacks a V-set immunoglobulin domain from its extracellular part but retains transmembrane and cytoplasmic domains (ΔIgV). It had highly variable inter-individual expression in the human brain (average frequency 10%; range 3.7-35%). We confirmed ΔIgV expression at RNA and protein levels in human myeloid cells. Using CRISPR/Cas9 we engineered TREM2 knockout (KO) in THP-1. Overexpression of full-length, but not ΔIgV TREM2 restored defective amyloid beta phagocytosis in KO cells. Conclusions: Because splicing of full-length and ΔIgV TREM2 transcripts are mutually exclusive events, increased level of ΔIgV isoform in the human brain will diminish the dosage of full-length receptor potentially weakening some TREM2 functions.

P366 / #1337


CHARACTERIZATION OF HUMAN DIFFERENTIATED MICROGLIA IN 2D AND 3D ENVIRONMENTS

Lecture Title:

A. Sabogal Guaqueta1, A. Dolga1, A. Thiruvalluvan2, F. Foijer3, L. Barazzuol4, B. Eggen2, E. Boddeke5 1University of Groningen, Molecular Pharmacology, Groningen, Netherlands, 2University of Groningen, University Medical Center Groningen, Biomedical Sciences Of Cells And Systems, Molecular Neurobiology, Groningen, Netherlands, 3University Medical Center Groningen, European Research Institute For The Biology Of Ageing (eriba), Groningen, Netherlands, 4University of Groningen, Biomedical Sciences Of Cells And Systems, Molecular Cell Biology, Groningen, Netherlands, 5University Medical Center Groningen, Biomedical Sciences Of Cells And Systems, Molecular Neurobiology, Groningen, Netherlands

Aims: Microglia, the brain-resident innate immune cells, are increasingly implicated in neurodegenerative diseases, such as Alzheimer’s and Parkinson’s disease. Our goal is to generate a human disease model to study how human microglia communicate with other brain cells in conditions of altered inflammatory signaling. Methods: First, we have generated and characterized microglia differentiated from an iPSC and ESC lines. Immunostaining of CD11b, TMEM119, and Iba-1 demonstrated the maturation of iPSC/ESC-derived microglia in culture. The capacity of phagocytosis was tested both in premature and mature microglia in culture, showing an increased phagocytic capacity in mature microglia compared to progenitor cells as measured by Incucyte live-cell imaging using pHrodo E. Coli BioParticles conjugates. In addition, to mimic inflammatory pathways and to test whether the human iPSC-derived microglia react differently to LPS compared to primary mouse microglia, we performed real-time cell impedance measurements using an xCELLigence system. Results: Microglia from both human and mouse increase cell index after LPS and alpha-syn stimuli. Finally, we investigated the integration of GFP-labeled microglia into brain organoids to mimic the 3D environment of the brain. Application of fluorescent-labeled mature microglia to 3-month-old cerebral organoids for a period of 10 days showed a significant integration based on the morphology of cells. We are currently determining the transcription profiles of microglia cultured in 2D and 3D in the presence of inflammatory stimuli (LPS & alpha-syn). Conclusions: These experiments will provide a characterization of human microglia in a “mini-brain” environment, which could be used to investigate underlying mechanisms and potential treatments of different neurological disorders.

P367 / #701

Topic: Theme B: Taupathies / B1.j. Disease Mechanisms, Pathophysiology: Astroglia

EVALUATION OF ASTROCYTE MORPHOLOGY IN TAUOPATHIES

Lecture Title:

E. Augustin1, A. Mate De Gerando1, C. Josephine1, G. Auregan1, M. Guillermier1, M. Gaudin1, C. Jan1, F. Petit1, M.-C. Gaillard1, E. Faivre2, D. Blum2, A. Bemelmans1, G. Bonvento1, K. Cambon1 1Institut de Biologie François Jacob / Commissariat à l'énergie atomique (CEA), Molecular Imaging Research Center (mircen), Fontenay-aux-Roses, France, 2Inserm UMR-S 1172 Lille Neuroscience & Cognition, Alzheimer & Tauopathies, Lille, France

Aims: Tauopathies are a heterogeneous group of neurodegenerative diseases characterized by intracellular deposits of different abnormal forms of tau in various cell types. Tau being mainly enriched in neurons and poorly expressed in astrocytes, the origin of astrocytic tau aggregates as well as the consequences of the presence of pathological tau in these glial cells remain elusive. Methods: Here, we report the presence of astrocytic tau inclusions in two distinct mouse models of tauopathy. The first model is based on AAV-mediated gene transfer inducing the expression of variants of human Tau (WT, P301L, Pro-aggregant) in neurons of the pyramidal layer of the hippocampus of adult mice. The second model is the Thy-Tau22 transgenic mouse expressing a human MAPT cDNA with 2 mutations (G272V, P301L) driven by the neuron-specific Thy1 promoter. Results: In the AAV tau model, GFAP-positive astrocytes were found immunopositive for phosphorylated (pSer422, AT8) and misfolded (AT100) tau forms. Similar tau-bearing astrocytes were detected in aged Thy-Tau22 male mice (12- and 18- months-old). These results suggest that astrocytic tau inclusions can be secondary to neuronal tauopathy in both models. To assess whether tauopathy had an impact on astrocytes' morphology, we injected intravenously Thy-Tau22 mice with a PHP.eB AAV expressing tdTomato under an astrocyte specific promoter. We are currently using this approach to perform a detailed analysis of their complex morphology, using confocal microscopy combined with 3-dimensional reconstruction. Conclusions: We expect to unravel to what extent different molecular species of tau modify astrocyte arborizations and could thereby alter their multiple functional interactions with neurons.

P368 / #321


ASTROCYTIC ADENOSINE A2A RECEPTOR DYSREGULATION HASTENS MEMORY LOSS, TAU PATHOLOGY AND NEUROINFLAMMATION IN A MOUSE MODEL OF TAUOPATHY

Lecture Title:

E. Faivre1, K. Carvalho1, V. Gomez-Murcia1, J. Coelho2, A. Launay1, B. Thiroux1, M. Besegher1, M. Hamdane1, L. Buee1,3, L. Lopes2, D. Blum1 1Inserm UMR-S1172 Lille Neuroscience & Cognition, Alzheimer & Tauopathies, Lille, France, 2Universidade de Lisboa, Instituto De Medicina Molecular, Faculdade De Medicina De Lisboa, Lisbon, Portugal, 3Inserm UMR-S 1172 Lille Neuroscience & Cognition, Alzheimer & Tauopathies, Lille, France

Aims: Epidemiological and experimental studies pointed out that chronic caffeine consumption reduces AD risk and associated cognitive deficits. These protective effects are thought to be ascribed to the blockade of adenosine A2A receptors (A2ARs). Interestingly, the latter are found abnormally upregulated in the brain of AD patient’s brains, notably in astrocytes, in correlation with Tau pathology development and cognitive deficits. These post-mortem observations suggest a link between A2AR dysregulation, Tau pathology and memory in AD. To get insights towards this relationship, we aimed at evaluating the pathophysiological impact of astrocytic A2AR upsurge in a transgenic model of AD-like Tauopathy (THY-Tau22 mice). Methods: To address the role of astrocytic upsurge, we have developed a conditional model (Tet-Off) allowing A2AR overexpression in GFAP-positive astrocytes. This model was crossed with THY-Tau22 mice, who develop a progressive hippocampal Tau pathology associated with cognitive decline. In the different groups of animals, we evaluated Tau pathological changes (phosphorylation, aggregation) and functional impairments (learning and memory) at 5-6 months of age, when pathology is present but not maximal in the THY-Tau22 model. Results: We found that astrocytic A2AR overexpression worsens spatial memory impairments of THY-Tau22 mice. These effects were associated to an increased Tau phosphorylation and aggregation as well as to the upregulation of hippocampal neuroinflammatory processes induced by Tau pathology Conclusions: Altogether, these data suggest that astrocytic A2AR dysregulation seen in the brain of AD patients contributes to the development of Tau-induced cognitive impairments by modulating Tau pathology and neuroinflammation.

P369 / #683


PLASMA GLIAL FIBRILLARY ACIDIC PROTEIN AND NEUROFILAMENT, AND THEIR COMBINATION AS BIOMARKERS IN FRONTOTEMPORAL DEMENTIA

Lecture Title:

N. Zhu1,2, M. Santos1,2, V. Montal1,2, T. Estellés1,2, I. Illán-Gala1,2, I. Barroeta1,2, M. Altuna1,2, O. Belbin1,2, I. Sala1,2, M.B. Sánchez-Saudinós1,2, A. Subirana1,2, L. Videla1,2, J. Clarimon1,2, M. Carmona-Iragui1,2, R. Blesa1,2, J. Fortea1,2, D. Alcolea1,2, A. Lleó1,2 1Centro de Investigación Biomédica en Red en Enfermedades Neurodegenerativas, Ciberned, Madrid, Spain, 2Institut d’Investigacions Biomèdiques Sant Pau - Hospital de Sant Pau, Universitat Autònoma de Barcelona, Sant Pau Memory Unit, Department Of Neurology, Barcelona, Spain

Aims: to investigate the diagnostic performance of plasma levels of Glial fibrillary acidic protein (pGFAP), plasma neurofilament (pNfL) in frontotemporal dementia (FTD) and Alzheimer disease(AD) and their correlation with cognition, cerebrospinal fluid (CSF) biomarkers and cortical thickness in FTD. Methods: We selected 239 participants from the Sant Pau Initiative on Neurodegeneration cohort (97 FTD/58 AD/84 HC). pGFAP and pNfL concentrations were measured using single molecule array (SIMOA). All subjects had CSF, 177 had a MRI. Biomarker levels across diagnostic groups were compared between groups by ANOVA and the diagnostic performance with ROC analyses. We used Spearman correlation to assess the relationship of pGFAP and pNfL with CSF biomarkers and cognition and cortical thickness was computed with Freesurfer. Results: pGFAP levels were lower in FTD (213±118pg/mL) compared to AD (322±153pg/mL)) and not different from HC (139±64pg/mL); pNfL levels were increased both in FTD (35±38pg/mL) and AD (27±14pg/mL) groups compared to HC (18±20pg/mL)(all p<0.01). The combination of pGFAP and pNfL showed better accuracy to differentiate FTD from AD (AUC: 0.78[CI 0.7-0.86]), superior to that of pGFAP (AUC: 0.73[CI 0.65-0.81]) or pNfL (AUC: 0.56[CI 0.47-0.65]) alone (p=0.04); pNfL and its combination with pGFAP differentiated FTD from HC with similar accuracy (AUC: 0.78[CI 0.71-0.85]/0.79[CI 0.72-0.85]). In FTD, levels of pGFAP correlated with MMSE score, CSF and plasma NfL (Rho: 0.44/0.3/0.51) and cortical-thickness in left-temporoparietal and right-posterior-medial-temporal regions (p<0.05). Conclusions: The combination of pGFAP and pNfL shows better accuracy than pNfL alone to distinguish FTD from AD. pGFAP could be used as a disease severity biomarker in FTD patients.

P370 / #1820

Topic: Theme B: Taupathies / B1.o. Disease Mechanisms, Pathophysiology: Neural networks & plasticity

TAU PATHOLOGY: PIECING THE PUZZLE TOGETHER

Lecture Title:

E. Hill1, T. Karikari2, J. Lantero Rodriguez2, H. Zetterberg2, K. Blennow2, M. Richardson3, M. Wall1 1University of Warwick, School Of Life Sciences, Coventry, United Kingdom, 2Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden, 3University of Warwick, School Of Mathematics, Coventry, United Kingdom

Aims: Tau modulates microtubule stability. When hyper-phosphorylated it forms small toxic oligomers, then fibrils and finally tangles, whose density in the brain correlates with the progression of Alzheimer’s disease. In our previous study (Hill et al, 2019; eNeuro), whole-cell patch-clamp recording was used to introduce oligomers made from full length human tau (oTau; 40-450 nM) into single hippocampal neurons in brain slices. Over a period of 30-40 minutes oTau increased neuronal input resistance, slowed action potential kinetics and reduced action potential amplitude. These effects were not observed with tau monomers or vehicle. Exactly how oTau mediated these changes in neuronal function is unclear. Methods: We have used specific recombinant truncations of oTau to elucidate the mechanisms underlying the changes in neuronal properties. We introduced the truncated versions of oTau into single hippocampal pyramidal cells in acute brain slices and measured the resultant changes in neuronal properties. Results: By using specific truncated forms of oTau we were able to dissect apart the effects of oTau on action potential waveform, excitability and input resistance. Thus, we have identified the parts of the Tau molecule that produce these specific effects and we have now begun defining the mechanisms involved. Conclusions: This simple, yet highly effective technique of introducing oligomeric proteins into single neurons allows unparalleled levels of detail and provides a unique opportunity to understand the underlying pathology for tauopathies. Hill et al (2019). eNeuro, 6(5), pp. eNEURO.0166-19.2019.

P371 / #668

Topic: Theme B: Taupathies / B1.p. Disease Mechanisms, Pathophysiology: Transcriptional & translational regulation, micro RNAs

UNRAVELLING THE REGULATORY MECHANISMS OF THE LNCRNA MAPT-AS1

Lecture Title:

R. Policarpo1,2, A. Sierksma1, A. Ebneth2, I. Royaux3, B. De Strooper1, C. D'Ydewalle2 1VIB-KU Leuven, Center For Brain & Disease Research, Leuven, Belgium, 2Janssen Pharmaceutica NV, Neuroscience, Janssen Research & Development, Beerse, Belgium, 3Janssen Pharmaceutica NV, Discovery Sciences, Janssen Research & Development, Beerse, Belgium

Aims: One strategy to reduce Tau pathology in Alzheimer’s Disease (AD) is to lower Tau transcription. Nonetheless, the regulatory mechanisms of Tau (encoded by the MAPT gene) expression remain largely unknown. Recently, long non-coding RNAs (lncRNAs) emerged as key regulators of protein-coding gene expression. MAPT antisense 1 (MAPT-AS1) has been identified as a lncRNA associated to MAPT. We aim to understand the mechanisms by which MAPT-AS1 regulates MAPT expression and evaluate whether targeting this lncRNA might be therapeutically relevant for AD. Methods: We measured MAPT-AS1 expression levels in brain samples from healthy and AD patients. We modulated MAPT-AS1 expression in a human neuroblastoma cell line and in human iPSC neurons using antisense oligonucleotides (ASOs), short interference RNAs (siRNAs) and lentiviral over-expression constructs. Additionally, we evaluated the effects of MAPT-AS1 knockdown and over-expression on the transcriptome of human iPSC neurons. Results: We confirmed that MAPT-AS1 is equally expressed in human control and AD brain samples, in human cancer cell lines and in human iPSC neurons. We identified ASOs that dose-dependently reduce MAPT-AS1 expression. siRNAs and lentiviral over-expression constructs dose-dependently changed MAPT-AS1 levels. In none of the conditions changes in Tau expression at the RNA or protein level were detected. A microarray analysis showed that alterations in MAPT-AS1 expression in human neurons did not elicit major transcriptional changes. Conclusions: Our data indicate that MAPT-AS1 is expressed in the human brain and that MAPT-AS1 knockdown and over-expression do not change Tau expression in human neurons. Our data suggest that neuronal MAPT-AS1 is not therapeutically relevant for AD.

P372 / #1508

Topic: Theme B: Taupathies / B1.q. Disease Mechanisms, Pathophysiology: Autophagy, apoptosis, cell death

ABERRANT ROLE OF ALK IN TAU PROTEINOPATHY THROUGH AUTOPHAGOSOMAL DYSREGULATION

Lecture Title:

J. Park, Y.-K. Jung Seoul National University, College Of Natural Sciences, Seoul, Korea, Republic of

Aims: Despite active identification of tau modifications responsible for tau aggregation, a critical modulator inducing tau proteinopathy by affecting its protein degradation flux is not known. Methods: Constitutive active form of ALK, ALK.Fc, was overexpression in neuronal cell lines. Lentivirus carrying ALK.Fc or ALK.Fc KD transduction into mouse primary cortical/hippocampal neurons. Human tau transgenic fly (gl-tau2.1 line) was employed to assess in vivo effect of ALK on tau-mediated neurotoxicity. TauC3 and 3xTg-AD mice were used to identify the role of ALK inhibition in mammals in vivo. Results: ALK caused abnormal accumulation of highly phosphorylated tau in the somatodendritic region of neurons through its tyrosine kinase activity. ALK induced LC3-positive axon swelling and loss of spine density, leading to tau-dependent neuronal degeneration. Notably, ALK activation in neurons impaired Stx-17-dependent autophagosome maturation and this defect was reversed by a dominant-negative Grb2. In a Drosophila melanogaster model, transgenic flies neuronally expressing active Drosophila Alk exhibited the aggravated tau rough eye phenotype with retinal degeneration and shortened lifespan. In contrast, expression of kinase-dead Alk blocked these phenotypes. ALK levels were significantly elevated in the brains of AD patients showing autophagosomal defects, and injection of an ALK.Fc-lentivirus exacerbated memory impairment in 3xTg-AD mice. Conversely, pharmacologic inhibition of ALK activity with inhibitors reversed the memory impairment and tau accumulation in both 3xTg-AD and tauC3 (caspase-cleaved tau) transgenic mice. Conclusions: Together, we propose that aberrantly activated ALK is a bona fide mediator of tau proteinopathy that disrupts autophagosome maturation and causes tau accumulation and aggregation, leading to neuronal dysfunction in AD.

P373 / #262

Topic: Theme B: Taupathies / B1.s. Disease Mechanisms, Pathophysiology: Aging

EXCITATORY NEURONS CONSTITUTE A PREDOMINANT SENESCENT CELL POPULATION IN HUMAN ALZHEIMER'S DISEASE BRAIN

Lecture Title:

M. Orr1, S. Kazempour Dehkordi2, H. Zare2, J. Walker2 1Salisbury VA Medical Center, Research, Salisbury, United States of America, 2University of Texas Health San Antonio, Cell Systems And Anatomy, San Antonio, United States of America

Aims: Cellular senescence contributes to tissue pathology, organismal aging and disease in laboratory animal models. The relevance of this complex stress response to human aging and chronic disease remains unknown. The aim of this study was to determine senescent cell abundance and cell type identity in human brain from older adults with or without Alzheimer's disease. Secondary aims included determining whether cellular senescence correlated with advanced age or Alzheimer's disease neuropathology. Methods: We interrogated single nucleus transcriptomic data (~140,000 cells) derived from 72 older adults with varying levels of Alzheimer’s disease pathology using three different gene sets representative of cellular senescence. Bioinformatics approaches were applied to determine the number, identity and profile of cells expressing transcriptomic signatures consistent with senescence. Histology was used to confirm results. Results: Approximately 2% of the total cellular population expressed transcriptomic signatures consistent with cellular senescence. The proportion of senescent cells within individuals varied (0-12%). Most of the senescent cells, >97%, were excitatory neurons and co-expressed transcriptomic profiles consistent with neurofibrillary tangles. The cell cycle inhibitor, CDKN2D, contributed most significantly to the senescence transcriptomic profile. Elevated levels of its protein product, p19, were confirmed in human Alzheimer’s disease tissue using immunohistochemistry.

Conclusions: These data indicate that excitatory neurons expressing elevated levels of CDKN2D/p19 and neurofibrillary tangle pathology constitute a prominent senescent cell population in Alzheimer’s disease dorsolateral prefrontal cortex.

P374 / #1012


EVIDENCE FOR LOCUS COERULEUS TAU PATHOLOGY, CASPASE 3 ACTIVATION AND IMPAIRMENT OF COGNITIVE FUNCTION BY BETA-BLOCKERS IN AGED MICE

Lecture Title:

H.H. Park, A. Evans, M. Shamloo Stanford University, Neurosurgery, Palo Alto, United States of America

Aims: Adrenergic signaling regulates cognition and inflammation. Noradrenergic neurons of the locus coeruleus (LC) provide the primary adrenergic input to the forebrain and are vulnerable to degeneration, which may contribute to age-related cognitive deficits and neuroinflammation. Additionally, adrenergic signaling may be compromised by the use of brain permeable beta-adrenergic antagonists, beta-blockers, prescribed for hypertension. This study was designed to examine age-related neuroinflammation and LC pathology and to determine the effect of beta-blockers on cognition and neuroinflammation in aged mice. Methods: Male mice, 3 and 18 months old, were administered a beta-blocker, propranolol or beta-adrenergic agonist, mabuterol, acutely, and a panel of behaviors were assessed. Inflammatory markers in plasma and hippocampus were assessed alongside evidence for tau pathology in the LC. Results: Hippocampal inflammation and tau pathology in the LC was increased in aged mice, with no effect of drug treatment. Aged mice had reduced activity in multiple tests. In Y-maze, aged mice showed preference for the familiar arm. Mabuterol enhanced and propranol reduced time spent in the familiar arm in old mice and time spent in the novel arm in young mice. Propranolol impaired Novel Object Recognition in both young and old mice. In Fear Conditioning, mabuterol enhanced trace learning. Aged mice froze more in response to context and less in response to cue than young mice. Propranolol attenuated contextual freezing in aged mice. Conclusions: In conclusion, aged mice show LC pathology and heightened impairment of behavior with acute propranolol administration and enhanced learning and memory by acute mabuterol dosing.

P375 / #1488


CORPORA AMYLACEA IN THE HUMAN BRAIN EXHIBIT NEOEPITOPES OF A CARBOHYDRATE NATURE

Lecture Title:

M. Riba1,2,3, E. Augé1,2,3, I. Tena3, L. Molina-Porcel4, T. Ximelis4, J. Vilaplana1,2,3, C. Pelegri1,2,3 1Centros de Biomedicina en Red de Enfermedades Neurodegenerativas, Ciberned, Madrid, Spain, 2Universitat de Barcelona, Institute Of Neurosciences, Barcelona, Spain, 3Universitat de Barcelona, Biochemistry And Physiology, Barcelona, Spain, 4Biobanc-Hospital Clinic-Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Neurological Tissue Bank, Barcelona, Spain

Aims: Corpora amylacea (CA) of the human brain are age-related polyglucosan bodies formed primarily by polymerized hexoses. They also contain residual products of different origins. As they are released from the brain to the cerebrospinal fluid (CSF), they may act as containers that remove waste products from the brain. Recent studies reveal that CA present neoepitopes (NEs) that can be recognized by natural IgMs. The nature of the NEs remains unknown, but several facts suggest that they can have a carbohydrate nature. Thus, the main objective of this study is to determine whether the NEs of CA have a carbohydrate nature. Methods: IgM preadsorptions with increasing concentrations of specific carbohydrates were performed to ascertain if the preadsorption produced a partial or total inhibition of CA staining by the IgMs. Another approach to study the interaction of the IgMs with CA involved staining the CA after their digestion with the peptidic enzyme pepsin. As controls, the same studies were performed by staining CA with concanavalin A, directed against carbohydrates, and with an antibody directed against the p62 protein. Results: The preadsorption of IgMs with carbohydrates show inhibitory effects on the interaction between IgMs and CA. The digestion of CA proteins has no effect on this interaction. Conclusions: The NEs recognized by the natural IgMs have a carbohydrate nature. Moreover, as the interaction of natural IgMs and the NEs can be blocked in vitro by sugars, further studies should now be carried out to assess the possible in vivo effect of glycemia on natural immunity.

P376 / #267

Topic: Theme B: Taupathies / B1.u. Disease Mechanisms, Pathophysiology: Other

MIDBRAIN STEREOTAXIC ROTENONE INFUSION MEDIATES SIMULTANEOUS HALLMARKS OF PARKINSON’S DISEASE, GUT DYSFUNCTION, SPERM MORPHOLOGICAL ABNORMALITY AND TESTICULAR PATHOLOGY

Lecture Title:

I.O. Awogbindin1, I. Adedara2, P. Adeniyi3, A.E. Agedah2, B.F. Oyetunde2, P.D. Olorunkalu3, E. Ogbuewu2, I.A. Akindoyeni2, Y.E. Mustapha2, O.G. Ezekiel2, E.O. Farombi2 1Neuroimmunology group, Molecular Drug Metabolism and Toxicology Unit, Department Of Biochemistry, University Of Ibadan, Ibadan, Nigeria, 2Molecular Drug Metabolism and Toxicology Unit, Department Of Biochemistry, University Of Ibadan, Ibadan, Nigeria, 3Cell Biology and Neurotoxicity Unit, Afe Babalola University, Ado - Ekiti, Ekiti State, Department Of Anatomy, Ado Ekiti, Nigeria

Aims: The asymptomatic and clinical stages of Parkinson’s disease (PD) are associated with comorbid non-motor symptoms. Herein, we aim to investigate a possible causal link between nigrostriatal degeneration and testicular/sperm abnormalities, in addition to the established gut dysfunction, in a rat stereotaxic rotenone model of PD. Methods: Wistar rats received 1 μL of DMSO or 3 μg/μL of rotenone (ROT) in DMSO directly injected into the right ventral tegmental area and substantia nigra pars compacta. Results: After 21 days, the lesioning induced nigrostriatal degeneration with attendant locomotion abnormalities and associated perturbations in the striatal GRP78 and XBP1 levels. Evidence of dopaminergic neuronal death was also observed in the hypothalamus. The rotenone infusion elicited reduced ileal occludin expression and impaired redox status. Epididymal sperm possesed few spermatozoa with myriads of morphological aberrances speculating impaired spermatogenic and spermiogenic events. Epididymal spermatozoa sparingly expressed acrosomal SPACA1. Seminiferous tubules possessed scanty luminal spermatozoa with a few spermatids and post-meiotic germ cells in the adluminal compartment suggesting hypospermatogenesis or maturation arrest. OCCLUDIN-expressing Sertoli cells were dispersed over a wide area suggesting compromised blood-testes barrier. Signal for activated caspase 3 expression was intense in Sertoli and germ cells. Immunoreactivity of spermatogenic-enhancing SRY and GADD45g was weak in the post-mitotic germinative elements. While serum follicle stimulating hormone level remain unaffected, the testicular pathology was associated with reduced serum testosterone level, testicular oxidative damage and inhibition of acetylcholinesterase activity, even when rotenone was not detected in the testes. Conclusions: Overall, testicular and sperm abnormalities are potential significant co-morbidities associated with PD via the brain-hypothalamic-testicular circuit independent of the pituitary.

P377 / #1073


DIFFERENTIAL EXPRESSION OF PROTEINS IDENTIFIED IN NEUROFIBRILLARY TANGLE MATURATION IN ALZHEIMER’S DISEASE USING NANOSTRING MULTIPLEX TECHNOLOGY

Lecture Title:

C. Moloney1, E. Lesser2, J. Kachergus3, C.-C. Liu1, M. Van Blitterswijk1, N. Graff-Radford4, D. Dickson1, E. Thompson3, M. Murray1 1Mayo Clinic, Department Of Neuroscience, Jacksonville, United States of America, 2Mayo Clinic, Department Of Health Sciences Research, Jacksonville, United States of America, 3Mayo Clinic, Department Of Cancer Biology, Jacksonville, United States of America, 4Mayo Clinic, Department Of Neurology, Jacksonville, United States of America

Aims: Neurofibrillary tangles are dynamic entities with a lifespan encompassing three maturity levels: pretangles, mature tangles, and ghost tangles. Proteins are differentially expressed throughout the tangle lifespan; however, past studies may be limited by spatial expression or number of proteins investigated. We sought to assess differences in protein expression across the tangle lifespan using GeoMxTM Digital Spatial Profiler (DSP) by NanoString, a new technology allowing for spatially derived multiplex investigation. Methods: We used DSP to multiplex protein expression in hippocampus from Alzheimer’s disease (n=6) and nondemented controls (n=2) using the “Human Neural Cell Profiling Core” and “Alzheimer’s Pathology” module. Regions of interest (ROIs) were classified by the major tangle maturity level (8 pretangle, 43 mature tangle, and 38 ghost tangle). Protein expression was normalized to the geomean of the three housekeeping genes. The tau-positive segment was specifically analyzed for comparisons. Results: Cytoskeletal and neuronal markers generally decreased through the tangle lifespan. The axonal marker was non-monotonically directed, decreasing from pretangles to mature tangles, then increasing in ghost tangles. Neurodegenerative markers generally increased through tangle maturity levels, except for TDP-43 which decreased. Glial markers were generally increased as tangles matured.

Conclusions: Our findings suggest that proteins are differentially expressed throughout the tangle lifespan and the majority followed a monotonically-directed pattern. Non-monotonically directed protein patterns may reflect the death of the neuron during the ghost tangle maturity level. Future studies will test the hypothesis that the microenvironment of the hippocampus is altered with increasing vulnerability to tangle pathology.

P378 / #1541


PROTEOMICS ANALYSIS OF THE SFPQ INTERACTOME REVEALS DISTINCT SUBTYPE SPECIFIC SIGNATURES IN ALZHEIMER’S DISEASE

Lecture Title:

N. Younas, S. Zafar, I. Zerr Prion Research Group, University Medical Centre Goettingen (UMG), Neurology, Goettingen, Germany

Aims: The main aim of the current study was to uncover pathological mechanisms linked to dysfunctional RNA-binding protein SFPQ (splicing factor proline and glutamine rich), leading to heterogeneous progression rates and phenotypes in Alzheimer’s disease. Methods: Co-immunoprecipitation coupled to mass spectrometry was used to establish differential signatures in rapidly and slowly progressive Alzheimer’s disease (AD). The targets identified from mass spectrometry analysis were translated into cellular models under oxidative stress condition. Immunofluorescence was used for visualization of stress granules. Differential expression of selected target proteins was analysed in untreated (control) and treated cells (stressed) and in human brain patients by immunoblotting. Results: Interestingly, the interactome mapping of stress granule associated RNA-binding protein SFPQ in the human brain of patients with classical AD, rapidly progressive AD and age-matched controls revealed subtype specific differences in the interacting partners of the SFPQ proteins. Differential pattern of post-translational modifications (methylation and phosphorylation) was evident in subtypes of AD. The subtype-specific proteomic differences in the interactome network of SFPQ protein were associated with distinct changes in the molecular function and biological processes, as aresult of disease stress. Furthermore, study of target candidates with the stress granule biology in the cellular models provided subtype specific mechanistic insights. Conclusions: Our findings highlight novel molecular pathways associated with subtypes of Alzheimer’s disease. Further characterization of the amyloidogenic proteins identified in the interactome network, at preclinical and clinical stages of the disease can provide mechanistic insights of the pathological aggregation processes.

P379 / #580

Topic: Theme B: Taupathies / B2.a. Therapeutic Targets, Mechanisms for Treatment: Tau, phosphorylation, truncation

HYDROMETHYLTHIONINE IMPROVES COGNITION AND FACILITATES BRAIN UTILIZATION OF ENERGY SUBSTRATES IN A MOUSE MODEL OF TAUOPATHY

Lecture Title:

R.X. Coelho Dos Santos1, V. Melis1, M. Leith2, T. Baddeley2, G. Riedel1, C. Wischik1,3, C. Harrington1,3 1University of Aberdeen, Institute Of Medical Sciences, Aberdeen, United Kingdom, 2University of Aberdeen, Department Of Chemistry, Aberdeen, United Kingdom, 3TauRx Therapeutics Ltd., N/a, Aberdeen, United Kingdom

Aims: Hydromethylthionine (LMTM), a known tau aggregation inhibitor, showed efficacy in slowing the progression of Alzheimer’s disease in phase 3 clinical trials. We aimed to explore changes in mitochondrial function and related energy production substrates in a mouse tauopathy model (L1) dosed with LMTM. Methods: Female L1 and control NMRI mice were gavaged with LMTM (5 and 15 mg/kg/day) or vehicle for 6 weeks and tested in the water maze. Following mice perfusion with saline, tissue was harvested, snap-frozen in liquid nitrogen and kept at -80° C. Protein levels of mitochondrial electron transport chain (ETC) subunits, activities of complexes I and IV (CI and CIV), levels of L-lactate and lactate dehydrogenase (LDH) subunits and brain concentration of methylthionine (MT) were measured. Results: LMTM improves spatial learning in L1 mice. LMTM did not change the levels of ETC subunits in NMRI or L1 mice, but enhanced CI and CIV activities in L1 mice, with a significant positive correlation between CI activation and MT levels in brain. CIV activity showed a similar dose-response profile. L1 mice have significantly elevated levels of L-lactate, providing a clinically-relevant model for metabolic changes in AD. Although not significant, LMTM decreased levels of brain L-lactate, accompanied by an increase in the levels of LDH-A, but not LDH-B. Conclusions: Our data suggest that LMTM facilitates the usage of central storage of energy substrates, such as L-lactate. Typically, this comes from astrocytes, contributing to reversal of spatial learning deficits in L1 mice by enhancing mitochondrial function.

P380 / #955


INSPECTING THE TAU TRUNCATION IN ALZHEIMER’S DISEASE EYE, A WINDOW TO BRAIN NEURODEGENERATION: TRANSLATIONAL IMPLICATIONS.

Lecture Title:

V. Latina1, G. Giacovazzo2, F. Cordella3, B. Balzamino4, A. Micera4, C. Marchetti5, F. Malerba6, R. Florio6, A. Atlante7, S. Middei8, R. Coccurello2,9, S. Di Angelantonio3,10, P. Calissano1, G. Amadoro1,11 1European Brain Research Institute (EBRI), Tau Protein, Tauopathies And Alzheimer's Disease (ad), Rome, Italy, 2IRCSS Santa Lucia Foundation, Neuroscience, Rome, Italy, 3University of Rome La Sapienza, Physiology And Pharmacology, Rome, Italy, 4IRCSS-G.B.Bietti Foundation, Research Laboratories In Ophtalmology, Rome, Italy, 5European Brain Research Institute (EBRI), Synaptic Signaling And Plasticity, Rome, Italy, 6European Brain Research Institute (EBRI), Nerve Growth Factor (ngf), Rome, Italy, 7National Research Council (CNR), Institute Of Biomembrane, Bioenergetic And Molecular Biotechnologies (ibiom), Bari, Italy, 8National Research Council (CNR), Institute Of Cellular Biology And Neurobiology (ibcn), Monterotondo (Rome), Italy, 9National Research Council (CNR), Institute For Complex System (isc), Rome, Italy, 10Istituto Italiano di Tecnologia (IIT), Center For Life Nanoscience, Rome, Italy, 11National Research Council (CNR), Institute Of Translational Pharmacology (ift), Rome, Italy

Aims: Retina and optic nerve are sites of early extra-cerebral manifestations of Alzheimer’s Disease (AD). Amyloid-beta (Aβ) plaques and neurofibrillary tangles of hyperphosphorylated tau protein are reported in eyes from AD patients and transgenic animals in correlation with inflammation, synaptic reduction, visual deficits, retinal ganglion cell and axonal loss in optic neuritis. However, the pathological relevance of other post-translational tau modifications -such as truncation with generation of toxic fragments- has not yet been investigated. Moreover, whether the neutralization of these deleterious tau-derived peptides exerts protective action on both AD retinal and cerebral neurodegeneration remains unclear. Methods: We have recently developed a monoclonal tau antibody (12A12mAb, 26-36 tau aminoacids) which selectively targets the AD-relevant neurotoxic peptide, derived from truncation of tau protein at its N-term domain, without cross-reaction towards its full-length normal form. 12A12mAb, when intravenously-injected into 6-month-old Tg2576 animal model, significantly improves the AD-like behavioural and neuropathological hippocampal syndrome (Corsetti et al., Brain Commun. 2020; 2(1): fcaa039, doi: 10.1093/braincomms/fcaa039). Results: By taking advantage of our well-established tau-directed immunization regimen, we found out that the 12A12mAb admininistration also exerts a neuroprotective action on some biochemical, morphological and functional parameters of ocular injury associated with animals’ AD phenotype. Conclusions: This study has important translational implications in the AD field by showing for the first time that tau cleavage plays a pivotal role in triggering the pathological changes occurring in vivo in affected retina and vitreous bodies.

P381 / #1397


MODULATORY EFFECT OF NEW TSPO LIGANDS ON THE MITOCHONDRIAL FUNCTION OF TAU CELLS

Lecture Title:

I. Lejri1, A. Grimm1, F. Hallé2, M. Schmitt2, J. Götz3, F. Bihel2, A. Eckert1 1University of Basel, Neurobiology Lab For Brain Aging And Mental Health, Basel, Switzerland, 2University of Strasbourg, Laboratoire D'innovation Thérapeutique, Strasbourg, France, 3University of Queesland, Queesland Brain Institute, Queesland, Australia

Aims: Translocator protein 18 kDa (TSPO) is a mitochondrial protein located in the outer membrane and involved in cholesterol translocation, a prerequisite for steroid biosynthesis. In the central nervous system, its expression is upregulated in neuropathology such as Alzheimer’s disease (AD). Previously, we demonstrated that two new TSPO ligands named 2a and 2b, stimulated pregnenolone synthesis and ATP production in a cellular model of AD overproducing b-amyloid peptide. We aim to evaluate the effect of these ligands to modulate the mitochondrial activity in tau cells. Methods: The present study evaluates the impact of the new TSPO ligands on mitochondrial dysfunction in a cellular model of AD-related tauopathy presenting mitochondrial impairments including a decreased ATP synthesis and mitochondrial membrane potential as well as a decrease in pregnenolone synthesis when compare to control cells. The effects of our new ligands were compared to those of TSPO ligands described in the literature (XBD173, SSR-180,575 and Ro5-4864). Results: The TSPO ligands 2a and 2b exerted beneficial mitochondrial modulatory effects by increasing ATP levels and mitochondrial membrane potential, paralleled with an increase of pregnenolone levels in mutant Tau cells as well as in control cells. The compounds 2a and 2b showed similar effects on mitochondrial activity than those obtained with the TSPO ligands of reference. Conclusions: These findings indicate that the new TSPO ligands modulate mitochondrial bioenergetic phenotype as well as the de novo synthesis of neurosteroids in a cellular model of AD-related tauopathy, suggesting that these compounds could be potential new therapeutic tools for the treatment of AD.

P382 / #748


APELIN-RECEPTOR AGONIST PROMOTES NEURONAL INSULIN SENSITIVITY—IMPLICATION OF THERAPEUTIC POTENTIAL OF ALZHEIMER’S DISEASE

Lecture Title:

K.F.O. Ma1, C.W.-K. Wong1, M.-Y.K. Leung2, K.-H. Chan1, R.C.-L. Ng1 1The University of Hong Kong, Medicine, Hong Kong, Hong Kong PRC, 2The City Univeristy of Hong Kong, Chemistry, Hong Kong, Hong Kong PRC

Aims: Insulin resistance has been considered as common molecular linkage between diabetes and Alzheimer’s Disease pathogenesis. Reports indicate that cerebral insulin resistance activates GSK3bwhich associates with increase of Abproduction and Tau phosphorylation. The G-protein-coupled receptor APJ and its endogenous ligand Apelin, are involved in the pathogenesis of metabolic disorders including diabetes. Recent studies have revealed that Apelin can enhance glucose uptake by promoting GLUT4 translocation and restore insulin sensitivity to inhibit GSK3bactivity by PI3K/AKT activation. Hence, we hypothesize that Apelin signaling can be a potential target to treat AD. In this project, a novel APJ agonist, CMF-019, is used to investigate if the cerebral insulin sensitivity can be restored through the activation of Apelin/APJ system. Methods: For in vitrostudy, insulin-induced insulin resistance (IR) hippocampal cells (HT22IR) were pre-treated with CMF-019 and cultured with 10 nmol/L insulin for 30 min. Insulin-signaling molecules and its corresponding phosphorylation were measured by western blot analysis. Using LC-MS/MS analysis, the pharmacokinetics of CMF-019 in mice plasma and brain were quantitatively determined after oral gavage. Results: Apelin and APJ are both expressed in cortical and hippocampal neurons. Recombinant Apelin and CMF-019 increased insulin-induced Akt phosphorylation in HT22 neurons. Moreover, CMF-019 pretreatment alleviated insulin sensitivity to increase Akt phosphorylation and inhibit GSK3bin HT22IRcells.CMF-019 was able to be detected by the LC-MS/MS in the plasma and the brain of CMF-019-oral adminstrated mice. Conclusions: CMF-019 enhances insulin sensitivity and inhibits GSK3bthrough PI3K/AKT activation. Orally active CMF-019 may implicate the thereuptic potential of CMF-019.

P383 / #534


NEUROPROTECTIVE PROPERTIES OF CENTRALLY ACTING ANOREXIGENIC PEPTIDES IN MOUSE MODELS OF NEURODEGENERATION

Lecture Title:

A. Popelová, B. Neprašová, M. Holubová, B. Železná, J. Kuneš, L. Maletínská IOCB AS CR, Biochemistry, Prague, Czech Republic

Aims: Despite immense search for treatment of Alzheimer’s disease (AD), there is still no effective drug suppressing broad spectrum of AD hallmarks, such as Aβ plaques, Tau hyperphosphorylation, neuroinflammation or decreased neurogenesis. Because of the described link between obesity and/or type 2 diabetes and increased risk of AD development, anorexigenic compounds with glucose-lowering properties, such as glucagon-like peptide 1 analog liraglutide, and palmitoylated analog of prolactin-releasing peptide (palm-PrRP) were repurposed as possible neuroprotective compounds. Methods: Liraglutide and palm-PrRP were tested in several mouse models for their prospective neuroprotective properties. MSG mice, model of prediabetes and obesity caused by injection of monosodium glutamate (MSG) to newborns, were 14 days subcutaneously (SC) injected twice daily with liraglutide (0.2mg/kg) or palm-PrRP (5mg/kg). Subsequently, double transgenic APP/PS1 mice, a model of AD-like Aβ pathology, were 2 months SCinjected once daily with liraglutide (0.2mg/kg) or with palm-PrRP (5mg/kg). The histopathological changes were measured using the method of Western blot or immunohistochemistry. Results: The treatment of MSG mice with liraglutide and palm-PrRP resulted in increased activation of hippocampal insulin signaling cascade, thus decreased activity of one of the main Tau kinase glucagon-synthase kinase 3β. Subsequently, attenuated Tau hyperphosphorylation at Thr212, Thr231 and Ser396 was observed. In APP/PS1 mice, both peptides reduced the hippocampal and cortical Aβ plaque load and microgliosis, and tended to reduce cortical astrocytosis. They also increased neurogenesis indicated by the number of doublecortin-positive cells. Conclusions: Anorexigenic peptides liraglutide and palm-PrRP have beneficial neuroprotective properties targeting the main hallmarks of AD.

P384 / #934

Topic: Theme B: Taupathies / B2.b. Therapeutic Targets, Mechanisms for Treatment: Immunotherapy

EVALUATION OF PNT001, A MONOCLONAL ANTIBODY DIRECTED TOWARD CIS-PT231 TAU, IN HUMAN TISSUE AND THE TG4510 TAU TRANSGENIC MOUSE

Lecture Title:

K. Foster1, V. Makani2, P. Koivula2, D. Havas3, S. Rind4, L. Goldstein4, K. Brunden2, M. Manca5, A. Kraus5, M. Ahlijanian1 1Pinteon Therapeutics, 1188 Centre St, Newton Centre, United States of America, 2Perelman School of Medicine, University of Pennsylvania, Center For Neurodegenerative Disease Research, Philadelphia, United States of America, 3PsychoGenics, Inc., 215 College Road, Parasmus, United States of America, 4Boston University School of Medicine, Molecular Aging & Development Laboratory, Boston, United States of America, 5Case Western Reserve University, School of Medicine, Department Of Pathology, Division Of Experimental Pathology, Cleveland, United States of America

Aims: The cis-conformer of tau phosphorylated at amino acid residue threonine-231 (cis-pT231 tau) is hypothesized to play a pathologic role in tauopathies. A humanized monoclonal antibody, PNT001, that recognizes cis-pT231 tau with high affinity and selectivity and may have benefit in tauopathies has been identified. PNT001 was characterized in human brain tissue and CSF, evaluated for its ability to reduce seeding activity in Alzheimer’s disease brain lysates, and tested in the Tg4510 mouse model of pathologic tau overexpression. Methods: Immunohistochemical and western blot analyses of PNT001 were assessed in brain tissue from PSP, AD, and CTE patients, and age-matched controls (AMCs). PSP, TBI and AD patient and AMC CSF was assessed for the presence of cis-pT231 tau by SIMOA-based ELISA. Published RT-QuIC methodology was used to assess seeding activity. A murine version of the antibody, mPNT001, was evaluated in vivo in the Tg4510 mice. Results: Immunohistochemical experiments with PNT001 revealed neurofibrillary tangle-like structures with no staining in AMCs in tauopathy disease patients. cis-pT231 tau was detected in CSF from PSP and TBI patients but not AMCs. PNT001 reduced tau seeding activity from AD brain lysates by ~90%. mPNT001 reduced neuroinflammatory marker mRNA, improved synaptic and behavioral outcomes, reduced serum NfL, and delayed NFT formation in Tg4150 mice. Conclusions: PNT001 selectively recognizes cis-pT231 tau in PSP, AD and CTE, reduces seeding activity from AD brain, and improves biochemical and functional endpoints in an aggressive model of pathologic tau overexpression. These data support clinical entry into PSP, AD, TBI and other tauopathies.

P385 / #414

Topic: Theme B: Taupathies / B2.c. Therapeutic Targets, Mechanisms for Treatment: Kinases, phosphatases, other enzymes

DIETARY (-)-EPICATECHIN INHIBITS TAU PHOSPHORYLATION INDEPENDENT OF DIRECT INHIBITION OF GSK3BETA

Lecture Title:

K. Hole1, L. Staniaszek2, J. Mason1, J. Brown2, R. Williams1 1University of Bath, Biology And Biochemisty, Bath, United Kingdom, 2University of Exeter, Medical School, Exeter, United Kingdom

Aims: Oral administration of the dietary flavonoid (-)-Epicatechin (EC) reduces tau hyperphosphorylation in the rTg4510 mouse model (unpublished). Although EC is bioavailable it is also metabolised and the human and rat metabolome have now been defined. The overall aim of this study was to determine the mechanisms by which EC or its metabolites inhibit tau phosphorylation. One such mechanism could be either direct or indirect inhibition of the key tau kinase glycogen synthase kinase 3 beta (GSK3beta). Methods: A luciferase-based in vitro GSK3beta activity assay was used to measure direct inhibition of GSK3beta activity. Primary cortical neurons were either transfected or transduced with GFP-tagged human tau constructs including the P301L tau mutant associated with Frontotemporal Dementia with Parkinsonism. Western blotting was undertaken with these cellular models of tauopathy to determine changes to kinase activity. Results: Neither EC, its metabolites, nor the structurally related anti-amyloidogenic flavonoid epigallocatechin gallate (EGCG) significantly inhibited GSK3beta activity directly at physiologically relevant concentrations (< 1μM). However, preliminary evidence suggests that EC could regulate GSK3beta indirectly by increasing activity of the Akt pathway. Conclusions: The reduction in tau phosphorylation observed in vivo following oral administration of EC does not appear to be due to direct inhibition of GSK3beta and is more likely a result of indirect modulation of tau kinases. Acknowledgements: Funded by the Bristol and Bath Alzheimer’s Research UK Network and Alzheimer’s Society (481). KLH is supported by a GW4 BioMed MRC PhD studentship.

P386 / #1403

Topic: Theme B: Taupathies / B2.g. Therapeutic Targets, Mechanisms for Treatment: Neurotrophic, synaptic plasticity, repair

HONEYBUSH COMPOUNDS EXERT BENEFICIAL EFFECTS ON BIOENERGETICS AND NEURITE OUTGROWTH AND IN A CELLULAR MODEL OF TAUOPATHY.

Lecture Title:

A. Agapouda1, I. Lejri1, V. Butterweck2, M. Hamburger3, D. De Beer4,5, E. Joubert4,5, A. Eckert1 1University of Basel, Transfaculty Research Platform, Molecular And Cognitive Neuroscience, Neurobiology Lab For Brain Aging And Mental Health, Basel, Switzerland, Basel, Switzerland, 2University of Applied Sciences and Arts Northwestern Switzerland (FHNW), School Of Life Sciences, Institute Of Pharmaceutical Technology, Muttenz, Switzerland, 3University of Basel, Division Of Pharmaceutical Biology, Department Of Pharmaceutical Sciences, Basel, Switzerland, 4Stellenbosch University, Department Of Food Science, Stellenbosch, South Africa, 5Agricultural Research Council (ARC) Infruitec-Nietvoorbij, Post-Harvest and Wine Technology Division, Plant Bioactives Group, Post-harvest And Agro-processing Technologies, Stellenbosch, South Africa

Aims: Honeybush is a plant endemic to South Africa. Honeybush extracts have been discussed to exert anti-diabetic, anti-obesity, anti-cancer and anti-oxidant properties. Previous studies of our group showed that the abnormal Tau protein (P301L) impairs mitochondrial function. This study aims to attest the potential capacity of lead compounds of honeybush (mangiferin, hesperidin) in enhancing the bioenergetics and in promoting neurite outgrowth in a cellular model of tauopathy. Methods: The control human neuroblastoma cell line SH-SY5Y and a neuroblastoma cell line bearing the human tau P301L mutation are used in this study. The capacity of mangiferin and hesperidin in increasing bioenergetics and in promoting neurite outgrowth is being investigated. The nerve growth factor (NGF) is used as positive control. Pictures are taken using confocal laser scanning microscopy and are analysed with ImageJ software. The soma and neurite count, the total neurite length, the attachment points and the endpoints of neurites and the extent of neurite branching are analysed. Results: Mangiferin and hesperidin were found to increase the ATP production in control neuroblastoma cells after 24 and 48 hours of treatment. Mangiferin (10 uM) increased respiration and promoted neurite outgrowth to the same extent as that caused by NGF. The majority of the neurite outgrowth parameters were increased by mangiferin. The effects of mangiferin and hesperidin are currently further being investigated in the cellular model of tauopathy. Conclusions: Mangiferin and hesperidin are promising agents in increasing bioenergetics and promoting neurite outgrowth in neurodegenerative disorders such as Alzheimer`s disease and tauopathies.

P387 / #522


SECRETED APP-ALPHA HAS THERAPEUTIC POTENTIAL IN A TAU TRANSGENIC MOUSE MODEL OF AD

Lecture Title:

C. Bold1, D. Baltissen1, S. Ludewig2, L. Salzburger2, D. Wolfer3,4, M. Korte2,5, U. Müller1 1Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Department Of Bioinformatics And Functional Genomics, Heidelberg, Germany, 2Zoological Institute, TU Braunschweig, Division Of Cellular Neurobiology, Braunschweig, Germany, 3Institute of Human Movement Sciences, Eth Zurich, Zurich, Switzerland, 4Institute of Anatomy and Zurich Center for Integrative Human Physiology, University Of Zurich, Zurich, Switzerland, 5Helmholtz Centre for Infection Research, Neuroinflammation And Neurodegeneration Group, Braunschweig, Germany

Aims: Previous evidence indicates a neuroprotective and neurotrophic function for APPsalpha not only in vitro, but also when expressed by AAV vectors in vivo such as in APP/PS1 transgenic AD model mice with Abeta-induced pathology. Previously we could show that APPsalpha rescued deficits of APP/PS1 in synaptic plasticity and spine density and also reduced plaque deposition. Furthermore, APPsalpha has been shown to reduce tau phosphorylation by regulating major tau kinases in vitro. Thus, it is crucial to test a more general potential of APPsalpha as a treatment for AD in particular for tau-induced pathology. Methods: Recombinant APPsalpha as well as AAV-mediated gene transfer of APPsalpha into the hippocampus of hTau.P301S mice was used to investigate its neuroprotective potential to improve structural, functional and biochemical deficits. Results: When assessing synaptic plasticity at the CA3/CA1 pathway in hTau.P301S mice we observed an increase in LTP induction and maintenance which was rescued to WT level by the application of recombinant APPsalpha. This increase in LTP was accompanied by a reduction of several subtypes of GABAergic interneurons in the hippocampus that was detectable as early as 4 weeks of age, before the onset of Tau pathology. To further analyze the neuroprotective potential of APPsalpha in vivo, we used intracranial AAV- APPsalpha injections into the hippocampus. Interestingly, this treatment fully rescued the spine density deficit of hTau.P301S mice to WT levels. Conclusions: Our results point towards a neuroprotective function of APPsalpha in a tau mouse model with aggressive tau pathology.

P388 / #1783


BRAIN PENETRANT TRKB NEUROTROPHIN RECEPTOR AGONIST ANTIBODIES - TRANSLATION TO NON HUMAN PRIMATES

Lecture Title:

F. Walsh1, P. Stocki1, L. Rutkowski1, P. Doherty2 1Ossianix, Research, Philadelphia, United States of America, 2King's College, Wolfson Centre, London, United Kingdom

Aims: Aims: We previously reported on TXB4 a transferrin receptor (TfR) single domain VNAR antibody that can transport TrkB agonist antibodies to the CNS at therapeutic concentrations and are active in PD mouse models. We now present a new shuttle called TXP1 for the delivery of similar product levels to the CNS in non-human primates (NHP). Methods: Methods: Brain accumulation the TfR targeting TXP1 VNAR fused to the human Fc domain was measure by ELISA 20 hr following iv injection at 1.3 mg/kg in NHPs. Multiple brain areas were isolated after cardiac perfusion and assessed for TfR VNAR levels compared to a control VNAR. Product accumulation in the brain was verified by immunohistochemistry. Results: Results: TXP1 Is a high affinity (1-3 nM) VNAR that binds primate and human TfR both as a soluble ectodomain and in live cells. TXP1 efficiently internalizes into hCMEC/D3 cells and does not compete with transferrin binding. After iv injection in NHPs TXP1 was found in multiple brain regions by ELISA with concentrations around 4 nM. Levels were up to 35-fold greater in regions such as the hippocampus when compared to a control VNAR. Brain/plasma ratios were around 2%. Immunohistochemistry confirmed the localization of TXP1 to capillaries, parenchyma and neurons. Conclusions: Conclusions: TXP1 is a high-capacity shuttle for delivery of antibodies and other products to the CNS. The shuttle will allow the delivery of TrkB agonist antibodies in NHPs and lead to clinical trials in neurodegenerative diseases.

P389 / #1048

Topic: Theme B: Taupathies / B2.h. Therapeutic Targets, Mechanisms for Treatment: Protein aggregation, NFT, misfolding, chaperones

EFFICACY OF A SMALL MOLECULE INHIBITOR TARGETING TAU SELF-ASSOCIATION IN THERAPEUTIC STUDIES IN MOUSE MODELS OF TAUOPATHY

Lecture Title:

J. Moe1, P. Lopez1, H. Jimenez-Bravar2, L. Adrien2, A. Wolin2, J. Eun2, J. Koppel2, P. Davies2, E. Davidowitz1 1Oligomerix, Inc., R&d, White Plains, United States of America, 2The Feinstein Institute For Medical Research, Northwell Health, The Litwin-zucker Research Center For The Study Of Alzheimer's Disease, Manhasset, United States of America

Aims: The objectives of these studies are to determine the therapeutic efficacy of the lead compound in the htau and JNPL3 mouse models of tauopathy. Measurements of therapeutic efficacy included reduction of insoluble and hyperphosphorylated tau that has already accumulated, inhibition of the continued progression of tau pathology, as well as amelioration of behavioral deficits. Methods: Therapeutic studies were independently performed in male htau and female JNPL3 transgenic mice. Mice were aged to 7 months (baseline) and treated for 4 months. Each study had 4 groups including baseline (n=20), vehicle (n=25), and two treatment groups (n=25, each) that were administered 40 or 80 mg/kg dose of lead compound formulated in feed. The htau baseline group was tested for working memory performance with the Barnes maze and the JNPL3 baseline group had open field behavior and Rotarod performance testing prior to sacrifice at 7 months; the vehicle and treatment groups had behavioral testing performed at 7 and 12 months. Samples of brain were taken for biochemical analysis of tau and phosphorylated tau, as well as levels and phosphorylation of insoluble, aggregated tau, and immunocytochemical examination. Results: The behavioral studies were completed; and the biochemical analyses are in progress and will be presented at the meeting. Conclusions: The results of the therapeutic studies build upon the successful preventive efficacy studies.

P390 / #1150

Topic: Theme B: Taupathies / B2.i. Therapeutic Targets, Mechanisms for Treatment: Gene and RNAi therapy

APOE4 REDUCTION BY ANTI-SENSE OLIGONUCLEOTIDES MITIGATES TAU PATHOLOGY AND NEURODEGENERATION IN THE P301S/APOE4 MOUSE MODEL OF TAUOPATHY

Lecture Title:

A. Litvinchuk1, T.-P.V. Huynh1, Y. Shi1, R. Jackson2, M.B. Finn1, M. Manis1, C. Francis1, A. Tran1, P. Sullivan3, J. Ulrich1, B. Hyman2, T. Cole4, D. Holtzman1 1Washington University School of Medicine, Hope Center For Neurological Disorders, St. Louis, United States of America, 2Harvard Medical School, Neurology, Boston, United States of America, 3Duke University Medical Center, Durham Veterans Health Administration Medical Center's Geriatric Research, Education and Clinical Center, Medecine, Durham, United States of America, 4Ionis Pharmaceuticals Inc., Neurodegeneration Unit, Carlsbad, United States of America

Aims: Apolipoprotein E (APOE) genotype is the strongest genetic risk factor for late-onset Alzheimer`s disease with the e4 allele increasing risk in a dose-dependent manner. In addition to apoE4 playing a crucial role in amyloid-β deposition, recent evidence suggests that it also plays an important role in tau pathology and tau-mediated neurodegeneration. It is not known, however, whether therapeutic reduction of apoE4 would exert protective effects on tau-mediated neurodegeneration. Methods: We utilized antisense oligonucleotides (ASOs) against human APOE to reduce apoE4 levels in the P301S/ApoE4 mouse model of tauopathy. We treated P301S/ApoE4 mice with apoE or control ASOs via intracerebroventriuclar injection at 6 months and 7.5 months of age and performed brain pathological assessments at 9 months of age. Results: Our results indicate that treatment with apoE ASO’s reduced apoE4 protein levels by ~50%, significantly protected against tau pathology and associated neurodegeneration, decreased neuroinflammation, and preserved synaptic density. These data were also corroborated by a significant reduction in levels of neurofilament light chain (NfL) protein in plasma of ASO-treated mice. Conclusions: Together, we conclude that reducing apoE4 levels should be further explored as a promising therapeutic approach for individuals with tauopathy including Alzheimer`s disease.

P391 / #659

Topic: Theme B: Taupathies / B2.m. Therapeutic Targets, Mechanisms for Treatment: Other

THE EXPRESSION OF MICROTUBULE ASSOCIATED PROTEIN TAU IN MICE PODOCYTE.

Lecture Title:

T. Miyasaka1,2, J. Chang2, Z. Yao2, S. Wada-Kakuda2, H. Tatebe3, T. Tokuda3 1Doshisha University, Center For Research In Neurodegenerative Disease, Kyotanabe-shi, Kyoto, Japan, 2Doshisha University, Neuropathology, Kyotanabe-shi, Kyoto, Japan, 3National Institutes for Quantum and Radiological Science and Technology, Department Of Functional Brain Imaging, Chiba, Japan

Aims: The microtubule-associated protein tau is known to be expressed in neurons and oligodendrocytes. However, the expression of tau in tissues other than nervous system is not well identified. In this study, we attempted to demonstrate tau-expressing cells in the peripheral tissues of mice. Methods: Paraffin-embedded or microslicer sections were prepared from the peripheral tissues of wild-type and tau-knockout mice and subjected to immunohistochemical staining using anti-tau antibodies. The phosphorylation and isoform of tau expressed in kidney were analyzed by Western blotting and RT-PCR. A mouse model of diabetic nephropathy was induced by intraperitoneal administration of streptozotocin, and the plasma p-tau was quantified by SIMOA. Results: We identified tau expression in mice podocyte in kidney. Super-resolution microscope analysis demonstrated that the expressed tau was distributed on microtubules. Biochemical analysis showed that the isoform component of tau expressed in podocyte is mainly 0N4R, that is indistinguishable with that in mice brains. The band shift of the tau on SDS-PAGE after protein phosphatase treatment indicated that the tau expressed in podocyte is highly phosphorylated. Now we are also verifying whether the degeneration of podocyte affects plasma p-tau level that is a biomarker of Alzheimer’s disease, using streptozotocin-induced diabetic nephropathy mouse model. Conclusions: Tau is also expressed in some peripheral tissues, and it is possible that it has a physiological function. It is also necessary to consider the effect of degeneration of peripheral cells expressing such neural proteins on blood biomarkers.

P392 / #704

Topic: Theme B: Taupathies / B3.d. Drug Development, Clinical Trials: Kinase inhibitors & phosphatase modulators

DISCOVERY OF SELECTIVE GSK-3Β INHIBITOR BY STRUCTURE BASED VIRTUAL SCREENING AND IN-VITRO KINASE ASSAY

Lecture Title:

B. Choudhary1, S. Sukanya1, S. Bach2, S. Ruchaud3, T. Robert2, B. Josselin2, R. Malik1 1Central University of Rajasthan, Department Of Pharmacy, Ajmer, India, 2Station Biologique de Roscoff, Kinase Inhibitor Specialized Screening Facility - Kissf, Roscoff, France, 3Station Biologique de Roscoff, Protein Phosphorylation And Human Diseases Unit, Roscoff, France

Aims: One of the characteristic hallmarks of Alzheimer's disease is accumulation of neurofibrillary tangles (NFT). These proteins are produced by hyperphosphorylation of tau protein by kinases. Among all kinases Glycogen synthase kinase - 3β (GSK-3β) catalyzes the rate-limiting step of the NFT generation, therefore GSK-3β has been proposed as a promising target for the treatment of AD. Inhibition of GSK-3β hyperphosphorylation activity would halt the formation of NFT at the very beginning, hence serving as a potential target for Alzheimer’s disease. Methods: In this study, we performed structure-based virtual screening of Asinex library of 6,92,982 compounds against GSK-3β using Glide module of Maestro (Schrodinger) and in-silico ADMET analysis using QikProp module of Maestro for best scoring hits by applying CNS filters to identify novel GSK-3β inhibitors. On basis of CNS filter 4 compounds was selected for further protein kinase assay against GSK-3β, CK-1δ, DYRK1A, CDK-5, CDK-9, PIM-1, CLK-1 and HASPIN. Results: New GSK-3β inhibitors were identified using a structure-based screening, ADMET analysis. These studies revealed that ZINC09036109, ZINC72371723, ZINC72371725 and ZINC01373165 approached optimal ADMET properties along with good MM-GBSA dG binding. During disease kinase profiling, ZINC09036109 emerged as selective GSK-3β inhibitor with 1.08 μM IC50 with more than 10-fold selectivity over other disease relevant kinases. Conclusions: From these studies novel selective GSK-3β inhibitor was identified, which may serve as tool in future for designing of new strategies for GSK-3β inhibition.

P393 / #275

Topic: Theme B: Taupathies / B3.e. Drug Development, Clinical Trials: Aggregation inhibitors

SELECTION AND CHARACTERIZATION OF NOVEL TAU SPECIFIC D-PEPTIDES AS A POTENTIAL THERAPEUTIC APPROACH FOR ALZHEIMER DISEASE

Lecture Title:

I. Aillaud1, S. Kaniyappan2,3, R.R. Chandupatla2, E. Mandelkow3,4,5, S. Funke1 1Coburg University of Applied Sciences and Arts, Institute Of Bioanalysis, Coburg, Germany, 2Deutsches Zentrum für Neurodegenerative Erkrankungen, (dzne), Bonn, Germany, 3University of Bonn, Department Of Neurodegenerative Diseases And Geriatric Psychiatry, Bonn, Germany, 4CAESAR, Research Center, Bonn, Germany, 5Deutsches Zentrum für Neurodegenerative Erkrankungen, (dzne) Bonn, Bonn, Germany

Aims: Alzheimer disease (AD), the most common form of dementia is a progressive neurodegenerative disorder of elderly. One of pathological hallmarks of the Alzheimer disease is abnormally aggregated tau protein. In AD, tau pathology correlates strongly with clinical symptoms, cognitive dysfunction and neuronal death. Thus, we aimed to develop novel therapeutic D-enantiomeric peptides as tau aggregation inhibitors that bind to tau monomers. This could provide an early point of intervention in the pathological fibrillization cascade. Methods: Using a phage display procedure against full-length tau, we selected novel D-peptides that can bind to tau monomers. The promising D-peptide’s ability in binding to tau, inhibition of tau aggregation, and disintegration of tau filaments were tested using ELISA, DLS and Thioflavin S (ThS). Using immunocytochemistry, the cellular uptake and localization of the novel D-peptides were investigated. Results: One of the selected peptides and its retro-inverso form inhibited in vitro fibrillization of wild type, and disease related mutant full-length Tau (TauFLΔK280, TauA152T, TauP103L) as well as pro-aggregant repeat domain Tau mutant (TauRDΔK280). Based on DLS and ThS results, it is evident that our D-peptide induced the formation of larger tau aggregates, presumably amorphous in nature without proper β-sheet conformation. N2a cells have taken up D-peptides and the peptides localized in cell organelles. Conclusions: Based on our results on novel D-peptide against tau, it appears that D-peptide could emerge as a promising therapy for early intervention of AD, presumably by inhibiting toxic tau oligomer formation.

P394 / #705

Topic: Theme B: Taupathies / B3.m. Drug Development, Clinical Trials: Medicinal chemistry approaches, drug repurposing

DISCOVERY OF NATURAL GSK-3Β INHIBITORS AS ANTI-ALZHEIMER’S AGENTS THROUGH STRUCTURE BASED VIRTUAL SCREENING AND IN-SILICO ADMET STUDIES

Lecture Title:

S. Sukanya, B. Choudhary, R. Malik Central University of Rajasthan, Department Of Pharmacy, Ajmer, India

Aims: Glycogen synthase kinase-3β (GSK-3β) phosphorylates many substrates that regulate various cellular processes like cell cycle progression, proliferation, metabolism, transcription and signaling, therefore it is an attractive therapeutic target for various human diseases like cancer, diabetes, Alzheimer’s disease, inflammation, mood disorders and other neurodegenerative diseases. Glycogen synthase kinase-3β (GSK-3β) is a serine/threonine kinase whose over activity results in increased tau phosphorylation, memory impairment, increased β-amyloid production and local plaque associated microglial-mediated inflammatory response. These all are the well known characterstics of Alzheimer’s disease (AD), hence making GSK-3β an attractive target for AD. Methods: Structure based virtual screening and in-silico ADMET studies were performed to identify novel scafford of GSK-3β inhibitors using biogenuix natural library. Structure based virtual screening was performed using Glide module of Maestro and for in-silico ADMET studies QikProp module of Maestro was used. Results: On the basis of structure based virtual screening and in silico ADMET studies 10 natural compounds were found to be promising GSK-3β inhibitors. Common key interaction were observed with Lys85, Val135 and Asp200 amino acid residues which are already reported in literature as catalytic interacting amino acid residues for GSK-3β inhibitors. Conclusions: Summarily novel GSK-3β inhibitors from natural sources were identified.

P395 / #924

Topic: Theme B: Taupathies / B4.a. Imaging, Biomarkers, Diagnostics: Structural MRI, MR spectroscopy

AUTOMATIC SEGMENTATION USING DEEP LEARNING FOR THE HIPPOCAMPUS

Lecture Title:

J. Weatheritt1, M. Reinwald1, B. Brito Vega1, R. Wolz1,2 1IXICO plc, R&d, London, United Kingdom, 2Imperial College London, Department Of Computing, London, United Kingdom

Aims: Hippocampal volume is used as a biomarker to track the development of Alzheimer’s disease (AD). Therefore, accurate hippocampal segmentation is of utmost clinical importance. Emerging deep-learning technologies can perform at state of the art for many computer vision problems and in this vain we present a fully automated approach that is able to accurately delineate the hippocampus. Methods: We present a deep-learning method, a 3D convolutional neural network (CNN), which can segment the whole brain or be tuned for specific regions. We compare against industry standard methodologies: FreeSurfer (FS) and a recently developed AI version called FastSurfer (F-FS). Manually edited hippocampi segmentations from 80 subjects from an AD cohort (ADNI) are used to develop the model. A further 9 are used for overlap validation scores. Predictions on unsegmented scans are then used to assess the discriminative capability of the method (disease vs control). Results: Figure 1 shows dice overlap scores, summarised in table 1. Figure 2 shows the hippocampi volumes split by cohort. One-sided t-tests for independent cohort means are described in table 1.

Table 1

Method Dice overlap (mean, std) T-test P-value

CNN 0.849, 0.0147 -5.048 2.05e-5

F-FS 0.790, 0.0267 -5.644 1.08e-5

FS 0.779, 0.0228 -4.437 1.37e-4

Conclusions: Our CNN has the highest dice score of the methods presented, with better consistency. Further, the AI methods are shown to have better cohort separation on the unsegmented subjects. These methods are an order of magnitude faster, in terms of compute time, than classical methods, which highlights their promise for AD clinical trials.

P396 / #1156

Topic: Theme B: Taupathies / B4.a. Imaging, Biomarkers, Diagnostics: Structural MRI, MR spectroscopy

THE DEVELOPMENT OF A VALID, RELIABLE, HARMONIZED SEGMENTATION PROTOCOL FOR MEDIAL TEMPORAL LOBE SUBREGIONS: A PROGRESS UPDATE

Lecture Title:

L. Wisse1, R. La Joie2, R. Olsen3, D. Berron4, K. Amunts5, J. Augustinack6, A. Bakker7, A. Bender8, M. Boccardi9, M. Bocchetta10, M. Chakravarty11, G. Chetelat12, R. De Flores12, J. Dekraker13, S.-L. Ding14, R. Insausti15, O. Kedo5, S. Mueller16, N. Ofen17, D. Palombo18, N. Raz19, C. Stark20, L. Wang21, P. Yushkevich22, Q. Yu17, V. Carr23, A. Daugherty19 1Lund University, Diagnostic Radiology, Lund, Sweden, 2University of California San Francisco, Memory And Aging Center, San Francisco, United States of America, 3University of Toronto, Rotman Research Institute, Toronto, Canada, 4Lund University, Department Of Clinical Sciences, Lund, Sweden, 5Forschungszentrum Jülich, Institute Of Neuroscience And Medicine (inm-1), Julich, Germany, 6Harvard University, Radiology, Boston, United States of America, 7Johns Hopkins University, Psychiatry And Behavioral Sciences, Baltimore, United States of America, 8Michigan State University, Neurology & Ophtalmology, East Lansing, United States of America, 9University of Rostock, German Center For Neurodegenerative Diseases, Rostock, Germany, 10Queens Square Institute of Neurology, Dementia Research Centre, London, United Kingdom, 11The Douglas Research Centre, Brain Imaging Centre, Montreal, Canada, 12University of Normandie, Inserm, Caen, France, 13Western University, The Brain And Mind Institute, London, Canada, 14Allen Brain Institute, Allen Brain Institute, Seattle, United States of America, 15University of Castilla La Manch, Human Neuroanatomy Laboratory, Albacete, Spain, 16University of California San Francisco, Radiology, San Francisco, United States of America, 17Wayne State University, Psychology, Detroit, United States of America, 18University of British Columbia, Psychology, Vancouver, Canada, 19Wayne State University, Institute Of Gerontology, Detroit, United States of America, 20University of California Irvine, School Of Biological Sciences, Irvine, United States of America, 21Northwestern University, Psychiatry And Behavioral Sciences And Radiology, Chicago, United States of America, 22University of Pennsylvania, Radiology, Philadelphia, United States of America, 23San Jose State University, Psychology, San Jose, United States of America

Aims: Neurodegenerative pathologies are known to affect medial temporal lobe (MTL) subregions differently, selectively, and in a complex progression. Characterizing MTL subregional changes using high-resolution MRI may provide more insight in disease processes and better biomarkers. However, substantial differences in MTL subregion definitions has hindered the ability to compare results across laboratories or draw robust conclusions. The Hippocampal Subfields Group (HSG) is an international group seeking to address this issue by developing a histologically-valid, reliable, and freely available segmentation protocol for high-resolution T2-weighted 3T MRI (http://www.hippocampalsubfields.com). Methods: Our workflow consists of four steps: 1) collecting histology samples labeled by multiple expert neuroanatomists to form a novel reference dataset to guide the development of the MRI segmentation protocol, 2) developing boundary definitions for each segment of the hippocampus, (head, body, and tail) and MTL cortices on MRI, 3) assessing HSG community agreement with boundary rules using online questionnaires and 4) testing reliability of the protocol definitions. Results: For the hippocampal body and head, we have developed a preliminary subfield segmentation protocol (i.e. completed steps 1-2). For the boundaries of the body with surrounding regions, 29 labs reached consensus for all rules (step 3). We are now administering additional questionnaires for assessing agreement of the hippocampal body and head subregion boundary rules. Upon completion, reliability testing of the protocol should begin end 2020 for the body (step 4). Conclusions: Once completed, the harmonized protocol will significantly facilitate cross-study comparisons thus advancing insight in the role of MTL subregions in aging and dementia.

P397 / #1405

Topic: Theme B: Taupathies / B4.b. Imaging, Biomarkers, Diagnostics: Functional MRI

PRODROMAL CHANGES IN HIPPOCAMPAL FUNCTIONAL CONNECTIVITY IN AD PATIENTS AND THY-TAU22 MICE: A TRANSLATIONAL EXPLORATION.

Lecture Title:

L. Degiorgis1, E. Chabran1, M. Sourty2, M. Karatas1,3, D. Blum4, L. Buee4, L. Harsan1, F. Blanc1,5 1ICube Laboratory - UMR 7357, CNRS, University of Strasbourg, Imis Team, Strasbourg, France, 2Inserm, U1114, Strasbourg, France, 3University Medical Center of Freiburg, Department Of Radiology, Medical Physics, Freiburg, Germany, 4Université de Lille, Inserm, CHU Lille, Umr-s 1172 - Jparc, Labex Distalz, Lille, France, 5Hôpitaux Universitaires de Strasbourg, Geriatrics And Neurology Units, Research And Resources Memory Center (cmrr), Strasbourg, France

Aims: As most of current animal models of AD are based on genetic modifications, their predictive potential for the sporadic form of the disease remains difficult to assess, especially at early stages. In this preliminary study, we compared resting-state functional connectivity (rsFC) changes in the hippocampus of human AD patients with Thy-Tau22 mice, a mouse model of tauopathy. Methods: We conducted seed-based rsFC analyses of hippocampal subregions in 32 prodromal AD patients (MMSE score >24) compared to 19 healthy elderly subjects, and performed the same analysis in 16 Thy-Tau22 mice compared to 13 wild-type mice, at an age when no memory impairment is yet detected in transgenic mice. We then carried out a qualitative comparison of the rsFC changes detected in human patients and Thy-Tau22 mice. Results: We found a similar increase in connectivity of somatosensory areas with the anterior hippocampus in human AD patients, and of olfactory-related regions with the ventral hippocampus in Thy-Tau22 mice. Both cohorts also exhibited a decreased connectivity of the anterior hippocampus with the limbic system. The posterior hippocampus in AD patients and the dorsal hippocampus in Thy-tau22 mice showed a comparable pattern of hyperconnectivity with hippocampal region, amygdala and putamen, as well as with frontal regions. However, several cortical clusters also had a decreased connectivity with the anterior hippocampus in human patients, that we didn’t find in Thy-Tau22 mice. Conclusions: Therefore, we highlighted several similar changes of the hippocampal rsFC between Thy-Tau22 mice at prodromal stages and early AD patients, strengthening the translational potential of this model.

P398 / #347

Topic: Theme B: Taupathies / B4.c. Imaging, Biomarkers, Diagnostics: PET – tau

REGIONAL DISTRIBUTION OF TAU AGGREGATES IN THE BRAIN IS STRONGLY INFLUENCED BY REGIONAL CORTICAL THICKNESS AND CONNECTIVITY

Lecture Title:

J.-P. Soucy1, F. Mohammadi2, P. Rosa-Neto2, T. Pascoal2, O. Ali1, A. Rahimabadi1, M. Savard2, F. Lussier2, M. Kang2, J. Therriault2, H. Benali1 1Concordia University, Perform Centre, Montréal, Canada, 2Douglas Hospital McGill University, Mcgill Centre For Studies In Aging, Montreal, Canada

Aims: Canonical progression of tau in Alzheimer’s is coherent with the hypothesis of its aggregates behaving like prions. Others and we (Soucy et al, HAI 2020) confirmed the impact of connectivity on progression, which also depends on continuing “upstream”, in addition to local, production of tau. Tau accumulation leads to loss of neurons and therefore decreasing upstream tau production, potentially altering the relationship between connectivity and spread. We evaluated the influence of connectivity on cortical thickness (a proxy for tissue loss) correlations and on tau PET SUVRs correlations across regions, looking for differences in the importance of connectivity for each. Methods: 35 ADNI AD subjects with 2 18F-florbetapir scans (Ta, Tb, mean interval 438 days, median 454), and a Ta T1 MRI were studied. To gauge disease severity, we segregated them as non-progressors (NP, 18) and progressors (P, 17) based on interval change of hippocampal SUVRs. We used Moran I statistic to assess impact of connectivity intensity (HCP) on measured autocorrelations between regional cortical thicknesses at Ta, and regional tau PET SUVR values at both times. Regions were defined by thresholding as poorly (< 0.01), moderately (0.01-0.021) or highly (> 0.021) connected. Results: Moran I statistic Z-scores for cortical thickness correlations were higher across regions with high than with moderate/low connectivity. For SUVR values, Z-score were lower for highly connected than for moderately/poorly connectivity regions.

Conclusions: Connectivity’s influence on cross-regional correlations of atrophy and tau SUVR is different, likely because atrophy is a constantly progressive phenomenon, while tau accumulation slows/drops over time.

P399 / #857

Topic: Theme B: Taupathies / B4.c. Imaging, Biomarkers, Diagnostics: PET – tau

18F-THK5351 PET IMAGING, NEUROPATHOLOGY AND CLINICAL PROGRESSION IN A TAU MOUSE MODEL.

Lecture Title:

L. Vegas-Gomez1, G. Edwards Iii2, O. Hasan2, N. Gamez2, J. Schulz2, C. Soto2, P. Schulz2, I. Moreno Gonzalez1,2 1University of Malaga/CIBERNED/IBIMA, Cell Biology, Genetics And Physiology, Malaga, Spain, 2The University of Texas Health Science Center at Houston, Neurology, HOUSTON, United States of America

Aims: Alzheimer’s disease (AD) and other associated dementias remain a consistent and unruly problem for the aging population and health. The neuropathology of AD is characterized by the extracellular deposition of beta-amyloid protein (Aβ) and the formation of intraneuronal neurofibrillary tangles (NFT) composed of hyperphosphorylated tau (ptau), along with neuroinflammation and neuronal loss that ultimately induces to noticeable cognitive impairments. Abnormal ptau leads to the formation of insoluble, beta-sheet rich amyloid aggregates in tauopathies such as AD. Positron emission tomography (PET) imaging is a promising avenue that may identify tau aggregates in vivo cross-sectionally and longitudinally in various dementia conditions. Methods: The goal of this study is to characterize the longitudinal assessment of the tau tracer 18F-THK5351 by in vivo tau PET imaging concomitantly to behavior and tau pathology by histology and biochemistry from 6 to 12 months of age in tau transgenic P301S mice, a mouse model of tauopathies. Results: Our results demonstrate an augmentation of overall gross brain tau pathology by in vivo PET imaging in P301S mice compared to age-matched wild-type (WT) animals accompanied by P301S-model associated pathological tau and phenotypic and behavioral deficits. Conclusions: This longitudinal study provides new insights on the relationship between imaging diagnostic tools, the in vivo neuropathological temporal pattern and the clinical signs observed in animal models of AD that could benefit early disease diagnosis. This work was partially funded by Department of Defense Peer Reviewed Alzheimer’s Research Program Convergence Science. Research Award grant AZ160106 and Alzheimer’s Association New Investigator Research Grant NIRG-394284 to IMG.

P400 / #1368

Topic: Theme B: Taupathies / B4.g. Imaging, Biomarkers, Diagnostics: Multimodal imaging

DETECTION OF POLYMORPHIC PROTEIN AGGREGATES IN NEURODEGENERATIVE DISEASES USING THIOPHENE-BASED LIGANDS

Lecture Title:

T. Klingstedt1, H. Shirani1, B. Ghetti2, R. Vidal2, P. Nilsson1 1Linköping University, Department Of Physics, Chemistry And Biology, Linköping, Sweden, 2Indiana University School of Medicine, Department Of Pathology And Laboratory Medicine, Indianapolis, United States of America

Aims: Luminescent conjugated oligothiophenes (LCOs) have been used to distinguish between alpha-synuclein assemblies in Parkinson’s disease and multiple system atrophy, as well as to reveal aggregate polymorphism in Alzheimer’s disease (AD). In addition, thiophene-based ligands selective for abeta or tau in AD have been presented. In this study, we have examined the structural determinants of ligands for binding to abeta or tau aggregates in AD. Methods: A library of novel thiophene-based ligands was synthesized by combining the thiophene backbone with nitrogen-containing heterocycles. In addition, an uncharged and charged variant of each ligand was included. The selectivity of the ligands towards abeta or tau was evaluated on human AD brain tissue sections by comparing ligand labelling with abeta or tau antibody staining. Results: Previously, by combining the thiophene backbone with the heterocyclic compound benzothiazole, ligands that selectively detected tau were achieved. In this study, the evaluation of the ligand library demonstrated a high selectivity towards abeta plaques. However, by shifting the position of nitrogen in the heterocyclic compounds or by adding charged groups, binding to tau inclusions could be also be seen, or there was a loss of binding to both aggregate types. Conclusions: When designing ligands towards aggregates in AD, the chemical nature of the heterocyclic compound, the position of nitrogen and the overall charge are important determinants for abeta and tau selectivity.

P401 / #1735

Topic: Theme B: Taupathies / B4.g. Imaging, Biomarkers, Diagnostics: Multimodal imaging

BIOLOGICAL SUBTYPES OF ALZHEIMER’S DISEASE: FROM CONCEPTUALIZATION TO OPERATIONALIZATION

Lecture Title:

R. Mohanty1, D. Ferreira2, E. Westman3 1Karolinska Institutet, Department Of Neurobiology, Caring Sciences And Society, Stockholm, Sweden, 2Karolinska Institutet, Division Of Clinical Geriatrics, Center For Alzheimer Research, Department Of Neurobiology, Care Sciences And Society, Stockholm, Sweden, 3Karolinska Institutet, Division Of Clinical Geriatrics, Department Of Neurobiology, Care Sciences And Society, Stockholm, Sweden

Aims: Biomarkers capture the heterogeneity within Alzheimer’s disease (AD). For the first time, a conceptual framework presented typicality and severity as two principal dimensions of biological subtypes (limbic-predominant, hippocampal-sparing, typical-AD, minimal-atrophy) of AD1 (Figure 1A). Here, we operationalize this framework for atrophy-based and tau-based subtypes. Methods: We included structural magnetic resonance imaging (MRI) and concurrent tau positron emission tomography (PET) from 368 individuals: 173 amyloid-beta positive individuals in the AD continuum (25 AD dementia, 50 prodromal AD, 98 preclinical AD) and 195 amyloid-beta negative healthy individuals. We quantified subtypes on a continuous scale: (1) typicality was computed by the hippocampus to cortex ratio in atrophy (MRI) and tau uptake (tau PET); (2) severity was computed as brain volume to cerebrospinal fluid ratio (MRI) and global tau uptake (tau PET). We also quantified the disease stage by estimating the Braak stage for each individual based on their regional tau uptake. Results: We operationalized the conceptual framework (Figure 1A) by measuring the typicality and severity dimensions on a continuous scale for atrophy-based and tau-based subtypes (Figure 1B) and observed: (a) subtypes based on atrophy and tau have notably distinct distributions across the two dimensions; (b) Braak stage corresponds more closely with severity than typicality; (c) subtypes, primarily found in AD, could potentially be extended to the AD continuum.

Conclusions: Operationalization of the conceptual framework for subtypes can: (a) compare correspondence of subtypes from different modalities; (b) differentiate subtypes from disease stage, especially based on typicality, (c) compare subtypes in AD and extend to the AD continuum.

P402 / #401

Topic: Theme B: Taupathies / B4.h. Imaging, Biomarkers, Diagnostics: CSF, blood, body fluid biomarkers

P. BOTELLA*, C. LODDER*, M. A GUTIERREZ DE RAVE*, JOYCE HABETS, SARAH VANHERLE, D. TERWEL, I.C. STANCU & I.DEWACHTER

Lecture Title:

P. Botella Lucena, C. Lodder, M.Á. Gutiérrez De Ravé, J. Habets, S. Vanherle, D. Terwel, I.-C. Stancu, I. Dewachter Hasselt University, Biomedical Research Institute - Neuroscience, Diepenbeek, Belgium

Aims: This study aims to characterize changes of Abeta and tau in the serum of APP.PS1(F)/tau(T) double transgenic mice (further denoted F+/T+) and their parental strains (F+, T+, F-/T-) in relation to the development of pathology in the brain. Methods: Serum samples were collected from F+, T+, F-/T- and F+/T+ mice at different time points to study the correlation with pathology. The parental T+ mice develop tau pathology and associated neurodegenerative phenotype from ~11 months onwards, while F+/T+ develops combined amyloid and tau-pathology at the age of 9 months. The parental F+ mice display robust amyloid pathology from 5-6 months onwards. To correlate blood based biomarkers concentrations of Ab and tau in serum were measured using Meso Scale Discovery (MSD). Results: t-tau levels in serum were significantly increased at 11 months of age in T+ mice, correlating with pathology. F+ mice showed significantly decreased Ab-42 levels at 5-6 months of age, correlating with robust amyloid pathology. Analysis in F+/T+ mice with combined pathology is ongoing, and detailed analysis of A-beta and tau dynamics is ongoing in all strains. Conclusions: Changes in A-beta and tau concentrations in serum reflect pathological processes in the brain, thereby serving as a blood-based biomarker.

P403 / #1759


TRANSCRIPTOMIC AND CLINICAL BIOMARKERS FOR AN EARLIER AND MORE ACCURATE DIAGNOSIS OF PROGRESSIVE SUPRANUCLEAR PALSY

Lecture Title:

M. D'Onofrio1, I. Arisi2, R. Brandi1, M. Sonnessa3, A. Cattaneo4, P. Bertolazzi5, M. Torti6, L. Vacca6, F. Stocchi6 1European Brain Research Institute (EBRI) Rita Levi-Montalcini, Genomics Facility, Roma, Italy, 2European Brain Research Institute (EBRI) Rita Levi-Montalcini, Bioinformatics, Roma, Italy, 3Bio-Fab Research srl, Bioinformatics, Roma, Italy, 4Scuola Normale Superiore (SNS), Laboratorio Di Biologia Bio@sns, Pisa, Italy, 5CNR-IASI, Bioinformatics, Roma, Italy, 6IRCCS San Raffaele Pisana, Center For Parkinson's Disease, Neurology Department, Roma, Italy

Aims: Progressive Supranuclear Palsy (PSP), atypical parkinsonism less common and usually more severe than classical PD, has been increasingly recognized to encompass a spectrum of clinical phenotypes involving behavioral, language, and movement abnormalities. Early and reliable diagnosis remains a major clinical challenge, crucial for prognosis, stratification, and therapeutic trials. Disease-specific biomarkers have yet to be identified. We aim at the identification of innovative transcriptomic biomarkers and clinical metrics to: -Improve diagnosis by integration of Next Generation Sequencing (NGS) data with clinical records; -Improve patients’ stratification. Methods: Subjects were enrolled after Informed Consent and Ethical approval at IRCCS San Raffaele La Pisana (Roma, Italy). The population is represented by 16 controls (CTRL) and 25 PSP subjects with Richardson Syndrome (PSP-RS) or Parkinsonism PSP (PSP-P). Peripheral blood samples were analyzed by RNA-seq. NGS transcriptomic data were analyzed by the Trimmomatic-Hisat-Stringtie pipeline and R-Bioconductor. Genomic variants (SNPs) were analyzed by the Opossum-GATK/Platypus/bcftools pipeline and clinical data by R-Bioconductor. Results: We identified tens of Differentially Expressed Genes (DEGs) in PSP-P vs PSP-RS, together with a set of SNPs significantly over-or under-represented in CTRL-PSP comparison. DEGs are related to Ubiquitin mediated proteolysis, Protein Kinase, and Transcription, while SNPs correspond mainly to immune system response genes. Specific cognitive (MOCA, NMS) and motor scales (ocular, limb) which discriminate PSP-RS and PSP-P are correlated with gene expression. Conclusions: We selected a brief list of potential transcriptomic biomarkers to discriminate PSP-RS and PSP-P subtypes and to better characterize the peripheral phenotype by integration with clinical metrics.

P404 / #423


SIMULTANEOUS MEASUREMENT OF SITE-SPECIFIC TAU PHOSPHORYLATIONS IN BLOOD FOR EARLY AND ACCURATE DIAGNOSIS OF ALZHEIMER´S DISEASE

Lecture Title:

L. Montoliu-Gaya1, N. Ashton1, M. Sauer1, J. Lantero Rodriguez1, G. Brinkmalm1, T. Karikari1, S. Kern2, H. Zetterberg1, K. Blennow1, J. Gobom1 1The Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Institute Of Neuroscience & Physiology, Mölndal, Sweden, 2Sahlgrenska Academy, Centre for Ageing and Health (AGECAP) at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Neuropsychiatric Epidemiology Unit, Institute Of Neuroscience & Physiology, Mölndal, Sweden

Aims: A reliable blood test could represent the first simple and scalable test for the diagnosis of Alzheimer´s disease (AD). Several studies have explored the potential of Aβ, t-tau and NfL as blood-based biomarkers for AD, but their implementation may be compromised by peripheral expression (Aβ), poor correlation with CSF (T-tau) and low specificity for AD (NfL). Recently, immunoassays targeting tau phosphorylated species p-tau181 and p-tau217 have shown promising results, proving to be reliable tools for AD diagnosis and correlating well with in vivo assessment of Aβ and tau pathology. However, the lack of sensitive methods that can simultaneously quantify phosphorylation at multiple positions of the tau protein, has limited understanding of the tau phosphorylation pattern in blood. Methods: Mass spectrometry (MS) techniques allow a broad characterization throughout the protein, with high specificity and quantitative precision. Results: We developed a highly sensitive and specific MS method using parallel reaction monitoring (PRM) to quantitatively assess the degree of phosphorylation at specific sites on tau from plasma. This highly optimized method, enables us to measure tau phosphorylation at amino acids T181, S202, T217 and T231, simultaneously in 1ml plasma samples. Conclusions: Concurrent characterization of plasma p-tau with our novel method will help to precisely determine which phosphorylation site is the earliest biomarker and most accurate indicator of tau pathology. Clinically, the method may prove useful to stage AD, distinguish among neurodegenerative diseases and monitor treatment effects, in a set affordable and available for large patient population.

P405 / #1818


BLOOD-BASED SIGNATURES OF ALZHEIMER’S AND PARKINSON’S DISEASES OBSERVED THROUGH UBIQUITIN PROTEASOME SYSTEM

Lecture Title:

K. Kadimisetty, J. Robustelli, B. Darnay, P. Gross, K. Sheets, A. Burriss, M. Zeleski LifeSensors Inc., Research And Development, Malvern, United States of America

Aims: Identify unique blood-based signatures for AD/PD could potentially facilitate early intervention delaying the onset of clinical symptoms and improve patients’ quality of life. Methods: Genetic and biochemical evidence suggest that dysfunction in the ubiquitin proteasome system (UPS) lies at the heart of several neurodegenerative diseases, including Alzheimer’s and Parkinson’s diseases (AD/PD). Ubiquitin-mediated signaling plays a central role in controlling protein degradation, regulating receptor recycling, and initiating mitophagy and autophagy. Neurons remove neurofibrillary tangles such as polymerized beta-amyloid, tau, or alpha-synuclein aggregates by autophagy/mitophagy. Signatures of AD/PD appear long before the disease manifests. LifeSensors has developed highly sensitive methods to detect signatures of ubiquitination from human blood and CSF. The breakthrough is achieved by engineering TUBEs (Tandem Ubiquitin Binding Entities) to recognize pan-poly-ubiquitinated proteins and linear, K6, K11, K33, K48, and K63 selective poly-ubiquitinated proteins. TUBE-based mass spectrometry data from patients’ blood serum and CSF reveals unique changes in levels of ubiquitylated proteins when compared to age-matched controls. Results: The data suggest, monitoring the levels of unique ubiquitylated proteins as screening panel, obtained from TUBE-based screening can potentially be used to diagnose AD/PD at early stages. We demonstrated unique changes in ubiquitylation signatures from mass spectrometry data obtained from pulldowns of chain selective (-K48 and -K63) and atypical TUBEs (-K6) to further characterize the role of the ubiquitylated markers identified. Conclusions: This study overall focuses on demonstrating evidence for the role of the UPS in the pathogenesis of AD/PD along with using the identified markers for accurate early testing.

P406 / #708

Topic: Theme B: Taupathies / B4.j. Imaging, Biomarkers, Diagnostics: Cognitive, psychometric & behavioral tests

COGNITIVE IMPAIRMENT IN PSP RELATES TO DEGENERATION OF THE NUCLEUS BASALIS OF MEYNERT

Lecture Title:

S. Rogozinski1, G. Respondek1, M. Grothe2,3, J.B. Pereira4,5, G. Höglinger1,6 1Hannover Medical School, Department Of Neurology, Hannover, Germany, 2Lund University, Clinical Memory Research Unit, Department Of Clinical Sciences, Malmo, Sweden, 3Karolinska Institute, Division Of Clinical Geriatrics, Department Of Neurobiology, Care Sciences And Society, Stockholm, Sweden, 4(DZNE), German Center For Neurodegenerative Diseases, Rostock, Germany, 5Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, Unidad De Trastornos Del Movimiento, Servicio De Neurología Y Neurofisiología Clínica, Instituto De Biomedicina De Sevilla, Seville, Spain, 6(DZNE), German Center For Neurodegenerative Diseases, Munich, Germany

Aims: To study the gray matter (GM) volumes of the nucleus basalis of Meynert (nbM) in different parkinsonian syndromes and assess their relationship with clinical variables. Methods: We included clinical data and T1-weighted magnetic resonance images from patients with progressive supranuclear palsy (PSP, N=43), multiple system atrophy - parkinsonian type (MSA-P, N=15), multiple system atrophy - cerebellar type (MSA-C, N=8), Parkinson´s disease (PD, N=26) and healthy controls (HC, N=29). T1-weighted images were analyzed using a voxel-based morphometry approach implemented in the VBM8 toolbox, and nbM volumes were extracted from the spatially normalized GM images using a cytoarchitectonically-defined nbM mask in stereotactic standard space. We compared nbM volumes between groups, while controlling for age, sex and intracranial volume. We performed correlation analyses between nbM volumes and the Mini Mental Status Examination (MMSE), Hoehn and Yahr stage, PSP Rating Scale, Unified Parkinson's Disease Rating Scale part III and Frontal Assessment Battery scores within each group. Results: We found significantly lower nbM volumes in patients with PSP and PD as compared to HC or patients with MSA. In PSP patients, we found a significant correlation between MMSE and nbM volumes. No other significant correlations were found between clinical scores and the NbM volumes in PSP or other groups. Conclusions: Our results suggest that cognitive impairment in PSP relates to degeneration of the nbM, which has multiple cholinergic projections to subcortical and cortical structures. This highlights the need for future clinical trials assessing the efficacy of cholinergic drugs for treating cognitive impairment in PSP.

P407 / #864

Topic: Theme B: Taupathies / B4.k. Imaging, Biomarkers, Diagnostics: Other

APPLICATION OF PROTEIN MISFOLDING CYCLIC AMPLIFICATION TO THE DETECTION OF TAU AGGREGATED SPECIES FROM TAU TRANSGENIC MOUSE MODELS

Lecture Title:

M. Bélondrade1, F. Almela1, R. Caillierez2, L. Buee2,3, M. Colin2, D. Bougard1 1Pathogenesis and Control of Chronic Infections UMR1058, Etablissement Français Du Sang, Montpellier, France, 2Univ. Lille, Inserm, CHU-Lille, Lille Neuroscience & Cognition, F-59000 Lille, France., Alzheimer & Tauopathies, Lille Cedex, France, 3Inserm UMR-S 1172 Lille Neuroscience & Cognition, Alzheimer & Tauopathies, Lille, France

Aims: The spreading mechanism of tau pathology is reminiscent of prion diseases in terms of misfolding induction and propagation. Largely used for diagnosis and understanding of prion diseases, this autocatalytic conversion process has been exploited in Protein Misfolding Cyclic Amplification (PMCA) that reproduces and accelerates this phenomenon in vitro. Our objective is to develop the amplification of misfolded tau proteins by adapting PMCA to the detection of tau seeding activity. Methods: We used a transgenic mouse line harbouring the double mutations P301S and G272V (THY-tau30) and presenting a progressive apparition of tau lesions during aging. Brain homogenates from 3 and 12 months THY-tau30 mice were submitted to PMCA using different physical and chemical parameters to establish the proof of principle of sonication-based aggregates amplification. Newly-formed tau aggregates were evaluated using Western blotting as well as a commercially aggregated tau-specific HTRF kit. Results: Our results showed an important increase of aggregated tau species from THY-tau30 mice after PMCA in comparison with KOTau and wild type C57Bl6 mice. This obtained effect of sonication/incubation cycles on tau aggregation was correlated with the age of mice. Western blot analysis of PMCA amplified tau proteins after protease digestion revealed two low molecular weight bands that appeared to be specific of tau aggregates. Conclusions: We have established the feasibility to induce aggregation of tau protein in vitro by using PMCA reaction in a cellular complex milieu, opening the possibility to detect a seeding activity from human samples from patients with tauopathies.

P408 / #1306

Topic: Theme B: Taupathies / B5.b. Genetics, Epidemiology: Disease-causing mutations

EXTENSIVE GENETIC AND PHENOTYPIC DESCRIPTION OF MAPT P.R406W IN THE FLANDERS-BELGIAN POPULATION

Lecture Title:

H. Gossye1,2,3,4, S. Van Mossevelde3,4, J. Van Der Zee4, Y. Vermeiren5, A. Sieben5,6,7, N. De Roeck7, P. Cras3, S. Engelborghs5,7,8,9, P. De Deyn4, C. Van Broeckhoven4,10 1Institute Born-Bunge, Laboratory For Neurogenetics, Antwerpen, Belgium, 2University Hospital Brussel and Center for Neurosciences, Department Of Neurology, Brussels, Belgium, 3University Hospital Antwerp, Neurology, Wilrijk, Belgium, 4VIB, Nbd, Antwerpen, Belgium, 5University of Antwerp, Laboratory Of Neurochemistry And Behaviour, Wilrijk, Belgium, 6Gent University Hospital, Neurology Department, Gent, Belgium, 7Antwerp, Institute Born-bunge, Antwerp, Belgium, 8University Hospital Brussels and University Center for Neurosciences VUB, Department Of Neurology, Brussels, Belgium, 9University of Antwerp, Department Of Biomedical Sciences, Antwerp, Belgium, 10University of Antwerp, Vib Center For Molecular Neurology, Antwerpen, Belgium

Aims: The missense mutation p.R406W in the MAPT gene is associated with frontotemporal lobar degeneration (FTLD) pathology and an atypical, Alzheimer’s disease (AD)-like phenotype. In our Flanders-Belgian patient cohort, we identified 10 p.R406W carriers. Of 3 index carriers, we sampled family members carrying the mutation, generating a total cohort of 55 p.R406W carriers. Our aim was to analyze phenotypical and genetic characteristics. Methods: From longitudinal follow-up over 19 years, we obtained data on clinical characteristics as well as neuropathology. We investigated the potential modifying effect of the MAPT H1/H2 and the APOE genotypes on the phenotype. Results: Of 55 carriers, 39 were symptomatic. Allele and haplotype sharing analysis confirmed a genetic kinship among all, suggesting a common ancestor. Average onset age and disease duration were 59.8 and 12.7 years (ranges 40-75, 5-25). The most frequent diagnoses were dementia (unspecified) (43.6%), AD (28.2%) and behavioral variant frontotemporal dementia (bvFTD) (25.6%). Presenting symptoms commonly included disinhibition and behavioral problems in all groups (72.7%). Remarkably, CSF biomarker profiles often showed decreased Aβ1-42 and Aβ1-42/ Aβ1-40 ratio, and elevated P-tau and T-tau. Neuropathology was FTLD-tau. We observed a significantly shorter disease duration in carriers of an APOE ε4 allele compared to non-carriers.

Conclusions: We observed a nonconforming clinical phenotype of MAPT p.R406W carriers in the Flemish-Belgian cohort with 25.6 % presenting a bvFTD phenotype. Contrary to previous reports, prominent behavioral symptoms were highly frequent in the entire cohort (72.7%). CSF biomarkers commonly showed AD-like abnormalities. The presence of an APOE ε4 allele shortened disease duration significantly.

P409 / #1196

Topic: Theme B: Taupathies / B5.g. Genetics, Epidemiology: Other

PHARMACOLOGICAL TREATMENT OF ALZHEIMER’S DISEASE PATIENTS IN CATALONIA DURING 2019

Lecture Title:

J.Á. Expósito Losada, M. Umbria Vivancos, M. Obach Cortadelles, A. Prat Casanovas, A. Vallano Ferraz, C. Pontes García Catalan Health Service, Medicine Area - Pharmacotherapeutic Harmonization Management, Barcelona, Spain

Aims: To describe the clinical, sociodemographic characteristics and treatment of Alzheimer's Disease (AD) patients in Catalonia during 2019. Methods: The data source was the Registry of Alzheimer’s Disease Treated Patients and the Pharmaceutical Reimbursement File of the Catalan Health Service. The study population was defined as patients who initiated, followed, switched or discontinued AD drugs throughout 2019. GDS-FAST score was collected at the initiation of antidementia drug and at the last follow-up visit. Results: A total of 51,350 patients (66.7% women) received treatment for AD and 84.7% were still treated by the end of the year. A prevalence of 674/100,000 was estimated. The proportion of treated patients increased with age, being 77.1% over 74 years old. Most of the patients received AChEI for at least two years, mainly as monotherapy (97%). Donepezil (41%) was the most widely drug used. A total of 15.3% patients discontinued AD treatment primarily due to exitus. The 62.1% of subjects presented an initial GDS-FAST score of 4, while an increased to 5 or 6 was observed at the last follow up visit. AD pharmacological expenditure was 34M€ which represents 2.05% of the overall drug prescription budget in 2019. Conclusions: AD treatment is mainly driven by the use of AChEI in monotherapy in elderly Catalan population. Even though the effectiveness of antidementia drugs is uncertain and often deemed modest, it represents a relevant expenditure within the overall pharmaceutical billing. A close monitoring of health outcomes in treated AD population may help to reassess the place in therapy and budget prioritization.

P410 / #362

Topic: Theme B: Taupathies / B6.a. Cell, Molecular and Systems Biology: Tau, tau isoforms

ISOFORM-SPECIFIC TAU ANALYSIS OF HUMAN BRAIN USING A HIGH-RESOLUTION MASS SPECTROMETRY MULTIPLEX ASSAY

Lecture Title:

G. Brinkmalm1, J. Lantero Rodriguez1, E. Camporesi1, L. Montoliu-Gaya1, T. Karikari1, M. Olsson1, I. Burmann1, T. Lashley2, H. Zetterberg1, K. Blennow1 1Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden, 2UCL Institute of Neurology, Department Of Neurodegenerative Disease, London, United Kingdom

Aims: Alzheimer’s disease (AD), progressive supranuclear palsy (PSP), Pick’s disease (PiD), and corticobasal degeneration (CBD), are diseases where the protein tau is affected. Neuronal tau exists in six major isoforms that are expressed to different degree during an individual’s development. Different tauopathies exhibit structures where the relative amounts of three repeat (3R) and four repeat (4R) tau vary; 3R tau dominates in PiD and 4R in PSP and CBD. Currently, there are no good isoform-specific assays and hence it is problematic to chemically characterize the diseases. Our aim was to develop a mass spectrometry-based assay for different tau isoforms in brain and cerebrospinal fluid (CSF). Methods: CSF, soluble (tris-buffered saline, TBS), and sarkosyl insoluble brain extracts were immunoprecipitated using 5 different antibodies. More than 20 tryptic peptides were monitored by liquid chromatography/high-resolution mass spectrometry with isotope-labelled protein standards for quantification. Results: In brain all tau isoforms were detected, while in CSF only N-terminal peptides were observed. In all samples, the 1N isoform was most abundant followed by 0N and 2N. In brain, 3R was more abundant in PiD, 4R in PSP and CBD, while in AD and controls they had similar abundance. In the TBS fraction, tau was markedly reduced in AD compared with controls, while in the sarkosyl insoluble fraction, tau was increased in AD, PSP, and CBD, but decreased for PiD compared with controls. Conclusions: The assay is reproducible and can distinguish isoform differencens in taopathies. Data from a larger study is under way and will be presented.

P411 / #558

Topic: Theme B: Taupathies / B6.a. Cell, Molecular and Systems Biology: Tau, tau isoforms

ELUCIDATING THE ROLE OF TAU ISOFORM EXPRESSION IN HUMAN IPSC-DERIVED TAUOPATHY MODELS

Lecture Title:

A. Dannert1,2, J. Klimmt1,2, D. Paquet1,2,3,4 1University Hospital, LMU Munich, Institute For Stroke And Dementia Research, München, Germany, 2Ludwig Maximilians Universität München, Graduate School Of Systemic Neurosciences, Planegg-Martinsried, Germany, 3Ludwig-Maximilians-Universität München, Munich Cluster For Systems Neurology (synergy), Munich, Germany, 4ISAR Bioscience GmbH, Neurodegeneration Program, Planegg, Germany

Aims: Malfunction of the protein Tau is a hallmark of neurodegenerative Tauopathies such as Alzheimer’s disease and Frontotemporal Dementia (FTD) with Tau mutations causing familial FTD. Tau expression and splicing are highly regulated and misregulation of the two classes of splice isoforms, 3R and 4R Tau, leads to FTD. Despite its importance in physiology and disease, the majority of current Tauopathy models does not recapitulate adult human Tau isoform expression at a 3R to 4R ratio of 1:1. Furthermore, it remains challenging to reproduce advanced hallmarks of Tauopathy in vitro. We therefore hypothesize that adult human 3R:4R isoform expression might be a prerequisite to recapitulate key pathological phenotypes of Tauopathy. Methods: We developed a CRISPR/Cas9 genome editing strategy to alter endogenous Tau isoform expression in induced pluripotent stem cell (iPSC)-derived cortical neurons. In addition, we included pathogenic Tau mutations to investigate disease phenotypes. We culture these neurons in 2D as well as in 3D culture. Results: We observed that our edited neurons indeed express a 1:1 ratio of 3R and 4R Tau isoforms on both protein and RNA level much earlier than iPSC-derived wild type neurons. Currently, we investigate the effect of pathogenic Tau mutations in a background of adult-like human 3R:4R Tau isoform expression. Conclusions: Using CRISPR/Cas9 genome editing, we can force expression of 4R Tau already in young iPSC-derived neurons. We will use this model to investigate isoform-specific differences of Tau biology and pathology.

P412 / #747

Topic: Theme B: Taupathies / B6.c. Cell, Molecular and Systems Biology: Posttranslational modifications

PHOSPHORYLATION OF MICROTUBULE-ASSOCIATED PROTEIN TAU REGULATES ITS ABILITY TO BIND AND POLYMERIZE MICROTUBULES IN MICE BRAINS

Lecture Title:

A. Hagita, S. Wada-Kakuda, M. Nobuhara, N. Kakuda, T. Miyasaka Doshisha University, Neuropathology, Kyotanabe-shi, Kyoto, Japan

Aims: Microtubule (MT) -associated protein tau constitutes a core of insoluble inclusions, formed in the affected neurons in the brains of dementia, tauopathy. The tau abnormally deposited in the inclusions were hyperphosphorylated and known to be lost its functions. So, it is considered that the phosphorylation of tau affects its functions onto MTs. Actually, there have been no evidence that phosphorylation regulates the physiological function of tau in vivo. Here, we tried to develop the procedure to quantify the binding of tau on MTs, and verified the effects of the tau phosphorylation on its physiological functions in mouse brain. Methods: We optimized the conventional fractionation method of tubulin/MT for brains to quantify stable-MTs, labile-MTs, and free tubulins. Dynamics of tubulins and MAPs were evaluated by Western Blotting. Anesthesia-induced-hypothermia model was used for the transient tau hyperphosphorylation model in adult mice. Postnatal brain was used as a model of physiological tau phosphorylation in vivo. Results: Our novel procedure showed that the nearly 90% of tau and tubulin were fractioned into labile- and stable-MTs in mice brains. Hypothermia-induced- phosphorylated tau became free from labile-MTs and lost its functions of MT-polymerization, however the amounts of tau/stable-MTs complex were not affected. During brain maturation in postnatal-stages, tau was phosphorylated in the first week of life, then dephosphorylated until P14. According with its dephosphorylation, tau gradually assembled into MTs simultaneously with stable-MTs formation. Conclusions: These findings suggest that phosphorylation regulates the functions of tau on MTs in mice brains.

P413 / #1041

Topic: Theme B: Taupathies / B6.f. Cell, Molecular and Systems Biology: Network biology, connectome, protein-protein interations

SERPINA5, A NOVEL TAU BINDING PARTNER, ACCUMULATES PROGRESSIVELY IN DYING TANGLE-BEARING NEURONS IN ALZHEIMER’S DISEASE

Lecture Title:

C. Moloney1, A. Crist1, K. Hinkle1, X. Wang2, E. Lesser2, N. Azu1, I. Frankenhauser1,3, S. Labuzan1, B. Matchett1, M. Deture1, R. Petersen4, R. Duara5, N. Graff-Radford6, M. Allen1, M. Carrasquillo1, H. Li7, O. Ross1, N. Taner1,6, D. Dickson1, Y. Asmann2, R. Carter2, M. Murray1 1Mayo Clinic, Department Of Neuroscience, Jacksonville, United States of America, 2Mayo Clinic, Department Of Health Sciences Research, Jacksonville, United States of America, 3Paracelsus Medical Private University, Neurology, Salzburg, Austria, 4Mayo Clinic, Department Of Neurology, Rochester, United States of America, 5Mount Sinai Medical Center, Wien Center For Alzheimer's Disease And Memory Disorders, Miami Beach, FL, United States of America, 6Mayo Clinic, Department Of Neurology, Jacksonville, United States of America, 7Mayo Clinic, Department Of Molecular Pharmacology And Experimental Therapeutics, Jacksonville, United States of America

Aims: Utilizing RNA-sequencing to uncover molecular underpinnings of hippocampal vulnerability in Alzheimer’s disease (AD), we identified the novel tau interactor, SERPINA5 to be upregulated in AD. Based upon preliminary findings, we hypothesized that SERPINA5 may play a role in neurofibrillary tangle maturity. Thus, we sought to objectively assess SERPINA5 immunoreactivity across cell types and within the context of tangle maturity (i.e. pretangles, mature tangles, and ghost tangles). Methods: Co-immunofluorescence was performed to examine SERPINA5 co-localization with cell type and tau accumulation in neurofibrillary tangles. Immunohistochemistry was performed in the hippocampus of AD cases (n=60) and nondemented controls (n=20). We developed a method to manually count tangle maturity levels in the hippocampus using digital scans of SERPINA5 immunohistochemical stains. Results: SERPINA5 co-localized with a neuronal marker, but not with other cell type markers. Additionally, SERPINA5 co-localized in tau-positive tangle-bearing neurons. Pretangles displayed diffuse or granular staining that was less intense than mature tangles. Mature tangles displayed intense staining throughout the entire neuron. Ghost tangles displayed loosely arranged bundles of fibers. Of all SERPINA5-positive tangles, 4% were pretangles, 52% were mature tangles, and 42% were ghost tangles (p<0.001). Co-immunopreciptiation experiments revealed evidence of tau-SERPINA5 binding in hippocampus and cortex.

Conclusions: SERPINA5 is present in neurons with neurofibrillary tangles and primarily associates with later neurofibrillary tangle maturity levels. Future studies will test the hypothesis that SERPINA5 acts as “tipping point” toward the disease state of a tangle-bearing neuron.

P414 / #967

Topic: Theme B: Taupathies / B6.g. Cell, Molecular and Systems Biology: Metabolomics, transcriptomics, lipidomics, proteomics

NOVEL PATHWAY ENRICHMENT AND NETWORK ANALYSIS METHODS FOR INVESTIGATING CELL-TYPE SPECIFIC SELECTIVE VULNERABILITY TO PATHOLOGICAL TAU IN ALZHEIMER'S DISEASE

Lecture Title:

E. Turkes, K. Duff University College London, Uk Dementia Research Institute, London, United Kingdom

Aims: Region-specific neuronal subpopulations known to be selectively vulnerable and resistant to tau pathology, as characterized in our previous work (Fu et al., 2019), were identified across several public single-cell/nucleus RNA seq datasets and analysed using novel bioinformatic methods to better understand the factors underlying selective vulnerability. Methods: In this poster, we show an analysis conducted using wild-type mouse SMART-Seq data from the Allen Brain Institute (Hodge et al. 2019) and human AD case/control 10X data from the Kampmann group (Leng et al. 2020). Following a standard preprocessing and exploratory analysis pipeline, we applied a custom differential pathway enrichment and network analysis pipeline that identifies features that covary with the vulnerability status of a cell-type. Results: We show that using known marker genes, we were able to identify highly specific clusters of cells that correspond to cell-types that exhibit selective vulnerability. Our novel pathway/network analysis pipeline also recapitulates known endpoints of AD pathology, such as the robust downregulation of synaptic pathways and genes. Most importantly, however, we demonstrate that through trajectory inference across multiple datasets, we can identify putative pathways and genes that underlie selective vulnerability to said endpoints. Conclusions: We present a novel pathway enrichment and network analysis pipeline and demonstrate its application in single cell/nucleus RNA seq data for uncovering pathways and genes that are associated with selective vulnerability in Alzheimer's Disease.

P415 / #1533


ELUCIDIATING CELLULAR CONTRIBUTIONS TO TAU-MEDIATED NEURODEGENERATION USING DROSOPHILA AND SINGLE CELL TRANSCRIPTOMICS

Lecture Title:

T. Wu1,2, C. Guo3, J. Shulman1,3,4,5 1Baylor College of Medicine, Genetics And Genomics, Houston, United States of America, 2Baylor College of Medicine, Medical Scientist Training Program, Houston, United States of America, 3Baylor College of Medicine, Neuroscience, Houston, United States of America, 4Baylor College of Medicine, Neurology, Houston, United States of America, 5Texas Children's Hospital, Jan And Dan Duncan Neurological Research Institute, Houston, United States of America

Aims: Characterizing longitudinal, age-related changes in Alzheimer’s dementia (AD) is of particular importance because clinical manifestation of AD is a decades-long process, and age is the strongest known risk factor. We use a highly tractable, longitudinal, Drosophila model of tauopathy and single cell transcriptomics to explore cell-specific contributions to tau-mediated gene expression while accounting for age-related changes. Methods: Whole brains of adult Drosophila pan-neuronally expressing a mutant variant of human Tau (R406W) and controls were dissociated for droplet single cell transcriptomics (10x Genomics Chromium) at 1-, 10-, and 20- days of age. Perturbations in gene expression and cellular composition were quantified. To address cross-species conservation of tau-mediated scRNAseq signatures, we leverage existing human AD brain scRNAseq data. Results: We observe tau-induced activation of Nuclear Factor kappa B (NFkB) innate immune signaling across distinct neuronal and glial subtypes. As expected, we highlight evidence for tau-mediated degeneration of Kenyon cells, a neuronal population in the Drosophila “mushroom body” that is required for learning and memory, along with a concomitant increase in glial cell numbers (e.g. gliosis). Gene set enrichment analysis reveal overrepresentation of acetylcholine receptor regulation pathways in vulnerable mushroom body neurons. Interestingly, regularized regression analyses highlight innate immune gene expression as predictive of tau-related neuronal reduction. Cluster-specific transcriptional signatures between flies and humans demonstrate strong correlations among inferred cell identities. Conclusions: Our results comprise a powerful single cell transcriptomic resource for studying tau-mediated disruption of gene expression and dynamic age-dependent changes at cellular resolution in a tractable model system.

P416 / #1762


DYSFUNCTIONAL GLUCOSE METABOLISM AND FAST PROGRESSION OF ALZHEIMER DISEASE.

Lecture Title:

S. Zafar1,2, I. Zerr2,3 1SMME, National University of Sciences and Technology (NUST), Islamabad, Pakistan, Biomedical Engineering And Sciences Department, Islamabad, Pakistan, 2Prion Research Group, University Medical Centre Goettingen (UMG), Neurology, Goettingen, Germany, 3DZNE, German Center For Neurodegenerative Diseases, Göttingen, Germany

Aims: Dysfunctional glucose metabolism and Type 2 diabetes mellitus involving insulin resistance and could be a main risk contributor in Alzheimer disease (AD).The early and accurate biomarker of neuronal atrophy and dysfunction in AD is a diminution of glucose uptake in the brain. Brain is one of the major energy-demanding organs in the human body and it constitutes only 2% of the total body mass.The essential energy substrate to sustain neuronal activity is glucose and is taken up via glucose transporters expressed in the brain endothelium, astrocytes and neurons. our main aim was to identified and characterized impaired cerebral glucose metabolism which is an invariant pathological feature of AD suggested by accumulating evidence. Before the occurrence of cognitive dysfunction and histopathological alterations, this phenomenon is proposed to contribute to disease progression and the manifestation of clinical symptoms, even decades. Methods: Here, we identified disease progressive variations in the sporadic AD (spAD), rapidly progressive AD (rpAD), as well as control cases utilization of advanced Sequential Window Acquisition of all Theoretical Mass Spectra (SWATH-MS) a specific variant of data-independent acquisition (DIA) method based proteomics. Results: A Total of 668 proteins were identified and quantitated and 17 proteins were specifically showed different levels specifically in rpAD subjects. Interestingly, in our rpAD dataset selectively, we identified an altered expression level of proteins involved in glucose metabolism in comparision with with spAD cases. Conclusions: We substantiate that the aberrant glucose linked networks are a specific phenotype of brain with rapid decline and fast progression of AD.

P417 / #1017

Topic: Theme B: Taupathies / B6.h. Cell, Molecular and Systems Biology: Epigenetics, histone modification, DNA methylation

THE INHIBITION OF LSD1 VIA SEQUESTRATION CONTRIBUTES TO TAU-MEDIATED NEURODEGENERATION

Lecture Title:

Y. Bai1, A. Engstrom1, A. Walker2, R. Moudgal3, D. Myrick1, S. Kyle1, M. Christopher4, D. Katz1 1Emory University, Cell Biology, Atlanta, United States of America, 2Boston Children's Hospital, Neurology Department, Boston, United States of America, 3Dartmouth–Hitchcock Medical Center, Department Of Internal Medicine, Lebanon, United States of America, 4Cell Care Therapeutics, Development And Quality, Monrovia, United States of America

Aims: Alzheimer’s disease (AD), a neurodegenerative disease resulting from the loss of neurons in the hippocampus and cortex, is characterized by the aggregation of β-amyloid plaques and neurofibrillary tangles of hyperphosphorylated tau (NFTs). However, the molecular mechanism underlying the neuronal cell death is poorly understood. Previously, our lab showed that the inhibition of LSD1 in adult mice induces cortical and hippocampal neurodegeneration, learning and memory deficits, and transcription alternations that match human AD cases. In addition, LSD1 is mis-localized with cytoplasmic NFTs in AD cases. Therefore, we hypothesized that NFTs contribute to neuronal cell death by sequestering LSD1 into the cytoplasm. Methods: We utilized the PS19 Tau mice expressing a mutated form of human tau. We generated mice with reduced LSD1, as well as overexpressed LSD1 to test if this would modulate the phenotype without altering tau pathology. Results: We found that LSD1 is depleted from the nucleus and mis-localized to the cytoplasm in the presence of NFTs. Additionally, reducing Lsd1 in PS19 Tau mice increases depletion of LSD1 from the nucleus; resulting in reduced survival, exacerbated paralysis, increased neurodegeneration, and aggravated transcription alternations compared to the PS19 Tau mice with wild type LSD1. Finally, we showed that overexpression of Lsd1 in hippocampal neurons of PS19 Tau mice after tau aggregates have formed, protects against neurodegeneration, blocks the associated gliosis, and counteracts the neurodegenerative transcription changes. Conclusions: Overall, these data suggest that NFTs induce neurodegeneration by functionally inhibiting LSD1. This novel molecular mechanism is a promising potential target for therapeutic treatments against AD.

P418 / #543

Topic: Theme B: Taupathies / B7.a. Animal Models: Transgenic rodents

CHARACTERIZATION OF THE PHENOTYPIC DEFICITS IN A TRANSGENIC MOUSE MODEL OF TAUOPATHY

Lecture Title:

L. Camargo1,2, D. Honold2, R. Bauer2, J. Kutzsche2, A. Willuweit3, D. Willbold1,2, S. Schemmert2 1Heinrich-Heine-Universität Duesseldorf, Institute Für Physikalische Biologie, Duesseldorf, Germany, 2Forschungszentrum Juelich, Institute Of Biological Information-7 (ibi-7), Juelich, Germany, 3Forschungszentrum Juelich, Institute Of Neuroscience And Medicine -4, Juelich, Germany

Aims: Translational research plays an essencial role in the study of Taupathies, including Alzheimer disease (AD). In this context, it is important to understand in detail the alterations in mousel models of Tauopathies and related diseases in order to clarify its physiology. Therefore, we performed an in-depth characterization of early phenotypic alterations in the transgenic TauP301L mouse model. Methods: Sex-matched Wild Type (WT) and homozygous TauP301L (Tau) mice were tested in different behavioural tests (Habituation/dishabituation Olfactory test, Pole test, SHIRPA test, Open field test) at different ages. Results: In the Olfactory test, Tau males smelled less the new aroma compared to the WT males at six months of age. In the SHIRPA test, the Tau mice (n=19) had higher scores compared to WT (n=19) at four months of age and Tau males had higher scores compared to females. In the Pole test, Tau males had higher sum scores compared to WT males at six months of age (SHIRPA and Pole: low score = no alterations, high score = phenotypic alterations). In the Open Field test, Tau males were slower and travelled less than the WT males (n=12), but Tau females (n=7) were similar to WT females (n=7) at two months of age. Conclusions: In conclusion, this study demonstrated that Tau mice have phenotypic alterations in the SHIRPA, Open Field, Pole and Olfactory test at early ages and a sex-difference in the performance of those test. Summarised, the TauP301L mouse model shows phenotypic alterations as earlier as two months of age.

P419 / #531


EFFECTS OF LEVETIRACETAM TREATMENT ON COGNITIVE AND NON-COGNITIVE IMPAIRMENTS IN A MOUSE MODEL OF TAUOPATHY.

Lecture Title:

A. Joly-Amado, M. Novoa, L. Alvarez, D. Chan, H. Paek, J. Willman, M. Willman, K. Nash university of south florida, Molecular Pharmacology And Physiology, Tampa, United States of America

Aims: The aim of this study was to test the effects of a Leviteracetam diet on cognitive and non-cognitive impairments in a mouse model of tauopathy, the Tg4510 mice. Methods: Three-month old Tg4510 transgenic and non-transgenic littermate mice were equally randomly assigned to treated (food containing 240mg/kg Levetiracetam) or placebo (normal diet) groups (n=10, 5F & 5M). All mice were single housed, and food intake and mice body weight were recorded once a week for 3 months. During the last 2 weeks before tissue collection the following behavioral tests were performed: open field, Y-maze, marble burying, RAWM with reversal, rotarod, novel object recognition (NOR) and fear conditioning. On the last day, the brain was dissected, blood sample was collected, and muscle mass, white fat, and brown fat were weighed. Results: Treatment with Leviteracetam did not improve cognition or activity in Tg4510 during open field, RAWM, and Ymaze. Leviteracetam improved performance during contextual fear conditioning in both non-transgenic and Tg4510 mice. Interestingly, Leviteracetam diet had significantly more impact in non-transgenic mice than Tg4510 mice. Indeed, we observed a decrease in body weight, food intake, increased time spent on rotarod and a decrease of anxiety like behavior in non-transgenic mice but not in Tg4510 mice treated with Leviteracetam when compared to chow diet. Conclusions: There was no significant improvement in cognitive and non-cognitive deficits in Tg4510 mice treated with Levetiracetam. Interestingly, Leviteracetam diet had significantly more impact in non-transgenic mice than Tg4510 mice, indicating that some of the Leviteracetam pathways could be disrupted in Tg4510 mice.

P420 / #612


KNOCK-IN HUMAN TAU MOUSE MODEL RECAPITULATES EARLY EVENTS OF TAU PATHOLOGY

Lecture Title:

S. Kraiem, S. Eddarkaoui, T. Boschetti, H. Benderradji, K. Carvalho, E. Faivre, N. Sergeant, D. Blum, L. Buee, V. Buée-Scherrer Univ. Lille, Inserm, CHU Lille, UMR-S1172 - Lille Neurosciences & Cognition, Alzheimer & Tauopathies, Lille, France

Aims: Alzheimer’s disease (AD) is the primary cause of cognitive deficits in aging. It is neuropathologically characterized by an intraneuronal aggregation of microtubule-associated Tau proteins which leads to neuronal death. Most of the existing animal models reconstituted Tau pathology by overexpressing human mutated Tau protein. Methods: Recently, our laboratory has developed a Knock In mouse model (Tau KI V5) by the insertion of a cDNA encoding 1N4R human Tau isoform (mutated at P301L and tagged with a V5 flag) in the mouse Mapt locus. A cohort of heterozygous, homozygous and their littermates at 16 months was established to evaluate Tau pathology. Results: This model shows a slight metabolic phenotype and a mild cognitive impairment, by using Y-Maze test. Expression of human Tau is detected both in central nervous system and some peripheral organs by using q-PCR and Western Blot. Localization of the transgene has also been evaluated by immunostaining. In the brain, an increase in some glial inflammatory markers has been detected by using q-PCR and Western Blot. Other synaptic and neuronal markers are currently under investigation. Conclusions: This Tau model is more likely to represent the early stages of AD. Since amyloid is the other hallmark of AD, challenging this model with amyloid might be interesting to study their interactions. Thus, this kind of model can help to explore pathophysiological mechanisms that lead to AD.

P421 / #596


SITE-SPECIFICALLY BLOCKING CASPASE-2-MEDIATED TAU CLEAVAGE DELAYS COGNITIVE IMPAIRMENT AND BRAIN ATROPHY IN PRECISELY ENGINEERED TAU TRANSGENIC MICE MODELING FRONTOTEMPORAL DEMENTIA

Lecture Title:

P. Liu, E. Steuer, L. Kemper, K. Leinonen-Wright, M. Montonye, C. Hlynialuk, J. Gamache, K. Ashe University of Minnesota, Neurology, Minneapolis, United States of America

Aims: In rTg4510 mice modeling frontotemporal dementia (FTD), memory deficits are linked to Δtau314, a soluble tau fragment generated by caspase-2 (Casp2) cleavage at aspartate 314 (D314). However, the disruption of Fgf14 resulting from random integration of tauP301L transgenes, and the lack of a Casp2-resistant control confound identification of mechanisms underlying the memory deficits. Here, we aim to better understand the contribution of this Casp2-mediated, site-specific tau cleavage to cognitive impairment and neurodegeneration in newly developed, precisely engineered tau transgenic mice. Methods: We used a mouse line, rT2, in which a single tauP301L transgene inserted into a neutral site promotes transgene expression without dysregulating endogenous gene expression, and an rT3 line harboring an additional tauD314E mutation that blocks Casp2-mediated tau cleavage at D314. Using a water maze, we assessed the spatial reference memory of rT2 and age-matched rT3 mice at 5, 8, and 12 months of age. We also measured age-related alterations in brain weights of rT2 and rT3 mice. Results: rT2 and rT3 mice expressed comparable levels of tau protein. Spatial reference memory was normal at 5 months, and became impaired at 8 months in rT2 mice, but not until 12 months in rT3 mice. Forebrain weights began to decrease at 15 months in rT2 mice, but not until 18 months in rT3 mice. Conclusions: Abolishing Casp2-mediated tau cleavage at D314 delays not only cognitive impairment but also brain atrophy in precisely engineered mice modeling FTD, confirming a pathogenic role for Casp2 cleavage of tau.

P422 / #1527


NON-INVASIVE HIGH-RESOLUTION WHOLE BRAIN OPTOACOUSTIC IMAGING OF TAUOPATHY IN P301L MICE

Lecture Title:

R. Ni1,2, X.L. Dean-Ben1, L. Mu3, Z. Chen1, B. Ji4, J. Klohs1, D. Razansky1, R. Nitsch2 1ETH Zurich & University of Zurich, Institute For Biomedical Engineering, Zurich, Switzerland, 2University of Zurich, Institute For Regenerative Medicine, Zurich, Switzerland, 3ETH Zurich, Institute Of Pharmaceutical Sciences, Zurich, Switzerland, 4National Institutes for Quantum and Radiological Science and Technology, Department Of Functional Brain Imaging Research, Chiba, Japan

Aims: Abnormal tauopathy accumulation play an important role in tauopathy diseases including Alzheimer’s disease and frontotemporal lobe dementia. There is a gap in high-resolution non-invasive whole brain imaging of tauopathy deposits in animal models, with the current small animal positron emission tomography reaching approx. 1 mm resolution. Here, we demonstrated transcranial 3D mapping of whole brain tauopathy in P301L mouse model of four-repeat tauopathy using optoacoustic tomography at a 110 μm spatial resolution. Methods: P301L and non-transgenic littermate mice (18 months-of age, both gender) underwent in vivo optoacoustic imaging using tau imaging probe PBB5 (i.v. injection), followed by structural T2-weighted magnetic resonance imaging (MRI) at 9.4T. The optoacoustic imaging data was reconstructed, multispectrally unmixed and coregistered with MRI data for volume-of-interest analysis. Immunohistochemical staining was performed with PBB5, anti-phosphorylated tau antibody AT-8 on mouse brain sections for validation of in vivo results. Results: Intravenous administration of PBB5 in P301L mice led to a retention of the probe in tau ladened cortex and hippocampus. Structural T2-weighted MRI showed atrophy in the cortical and hippocampal regions in P301L mice compared to non-transgenic littermates. The specificity of PBB5 was validated by immunohistochemistry with anti-phosphorylated tau antibody AT-8. Conclusions: Optoacoustic imaging with PBB5 facilitates preclinical high-resolution non-invasive whole brain imaging of tauopathy, for studying spread and accumulation of tauopathy in animal models, and for monitoring of putative treatments targeting tau.

P423 / #1731


IDENTIFYING TAU SPECIES RESPONSIBLE FOR BEHAVIORAL IMPAIRMENTS IN A MOUSE MODEL OF TAUOPATHY

Lecture Title:

H. Patel, P. Martínez C., A. Perkins, C. Lasagna-Reeves, D. Mckinzie IUPUI, Stark Neuroscience Research Institute, Indianapolis, United States of America

Aims: Objectives: Abnormal aggregation of the protein tau is the underlying cause of several neurodegenerative diseases, termed as tauopathies, which are associated with motor and cognitive impairments. To identify the association between tau pathology in the brain and the functional deficits manifested, we assessed multiple in vivo phenotypes in a transgenic tauopathy mouse model, overexpressing human tau with P301S mutation. Methods: We tested 9-month-old male and female PS19 and WT littermates through a battery of behavioral and physiological assessments: Y-maze spontaneous alternation, rotarod performance, frailty clinical observations, body temperature, grip strength, open-field activity, fear conditioning, and active place avoidance learning. Following in vivo testing, brains were collected for analyzing tau seeding load and other biochemical changes. Results: PS19 mice exhibited significant hyperactivity, reduced body weight and body temperature, muscle weakness, and impaired spatial learning during a place reversal phase. Moreover, sex differences emerged in the magnitude of impairment in some of these measures. We are currently performing correlational analysis to determine the associative strength of a diverse set of tau pathological species with the functional impairment observed in this tauopathy mouse model. Conclusions: Establishing an association between tau deposition and functional deficits is critical for determining which measures are most strongly mediated by tau pathology. This information can facilitate future treatment studies by using in vivo measures as surrogate efficacy biomarkers prior to post-mortem histopathological and biochemical analyses. Further, the establishment of robust in vivo phenotypes can be used to identify those tau species most associated with functional decline.

P424 / #459


HUMAN MAPT KNOCK-IN MICE THAT HARBOR FAMILIAL TAUOPATHY-CAUSING MUTATIONS

Lecture Title:

N. Watamura, S. Hashimoto, H. Sasaguri, T. Saido RIKEN, Center For Brain Science, Wako-shi, Japan

Aims: Tauopathy is characterized by the accumulation of pathological tau in brain. To clarify the disease mechanisms, we aimed to develop mouse models that reproducibly display specific tau pathology without artifacts because the current tauopathy models depend on the overexpression paradigm. The purpose of this study is to develop new tauopathy mouse models by knock-in strategy. Methods: Previously we generated human MAPT knock-in (hTau-KI) mice, in which the entire Mapt gene was humanized. The hTau-KI mice express all six isoforms (Hashimoto et al.,2019; Saito et al., 2019). In this study, several tauopathy-causing mutations were introduced into hTau-KI mice by Base Editor, a novel CRISPR/Cas9-based genome editing technology. Results: We obtained several tauopathy mouse models (hTau-KI P301L, hTau-KI P301S, hTau-KI P301V, hTau-KI Intron10+3 G>A, hTau-KI P301L; Intron10+3 G>A, hTau-KI P301S; Intron10+3 G>A, hTau-KI S305N; Intron10+3 G>A). Immunohistochemical analysis showed phospho-tau positive signals in the entorhinal cortex in some of these lines. In addition, Biochemical analysis demonstrated that the Intronic mutation altered the expression patterns of 3- and 4- repeat tau. Conclusions: We successfully introduced tauopathy-causing mutations in hTau-KI mice and detected the effect of mutations on MAPT gene. We expect these models to contribute to elucidating the mechanism of tauopathies and plan to make them available to the research community worldwide.

P425 / #1195

Topic: Theme C: α-Synucleinopathies / C1.a. Disease Mechanisms, Pathophysiology: Αlpha-synuclein aggregation

GENERATING A FIRST IN CLASS INHIBITOR TO TREAT PARKINSON’S DISEASE BY TARGETING INTRACELLULAR ALPHA-SYNUCLEIN PATHOLOGY

Lecture Title:

N. Aït-Bouziad, A. Davranche, E. Tsika, N. Dreyfus, C. Boudou, V. Darmency, H. Kroth, K. Piorkowska, A. Ouared, M. Van Campenhoudt, L. Rey-Bellet, A. Pfeifer, M. Kosco-Vilbois AC Immune SA, Research, Lausanne, Switzerland

Aims: Synucleinopathies, such as Parkinson’s Disease (PD), are characterized by the progressive accumulation of alpha-synuclein (a-syn) inclusions in the brain which correlates with disease severity. Using our unique MorphomerTM library, we aim to discover and develop brain- and cell-penetrant small molecules that can interfere and potentially prevent the formation of a-syn aggregates and the downstream pathological and neuroinflammatory processes leading to these devastating diseases Methods: The AC Immune MorphomerTM library, designed to target and modify β-sheet structures of aggregation-prone proteins, rendering them non-pathological, was employed. A human embryonic kidney 293 (HEK) cell line overexpressing a-syn fused to eGFP was used in a screen to identify compounds that inhibit a-syn de novo aggregate formation in vitro. Chemically diverse primary hits were subsequently validated using aggregation assays combining Thioflavin-T (ThT) dye, as fluorescence reporter of amyloid fibril formation, with sedimentation analysis, confirming whether compounds inhibited the structural transition of a-syn into a fibrillar state Results: Several series of small molecules were identified. In a cellular environment, a-syn aggregates were significantly decreased upon treatment. The identified inhibitors reduced ThT signal, prolonged the lag phase and prevented structural transitions of a-syn into aggregates. Using medicinal chemistry iterative optimization, we successfully identified active optimized hits with physico-chemical CNS properties suitable for in vivo evaluation Conclusions: Several chemical series interfere with the fibrillation process of a-syn and hold promise in providing therapeutic benefit for PD and related synucleinopathies. These compounds will be progressed into in vivo proof of concept in a mouse model recapitulating the formation of pathological a-syn aggregates

P426 / #1035


NEUROPROTECTIVE EFFECTS OF EZEPROGIND® (AZP2006) IN IN VITRO AND IN VIVO MODELS OF PARKINSON’S DISEASE.

Lecture Title:

N. Callizot1,2, C. Estrella2, S. Burlet2, A. Henriques3, P. Verwaerde2 1Neuro-Sys, Pharmacology, Gardanne, France, 2Alzprotect, Drug Development, Loos, France, 3Neuro-Sys, Pharmacology, GARDANNE, France

Aims: Parkinson disease (PD) is the most common neurodegenerative movement disorder. The hallmarks of PD are loss of dopaminergic neurons in the substantia nigra (SN) and accumulation of misfolded α‐synuclein in Lewy body intra‐cytoplasmic inclusions. Risk factors include age and environmental factors. The etiology of the disease is unknown, but different genetic causes have been identified (5%–15% of cases). Progranulin (PRGN) is a growth factor with multiple activities acting as a neurotrophic, anti-inflammatory factor, cytokine and regulator of lysosomal function. In brain, it has been demonstrated to regulate neuroinflammation, neurite branching and outgrowth and lysosomal function. Ezeprogind is a small molecule clinically tested in Progressive supranuclear palsy patients (phase 2a). It has been demonstrated to increase the half-life of PRGN. In addition, Ezeprogind was also shown to massively decrease the neuroinflammation via a direct inhibition of Toll like receptor subtype 9. This dual mode of action makes Ezeprogind a promising treatment for neurodegenerative diseases. Methods: Using cellular models of PD, we showed that Ezeprogind was able to prevent the neurodegeneration of primary dopaminergic neurons induced by mitochondrial toxins. Results: . A reduction of a-synuclein aggregation was also observed in a dose dependent manner. In parallel, Ezeprogind was able to prevent the neuronal death induced by asyn toxicity and was able to fully abolish the associated neuroinflammation. In in vivo model of PD, Ezeprogind orally administrated was able to prevent the SN lesions and to reduce the asyn aggregation. Conclusions: Altogether these results are in favor of a development of Ezeprogind in PD and synucleinopathies.

P427 / #1283


PHOTODYNAMIC STUDIES OF ΑLPHA-SYNUCLEIN PROTEIN SEEDING, CLEARANCE AND PRODUCTION

Lecture Title:

C. Croft, G. Paterno, A. Vause, M. Goodwin, D. Ryu, C. Moran, B. Giasson, T. Golde McKnight Brain Insitute, Department Of Neuroscience, Gainesville, United States of America

Aims: Alpha-synuclein accumulation is associated with the development of Lewy body pathology in neurodegenerative diseases such as Parkinson’s disease and Lewy Body Dementia. There is still limited understanding of how alpha-synuclein and Lewy bodies are associated with cellular dysfunction and degeneration. Methods: We have recently established ex vivo models of intrinsic α-synuclein inclusion pathology in three-dimensional, murine brain slice cultures (BSCs), transduced with recombinant adeno-associated viruses (rAAVs) to express human wild-type and mutant α-synuclein. These models develop pathology which is highly recapitulative of Lewy body pathology relevant to Parkinson’s disease and other α-synucleinopathies. We now extend these findings using photoswitchable versions of these rAAVs with long-term live imaging to understand the protein clearance and production dynamics of α-synuclein in these models, as well as in seeded models of α-synuclein accumulation. Results: Using optical pulse-chase methodology over periods of several weeks in culture we are able to follow individual cells expressing α-synuclein and follow new and old populations of protein in these cells. We identify similarities and differences between the production and turnover of aggregated wild-type and mutant α-synuclein in the presence or absence of pre-formed fibrils. Conclusions: These studies highlight the value of this amenable system to explore the dynamics of α-synuclein inclusions over long-term culture periods. This system will provide a useful platform to provide mechanistic insight on how therapeutics, other genes and different mutations can affect α-synuclein protein dynamics.

P428 / #547


DIFFERENTIAL IN-VITRO DIAGNOSTICS OF NEURODEGENERATIVE DEMENTIAS

Lecture Title:

V. Gupta University of Oxford, Department Of Clinical Neurosciences, Oxford, United Kingdom

Aims: Background: There is a vital unmet need for objective and sensitive diagnostic methods that would allow early and differential diagnosis of neurodegenerative dementias (NDs). RT-QuIC is one very promising method that can detect various aggregating proteins with high accuracy in patient’s cerebrospinal fluid (CSF) as a surrogate of brain pathology. RT-QuIC was initially developed to detect prion diseases but it has been successfully applied to detect alpha-synuclein (αSyn) aggregation in Parkinson’s and Lewy body dementia and tau aggregation in Alzheimer’s disease and other tauopathies. Objective: To compare the various protocols for real-time quaking-induced-conversion (RT-QuIC) assay from different laboratories and assess their strengths and weaknesses. Methods: RT-QuIC assays are multi-well plate-based reactions where intermittent shaking enhances the conversion of soluble recombinant protein into amyloid fibrils in the presence of pathological seed from patient-derived sample (e.g. CSF). These fibrils form bonds with Thioflavin T generating an easily measurable fluorescent signal. Results: Among different laboratories, αSyn RT-QuIC exhibits specificity of 82 -100% and sensitivity of 70-100%. RT-QuIC protocols differ in terms of substrates, temperature, shaking/incubation cycles and buffers that all have an effect on the assay performance and make it difficult to compare the results. Moreover, RT-QuIC may hold potential to detect different conformational strains of the protein that would allow a better stratification of patients with different NDs. Although RT-QuIC is sensitive and specific, its quantitative nature needs to be established. Conclusions: Conclusion: We recommend one standardised RT-QuIC procedure to be established as a routine diagnostic assay for NDs in the surveillance-centres worldwide.

P429 / #1027


EARLY AGGREGATION STEPS OF ALPHA-SYNUCLEIN FROM LARGE-SCALE IN SILICO SIMULATIONS

Lecture Title:

A. Guzzo Laboratoire Interdisciplinaire Carnot de Bourgogne (ICB), UMR UB-CNRS 6303, Nanosciences, Dijon, France

Aims: The hallmark of the neurodegeneration is the conversion of soluble proteins in insoluble aggregate in neurons. In the Parkinson disease, the largest protein aggregates, amyloids, were identified as composed of alpha-synuclein (aS). Deciphering the mechanism of aggregation of aS is a pre-requisite to develop identification methods of the early onset of the Parkinson disease. The monomer of aS is an intrinsically disordered protein composed of 140 amino-acids characterized by a repeat motif of 6 amino-acids (KTKEGV). The repeat motif in aS sequence plays a role in its aggregation and can disturb the equilibrium between monomers and multimers which seems to be a key to understand the disease. Methods: The aim of the present work is to identify the aggregation mechanism of aS using in silico simulations and high-performance computer resources. We apply a coarse-grained molecular dynamics model, UNRES (UNited RESidue Model), to simulate the aS dynamics at the atomic scale on an impressive timescale of hundreds of milliseconds, relevant for the aggregation mechanism. Results: We simulated the structural properties of wild-type (WT) aS monomer and mutants (A30P, E46K, A53T) and found agreement with the scarce experimental data. Simulations of aS dimers were performed and differences between WT and mutants were identified. Preliminary data for the classification of the dimeric structures are presented. Conclusions: The present results could be used in the future to identify relevant physical properties for early detection of the disease and to identify a possible way of inhibition.

P430 / #861


IN VITRO AND IN VIVO MODELING OF MUTATED A53T ALPHA-SYNUCLEIN PATHOLOGY

Lecture Title:

T. Loeffler, R. Rabl, I. Schilcher, M. Daurer, S. Flunkert, B. Hutter-Paier QPS Austria GmbH, Neuropharmacology, Grambach, Austria

Aims: Aggregation of α-Synuclein (α-Syn) plays a central role in Parkinson’s disease (PD). Mutations in the α-Syn gene have been identified in rare forms of familial PD and are reported to accelerate its oligomerization and aggregation. In subjects affected by the α-Syn A53T mutation, the age of onset is earlier compared to sporadic PD. The development of new PD drugs halting the production of α-Syn aggregates and the resulting neurodegeneration is thus the main focus of PD research. To be able to test these new drugs, appropriate in vitro and in vivo models are needed. Methods: We therefore evaluated toxicity and seeding properties of different recombinant human α-Syn peptides and fibrils in primary cortical neurons. Additionally, human A53T α-Syn transgenic mice with murine Thy-1 promoter on a C57BL/6J background and age-matched non-transgenic littermates are currently characterized for general health and motor deficits. Animals of both sex and different age groups are evaluated. Results: Our in vitro results show that monomeric human α-Syn has no impact on cell viability, while preformed wild type and A53T human α-Syn fibrils have toxic effects on primary cortical neurons. Only A53T α-Syn fibrils of the tested isotypes showed seeding properties. Conclusions: It can be concluded that primary cortical neurons treated with preformed A53T α-Syn fibrils are a valuable in vitro model to analyze α-Syn aggregation and toxicity. Results of hA53T α-Syn transgenic mice are expected to validate the value of this mouse model for PD research.

P431 / #1816


HSP70 CHAPERONE BLOCKS Α-SYNUCLEIN OLIGOMER FORMATION VIA A NOVEL ENGAGEMENT MECHANISM

Lecture Title:

L. Mcconlogue1, J. Tao1, A. Berthet2, Y.R. Citron1, R. Stanley2, J. Gestwicki3, D. Agard1 1University of California, San Francisco, Biochemistry And Biophysics, San Francisco, United States of America, 2The Gladstone Institutes, Institute Of Neurological Disease, San Francisco, United States of America, 3University of California, San Francisco, Pharmaceutical Chemistry, San Francisco, United States of America

Aims: Over-expression and aggregation of alpha-synuclein (ASyn) are linked to the pathology of the synucleinopathies. Elevated levels of the stress-induced chaperone Hsp70 protects against ASyn misfolding and ASyn-driven neurodegeneration in cell and animal models, yet there is minimal mechanistic understanding of this important protective pathway. It is generally assumed that Hsp70 binds to ASyn using its canonical and promiscuous substrate-binding cleft to limit aggregation but this has not been tested. Indirect methods show that Hsp70 blocks oligomerization, the earliest stage of aggregation. We aim to test whether 1) Hsp70 blocks ASyn oligomerization using direct methods and 2) Hsp70 blocks ASyn oligomerization using its canonical mode. Methods: Using biochemical assays of ASyn oligomerization we tested Hsp70 for impact. We tested the requirement of ATP cycling and the impact of active site inhibitors to probe the requirement for Hsp70 canonical action. We further tested the requirement for canonical action in H4 neuroglioma cells. Results: We found that Hsp70 directly impairs ASyn oligomerization in an ATP cycling independent mechanism both in vitro and in cells. Importantly, this activity was not inhibited by an active site inhibitor indicating a site separate from the canonical Hsp70 active site is responsible. Conclusions: We report that Hsp70 blockage of ASyn oligomerization is due to a novel and unexpected mode of Hsp70 action, involving neither ATP nor its substrate-binding cleft. Together, these findings suggest that new chemical approaches will be required to target the Hsp70-ASyn interaction in synucleinopathies. Such approaches are likely to be more specific than targeting Hsp70’s canonical action.

P432 / #724


TRACTABLE HUMAN STEM CELL-BASED MODELS FOR MODELING ADVANCED AGGREGATION PATHOLOGIES IN ALPHA-SYNUCLEINOPATHIES

Lecture Title:

A. Ndayisaba1, I. Lam2, L. Zaccagnini3, R. Sanz2, A. Lewis4, J. Sandoe2, A. Vahdatsoar2, N. Ramalingam1, X. Jiang5, P. Xu5, T. Martin6, S. Swarup7, J. Zhang7, A. Verma1, C. Park1, I. Ferrer8, G. Halliday9, H. Stahlberg4, U. Dettmer1, C. Chung10, W. Harper7, S. Elledge11, T. Bartels3, V. Khurana1,5,12 1Brigham and Women's Hospital/Harvard Medical School, Neurology, Boston, United States of America, 2Brigham and Women's Hospital/Harvard Medical School, Ann Romney Center For Neurological Diseases, Boston, United States of America, 3Dementia Research Institute / University College London, Neurology, London, United Kingdom, 4Ecole Polytechnique Fédérale de Lausanne, Laboratory Of Biological Electron Microscopy, Lausanne, Switzerland, 5Yumanity Therapeutics, Discovery Biology, Boston, United States of America, 6Brigham and Women's Hospital/Harvard Medical School, Medicine, Boston, United States of America, 7Harvard Medical School, Cell Biology, Boston, United States of America, 8Institut de Neuropatologia, Institut D'investigació Biomèdica De Bellvitge-hospital Universitari De Bellvitge, Hospitalet de Llobregat, Spain, 9Sydney Medical School, The University of Sydney, & Faculty of Medicine, University of New South Wales & Neuroscience Research Australia, Brain And Mind Centre, Sydney, Australia, 10Yumanity Therapeutics, Yumanity Therapeutics, Boston, United States of America, 11Brigham and Women's Hospital/Harvard Medical School, Genetics, Boston, United States of America, 12MIT, Broad Institute Of Harvard And Mit, Cambridge, United States of America

Aims: Proteinaceous aggregation, comprising both membrane- and amyloid fibril-rich alpha-synuclein (aS) inclusions, is the hallmark brain pathology of synucleinopathies like Parkinson’s disease (PD). However, disease-relevant aggregation pathologies do not occur in reasonable timeframes in patient iPSC models. We aimed to develop tractable human iPSC-based models that rapidly develop aS aggregation pathologies. Methods: We developed an all-in-one PiggyBac-based system that enables tetracycline-inducible expression of the Ngn2 transcription factor in familial PD iPSC, with or without concomitant transgenic over-expression of aS. Scalable and virus-free generation of cortical glutamatergic neurons was achievable with 8 days. We characterized i) a fibrillar synucleinopathy model in which neurons were seeded with synthetic or brain lysate-templated aS pre-formed fibrils (PFFs) and ii) a membrane-rich synucleinopathy model in which transgenic expression of an amplified aS (E46>K) mutation resulted in rapid development of vesicle-rich intracellular inclusions. Results: These iPSc-derived neuron models recapitulate both immunohistochemical markers of pathologic aS (e.g., phospho-Ser129, ubiquitination and SDS-insolubility) and ultrastructual characteristics of advanced intracellular pathologies found in postmortem synucleinopathy brains. We are dissecting differences and similarities between these two types of inclusions, including examining their effect on neuronal survival through longitudinal live-cell tracking, and CRISPR/Cas9-based screens to identify genetic determinants of cellular toxicity. Conclusions: Our technology enables us to dissect the cellular consequences of distinct aS inclusion formation in a rapid and scalable human stem-cell model system. We envisage these models will be useful for biological and drug discovery in PD and related synucleinopathies.

P433 / #371


DEVELOPMENT OF A NOVEL OPTOGENETIC BASED MODEL OF ΑLPHA-SYNUCLEIN AGGREGATION TO STUDY PARKINSON’S DISEASE

Lecture Title:

R. Sheta, M. Bérard, R. Rodriguez Aller, M. Teixeira, A. Oueslati Université Laval, Department Of Molecular Medicine & Chu De Quebec Research Center, Québec, Canada

Aims: Parkinson’s disease (PD) is characterized by dopaminergic neuronal loss and presence of proteinecious inclusions Lewy bodies. These inclusions are constituted of a pre-synaptic protein, α-synuclein (α-syn). Evidence suggest for a central role of α-syn aggregation in PD. However, how these aggregates precipitate DA neuronal loss remain elusive. This is due to the lack of proper models to undertake such investigations. To overcome this limitation, our group created a cellular and animal model of PD mimicking authentic LBs features. Using our new optogenetic-inducible α-syn aggregation, we aim to dissect how these inclusions interfere with physiological functions of DA neurons leading to neuronal loss. Methods: Our optogenetic versatile strategy allows for spatiotemporal control of α-syn aggregation both in vivo and in living cells. This approach is based on the use of a mutant form of the Arabidopsis thaliana photoreceptor cryptochrome 2 (CRY2). When stimulated with blue light, CRY2 undergoes reversible and robust protein clustering. Fusing this system to α-syn, CRY2 clustering triggered aggregation of α-syn prompting formation of LB-like inclusions in living cells. We refer to this system as light-inducible protein aggregation (LIPA). Results: LIPA has allowed for real-time induction of α-syn inclusions with remarkable spatial and temporal resolution both in vitro and in vivo. Results showed that LIPA-induced aggregates auto-perpetuate for several days, faithfully mimicing authentic features of LBs. Optogenetically induced α-syn aggregation in mice induced significant dopaminergic neuronal loss and behavioural impairment. Conclusions: LIPA provides a dependable and invaluable tool to generate, visualize and dissect the role of protein aggregates in neurodegenerative disorders.

P434 / #1682


ALPHA-HELICAL PEPTIDIC SCAFFOLDS TO TARGET ALPHA-SYNUCLEIN PATHOGENIC SPECIES WITH HIGH AFFINITY AND SELECTIVITY.

Lecture Title:

S. Ventura Universitat Autonoma de Barcelona, Institut De Biotecnologia I Biomedicina, Bellaterra-Barcelona, Spain

Aims: α-Synuclein aggregation is a key driver of neurodegeneration in Parkinson’s disease and related syndromes. Accordingly, obtaining a molecule that targets α-synuclein pathogenic assemblies with high affinity and selectivity is a long-pursued objective. Methods: Here, we have exploited the biophysical properties of toxic oligomers and amyloid fibrils to identify a family of α-helical peptides that bind selectively to these α-synuclein species with low nanomolar affinity, without interfering with the monomeric functional protein. Results: This activity is translated into an unprecedented anti-aggregation potency and the ability to abrogate the oligomers toxicity. With a structure-function relationship in hand, we identified a human peptide expressed in the brain and in the gastrointestinal tract with exceptional binding, anti-aggregation, and detoxifying properties, which suggests it might play a protective role against synucleinopathies. Conclusions: The chemical entities we describe here represent a new therapeutic paradigm and are promising tools to assist diagnosis by selectively detecting α-synuclein pathogenic species in biofluids.

P435 / #433


STEREOLOGICAL STUDY OF HUMAN ANTERIOR OLFATORY NUCLEUS IN PARKINSON’S DISEASE

Lecture Title:

S. Villar-Conde, M. Gonzalez-Rodriguez, V. Astillero-Lopez, P. Villanueva-Anguita, D. Saiz-Sanchez, I. Ubeda-Banon, A. Martinez-Marcos, A. Flores-Cuadrado Ciudad Real Medical School. University of Castilla-La Mancha, Medical Sciences, Ciudad Real, Spain

Aims: Neuropathologically Parkinson’s disease (PD) is characterized by aggregations of α-synuclein, which can spread through neuronal connections in a predictable staging. These aggregates appear first in the dorsal nucleus of the vagus nerve and the olfactory bulb, particularly in the anterior olfactory nucleus (AON). Elevated synucleinopathy reported in human AON during PD could alter cell populations. But studies focused on neurodegeneration show controversial data. The best of our knowledge only one stereological study has analyzed microglial and astroglial populations indicating microgliosis in human AON in PD. This work aims analyzing α-synucleinopathy and neural and glial changes among AON bulbar (AONb), retrobulbar (AONrb), anterior-cortical (AONca) and posterior-cortical (AONcp). Methods: Brains from PD (Braak 5-6) and non-PD age-matched subjects were obtained from Spanish Biobank network (AONb: PD n=12, NPD n=9; AONrb: PD n=6, NPD n=7; AONca: PD n=6, NPD n=8; AONcp: PD n=4, NPD n=5). Procedures were approved by the Ethical Committee for Clinical Research at the University Hospital of Ciudad Real (SAF2016-75768-R and PID2019-108659RB-I00). Area fraction fractionator was used to determine the area occupied by α-synuclein. Volume changes were estimated by Cavalieri’s method. Optical Fractionator method was used to analyze cell types (Neu-N, Iba-1 and GFAP). Results: AON in PD present a conspicuous α-synucleinopathy that influences neuronal and glial populations, although volume remains unaltered. Conclusions: This work is supported by UCLM/ERDF (2020-GRIN-2145 to NPND), Spanish Ministries of Economy and Competitiveness/ERDF (SAF2016-75768-R) and Science and Innovation (PID2019-108659RB-I00) to AMM and Autonomous Government of Castilla-La Mancha/ERDF (SBPLY/17/180501/000430) to AMM and DSS. SVC received a UCLM-FEDER predoctoral fellowship.

P436 / #183


INCREASED ALPHA-SYNUCLEIN OLIGOMERIZATION IS ASSOCIATED WITH DECREASED ACTIVITY OF GLUCOCEREBROSIDASE IN THE AGING HUMAN STRIATUM AND HIPPOCAMPUS

Lecture Title:

W. Yang, X. Li, N. Yin Xuanwu Hospital, Capital Medical University, Neurobiology, Beijing, China

Aims: Aging is associated with an increased risk for Parkinson’s disease and dementia with Lewy bodies, in which α-synuclein (α-syn) oligomerization plays key pathogenic roles. Here, we show that oligomeric α-syn levels increase with age in the human brain and are accompanied by a decrease in the activity of glucocerebrosidase (GCase), a lysosomal enzyme whose dysfunction is linked to the accumulation of oligomeric α-syn. Methods: The oligomeric α-syn concentration in brain tissues and cell lysates was measured by ELISA. GCase activity was determined using the QuantiChrom β-Glucosidase Assay kit. The lysosome-enriched fraction of brain tissue was prepared as previously described for GCase activity detection. The cells were transferred to 35-mm dishes for immunocytochemistry and 25-cm2 flasks for the GCase activity assay and oligomeric α-syn detection. Results: The inverse relationship between oligomeric α-syn levels and GCase activity was more evident in brain regions susceptible to neurodegeneration (i.e., the striatum and hippocampus) than those that are less vulnerable (i.e., the cerebellum and occipital cortex). GCase could potentially regulate α-syn oligomerization, as demonstrated by the decrease in oligomeric α-syn levels caused by a GCase agonist. In vitro experiments showed that GCase activity was more potently inhibited by oligomeric than monomeric α-syn in the lysosome-enriched fractions isolated from brain tissues and cultured neuronal cells. Conclusions: Alterations in oligomeric α-syns and their association with GCase in aging brains may explain the vulnerability of certain brain regions to neurodegeneration in Parkinson’s disease and dementia with Lewy bodies.

P437 / #1038


QUANTIFY, ISOLATE AND AMPLIFY THE PRION-LIKE ALPHA-SYNUCLEIN FOR STRUCTURAL ANALYSIS USING PMCA ASSAY

Lecture Title:

L. Zaccagnini1, R. Gallardo2, N. Ranson2, S. Radford2, T. Bartels1 1Dementia Research Institute / University College London, Neurology, London, United Kingdom, 2University of Leeds, Structural Molecular Biology, Leeds, United Kingdom

Aims: The aims of our research are: (i) compare the αSyn-PMCA products from different brain regions between PD, DLB and MSA patients, in order to assess if different brain regions are affected by different αSyn-aggregates. (ii) purify and amply peripheral and central αSyn aggregates from PD patients by PMCA to analyze their structures, since it has been demonstrated that aggregates of αSyn fibrils are also found in neural tissue located outside the central nervous system (CNS) of PD patients, in both the autonomic and enteric nervous system (ENS). Methods: α-synuclein insoluble fractions extraction from human brain and colon tissues, Protein Misfolding Cyclic Amplification (PMCA), proteinase digestion, western blotting, cryo EM Results: In the last year, our laboratory has optimized the PMCA assay for either αSyn preformed fibrils (PFF) and insoluble fraction of αSyn deriving from human brain material. Now, we are amplifying brain material from PD, DLB and MSA patients in order to analyze the product of αSyn-PMCA using a combination of biochemical, biophysical and biological methods, such as proteinase digestion and cryo EM. Conclusions: In conclusion, we think use PMCA assay to amply the αSyn fibrils and cryo EM for the structural analysis is a perfect strategy in order to understand the region specificity in the synucelinopathies.

P438 / #927

Topic: Theme C: α-Synucleinopathies / C1.b. Disease Mechanisms, Pathophysiology: LRKK2, parkin, PINK1, DJ-1

DELINEATION OF A MOLECULAR CASCADE LINKING XBP1 TO MITOPHAGY CONTROL IN PARKINSON’S DISEASE

Lecture Title:

C. Alves Da Costa, W. El Manaa, E. Duplan, I. Lauritzen, L. Vaillant-Beuchot, M. Chami, F. Checler Université Côte d'Azur, CNRS-UMR 7275, Institut De Pharmacologie Moléculaire Et Cellulaire, Valbonne, France

Aims: Parkinson's disease (PD) is a neurodegenerative disease characterized by progressive loss of dopaminergic neurons. PD-affected brains show consistent endoplasmic reticulum (ER) stress and mitophagic dysfunctions. The mechanisms underlying these perturbations and how they are directly linked remain poorly understood. XBP1 is a transcription factor activated upon ER stress after unconventional splicing by the nuclease IRE1 thereby yielding XBP1S while PINK1 is a kinase considered as the sensor of mitochondrial physiology and a master gatekeeper of mitophagy process. Our main objective was to test and demonstrate the existence of a forward loop cascade linking XBP1 and PINK1 in normal and PD pathological conditions. Methods: We have used pharmacological and genetic modulation approaches to modulate XBP1 and PINK1 levels in various cellular and animal models Results: We show that XBP1S transactivates PINK1 in human cells, primary cultured neurons and mice brain and triggers a pro-mitophagic phenotype that is fully dependent of endogenous PINK1. We also demonstrate a PINK1-dependent phosphorylation of XBP1S that conditions its nuclear localization andthereby, governs its transcriptional activity. PINK1-induced XBP1S phosphorylation occurs atresidues reminiscent of those phosphorylated in substantia nigra of PD-affected brain. Conclusions: Overall, our study delineates a functional crosstalk between XBP1S and PINK1 governing mitophagy that is disrupted in PD conditions.

P439 / #1494


OXIDIZED PARKIN, ITS MISFOLDING AND AGGREGATION VIA THE FORMATION OF ABERRANT INTRA- AND INTERMOLECULAR DISULFIDE BONDS PRODUCE CELLULAR TOXICITY

Lecture Title:

A. Esmaeili, M. Duennwald Western University, Pathology, London, Canada

Aims: Objective: In this study, we hypothesized that oxidative stress damages parkin resulting in its misfolding and altered cellular localization, which contributed to UPS and autophagy dysfunction and thus to PD pathogenesis. Methods: We obtained key results from studies in HEK 293 cells stably expressing parkin and SH-SY5Y cells expressing endogenous parkin, We performed western blot analyses with protein lysates from cells to evaluate that parkin is modified by oxidative damage under normal growth conditions and under oxidative stress. Results: . Untreated SH-SY5Y cells expressing endogenous parkin and N2a cells transiently transfected with parkin-YFP have several small dots and crescent-shaped of parkin observing throughout of cytoplasm. However, upon stress treatment, endogenous parkin and parkin-YFP accumulates excessively in foci, which often appears interconnected with one another, localized throughout of cytoplasm and nucleus not necessarily on or near mitochondria. More than 80% of our analyzed cells, rather than showing normal diffuse parkin fluorescence, exhibited a large or several smaller discrete parkin spots in untreated or exposure to stress treatment. Under these experimental conditions, analysis performed with the software image J showed that roughly 60 % of these spots colocalized with the mitochondrial protein TOM20, and the rest appeared throughout of the cells. Conclusions: We found that alteration of parkin solubility induced by various extrinsic and intrinsic stressors provides a mechanism for parkin misfolding and dysfunction may be relevant to the pathogenesis of sporadic PD.

P440 / #428


ANALYSIS OF INTRACELLULAR MEMBRANE TRAFFIC USING IPS CELL-DERIVED NEURONS FORM FAMILIAL PARKINSON'S DISEASE PATIENT

Lecture Title:

A. Hattori1, E. Ohta2, M. Nagai3, K. Iwabuchi1, H. Okano4 1Graduate School of Medical Science Kitasato University, Program In Cellular Immunology, Sagamihara, Japan, 2Graduate School of Medical Science Kitasato University, Division Of Clinical Immunology, Sagamihara, Japan, 3Kitasato University School of Medicine, Department Of Neurology, Sagamihara, Japan, 4Keio University School of Medicine, Department Of Physiology, Tokyo, Japan

Aims: Parkinson's disease (PD) is a progressive neurodegenerative disorder which causes tremor, rigidity, postural instability, bradykinesia, and non-motor symptoms. Mutations in LRRK2 gene are the most prevalent cause of autosomal dominant PD. Our university previously reported that induced pluripotent stem cells (iPSC) derived from a patient with I2020T mutation in LRRK2 (I2020T LRRK2-iPSC) in a Japanese family (the Sagamihara family) replicate the pathologic phenotype evident in the PD brain to some extent. Recently, the pathogenic mutation in LRRK2 have been shown to enhance phosphorylation of Rab proteins, which is a related molecule to early or late or recycling endosome. The purpose of this study is to investigate whether the I2020T mutant LRRK2 affects intracellular membrane traffic via the Rab protein. Methods: We analyzed these protein levels and its intracellular localization using the differentiated neurons from I2020T LRRK2-iPSC and isogenic-iPSC. In addition, to evaluate the puncta of Rab endosome of iPSC-derived neurons, we developed combined new method of ImageJ and CellProfiler. Results: By using our method, we found that I2020T LRRK2 iPSC-derived neurons differed in their localization, protein levels, and number in the early, late, and recycling endosomes compared to isogenic-iPSC-derived neurons. Conclusions: These results suggest that I2020T LRRK2 mutation may affect regulation of intracellular membrane traffic in PD.

P441 / #505


ANALYSIS OF THE RAB29-LRRK2-RAB8/10 CASCADE UNDER LYSOSOMAL OVERLOAD STRESS

Lecture Title:

T. Kuwahara, T. Komori, K. Funakawa, M. Sakurai, T. Iwatsubo Graduate School of Medicine, The University of Tokyo, Department Of Neuropathology, Tokyo, Japan

Aims: LRRK2, a causative kinase for autosomal dominant Parkinson’s disease, phosphorylates a subset of Rab GTPases including Rab8 and Rab10 in cells. We have previously shown that the treatment with a lysosomotropic drug chloroquine upregulates the Rab29-LRRK2-Rab8/10 cascade to maintain lysosomal homeostasis. Here we investigated the effects of other lysososomotropic drugs and the molecular mechanisms induced by these drugs. Methods: IFN-gamma-activated RAW264.7 cells and LRRK2-overexpressing HEK293 cells were exposed to various lysosomal stressors, and the alterations in Rab phosphorylation, LRRK2 autophosphorylation and other lysosomal properties were analyzed by immunoblotting or immunocytochemistry. We also investigated the alteration in Rab29 phosphorylation that is induced under lysosomal stress. Results: We confirmed that the treatment with a set of clinically used lysosomotropic drugs commonly induced Rab10 phosphorylation without increasing LRRK2 autophosphorylation at pSer1292, the latter being an indicator of LRRK2 enzymatic activity. These drugs caused an increase in the molecular proximity between LRRK2 and Rab8/10, whereas Rab29-mediated LRRK2 activation was required for the induction of Rab10 phosphorylation. Lysosomotropic drug treatment also induced Rab29 phosphorylation that was not mediated by LRRK2. The expression of phosphomimetic mutant Rab29 suppressed the lysosomal enlargement induced by lysosomotropic drugs. Conclusions: These results implicate an essential role of the Rab29-LRRK2-Rab8/10 cascade in the response to lysosomal overload stress. Establishing the way to regulate this cascade may provide novel strategies to interfere with the pathogenetic process involving LRRK2.

P442 / #469


ALPHA-SYNUCLEIN AND LRRK2’S COOPERATION IN PARKINSON’S DISEASE: A FOCUS ON AUTOPHAGY

Lecture Title:

F. Lebourg, C. Gardier, M.-C. Gaillard, N. Dufour, E. Brouillet, G. Liot Université Paris-Saclay, CEA, CNRS, Laboratoire des Maladies Neurodégénératives : mécanismes, thérapies, imagerie, Molecular Imaging Research Center, Fontenay-aux-Roses, France

Aims: Parkinson’s Disease (PD) is characterized by a loss of dopaminergic neurons in the substantia nigra and the formation of Lewy bodies containing α-synuclein (α-syn). Mutations on genes, such as A53T on α-syn and G2019S on Leucine-Rich Repeat Kinase 2 (LRRK2), have been described to take part in the pathogenic mechanisms underlying PD. Moreover, it has been suggested that α-syn and LRRK2 act jointly in PD pathogenesis. This idea is supported by one of our previous in vivo study showing that LRRK2-G2019S potentiates the dopaminergic cell death induced by α-syn-A53T in rats. Thus, our aim is to determine how α-syn and LRRK2 act together on neuronal stress. Methods: Thereby, we use rat primary neuronal culture transduced with lentiviral to express α-syn and LRRK2. Results: We recently showed that LRRK2-G2019S induces a significant increase in the number of neurons expressing a pathological form of α-syn (phospho-Ser129) without inducing any cell loss. In addition, this effect is dependent on LRRK2 kinase activity (genetic dead kinase form and LRRK2 inhibitor, MLi2, treatment) and on the proteostasis pathways such as autophagy. We particularly focus on autophagy since this process appears to be altered in PD, and individually α-syn and LRRK2 have both been found to modulate it. We starve the neurons and analyze their autophagic flux by treating them with BafilomycinA1. Conclusions: Finally, our objective is to find which autophagic step is impacted by α-syn and LRRK2 to identify new molecular targets and thereafter develop new therapeutic strategies for PD.

P443 / #1740


WNT AND NFAT SIGNALLING CHANGES IN LRRK2 PARKINSON’S DISEASE MODELS

Lecture Title:

S.H. Lei, A. Wetzel, M. Hughes, T. Liu, A. Rahim, K. Harvey University College London, School Of Pharmacy, London, United Kingdom

Aims: LRRK2 mediated Wnt and Ca2+ cell signalling changes were reported previously. These include Ca2+-dependent immune responses and autophagy. In addition, LRRK2 protective and pathogenic genetic PD variants affect canonical Wnt signalling in opposite directions showing a clear correlation between Wnt signalling activity and neurodegenerative disease susceptibility. As most evidence to date was based on in vitro experiments, this project is aimed to obtain a better understanding of LRRK2 signalling function and dysfunction in vivo. Methods: We observed Wnt and NFAT signalling activities in different brain regions of wild type (WT), LRRK2 knock-out (KO), and G2019S knock-in (KI) mouse models. TCF/LEF-GFP lentiviral biosensors were transduced at P0 and signalling activity distribution in the brain investigated using immunohistochemistry. This data was complemented by measuring mRNA and protein expression changes, and signalling activity under basal and stimulated conditions in primary neuronal cultures from different brain regions. Results: We observed Wnt signalling pathway activation in different brain regions. Localised differences in Wnt pathway mediators were also confirmed using mRNA and protein expression analysis. Wnt signalling activity differed between investigated brain regions and cell types and was dependent on LRRK2 genotype and sex. Conclusions: LRRK2 KO and LRRK2 G2019S KI alter gene and protein expression of Wnt and NFAT signalling cascades. The observed signalling dysregulation was most prominent in cortex and striatum, and also suggested changes in glia. Further investigation into detailed regulatory effects of LRRK2 mediated Wnt signalling activities under physiological and pathological conditions is ongoing.

P445 / #1768

Topic: Theme C: α-Synucleinopathies / C1.c. Disease Mechanisms, Pathophysiology: Cell to cell transmission, spreading of pathology, prion-like

EXOSOMES MODIFY Α-SYNUCLEIN CATABOLIC PROCESSING AND INFLAMMATORY RESPONSE

Lecture Title:

A. Lamprokostopoulou, M. Pantazopoulou, L. Stefanis, K. Vekrellis Biomedical Research Foundation of the Academy of Athens, Translational Medicine, athens, Greece

Aims: Exosomes contribute to the secretion of α-synuclein (α-syn) however the connection between aggregation and propagation of toxic α-syn forms via exosomes has not been clarified. The aim of the present study was to examine the process of exosome-associated α-syn pathology transmission in mouse primary neurons and microglia. Methods: Mouse cortical neurons and microglia-enriched cultures were cultured with mouse hypersonicated α-syn preformed fibrils (PFF seeds) in the presence or absence of exosomes. The effect of the exosome-PFF association on the uptake and consequent aggregation and/or degradation of α-syn, in the recipient cells, was analyzed by immunoblotting and immunofluorescence. Results: Association of exosomes with PFFs, did not seem to alter the propensity of PFFs to be uptaken by all cell types. In the case of mouse cortical neurons, association of exosomes with PFFs led to lower levels of insoluble α-syn and prevented the appearance of α-syn truncated species. The levels of α-syn phosphorylated (pS129) products were not altered. PFFs can readily enter microglia, either alone or in association with exosomes, triggering delayed microglial activation when PFFs enter in association with exosomes. Additionally, treatment of microglial cells with PFF-associated exosomes induced the appearance of truncated as well as aggregated α-syn species. Conclusions: The association of exosomes with PFFs appears to alter α-syn truncation and aggregation products depending on the recipient cells, depicting that this association per se may elicit different α-syn catabolic processing and inflammatory response in cells.

P446 / #874

Topic: Theme C: α-Synucleinopathies / C1.d. Disease Mechanisms, Pathophysiology: Autophagy, lysosomes, ubiquitin, proteasome

PARKINSON’S DISEASE-LINKED LDL RELATED RECEPTOR 10 (LRP10) IS IMPLICATED IN REGULATION OF AUTOPHAGY-LYSOSOME AND SECRETORY PATHWAY FUNCTION

Lecture Title:

A. Carreras Mascaro, M. Grochowska, V. Boumeester, V. Bonifati, W. Mandemakers Erasmus Medical Center, Clinical Genetics, Rotterdam, Netherlands

Aims: Pathogenic loss-of-function variants in the LDL related receptor protein 10 gene (LRP10) have been recently identified in families with late-onset Parkinson’s disease (PD), Parkinson’s disease dementia (PDD) and dementia with Lewy bodies (DLB). Although LRP10 has been suggested to function in intracellular transport, mechanistic insight into how LRP10 variants cause these diseases is still unknown. In this work, we aim to determine the potential role of LRP10 deficiency in neurodegeneration, focusing on two pathways known to be central in PD pathogenesis: autophagy-lysosome pathway (ALP) and secretory pathway. Methods: Biochemical analyses, Western blotting and immunocytochemistry were used to study ALP and secretory pathway function in wild type, LRP10 knock-out or LRP10 overexpressing HEK-293T cells, as well as in iPSC derived brain cells. Results: Drug-induced modification of ALP function leads to dramatic changes in endogenous LRP10 protein levels in HEK-293T and iPSC derived brain cells. Interestingly, knock-out of LRP10 in HEK-293T cells leads to changes in expression levels of proteins associated with the ALP. Furthermore, immunocytochemistry experiments showed partial overlap of endogenous LRP10 and ALP markers, especially after blocking lysosomal acidification via the addition of Bafylomycin A1. Finally, association of LRP10 with the secretory pathway was also demonstrated, for instance, by co-localization of LRP10 with markers of multivesicular bodies (MVBs). Conclusions: In this study, we show that LRP10 is associated with ALP and secretory pathway function , indicating that deregulation of these pathways might be important disease mechanisms in patients carrying LRP10 pathogenic variants.

P447 / #977


NEUROTOXICITY OF Α-SYNUCLEIN PROTOFIBRILS IS EXACERBATED BY INHIBITION OF GBA, A LYSOSOMAL ENZYME, IN AN IN VITRO MODEL OF PARKINSON’S DISEASE.

Lecture Title:

A. Henriques1, L. Rouvière1, C. Farrugia1, P. Poindron1, N. Callizot2 1Neuro-Sys, Pharmacology, GARDANNE, France, 2Neuro-Sys, Pharmacology, Gardanne, France

Aims: Motor symptoms in Parkinson’s disease are caused by the loss of dopaminergic neurons in the substantia nigra. Neurodegeneration is due to mitochondrial stress and α-synuclein aggregation. Multiple lines of evidence suggest that lysosomes are involved in the degradation of toxic α-synuclein in dopaminergic neurons. Mutations on the gene coding for the lysosomal protein GBA is a strong risk factor for Parkinson’s disease. We investigated the link between the toxicity of α-synuclein protofibrils and the loss-of-function of GBA. Methods: Dopaminergic neurons were intoxicated with a preparation of α-synuclein (250 nM, up to 96 hours) containing protofibrils (30% determined by automated protein analysis), in presence or absence of conduritol B epoxide (CBE, inhibitor of GBA). Immunocytochemistry was applied to study the survival and neurite network of dopaminergic neurons (positive for TH, tyrosine hydroxylase), mitochondrial stress (ROS production) and lysosomes (positive for Lamp2). Results: We showed that α-synuclein protofibrils induced progressive mitochondrial stress and lysosomal dysfunction in dopaminergic neurons, starting 3 days after application. The slow and discrete direct toxicity led to late neuronal death of TH neurons. The application of CBE was also toxic for dopaminergic neurons and triggered dose-dependent lysosomal pathology. The co-application of α-synuclein with low doses of CBE induced a synergistic toxicity, demonstrated by enhanced mitochondrial stress and enlargement of lysosomes. Conclusions: Altogether, these results show that the toxicity of α-synuclein is exacerbated in dopaminergic neurons when lysosomal GBA activity is impaired. It reinforces the tight association between lysosomal dysfunction and α-synucleinopathy in the neurodegenerative process of Parkinson’s disease.

P448 / #1645


PATHOLOGICAL MECHANISM OF FAF1 INDUCING DOPAMINERGIC NEURONAL DEATH

Lecture Title:

B.-S. Kim, E. Kim Chungnam national university, Bioscience & Biotechnology, Daejeon, Korea, Republic of

Aims: Impairment of protein clearance mechanisms leads to α-synuclein accumulation in dopaminergic neurons, contributing to the pathogenesis of Parkinson’s disease (PD). Based on the finding that Fas-associated factor 1 (FAF1), a positive modulator of PD, colocalizes with α-synuclein in PD patient brains (Neurobiol Dis. 31, 309-15, 2008). Therefore, we studied pathological mechanism manifested by FAF1. Methods: To examine the pathological function of FAF1 as an autophagic suppressor, we performed experiments using SH-SY5Y cells and mouse model. Results: α-Synuclein-mediated mitochondrial depolarization was even further intensified in cells cotransfected with FAF1 and α-synuclein, implying that FAF1 enhanced α-synuclein-mediated mitochondrial depolarization. Moreover, cotransfection of α-synuclein with FAF1 synergistically increased mitochondrial fragmentation. In addition, FAF1 overexpression using stereotaxic injection of adeno-associated virus (AAV) led to α-synuclein accumulation by suppressing autophagic flux. Furthermore, FAF1-mediated impairment of autophagy resulted in dopaminergic neuronal loss in the PD model mice. Conclusions: We identified a novel function of FAF1 associated with potentiation of α-synuclein accumulation leading to mitochondrial dysfunction. [This research was supported by a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (grant number : H I 1 6 C 0 9 4 7 )]

P449 / #1401


CHARACTERIZING GLUCOCEREBROSIDASE ACTIVITY AND THE SUSCEPTIBILITY OF DEVELOPING ALPHA-SYNUCLEIN PATHOLOGY IN DIFFERENT BRAIN REGIONS OF YOUNG AND AGED MICE

Lecture Title:

Y. Malki, Y.K. Choi The Hong Kong University, Li Ka Shing Faculty Of Medicine, Division Of Neurology, Hong Kong, Hong Kong PRC

Aims: Parkinson’s Disease (PD) is characterized by the progressive aggregation and inter-neuronal propagation of alpha-synuclein. It is widely believed that the activity of the lysosomal enzyme, glucocerebrosidase, plays a role in regulating alpha-synuclein accumulation. Here, we aim to establish a brain region profile for the level of phosphorylated serine 129 (pSer129) alpha-synuclein, the fibril promoting form of alpha-synuclein, as well as the glucocerebrosidase activity, in both young and aged mice. Methods: Isolated brain regions from young and aged wild-type mice were subjected to Western Blotting, where we assessed the levels of pSer129 and total alpha-synuclein. Glucocerebrosidase activity was measured through an established fluorescence assay. Results: We observed relatively higher levels of pSer129 alpha-synuclein in the olfactory bulb and cortex of both young and aged mice, compared to other brain regions; while only aged mice had a marked increase of pSer129 alpha-synuclein in the striatum. Both young and aged mice had significantly elevated levels of glucocerebrosidase activity in the mid-brain and olfactory bulb, while the other brain regions were similar in activity levels. However, the levels of glucocerebrosidase activity in aged mice were generally lower. Conclusions: Our results suggest that the age-dependent striatal accumulation of pSer129 alpha-synuclein, and the general decrease of glucocerebrosidase activity, may contribute to an increased risk of PD pathology. Furthermore, our data also implies that both the levels of pSer129 alpha-synuclein and glucocerebrosidase activity are modulated in a brain-region, or possibly neuronal type, dependent manner.

P450 / #1286


ABERRANT TFEB SUBCELLULAR LOCALIZATION IN NIGRAL NEURONS OF PATIENTS WITH GBA-RELATED AND SPORADIC PARKINSON’S DISEASE AND DEMENTIA WITH LEWY BODIES

Lecture Title:

M.L. Morella1, T. Moors1, H. Geut1, F. Bertran-Cobo1, E. Timmermans1, A. Ingrassia1, V. Udayar2, V. Bonifati3, R. Jagasia2, W. Van De Berg1 1Amsterdam UMC - Location VUmc, Anatomy And Neurosciences, Amsterdam, Netherlands, 2F-Hoffmann-La Roche Ltd, Roche Pharma Research And Early Development, Basel, Switzerland, 3Erasmus Medical Center, Clinical Genetics, Rotterdam, Netherlands

Aims: Transcription factor EB (TFEB) controls autophagosome and lysosomal homeostasis, processes implicated in the pathogenesis of Parkinson’s disease and dementia with Lewy bodies, both sporadic (sPD/DLB) and GBA-related (GBA-PD/DLB). TFEB regulates target genes of the Coordinated Lysosomal Expression and Regulation (CLEAR) network. We aimed at studying whether TFEB subcellular localization is impaired in Ser129-phosphorylated alpha-Synuclein (pSer129-aSyn) immunoreactive nigral neurons in GBA-PD/DLB and sPD/DLB and whether it is associated with changes in the expression of CLEAR genes. Methods: TFEB and pSer129-aSyn subcellular localization was analyzed in substantia nigra (SN) of age-matched individuals with incidental Lewy body disease (iLBD, N=3), GBA-PD/DLB (N=9), sPD/DLB (N=9), and non-demented controls (N=7). 441 nigral dopaminergic neurons (≥98 per group) were scanned using confocal and STED microscopy. Expression of CLEAR genes (i.e. GALC, HEXA, MAP1LC3A, SQSTM1, UGCG, VPS35 and GBA) was measured using qPCR. Results: Reduced nuclear TFEB immunoreactivity was observed in iLBD, GBA-PD/DLB and sPD/DLB compared to controls, and was most pronounced in GBA-PD/DLB and in pSer129-aSyn positive neurons. Furthermore, we observed previously unidentified TFEB-immunopositive clusters localized at the trans-Golgi network. Notably, the proportion of neurons displaying TFEB clusters was increased in the disease groups compared to controls, both in presence and absence of pSer129-aSyn. Expression of CLEAR genes was downregulated in both PD/DLB groups compared to controls in SN (-10%) and frontal cortex (-22%).

Conclusions: This study demonstrates a role of aberrant TFEB localization and reduced CLEAR gene expression in GBA- and sPD/DLB. The results suggest that impaired TFEB homeostasis acts upstream of pSer129-aSyn accumulation in PD/DLB.

P451 / #1678


INVESTIGATION OF FIBRIL-INDUCED AGGREGATION, DEGRADATION AND RELEASE OF ΑLPHA-SYNUCLEIN IN INDUCIBLE NEUROBLASTOMA CELLS

Lecture Title:

M. Pantazopoulou, A. Lamprokostopoulou, K. Vekrellis, L. Stefanis Biomedical Research Foundation of the Academy of Athens, Translational Medicine, Athens, Greece

Aims: A major pathological feature of PD is the accumulation and transmission of misfolded assemblies of alpha-synuclein (AS). The current study aims at deciphering the mechanisms governing aggregation, degradation and release of AS aggregates in neuronal cells. Methods: Using recombinant fibrils (PFFs) in neuronally differentiated SH-SY5Y cells with inducible expression of AS, we examined aggregated AS turnover and release through exosomes by immunoblotting and immunocytochemistry assays. Results: PFF incubation leads to the seeding and aggregation of endogenous AS and to the release of exosomes loaded with AS oligomeric species. Lysosomal inhibition resulted in the accumulation of PFF-induced SDS-soluble AS but not to increased oligomeric AS load in exosomes. Inhibition of the Sumoylation Pathway increased AS aggregation and release through exosomes, that both were more prominent when combined with lysosomal inhibition. Phosphorylated at S129 AS oligomers were only apparent at 7 days of high-dose PFF incubation. Proteasomal inhibition led to further accumulation of phosphorylated AS aggregates, without affecting their release through exosomes. Conclusions: Our data suggest the involvement of both the Lysosome and the Proteasome in the clearance of AS assemblies. The Lysosomal Pathway is responsible for the turnover of fibrillar AS and Sumoylation seems to play a role in this process. However, inhibition of the Sumoylation Pathway, and not of the Lysosome, enhances exocytosis of AS oligomers through exosomes suggesting a role of this post-translational modification in exosome release. On the other hand, the Proteasome System is responsible, at least in part, for the selective clearance of phosphorylated AS aggregates.

P452 / #457


RESTORATION OF BEHAVIORAL ALTERATIONS BY ACTIVATING AUTOPHAGY IN MOUSE MODELS SIMULATING PARKINSON'S DISEASE WITH WEAKENED NEUROINFLAMMATION

Lecture Title:

A. Pupyshev, M. Tenditnik, M. Ovsyukova, A. Akopyan, N. Dubrovina, M. Tikhonova Scientific Research Institute of Physiology and Basic Research, Laboratory Of Experimental Models Of Neurodegenerative Processes, Novosibirsk, Russian Federation

Aims: In Parkinsons disease (PD) models induced by acute neurotoxin treatment, combined damage of neurons and microglia in the nigrostriatal system leads to behavioral impairments. However, chronic course of PD in humans is not accompanied by pronounced neuroinflammation. Possibilities for correcting the behavioral disorders of PD models with weakened neuroinflammation by activating autophagy through the mTOR-dependent (rapamycin) and mTOR-independent (trehalose) pathways were studied. Methods: We applied autophagy-stimulating therapy to MPTP-induced disease model in C57Bl/6J mice at the "postponed" mode (7 days after intoxication) and transgenic model at early stage of the disease (B6.Cg-Tg (Prnp-SNCA * A53T) 23Mkle / J mice, 5 m.o.). Rapamycin was injected i.p. at a dose of 10 mg/kg body weight seven times every other day for two weeks. Trehalose was added to the drinking water (2% solution) for the same period. Behavior was estimated in the open field test, passive avoidance learning test and Rotarod. Results: The passive avoidance test showed a sharp decrease in avoidance latency and restoration of cognirive function under action of rapamycin, trehalose and their combination in the MPTP-induced model with weakened inflammation. However the recovery was some weaker than in an "acute" model (1 day after intoxication) of PD. In the transgenic PD model, a motor-cognitive activity at variable speed of rod rotation in the Rotarod test was restored in the drug-treated mice. The recovery was suppressed by autophagy inhibitor 3-methyladenine. Conclusions: Rapamycin and trehalose restored partially behavior and cognitive altersations in MPTP-induced models of PD with weakened inflammation. The recovery did not depend on a pathway of autophagy stimulation but was sensitive to autophagy inhibition.

P453 / #1386


RETROMER REGULATES SYNAPTIC TRANSMISSION AND IS REQUIRED FOR ENDOLYSOSOMAL FUNCTION IN THE AGING BRAIN

Lecture Title:

H. Ye1, S. Ojelade1, D. Li-Kroeger1, Z. Zuo2, N. Seyfried3, H. Bellen2, J. Shulman1 1Baylor College of Medicine, Neurology, Houston, United States of America, 2Baylor College of Medicine, Molecular And Human Genetics, Houston, United States of America, 3Emory University School of Medicine, Department Of Neurology, Atlanta, United States of America

Aims: To understand the requirements of retromer in neurons and particularly within the aging nervous system, and its link to Parkinson’s disease (PD) susceptibility. Methods: The retromer, including VPS35, VPS26, and VPS29, is a conserved protein complex responsible for recycling proteins within the endolysosomal pathway. In Drosophila, we generated a Vps29 null allele using CRISPR/Cas9 technology. Since Vps29 null homozygotes are adult viable, we evaluated age-dependent survival, startle-induced negative geotaxis, electrophysiology, and Transmission Electron Microscopy to reveal evidence of potential neuronal dysfunction. Western blotting and immunofluorescence were also performed to evaluate other retromer components and the integrity/function of the endolysosomal system. Results: We find that Vps29 is dispensable for embryogenesis but is required for proper retromer function in aging adults, including for synaptic transmission, survival, and locomotion. Unexpectedly, in the absence of Vps29, Vps35 and Vps26 proteins were normally expressed and associated, but retromer was mislocalized from neuropil to soma in association with the Rab7 GTPase. With aging, retromer insufficiency triggered progressive endolysosomal dysfunction, with ultrastructural evidence of impaired substrate clearance and resulting lysosomal stress. Conclusions: Loss of Vps29 in Drosophila causes progressive synaptic dysfunction, impaired locomotor behavior, and toxic neuronal accumulation, thus recapitulating salient features of PD, and establishing a valuable model for studying the role of the retromer and associated endolysosomal biology in the aging, adult nervous system. We are currently incorporating comparative quantitative proteomics to further Investigate the role of retromer in synaptic vesicle recycling and lysosomal homeostasis during aging following retromer insufficiency.

P454 / #1393

Topic: Theme C: α-Synucleinopathies / C1.e. Disease Mechanisms, Pathophysiology: Lipids, lipoproteins and membrane trafficking

CONTRASTING THE PATHOMECHANISMS OF MEMBRANE VERSUS CYTOSOL ALPHA-SYNUCLEIN EXCESS

Lecture Title:

A. Tripathi, E. Kantor, H. Alnakhala, N. Ramalingam, L. Liu, D. Selkoe, U. Dettmer ARCND-Brigham and Women's Hospital-Harvard Medical School, Dept Of Neurology, Boston, United States of America

Aims: Parkinson’s disease (PD) is characterized by neuronal loss and associated with aggregation of the vesicle membrane-associated protein alpha-synuclein (αS). Certain familial PD-linked missense mutations in αS lead to increased membrane interaction (E46K) while others (G51D) lead to decreased membrane association, yet both cause PD. We are interested in understanding the different stress signatures elicited by E46K and G51D to gain insight into their role in disease etiology. Methods: We are using a novel protein engineering strategy based on the KTKEGV repeat structure of aS to create mutants that exacerbate the effects of single point mutants. These mutants were expressed in human neural cells and neurons to establish toxicity models. Results: Expression of both 3D (strategic amplification of G51D) and 3K (strategic amplification of E46K) variants in human neural cells invariably leads to reduced growth and viability. In neurons, both cause decreased neurite outgrowth, and 3D and 3K exacerbate subtle effects seen with single mutant 1D and 1K, respectively. Conclusions: Our novel strategically engineered mutants, 3D and 3K, elicit stress and characteristics of neurodegeneration in cellular models. We will present novel insight into stress signatures that are shared between or specific for membrane vs. cytosol αS accumulation and discuss implications for targeted drug discovery.

P455 / #1467

Topic: Theme C: α-Synucleinopathies / C1.f. Disease Mechanisms, Pathophysiology: Inflammation

CXCR4 REGULATES TH17 T CELL BRAIN HOMING IN PARKINSON’S DISEASE

Lecture Title:

D. Gate, T. Wyss-Coray Stanford University, Chem-h/neuro Research Complex, Stanford, United States of America

Aims: Recent studies indicate a role of the adaptive immune system in Parkinson’s disease (PD). However, the mechanisms regulating T cell brain homing remain unknown in PD. We demonstrate a physical interaction of T cells with phosphorylated α-synuclein and with dopaminergic neuronal processes in the substantia nigra of post-mortem PD brains. Methods: Using single cell RNA sequencing of cerebrospinal fluid (CSF), we identified the G protein-coupled receptor C-X-C Motif Chemokine Receptor 4 (CXCR4) as differentially expressed by clonally expanded CD4+ T cells in PD. Results: Protein levels of the CXCR4 ligand, C-X-C Motif Chemokine Ligand 12 (CXCL12) were associated with cognitive decline in PD. Within the CSF, clonally expanded CD4+ T cells expressed marker genes corresponding to Th17 cells. Finally, PD patient T cells upregulated IL-17 upon stimulation with a phosphorylated α-synuclein peptide ex vivo. Conclusions: Cumulatively, these results indicate the CXCR4-CXCL12 signaling axis as a mechanistic target for inhibiting pathological Th17 T cell entry into the PD brain.

P456 / #969

Topic: Theme C: α-Synucleinopathies / C1.f. Disease Mechanisms, Pathophysiology: Inflammation

DECREASED ALPHA-SYNUCLEIN IN CEREBROSPINAL FLUID COINCIDES WITH ALTERED CYTOKINE LEVELS IN MILD COGNITIVE IMPAIRMENT AND ALZHEIMER’S DISEASE PATIENTS

Lecture Title:

C. Mcdonnell, T. Burke, E. Sokolowska, J. Prenderville Transpharmation Ireland Ltd, Transpharmation Ireland Ltd, Dublin, Ireland

Aims: α-synuclein (a-syn) pathology and inflammation are a hallmarks of Alzheimer’s diseases (AD). Here, we assess cerebrospinal fluid (CSF) levels of a-syn in conjunction with inflammatory markers (IFN-γ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, TNF-α), total tau, phosphorylated tau (threonine 181), and amyloid-β-42. Methods: CSF samples from control, mild cognitive impairment (MCI), and AD donors were acquired from a private biobank. All targets were measured by Meso Scale Discovery assays. One-way ANOVA with Fisher’s least significant difference test was carried out for each target in Graphpad Prism 8. Results: a-syn is significantly lower in MCI (p<0.01) and AD (p<0.05) individuals compared to healthy controls. Total tau is significantly higher in MCI (p<0.001) and AD (p<0.001) individuals compared to healthy controls. Phosphorylated tau is significantly higher in AD (p<0.001) individuals compared to healthy controls. Amyloid-β-42 is significantly lower in MCI (p<0.001) and AD (p<0.001) individuals compared to healthy controls. IL-12 is significantly lower in AD (p<0.05) individuals compared to healthy controls. IL-10 (p<0.01), IL-12 (p<0.01), and IL-8 (p<0.05) are significantly higher in AD individuals compared to healthy controls. TNF-alpha is significantly higher in MCI (p<0.01) and AD (p<0.001) individuals compared to healthy controls. Conclusions: The data presented here supports previous work indicating both altered cytokine levels and decreased a-syn in the CSF of MCI and AD individuals. Decreased amyloid-β-42 and a-syn in the CSF may coincide with increased aggregation in the brain that drives inflammation, driving inflammation as MCI progresses to AD.

P457 / #402

Topic: Theme C: α-Synucleinopathies / C1.g. Disease Mechanisms, Pathophysiology: Microglia

IMMUNE AND PHAGOCYTIC DYSFUNCTIONS IN HUMAN MICROGLIA CELLS DERIVED FROM LRRK2-G2019S PARKINSON’S DISEASE IPSCS

Lecture Title:

L. Blasco-Agell1,2, M. Pons-Espinal1,2, G. Carola1,2, C. Calatayud1,2, I. Fernandez-Carasa1,2, M. Juan-Otero3,4, E. Tolosa4,5, A. Raya6,7,8, A. Consiglio1,2,8,9 1Bellvitge Biomedical Research Institute (IDIBELL), Pathology And Experimental Therapeutics (university Barcelona), Hospitalet de Llobregat, Spain, 2Institute of Biomedicine of the University of Barcelona (IBUB), University Of Barcelona, Barcelona, Spain, 3Hospital Clinic, 4. department Of Immunology, Barcelona, Spain, 4August Pi i Sunyer Institute for Biomedical Research, University Of Barcelona, Barcelona, Spain, 5Hospital Clinic, Deparment Of Neurology, Barcelona, Spain, 6Bellvitge Biomedical Research Institute, Program For Advancing The Clinical Translation Of Regenerative Medicine Of Catalonia, Hospitalet de Llobregat, Spain, 7Centre for Networked Biomedical Research on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), (ciber-bbn), Madrid, Spain, 8Institucio Catalana de Recerca i Estudis Avançats, (icrea), Barcelona, Spain, 9University of Brescia, Department Of Molecular And Translational Medicine, Brescia, Italy

Aims: Parkinson’s disease (PD) is a neurodegenerative disease characterized by a progressive loss of dopaminergic neurons in the substantia nigra and misfolded α-synuclein deposition inside neurons. Associated with these pathological hallmarks, neuroinflammatory alterations are also present including microglial activation, astrocyte reactivity and lymphocyte infiltration. However, whether glial cells contribute of PD pathogenesis remain poorly understood. Methods: We generated human Microglia cells (hMG) from induced Pluripotent Stem Cells (iPSC) of PD patients carrying the G2019S mutation in the LRRK2 gene or from isogenic corrected lines. Results: We report that iPSC derived hMG expressed key microglial-specific markers, such as TMEM119, which strongly co-localized with the myeloid marker IBA-1, confirming identity of brain-resident-like microglia. Under non-stimulated condition, LRRK2 hMG expressed similar pro-inflammatory cytokines as their gene-edited isogenic counterparts. In contrast, stimulation of LRRK2 hMG with LPS elicits a strong pro-inflammatory response, compared with isogenic hMG cells. Moreover, when we assessed phagocytic activity using pHrodo red-labeled synaptosomes, LRRK2 hMG was found to be highly phagocytic under non-stimulation. However, upon LPS stimulation, PD hMG failed to respond as the isogenic hMG, since we did not detect any increase in their phagocytic capacity. Finally, to evaluate whether and how neuroinflammation contributes to PD, we are currently evaluating the neuronal response to neuroinflammatory signaling using an advanced co-culture system in which astrocytes, microglia and neurons cultured together in 2D culture or in complex 3D organoids. Conclusions: So far, our data suggest a misbalanced microglial pro-healing and pro-killing properties within LRRK2 PD hMG, that could help to identify potential novel therapeutic approaches.

P458 / #572

Topic: Theme C: α-Synucleinopathies / C1.g. Disease Mechanisms, Pathophysiology: Microglia

GLIAL-NEURONAL INTERACTION PROMPTED BY RECOMBINANT FIBRILLAR PROTEIN CONSTRUCTS OF ALPHA-SYNUCLEIN – AN IN VITRO CHARACTERISATION

Lecture Title:

C. Tyndall1, T. Burke1,2, D. Kozareva1,2, J. Prenderville1,2, E. Sokolowska2, A. Netter-Glangeaud3, A. Louwrier3, A. Harkin1 1Neuropsychopharmacology Research Group, Trinity College Institute Of Neuroscience, Dublin, Ireland, 2Transpharmation Ireland Ltd., Trinity College Institute Of Neuroscience, Dublin, Ireland, 3StressMarq Biosciences Inc., Suite 118, 1537 Hillside Ave, Victoria, Canada

Aims: Recombinant fibrillar protein constructs of alpha-synuclein have recently been developed in efforts to establish new cell and animal model systems in which representative synucleinopathy-related pathology is generated more rapidly than current models. In the present investigation, we explore in primary cultures the glial-neuronal interactions triggered by the administration of a variety of monomeric, oligomeric, and fibrillar alpha-synuclein preparations provided by StressMarq Biosciences Inc. Methods: Primary rat cortical neuronal, mixed glial, enriched microglial, and astroglial cultures are treated for 14 days with monomer-seededSPR-321 preformed fibrils (PFFs); type 1SPR-322, type 2SPR-317, or type 3SPR-

448 PFFs; A53T mutant alpha-synuclein PFFsSPR-326; soluble filamentsSPR-450; or dopamine-stabilised oligomersSPR-466. Glial conditioned media (CM) is collected, filtered, and used to treat neurons for 48 hours. Delivery of the alpha-synuclein gene using an adeno-associated viral (AAV) vector is an effective means to drive protein expression within neurons over time. Neurons are treated with the AAV5 serotype encoding human wildtype alpha-synuclein (AAV5-CBA-alpha-synuclein1x10^13μg/ml). Following transfection of neurons, microglia are treated with CM of alpha-synuclein-expressing neurons. Immunocytochemistry, qPCR, and multiplex infrared immunoassays are performed to determine cell morphology, activation state, viability, and inflammatory profile. Results: These preparations have shown to generate representative synucleinopathy-related pathology in vitro, informing the study of glial-neuronal interactions associated with alpha-synuclein aggregation. Conclusions: Alpha-synuclein fibrils are shown to “seed” their own monomers into more complex structures during aggregation. As such, we propose the in vitro cellular investigation of fibrillar alpha-synuclein protein constructs to be the optimal approach for exploring in a time frame of days what occurs over years in humans.

P459 / #587

Topic: Theme C: α-Synucleinopathies / C1.h. Disease Mechanisms, Pathophysiology: Astroglia

CHARACHTERIZATION OF LRP10 EXPRESSION IN LEWY BODY DISORDERS AND IPSC-DERIVED BRAIN CELLS

Lecture Title:

M. Grochowska1, A. Carreras Mascaro1, V. Boumeester1, H. Geut2,3, E. Sammler4,5, I. Huitinga3, M. Ghazvini6, P. Parchi7,8, P. Cortelli8,9, W. Van De Berg2, D. Alessi4, V. Bonifati1, W. Mandemakers1 1Erasmus Medical Center, Clinical Genetics, Rotterdam, Netherlands, 2VU University Medical Center, Anatomy And Neurosciences, Section Clinical Neuroanatomy Ao2|m, Amsterdam, Netherlands, 3Netherlands Institute for Neuroscience, Netherlands Brain Bank, Amsterdam, Netherlands, 4University of Dundee, School of Life Sciences, Medical Research Council (mrc) Protein Phosphorylation And Ubiquitylation Unit, Dundee, United Kingdom, 5University of Dundee, School of Medicine, Ninewells Hospital, Neurology, Dundee, United Kingdom, 6Erasmus Medical Center, Developmental Biology, Rotterdam, Netherlands, 7University of Bologna, Experimental, Diagnostic And Specialty Medicine, Bologna, Italy, 8Institute of Neurological Sciences of Bologna (ISBN), Istituto Di Ricovero E Cura A Carattere Scientifico (irccs), Bologna, Italy, 9Alma Mater Studiorum-University of Bologna, Dipartimento Di Scienze Biomediche E Neuromotorie (dibinem), Bologna, Italy

Aims: Loss-of-function mutations in LRP10 (low-density lipoprotein receptor-related protein 10) have been implicated in the development of Parkinson’s disease (PD), Parkinson’s Disease Dementia (PDD), and Dementia with Lewy Bodies (DLB). Yet, little is known about LRP10 function and expression in the central nervous system in health and disease. Our objective is to perform an in-depth characterization of endogenous LRP10 expression in human iPSC-derived brain cell types and adult human brains from healthy individuals and Lewy Body Disorder (LBD) patients. Methods: Generation and validation of LRP10 antibodies. Generation of human iPSC-derived progenitors, ventral midbrain dopaminergic neurons, and astrocytes. Immunocytochemistry (ICC), immunohistochemistry (IHC), and biochemical analysis to study LRP10 (sub-)cellular distribution in iPSC-derived cells, and in healthy and LBD adult human brains. Results: Using two knockout validated antibodies binding to different LRP10 protein domains, we show that iPSC-derived astrocytes have higher LRP10 protein expression when compared to neural progenitors or ventral midbrain dopaminergic neurons. Furthermore, we demonstrate by IHC that LRP10 expression is mainly restricted to astrocytes and neurovasculature in adult human brain. Finally, we show that at the subcellular level, endogenous LRP10 is localized to vesicular structures, including trans-Golgi network, endosomes, multivesicular bodies, as well as the plasma membrane. Conclusions: High vesicular expression in astrocytes and lack of expression in dopaminergic neurons points to potential involvement of LRP10 in LBD through non-cell-autonomous mechanisms. Future research will explore the crucial roles of astrocytes and more specifically LRP10 in disease mechanisms of LBD.

P460 / #1044

Topic: Theme C: α-Synucleinopathies / C1.i. Disease Mechanisms, Pathophysiology: Cellular signaling, kinases, phosphatases, calcium

HUMAN-SPECIFIC LEVELS OF CDNF, MANF AND ER STRESS IN THE PATHOLOGICAL PROGRESSION OF PARKINSON’S DISEASE

Lecture Title:

L. Parkkinen1, D. Hrabos1, P. Pulkkila2, I. Poggiolini1, M. Saarma2 1University of Oxford, Nuffield Department Of Clinical Neurosciences, Oxford, United Kingdom, 2University of Helsinki, Institute Of Biotechnology, Helsinki, Finland

Aims: To establish whether levels of Endoplasmatic Reticulum (ER)-stress responsive neurotrophic factors, Cerebral Dopamine Neurotrophic Factor (CDNF) and Mesencephalic Astrocytes Derived Neurotrophic Factor (MANF) are altered during the progression of pathology in Parkinson’s and whether these alterations are related to a-synuclein (aSyn) or ER-stress. The expression and secretion of CDNF and MANF increase with ER-stress, they protect dopamine neurons in animal models of Parkinson’s and CDNF also decreases intracellular aSyn aggregation. Methods: CDNF and MANF were analysed by sandwich ELISA, aSyn and phosphorylated/total eIF2alevels with automatized Western blotting and aSyn seeding with RT-QuIC. In addition, we analysed mRNA levels of above and other ER-stress targets. The levels were analysed 45 subjects selected from Oxford Brain bank in four brains regions: substantia nigra (SN), striatum, entorhinal cortex and occipital cortex (OccCx) and compared between three groups: 12 healthy controls without pathology, 19 cases with incidental Lewy body pathology (Braak stages 1-4) and 14 Parkinson’s patients (Braak stages 5-6). Results: Although we could not detect significant differences in MANF and CDNF protein levels between study groups, we were able to show that the levels of MANF significantly decreased in SN with immunohistochemistry during Parkinson’s disease progression. aSyn seeding (but not total levels) significantly increased with the progression of Parkinson’s pathology. MANF levels correlated with phosphorylated/total eIF2abut not with aSyn. Conclusions: MANF but not CDNF levels are related to the extent of ER-stress in Parkinson’s brain but not to aSyn levels.

P461 / #348

Topic: Theme C: α-Synucleinopathies / C1.i. Disease Mechanisms, Pathophysiology: Mitochondrial dysfunction, oxidative damage

Α-SYNUCLEIN HAS DISTINCT IN VIVO ROLES IN MITOCHONDRIAL HOMEOSTASIS-A MITOCHONDRIAL PATHOGENIC PATHWAY FOR THE INITIATION OF PARKINSON’S DISEASE

Lecture Title:

S. Gunawardena, T. Krzystek, R. Banerjee The State University of New York at Buffalo, Biological Sciences, Buffalo, United States of America

Aims: The balance between fission (via dynamin-related protein, DRP1) and fusion (via mitofusin, MFN), which occur in concert with mitochondrial motility and organelle turnover mediated by PTEN-induced kinase 1 (PINK)/ E3 ubiquitin ligase (Parkin) is essential for mitochondrial homeostasis with dysfunction causing mitochondrial disease and neurodegeneration. Methods: Here we evaluate the physiological relevance of α-syn during mitochondrial homeostasis in a humanized Drosophila model of Parkinson’s disease (PD). Results: We show that excess α-syn cause fragmented mitochondria, which persist with either truncation of the C-terminus (α-syn1-120) or deletion of the NAC region (α-synΔNAC). α-syn-mediate fragments were oxidized as measured by in vivo oxidation markers Mito-roGFP2-ORP1/GRX1 and MitoTimer, but fragments induced by α-syn1-120 were healthy. α-syn-mediated oxidized fragments showed biased retrograde motility, but α-syn1-120-mediated healthy fragments did not. Depletion/inhibition or excess DRP1 rescued α-syn-mediated fragmentation and oxidation. Excess PINK1/Parkin rescued α-syn-mediated fragmentation and membrane depolarization, likely via functional associations with the C-terminus. Conclusions: Taken together, our findings identify distinct physiological roles for the N and C-terminus of α-syn in mitochondrial dynamics, highlighting a previously unknown pathogenic pathway for the induction of Parkinson’s Disease.

P462 / #965


UNCOVERING THE MECHANISMS OF MITOCHONDRIAL RESCUE WITH THE USE OF A NOVEL THERAPEUTIC COMPOUND FOR THE TREATMENT OF SPORADIC PARKINSON’S

Lecture Title:

R. Hughes1, C. Hastings1, G. Packer2, A. Weymouth-Wilson2, O. Bandmann1, H. Mortiboys1 1Sheffield Institute for Translational Neuroscience, Neuroscience, Sheffield, United Kingdom, 2NZP UK Ltd, Nzp Uk Ltd, Reading, United Kingdom

Aims: There is strong evidence of mitochondrial dysfunction in sporadic and familial Parkinson’s disease (PD). Previous work highlighted mitochondrial restoration properties of ursodeoxycholic acid (UDCA) in fibroblasts from people with sporadic PD (sPD) which led to a collaboration with an industrial partner. Following a comprehensive drug screen, we identified a lead compound, compound A, which showed potent mitochondrial restorative properties in sPD fibroblasts. We aim to investigate cellular mechanisms altered by treatment with compound A in dopaminergic neurones derived from sPD fibroblasts. Methods: Fibroblasts from sPD patients and neurologically healthy donors (3 and 3) were re-programmed, generating induced neuronal progenitor cells, which underwent a 27-day differentiation, producing induced dopaminergic neurones. Neuronal and dopaminergic markers and mitochondrial membrane potential (MMP) were assessed. Basal mitophagy was assessed using live time-lapse fluorescent microscopy while induced mitophagy and alpha synuclein were investigated using immunocytochemistry. Experiments were conducted on iNeurones treated with vehicle, Compound A or UDCA. Results: iNeurones achieved approximately 90% tyrosine hydroxylase positive cells with no effect of compound A or UDCA. MMP increased following treatment with UDCA or compound A (73% and 100% respectively). Alterations were found between control and Parkinson’s iNeurones in mitophagy and total alpha synuclein content, however no drug effect was seen. Conclusions: Compound A does not elicit its effect via mitophagy or changes in total alpha synuclein. We continue to study the mechanism of compound A and future work will knockdown a potential target in the iNeurones to investigate whether the rescue effects of compound A are ablated.

P463 / #974


SINGLE-CELL ANALYSIS OF ALPHA-SYNUCLEIN-INDUCED MITOCHONDRIAL DYSFUNCTION IN-VIVO

Lecture Title:

S. O'Sullivan, S. Lee, A. Ulusoy, B. Jevans, D. Di Monte The German Center for Neurodegenerative Diseases (DZNE), Fundamental Research, Bonn, Germany

Aims: Parkinson’s disease (PD) is characterised by progressive degeneration of discrete neuronal populations, in particular, dopaminergic cells in the substantia nigra pars compacta (SN). PD pathogenesis is also linked to accumulation and aggregation of alpha-synuclein within neurons of various brain regions, including the SN. Clinical and experimental evidence supports a vital role for mitochondrial dysfunction in the pathogenesis of both familial and sporadic PD. Our aim was to develop a quantitative assessment of potential mitochondrial dysfunction at the single-cell level in a mouse model of alpha-synuclein overexpression. Methods: To over-express alpha-synuclein, AAVs carrying the DNA for human alpha-synuclein were injected into the SN. After 12 weeks, animals were perfused, and quadruple immunofluorescent staining was carried out using anti-tyrosine hydroxylase (TH, a marker of dopaminergic cells), anti-human alpha-synuclein, anti-Grp75 (a mitochondrial marker) and anti-GRIM19 (a marker of mitochondrial complex I). Results: Alpha-synuclein overexpression caused the degeneration of approximately 40% of TH-immunoreactive SN neurons. Quantitative fluorescence measurements were done within surviving cells that were positive for both TH and alpha-synuclein. Data showed a significant decrease in GRIM19:Grp75 ratio. We also found a negative correlation between levels of alpha-synuclein expression and GRIM19:Grp75 ratio. Conclusions: These results indicate that higher levels of alpha-synuclein expression are associated with mitochondrial impairment and, in particular, a loss of mitochondrial complex I. Ongoing work will determine the progression of these changes by measuring them at earlier and later time points. Further assessment of specific mitochondrial damage will involve quantitative analyses using markers of other respiratory chain complexes (e.g., complex IV).

P464 / #710

Topic: Theme C: α-Synucleinopathies / C1.k. Disease Mechanisms, Pathophysiology: Synapse pathology, neural networks, plasticity, neurogenesis

HIGH-THROUGHPUT ELECTROPHYSIOLOGICAL PHENOTYPING OF IN VITRO-RECONSTITUTED PD PATIENT-SPECIFIC NIGROSTRIATAL CIRCUITS

Lecture Title:

C. Calatayud Aristoy1, A. Chouhan1, L. Heinrich2, D. Lambrechts3, B. Miccoli3, V. Uytterhoeven1, O. Krylychkina3, A. Geens1, W. Vandenberghe4, B. Schüle2, D. Braeken3, P. Verstreken1 1Center for Brain & Disease Research, VIB-KU Leuven, Laboratory For Neuronal Communication, Leuven, Belgium, 2Stanford University School of Medicine, Department Of Pathology, Stanford, United States of America, 3Imec, Life Sciences, Leuven, Belgium, 4UZ Leuven - Faculty of Medicine, KU Leuven, Department Of Neurology, Leuven, Belgium

Aims: The main objective is to link the molecular etiology of idiopathic Parkinson´s disease (PD) cases to that of monogenic cases. A second objective is to elaborate a catalogue of the electrophysiological signatures associated with the different molecular pathways defective in PD. Methods: Cortical excitatory (CN), striatal projection (SPN), and nigral dopamine neurons (DAN) were generated on top of CMOS high-density multi-electrode (MEA) microchips by differentiating stem cells from control and patient-specific human stem cells through the forced expression of lineage-specific transcription factors. Different electrophysiological and network parameters were recorded using the chip's in-built electrodes. Results: Neurons generated on the CMOS-MEA chips expressed markers typical of CN, SPN, and DAN, and established functional connections with each other, hence forming synthetic nigrostriatal-like microcircuits. These circuits showed electrophysiological features reminiscent of their in vivo brain counterparts. Conclusions: We have built an experimental platform capable of capturing fine electrophysiological and network defects in human stem cell-derived nigrostrital circuits. This technology will be used to define electrophysiological signatures attributable to genetic defects associated with PD.

P465 / #1331


MOTOR RESERVE IN PARKINSON’S DISEASE: MODEL OPTIMIZATION AND CORTICAL CORRELATES

Lecture Title:

V. Dzialas1, M. Hönig1,2, G. Bischof1, A. Drzezga1,2,3, T. Van Eimeren1,3,4, P.P.M.I. (Ppmi)5 1University of Cologne, Nuclear Medicine, Köln, Germany, 2Research Center Jülich, Neuroscience And Medicine Ii, Jülich, Germany, 3German Center for Neurodegenerative Diseases, Positron Emission Tomography (pet), Bonn, Germany, 4University of Cologne, Neurology, Köln, Germany, 5The Michael J. Fox Foundation, Ppmi/institute For Neurodegenerative Disorders, New York, United States of America

Aims: To elucidate the emerging concept of motor reserve (MR) regarding its quantification (i.e. residual approach) and neurobiological underpinnings, we assessed the association between UPDRS-III sub-scores, regional dopamine transporter (DaT) loss, and structural network integrity in de novo Parkinson’s disease (PD) patients. Methods: Data of 151 PD patients (Mage 58.7±4.5) were included, for whom a MRI, DaT-SPECT and clinical information were available at the PPMI database. First, using a correlational approach, we determined the best-fit model between regional DaT loss and UPDRS-III sub-scores, considering hemisphere, brain region (i.e. putamen, caudate, striatum), bodyside and symptom class (i.e. limb-akinetic, axial, tremor) based differences. Subsequently, MR-residuals were derived from the best-fit model and split into a high (n=50) and low (n=45) MR group. To validate the MR-residuals, we compared the physical activity score of the elderly (PASE) between groups. Further, the neurobiological substrate of MR was assessed by voxel-wise comparison of gray-matter-volume (GMV) maps (p<.005, uncorrected) with the CAT12 toolbox and derived structural connectivity networks for the respective groups using the GAT toolbox. Results: Putaminal DaT signal and akinetic-rigid (limb and axial) UPDRS-III items of the less-affected hemisphere and contralateral bodyside showed the closest association compared to all other models. The high MR group presented a significantly higher PASE score (p=.045), larger GMV (Figure1) and network nodes in motor-associated brain regions.

Figure1 GMV comparison with high>low MR marked in blue.

Conclusions: As indicated by structural differences of underlying motor networks, MR can sensitively be quantified by the association between less-affected putaminal DaT signal and akinetic-rigid motor signs.

P467 / #1377


ΑLPHA-SYNUCLEIN DISRUPTS NEURON NETWORK BURSTS WHEN OVEREXPRESSED IN CULTURED NEURONS

Lecture Title:

K. Leite, S. Kügler, P. Garg, M. Bähr University Medical Center Göttingen, Department Of Neurology, Göttingen, Germany

Aims: 1.) The first aim was to determine if α-synuclein overexpression affects the network bursting activity of cultured neurons. 2.) The second aim was to determine if network bursting is affected more by extracellular or intracellular α-synuclein. 3.) The third aim was to determine the pathways that are used by α-synuclein overexpression to affect network bursting. Methods: 1.) For modelling, cultured embryonic rat cortical neurons were used. 2.) Calcium sensors were used to visualize the network bursts, following which the calcium changes of individual cells were used to identify and analyze network bursting. 3.) Immunocytochemistry and fluorescent sensors were used to characterize the effect of α-synuclein overexpression on neurites, synapses and the signaling molecule cyclic AMP. Results: 1.) α-synuclein overexpression affects the frequency of neuron network bursting. 2.) The effect of α-synuclein overexpression appears to be mediated by intracellular α-synuclein. 3.) α-synuclein overexpression also affects neurites, synapses and cytosolic levels of cyclic AMP. Conclusions: α-synuclein overexpression affects neuron network bursting activity and also damages neurites, synapses and cytosolic cyclic AMP levels, with the effect driven more by intracellular α-synuclein.

P468 / #1295


LIGHT-INDUCIBLE ALPHA-SYNUCLEIN AGGREGATION IN THE MIDBRAIN IMPAIRS NIGROSTRIATAL DOPAMINERGIC TRANSMISSION

Lecture Title:

R. Rodriguez Aller1, S. Malvaut2, M. Bérard1, R. Sheta1, A. Saghatelyan2, A. Oueslati1 1Université Laval, Department Of Molecular Medicine & Chu De Quebec Research Center, Québec, Canada, 2CERVO Brain Research Centre, Department Of Psychiatry And Neurosciences, Quebec, Canada

Aims: Parkinson’s disease (PD) is characterized by intracellular inclusions of misfolded alpha-synuclein, known as Lewy bodies, and by the loss of dopaminergic neurons in the substantia nigra pars compacta (SNc) that leads to dopamine (DA) depletion at the striatum. However, it remains elusive how the aggregates of alpha-synuclein can affect the normal function of dopaminergic projections, especially due to the absence of proper models that can reproduce the features of PD. In this context, we have recently developed an in vitro and in vivo model of PD based on the optogenetics technology named LIPA (light-inducible protein aggregation) that controls the aggregation of alpha-synuclein under the control of blue light. We showed that LIPA mimics the histopathological characteristics of PD, and allow thus to study how the aggregation of alpha-synuclein in dopaminergic cells of SNc can cause a progressive disruption in the nigrostriatal pathway. Methods: To investigate the physiological impact of LIPA-induced alpha-synuclein aggregation on the dopaminergic projections, we assessed the activity of striatal cells. Briefly, we implanted mini-endoscopes coupled with an optic fiber to induce alpha-synuclein aggregation in the SNc, and analyzed the neuronal activity using the calcium indicator GCaMP6s in the striatum of freely moving mice. Results: Our results show a progressive decrease in the synchronized activity of striatal cells caused by the aggregation of alpha-synuclein. Conclusions: Altogether, our data showed that the use of this new LIPA-alpha-synuclein system offers a unique tool to elucidate the morphological, and physiological changes occurring in the dopaminergic projections in the context of PD.

P470 / #833

Topic: Theme C: α-Synucleinopathies / C1.l. Disease Mechanisms, Pathophysiology: Transcriptional & translational regulation, micro RNAs

IMPLICATIONS FOR ALPHA-SYNUCLEIN ANTISENSE TRANSCRIPT SNCA-AS1 IN PARKINSON’S DISEASE: A FOCUS ON SYNAPSES- AND AGING-RELATED PATHWAYS

Lecture Title:

F. Rey1,2, C. Pandini3,4, L. Messa5, R. Zangaglia6, G.V. Zuccotti1,2, S. Gagliardi4, S. Carelli1,2, C. Cereda4 1University of Milan, Department Of Biomedical And Clinical Sciences "l. Sacco", Milan, Italy, 2University of Milano, Pediatric Research Center “romeo Ed Enrica Invernizzi”, Milano, Italy, 3University of Pavia, Department Of Biology And Biotechnology, Pavia, Italy, 4IRCCS Mondino Foundation, Genomic And Post-genomic Center, Pavia, Italy, 5Politecnico di Milano, Department Of Chemistry, Materials And Chemical Engineering "giulio Natta", Milan, Italy, 6IRCCS Mondino Foundation, Parkinson’s Disease And Movement Disorders Unit, Pavia, Italy

Aims: In this work, we aim to characterize SNCA-AS1, antisense transcript to the SNCA gene, and its implications in cellular processes. Methods: SH-SY5Y cells were stably transfected with either SNCA-AS1 or SNCA and their transcriptional signature was investigated via RNA-sequencing. Real Time-PCR and western blot were used to verify SNCA-AS1’s effect on SNCA’s expression, and neurite extension was assessed via immunofluorescence analysis. Results: The overexpression of SNCA-AS1 upregulates SNCA mRNA and protein, and both genes appear to strongly impact neurite extension and synapses’ biology, through specific molecular signatures. We report a reduced expression of markers associated with synaptic plasticity, and we specifically focus on GABAergic and dopaminergic synapses, for their relevance in aging processes and PD, respectively. As part of this signature is co-regulated by the two genes, we discriminate between functions elicited by genes specifically altered by SNCA-AS1 or SNCA’s overexpression, observing a highly relevant role for solely SNCA-AS1. We also highlight how numerous deregulated pathways are implicated in aging-related processes, suggesting that SNCA-AS1 could be a key player in cellular senescence, with implications for aging-related diseases. Indeed, the upregulation of SNCA-AS1 leads to alterations in numerous PD specific genes, with an impact highly comparable to that of SNCA’s upregulation. SNCA/SNCA-AS1 ratio is also significantly altered in PBMCs from PD patients versus healthy controls. Conclusions: Our results show that SNCA-AS1 elicits its cellular functions not only through the regulation of SNCA, but also through a selective and specific modulation of synaptogenesis, senescence and presenting significant implications in PD pathology.

P471 / #1411

Topic: Theme C: α-Synucleinopathies / C1.l. Disease Mechanisms, Pathophysiology: Transcriptional & translational regulation, micro RNAs

THE ROLE OF RNA IN SYNAPSE PHYSIOLOGY AND NEURODEGENERATION IN MODELS OF PARKINSON’S DISEASE

Lecture Title:

M. Xylaki1, B. Atzler1, I. Paiva1, G. Jain1, R. Islam2, Z. Wassouf1, E. Vasili1, M. Al-Azzani1, J. Schulze‐Hentrich3, A. Fischer2, T. Outeiro1 1University Medical Center Goettingen, Experimental Neurodegeneration, Goettingen, Germany, 2German Center for Neurodegenerative Diseases (DZNE), Department For Epigenetics And Systems Medicine In Neurodegenerative Diseases, Göttingen, Germany, 3University of Tübingen, Institute Of Medical Genetics And Applied Genomics, übingen, Germany

Aims: Objectives. Synucleinopathies are characterized by the accumulation of α-synuclein (αsyn), linked with alterations of synaptic function. microRNAs can translocate to synapses and regulate mRNA expression by complementary binding contributing to synaptic remodelling. Here, we sought to identify microRNAs associated with synaptic processes linked to synapse degeneration. Methods: . We performed RNA- and small RNA-Seq of the midbrain of 6 month-old transgenic mice expressing A30P mutant αsyn. A negative correlation between deregulated microRNAs and target genes highlighted the top interacting microRNAs. The most prominent microRNA and its targets were validated by qPCR and immunoblotting. The microRNA or recombinant αsyn species were introduced in primary cortical neurons to study their interactions and effects on synapses. Finally, immunofluorescence and confocal microscopy were used to assess the neuronal morphology. Results: . We identified deregulated biological processes linked with the synaptic compartment and mir101a-3p as a prominent regulator of synaptic plasticity. Mir101a-3p was validated by qPCR in the mouse and in the cortex of Dementia with Lewy Bodies patients. Primary cortical neurons overexpressing mir101a-3p showed altered dendritic and spine morphology. Further correlation with synaptic plasticity was provided by wild-type mice exposed to enriched environment showing reduced levels of mir101a-3p. Finally, exposure of primary cortical neurons to recombinant αsyn species showed a direct effect of αsyn on mir101a-3p levels. Conclusions: . Our data support the emerging role of microRNAs as key regulators of epigenetic alterations associated with αsyn. Identification of RNA-based processes leading synaptic compromise may reveal novel targets for the development of biomarkers and therapeutic interventions in synucleinopathies.

P472 / #1302

Topic: Theme C: α-Synucleinopathies / C1.m. Disease Mechanisms, Pathophysiology: Apoptosis, cell death

IPLA2Β PROTECTS CELLS AGAINST FERROPTOSIS BY HYDROLYZING THE LIPID DEATH SIGNAL, 15-HPETE-PE: RELEVANCE TO PARKINSON’S DISEASE

Lecture Title:

W. Sun1, V. Tyurin2, S. Korolev3, A. Abramov4, P. Angelova4, I. Miller3, G.-W. Mao2, A. Kapralov2, T. Hastings5, J.T. Greenamyre5, C. Chu6, H. Bayır2, Y. Tyurina2, R.-R. He1, V. Kagan2 1Jinan University, Guangdong Engineering Research Center Of Chinese Medicine & Disease Susceptibility, College Of Pharmacy, Guangzhou, China, 2University of Pittsburgh, Department Of Environmental And Occupational Health And Center For Free Radical And Antioxidant Health, Pittsburgh, United States of America, 3Saint Louis University School of Medicine, Department Of Biochemistry And Molecular Biology, St. Louis, United States of America, 4University College London, Ucl Queen Square Institute Of Neurology, Department Of Clinical And Movement Neurosciences, London, United Kingdom, 5University of Pittsburgh, Department Of Neurology, Pittsburgh, United States of America, 6University of Pittsburgh, Department Of Pathology, Pittsburgh, United States of America

Aims: As Ca2+-independent phospholipase PLA2 (iPLA2β), a product of the PNPLA9 gene, can preferentially hydrolyze peroxidized phospholipids, we hypothesized that iPLA2β guards from the ferroptotic 15-hydroperoxy-arachidonoyl-PE (15-HpETE-PE) death signaling. Methods: We purified recombinant iPLA2β and accessed its hydrolytic activity towards ETE-PE and 15-HpETE-PE. As several types of Parkinson’s diseases (PD) are related to mutations in the PNPLA9 gene, we examined iPLA2β hydrolytic activity and sensitivity to ferroptosis of fibroblasts from a patient with a Parkinson’s disease (PD)-associated mutation fPDR747W. LC-MS-based redox phospholipidomics was performed to measure the accumulation of oxidized phospholipids in these cells. To investigate the relationship between iPLA2β deficiency and PD, we generated CRISPR-CAS9-engineered PNPLA9R748W/R748W mice (corresponding to PNPLA9R747W mutation in human), monitored their motor abilities, and measured 15-HpETE-PE level in midbrain. Further, we accessed the level of 15-HpETE-PE and iPLA2β hydrolytic activity in midbrain of SNCAA53T mice and rotenone-infused parkinsonian rats. Results: iPLA2β preferentially hydrolyzes 15-HpETE-PE vs. ETE-PE. fPDR747W cells showed selective loss of 15-HpETE-PE activity, 15-HpETE-PE accumulation and elevation of sensitivity to ferroptosis. CRISPR-CAS9-engineered PNPLA9R748W/R748W mice exhibited progressive parkinsonian motor deficits along with 15-HpETE-PE accumulation. Elevated 15-HpETE-PE levels were detected in midbrain of SNCAA53T mice and rotenone-infused parkinsonian rats.

Conclusions: iPLA2β is a new ferroptosis regulator and its mutations may be implicated in PD pathogenesis.

P473 / #879

Topic: Theme C: α-Synucleinopathies / C1.n. Disease Mechanisms, Pathophysiology: Protein aggregation, misfolding, chaperones

ULTRA-EFFICIENT PMCA OF ALPHA-SYNUCLEIN AGGREGATES FROM PATIENTS WITH MULTIPLE SYSTEM ATROPHY

Lecture Title:

L. Bruyere-Ostells1, J. Verchere2, D. Betemps2, M. Bélondrade1, T. Baron2, D. Bougard1 1Pathogenesis and Control of Chronic Infections, Etablissement Français Du Sang, Inserm, Université De Montpellier, Montpellier, France, 2French Agency for Food, Environmental and Occupational Health and Safety (ANSES), Unité De Maladie Neuro-dégénératives, Lyon, France

Aims: Misfolding and aggregation of alpha-synuclein (a-syn) are key-events of pathogenesis of synucleinopathies. Protein Misfolding Cyclic Amplification (PMCA) is a well-known method for prion seeding activity detection. Considering the emerging knowledge about prion-like properties of a-syn, we aimed at adapting PMCA to the detection of human disease-associated alpha-synuclein from brain samples. Methods: Brain homogenates from patients with Parkinson’s disease (PD) or Multiple System Atrophy (MSA) were serially tenfold diluted from 10-4 to 10-10 (w/v) and submitted to asynPMCA using healthy C57Bl/6 brain homogenate as a substrate. Newly formed a-syn aggregates were identified using a homemade specific ELISA and Western blot methods, as previously described (Nicot et al., 2019). Results: A-syn aggregates could be detected down to the 10-7 dilution of a PD brain sample and down to the 10-10 dilution of MSA brain samples without any false positive result on substrate or negative controls. Conclusions: MSA associated a-syn aggregates appeared to be more prone to misfold C57Bl/6 a-syn than PD ones during our asynPMCA. The proof-of-principle of ultrasensitive detection of disease-associated a-syn in human brain obtained in this study is very encouraging and promising for the possible detection of minute amounts of a-syn aggregates during early stages of the disease or in peripheral tissues.

P474 / #1399

Topic: Theme C: α-Synucleinopathies / C1.o. Disease Mechanisms, Pathophysiology: Metal ions

LABEL-FREE ANALYSIS OF NEUROMELANIN AND ASSOCIATED IRON DEPOSITS IN PARKINSON’S DISEASE BRAIN TISSUE BY SYNCHROTRON X-RAY SPECTROMICROSCOPY

Lecture Title:

J. Brooks1, J. Everett1,2, F. Lermyte1, V. Tjendana-Tjhin1, S. Banerjee3, P. O'Connor3, C. Morris4, P. Sadler3, N. Telling2, J. Collingwood1 1University of Warwick, School Of Engineering, Coventry, United Kingdom, 2Keele University, School Of Pharmacy And Bioengineering, Stoke-on-Trent, United Kingdom, 3University of Warwick, Department Of Chemistry, Coventry, United Kingdom, 4Newcastle University, Newcastle Brain Tissue Resource, Newcastle-upon-Tyne, United Kingdom

Aims: Neuromelanin-pigmented neurons, which are highly susceptible to neurodegeneration in the Parkinson’s disease substantia nigra, harbour elevated iron in the diseased state. Non-disruptive analysis herein is inherently challenging since conventional fixation and staining can significantly influence native tissue chemistry. State-of-the-art synchrotron x-ray spectromicroscopy was applied to label-free characterisation of organic and inorganic material in Parkinson’s disease substantia nigra, providing remarkable specificity and sensitivity at intracellular spatial resolution. Methods: Post-mortem fresh-frozen substantia nigra from three Parkinson’s disease cases were resin embedded, ultra-microtome-sectioned and analysed using scanning transmission x-ray microscopy (STXM) at the carbon and oxygen K-edges, and the iron L3-edge, using the I08 beamline, Diamond Light Source synchrotron (UK). Results: Using STXM, a unique feature in the neuromelanin absorption spectrum was discovered at 287.4 eV, facilitating direct, label-free visualisation of neuromelanin in situ1. STXM also revealed evidence for a range of oxidation states in neuromelanin-associated iron deposits in Parkinson’s disease substantia nigra. The excellent sensitivity, specificity and spatial resolution of these STXM measurements demonstrated variation in iron oxidation state on sub-micron length scales.

Conclusions: STXM offers a label-free approach to characterising distributions of both organic and inorganic components in post-mortem human brain tissue, with particular utility for non-destructive analysis of metals associated with neuromelanin. Reference: [1] Brooks 2020 Angewandte Chemie https://doi.org/10.1002/anie.202000239

P475 / #544

Topic: Theme C: α-Synucleinopathies / C1.o. Disease Mechanisms, Pathophysiology: Metal ions

WIDESPREAD COPPER DECREASES COMMON TO PARKINSON'S DISEASE DEMENTIA AND ALZHEIMER'S DISEASE BRAINS

Lecture Title:

M. Scholefield1, G. Cooper2,3, S. Church1, S. Patassini3, F. Roncaroli4, N. Hooper4, R. Unwin2,5 1University of Manchester, Division Of Cardiovascular Sciences, Manchester, United Kingdom, 2University of Manchester, Division of Cardiovascular Sciences, Division Of Cardiovascular Sciences, Manchester, United Kingdom, 3University of Auckland, School Of Biological Sciences, Auckland, New Zealand, 4University of Manchester, Division Of Neuroscience & Experimental Psychology, Manchester, United Kingdom, 5University of Manchester, Stoller Biomarker Discovery Centre & Division Of Cancer Sciences, Manchester, United Kingdom

Aims: Studies of metals in the brains of individuals with Parkinson’s disease (PD) have repeatedly reported metal dysregulations, including decreases in copper and increases in iron. However, most of these studies have focused on the heavily-affected substantia nigra of the PD brain, and few distinguish between PD cases with and without dementia. This study aimed to address both of these issues with a multi-regional analysis of essential metals in the Parkinson’s disease dementia (PDD) brain. Methods: Tissues from nine different regions were investigated to allow coverage of regions with varying levels of neuronal loss in PDD. Levels of nine essential metals were determined using inductively-coupled plasma mass spectrometry (ICP-MS) and compared between cases (n = 9) and controls (n = 9). Findings were then compared to a previous study (Xu et al., 2017) of AD which employed comparable methods to look for differences and similarities between the two diseases. Results: Widespread copper decreases were observed in PDD cases within seven of the nine regions investigated, including five which also showed decreases in the previous AD study (Xu et al., 2017). Decreased potassium and manganese were also observed across multiple regions, as were decreases in magnesium, zinc, and selenium in a single region each. Conclusions: These results indicate the presence of several perturbations in essential metals across multiple regions of the PDD brain, the most common being widespread decreases in copper, which are also present in AD. Such findings may indicate common metallic dysfunctions contributing to cognitive decline in both of these dementia diseases.

P476 / #1807

Topic: Theme C: α-Synucleinopathies / C1.p. Disease Mechanisms, Pathophysiology: Modeling of disease progression

LONGITUDINAL SYNTHETIC DATA GENERATOR FOR REPRODUCIBLE RESEARCH

Lecture Title:

R. Couronne1, A. Valladier1,2, S. Durrleman1,2 1ICM, Institut Du Cerveau, Paris, France, 2INRIA Paris, Aramis, Paris, France

Aims: We aim at providing a longitudinal synthetic data generator for neurodegeneration biomarkers. This generator can be first fitted on any set of progressing biomarker in a cohort, to then be able to generate synthetic data that mimic the real data, while preserving privacy. Although not directly usable for clinical research, this lightweight, easily sharable generator fosters reproducibility by providing plausible data on demand for educational, visualization or benchmarking purposes. Methods: We included 404 idiopathic Parkinson’s disease subjects from the PPMI dataset with six clinical scores, the MDS-UPDRS part I, II and III, MoCA, SCOPA-AUT and RBDSQ, for an average of 13.8+/-2.7 visits over 6.4 +/- 1.7 years. We then used a generative mixed-effect model [Schirrati et al 17] to fit the longitudinal progression of these clinical scores. We ensured privacy by using a bayesian gaussian mixture model to fit the empirical individual parameters distribution, so that no individual parameters from real data can be retrieved from the generator. Results: We fitted a model on PPMI and generated 404 synthetic patients. To assess the quality of synthesis, we trained two classifiers with default parameters to distinguish synthetic visits from real ones, leading to low test accuracy : 60% for Random Forest and 52% for Logistic Regression. We then showed that biomarker’s correlation structure is consistent in synthetic data. Conclusions: We proposed a longitudinal synthetic data generator for reproducible research. Synthetic data are difficult to distinguish from real data, and preserve the correlation structure.

P477 / #821


AGE AT ONSET AND PARKINSON’S DISEASE PROGRESSION

Lecture Title:

G. D'Urso1, K. Taylor1,2, T. Kustermann1, N. Pavese3,4, G. Pagano1,5 1F. Hoffmann - La Roche Ltd, Roche Pharma Research And Early Development, Neuroscience And Rare Diseases Discovery And Translational Area, Basel, Switzerland, 2University of Basel, Faculty Of Psychology, Basel, Switzerland, 3University of Newcastle, Clinical Ageing Research, Newcastle Upon Tyne, United Kingdom, 4Aarhus University Hospital, Department Of Clinical Medicine- Nuclear Medicine And Pet, Aarhus, Denmark, 5King's College, Institute Of Psychiatry, Psychology And Neuroscience (ioppn),, London, United Kingdom

Aims: Objective: Explore whether age at onset of Parkinson ’s disease influences early disease progression. Methods: We analysed data from the Parkinson’s Progression Markers Initiative Database, including 423 patients with a diagnosis of PD confirmed by DaTSCAN imaging. Over a 60-month period, we investigated whether older age at onset was associated with faster motor progression, development of motor complications, cognitive decline and loss of striatal DaTSCAN binding. We used multivariate linear regression models to predict changes in each of the following outcome variables: severity of motor, nonmotor symptoms, motor complications and nigrostriatal function. We included in the models the following covariates: age at onset, gender and years of education, H&Y stage, MDS-UPDRS, MoCA, REM Behaviour disorder, DaTSCAN binding in putamen contralateral to most affected clinical side, CSF levels of total alpha-synuclein, tau, p-tau and amyloid-beta1-42. Results: Older age at onset was associated with faster motor progression (change in MDS-UPDRS part III: p=0.011; change in MDS-UPDRS total: p=0.021), development of motor complications (change in MDS-UPDRS part IV: p=0.001), cognitive decline (change in MoCA: p<0.0001) and faster decline of nigrostriatal dysfunction (change in DaTSCAN putamen ipsilateral binding: p=0.002). Older age at onset was not associated with progression of motor or non-motor patient-reported activities of daily living (changes in MDS-UPDRS part II = p>0.10 and part I = p>0.10). Conclusions: Our findings confirm a direct contribution of aging to the progressive neurodegeneration of PD, which suggests that age at onset should be included as stratification factor in future clinical trials in patients with early PD.

P478 / #997


INTRA-NIGRAL INJECTIONS OF ALPHA-SYNUCLEIN PROTOFIBRILS AND INHIBITION OF GBA INDUCE PROGRESSIVE LOSS OF DOPAMINERGIC NEURONS: CHARACTERIZATION OF A NOVEL ANIMAL MODEL OF PARKINSON’S DISEASE.

Lecture Title:

A. Henriques1, B. Calvo-Flores Guzman1, C. Filippi2, P. Jego2, P. Poindron2, N. Callizot2 1Neuro-Sys Vivo, Pharmacology, GARDANNE, France, 2Neuro-Sys Vivo, Pharmacology, Gardanne, France

Aims: Patients with Parkinson’s disease are usually diagnosed at 60 years of age and present with resting tremor and bradykinesia, caused by the degeneration of the nigrostriatal dopaminergic pathways. Accumulation of misfolded alpha-synuclein, forming Lewy-bodies, causes mitochondrial stress and impairs autophagy-lysosomal pathway. Mutations on the gene coding for the lysosomal protein GBA is a strong risk factor for Parkinson’s disease. Animal models of Parkinson’s disease often rely on one insult despite the multifactorial pathophysiology of this disease. Here, we aimed to develop and characterize a novel and inducible in vivo model of Parkinson’s disease based on three of its key features: the toxicity of α-syn protofibrils, the impairment of lysosomal function and ageing. Methods: Mice at 18-month of age were bilaterally injected with a preparation of alpha-synuclein (50 µM) containing protofibrils (30% determined by automated protein analysis). Inhibition of GBA was achieved with conduritol-B-epoxide (50 mg/kg/2days; an inhibitor of GBA). Experiment was stopped 2-, 4-, 6- and 8-weeks post-injection for histological analysis. Results: Progressive loss of dopaminergic projections and neurons in the substantia nigra was observed after intra-nigral injections of alpha-synuclein. Neuronal loss was associated with a clear accumulation of alpha-synuclein in dopaminergic neurons and activation of IBA-1 positive microglial cells. Conclusions: Altogether, intra-nigral injections of alpha-synuclein protofibrils, combined with a chronic inhibition of GBA, led to histological phenotype highly similar to those observed in patients. This new animal model of Parkinson’s disease represents a valuable tool for studying the combined toxicity of alpha-synuclein protofibrils and GBA-linked lysosomal dysfunction.

P479 / #1221


MODELLING PARKINSON’S DISEASE PATHOLOGY IN THE RAT BRAIN USING VIRAL-MEDIATED HUMAN ALPHA-SYNUCLEIN OVEREXPRESSION COMBINED WITH FN075-MEDIATED ALPHA-SYNUCLEIN AGGREGATION.

Lecture Title:

R. Kelly1, V. Alamilla1, S. Jarrin1, T. Patton1, A. Cairns2, J. Ådén2, F. Almqvist2, A. Bemelmans3, E. Brouillet3, D. Mckernan1, E. Dowd1 1National University of Ireland, Galway, Pharmacology & Therapeutics, Galway, Ireland, 2Umeå University, Department Of Chemistry, Umeå, Sweden, 3Institut de Biologie François Jacob / Commissariat à l'énergie atomique (CEA), Molecular Imaging Research Center (mircen), Fontenay-aux-Roses, France

Aims: Parkinson’s disease models which overexpress human alpha-synuclein in the rat brain are considered some of the most valid currently available. However, they are limited by their slowly developing pathology and high degree of variability. We have recently developed a novel small molecule (FN075) that is capable of promoting alpha-synuclein oligomerisation and progressive fibril formation1 and inducing pathology after intra-nigral injection in mice2 and rats3. Therefore, we sought to determine if combining viral-mediated overexpression of alpha-synuclein with FN075-mediated aggregation of alpha-synuclein resulted in a more rapidly developing and consistent model of PD. Methods: 40 female Sprague-Dawley rats were given unilateral intra-nigral injections of AAV-α-synuclein or AAV-GFP. Four weeks later, the rats were given unilateral intra-nigral injections of FN075 or vehicle. Motor function was assessed using lateralised tests of spontaneous motor function. After sacrifice, immunohistochemical analyses were used to assess alpha-synuclein expression and aggregation as well as nigrostriatal degeneration. Results: At post mortem, expression of alpha-synuclein was seen throughout the ventral midbrain and the striatum on the ipsilesional side in both groups which received AAV-alpha-synuclein infusions. Degeneration of nigral neurons was seen in the combined AAV-alpha-synuclein & FN075 group. In the Corridor test of lateralised neglect, this group also exhibited mild motor impairment. Conclusions: AAV-alpha-synuclein combined with FN075 is a promising model for Parkinson’s disease which warrants further investigation. References: 1. Cegelski L et al., 2009. Nat Chem Biol. 5:913-9. 2. Chermenina M et al., 2015. NPJ Parkinson's disease; 1: 15024. 3. Olsen L et al., 2019. Brain Behav Immun; 80: 525-535

P480 / #1763


THE EFFECT OF CHRONIC CORTICOSTERONE ADMINISTRATION IN THE DEVELOPMENT OF A PARKINSONIAN PHENOTYPE IN BAC TRANSGENIC RATS OVEREXPRESSING HUMAN Α-SYNUCLEIN

Lecture Title:

M. Nakos-Bimpos1, D. Palermos1, G. Chrousos1,2, L. Stefanis1,3, A. Polissidis1 1Biomedical Research Foundation Academy of Athens, Center Of Clinical, Experimental Surgery And Translational Research, Athens, Greece, 2National and Kapodistrian University of Athens, 1st Department Of Pediatrics, Aghia Sophia Children's Hospital, Medical School, Athens, Greece, 3National and Kapodistrian University of Athens, 1st Department Of Neurology, Eginition Hospital, Medical School, Athens, Greece

Aims: Accumulating evidence supports a role of chronic stress and HPA axis activation in the pathogenesis of Parkinson’s disease (PD). We assessed the effects of chronic corticosterone in a prodromal PD rat model by examining non-motor and motor behaviors, brain-region specific monamine profiles and dopaminergic system integrity. Methods: 9 mo male BAC-transgenic Sprague Dawley rats overexpressing human alpha-synuclein (AS-BAC) and their wild type (WT) littermates were administered corticosterone (50 μg/ml in drinking water, 14 days). Subsequently, subjects underwent a behavioral test battery followed by brain dissection (striatum, hippocampus, hypothalamus and amygdale) for neurochemical analysis using HPLC or perfusion for tyrosine hydroxylase positive (TH+) stereological cell counts in the substantia nigra and VTA. Results: Corticosterone administration reduced weight gain and adrenal gland weights, increased thymus gland weights, and decreased olfactory discrimination and open arm time in the elevated plus maze in WT rats. These alterations were present in AS-BAC rats at baseline and not further affected by corticosterone. In the open field, previously reported novelty-induced hyperactivity in AS-BAC rats (Polissidis et al., in press) was reversed and postural instability was exacerbated following corticosterone. Noradrenaline levels were increased in the hypothalamus of corticosterone-administered WT and AS-BAC rats and reduced in the hippocampus of AS-BAC rats, irrespective of corticosterone treatment. Finally, corticosterone reversed enhanced dopamine turnover in the hypothalamus of AS-BAC rats. TH+ stereological measurements are currently underway. Conclusions: Chronic corticosterone recapitulates key non-motor behavioural features of PD and exacerbates motor symptoms in AS-BAC rats, indicating that chronic stress may contribute to and/or exacerbate PD pathology.

P481 / #463


AGE-DEPENDENT MOTOR AND NON-MOTOR PARKINSONIAN FEATURES IN A NOVEL NEUROMELANIN-PRODUCING TRANSGENIC MOUSE MODEL OF PARKINSON´S DISEASE AND BRAIN AGING

Lecture Title:

N. Peñuelas1, A. Laguna1, M. Gonzalez-Sepulveda1, L. Miquel-Rio2, N. Benseny-Cases3, H. Xicoy1, J. Compte1, M. Lorente-Picón1, E. Álvarez-Marimon4, J. Romero-Giménez1, A. Parent1, T. Cuadros1, B. Rodríguez-Galván1, F. Cacho-Nerin5, J. Cladera4, A. Bortolozzi2, I. Carballo-Carbajal1, M. Vila1,6,7 1Vall d’Hebron Research Institute (VHIR)–Center for Networked Biomedical Research on Neurodegenerative Diseases (CIBERNED), Neurodegenerative Diseases Research Group, Barcelona, Spain, 2IIBB–CSIC, August Pi i Sunyer Biomedical Research Institute (IDIBAPS)-Center for Networked Biomedical Research on Mental Health (CIBERSAM), Department Of Neurochemistry And Neuropharmacology, Barcelona, Spain, 3ALBA, Synchrotron Light Source, Cerdanyola del Vallès, Spain, 4Biophysics Unit and Centre of Studies in Biophysics, Universitat Autònoma de Barcelona, Department Of Biochemistry And Molecular Biology, Bellaterra, Spain, 5Harwell Science and Innovation Campus, Diamond Light Source, Didcot, United Kingdom, 6Autonomous University of Barcelona, Department Of Biochemistry And Molecular Biology, Cerdanyola del Vallès, Spain, 7Catalan Institution for Research and Advanced Studies (ICREA), Na, Barcelona, Spain

Aims: Parkinson's disease (PD) is characterized by a preferential degeneration of neurons that accumulate with age the pigment neuromelanin (NM), especially neurons from substantia nigra (SN) and locus coeruleus. We previously generated a rat model exhibiting progressive unilateral SN production of human-like NM that, when reaching a certain threshold of accumulation, triggered PD-like neuropathology and motor deficits (Carballo-Carbajal, Laguna et al. 2019 Nat Commun). However, pigmentation in the human brain is not limited to the SN but is also present, at varying degrees, in most catecholaminergic neuronal groups. Here we generated a new NM-producing rodent model that mimics the bilateral distribution of pigmentation within the whole human brain (i.e. cathecholaminergic groups A1-A14) in order to assess the potential effects of multisystemic NM accumulation, as it occurs in aging humans. Methods: We generated and characterized tissue-specific transgenic mice with constitutive expression of human melanin-producing enzyme tyrosinase (hTyr) under the tyrosine hydroxylase (TH) promoter (Tg-TH-hTyr). Results: Tg-TH-Tyr mice recapitulated the neuronal production, distribution and age-dependent accumulation of NM seen in the human brain. In parallel to NM intracellular buildup, Tg-TH-hTyr mice exhibited in an age-dependent manner major PD features, including both motor and non-motor behavioral alterations, changes in dopamine metabolism and degeneration of specific catecholaminergic neuronal groups. Transcriptomic analysis of NM-accumulating cells revealed alterations in PD-related biological pathways. Conclusions: Our results show that progressive NM accumulation leads to molecular and functional alterations within specific catecholaminergic neuronal groups resulting in PD-like motor and non-motor behavioral alterations relevant to both PD and brain aging.

P482 / #1461


PLASMA INFLAMMATORY AND NEURODEGENERATION MARKERS AND DISEASE PROGRESSION IN LEWY BODIES DISORDERS

Lecture Title:

A. Pilotto1, A. Imarisio2, E. Garraffa3, F. Conforti1, A. Scalvini1, S. Masciocchi2, S. Nocivelli1, R. Turrone1, S. Gipponi1, E. Cottina1, B. Borroni1, M. Rizzetti4, M. Pizzi5, A. Bellucci6, L. Bonanni7, H. Zetterberg8, N. Ashton9, A. Hye10, A. Padovani1 1university of Brescia, Department Of Clinical And Experimental Sciences, Brescia, Italy, 2University of Brescia, Department Of Clinical And Experimental Sciences, Brescia, Italy, 3University of bresica, Department Of Biochemical Chemistry, Department Of Molecular And Translational Medicine, brescia, Italy, 4FERB ONLUS, Department Of Parkinson's Disease Rehabilitation, Trescore Balneario, Italy, 5University of Brescia, Department Of Molecular And Translational Medicine, Brescia, Italy, 6University of Brescia, Molecular And Translational Medicine, Brescia, Italy, 7University G. d'Annunzio of Chieti-Pescara, Neuroscience, Imaging And Clinical Sciences, Chieti, Italy, 8Sahlgrenska University Hospital, Clinical Neurochemistry Laboratory, Mölndal, Sweden, 9The Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Institute Of Neuroscience & Physiology, Mölndal, Sweden, 10king' college london, Institute Of Neurology, london, Italy

Aims: to evaluate the different distribution of peripheral markers in PD and DLB and their relationship with motor and non-motor symptoms, neurodegenerative markers and clinical progression over time. Methods: Plasma IL6, CRP, cystatin C and NfL were assessed in a longitudinal study including 68 non demented PD and 24 DLB patients who underwent an extensive motor and cognitive assessment at baseline and after 2 years of follow-up. The association with disease severity was evaluated in a multilinear model adjusted for the effect of age, sex and disease duration. Results: DLB patients exhibited higher levels of plasma IL6 (p = 0.050) and cystatin C (p = 0.022) compared to PD. In PD, IL6 correlated with disease duration (r = 0.315, p = 0.009), whereas Cystatin C correlated with age and dysautonomic impairment (p = 0.017). Plasma NfL correlated with cystatin C, but not with CRP and IL6 levels. In DLB, no associations between peripheral markers and cognitive/behavioral abnormalities and progression were found. In PD, a multivariate model showed that patients with higher Cystatin levels exhibited faster motor and cognitive progression at two years of follow-up (p = 0.030 and p = 0.046, respectively). Conclusions: Our study found evidences of a pro-inflammatory immune system activation in DLB compared to PD patients. In PD, Cystatin C was associated with major non-motor symptoms, higher NfL levels and a remarkably faster motor and cognitive progression. Further studies are needed in order to understand the mechanism underlying the role of Cystatin C as potential modulator of disease progression in PD.

P483 / #1482


ORTHOSTATIC HYPOTENSION IS ASSOCIATED WITH CEREBRAL ATROPHY IN LEWY BODY DISORDERS

Lecture Title:

A. Pilotto1, A. Romagnolo2, A. Scalvini1, M. Masellis3, Y. Shimo4, L. Bonanni5, R. Camicioli6, L. Wang7, D. Alok8, K. Longardner9, F. Rodriguez-Porcel10, M. Di Francesco7, J. Viczarra11, E. Montanaro2, S. Maule2, A. Lupini1, C. Oyeda3, S. Delli Pizzi5, M. Gee6, B. Borroni1, M. Rizzetti12, N. Hattori4, L. Lopiano2, I. Litvan13, A. Espay7, A. Padovani1, A. Merola14 1university of Brescia, Department Of Clinical And Experimental Sciences, Brescia, Italy, 2University of Turin, Neuroogy, Turin, Italy, 3university of Toronto, Sunnybrook Research Institute, toronto, Canada, 4Juntendo University, Department Of Research And Therapeutics For Movement Disorders, tokyo, Japan, 5University G. d'Annunzio of Chieti-Pescara, Neuroscience, Imaging And Clinical Sciences, Chieti, Italy, 6University of Alberta, Medicine, Alberta, Canada, 7university of Cincinnati, Neurology, cincinnati, United States of America, 8Texas Tech University Health Sciences Center, Department Of Molecular And Translational Medicine, El paso, United States of America, 9San Diego helath system, Department Of Neuroscience, san diego, United States of America, 10Medical univeristy of South Carolina, Department Of Neurology, Charleston, United States of America, 11Emory University, Department Of Neurology, Atlanta, United States of America, 12FERB ONLUS, Department Of Parkinson's Disease Rehabilitation, Trescore Balneario, Italy, 13universty of california, Department Of Neurology, san francisco, United States of America, 14Ohio state university, Neurology, columbus, United States of America

Aims: To evaluate whether orthostatic hypotension (OH) or supine hypertension (SH) is associated with global and regional brain atrophy and white matter hyperintensities (WMH) in patients with Parkinson’s disease (PD) and dementia with Lewy bodies (DLB). Methods: Supine and orthostatic blood pressure (BP) and structural magnetic resonance imaging (MRI) data were extracted from a multicenter repository of PD and DLB patients evaluated at eight tertiary-referral centers in the USA, Canada, Italy, and Japan. OH was defined as a systolic/diastolic BP fall ³ 20/10 mm/Hg within 3 minutes of standing (severe, ³30/15 mm/Hg) and SH as a BP ≥140/90 mmHg with normal sitting hypertension. Diagnosis-, age-, sex-, and disease duration-adjusted differences in global and regional cerebral atrophy, as well as WMH were appraised using validated semi-quantitative rating scales. Results: A total of 384 patients (310 with PD, 74 with DLB) were included in the analyses, of whom 44.3% (n=170) had OH, 10.9% (n=42) severe OH, and 18.5% (n= 71) SH. OH was associated with global cerebral atrophy (p=0.016) and regional atrophy involving the orbito-frontal (p=0.012), anterior-temporal (p=0.001), and medio-temporal (p=0.006) regions. Severe OH was associated with greater anterior-temporal and medial temporal (p=0.001) atrophy than non-severe OH. The WMH burden was similar in those with and without OH (p>0.05). SH was not associated with brain atrophy or WMH (both, p>0.05). Conclusions: OH, but not SH, is associated with global and regional cerebral atrophy in Lewy body disorders. Neither OH nor SH were associated with WMH.

P484 / #284

Topic: Theme C: α-Synucleinopathies / C1.q. Disease Mechanisms, Pathophysiology: Other

CHARACTERIZATION OF DEMENTIA WITH LEWY BODIES (DLB) AND MILD COGNITIVE IMPAIRMENT USING THE DLB MODULE (DLB-MOD)

Lecture Title:

J. Galvin1, S. Chrisphonte1, L.-C. Chang2, M. Tolea1 1University of Miami Miller School of Medicine, Neurology, Miami, United States of America, 2Florida Atlantic University, Mathematical Sciences, Boca Raton, United States of America

Aims: The NIA Alzheimer Disease Center program created the Dementia with Lewy bodies (DLB) module (DLB-MOD) to facilitate characterization of DLB and related disorders and distinguish DLB from Alzheimer’s disease (AD). We tested the performance of the DLB-MOD. Methods: The DLB-MOD was studied in 342 patients: 53 controls, 78 AD, and 110 DLB, 79 mild cognitive impairment due to AD (MCI-AD) and 22 MCI-DLB. Results: DLB differed from AD in extrapyramidal symptoms, hallucinations, activities in daily living, apathy, autonomic features, REM sleep behaviors, daytime sleepiness, cognitive fluctuations, timed attention tasks, episodic memory, and visual perception. MCI-DLB differed from MCI-AD in extrapyramidal features, mood, autonomic features, fluctuations, timed attention tasks, and visual perception. Descriptive data on DLB-MOD measures are provided for reference. Conclusions: The DLB-MOD provided excellent characterization of core, supportive, and suggestive features to differentiate DLB from AD and cognitively normal controls while also providing important characterizing features of MCI-DLB.

P485 / #486


A COMPREHENSIVE ANALYSIS OF DEMENTIA CEREBROSPINAL FLUID BIOMARKERS USING GWAS, POLYGENIC RISK SCORES AND MENDELIAN RANDOMIZATION IN PARKINSON’S DISEASE

Lecture Title:

L. Ibanez1, J. Bahena1, U. Dube1, F. Farias2, C. Yang3, O. Harari3, C. Cruchaga2, B. Benitez3 1Washington University School of Medicine, Psychiatry, Saint Louis, United States of America, 2Washington University School of Medicine, Psychiatry, St Louis, United States of America, 3Washington University in St. Louis, Psychiatry, St. Louis, United States of America

Aims: We aimed to investigate the relationship between dementia biomarkers and PD using , GWAS, polygenic risk scores and Mendelian Randomization approaches. Methods: We used publicly available summary statistics from our genome-wide association study on amyloid beta, tau and p-tau and an in-house dataset for alpha synuclein to select the instrumental variables and the summary statistics from the largest PD dataset todate. Instrumental variables for each biomarker were limited to those variants with p<10-05 and uncorrelated. Additionally, we evaluated the genetic architecture of dementia biomarkers in PD using an in-house dataset (N=700). Results: GWAs analyses reveal the APOE locus is associated with CSF Ab levels in PD cohorts. PRS analysis showed that CSF Ab genetic architecture was correlated to that of PD risk maximizing when collapsing independent SNPs with p value < 0.01 (p=2.5×10-11) with an R2 of 2.29%. In addition, Mendelian Randomization methods that account for pleiotropy and outliers, found that CSF Ab have a causal role in PD (p=1.44×10-05) and effect mediated by a SNP near the APOE gene (rs769449). A trend toward a causal association was found for a-Syn and PD (p=0.06). However, we found not GWAS hit for CSF a-Syn, tau or p-Tau in PD cohorts. Conclusions: Our results suggest that Ab is involved in the causal pathway of PD, even though APOE does not seem to be related to CSF Ab levels in PD cohorts. a-Syn also seems to have a causal relation with PD, but studies with a larger number of samples are needed.

P486 / #1422


THE RELATIONSHIP BETWEEN BETA-GLUCOCEREBROSIDASE ACTIVITY LEVELS AND DEMENTIA SEVERITY IN INDIVIDUALS WITH PARKINSON’S DISEASE WITHOUT GBA1 MUTATION

Lecture Title:

C. Silveira1, R. Hegele2, J. Baker1, R. Tirona3, G. Zou4, R. Bartha5, P. Macdonald6, M. Jenkins6, M. Jog6, E. Finger6, S. Morrow6, J. Wells7, M. Borrie7, K. Coleman1, S. Pasternak6 1Parkwood Institute, Neurology, London, Canada, 2Western University, Department Of Medicine, London, Canada, 3Western University, Department Of Physiology And Pharmacology, London, Canada, 4University of Western Ontario, Epidemiology And Biostatistics, London, Canada, 5Western University, Department Of Medical Biophysics, London, Canada, 6Western University, Department Of Clinical Neurological Sciences, London, Canada, 7Parkwood Insitute, Geriatrics, London, Canada

Aims: This study examined whether β-Glucocerebrosidase (GCase) activity level is a predictor of dementia severity in individuals with Parkinson’s disease (PD) without GBA1 mutation. Decreased activity level of GCase has been demonstrated in individuals with PD carriers and non-carriers of GBA1 mutation, and linked to α-synuclein accumulation. It is currently known that PD patients carrying a GBA1 mutation show greater prevalence of and more severe cognitive decline. Yet, it remains unclear whether a link between GCase activity level and the severity of cognitive decline exists in PD non-carriers. Methods: Baseline data from 31 individuals with PD dementia enrolled in a clinical trial were used for this study. Demographic and clinical information including age, age at PD onset, disease duration, disease severity, sex, years of education, and general cognitive status were collected. The severity of dementia was assessed through the Clinical Dementia Rating Scale (CDR) and GCase activity levels were measured in blood lymphocytes. Results: Hierarchical Multiple Regression analysis showed that global CDR scores were predicted by GCase activity levels even after controlling by the severity of motor symptoms (R2 change= 0.17; p=0.032; GCase Stand. b= -0.44, p=0.032; MDS UPDRS-III Stand. b= 0.051, p=0.79). Conclusions: This result supports the role of GCase activity levels in the severity of cognitive symptoms in PD even for non-carriers of GBA1 mutation. Findings from this research have important implications to current clinical trials testing medications targeting GCase in PD.

P487 / #1199


DOES HEAVY ALCOHOL CONSUMPTION PREVENT FROM AGE-RELATED NEURODEGENERATION?

Lecture Title:

F. Tamsen, I. Alafuzoff Akademiska sjukhuset, Klinisk Patologi, Uppsala, Sweden

Aims: To assess whether liver pathology (LP) seen in association with heavy alcohol consumption (HAC) correlates with the type and/or extent of neurodegenerative alterations (NA) in the brain. Methods: 94 deceased with a standardized neuropathological investigation parallel with assessment of LP, i.e. fibrosis, inflammation and fatty changes, were identified from hospital records. A score of LP was created based on the sum of the alterations listed ranging from 0 to 9. Mean age ± standard error of means (SE) of the cohort was 73 ± 0.8 years at death, 27 females and 68 males. 50 subjects with HAC, 15 had during life shown neurological impairment (NI) and 29 were neurologically unaffected. The standardised neuropathological investigation included assessment of NA such as hyperphosphorylated tau (HPtau), β-amyloid (Aβ), α-synuclein (αS) and Tar DNA binding protein 43 (TDP43). Results: The mean ± SE for LP was 3.7 ± 0.2. The score was highest 4.1 ± 0.3 in the HAC group and lowest 2.6 ± 0.6 in the NI group. A significant positive correlation was observed between the extent of HPtau/Aβ, HPtau/TDP43, HPtau/αS and Aβ/TDP43. The correlation between the extent of LP and NA was always negative and reached significance (r = -0.25, p = 0.01) between LP and the extent of αS. Conclusions: It has been suggested that alcohol consumption might be preventive regarding NA. When assessing alterations in the end organs, liver and brain, we noted a negative correlation between LP and NA, and significantly so between LP and αS pathology.

P488 / #687


PLASMA TOTAL ALPHA-SYNUCLEIN AND NEUROFILAMENT LIGHT CHAIN: CLINICAL VALIDATION FOR DISCRIMINATING PARKINSON’S DISEASE FROM NORMAL CONTROL

Lecture Title:

W.-C. Lin1, C.S.Y. Yang2, C.-H. Lu3, P.-Y. Chiu4 1Kaohsiung Chang Gung Memorial Hospital, Department Of Diagnostic Radiology, Kaohsiung, Taiwan, 2MagQu Co., Ltd., President Office, New Taipei City, Taiwan, 3Kaohsiung Chang Gung Memorial Hospital, Department Of Neurology, Kaohsiung, Taiwan, 4Chang Bin Show Chwan Memorial Hospital, Department Of Neurology, Chunghwa, Taiwan

Aims: A previously published paper (referred to as the original cohort) showed that using a cut-off value of 116.1 fg/ml for the plasma total α-synuclein concentrations could discriminate Parkinson’s disease (PD) patients from normal controls (NCs). In this study, another independent cohort (referred to as the validation cohort) was recruited to validate the agreement between the clinical diagnosis and the use of plasma total α-synuclein to identify PD patients. In addition to total α-synuclein, plasma neurofilament light chain (NfL) in the validation cohort was detected. Methods: Seventy PD patients and thirty-three NCs were enrolled in the validation cohort. A clinical diagnosis and the immunomagnetic reduction (IMR) assay for plasma total a-synuclein were performed for each participant. Thirty-three of seventy PD patients and 23 of 33 NCs were subjected to the plasma NfL assay via IMR. Results: The positive, negative and overall percentages of agreement between the clinical diagnosis and plasma-total-a-synuclein diagnosis determined based on 116.1 fg/ml as the cut-off value were found to be 0.943, 0.818 and 0.903, respectively. The PD patients and NCs showed plasma NfL levels of 8.38±4.19 pg/ml and 17.6±7.95 pg/ml (p < 0.001), respectively. The cut-off value of the plasma NfL level used to differentiate PD patients from NCs was 12.8 pg/ml, with sensitivity and specificity values of 0.788 and 0.870, respectively. Conclusions: The results demonstrate the usefulness of the plasma total α-synuclein concentration to discriminate PD patients from NCs and reveal the elevation of the plasma NfL level in PD patients.

P489 / #1656

Topic: Theme C: α-Synucleinopathies / C2.a. Therapeutic Targets, Mechanisms for Treatment: Αlpha-synuclein

DISEASE-MODIFYING APPROACH THROUGH ENHANCEMENT OF LYSOSOMAL CATHEPSIN PROTEASE REDUCES PROTEIN ACCUMULATION EVENTS AND ASSOCIATED SYNAPTOPATHOLOGY OF ALZHEIMER’S AND PARKINSON’S DISEASES

Lecture Title:

M. Almeida1, J. Hwang1, D. Butler2, M. Pait1, K. Rentschler1, J. Locklear1, K. Farizatto1, B. Bahr1 1University of North Carolina– Pembroke, William C. Friday Laboratory, Biotechnology Research And Training Center, Pembroke, United States of America, 2Northeastern University, Center For Drug Discovery, Boston, United States of America

Aims: Disruption of lysosomal protein clearance functions has been linked to neurodegenerative disorders (Nixon 2017–FASEB J 31:2729; Lowry & Klegeris 2018–Brain Res Bulletin 139:144; Wang et al. 2018–Curr Opin Neurobiol 48:52). Amyloidogenic events and the spreading of toxic α-synuclein are thought to be influenced by lysosomal activity in neurons (Hwang et al. 2019–International J Mol Sci 20:4432), astrocytes (Tsunemi et al. 2020–J Neurosci 40:8618), and microglia (Lowry & Klegeris 2018; Bartels et al. 2020–Science 370:66). Here, our objective was to investigate positive lysosomal modulation for affecting protein accumulation events of Alzheimer’s disease (AD), pre-AD, and Parkinson’s disease (PD). Methods: utilized included immunocytochemistry in brains from wild-type mice, APP/PS1 AD mice, mild cognitive impairment (MCI) models, and A53T PD transgenic mice. Results: included Aβ42 detoxification induced by Z-Phe-Ala-diazomethylketone (PADK) and other small molecules that enhance the active form of lysosomal cathepsin B (CatB). The CatB induction also increased clearance of APP-βCTF, improved integrity of pyramidal neurons and interneurons, and corresponding synaptic and behavioral protection in the APP/PS1 and MCI animals. In the A53T PD mice, CatB enhancement led to clearance of human α-synuclein (>60% reduction), and within-animal analyses found a correlation between α-synuclein levels and synaptic marker recovery (p<0.01). Conclusions: Conclusion: Multiple cell types play a part in protein accumulation pathogenesis, and positive cathepsin modulation represents a disease-modifying therapy to slow the MCI to dementia continuum. Therapeutic focus needs to be directed at a safe route to enhance protein clearance in both glial and neuronal cells.

P490 / #1237


THE SECRETOME FROM INDUCED MESENCHYMAL STROMAL CELLS REDUCES ALPHA-SYNUCLEIN OLIGOMERS AND RESCUES ASSOCIATED CYTOTOXICITY

Lecture Title:

J. Burgess1,2,3, D. Amerna2, A. Faroqi1,2, T. Parsons2,4, R. Al-Kharboosh3, M. Delenclos2, A. Quiñones-Hinojosa5, T. Kanekiyo2,4, P. Mclean1,2 1Mayo Clinic Graduate School of Biomedical Sciences, Neuroscience Phd Program, Jacksonville, United States of America, 2Mayo Clinic, Neuroscience, Jacksonville, United States of America, 3Mayo Clinic, Regenerative Sciences Training Program, Jacksonville, United States of America, 4Mayo Clinic, Center For Regenerative Medicine, Jacksonville, United States of America, 5Mayo Clinic, Neurologic Surgery, Jacksonville, United States of America

Aims: Mesenchymal stromal cells (MSCs) have demonstrated promising regenerative effects in a number of disease states, including models of Parkinson’s disease (PD). These effects are believed to be mediated through paracrine secretions. Here we test the hypothesis that iPSC-derived MSCs (iMSCs) cultured in a synucleinopathy-mimicking environment will secrete factors that specifically target asyn-associated pathology and dysfunction. Additionally we will establish tools to enable identification of secreted factors. Methods: A FRET based αsyn-biosensor HEK cell line was treated for 24 hours with αsyn PFFs or human brain homogenate to simulate a PD environment. Subsequently, cells were co-cultured with iMSCs generated from iPSCs in transwell inserts for 48 hours. Synuclein aggregation was quantified using flow cytometry.

Experiments to genetically modify iPSCs with mutant MetRS to facilitate isolation and identification of iMSC originating proteins from co-cultures are ongoing. Results: 24 hours treatment with brain homogenate or PFFs followed by 48 hours incubation induces αsyn oligomers in ~3% and ~30% of biosensor cells, respectively. Co-culturing with iMSCs during the 48 hour incubation mediates a dose-dependent decrease in oligomers, peaking at ~20% decrease in optimal

conditions. These results are accompanied by an iMSC dose-dependent rescue of simultaneously measured cytotoxicity, according to intensity of fixable viability stain.

These findings mirror those achieved in similar experiments using adipose-derived MSCs, further supporting iMSCs as a readily renewable and modifiable potential alternative. Conclusions: This study supports the use of autologous iMSC therapy in ameliorating αsyn oligomers and provides a potential toolkit to enable identification of secreted factors specifically targeting αsyn-associated dysfunction.

P491 / #1464


ZINC FINGER PROTEIN TRANSCRIPTION FACTOR-MEDIATED REPRESSION OF ALPHA-SYNUCLEIN EXPRESSION AS A THERAPEUTIC APPROACH FOR PARKINSON’S DISEASE

Lecture Title:

A. Hatami, H.-O. Nguyen, Q. Yu, K. Marlen, D. Paschon, E. Mutter-Rottmayer, K. Van, Y. Lu, E. Tait, S. Lam, S. Hinkley, A. Goodwin, R. Surosky, E. Rebar, K. Meyer, B. Zeitler, A. Pooler Sangamo Therapeutics, Discovery And Translational Research, Richmond, United States of America

Aims: Molecular and genetic evidence implicate alpha-synuclein as a key mediator of PD pathogenesis. Reducing alpha-synuclein expression in neurons thus has the potential to halt or slow PD progression. Zinc finger proteins (ZFPs) are DNA-binding proteins that can be engineered to specifically bind any genomic sequence. When fused to transcriptional regulatory domains to form ZFP transcription factors (ZFP-TFs), they can modulate the expression level of the targeted gene. We engineered ZFP-TFs with human SNCA repression activity for use as potential therapeutics for PD. Methods: ZFP-TFs were identified in a screen of human neuroepithelial cells. On-target activity was assessed in human iPSC-derived neurons transduced with AAVs expressing the ZFP-TFs (AAV-ZFP-TFs). Transcriptome-wide specificity analyses using Affymetrix microarrays were performed in human iPSC-derived and mouse primary neurons. ZFP-TFs with minimal off-target activity in both human and mouse neurons were administered to PAC synuclein mice, which express the full human SNCA sequence on a mouse SNCA-null background. Animals were euthanized after 3 weeks and their brains were collected for molecular analyses. Results: The initial screen identified ZFP-TFs with human alpha-synuclein repression activity ranging from ~40% to >99%. Experiments in human iPSC-derived neurons confirmed the on-target activity of the constructs and the durability of the response for 30 days. Alpha-synuclein AAV-ZFP-TFs with a range of repression activity and minimal off-target activity were well tolerated in vivo and led to significant downregulation of aSyn mRNA expression in the mouse brain. Conclusions: The data presented here support continued development of ZFP-TFs as a potential therapy for PD.

P492 / #831


ENCAPSULATED CELL-BASED BIODELIVERY OF PROGRANULIN IMPROVES BEHAVIOUR IN A RAT MODEL OF PARKINSON’S DISEASE

Lecture Title:

J. Lundkvist1, Y. He2, M. Rossini3, J. Sandin1, H. Biverstål4, T. Pollare1, X. Zhang2, P. Svenningsson2, G. Paolone3, L. Wahlberg5, D. Emerich5, R. Blomquist6 1Sinfonia Biotherapeutics AB, R&d, Huddinge, Sweden, 2Karolinska Institute, Department Of Clinical Neuroscience, Solna, Sweden, 3Universita degli Studi di Verona, Biotecnologie, Verona, Italy, 4Karolinska Institute, Nvs, Huddinge, Sweden, 5Gloriana Therapeutics Inc, R&d, Warren, United States of America, 6Sinfonia Biotherapeutics, Business And Finance, Boulder, United States of America

Aims: The secreted factor progranulin exhibits a broad repertoire of signaling events essential for neuronal health and has been genetically associated with frontotemporal dementia (FTD), Alzheimer’s disease (AD) and Parkinson’s disease (PD). To increase brain progranulin levels has therefore emerged as a prioritized therapeutic strategy to combat several neurodegenerative disorders. The aim of this project was to develop encapsulated cell-based biodelivery (ECB) of progranulin to the brain as a novel therapy for multiple neurodegenerative disorders. Methods: ECB is a removable and replaceable clinical stage medical device, which contains human cells that secrete therapeutic factors at a therapeutic dose at the site needed within the diseased brain. ECB-progranulin devices were generated and progranulin diffusion, device functionality, treatment impact on brain tissue, inflammation, body weight and general exploratory behaviour, were assessed upon treatment of the striatum of naive rats. Furthermore, the therapeutic activity of ECB-progranulin was evaluated in the rat AAV-asynuclein model of Parkinson disease. Results: Functional ECB-progranulin treatment up to 6 months was monitored with no overt tissue reactions, inflammatory reactions or effect on body weight or general exploratory behaviour, detected. Moreover, 12 weeks of striatal ECB-progranulin therapy improved motor function in the rat AAV- asynuclein rat model, an effect which was accompanied by an increase in dopamine receptor type 1 expression levels. Furthermore, a broad progranulin tissue distribution was monitored in minipigs treated with ECB-progranulin. Conclusions: Combined these data validate the use of ECB-progranulin as an attractive and efficient brain progranulin therapy and supports its further development for Parkinson´s disease and other neurodegenerative disorders.

P493 / #1835


ALPHA SYNUCLEIN OLIGOMERIC POLYMORPHISMS: THERAPEUTIC TARGETS IN SYNUCLEINOPATHIES

Lecture Title:

K. Moore University of Texas Medical Branch, Neurology, Galveston, United States of America

Aims: Neurodegenerative diseases such as Parkinson’s disease (PD) and Dementia with Lewy Bodies (DLB) can be characterized as synucleinopathies due to their pathological hallmark, the accumulation of alpha synuclein into toxic proteinaceous aggregates. Aggregates of alpha synuclein have been long studied, however an intermediate state of aggregates, oligomers, are now considered to be the most toxic species. Our lab recently showed alpha synuclein oligomers exhibit polymorphism, distict seeding potency, and differ in toxicity. We have developed novel synuclein oligomer-specific monoclonal antibodies (SOMAs) to evaluate antibody biologics as therapeutic agents targeting polymorphic alpha synuclein oligomers. Methods: By utilizing the SOMAs in a combination of methods, both in vtiro and ex vivo, I can characterize the interaction between SOMAs and alpha synuclein oligomers. Immunostaining, immunoblotting, and cell based assays are utilized to evaluate SOMA specificity, immunoreactivity, and ability to prevent alpha synuclein oligomer-mediated toxicity. Results: SOMAs demonstrate specificity and immunoreactivity for distinct ɑ-Syn oligomers both in vitro and ex vivo. SOMAs detect disease-associated oligomers in both PD and DLB brain tissue. In a synucleinopathy mouse model, SOMAs detect oligomers in an age-dependent manner. Together with determining SOMAs ability to prevent toxicity in cellular conditions, establishes parameters for future in vivo studies. Conclusions: Many studies have investigated the multiple mechanisms explaining alpha synuclein oligomer-mediated toxicity; however, it is still unclear the role alpha synuclein oligomers play in synucleinopathy disease progression. Utilizing our SOMAs in a combination of methods, both in vitro and ex vivo, will evaluate the effects of utilizing antibody biologics to therapeutically targeti alpha synuclein oligomeric polymorphisms.

P494 / #1068


GENERATION OF MACROPHAGES AND MICROGLIA FROM SNCA A53T HOMOZYGOUS IPSC FOR DISEASE MODELLING APPLICATIONS

Lecture Title:

D. Rajesh1, C. Munn2, S. Burton2, S. Dickerson3 1FujiFilm Cellular Dynamics International, R&d Life Science, Madison, United States of America, 2Fujifilm Cellular Dynamics Inc., R&d Life Science, Madison, United States of America, 3Fujifilm Cellular Dynamics International, R&d Life Science, Madison, United States of America

Aims: Objective: Alpha-synuclein (SNCA) aggregation is a hallmark of Parkinson’s disease (PD). A substitution of alanine to threonine at position 53 (A53T) of SNCA results in an earlier onset PD. The A53T mutation promotes mitochondrial dysfunction, impaired phagocytosis and expression of pro-inflammatory cytokines in non-neuronal cell types. The present study validates the role of A53T on hematopoietic precursor cells (HPCs), macrophages and microglia derived from isogenic human induced pluripotent stem cells (iPSCs) for genotype-specific programs for PD therapy. Methods: Episomally reprogrammed iPSCs were subjected to nuclease-mediated homologous recombination to generate SNCA A53T homozygous iPSC line (SNCA A53T HO). Parental and SNCA A53T HO and isogenic control iPSCs were first differentiated to HPCs and further differentiated to macrophages and microglia under defined conditions. Results: HPCs were characterized by the expression of CD34, CD43, CD45, CD41, CD235 and functional assays to mark pluripotency. Macrophages were characterized by expression of CD68, CD64, CD80 CD33, CD11b, CD11c, CD1a, CD169, CD206, CD340, CD45, CD163, HLA-DR and CD86. Microglia were characterized by the presence of CD45, CD11b, CD33, P2RY12, TREM-2, CX3CR1 and IBA expression. SNCA A53T HO iPSC derived HPCs revealed a lower multilineage potential and efficiency for generating Macrophages and Microglia. SNCA A53T HO microglia exhibited impaired phagocytosis while SNCA A53T HO macrophages revealed an altered kinetics of phagocytic function. SNCA A53T HO macrophages and microglia revealed polarization to a pro-inflammatory subtype upon stimulation. Conclusions: Cryopreserved HPCs, macrophages and microglia with neural cell types can be used to mimic the onset of PD in a dish.

P495 / #1404


AMBROXOL AS A DISEASE-MODIFYING TREATMENT FOR PARKINSON’S DISEASE DEMENTIA: A PHASE 2, RANDOMIZED, DOUBLE-BLIND, PLACEBO-CONTROLLED TRIAL.

Lecture Title:

S. Pasternak1, C. Silveira2, K. Colman2, E. Finger1, M. Mayich3, M. Sharma3, S. Pandey3, R. Bartha4, S. Morrow5, J. Wells6, M. Borrie6, R. Hegele7, T. Rupar8, G. Zou9, P. Macdonald5, M. Jenkins5, M. Jog5 1University of Western Ontario, Clinical Neurological Sciences, London, Canada, 2Parkwood Institute, Neurology, London, Canada, 3London Health Science Center, Neuroradiology, London, Canada, 4Robarts Research Institute, Medical Physics, London, Canada, 5London Health Science Centre, Neurology, London, Canada, 6Parkwood Insitute, Geriatrics, London, Canada, 7Robarts Research Institute, Genetics, London, Canada, 8London Health Science Centre, Medical Biochemistry, London, Canada, 9University of Western Ontario, Epidemiology And Biostatistics, London, Canada

Aims: Currently, there are no disease-modifying treatments for Synucleinopathies such as Parkinson’s disease dementia (PDD).. A potential target for treatment target is the enzyme α-Glucocerebrosidase (GCase). Raising GCase levels lowers the levels of α-synuclein in cell and animal models and improve Parkinsonian symptoms. Ambroxol is an over the counter expectorant that is a pharmacological chaperone for GCase; a small molecule which stabilizes and increases the levels of GCase. This study aims to assess the safety and tolerability of Ambroxol in individuals with PDD, and to examine the effects of Ambroxol on cognitive, biochemical, and neuroimaging measures. Methods: Fifty individuals with mild to moderate PDD are being randomly assigned to Ambroxol 1050mg/day or placebo for 1 year. Primary outcome measures are the Alzheimer’s Disease Assessment Scale-cognitive subscale and the ADCS Clinician’s Global Impression of Change. Secondary outcomes include the UPDRS, an extended set of cognitive and clinical assessments, and plasma/CSF and neuroimaging biomarkers. Pharmacokinetics and pharmacodynamics data for use in future trials will also be acquired. Results: To date, 37 participants have been randomized, 33 titrated to full dose (Ambroxol/placebo), 29 passed 6 months, and 23 completed a year. Ambroxol appears to be well tolerated There have been no SAE's related to Ambroxol.. Blood levels of 10-12 micromolar are being achieved. Blinded pharmacological analyses shows GCase levels are raise by 1.6 fold in white blood cells. Penetration of Ambroxol in CSF was also demonstrated. Conclusions: If found effective, Ambroxol will be the first disease-modifying treatment for PDD. Clinical Trial Registration NCT02914366. Funded by the Weston Brain Institute.

P496 / #1130


ENZYMATIC REPLACEMENT THERAPY OF NANOCONJUGATED GBA AS A PROMISING THERAPEUTIC TREATMENT FOR PARKINSON’S DISEASE

Lecture Title:

E. Pradas1, D. Montpeyó1, A. Navarro-Romero2, P. Sarlé1, M. Montpeyó2, I. Fernandez-Gonzalez2, D. Ruiz-Molina3, F. Novio3, J. Lorenzo1, M. Martinez-Vicente2 1Institut de Biotecnologia i de Biomedicina (IBB), Enzymology Group, Bellaterra (Barcelona), Spain, 2Vall d'Hebron Institut de Recerca (VHIR), Neurodegenerative Diseases, Barcelona, Spain, 3Catalan Institute of Nanoscience and Nanotechnology (ICN2), CSIC and BIST, Nanostructured Functional Materials Group, Bellaterra (Barcelona), Spain

Aims: The objective of this study is the restoration of lysosomal glucocerebrosidase (GCase) activity in neurons through Enzymatic Replacement Therapy (ERT) using enzyme-polymer nanoconjugation of GBA recombinant enzyme. Methods: Recombinant GBA was conjugated to polypeptides polymers through a reduction-sensitive disulfide bond-bearing linker to improve protein stabilization and plasma protease resistance. Nanoconjugates were validated in vitro in a new cellular model of differentiated dopaminergic-like neurons expressing GBA knock out (GBA-KO) developed by our team. Results: GBA nanoconjugate presented GCase activity and high stability. Efficacy in vitro was tested in our neuronal GBA-KO model where GBA nanoconjugates were properly internalized and delivered to lysosomes and were able to restore lysosomal GCase activity, reduce sphingolipids accumulation and reduce neurotoxic forms of alpha-synuclein levels. Conclusions: The use of nanotechnological approximations to improve Enzymatic Replacement Therapy is a promising strategy to restore lysosomal GBA activity and a new therapeutic opportunity for Parkinson’s disease treatment.

P497 / #942


A THREE-DIMENSIONAL PATIENT-DERIVED CORTICAL NEUROSPHERE MODEL OF PARKINSONS’ DISEASE

Lecture Title:

W. Raja1, E. Neves1, C. Burke1, X. Jiang1, P. Xu1, V. Khurana2, C. Chung1, R. Scannevin1 1Yumanity Therapeutics, Discovery Biology, Boston, United States of America, 2Brigham and Women’s Hospital, Neurology, Boston, United States of America

Aims: To generate three-dimensional (3D) cortical neurosphere cultures from patient-derived induced pluripotent stem cells (iPSCs) to study the underlying cellular pathology of Parkinson’s disease (PD) and facilitate the development of small molecule therapeutics. Methods: iPSCs derived from a patient harboring a point mutation (A53T) in the alpha-synuclein (a-syn) gene and an isogenic mutation-corrected iPSC line were utilized in these studies. The A53T patient was associated with early-onset PD and a strong dementia phenotype. Neural progenitor cells from both lines were generated and further differentiated into three-dimensional (3D) cortical neurospheres to better recapitulate the microenvironment of the brain. An arrayed 384-microwell format was used. The differentiation process was extensively characterized using quantitative PCR, Western immunoblotting, and immunostaining. Disease-relevant phenotypes were investigated using Western immunoblotting and fatty acid profiling. Results: Our differentiation protocol generated appropriately patterned 3D cortical neurospheres. These cultures were consistent across multiple preparations based on quality-control measures and disease-relevant phenotypes. The latter included elevated levels of desaturated C16 and C18 fatty acids and elevated total and pathogenic phospho-a-syn in the A53T line as compared to isogenic controls. Furthermore, the treatment of the neurospheres with a stearoyl-CoA desaturase (SCD) inhibitor attenuated aberrant fatty acid desaturation and reduced accumulation of pathological a-syn. Conclusions: 3D cortical neurospheres appear to model pathological phenotypes relevant to neurodegenerative disorders such as Parkinson’s disease that are not consistently observed in other cell-based model systems. Based on consistent and reproducible phenotypes and scalability, neurospheres also have demonstrated utility in evaluating the potential of small molecule therapeutics.

P498 / #741


IMPROVED DELIVERY OF ABL301 INTO BRAIN PARENCHYMA OF PARKINSON’S DISEASE MOUSE BRAINS VIA GRABODY B, ABL BIO’S PROPRIETARY BBB SHUTTLE

Lecture Title:

S. An, J.-W. Shin, D. Kim, D. Kim, H. Yun, H. Lee, J. Ahn, J. Eom, S.H. Lee ABL bio, Abl Bio, Seongman-si, Korea, Republic of

Aims: ABL301 is a bispecific antibody composed of an anti-α-synuclein antibody (M30103) with Grabody B, an anti-IGF1R as the ABL Bio’s proprietary BBB technology. Previous study demonstrated improved efficacy of ABL301 than M30103 in a mouse model of Parkinson’s disease. The current study aims to identify 1) temporal and spatial movement of ABL301 out of brain vessel in a Parkinson’s disease animal model, 2) its PK profile, and 3) expression profile of IGF1R in postmortem brains. Methods: Human IgG was visualized by immunostaining of fixed mouse brain sections or in real time using intravital two-photon microscopy. Serum PK was analyzed using ELISA specific to human IgG. Postmortem human brains were analyzed using immunostaining and western blot. Results: ABL301 induced remarkably increased localization in parenchyma of various brain areas and brain cells in mThy-1 hu α-syn mouse brain within 24 hours after dosing. In contrast, M30103 showed signal mainly confined in brain vessel. ABL301 showed comparable serum PK with M30103 in both rodent and monkeys. Clear IGF1R expression was visible in neurons, capillaries and endothelial cells in postmortem brains. Conclusions: ABL301 is localized into brain parenchyma, possibly to its target, aggregated α-synuclein with higher degree than M30103. Its superior serum PK comparable to mAb might have contributed to its sustained BBB penetration over time. IGF1R expression in capillary and endothelial cells in postmortem brains partially validates Grabody B’s usage as a BBB shuttle in clinical studies.

P499 / #1463


NON-CLINICAL PHARMACOKINETIC AND PHARMACODYNAMIC PROPERTIES OF YTX-7739, A CLINICAL STAGE STEAROYL-COA DESATURASE INHIBITOR FOR PARKINSON’S DISEASE

Lecture Title:

D. Tardiff1, M. Lucas2, I. Wrona2, B. Chang1, C. Chung1, B. Le Bourdonnec3, K. Rhodes4, R. Scannevin1 1Yumanity Therapeutics, 40 Guest Street, Suite 4410, Boston, United States of America, 2Black Diamond Therapeutics, Chemistry, Cambridge, United States of America, 3Deciphera Pharmaceuticals, Chemistry, Lawrence, United States of America, 4Wave Life Sciences USA, Inc., 733 Concord Avenue, Cambridge, United States of America

Aims: To characterize the pharmacokinetic properties of YTX-7739 in animals and the corresponding pharmacodynamic changes in fatty acid profiles. Methods: YTX-7739 was administered orally once-daily to multiple animal species for up to 31 days. YTX-7739 levels and fatty acid profiles were assessed in several biofluids and tissues, including determination of the desaturation index (ratio of monounsaturated to saturated fatty acid). The relationships among tolerability, compound concentrations, and fatty acid changes were evaluated. Results: YTX-7739 exhibited activity in vitro, and acceptable bioavailability and brain penetration in vivo. In rats, YTX-7739 was well-tolerated when administered for 15 days at doses achieving concentrations that reduced the desaturation index up to 50% in brain, plasma, and skin. Secondary changes in fatty acids in plasma and skin were evident at the highest dose levels. In cynomolgus macaques, YTX-7739 reduced the desaturation index in plasma and brain after 15 days of dosing. Kinetic analyses of the onset and duration of the pharmacodynamic response in rats and cynomolgus macaques indicated that dosing greater than 5 days was required to achieve a maximal response, and that this reduction persisted for at least 2 days after cessation of dosing. Conclusions: YTX-7739 is a potent, brain-penetrant, well-tolerated stearoyl-CoA desaturase inhibitor that reduces levels of unsaturated fatty acids in a dose-dependent manner in the brains of multiple animal species. In conjunction with efficacy measures described elsewhere, these data support the on-going clinical evaluation of YTX-7739 as a disease-modifying drug for synucleinopathies such as Parkinson’s disease.

P500 / #307

Topic: Theme C: α-Synucleinopathies / C2.c. Therapeutic Targets, Mechanisms for Treatment: Kinases, other enzymes

POLO-LIKE KINASE 2 INHIBITION REDUCES PHOSPHORYLATION OF SERINE 129 ON ALPHA-SYNUCLEIN IN THE NUCLEUS BUT NOT IN LEWY BODY-LIKE AGGREGATES

Lecture Title:

S. Elfarrash1,2, N. Jensen2, N. Ferriera2, S. Schmdit3, E. Gregersen2, M. Meyer3, P.H. Jensen2 1Faculty of Medicine, Mansoura University, Medical Physiology, Mansoura, Egypt, 2Aarhus University, Department Of Biomedicine / Danish Research Institute Of Translational Neuroscience - Dandrite, Aarhus C, Denmark, 3Institute of Molecular Medicine, University of Southern Denmark, Department Of Neurobiology Research, Odense, Denmark

Aims: Accumulation of aggregated and phosphorylated alpha-synuclein (α-syn) is believed to play an important role in the pathophysiology of Parkinson’s disease (PD) and other synucleinopathies. Many studies are carried out to depict the relevance of phosphorylation at serine-129 (pS129) with a considerable controversy. In this study, we investigated the influence of Polo-like kinase 2 (PLK2) inhibition using α-syn recombinant pre-formed fibrils (PFFs) induced synucleinopathy models. Methods: Organotypic hippocampal slice cultures (OHSCs) are made from C57BL/6 pups as described in (Elfarrash et al.,2019). α-syn aggregation is induced by microinjection of PFFs. 24 hours before microinjection, OHSCs are treated with PLK2 inhibitor and replenished thrice/week. At 7 days post-injection, the influence of PLK2 inhibition on α-syn aggregation and spreading are evaluated biochemically and immunohistopathologically. Short-term PLK2 inhibition is also studied using following PFFs models; M83+/− transgenic mice, OHSCs and human stem cell derived dopaminergic neurons where PLK2 inhibition continued for 48, 24 or 4 hours respectively. Results: PLK2 inhibition doesn't affect PFFs induced α-syn aggregation, S129 phosphorylation or spreading in OHSCs. Instead, PLK2 inhibiton reduces the non-aggregate specific phosphorylation in nuclear α-syn. Short-term PLK2 inhibition is removing nuclear S129 phosphorylation in animal model, OHSCs and cell model, increasing signal to noise ratio when assessing aggregate-specific α-syn phosphorylation. Conclusions: PLK2 phosphorylates α-syn at S129 in the nucleus as revealed by the dramatic reduction of pS129 level following PLK2 inhibition. Role of PLK2 in S129 phosphorylation in Lewy body like aggregates is questioned, as inhibition of PLK2 has not affected the phosphorylation of PFFs induced aggregates formation or inter-neuronal spreading.

P501 / #606


DISCOVERY OF SMALL MOLECULES FOR THE TREATMENT OF PD-GBA

Lecture Title:

N. Khatri1, M. Wozniak1, J. Brendel1, C. Holler1, J. Lanter2, D. Burnett2, G. Koenig1, J.-F. Blain1 1Arkuda Therapeutics, Biology, Watertown, United States of America, 2Arkuda Therapeutics, Chemistry, Watertown, United States of America

Aims: Heterozygous mutations in the GBA1 gene, encoding for the lysosomal enzyme glucocerebrosidase (GCase), are the strongest genetic risk factor for Parkinson’s disease (PD). Furthermore, GCase enzymatic activity is reduced in PD-GBA patients, suggesting that loss of GCase activity contributes to the pathogenesis of Parkinson’s disease. Therefore, therapeutics targeted at increasing the activity of GCase may prove efficacious in the treatment of PD. Saposin C, a well-characterized co-factor for GCase activity, is a lysosomal cleavage product of its precursor, prosaposin (PSAP). Methods: We previously discovered novel small molecules that increase progranulin (PGRN) as a potential treatment for frontotemporal dementia, and given the interrelationship between PGRN and PSAP in lysosomal biology, we interrogated the effect of these molecules on PSAP. Results: Here we report the characterization of one of these molecules which promotes increases in secretion and intracellular expression of PSAP, along with increases in intracellular saposins and GCase protein levels. Using a live-cell GCase activity assay, we also show this compound leads to a concentration-dependent increase in endogenous GCase enzymatic activity. These data suggest the effect on GCase activity may be occurring either via the increased expression in GCase, saposin C, or a combination of both. Conclusions: In conclusion, we have discovered small molecules that increase WT GCase activity which can be developed further as potential therapeutics for Parkinson’s disease.

P502 / #639


THE TALE OF PROTEOLYSIS TARGETING CHIMERAS (PROTACS) AND LEUCINE-RICH REPEAT KINASE 2 (LRRK2)

Lecture Title:

A. Oun1,2, M. Konstantinidou3, P. Pathak2, B. Zhang3, Z. Wang3, F. Ter Brake3, A. Dömling3, A. Kortholt2, A. Dolga1 1University of Groningen, Molecular Pharmacology, Groningen, Netherlands, 2University of Groningen, Cell Biochemistry, Groningen, Netherlands, 3University of Groningen, Drug Design, Groningen, Netherlands

Aims: Enhanced kinase function of leucine-rich repeat kinase 2 (LRRK2) has been reported in familial PD and postmortem brain tissue from idiopathic PD patients. Therefore, targeting the kinase function of LRRK2 is beneficial for PD pathogenesis. One promising technique is using Proteolysis targeting chimeras (PROTACs). PROTACs have been designed based on two highly potent, selective and brain-penetrant kinase inhibitors. Biological evaluation for the synthesized PROTACs was performed. Methods: In-vitro kinase assay and microscopy were utilized to check the potency and the cell permeability of the PROTACs, respectively. Immunoblotting was performed to test the ability of different PROTACs to degrade endogenously expressed LRRK2 in RAW 264.7 macrophages. Immunoprecipitation of GFP-tagged LRRK2 was implemented to detect the ubiquitination level of LRRK2 in presence of the PROTACs. Results: All the PROTACs inhibit LRRK2 kinase activity in-vitro with IC50 values in nanomolar range. Furthermore, most of the PROTACs were able to permeate the cells and inhibit LRRK2 kinase activity. Similar to all ATP-competitive inhibitors, the PROTACS induced LRRK2 filament formation on microtubules. However, the PROTACs couldn’t significantly induce ubiquitination nor reduce the degradation level of LRRK2 compared to the original kinase inhibitors. Conclusions: The PROTACs were unable to induce LRRK2 degradation, possibility due to the relocalization of LRRK2 to the microtubules and the inaccessibility to the E3 ligase components. Moreover, the proximity of lysine residues suitable for ubiquitination might not be optimal since the full-length structure of LRRK2 is not resolved. The use of non-ATP-competitive inhibitors or targeting other domains of LRRK2 might be a solution.

P503 / #697

Topic: Theme C: α-Synucleinopathies / C2.d. Therapeutic Targets, Mechanisms for Treatment: Dopamine, neurotransmitters

COGNITIVE FLUCTUATIONS ON DOPAMINERGIC THERAPY IN PATIENTS WITH PARKINSON’S DISEASE

Lecture Title:

S.B. Ismailova, S. Prokopenko, D. Pokhabov Professor V. F. Voino-Yasenetsky Krasnoyarsk State Medical University, Department Of Nervous Diseases With A Course Of Postgraduate Education, Krasnoyarsk, Russian Federation

Aims: Assessment of the dynamics of the cognitive function (CF) during drug treatment with levodopa Methods: The study included 65 patients, which were randomized into 2 groups. In the I group evaluation of CF was conducted 1 hour after receiving levodopa and after 6 months of an hour before the next dose and vice versa in the II group. The inclusion criteria were: verified diagnosis of Parkinson's disease, disease stages 2-3.5 according to the Hoehn-Yahr scale, a stable anti-parkinsonian therapy during the study, complaints of cognitive decline confirmed by the Mini-Cog test, voluntary signed informed consent by the patient. The exclusion criteria were: severe dementia, affective disorders (depression requiring medical correction), somatic pathology in the decompensation stage, severe loss of vision and hearing, severe dysarthria. We used: neurological examination, assessment of the stage of PD using a modified Hoehn and Yahr scale, assessment of the state of CF using the Montreal Cognitive Assessment Scale (MOCA), the mini-mental state examination test (MMSE), the frontal assessment battery (FAB), the scale for assessing the cognitive status in patients with PD (SCOPA-Cog), the geriatric depression scale (GDS). Results: The state of CF in PD patients in the "off" phase of levodopa drugs is statistically significantly worse according to the MMSE, MoCA, FAB, Scopa-Cog scales than in the "on" phase and in the “on” phase of levodopa drugs CF significantly better presented (due to visual-spatial and executive functions). Conclusions: The developed approach to the dynamic assessment of cognitive impairments in PD patients makes it possible to identify cognitive fluctuations in patients taking levodopa drugs.

P504 / #1176

Topic: Theme C: α-Synucleinopathies / C2.d. Therapeutic Targets, Mechanisms for Treatment: Dopamine, neurotransmitters

HUMAN IPSC-BASED MODEL OF TYROSINE HYDROXILASE DEFICIENCY UNCOVERS PATIENT AND CELL TYPE- SPECIFIC DISEASE PHENOTYPES

Lecture Title:

A. Tristán-Noguero1, I. Fernandez-Carasa2,3, C. Calatayud2,3,4, R. Domingo-Jiménez5, M. Pineda6, S. Alcántara3, A. Raya4,7,8, R. Artuch9,10, À. García-Cazorla1,10, A. Consiglio2,3,8,11 1Fundació Sant Joan de Déu, Neurometabolic Unit And Synaptic Metabolism Lab, Neurology Department, Esplugues de LLobregat, Spain, 2Institute of Biomedicine of the University of Barcelona (IBUB), Ibub, Barcelona, Spain, 3Bellvitge Biomedical Research Institute (IDIBELL), Pathology And Experimental Therapeutics (university Barcelona), Hospitalet de Llobregat, Spain, 4Bellvitge Biomedical Research Institute, Program For Advancing The Clinical Translation Of Regenerative Medicine Of Catalonia, Hospitalet de Llobregat, Spain, 5Hospital Clínico Universitario Virgen de la Arrixaca, Departamento De Neuropediatria, Murcia, Spain, 6Fundació Sant Joan de Déu (FSJD) and Hospital Sant Joan de Déu (HSJD), Department Of Neurology, Esplugues de Llobregat, Spain, 7Instituto Carlos III Spain, Centre For Networked Biomedical Research On Bioengineering, Biomaterials And Nanomedicine, Barcelona, Spain, 8Institucio Catalana de Recerca i Estudis Avançats, (icrea), Barcelona, Spain, 9Hospital San Joan de Déu, Department Of Biochemistry, Institut Pediàtric De Recerca, Esplugues de Llobregat, Spain, 10Instituto de Salud Carlos III, Ciberer (centro De Investigación Biomédica En Red Enfermedades Raras), Madrid, Spain, 11University of Brescia, Department Of Molecular And Translational Medicine, Brescia, Italy

Aims: Tyrosine hydroxylase deficiency (THD) is an inherited metabolic disorder caused by mutations in TH gene, leading to dopaminergic depletion and early-onset parkinsonism. Some of affected patients have a good response to L-Dopa (Type A) while others show a severe phenotype with complex early-onset parkinsonism and intellectual disability without response to L-Dopa (Type B). Although the genetic mutations are known, the effect of TH mutation in relevant neurons is poorly elucidated. Methods: Here, we generated induced pluripotent stem cell (iPSC) lines from Type A and B patients, age-matched healthy individuals or isogenic-corrected counterparts. We used non-integrative episomal vectors to deliver factors to generate several independent iPSC lines per individual, that showed a fully pluripotency. Results: Upon neuronal differentiation, iPSC-derived mutant neurons showed a phenotype which recapitulates human disease phenotype: decrease in TH protein expression, low levels of DA metabolites and reduced expression of DA-related genes when compared to control iPSC-derived neurons. Interestingly, in addition we identified specific defects unique for each type of phenotype as shown by reduced neurite arborisation in Type B mutant neurons and reduced axonal TH localization in Type A. The impact of TH expression (achieved by CRISPR) in reverting the pathological phenotype and its potential impact on human neurons’ biology were also assessed. Finally, by treating the cells with L-DOPA, we were able to rescue the phenotypic alterations only in THD-A neurons. Conclusions: Overall, our findings show that dysfunctional neurons play a crucial role during THD pathogenesis and may have broad implications for future intervention in early stages of THD.

P505 / #988

Topic: Theme C: α-Synucleinopathies / C2.e. Therapeutic Targets, Mechanisms for Treatment: Cell transplantation

A BIOENGINEERED NICHE STIMULATES PLURIPOTENCY OF NEURAL PRECURSORS CELLS DURING EXPANSION AND ENHANCES THEIR THERAPEUTIC EFFICACY

Lecture Title:

B. Barzaghini1, T. Giallongo2, F. Rey2, T. Zandrini3, A. Pulcinelli1, R. Nardomarino1, G. Cerullo3, R. Osellame3, C. Cereda4, G.V. Zuccotti2, M.T. Raimondi1, S. Carelli2 1Politecnico di Milano, Department Of Chemistry, Materials And Chemical Engineering "giulio Natta", Milan, Italy, 2University of Milan, Dept. Biomedical And Clinical Sciences "l. Sacco", Pediatric Clinical Research Center Fondazione “romeo Ed Enrica Invernizzi”, University Of Milan, Milan, Italy., Milano, Italy, Italy, 3Politecnico di Milano, Istituto Di Fotonica E Nanotecnologie (ifn)-cnr And Department Of Physics, Milan, Italy, 4IRCCS Mondino Foundation, Genomic And Postgenomic Lab,, Pavia, Italy

Aims: OBJECTIVES: Biomaterials have been used to create micro-scaffolds, allowing the generation of active biophysical signals for directing stem cell fate. The Nichoid, has been reported to maintain the stemness capacity in cells of different origins [1]. The aim of this study was to investigate: i) the proliferation, differentiation and stemness properties of neural precursor cells expanded inside the Nichoid (NIC-NPCs); ii) the therapeutic potential of NIC-NPCs in preclinical experimental model of Parkinson’s Disease (PD) [2,3]. Methods: METHODS: Nichoids were fabricated by two-photon laser polymerization using an organic-inorganic photoresist. NPCs expanded inside the Nichoid were counted and analyzed through RNA-seq, Real Time PCR, western blot and immunofluorescence. Parkinsonism was induced by the administration of MPTP in C57/black[3] and the recovery of function was investigated with behavioral tests. Results: RESULTS: 7 days after plating, the NIC-NPCs show a significantly higher proliferation and viability than in standard floating conditions and NIC-NPCs show an increase in pluripotency. Moreover, NIC-NPCs showed a potentiation in their therapeutic features by counteracting neurodegeneration and promoting recovery of function in vivo. These evidences are supported by histological analysis of specific molecular markers. Moreover, NIC-NPCs downregulated gliosis. Conclusions: CONCLUSIONS: Here we provide evidences that NIC-NPCs both favors their pluripotency and potentiates their efficacy showing great promise in the field of regenerative medicine. REFERENCES: Raimondi et al. 2013; 10.1016/j.actbio.2012.08.022 Carelli et al. 2017; 10.1016/j.neuropharm.2017.03.035 Carelli et al. 2018; 10.1186/s12974-018-1375-2 ACKNOWLEDGEMENTS: Financial support was received from “Neurogel-en-Marche” Foundation; Fondazione “Romeo and Enrica Invernizzi”; ERC grant NICHOID, G.A. 646990 and NICHOIDS, G.A. 754467.

P506 / #354

Topic: Theme C: α-Synucleinopathies / C2.f. Therapeutic Targets, Mechanisms for Treatment: Deep brain stimulation

NON-INVASIVE TRANSCRANIAL DIRECT CURRENT STIMULATION AFFECTS ON THE AMOUNT OF DAILY ACTIVITY AND MOTOR FUNCTIONS IN PARKINSON’S DISEASE PATIENTS.

Lecture Title:

D.-Y. Kwon1, M.H. Park1, H.K. Yoon2 1Korea university, Neurology, Ansan-city, Korea, Republic of, 2Korea university, Psychiatry, Ansan-city, Korea, Republic of

Aims: To investigate the effects of transcranial direct current stimulation on the ADL, sleep and motor functions in Parkinson's disease measured with individual smartband recordings. Methods: All PD participants who met UK Parkinson Disease Society Brain Bank criteria were evaluated their severity using part III of the UPDRS. PD patients who taking antiparkinsonian therapy without medication change during the recent 3 months were recruited. Total 10 times of tDCS was applied for 20 minutes with 2mA current over left dorsolateral prefrontal cortex. The primary outcome was assessed by average number of total daily steps and the amount of sleep time on the smart-band recording checked for 7days before and after tDCS. UPDRS part III score and the change of depression rating score were analyzed as secondary outcomes. Results: Total 20 PD patients were completed the study. We found that the total step counts were significantly increased after stimuli.(p=0.012) The total activity time, sleep time, deep sleep time were improved without statistical significance. UPDRS part III score and patient global impression score were significantly improved (p<0.01). Clinical improvement last for over one month after stimulation finished. Conclusions: Short-term active tDCS on left DLPFC was well-tolerated and resulted in improvements in activity of the subjects with PD, indicating a possible adjunctive therapeutic option for motor symptoms and improve the activity of daily living function in real world situations. Further investigation involving large group with randomized double-blind design is recommended to exploring tDCS effects as an adjuvant therapy for specific drug-resistant motor symptoms and activity in PD.

P508 / #1116

Topic: Theme C: α-Synucleinopathies / C2.g. Therapeutic Targets, Mechanisms for Treatment: Anti-inflammatory, anti-oxidant

PARAQUAT NEUROPROTECTION INDUCED BY ETHANOLIC EXTRACTS FROM SOME ZANTHOXYLUM SPECIES (RUTACEAE) NATIVE TO COLOMBIA IN DROSOPHILA MODEL

Lecture Title:

A. Ceballos Torres1, A. Vega Quiñones1, G. Arboleda-Bustos1,2, A. Sandoval-Hernández3, M. Ávila3, L. Cuca Suarez3 1Universidad Nacional de Colombia, Instituto De Genética, Bogota, Colombia, 2Universidad Nacional de Colombia, Facultad De Medicina, Bogotá, Colombia, 3Universidad Nacional de Colombia, Facultad De Ciencias - Departamento De Química, Bogotá, Colombia

Aims: Parkinson's disease (PD) is the second cause of dementia worldwide, the cause of the disease has been associated to oxidative stress and mitochondrial dysfunction inducing apoptosis of dopaminergic neurons of substantia nigra pars compacta. The exposition of the herbicide Paraquat (PQ) has been linked to PD in humans, as PQ is an inhibitor of mitochondrial Complex-I producing oxidative stress and mitochondrial dysfunction. Plants from Zanthoxylum genus are characterized for the presence alkaloids, amides, coumarins, flavonoids, lignans, sterols and terpenes, among others antioxidant agents potentially used for prevents neurodegeneration. The present paper aims to assess the potential neuroprotective of native Colombian natural products from Zanthoxylum genus (Rutaceae) against PQ used as model of PD in Drosophila melanogaster. Methods: We used male adult wild-type Drosophila melanogaster flies 0-2 days old that were exposed to PQ and/or ethanolic extracts for 7 days. Survival was monitored counting death flies per day, and negative geotaxis assay was performed every 2 days to assess locomotor activity. Results: Significant Improvements in motor activity and survival rates were observed in flies treated with Zanthoxylum extracts compared to PQ treatment. Experiments showed that Z. caribeum and Z. schreberi coadministered with PQ significantly improves survival rates compared with PQ only treatment. In functional analysis we found that motor impairment produced by PQ was reduced by Z. caribeum, Z. schreberi and Z. rigidium roots. Conclusions: Our findings suggest that the extracts obtained from native Colombian plants possess therapeutic potential for the prevention of PD as inhibitors of oxidative stress potentially used for neuroprotection in PD.

P509 / #491


NEUROPROTECTION OF LOW DOSE CARBON MONOXIDE IN GENETIC AND TOXIN MODELS OF PARKINSON’S DISEASE

Lecture Title:

S. Gomperts1, K. Rose1, M. Zorlu1, X. Xue1, E. Gomperts2, W. Cai1, M. Schwarzschild1, X. Chen1 1Massachusetts General Hospital, Neurology, Charlestown, United States of America, 2Hillhurst Biopharmaceuticals, Inc., Clinical Development And Scientific Affairs, Montrose, United States of America

Aims: Exposure to low doses of carbon monoxide (CO) may underlie the reduced risk of Parkinson’s disease (PD) among smokers. The goal of this research is to evaluate the neuroprotective potential of low-dose CO treatment in PD models. Methods: In the AAV-alpha-synuclein (aSyn) model, rats underwent right nigral injection of AAV1/2-asynA53T, with left injection of empty AAV and were treated with oral CO drug product (HBI-002 10ml/kg, daily by gavage) or vehicle from days 5-21. In the short-term MPTP model, mice were treated with inhaled CO (iCO) (250ppm) or air for 1 hour, 1 hour after MPTP injection (40mg/kg, i.p.), and sacrificed at day 6. HPLC measurement of striatal dopamine and immunohistochemistry for nigral aSyn and tyrosine hydroxylase for stereological cell counting were conducted blinded to treatment condition. Results: Each HBI-002 treatment increased carboxy-hemoglobin to 9%. AAV-aSyn rats treated with HBI-002 retained 74±12% (SD) of right striatal dopamine versus left, compared to 36±7% in vehicle-treated rats (p=0.03). HBI-002 treatment was associated with increased preservation of right-sided TH+ cell counts, to 74±4% versus left, compared to 38±3% with vehicle (p=0.0001). HBI-002 reduced aSyn aggregates (p=0.02) and S129 phosphorylation (p=0.005). MPTP-exposed mice treated with iCO had 46% higher dopamine levels (p=0.03) and 22% more TH+ neurons (p=0.04) than those treated with air. In saline-treated mice, iCO had no effect on striatal dopamine levels or TH+ cell counts. Conclusions: Low-dose CO protects nigral dopamine neurons, preserves striatal dopamine content, and reduces aSyn pathology in animal models of PD. Low dose CO may have disease-modifying activity in PD.

P510 / #1186


THE REVERSAL OF THE DOPAMINERGIC NEURON LOSS IN THE EX VIVO PARKINSON’S DISEASE MODEL BY THE ANTI-INFLAMMATORY ACTION OF BILIRUBIN

Lecture Title:

S. Jayanti1,2,3, R. Moretti4, C. Tiribelli1, S. Gazzin1 1Italian Liver Foundation, Fondazione Italiana Fegato, Trieste, Italy, 2University of Hasanuddin, Faculty Of Medicine, Makassar, Indonesia, 3University of Trieste, Molecular Biomedicine Ph.d. Program, Trieste, Italy, 4University of Trieste, Neurology Clinic, Department Of Medical, Surgical, And Health Sciences, Trieste, Italy

Aims: Investigating the dopaminergic neuron protection in the ex vivo Parkinson’s disease (PD) model by unconjugated bilirubin (UCB) challenging and identifying the protective UCB concentrations threshold. Methods: The ex vivo PD model has been developed by using the organotypic brain cultures (OBCs) of substantia nigra obtained from Wistar rat at day 5 post-natal exposure to rotenone (Rot). The OBCs were co-challenged with Rot and UCB (0.5-4 μM) for 24 hours. The number of dopaminergic neurons (DOPAn) was assessed by tyrosine hydroxylase (TH+) immunofluorescence staining. RT-qPCR was used to monitor oxidative stress marker genes (Srnx1), inflammation-related genes (Tnfα, Il6, Cox2), and neurotrophic genes (Bdnf). Results: The DOPAn number was significantly reduced in Rot-treated OBCs (-35%, p < 0.01). The full reversal of DOPAn number was obtained by challenging 0.5 μM and 1 μM UCB (p<0.05 vs. Rot). Inflammation (Tnfα, Il6, Cox2), redox imbalance (Srnx1), and neurotrophic factor (Bdnf) mRNA expressions were up-regulated in the PD model (p<0.05). 0.5 μM UCB reduced Tnfα expression (p<0.01), 1 μM UCB treatment resulted in downregulation of the expression of Tnfα, Il6, and Bdnf (p<0.05, p<0.01, and p<0.05, respectively). 2 μM UCB decreased the expression of Il6 (p<0.01), while 4 μM UCB increased the mRNA level of all the selected markers above the expression in Rot challenged slices. Conclusions: The dopaminergic neuron loss in the ex vivo PD model has been reversed by the anti-inflammatory action of UCB at low concentrations (0.5 μM and 1 μM).

P511 / #727


SYSTEMIC MC1R ACTIVATION MODULATES IMMUNE RESPONSE AND PROTECTS THE NIGROSTRIATAL DOPAMINERGIC SYSTEM IN MPTP+LPS MODEL OF PD

Lecture Title:

P. Srivastava, W. Cai, T. Stimpson, M. Schwarzschild, X. Chen Massachusetts General Hospital, Neurology, Charlestown, United States of America

Aims: We previously identified a role of Melanocortin receptor 1 (MC1R) in dopaminergic neuron survival. MC1R is present on immune cells and can regulate the immune system. The present study explores the effects of systemically-administered NDP-MSH, an MC1R agonist and existing drug that does not cross BBB, in modifying the immune response and protecting the nigrostriatal dopaminergic system in the MPTP+LPS mouse model of PD. Methods: Male C57BL/6 mice were treated intraperitoneally with MPTP (17 mg/kg) and LPS (1 mg/kg) from day 1 to day 4 and NDP-MSH (400 µg/kg) or vehicle from day 1 to day 12 following which the mice were sacrificed. Immune cells in the blood, striatal dopamine, and nigral TH+ neurons and glia, and expression of occludin, a marker for BBB integrity were assessed. Results: NDP-MSH treatment significantly attenuated striatal dopamine depletion (by 37%, p<0.001) and nigral TH+ neuron loss (by 33%, p<0.01) induced by MPTP+LPS. NDP-MSH reduced microglia activation and IBA1+ cells in the nigral region. No significant difference in occludin expression was observed in NDP-MSH treated mice vs controls in the striatum. NDP-MSH treatment in MPTP+LPS mice significantly reduced blood CD4+ T cells (p=0.006) and B cells (p=0.04). NDP-MSH did not show any protective effect in MC1R mutant mice following MPTP+LPS. Conclusions: A peripherally acting MC1R agonist confers protection on dopaminergic nigrostriatal neurons and reduces hyperactivate states of microglia. Whether the neuroprotection by NDP-MSH results, at least in part, from its immune-modulating effects remains to be determined.

P512 / #1233

Topic: Theme C: α-Synucleinopathies / C2.j. Therapeutic Targets, Mechanisms for Treatment: Protein aggregation, misfolding, chaperones

NANOPARTICULATE COMPLEX OF NICOTINE AND CERIUM OXIDE (NANOCERIA) PRESERVES ELECTROPHYSIOLOGICAL NEURONAL FUNCTION IN AN IN VITRO PARKINSON’S DISEASE MODEL.

Lecture Title:

S. Imam1, Z. Zhang1, A. Guzman-Lopez2, E. Cuevas1, M. Ramirez-Lee1 1US FDA/NCTR, Div Of Neurotoxicology, Jefferson, United States of America, 21Escuela Nacional Preparatoria-UNAM, Biochemistry, Mexico, Mexico

Aims: The pathophysiology of dopaminergic (DA) loss in Parkinson’s disease (PD) is still unclear. Here, we report that a combination of Nicotine and NanoCeria (Nic-NANO) results in efficient prevention of PD progression in a 1-methyl-4-phenylpyridinium (MPP+) induced in vitro model of PD. Methods: Human primary dopaminergic neuronal precursor cells (HPDNPCs) were seeded on poly-L-lysine (PLL)-coated Multi-Electrode Array (MEA) and maintained in differentiation media for 7 days. HPDNPCs were exposed to nicotine (0.1 μM), NanoCeria (CNP) or Nic-NANO conjugate (200 nM), as well as to PLGA encapsulated Nic-NANO (5 and 10 μg/mL). HPDNPCs were treated with 1 mM MPP+ after 6 h of nanomaterial (NM) exposure. MEA recordings of HPDNPCs were performed prior to and 6, 24, 48 and 72 h after exposure in the absence or presence of MPP+. Action potentials were distinguished from noise using voltage threshold 5 times the standard deviations. Results: MPP+ decreased electrophysiological activity at all time points, modeling the altered neural activity observed in PD. Basal neural activity was not altered by CNP, Nic-NANO, or PLGA in the absence of MPP+. However, all nanoparticulate conjugates significantly prevented the MPP+-induced altered neural activity, even at the 72 h timepoint. Conclusions: We report for the first time that this therapeutic combination of Nic-Nano prevents MPP+ induced loss of neuronal function. A correlation of this functional recovery with preservation of dopaminergic phenotype was also observed. In summary, our results suggest that the combination of nicotine and NanoCeria might be a successful approach to prevent PD progression and preserve dopaminergic function during PD.

P513 / #1168

Topic: Theme C: α-Synucleinopathies / C2.j. Therapeutic Targets, Mechanisms for Treatment: Protein aggregation, misfolding, chaperones

THERAPEUTIC TARGET OF GBA IN PARKINSON’S DISEASE WITH NOVEL PHARMACOLOGICAL CHAPERONES

Lecture Title:

M. Montpeyó1, A. Navarro-Romero1, A.M. Garcia-Collado2, N. Perez-Carmona2, I. Fernandez-Gonzalez1, J. Riera1, A. Ruano2, A. Delgado2, S. Morales2, E. Cubero2, X. Barril2, M. Martinez-Vicente1 1Vall d'Hebron Institut de Recerca (VHIR), Neurodegenerative Diseases, Barcelona, Spain, 2Gain Therapeutics Switzerland, Barcelona Branch, Barcelona, Spain

Aims: To apply a novel in vitro neuronal model to validate the use of new pharmacological chaperones aimed to restore GBA activity as a therapeutic strategy in Parkinson’s disease. Methods: A new in vitro model consisting of differentiated dopaminergic-like neurons expressing GBA carrying N370S, L444P and D409V mutations was used as an in vitro disease model to validate new pharmacological GBA chaperones. The compounds were assayed for their ability to increase neuronal GCase activity, reduce neurotoxic forms of alpha-synuclein and reduce the accumulation of toxic substrate. The non-inhibitory compounds used in the assay were identified by Gain Therapeutics using its proprietary SEE-Tx technology. Compound selection was done based on the following compound characteristics: i) increasing GCase activity in vitro, ii) crossing the BBB, iii) lack of toxicity and iv) low metabolism. Results: The outcome of the novel neuronal in vitro assay confirmed that Gain’s compounds increase GBA protein levels and activity, reduce the accumulation of total and phosphorylated alpha-synuclein species and reduce the accumulation of glucosylsphingosine. Conclusions: Our neuronal in vitro system is a useful Parkinson/GBA model for pharmacological screening and validation of GBA chaperones. The brain penetrant Gain Therapeutics compounds show a big therapeutic potential for the treatment of Parkinson’s and Gaucher’s diseases. The results will be confirmed in future in vivo studies.

P514 / #322

Topic: Theme C: α-Synucleinopathies / C2.m. Therapeutic Targets, Mechanisms for Treatment: Neurogenesis and iPSC

THE NOVEL MECHANISTIC ROLE OF ALPHA-SYNUCLEIN IN THE NUCLEUS: IMPAIRED NUCLEAR FUNCTION CAUSED BY FAMILIAL PARKINSON’S DISEASE SNCA MUTATIONS

Lecture Title:

B. Kantor1, V. Chen2, M. Moncalvo3, W. Dong3, O. Chiba-Falek2 10702356, Neurology, Durham, United States of America, 2Duke University, Neurology, Durham, United States of America, 3Duke University, Neurobiology, Durham, United States of America

Aims: Here, we aimed to investigate the mechanistic role of SNCA in the nucleus utilizing isogenic hiPSC-derived neurons from PD patients with autosomal dominant mutations, A53T and SNCA-triplication, and their corresponding corrected lines by genome- and epigenome- editing. Methods: In this study, we employed a range of physiological-stress tests paired with immunohistochemistry (IHC), co-IP and real-time PCR analysis to elucidate the mechanistic role of SNCA in the nucleus utilizing isogenic human induced Pluripotent Stem Cells (hiPSC)-derived neurons from PD patients and their corresponding genome-edited lines with the corrected mutations. Results: We investigated the mechanistic role of SNCA in the nucleus utilizing isogenic hiPSC-derived neurons from PD patients with autosomal dominant mutations, A53T and SNCA-triplication, and their corresponding corrected lines by genome- and epigenome- editing. Comparisons of shape and integrity of the nuclear envelope and its resistance to stresses found that both mutations result in similar nuclear envelope perturbations that were reversed in the isogenic mutation-corrected cells. Further mechanistic studies showed that SNCA mutation has adverse effects on the nucleus by trapping Ras-related nuclear protein (RAN) and preventing it from transporting key nuclear proteins such as, DNMT3A, for maintaining normal nuclear function. Conclusions: For the first time, we proposed that a-syn interacts with RAN and normally functions in the nucleocytoplasmic transport while exerts its dominant-negative pathogenic effect by sequestering RAN. We suggest that defects in the nucleocytoplasmic transport components may be a general pathomechanistic driver of neurodegenerative diseases.

P515 / #1826

Topic: Theme C: α-Synucleinopathies / C2.n. Therapeutic Targets, Mechanisms for Treatment: Other

EVALUATION OF A NEUROPROTECTIVE PACAP GLYCOPEPTIDE TO TREAT MOTOR AND COGNITIVE SYMPTOMS IN TWO RODENT MODELS OF PARKINSON’S DISEASE

Lecture Title:

K. Bernard1, M. Bartlett2, C. Liu3, G. Molnar4, C. Apostol3, L. Szabos3, S. Sherman2, L. Madhavan2, R. Polt3, J. Streicher4, M. Heien3, T. Falk2 1University of Arizona, Physiological Sciences Gidp, Tucson, United States of America, 2University of Arizona, Department Of Neurology, Tucson, United States of America, 3University of Arizona, Chemistry And Biochemistry, Tucson, United States of America, 4University of Arizona, Medical Pharmacology, Tucson, United States of America

Aims: We have designed and synthesized a glycosylated PACAP analog (PACAP1-27S-Lac) in order to increase blood-brain barrier (BBB) penetrance and promote survival and inhibit inflammation in neurodegenerative disease. In order to test for protective ability of the PACAP1-27S-Lac, we conducted a study using a mild progressive unilateral 6-hydroxydopamine rat PD model. We have also begun a pilot to determine if PACAP1-27S-Lac is able to reduce cognitive symptoms in the Thy1-Asyn mouse model of PD. Methods: Using in vivo microdialysis in rats coupled with LC-MS3 we show PACAP1-27S-Lac (n=3, 15mg/kg, i.v.) is able to penetrate the BBB. To test for protective ability of the PACAP1-27S-Lac (15mg/kg; i.p.), we used a mild progressive unilateral 6-hydroxydopamine (6-OHDA; 2 x 13.75 μg in striatum) rat PD model (n=16/group). Unbiased stereology on cells with the substantia nigra as well as motor behavior tasks in the 6-OHDA lesioned rats and cognitive tasks in the Thy1-Asyn mouse PD model are employed to test the efficacy in vivo. Results: PACAP1-27S-Lac stimulated cAMP production in vitro using CHO cell lines expressing PACAP receptors. Preliminary analysis using unbiased stereology of a subset of the 6-OHDA-lesioned rats indicates a lesion size of 20% in the PACAP-treated compared to 30% in the vehicle-treated groups. Video analysis of motor behavior is ongoing to determine the exact extent of neuroprotective activity. Conclusions: Our glycosylated PACAP peptide is able to cross the BBB and stimulate cAMP production through its endogenous receptors, which could indicate translational potential for PACAP1-27S-Lacas a novel therapeutic to treat PD.

P516 / #996


VMAT2 OVEREXPRESSION REDUCES INTRACELLULAR NEUROMELANIN LEVELS AND ATTENUATES PARKINSON-LIKE PATHOLOGY IN NEUROMELANIN-PRODUCING PARKINSONIAN RATS.

Lecture Title:

J. Compte Barron1, M. Vila2,3,4, M. Gonzalez-Sepulveda4, T. Cuadros4, J. Romero-Giménez4, A. Parent4, A. Laguna4 1Vall d´Hebron Research Institute, Neurodegenerative Diseases Group, Barcelona, Spain, 2Neurodegenerative Diseases Research Group, Vall d’Hebron Research Institute (VHIR)-Center for Networked Biomedical Research on Neurodegenerative Diseases (CIBERNED), Department Of Biochemistry And Molecular Biology, Autonomous University Of Barcelona (uab), Barcelona, Spain, 3Catalan Institution for Research and Advanced Studies (ICREA), Na, Barcelona, Spain, 4Vall d’Hebron Research Institute (VHIR)–Center for Networked Biomedical Research on Neurodegenerative Diseases (CIBERNED), Neurodegenerative Diseases Research Group, Barcelona, Spain

Aims: In Parkinson’s disease (PD), there is a preferential degeneration of neurons that contain the pigment neuromelanin, especially dopaminergic neurons of the substantia nigra (SN), the loss of which leads to classical motor PD symptoms. We recently developed the first rodent model of human-like neuromelanin production based on the viral vector-mediated nigral expression of melanin-producing enzyme tyrosinase (AAV-hTyr). This has revealed that neuromelanin can trigger PD pathology when accumulated above a specific pathogenic threshold. Because neuromelanin derives from the oxidation of free cytosolic dopamine, we hypothesized that enhancing dopamine vesicular encapsulation with vesicular monoamine transporter 2 (VMAT2) will decrease cytosolic dopamine that can convert to neuromelanin. This approach should slow down the intracellular buildup of neuromelanin that occurs with age and thus prevent, delay or attenuate PD pathology. Methods: Adult male Sprague-Dawley rats received single unilateral stereotaxic co-injections of AAV-hTyr and AAV-VMAT2 immediately above the SN. Control animals received equivalent amounts of either vehicle, AAV-hTyr or AAV-VMAT2, separately. At selected times post-injection animals were assessed for motor asymmetry and their brains processed for histological analyses. Results: VMAT2 overexpression in hTyr-expressing rats reduced intracellular neuromelanin to levels below its pathogenic threshold by lowering the production of dopamine-oxidized neuromelanin precursors. Reduction of intracellular neuromelanin levels was associated with an attenuation of PD-like motor deficits and nigrostriatal denervation in these animals. Conclusions: Our results demonstrate the feasibility and therapeutic potential of modulating intracellular neuromelanin levels in vivo.

P517 / #1517


TREMOR QUANTIFICATION THROUGH EVENT-BASED MOVEMENT TRAJECTORY MODELING BEFORE AND AFTER UNILATERAL RADIOFREQUENCY SUB-THALAMOTOMY

Lecture Title:

W.O. Contreras Lopez1, L. Guayacan2, F. Martinez2, E. Fonoff3, P. Navarro1 1Clinica Foscal Internacional/NEMOD, Neurosurgery/translational Research, Floridablanca, Colombia, 2UIS, System Engineering, Bucaramanga, Colombia, 3University of Sao Paulo, Neurosurgery/functional Neurosurgery, Sao Paulo, Brazil

Aims: To describe the effect of bilateral subtalamotomy by radiofrequency (RF) by quantifying the tremor in hand from trajectories of movement. Methods: An 83 years-old female patient with a history of chronic obstructive pulmonary disease, low weight (BMI: 16) and Parkinson's Disease (PE) Hoehn & Yahr II. Presented 5 years of tremor at rest of high frequency and great amplitude of predominance in right hand with significant limitation of quality of life. It was decided to perform a unilateral subtalamotomy for RF (2 lesions at 75 ° C, 1:00 min each). The tremor was measured by an original computational method measuring a set of motion trajectories, calculated using video. The trajectories were characterized by a set of kinematics, which with a statistical analysis allowed to quantify the tremor. Additionally, it was followed with part 3.16 of the UPDRS, the quality of life questionnaire (PDQ-39) and the Fahn-tolosa tremor scale before and two years after the procedure. Results: There were no complications. The tremor stopped intra-surgically and this benefit was maintained until the last visit. The scores in UPDRS-3.16 decreased by 47.6%, in the Fahn-Tolosa tremor scale by 60% and a concomitant improvement in quality of life was also observed with the PDQ-39 questionnaire in 66%. Conclusions: The subtalamotomy by RF provided significant control of tremor and rigidity, significantly improving the quality of life of an 83-year-old patient whose nutritional status was not a candidate for DBS. The software allowed an accurate evaluation of tremor outcome.

P518 / #947


A RANDOMIZED, DOUBLE-BLIND, PLACEBO-CONTROLLED TRIAL ASSESSING THE SAFETY AND TOLERABILITY OF GRF6021 IN SUBJECTS WITH PARKINSON’S DISEASE AND COGNITIVE IMPAIRMENT

Lecture Title:

E. Rawner1, K. Koborsi1, V. Klutzaritz1, J. Duncan1, K. Nikolich1, K. Marek2, S. Braithwaite1, J. Hannestad1 1Alkahest, Clinical Development, San Carlos, United States of America, 2INDD, Clinical, New Haven, United States of America

Aims: The plasma fraction GRF6021 reverses age-related changes in the mouse brain, including impaired memory, reduced synaptic density, and neuroinflammation. In this trial, we evaluated the safety and tolerability of GRF6021 in subjects with Parkinson’s Disease (PD) and cognitive impairment. Methods: In this 24-week trial, subjects aged 40-85 with a diagnosis of PD with mild cognitive impairment or dementia were randomized in a 2:1 ratio to receive 250 mL GRF6021 or placebo IV daily for 5 consecutive days during weeks 1 and 13. Study drug allocation was blinded to subjects, raters, and investigators. The primary endpoint was safety and tolerability. Secondary endpoints included the Montreal Cognitive Assessment (MoCA), the Cognitive Drug Research system, the Delis-Kaplan Executive Function System verbal fluency, the Movement Disorders Society Unified PD Rating Scale (MDS-UPDRS), the PD Questionnaire 39, and the DCTclock digital clock drawing. Results: 79 subjects (26 women, 53 men) with a mean age of 67, an MDS-UPDRS Total score of 65 (range 20-123) and a MoCA score of 22 (range 14-25) received GRF6021 (n=53) or placebo (n=26). GRF6021 was safe and well tolerated compared to placebo. The trial was not statistically powered to detect differences in secondary efficacy endpoints; however, numerical superiority of GRF6021 over placebo was observed on multiple cognitive endpoints. Conclusions: This trial met its primary endpoint by demonstrating that daily infusions of GRF6021 for 5 consecutive days at 3-month intervals were safe and well tolerated. Secondary endpoints indicated that GRF6021 may have cognitive benefits and future development in PD with cognitive impairment is planned.

P519 / #260

Topic: Theme C: α-Synucleinopathies / C3.b. Drug Development, Clinical Trials: Vitamins, antioxidants, neuroprotective compounds

PARKINSON’S DISEASE RELATED DEMENTIA AND VITAMIN D CORRELATION

Lecture Title:

S. Bejjani, S. Gebeily Lebanese University, Neuroscience Research Center, Beirut, Lebanon

Aims: Dementia is a frequent co-morbid disorder of Parkinson’s disease (PD) and has been included in the recent definition of PD updated by the DSM. The relationship between vitamin D (vit-D) and PD-related dementia (PDRD) is still not well understood.Determinate if patients with PDRD have lower vit-D level than PD patients with no dementia, and investigate the evidence of a possible correlation between vit-D serum levels and PDRD, especially Parkinson’s disease dementia (PDD) and Dementia with Lewy bodies (DLB). Methods: A retrospective study was conducted on 22 Lebanese outpatients who performed a vit-D serum level measurement. Patients with advanced cognitive deterioration according to the Unified Parkinson Disease Rating Scale (UPDRS) Part I who had a mentation score of 10/16 and higher were recognized as having PDRD. In reference to the Food and Nutrition Board (2010), we considered a vit-D deficiency for a level lower than 20ng/mL. SPSS software was used to compare the data from our PDRD sample with those from two Lebanese control groups and a PD group. Results: The mean serum vit-D level in our PDRD group patients was significantly lower than the non-demented PD-related group (14.5±8.42 ng/mL vs 17.61±11.17 ng/mL), whereas 72.72% of the total PD-related population were also in the range of vit-D deficiency. Conclusions: PDRD patients tend to have lower serum 25(OH)D than the Lebanese control groups. No significant association was found between vit-D levels and dementia. Longitudinal observational and interventional large scale studies are needed to confirm the correlation between low vit-D level and PDRD.

P520 / #537

Topic: Theme C: α-Synucleinopathies / C3.b. Drug Development, Clinical Trials: Vitamins, antioxidants, neuroprotective compounds

PHASE I-II FIRST-IN-MAN CLINICAL TRIAL OF INTRAPUTAMENAL CDNF IN PARKINSON’S DISEASE: EXPLORATORY FLUID-BASED BIOMARKER ENDPOINTS OF THE 12-MONTH TREATMENT PERIOD

Lecture Title:

H. Huttunen1, S. Booms1, N. Majbour2, H. Abdesselem2, O. El-Agnaf2, I. Poggiolini3, L. Parkkinen4, A. Cryar5, J. Harris5, E. Sirka5, M. Dove5, M. Sjogren1, M. Woolley6, B. Muprhy6, P. Fielder6, J. Baker6, P. Skinner6, M. Andreasson7, G. Paul-Visse8, R. Kivisaari9, H. Bjartmarz8, G. Lind10, P. Almqvist10, F. Scheperjans11, H. Widner8, P. Svenningsson7 1Herantis Pharma Plc, N/a, Espoo, Finland, 2Qatar Biomedical Research Institute, Neurological Disorders Research Center, Doha, Qatar, 3University of Oxford, Oxford Parkinson's Disease Center, Oxford, United Kingdom, 4University of Oxford, Nuffield Department Of Clinical Neurosciences, Oxford, United Kingdom, 5Inoviv Ltd, N/a, London, United Kingdom, 6Renishaw Neuro Solutions Ltd, N/a, Gloucestershire, United Kingdom, 7Karolinska University Hospital, Department Of Neurology, Stockholm, Sweden, 8Lund University Hospital, Department Of Neurology, Lund, Sweden, 9Helsinki University Central Hospital, Department Of Neurosurgery, Helsinki, Finland, 10Karolinska University Hospital, Department Of Neurosurgery, Stockholm, Sweden, 11Helsinki University Central Hospital, Department Of Neurology, Helsinki, Finland

Aims: Cerebral dopamine neurotrophic factor (CDNF) is an unconventional neurotrophic factor that in preclinical models of Parkinson’s disease (PD) protects dopamine neurons via a unique multi-modal mechanism of action. As part of the first-in-man clinal study of intermittent bilateral intraputamenal monthly infusions of CDNF in subjects with moderately advanced PD, we explored cerebrospinal fluid (CSF) biomarkers before and after treatment. Methods: A randomized, placebo-controlled, double-blind phase I-II trial in 17 patients with moderate PD, comprising placebo or incremental CDNF dosing for six-month followed by an active treatment six-month extension study. Lumbar CSF samples were collected at baseline and at 20 and 45 weeks after first dosing. As an exploratory endpoint, the levels of total, oligomeric and Ser129-phosphorylated α-synuclein were determined by ELISA. The aggregation propensity of CSF α-synuclein was determined by RT-QuiC assay. A proteomic biomarker screen was performed using mass spectrometric methods. Results: The data for the α-synuclein analyses and proteomics screen will be available in Q4 2020. In the discovery phase of the proteomic screen, the PD subjects at baseline (n=15) were compared to healthy, age-matched controls (n=14). Based on the results, a panel 50 proteins were selected for a targeted proteomics screen to be conducted with CSF samples collected at different timepoints. In addition, a method was developed for enrichment of five cytokines (TNF-α, IL-1β, IL-6, IL-10 and CCL2) from CSF for quantitative proteomic analysis. Conclusions: The results from these exploratory CSF analyses are expected to provide further insight into the biological responses to intraputamenal CDNF infusions in PD patients.

P521 / #1051

Topic: Theme C: α-Synucleinopathies / C3.c. Drug Development, Clinical Trials: Neurotransmitter- and receptor based modulators

RESULTS OF THE PRESENCE STUDY EVALUATING MEVIDALEN FOR THE TREATMENT OF LEWY BODY DEMENTIA (LBD)

Lecture Title:

K. Biglan1, L. Munsie1, K. Svensson2, P. Ardayfio3, M. Pugh4, J. Sims5, M. Mintun1 1Eli Lilly, Neuroscience Research, Indianapolis, United States of America, 2Eli Lilly, Neuroscience Discovery, Indianapolis, United States of America, 3Eli Lilly, Global Patient Safety, Indianapolis, United States of America, 4Eli Lilly, Statistics Neuroscience, Indianapolis, United States of America, 5Eli Lilly, Neurodegeneration, Indianapolis, United States of America

Aims: To assess the safety and efficacy of mevidalen, a D1 receptor positive allosteric modulator, for the treatment of cognition in patients with LBD. Methods: PRESENCE (NCT03305809) was a Phase 2, 12-week study in LBD participants with mild-to-moderate dementia randomized (1:1:1:1) to once-daily doses of mevidalen (10 mg, 30 mg, or 75 mg) or placebo. Key study eligibility requirements were age of 40-85 years, Hoehn and Yahr score of 0-4, and Montreal Cognitive Assessment (MoCA) score of 10 to 23. The primary outcome measure was change from baseline on the Cognitive Drug Research Continuity of Attention (CDR-CoA) composite score using a Bayesian mixed model repeated measures (MMRM) model. Success criterion was defined as at least a 67% probability of an effect size (ES)≥0.2. Secondary outcomes included Alzheimer’s Disease Assessment Scale cognitive subscale 13 (ADAS-cog13), Alzheimer’s Disease Cooperative Study – Clinical Global Impression of Change (ADCS-CGIC), Movement Disorder Society Unified Parkinson’s Disease Rating Scale (MDS-UPDRS), and Epworth Sleepiness Scale (ESS). Numerous safety measures were collected. Results: Primary and secondary outcomes measures and key safety outcomes will be presented. Conclusions: The PRESENCE study assessed the safety and efficacy of mevidalen as a potential symptomatic cognitive treatment in Lewy Body Dementia. Secondary measures across several domains potentially affected by mevidalen and relevant to LBD were also assessed including cognition, function, motor, and sleep.

P522 / #1062

Topic: Theme C: α-Synucleinopathies / C3.c. Drug Development, Clinical Trials: Neurotransmitter- and receptor based modulators

ISTRADEFYLLINE EFFECTS ON TREMOR DOMINANCE (TD) AND POSTURAL INSTABILITY AND GAIT DIFFICULTY (PIGD)

Lecture Title:

F. Pagan1, J. Parno2, R. Ristuccia3, A. Mori4, Y. Torres-Yaghi1 1MedStar Georgetown University Hospital, Department Of Neurology, Washington, United States of America, 2Kyowa Kirin Pharmaceutical Development, Inc., Biostatistics, Princeton, United States of America, 3Kyowa Kirin, Inc., Medical Communications, Bedminster, United States of America, 4Kyowa Kirin Co., Ltd., Medical Affairs, Tokyo, Japan

Aims: Istradefylline, a selective adenosine A2A receptor antagonist, is indicated in the USA and Japan as adjunctive treatment to levodopa/decarboxylase inhibitors in adults with Parkinson’s disease (PD) experiencing OFF time. Two distinct motor subtypes, tremor dominant (TD) and postural instability and gait difficulty (PIGD), can be differentiated using Unified Parkinson’s Disease Rating Scale (UPDRS) sub-scores. This post hoc analysis of pooled data from 6 pivotal studies examined the effect of istradefylline in these subtypes. Methods: In 6 randomized, placebo-controlled phase 2b/3 trials, patients on baseline anti-PD medications received istradefylline (20 or 40mg/day) or placebo for 12 or 16 weeks. TD subtype was defined by the UPDRS II/III items kinetic and postural tremor in right and left hand and tremor (at rest) of face/lips or chin/arms/legs; PIGD items were freezing, walking, posture, gait, and postural instability. The ratio of mean scores from TD:PIGD items determined subtype (TD [TD:PIGD ratio≥1.5], PIGD [TD:PIGD ratio≤1.0], mixed-type [ratio 1-1.5]). Results: 2284 patients were included (PIGD n=1848, TD n=101, mixed-type n=162, not evaluable n=173). Patients identified as PIGD subtype at baseline showed a statistically significant LS mean change from baseline (CFB) in UPDRS II/III PIGD total score for istradefylline 40mg vs placebo (Table) Similarly, patients with TD subtype at baseline showed a significant LS mean CFB in UPDRS II/III TD total score for both istradefylline doses vs placebo (Table). Conclusions: In this analysis of UPDRS-based motor subtypes, istradefylline significantly improved these difficult-to-treat symptoms, although further research is needed to characterize the effects of istradefylline on tremor. Supported by: Kyowa Kirin

P523 / #626

Topic: Theme C: α-Synucleinopathies / C3.d. Drug Development, Clinical Trials: Deep brain stimulation

LONG-TERM OUTCOME OF SUBTHALAMIC NUCLEUS DEEP BRAIN STIMULATION FOR PARKINSON’S DISEASE: 10 YEARS AND BEYOND

Lecture Title:

H.R. Park1, S.H. Paek2 1Soonchunhyang University Seoul Hospital, Neurosurgery, Seoul, Korea, Republic of, 2Seoul National University Hospital, Neurosurgery, Seoul, Korea, Republic of

Aims: Although there have been some reports of long-term efficacy and safety after deep brain stimulation (DBS) surgery for Parkinson’s disease (PD), little is known about the fate of all patients. We performed a follow-up of all patients who undergone bilateral subthalamic nucleus (STN) DBS surgery for PD before more than 10 years, and assessed the survival rate and long-term outcome of DBS. Methods: We included all 81 patients including 37 males and 44 females who underwent bilateral STN DBS from March 2005 to Mar 2008 at single institution. Whether or not the patients were alive or dead was investigated, and pre- and postoperative follow-up assessments including UPDRS score were analyzed. Results: The mean age at the time of surgery was 72 years (range, 27-82). Twenty-two patients (27.2%) died during the follow-up period. The median duration from the DBS surgery and to death of the 22 patients who died was 86 months (range, 0-155). Nine patients (40.9%) died because of disease progression, followed by 5 patients died due to pneumonia. The cumulative survival rate was as follows: 97.5 ± 1.7% (3 yr), 95.1 ± 2.4% (5 yr), 86.5% ± 3.8% (7 yr), and 80.2 ± 4.4% (10 yr). Conclusions: STN DBS is a safe and effective treatment in selective patients with PD. This study was based on the long-term follow-up of large-scale patients, and would contribute to elucidate the long-term fate of patients who underwent bilateral STN DBS for PD.

P524 / #1515

Topic: Theme C: α-Synucleinopathies / C3.g. Drug Development, Clinical Trials: Medicinal chemistry approaches, drug repurposing

IDENTIFICATION OF POTENTIAL NATURAL NEUROPROTECTIVE MOLECULES FOR PARKINSON’S DISEASE BY USING CHEMOINFORMATICS AND MOLECULAR DOCKING.

Lecture Title:

W.O. Contreras Lopez1, P. Rondon Villamizar2 1Clinica Foscal Internacional, Neurosurgery/translational Research, Floridablanca, Colombia, 2UIS, Biomol, Floridablanca, Colombia

Aims: To identify molecules that may lead to the design or discovery of new effective neuroprotective treatments for Parkinson’s disease. Methods: We used chemoinformatics tools and molecular docking simulations to analyze molecules that have been experimentally tested and bound to a-synuclein, causing neuroprotective or neurotoxic activity, and whose results have been used to select potential natural neuroprotective molecules. The first step in the selection of the molecules was based on the number of hydrogen bonds with the a-synuclein, obtained in the 50 binding poses. Due to potential neuroprotective molecules should also have Blood-Brain Barrier (BBB) permeability and good druglike properties, we used the free web tool SwissADME to obtain the predicted BBB permeation and drug-likeness of the candidate molecules. Results: In total, 156 molecules showed a number of hydrogen bonds with a-synuclein, higher than 80 in the 50 binding poses obtained in the molecular docking simulations. From this set of molecules, Six (06314, 69974, 17542, 04175, 16958, and 19925) had a positive BBB permeability and good druglike properties according to SwissADME tool. Conclusions: In this study, we found 6 natural molecules with a potential neuroprotective activity based on their similarity in the chemical structure with recognizedneuroprotective molecules and their theoretical binding with a- synuclein. The identification of potential neuroprotective compounds that bind themselves to a-synuclein may lead to the discovery of new therapeutics that prevents the a-synuclein oligomerization, and therefore, the development of PD.

P525 / #867

Topic: Theme C: α-Synucleinopathies / C3.g. Drug Development, Clinical Trials: Medicinal chemistry approaches, drug repurposing

REPURPOSING OF THE ANTI-ASTHMATIC DRUG MONTELUKAST FOR THE TREATMENT OF PARKINSON’S DISEASE

Lecture Title:

K. Strempfl1,2,3, M. Unger2,3, L. Aigner2,3, J. Neddens1, S. Flunkert1, B. Hutter-Paier1 1QPS Austria GmbH, Neuropharmacology, Grambach, Austria, 2Paracelsus Medical University, Institute Of Molecular Regenerative Medicine, Salzburg, Austria, 3Paracelsus Medical University, Spinal Cord Injury And Tissue Regeneration Center Salzburg (sci-trecs), Salzburg, Austria

Aims: The neurodegenerative disorder Parkinson’s disease (PD) affects over seven million people worldwide. Its major hallmark is an abnormal aggregation of the α-synuclein protein (α-syn) in neurons and/or glial cells of the substantia nigra and the deep brainstem, impairing movement. The complex pathomechanisms underlying α-syn aggregation and neurodegeneration are still not fully understood which limits available treatment options. Neuroinflammation, which is mainly driven by microglia and is also caused by a dysregulated leukotriene signaling pathway, might play a critical role in evoking neurodegenerative conditions. A previous study showed that blocking of leukotriene signaling via the leukotriene receptor antagonist Montelukast (MTK) improved learning and memory deficits in the α-syn transgenic D-Line mouse model. This study aims to evaluate whether MTK treatment can additionally alleviate motor symptoms in the α-syn transgenic Line 61 mouse model of PD. Methods: Male transgenic Line 61 mice and wild type littermates were daily treated with 10 mg/kg MTK or placebo orally from 2 weeks of age for 10 weeks. Motor tests were performed in the 2nd, 5th and 10th treatment week. Brains were collected for immunohistochemical and biochemical analyses of neuroinflammation markers and α-syn load. Results: Preliminary results show motor deficits already in 4-weeks old Line 61 mice as compared to age-matched wild type littermates. The beneficial effects of MTK on motor functions and mouse behavior as well as immunohistochemical stainings are currently under investigation. Conclusions: This study will provide insight whether Montelukast can be repurposed for the treatment of PD and related neurodegenerative diseases.

P526 / #883

Topic: Theme C: α-Synucleinopathies / C3.h. Drug Development, Clinical Trials: Drug delivery systems

SUBCUTANEOUS INFUDOPA (LEVODOPA/CARBIDOPA) INFUSION COULD BE SUITABLE MONOTHERAPY IN PATIENTS WITH ADVANCED PARKINSON’S DISEASE

Lecture Title:

N. Dizdar1, M. Ehrnebo2, L. Bring3, E. Eriksson4, F. Bergquist5 1Linkoping University, Department Of Biomedical And Clinical Sciences, Linkoping, Sweden, 2Pharm Assist AB, ., Uppsala, Sweden, 3Dizlin Pharmaceuticals AB, ., Vimmerby, Sweden, 4University of Gothenburg, Department Of Pharmacology, Gothenburg, Sweden, 5Sahlgrenska University Hospital, Neurology, Gothenburg, Sweden

Aims: The idea of continuous dopaminergic stimulation has led to new therapies for Parkinson’s disease. We performed a clinical trial on levodopa/carbidopa solutions for both intravenous and subcutaneous infusion (Infudopa). The primary objective was to demonstrate that both the infusions of Infudopa yield steady state plasma concentrations of levodopa that are equivalent with those of levodopa/carbidopa intestinal gel (Duodopa). Methods: In a prospective, open-label, randomized, multicentre, three-period crossover study, eighteen patients on Duodopa were randomized to the three infusion treatments, given for 16 hours. Their normal daily Duodopa dose was administered as constant infusion preceded by a standardized morning bolus. Estimated equivalent doses were given as intravenous and subcutaneous infusions. No oral levodopa medication was allowed. PK samples were taken for 24 hours. Results: All treatments reached mean levodopa concentrations of 2000 ng/ml simultaneously and they had comparable mean steady state levels of about 2500 ng/ml. Levodopa levels were more sustained after subcutaneous infusion. During the bolus dose mild infusion site pain of short duration occurred in 30% and 61% following i.v. and s.c. administration respectively. Conclusions: Bioequivalence to Duodopa treatment was shown for subcutaneous and intravenous administration, with equal early and steady-state levodopa plasma concentrations. Slower elimination of levodopa after subcutaneous infusion may give better night time sleep for the patients reducing complementary oral medication. Subcutaneous Infudopa infusion is a well-tolerated and clinically efficient treatment that is feasible as monotherapy also for patients requiring high doses, easy to perform, and requries no surgical intervention.

P527 / #1187

Topic: Theme C: α-Synucleinopathies / C3.h. Drug Development, Clinical Trials: Drug delivery systems

FIRST RESULTS FROM AN OPEN-LABEL STUDY EVALUATING LONG-TERM SAFETY OF ND0612 IN PD PATIENTS EXPERIENCING MOTOR FLUCTUATIONS

Lecture Title:

W. Poewe1, F. Stocchi2, N. Giladi3, L. Adar4, O. Rosenfeld4, L. Salin4, C.W. Olanow5 1Innsbruck Medical University, Department Of Neurology, Innsbruck, Austria, 2IRCCS San Raffaele Pisana, Institute Of Neurology, Rome, Italy, 3Tel Aviv Sourasky Medical Center, Neurology, Tel Aviv, Israel, 4NeuroDerm, Clinical Development, Rehovot, Israel, 5Clintrex, LLC, ., Sarasota, United States of America

Aims: ND0612 is a continuous, subcutaneous (SC) delivery system of levodopa/carbidopa that provides therapeutic plasma levodopa levels with less variability than oral levodopa. This phase 2b, international, open-label study assessed the long-term safety (systemic and local) and tolerability of continuous infusion with two regimens of ND0612 (24-hour delivery and 16-hour delivery + morning oral LD dose). Here we present 1 year data. Methods: This study (NCT02726386) was conducted in PD patients (aged ≥ 30y, H&Y ≤3 during ON), taking ≥4 levodopa doses/day and ≥1 other PD medications, and experiencing ≥2 hours of OFF time/day with predictable early-morning OFF periods. Patients were assigned to receive ND0612 for a regimen of either 16-hours/day or 24-hours/day. Results: 214 patients were enrolled (24-hour regimen: n=90; 16-hour regimen: n=124) at 46 sites in 8 countries. Over the course of the one-year treatment period, 67% of patients experienced treatment-related TEAE’s with 5.6% of subjects experiencing serious treatment-related TEAE’s. Systemic safety was typical for PD patients treated with LD/CD. The most frequent AEs were local infusion site reactions (57.9% patients) which are typical for a continuous subcutaneous mode of drug administration. Overall, 17.8% patients discontinued due to AEs. Conclusions: ND0612 infusion was found to be safe with generally mild to moderate local AEs which were reversible and manageable. No unanticipated systemic TEAEs for this patient population on levodopa treatment occurred. Long-term data will continue to be collected in patients enrolled in the study extension, some of whom are now in their fifth year of ND0612 treatment.

P528 / #1081

Topic: Theme C: α-Synucleinopathies / C3.i. Drug Development, Clinical Trials: Non-pharmacological interventions, neurosurgery

THE IMPACT OF COGNITIVE INTERVENTION ON COGNITION AND QUALITY OF LIFE IN IDIOPATHIC PARKINSON’S DISEASE: A RANDOMIZED AND CONTROLLED STUDY

Lecture Title:

N. Sousa1, A. Neri2, S. Brucki3 1University of Sao Paulo, Neurology, Sao Paulo, Brazil, 2SARAH Network of Rehabilitation Hospitals, Neurological Reahabilitation, Salvador, Brazil, 3University os Sao Paulo, Neurology, Sao Paulo, Brazil

Aims: To investigate the effects of cognitive training (CT) on cognitive and quality of life measures in PD-MCI patients. Methods: A total of 39 participants was randomly to 2 groups: control (CG=15) and experimental (CTG=24) according to UK PD Brain Bank criteria. The inclusion criteria: (i) an age over 50 years, (ii) Hoehn and Yahr stage (H&Y) until III stage; (iii) Beck Depression Inventory ≤16. The treatment group received an CT program, and the CG received speech therapy sessions and physical exercise. Addenbrooke’s Cognitive Examination (ACE-III), Digit Span, Trail Making Test (TMT-A and B) and Parkinson's Disease Questionnaire (PDQ-39) were used in each patient at baseline (T0) and final (T1). Descriptive statistic, and t test for independent samples and paired-sample t test to evaluate the effect of group (CTG and CG) and time (T0 and T1), respectively. Results: The CTG had an average of 60 (7.5) years of age, schooling of 12.4 (3.1) years, 5.7 (3.3) years of disease evolution and 87.5% were in stage I-II on the H&Y. The CG had 58.5 (9.8) of age, schooling 12.8 (3.4), 6.8 (8.8) duration of disease and 93.3% in stage I-II H&Y. There was a statistically significant in the total score, attention/orientation, memory, verbal fluency and visuospatial of the ACE-III, TMT-B and TMT-A. Regarding PDQ values, there was an improvement in the total score, mobility, activities of daily living and body discomfort in the CTG. Conclusions: Our findings suggest that intensive CT may an effective tool for improving cognitive functions and quality of life in patients.

P529 / #1596

Topic: Theme C: α-Synucleinopathies / C3.k. Drug Development, Clinical Trials: Personalized medicines

POLYMORPHISM OF CATECHOL-O-METHYLTRANSFERASE CHANGES PLASMA CONCENTRATION OF L-DOPA AND 3-O-METHYLDOPA

Lecture Title:

J. Yamamoto1, T. Omura2, S. Kasamo3, S. Yamamoto4, M. Kawata4, A. Yonezawa4, Y. Taruno5, H. Endo6, H. Aizawa7, N. Sawamoto5, K. Matsubara8, R. Takahashi5, Y. Tasaki1 1Asahikawa Medical University, Department Of Hospital Pharmacy & Pharmacology, asahikawa, Japan, 2Kobe University Hospital, Department Of Pharmacy, Kobe, Japan, 3Asahikawa Medical University, Institutional Research Office, Asahikawa, Japan, 4Kyoto University Hospital, Department Of Clinical Pharmacology And Therapeutics, Kyoto, Japan, 5Graduate School of Medicine, Kyoto University, Department Of Neurology, Kyoto, Japan, 6Asahikawa Medical University, Division Of Neurology, Department Of Internal Medicine, Asahikawa, Japan, 7Tokyo Medical University, Department Of Neurology, Tokyo, Japan, 8Wakayama Medical University, Department Of Pharmacy, Wakayama, Japan

Aims: In Parkinson’s disease (PD), inter-individual variations in the effectiveness of entacapone on the pharmacokinetics of L-dopa exist between patients with high-activity (H/H) and low-activity (L/L) catechol-O-methyltransferase (COMT) Val158Met polymorphisms while its effects in the heterozygous (H/L) patients are unclear. Moreover, the elevated levels of 3-O-methyldopa(3-OMD) have been associated with a poor response to L-dopa treatment; however, the significance of this polymorphism on the 3-OMD production remains unknown. Therefore, we aimed to evaluate the effect of the COMT Val158Met polymorphism on the plasma level of L-dopa and 3-OMD in the presence of entacapone. Methods: This was a single-sequence, single-period, open-label, crossover study at two hospitals in Japan. Participants were 54 Japanese patients with PD, who underwent an acute L-dopa administration test with and without 100 mg entacapone on two separate days. Results: Entacapone increased the L-dopa area under the concentration curve (AUC) 0-infinity by 1.59 ± 0.26-fold in the H/H group, which was significantly higher than that in the H/L and L/L groups (p<0.05). There was no difference in 3-OMD AUC0-4h after L-dopa dosing without entacapone among three groups; however, 3-OMD AUC0-4h / L-dopa daily dose and ΔC3-OMD AUC0-4h tended to be higher in the H/H group than those of the other two groups. The co-administration of L-dopa and entacapone suppressed the increase in 3-OMD levels compared with L-dopa alone in all genotypes. Conclusions: Based on our findings, COMT Val158Met polymorphism is a potential biomarker for individualized dose selection of COMT inhibitors in PD treatment.

P530 / #823

Topic: Theme C: α-Synucleinopathies / C4.b. Imaging, Biomarkers, Diagnostics: Functional MRI

NEUROIMAGING-GENETIC ASSOCIATIONS IN PARKINSON’S DISEASE STAGING VIA STEPWISE FUNCTIONAL CONNECTIVITY

Lecture Title:

S. Basaia1, F. Agosta2, I. Diez3, E. Bueichekú3, F. D’Oleire Uquillas4, M. Delgado-Alvarado5, C. Caballero-Gaudes6, M. Rodriguez-Oroz7, T. Stojkovic8, V. S. Kostic8, M. Filippi9, J. Sepulcre3 1IRCCS San Raffaele Scientific Institute, Division Of Neuroscience, Milano, Italy, 2IRCCS San Raffaele Scientific Institute and Vita-Salute San Raffaele University, Division Of Neuroscience, Milano, Italy, 3Gordon Center for Medical Imaging, Department Of Radiology, Massachusetts General Hospital, Boston, United States of America, 4Massachusetts General Hospital, Harvard Medical School, Department Of Neurology, Boston, United States of America, 5Sierrallana Hospital, Neurology Department, Torrelavega, Spain, 6Basque Center on Cognition, Brain and Language, Basque Center On Cognition, Brain And Language, Donostia, Spain, 7Clínica Universidad de Navarra, Neurology Department, Navarra, Spain, 8University of Belgrade, Clinic Of Neurology, Belgrade, Serbia, 9IRCCS San Raffaele Scientific Institute; Vita-Salute San Raffaele University, Division Of Neuroscience; Unit Of Neurology, Milano, Italy

Aims: The aim of this study was to investigate the topological similarity between large-scale connectivity networks –in healthy controls (HC)– and gene expression patterns of SNCA gene across the human cortex. Methods: 60 HC performed clinical and cognitive evaluations and resting state functional MRI. A graph theory metric based on Stepwise Functional Connectivity (SFC) analysis was applied in order to map connectivity patterns of medulla (known as to be a pathological epicenter in PD). The genetic intersection of SFC patterns and genetic transcription profiles was built using the Allen Human Brain Atlas. We computed the cortical spatial similarity among PD-related genes’ expression and SFC maps in HC. Next, we investigated whether the data-driven genetic profiles displayed specific biological functionality. Using postmortem PD patients’ data, the association between alpha-synuclein histopathology density scores in 10 brain regions of PD patients and the SFC maps in HC was evaluated. Results: SNCA gene expression is associated with SFC patterns in HC (r=-0.29; p=0.006). Gene expression levels of 784 genes (including SNCA) were distributed along the cortical mantle similarly as SFC maps in HC. These genes displayed also an association with the regulation of dopamine secretion. Furthermore, the analysis of spatial similarity among SFC and alpha-synuclein histopathology density scores demonstrated that regions closer to the epicenter present higher density of alpha-synuclein. Conclusions: This study may offer novel avenues for enhancing PD detectability via the evaluation of in vivo trajectories of connectivity changes in the human brain based on neuroimaging-genetic integration strategies. Funding: Ministry of Education and Science Republic of Serbia(Grant#175090).

P531 / #868

Topic: Theme C: α-Synucleinopathies / C4.c. Imaging, Biomarkers, Diagnostics: PET

[18F]ACI-12589, A NOVEL ALPHA-SYNUCLEIN RADIOTRACER AS A BIOMARKER IN PATIENTS WITH PARKINSON’S DISEASE AND OTHER SYNUCLEINOPATHIES

Lecture Title:

F. Capotosti, E. Vokali, J. Molette, M. Ravache, C. Delgado, J. Kocher, K. Piorkowska, M. Chauhan, T. Touilloux, H. Kroth, R. Luthi-Carter, O. Sol, A. Pfeifer, M. Kosco-Vilbois AC Immune, Research, Lausanne, Switzerland

Aims: Synucleinopathies are characterized by progressive accumulation of alpha-synuclein (a-syn) inclusions in the brain which correlates with disease severity. Interventional studies targeting a-syn would strongly benefit from biomarkers that support accurate diagnosis and following disease progression. Due to co-pathologies and low density of a-syn pathology, an a-syn PET tracer must display strong selectivity, high binding affinity and minimal non-specific binding. Methods: Initial screening efforts of our proprietary MorphomerTM library were followed by an iterative medicinal chemistry optimization program using a-syn aggregates derived from human diseased brain samples. Autoradiography and radiobinding experiments on synucleinopathy-containing tissue samples were used to evaluate affinity and target occupancy. Those techniques were then employed to assess selectivity using tissue samples from Alzheimer’s disease or Frontotemporal lobar degeneration-TDP donors. Selected compounds were then 18F radiolabeled and evaluated for their pharmacokinetic (PK) profiles in non-human primates (NHPs). Results: Recent optimization efforts have led to the identification of several new, highly promising chemical scaffolds. Particularly, the compound, ACI-12589, demonstrates excellent target occupancy and signal specificity on tissue samples from different synucleinopathies including Parkinson’s disease, multiple system atrophy and dementia with Lewy bodies. The compound demonstrates selectivity versus Abeta, Tau and TDP-43 aggregates. Studying [18F]ACI-12589 in NHPs reveals that this compound has a desirable brain-PET ligand PK profile. Conclusions: [18F]ACI-12589 has been identified as a novel clinical candidate displaying the characteristics deemed necessary for a successful radiotracer for a-syn inclusions. [18F]ACI-12589 is intended to soon enter a FiH study in patients with Parkinson’s disease followed by evaluation in other synucleinopathies.

P532 / #1193


PERFORMANCE OF THE FASTER REGION-BASED CONVOLUTIONAL NEURAL NETWORK-BASED 18F-FP-CIT SCORING ALGORITHM FOR DIFFERENTIATING BETWEEN PATTERN OF NORMAL AND PARKINSON’S DISEASE

Lecture Title:

B.W. Choi1, S. Kang1, H. Vu2, S.W. Youn2, H.W. Kim3 1Daegu Catholic University Medical Center/Catholic University of Daegu School of Medicine, Nuclear Medicine, Daegu, Korea, Republic of, 2Daegu Catholic University Medical Center/Catholic University of Daegu School of Medicine, Radiology, Daegu, Korea, Republic of, 3Keimyung University Dongsan Hospital/Keimyung University School of Medicine, Nuclear Medicine, Daegu, Korea, Republic of

Aims: The aim of this study was to evaluate the performance of faster Region-based Convolutional neural network (fR-CNN) for differential diagnosis between Parkinson's disease (PD) and secondary parkinsonism based on maximum intensity projection (MIP) image of 18F-FP-CIT PET. Methods: A total of 139 images of PD pattern and 388 images of non-PD pattern were used for analysis. Non-PD pattern images were further classified into 23 vascular parkinsonism with prominently decreased dopamine transporter binding (dVP) pattern and 365 normal pattern images. Diagnostic performance of the fR-CNN and three NM physicians was assessed among three clinically relevant pairs of image pattern groups (PD/non-PD, PD/dVP, and PD/normal). Diagnostic performance was assessed using receiver operator characteristic (ROC) curve analysis. And the difference of diagnostic performance and inter-rater agreement were accessed between fR-CNN and three NM physicians. Results: fR-CNN achieved a range of 94.7−100% classification sensitivity, 71.4−100% specificity, and 92.3−98.6% accuracy. Those of the three NM physicians were 89.5%, 100%, and 92.3−97.2%; 100%, 95.6−100%, and 96.9−100%; and 94.7%, 85.7−97.8%, and 92.3−96.9%, respectively. There were no significant differences in area under the curve values between the fR-CNN and three NM physicians. There was no significant difference in concordance assessment using Cochran's Q test, and the overall inter-rater agreement among fR-CNN and NM physicians evaluated by Fleiss k coefficient showed almost perfect to substantial agreements. Conclusions: In differentiating between PD and non-PD image patterns using MIP image of 18F-FP-CIT PET, the performance of the fR-CNN was comparable to that of three NM physicians.

P533 / #1510


EARLY DETECTION OF DLB: DECREASED UPTAKE IN 123I-MIBG SCINTIGRAPHY AND 18F FP-CIT-PET IMAGING IN PRODROMAL DLB

Lecture Title:

M.Y. Park1, G. Lee2 1Yeungnam University Medical Center, Neurology, Daegu, Korea, Republic of, 2Danguk university medical hospital, Neurology, Cheonan, Korea, Republic of

Aims: Because of the time course of detecting DLB symptoms and signs, DLB is poorly diagnosed and hardly differentiate from AD, especially in early stage of dementia without the core clinical features of DLB. We investigated patients with a clinical diagnosis of amnestic mild cognitive impairment (MCI) and early AD whether they had cardiac sympathetic denervation, detected by cardiac 123I-MIBG scintigraphy. And we also assessed presynaptic nigrostriatal dopaminergic system by 18F FP-CIT-PET imaging to distinguish between DLB and AD. Methods: Twenty consecutive patients with a clinical diagnosis of amnestic MCI (n=8) and early AD (n=12, CDR 0.5/SOB 2.5) who have been had visual hallucination and/or suspicious fluctuating cognition history without parkinsonism were enrolled in this study; patients were evaluated neuropsychologic test. Metaiodobenzylguanidine (MIBG) uptake was assessed using the ratio of the heart to the upper mediastinum (H/M ratio), and we also assessed presynaptic nigrostriatal dopaminergic system by 18F FP-CIT-PET imaging to distinguish between DLB and AD. Autonomic function tests and orthostatic vital signs were recorded. Results: The mean H/M ratio was significantly lower (1.6±0.18) and presynaptic nigrostriatal dopaminergic deficit were founded in all enrolled patients. Orthostatic hypotension was evident in all patients regardless of spontaneous Parkinsonism. Conclusions: Myocardial postganglionic sympathetic denervation and autonomic dysfunctions including orthostatic hypotension can be good biomarkers to predict DLB before core clinical symptoms appear, and may useful to distinguish from AD even in early stage.

P534 / #1382

Topic: Theme C: α-Synucleinopathies / C4.e. Imaging, Biomarkers, Diagnostics: Multimodal imaging

PROGRESSION OF CORTICAL THINNING IN PARKINSON’S DISEASE

Lecture Title:

A. Vo, C. Tremblay, S. Rahayel, Y. Yau, A. Dagher Montreal Neurological Institute and Hospital, McGill University, Department Of Neurology And Neurosurgery, Montreal, Canada

Aims: We sought to validate cortical thickness as an objective biomarker of Parkinson’s disease (PD) progression. Cortical thickness analysis has been proposed as a more sensitive measure of age- and disease-related changes in cortical integrity than voxel-based morphometry. The application of this technique in PD progression has yielded inconsistent findings, however. This discrepancy is explained by studies with small patient samples and cross-sectional designs that fail to account for disease heterogeneity. Methods: In the present study, we investigated the trajectory of cortical thinning in a large, longitudinal, and multicentre cohort of PD patients. T1-weighted MRI scans of de novo PD patients from the Parkinson Progression Marker Initiative (PPMI) were acquired at baseline, 1-, 2-, and 4-year follow-up. For each scan, we computed a W-score map to control for age-related changes. We modeled longitudinal changes in cortical thickness across time points, covaried with age and sex. Results: One-year post-diagnosis, early regional thinning was observed in the left inferior frontal lobe, left temporal pole, and right somatomotor cortex. By 2-year follow-up, cortical atrophy had become relatively widespread. Partial correlations showed a negative relationship between atrophy in a frontal cluster and baseline measure of CSF beta-amyloid and phosphorylated tau. Conclusions: PD patients showed early regional thinning in frontal and parietal areas, replicating earlier findings. This cortical atrophy continued to rapidly spread before appearing to level off in a non-linear trajectory. Our results demonstrate the use of cortical atrophy as a potential biomarker for PD and PD progression.

P535 / #1344

Topic: Theme C: α-Synucleinopathies / C4.f. Imaging, Biomarkers, Diagnostics: CSF, blood, body fluid biomarkers

APTAMER-BASED BIOMARKER DISCOVERY IN PERIPHERAL PLASMA IDENTIFIES NEW CANDIDATES FOR EARLY PARKINSON’S DISEASE

Lecture Title:

M. Bartl1, I. Yahya Abdi2, M. Dakna1, C. Trenkwalder3,4, O. El-Agnaf2, B. Mollenhauer4 1University Medical Center Göttingen, Dept. Of Neurology, Göttingen, Germany, 2Qatar Biomedical Research Institute, Neurological Disorders Research Center, Doha, Qatar, 3University Medical Center Goettingen, Department Of Neurosurgery, Göttingen, Germany, 4Paracelsus-Elena-Klinik, Paracelsus-elena-klinik, Kassel, Germany

Aims: To identify blood-based biomarkers and enable pathway analyses in Parkinson’s disease (PD) to elucidate underlying pathophysiology, optimize early diagnosis and evaluate potential predictive markers for different progression-/ phenotypes. Methods: Aptamer technology was applied in plasma samples from 85 recently diagnosed, drug-naïve PD patients and 93 healthy controls (HC) from de novo Parkinson’s (DeNoPa) cohort at baseline. Longitudinal follow-up of clinical symptoms was carried out biannually and finalized for the 8th year after diagnosis. Protein levels were quantified using Somalogic modified based approach (SOMAscan platform, 1.3K assay). Results: After multiple testing corrections out of 1137 targeted proteins, 886 passed quality check and 150 passed a false discovery rate (FDR) threshold of 0.05, including markers of immune response, metabolism, extracellular matrix and endothelial function. Clustering and pathway analysis showed decreased cardiovascular markers (CM) here (e.g. uPA, PCSK9, CK-MB), contrasting previous findings, that CM are relevant in PD progression (Mollenhauer et al. 2019). In congruency to other reports, the analysis revealed pro- (e.g. Calcineurin) and anti-(e.g. ILT4) inflammatory markers (IM). Conclusions: Since hypothesis driven, mainly antibody-based analyses in PD was of limited success, newly developed (antibody-independent) unbiased multiplex approaches are now applied. With increasing evidence for peripheral pathology in PD, peripheral matrices like blood become more important as early diagnostic and possibly also progression marker. IM and CM seem to play a role in PD progression and pathophysiology, which is not fully elucidated yet. To evaluate predictive potential and possible therapeutic targets, further investigation of longitudinal data is needed.

P536 / #1122


DIFFERENTIAL DIAGNOSTIC VALUE OF TOTAL ALPHA-SYNUCLEIN ASSAY IN THE CEREBROSPINAL FLUID BETWEEN ALZHEIMER'S DISEASE AND DEMENTIA WITH LEWY BODIES FROM THE PRODROMAL STAGE

Lecture Title:

O. Bousiges1, N. Philippi1,2, T. Lavaux3, A. Perret-Liaudet4, I. Lachmann5, C. Schaeffer-Agalède3, P. Anthony2, A. Botzung6, L. Rauch2, B. Jung6, P. Loureiro De Sousa1, C. Demuynck2, C. Martin-Hunyadi6, B. Cretin2, F. Blanc1 1iCube, Imis Team - Institut De Physique Biologique (ipb), Strasbourg, France, 2University Hospital of Strasbourg, Geriatrics And Neurology Services, Research And Resources Memory Center (cmrr), Strasbourg, France, 3Hôpitaux Universitaires de Strasbourg, Biochemistry Laboratory, Strasbourg, France, 4Hospices Civils de Lyon, Biochemistry And Molecular Biology, Bron Cedex, France, 5AJ Roboscreen GmbH, Roboscreen, Leipzig, Germany, 6Hôpitaux Universitaires de Strasbourg, Geriatrics And Neurology Units, Research And Resources Memory Center (cmrr), Strasbourg, France

Aims: To determine the value of alpha-synuclein assay in the cerebrospinal fluid (CSF) of Alzheimer’s disease (AD) and dementia with Lewy bodies (DLB) patients at the MCI stage. Methods: All patients were enrolled under a hospital clinical research protocol by an experienced team of clinicians. A total of 166 patients were included in this study: 21 control subjects (CS), 51 patients with DLB at the prodromal stage (pro-DLB), 16 patients with DLB at the demented stage (DLB-d), 33 AD patients at the prodromal stage (pro-AD), 32 AD patients at the demented stage (AD-d) and 13 patients with mixed pathology (AD+DLB). CSF levels of total alpha-synuclein were assessed using a commercial enzyme-linked immunosorbent assay (ELISA) for alpha-synuclein (AJ Roboscreen). Alzheimer’s biomarkers (t-Tau, P-Tau, Aβ42 and Aβ40) were also measured. Results: The alpha-synuclein assays showed a significant difference between the AD and DLB groups. Total alpha-synuclein levels were significantly higher in AD patients than in DLB patients. However, the ROC curves show a moderate discriminating power between AD and DLB (AUC = 0.78) which does not improve the discriminating power of the combination of Alzheimer biomarkers (AUC = 0.95 with or without alpha-synuclein). Interestingly, the levels appeared to be altered from the prodromal stage in both AD and DLB. Conclusions: The modification of total alpha-synuclein levels in the CSF of patients occurs early, from the prodromal stage. The adding of alpha-synuclein total to the combination of Alzheimer’s biomarker does not improve the differential diagnosis between AD and DLB.

P537 / #424


ANALYSIS OF SMALL RNAS AND PROTEINS FROM CEREBROSPINAL FLUID REVEALS A NOVEL MIRNA SIGNATURE FOR PARKINSON’S DISEASE AND IDENTIFIES DISEASE-RELEVANT PATHWAYS

Lecture Title:

L. Caldi Gomes1, A.-E. Roser2, G. Jain3, T. Pena-Centeño4, F. Maass2, L. Schilde5, M. Bähr2, K. Marcus5, A. Fischer4, P. Lingor1 1Rechts der Isar Hospital - Technical University of Munich, Department Of Neurology, Munich, Germany, 2University Medical Center Göttingen, Neurology, Göttingen, Germany, 3Technical University of Munich, Translatum, Munich, Germany, 4German Center for Neurodegenerative Diseases, Department For Epigenetics And Systems Medicine In Neurodegenerative Diseases, Göttingen, Germany, 5Ruhr-Universität Bochum, Medical Proteom-center, Bochum, Germany

Aims: Identification of differentially expressed (DE) miRNAs and proteins in cerebrospinal fluid (CSF) samples of patients with Parkinson’s disease (PD) and age-correlated control patients using small-RNA-sequencing/mass spectrometry, aiming for a miRNA-biomarker signature that differentiates PD patients and controls, and to reveal disease-relevant pathways. Methods: CSF samples were collected from a total of 76 PD patients and 79 age-correlated control subjects. RNA-sequencing was performed in a discovery cohort (42 PD/43 Ctr) on Illumina HiSeq4000. Signature miRNAs were identified using Measure of Relevance/reliability analysis, followed by machine learning-based variable ranking for further dimensionality reduction. The performance of the identified miRNAs was evaluated in a validation cohort (9 PD/11 Ctr), using the random forest algorithm in R, and in a second validation cohort (25 PD/25 Ctr) by qRT-PCR . The performance of the model was estimated by receiver operating characteristic (ROC) curve analysis. Furthermore, mass spectrometry analyses were conducted on a subgroup of subjects (23 PD/35 Ctr) using a Q-Exactive mass spectrometer in data-independent acquisition mode to obtain the proteome composition. Results: DE analysis of miRNAs from CSF identified miR-126-5p, miR-99a-5p, and miR-501-3p as a novel miRNA signature for the diagnosis of PD, able to classify PD and Ctr subjects with an AUC of 0.85. Pathway-enrichment analysis of target genes of DE miRNA and DE proteins identified biological processes involved in neuronal growth, synaptic function and immune response. Conclusions: The identification of three signature miRNAs widens the range of potential CSF biomarkers for PD. Pathway analysis identified biological processes that may represent novel therapeutic targets in PD.

P538 / #1227


DETECTION OF DIAGNOSTIC RELEVANT BIOMARKER PROTEINS IN PLASMA AND CSF OF ALPHA SYNUCLEINOPATHIES BY SINGLE MOLECULE ARRAY (SIMOA)

Lecture Title:

S. Canaslan1,2, M. Schmitz1,2, I. Zerr2 1Universitätsmedizin Göttingen, Neurology, Göttingen, Germany, 2DZNE, German Center For Neurodegenerative Diseases, Göttingen, Germany

Aims: Synucleinopathies, such as Parkinson’s disease, dementia with Lewy bodies, and multiple system atrophy, are a class of neurodegenerative diseases and characterized by the presence of alpha-synuclein aggregates in brain tissue. Differential diagnosis of alpha-synucleinopathies is challenging because of the clinical overlap of clinical symptoms and a lack of an accurate diagnostic test system. Our aim is to analyze the levels of different marker proteins’ in cerebrospinal fluid (CSF) and blood in synucleinopathies. Methods: CSF and blood are valuable resources for biomarker research. A small change in the protein concentration can be specific for a certain diagnosis. To detect these changes, we have established the ultrasensitive Single Molecule Array (SIMOA) method which is able to detect protein concentration in femtomolar amount instead of colorimetric ELISAs which are less sensitive. The underlying reason for its ultra-sensitivity is that target proteins are grabbed with capture antibody-coated microscopic magnetic beads and the machine has the ability to detect every single signal coming from the bead-antibody-target complex. Results: We have analyzed the level of alpha-synuclein, Neurofilament-light, Glial Fibrillary Acidic Protein in plasma and CSF from patients with different types of alpha synucleinopathies in comparison to healthy individuals. We observed a subtype-specific regulation of certain markers which may be helpful to distinguish between different types of alpha synucleinopathies. Conclusions: The investigation of biomarkers is important for an accurate diagnosis. We identified regulation of these proteins which might be useful not only for the diagnostic of an alpha synucleinopathies but also to distinguish between different subtypes.

P539 / #1481


BEYOND THE BRAIN: PERIPHERAL MICRORNA EXPRESSION AND PARKINSON’S DISEASE

Lecture Title:

M. Cressatti1, W. Song1, L. Juwara2, J. Galindez1, N. Orlovetskie1, A. Velly3, M. Gornitsky3, H. Schipper1 1McGill University/Lady Davis Institute, Department Of Neurology And Neurosurgery, Montreal, Canada, 2McGill University, Quantitative Life Sciences, Montreal, Canada, 3Jewish General Hospital, Department Of Dentistry, Montreal, Canada

Aims: Dysregulated microRNAs (miRNAs) play a major role in normal aging and various neurodegenerative conditions, including Parkinson’s disease (PD). We previously showed that altered expression profiles of salient miRNAs and their mRNA targets contribute to neural damage accruing from the overexpression of glial heme oxygenase-1 (HO-1). HO-1 is increased in PD brain and has been implicated in its pathogenesis. Objective: To assess the diagnostic potential of miR-153 and miR-223 in mice and humans with parkinsonism, and elucidate the mechanism that links peripheral and neurological pathologies in neurodegenerative disease. Methods: Parkinsonian transgenic (TG) GFAP.HMOX1 mice, engineered to overexpress the human HO-1 gene (HMOX1) in astrocytes between 8.5 and 19 months of age, were evaluated. MiRNA levels were measured by RT-qPCR in mice as well as human saliva. Peripheral extracellular vesicle (EV) content in mice and humans was assayed using polymer- and immunoaffinity-based isolation methods, followed by RT-qPCR, Western blot and ELISA. Results: Downstream of HO-1 overexpression, miR-153 and miR-223 (regulators of α-synuclein) were significantly downregulated in TG basal ganglia and serum compared to wild-type controls. Moreover, miR-153 and miR-223 were similarly decreased in the saliva of human PD subjects (n = 83) compared to healthy controls (n = 77). Intriguingly, EVs from GFAP.HMOX1 mouse serum and human saliva exhibit similar patterns of HO-1, miR-153 and miR-223 expression. Conclusions: HO-1-mediated (and potentially EV HO-1-mediated) perturbations in brain and peripheral miRNA expression profiles may drive PD neuropathology and serve as diagnostic markers for this condition.

P540 / #417


EVALUATING SERUM NEUROFILAMENT LIGHT AS A MEASURE OF TREATMENT RESPONSE IN STANDARD SIZED PH2 PARKINSON’S DISEASE TRIALS

Lecture Title:

K. Fraser1, W. Wang2, F. Gao2, T.-Y. Liu2, D. Graham1 1Biogen, Biomarkers, Cambridge, United States of America, 2Biogen, Biostatistics, Cambridge, United States of America

Aims: Recent analysis from the PPMI cohort demonstrated a higher annual rate of increase in serum Neurofilament Light (NfL) levels in idiopathic Parkinson’s disease (PD) compared with healthy controls (HC). This suggests that serum NfL levels could be utilized to monitor treatment effects in Ph1/Ph2 trials and inform which drugs merit development in registrational studies. Leveraging the PPMI dataset we sought to model a typical sized Ph2 trial to assess the feasibility and power of serum NfL as a treatment-response biomarker. Methods: 90 subjects each were randomly selected with replacement from the PPMI PD cohort for inclusion in placebo arm and treatment arm simulations. NfL progression rates were simulated from baseline and month 12 serum NfL data with modeling considerations for various true treatment effect sizes and progression rate assumptions. Linear model was fitted to estimate the treatment effect on NfL by adjusting for age and gender. 1000 simulations per treatment condition were evaluated and data was summarized. Results: Assuming the true underlying treatment effect is a full restoration as defined by shifting the annual rate in serum NfL increase from PD to HC levels, all treatment conditions and assumptions had a less than 50% probability of detecting a treatment response with this measure. This observation was consistent whether using stringent (p<0.05) or more relaxed (p<0.2) statistical definitions. Conclusions: The magnitude of the annual rate of change in serum NfL levels in PD is not sufficiently powered to observe a treatment effect in an N=90 per arm 1-year Ph2 PD study.

P541 / #624


DEVELOPMENT OF SANDWICH ELISA FOR MEASUREMENT OF OLIGOMERIC A-SYNUCLEIN IN BIOLOGICAL SAMPLES

Lecture Title:

W.-C. Liao, P. Hsueh, L. Wang, S. Zou, J. Ni BioLegend, Immunoassay, San Diego, United States of America

Aims: Measurement of ‘total α -synuclein’ has not yielded a robust biomarker to diagnose or monitor progression of Parkinson’s disease (PD). We hypothesize that oligomeric human α -synuclein protein is a more robust biomarker for PD, as it is the true toxic species inducing neuronal toxicity and cell death in PD. Our goal is to develop a sensitive and specific sandwich ELISA for measuring human α-synuclein oligomers in biological fluids, such as cerebrospinal fluid (CSF), to aid diagnosis of PD and monitoring disease progression. Methods: Panels of oligomer-specific α-synuclein monoclonal antibodies were developed at BioLegend. Specificity of these clones were initially evaluated by dot blots using monomeric and aggregated recombinant human a-synuclein protein and by immunohistochemistry using samples derived from Drosophila expressing a-synuclein vs non-expressing controls. Subsequently, we screened these oligomer-specific α-synuclein monoclonal antibodies in combination with total a-synuclein antibodies to identify an antibody pair that specifically detects α -synuclein oligomers in the sandwich ELISA with pg/mL ranges of sensitivity. Results: Firstly, we established an ELISA platform that is specific to oligomeric α-synuclein. We further examined our assay using biological samples. We used lysates derived from Drosophila model of human α -synucleinopathy to demonstrate assay specificity and sensitivity. In addition, we observed that expression of α-synuclein oligomers in PD brain lysates is higher than normal control brain lysates. We also observed promising results in spike linearity and recovery experiments using CSF samples. Conclusions: In summary, our sandwich ELISA has satisfied several key parameters for an immunoassay, including specificity, sensitivity, dilutional linearity, and spike recovery.

P542 / #1177


PROFILING OF LIPOPROTEINS AND ASSOCIATED LIPIDS IN SERUM SAMPLES FROM LRRK2-MUTATION CARRIERS

Lecture Title:

A. Laguna1, H. Xicoy1, D. Vilas2, A. Garrido3, A. Sánchez4, M. Fernández3, N. Amigó5, O. Yanes6, J. Infante4, M.J. Marti7, E. Tolosa8, M. Vila1 1Vall d’Hebron Research Institute (VHIR)–Center for Networked Biomedical Research on Neurodegenerative Diseases (CIBERNED), Neurodegenerative Diseases Research Group, Barcelona, Spain, 2Research Institute Germans Trias i Pujol, Neurology, Badalona, Spain, 3Hospital Clinic de Barcelona, Parkinson Disease And Movement Disorders Unit, Neurology Service,, Barcelona, Spain, 4Hospital Marqués de Valdecilla, Neurology, Santander, Spain, 5Biosfer Teslab, Ceo, Reus, Spain, 6Universitat Rovira i Virgili, Metabolomics Platform, Tarragona, Spain, 7Hospital Clínic de Barcelona - IDIBAPS - Universitat de Barcelona, Movement Disorders Unit - Neurology Service, Barcelona, Spain, 8Hospital Clínic de Barcelona, Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), University of Barcelona, Department Of Neurology, Barcelona, Spain

Aims: To perform a complete lipidomic profile in serum samples from leucine-rich repeat kinase 2 (LRRK2) gene-mutation carriers to identify new pathophysiological pathways and possible candidate biomarkers. Background: Mutations in LRRK2 gene are the most common genetic cause of Parkinson’s disease (PD). Asymptomatic subjects with mutations in LRRK2 (LRRK2-NMC) present a higher risk of developing LRRK2-PD and represent a unique opportunity to investigate early pathophysiology. Metabolomic profiling has proved useful to study disease mechanisms and to identify biomarkers for a number of diseases, including PD. Methods: We have profiled a total of 225 serum samples of individuals in different subgroups (n=25 samples per group). For the discovery cohort: LRRK2-NMC, LRRK2-PD, idiopathic PD (iPD) and sex- and aged-matched control subjects (HC). For the validation cohort: same groups with an additional follow-up sample for LRRK2-NMC subjects. Mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy techniques have been used for metabolomic profiling, and univariate analysis and machine learning methods for data analysis. Results: Significant alterations of the lipid metabolism were detected in subjects carrying LRRK2 mutations. Our analyses point to certain metabolites as possible biomarkers of PD state. We also incorporated the gender perspective into our analyses and identified metabolites with different profiles according to the sex of the subject/patient. Conclusions: We provide evidence of altered lipidomic profiles associated with prodromal or manifest LRRK2-PD. Some of the identified changes are sex-specific. Our data also indicates that profiling by MS/NMR may be a useful tool for identifying high-risk LRRK2-NMC for PD conversion.

P543 / #442


MASS SPECTROMETRY BASED ANALYSIS OF THE BIOPHYSICAL PROPERTIES OF RECOMBINANT ΑLPHA-SYNUCLEIN FOR THE PROTEIN QUALITY CONTROL AND ROBUST RT-QUIC

Lecture Title:

S. Lee, L. Parkkinen University of Oxford, Nuffield Department Of Clinical Neurosciences, Oxford, United Kingdom

Aims: Real-time quaking-induced conversion (RT-QuIC) is a protein amplification assay that can detect pathological α-synuclein in CSF and brain from patients with Parkinson’s disease. Recombinant α-synuclein, the aggregation substrate, is an important variable in the RT-QuIC assay. Methods of producing recombinant α-synuclein vary significantly among different studies, possibly creating variability in the RT-QuIC outcome. The objective is to develop a quality control method for recombinant α-synuclein to be used for RT-QuIC, which will provide a more rigorous RT-QuIC for the diagnosis of Parkinson’s disease and α-synuclein research. Methods: We purified monomeric recombinant α-synuclein and subjected it to repeated freeze/thaw cycles and different salt (NaCl) concentration. These recombinant α-synuclein were tested in RT-QuIC and analyzed using native size exclusion chromatography - mass spectrometry (SEC-MS) to show that working recombinant α-synuclein in RT-QuIC exists in distinct charge states. Based on the abundance of each distinct charge state, we generated a potential reference for the quality control of recombinant α-synuclein. Results: Recombinant α-synuclein that results in a successful RT-QuIC reaction has a specific charge distribution pattern in the MS raw data. Repeats of freeze/thaw cycles and increasing the salt concentration of recombinant α-synuclein solution disrupted the charge distribution pattern and resulted in RT-QuIC false positives. Conclusions: Specification of a detailed biophysical profile of working recombinant α-synuclein for RT-QuIC can set a criteria for rigorous α-synuclein RT-QuIC. Improved robustness of RT-QuIC will enable more reliable and accurate diagnosis of Parkinson’s disease. Furthermore, studying the α-synuclein fibril products generated by RT-QuIC will provide a clearer understanding of the pathogenesis of Parkinson’s disease.

P544 / #1250


THE A/T/(N) PROFILE IN CSF OF COGNITIVELY UNIMPAIRED PD, PD-MCI, PDD AND DLB PATIENTS

Lecture Title:

F. Paolini Paoletti1, G. Bellomo2, E. Chipi1, M. Petricciuolo2, S. Simoni1, N. Tambasco1, L. Parnetti2 1Department of Medicine, University of Perugia, Section Of Neurology, Perugia, Italy, 2Department of Medicine, University of Perugia, Lab. Of Clinical Neurochemistry, Section Of Neurology, Perugia, Italy

Aims: The A/T/(N) system may be helpful in detecting an underlying Alzheimer’s disease (AD)-pathology in synucleinopathies. We measured cerebrospinal fluid (CSF) Aβ42/Aβ40 ratio, p-tau and t-tau as markers of amyloidosis (A), tauopathy (T) and neurodegeneration (N), respectively, in a cohort of cognitively unimpaired Parkinson’s disease (PD), PD with mild cognitive impairment (PD-MCI), Parkinson’s disease-dementia complex (PDD) and dementia with Lewy bodies (DLB) patients. Our aim was to assess a possible relationship between A/T/(N) positivity and cognitive impairment in synucleinopathies. Methods: CSF samples were collected from 50 PD, 48 PD-MCI, 14 PDD and 15 DLB patients. A/T/(N) profile was considered for each group and A/T/(N)-related biomarkers were classified as + or -, according to the cut-off values used in our Laboratory. Statistical analysis was performed by binomial logistic regression (for differences in prevalences of A+, T+, N+) and two-way ANCOVA (for differences in biomarkers levels) by assuming age as a covariate variable. Results: CSF AD biomarkers did not differ between PD and PD-MCI which showed a prevalence of A-/T-/N-. AD-like CSF profile was significantly more frequent in DLB (40%) and PDD (29%). Prevalence of A+ was higher in DLB and PDD, as well. Remarkably, DLB group showed the lowest value of Aβ42/Aβ40 ratio. Conclusions: Our results are consistent with neuropathological studies describing an amyloid burden hierarchy DLB > PDD > PD. Notably, MCI in PD seems not to be associated to AD-pathology.

P545 / #175


INCREASED CSF NFL IN NON-DEMENTED PARKINSON’S DISEASE PATIENTS REFLECTS EARLY WHITE MATTER DAMAGE

Lecture Title:

E. Papuć, K. Rejdak Medical University of Lublin, Poland, Department Of Neurology, Lublin, Poland

Aims: Parkinson’s disease (PD) is a chronic neurodegenerative disorder with different underlying pathological processes. Until now, no biochemical biomarkers have been established for PD. Given recent biochemical and neuroimaging evidence for the presence of white matter damage in PD, which may even precede neuronal loss, we checked whether neurofilament light (NFL) was increased in the cerebrospinal fluid (CSF) of PD patients in comparison to controls. NFL is located mainly in large myelinated axons, and increased CSF levels of this protein reflect axonal injury. Methods: CSF levels of NFL in 58 early PD patients and 28 controls were quantified by ELISA (Uman Diagnostics). Measures of PD severity included disease duration, UPDRS-III, and Hoehn-Yahr stage. Results: Statistically significant differences in CSF NFL levels were found between PD patients and controls [median with interquartile range 524.82 (393.28–678.34) vs 271.84 (198.09–335.24) ng/L; p<0.05)]. In PD patients, there were no correlations between CSF NFL level and the measures of disease severity. The CSF NFL turned out to have a high discriminatory value (AUC 0.850) for differentiating between PD subjects and healthy controls, with 84 % sensitivity and 85.2 % specificity. Conclusions: The study demonstrates that axonal damage is present in early PD in addition to neuronal loss. Interestingly, white matter damage was observed in non-demented PD patients. In the light of the results of recent MRI studies which confirm early white matter damage in PD, our data may turn out to be potentially useful in the diagnosis of early, or even preclinical, stages of the disease.

P546 / #1795


IDENTIFICATION OF NEW BIOMARKER SIGNATURES FOR NEURODEGENERATIVE DISEASES

Lecture Title:

M. Schulz1, C. Schroeder2, R. Schmidt2, M. Richter1 1AbbVie Deutschland, Dmpk-ba, Ludwigshafen, Germany, 2Sciomics, R&d, Neckargemuend, Germany

Aims: We have investigated CSF and plasma samples from Alzheimer Disease (AD), Parkinson Disease (PD) as well as Multiple Sclerosis (MS) patients and compared the protein expression profile to samples from healthy humans in order to identify new biomarker signatures. In contrast to other areas of medicine diagnosis of neurodegenerative disease is still lacking reliable fluid biomarkers, especially since now disease modifying therapies are in clinical development. New biomarkers should increase the diagnostic accuracy and allow earlier diagnosis, better participant selection and disease activity and treatment effect monitoring. Methods: Antibody Microarrays for immuno-based protein biomarker detection developed by Sciomics cover key pathway proteins as well as secreted proteins. With sample consumption of less than 100µL a spectrum of about 1300 proteins is detected covering signaling pathway proteins, transcription factors, apoptosis marker, markers for oxidative stress, cell surface markers and cytokines. Results: Proteins showing different abundance between AD, MS and PD compared to controls were identified in both body fluids. In AD compared to control subjects 108 differential proteins were identified in CSF samples whereas 21 proteins were differential in plasma samples with some proteins showing an overlap between the body fluids. This will be verified in a larger cohort with focus on proteins significantly changed on both matrices or proteins contributing to selected functions. Conclusions: The comparison of CSF and plasma protein expression patterns allow a conclusion on the capability to identify pathways affected by the disease in the same way in both fluids. This may guide the search for new blood-based biomarkers for neurodegenerative disease.

P547 / #1240


PLASMA INSULIN-LIKE GROWTH FACTOR 2 (IGF2) IN PARKINSON DISEASE: POTENTIAL PREDICTOR MARKER.

Lecture Title:

D. Sepulveda1,2,3, F. Grünenwald1,2,3, M. Cisternas-Olmedo1,2,3, R. Vidal1,2,3 1UNIVERSIDAD DE CHILE, Biomedical Neuroscience Institute, Santiago, Chile, 2Center for Geroscience, Brain Health and Metabolism, Gero, Santiago, Chile, 3UNIVERSIDAD MAYOR, Center For Integrative Biology, Santiago, Chile

Aims: Introduction: The search to identify non-invasive early markers is a current challenge in the neurodegeneration field. Insulin-like growth factor 2 (IGF2) have been proposed as an interesting biomolecule related to Idiopathic Parkinson’s disease (PD). IGF2 is a secreted factor with interesting neuroprotective properties in various disease models and recent reports suggest that IGFs are possible targets for the treatment of neurodegenerative diseases, particularly considering two roles (i) neurogenic agents and (ii) their apparent neuroprotective effects. Methods: Materials & Methods: Here we determine the IGF2 and IGF1 levels in plasma and polymorphonuclear blood cells (PMBCs), from Chilean patients with moderately advanced PD, by ELISA and real time PCR methodologies and we correlate with clinical parameters of the disease severity (Hoehn and Yahr modified and UPDRS scale). Results: The results shown a significative decrease of IGF2 plasma levels in PD patients against health control group. In addition, we report a dramatic diminished of IGF2 mRNA and protein levels in PMBCs from PD patients. We found no correlations between IGF2 plasma levels and motor symptoms related with the severity of the disease. Conclusions: Furthermore, IGF2 may be involved in the underlying pathophysiology of PD and a potential biomarker by this disease.

P548 / #773


EXTRACELLULAR VESICLES IMMUNE-PROFILING IN PLASMA AND CSF: A DIAGNOSTIC TOOL FOR PARKINSON’S DISEASE

Lecture Title:

E. Vacchi1,2, J. Burrello3, A. Burrello4, S. Bolis3, L. Barile1,3, A. Kaelin-Lang1,2,5,6, G. Melli1,2,5 1Università della Svizzera Italiana (USI), Faculty Of Biomedical Sciences, Lugano, Switzerland, 2Ente Ospedaliero Cantonale (EOC), Laboratory For Biomedical Neurosciences, Lugano, Switzerland, 3Cardiocentro Ticino Foundation, Cellular And Molecular Cardiology Laboratory And Laboratory For Cardiovascular Theranostics, Lugano, Switzerland, 4University of Bologna, Department Of Electrical, Bologna, Italy, 5Ente Ospedaliero Cantonale (EOC), Neurology Department, Lugano, Switzerland, 6University of Bern, Department Of Neurology, Bern, Switzerland

Aims: Extracellular vesicles (EVs) play a central role in intercellular communication which is highly relevant for inflammatory processes implicated in neurodegenerative proteinopathies disorders, such as Parkinson’s Disease (PD). Thus, we characterized and compared distinctive EV subpopulations from plasma and CSF of PD and atypical parkinsonisms (AP), with the aim to build a diagnostic model via artificial intelligence. Methods: Plasmatic EVs were collected from 27PD, 19 matched healthy controls (HC), 9AP with multiple system atrophy (MSA) and 9AP with tauopathies (AP-Tau). CSF-derived EVs were collected from 4PD, 4MSA, 4AP-Tau. The expression of 37 EV-surface markers, related to inflammatory and immune cells, were measured by MACSPlex in plasm and CSF. Random forest (RF) diagnostic models based on EV markers expression were built via supervised machine learning algorithms. Results: In plasma, 17 EV-derived markers resulted statistically different between HC and PD. Among these, 8 were overexpressed also in CSF of a subset of PD, 10 in a subset of MSA and 6 in a subset of AP-TAU. A RF model based on the 17 differentially expressed, EV-derived markers in plasma discriminates patients from HC with high sensitivity (100%) and specificity (83.3%). Integrating CSF-derived data a potentiated model was able to differentiate PD from not-PD patients with 96.6% of sensitivity and 88.0% specificity. Conclusions: Multiple immune surface-markers profiling of EVs in plasma allows the stratification of patients with PD, MSA and AP-Tau. Furtherly it suggests a different immune dysregulation in PD and MSA vs. AP-Tau, to be confirmed by functional analysis in experimental models of disease.

P549 / #765

Topic: Theme C: α-Synucleinopathies / C4.g. Imaging, Biomarkers, Diagnostics: EEG, brain mapping, MEG

IDENTIFICATION OF A NEW PRODROMAL MARKER OF PARKINSON’S DISEASE USING TRANSLATIONAL EEG BIOMARKERS IN PRECLINICAL MODELS

Lecture Title:

V. Duveau1, J. Volle1, G. Porras2, E. Pioli2, A. Evrard1, C. Ruggiero1, C. Roucard1, E. Bezard2, Y. Roche1 1SynapCell SAS, Synapcell Sas, Saint Ismier, France, 2Motac Neuroscience, Motac Neuroscience, Bordeaux, France

Aims: Whereas the nature of the etiology of the process underlying clinical deterioration remains unknown, Parkinson’s disease (PD) is characterized by the progressive loss of Substantia Nigra neurons with two main phases: a prodromal and a symptomatic phase. Despite years of research, surrogate biomarkers of disease progression are still lacking. Cumulative evidences have demonstrated the presence of aberrant oscillatory synchronization of neuronal activity across the cortico-basal ganglia-thalamo-cortical circuit in both PD patients and experimental animal models during the symptomatic phase. Using state-of-the-art EEG methodologies, we here investigated the progressive development of β-band (~30Hz) in a rat model of PD as well as its response to the reference dopamine replacement therapy, L-DOPA, once degeneration is established. Methods: We used the bilateral AAV-alpha-synuclein (α-Syn) rat model that allows investigating the progression of the degeneration over several weeks combined with the recording of EEG activities in the motor cortex. Results: In the early stage we observed the progressive rise in β-band power from the prodromal phase onwards in the motor cortex. In the late stage of the disease, a prominent β-band (~30Hz) was observed in the motor cortex (inexistent in control rats) in a subset of rats. The pharmacological challenge with L-DOPA, shows a reduction of this β-band. Conclusions: Since the late stage data fit with the clinical literature, we propose that the β-band rise may represent a predictive, reliable, objective and clinically-relevant biomarker for the validation of disease modifying experimental therapeutics. In addition, our data call for a clinical validation of this biomarker.

P550 / #1216


RESTING-STATE QEEG ALPHA/THETA RATIO RELATED TO NEUROPSYCHOLOGICAL TEST PERFORMANCE IN PARKINSON’S DISEASE

Lecture Title:

A. Jaramillo-Jimenez1, J. Suarez-Revelo2, J. Ochoa-Gomez2, J. Carmona2, Y. Bocanegra2, F. Lopera2, O. Buriticá2, D. Pineda Salazar2, L. Moreno Gómez3, C. Tobón Quintero4, M. Borda1, L. Bonanni5, D. Ffytche6, K. Brønnick1, D. Aarsland1 1Stavanger University Hospital, Centre For Age-related Medicine (sesam), Stavanger, Norway, 2Universidad de Antioquia, School of Medicine, Grupo De Neurociencias De Antioquia, Medellín, Colombia, 3Pablo Tobón Uribe Hospital, Neurology Unit, Medellín, Colombia, 4Universidad de Antioquia, School of Medicine, Grupo Neuropsicología Y Conducta (gruneco), Medellín, Colombia, 5University G. d'Annunzio of Chieti-Pescara, Neuroscience, Imaging And Clinical Sciences, Chieti, Italy, 6King’s College London, Department Of Old Age Psychiatry, Institute Of Psychiatry, Psychology, And Neuroscience, London, United Kingdom

Aims: To determine possible associations of hemispheric-regional alpha/theta ratio (α/θ) with neuropsychological test performance in Parkinson’s Disease non-demented patients (PD). Methods: 36 PD were matched to 36 Healthy Controls (HC). Resting-state quantitative electroencephalograms (qEEG) were recorded, α/θ in eight hemispheric-regions was computed from relative power spectral densities. Correlations between α/θ and performance in several neuropsychological tests were conducted, significant findings were included in a moderation analysis. Results: The α/θ in all regions was lower in PD than in HC, with larger effect sizes in the posterior regions. Right parietal, and right and left occipital α/θ had significant positive correlations with performance in Judgement of Line Orientation Test (JLOT) in PD. Adjusted moderation analyses indicated that right, but not left, occipital α/θ influenced the JLOT performance related to PD. Conclusions: Reduction of the occipital α/θ, in particular on the right side, was associated with visuospatial performance impairment in PD. Visuospatial impairment in PD, which is highly correlated with the subsequent development of dementia, is reflected in α/θ in the right posterior regions. The right occipital α/θ may represent a useful qEEG marker for evaluating the presence of early signs of cognitive decline in PD and the subsequent risk of dementia. A hemispheric approach of occipital α/θ must be considered for further research.

P551 / #622


ANALYSIS OF EEG COHERENCE IN NON-MOTOR SYMPTOMS WITH PARKINSON'S DISEASE.

Lecture Title:

K. Kinugawa, T. Mano, K. Sugie Nara Medical Univercity, Neurology, Kashihara, Japan

Aims: The underlying pathophysiology of the occurrence of non-motor symptoms (NMS) in Parkinson's disease (PD) is unknown. This aim to the present study is to evaluate the association between NMS and functional connectivity in brain networks quantified by electroencephalography (EEG) coherence. Methods: A total of fourteen sporadic PD patients were included in the present study and evaluated for motor symptoms and NMS, including hallucination, rapid eye movement sleep behavior disorder (RBD), hyposmia, pain and cognitive dysfunction. We performed time frequency analysis and coherence analysis on the resting state EEG in the theta (4-7 Hz), alpha (8-12 Hz), beta (13-30 Hz) and gamma (31-45 Hz) ranges. We calculated whole-brain coherence and regional coherence based on the coherence for all electrode pairs. Results: The coherence values was correlated with disease duration rather than with age. EEG coherence values were compared between NMS positive and negative patients; RBD positive PD showed low coherence in theta and alpha parietal-occipital regions compared to RBD negative. Hallucination positive PD showed low coherence in the alpha occipital regions compared to negative patients. The mini mental state examination (MMSE) scores, especially orientation and memory items, showed a positive correlation with the whole-brain and regional coherences in the alpha range. Conclusions: The NMS indicated the decreased brain efficiency as well as impaired neural synchronisation. The discrimination of coherence across NMS may reflect a compensatory response of the brain to the neuropathology in PD.

P552 / #904

Topic: Theme C: α-Synucleinopathies / C4.h. Imaging, Biomarkers, Diagnostics: Cognitive, psychometric, behavioral and motor tests

NEUROANATOMICAL SUBSTRATES OF ATTENTION PROCESSING SPEED IN EARLY DEMENTIA WITH LEWY BODIES (DLB): A WHITE-MATTER VOXEL-BASED MORPHOMETRY AND DIFFUSION TENSOR IMAGING STUDY

Lecture Title:

A. Botzung1, M. Mondino2, V. Noblet2, N. Philippi1,2, F. Blanc1,2 1Hôpitaux Universitaires de Strasbourg, Geriatrics And Neurology Units, Research And Resources Memory Center (cmrr), Strasbourg, France, 2CNRS, Icube Laboratory, Umr 7357, Strasbourg, France

Aims: Based on a previous study (Botzung et al., 2019) that revealed the importance of grey matter (GM) integrity in the striatum for attention processing speed, the purpose of this study was now to examine, in early DLB patients, the relationship between impaired attentional speed performances and white matter changes. Methods: We administered the Trail Making Test A (TMTA) to 73 prodromal to moderate DLB patients (mean MMSE = 26.4) and 30 control subjects (mean MMSE = 28.9) to assess attention processing speed. Three-dimensional (3D) MRI and Diffusion-weighted images (DTI) were acquired for all participants and correlational analyses were performed in the patient group using WM-VBM, fractional anisotropy (FA) and mean diffusivity (MD). Results: Behavioral results showed significantly impaired performances in patients in comparison to control subjects (p = .004). In addition, correlational analyses using WM-VBM revealed negative WM correlations in the anterior corpus callosum and in the anterior cingulum (p < .05 FDR). Using DTI (p < .05 FDR), correlated with TMTA scores, we found reduced FA in bilateral frontal and posterior regions, and increased MD in bilateral anterior and posterior networks. Conclusions: WM-VBM analysis revealed the involvement of anterior corpus callosum and cingulum, which could indicate a disruption of the loops connecting striatal regions and the prefrontal cortex. Changes in both FA and MD also correlate with impaired attentional speed: further DTI analyses are still ongoing to question the degree of which white matter tracts are disrupted between the striatum and cortical anterior and posterior networks, respectively.

P553 / #1157


MOTOR AND PERCEPTUAL IMPAIRMENTS IN A PRODROMAL STAGE OF SYNUCLEINOPATHIES

Lecture Title:

M. Brillon-Corbeil1,2, L. Bernier-Lalonger1,2, R. Postuma1,3, J. Montplaisir1,4, J. De Roy1,2, A. Pelletier1,5, J.-F. Gagnon1,2 1Hôpital du Sacré-Coeur de Montréal, Center For Advanced Research In Sleep Medicine, Montreal, Canada, 2Université du Québec à Montréal, Psychology, Montreal, Canada, 3McGill University, Department Of Neurology, Montreal, Canada, 4Université de Montréal, Psychiatry, Montreal, Canada, 5Montreal General Hospital, Neurology, Montreal, Canada

Aims: Idiopathic rapid eye movement sleep behavior disorder (iRBD) is a parasomnia and a prodromal stage of Parkinson’s disease and dementia with Lewy bodies. Mild cognitive impairment (MCI), motor symptoms and perceptual alterations are frequent in iRBD. However, the proportion of patients with clinically significant motor and perceptual alterations and their links with MCI remain poorly studied. We aimed to investigate motor and perceptual impairments in iRBD patients according to their cognitive status. Methods: Patients with polysomnography-confirmed iRBD (n=107, age=66.05±7.06, 78% male) were classified into two groups according to their cognitive performance: patients with normal cognition (iRBD-NC) and patients with MCI (iRBD-MCI). All participants had a neurological examination including four motor measures (Unified Parkinson Disease Rating Scale part III, Purdue Pegboard, Alternate Tap test, and Timed Up and Go) and two perceptual (olfaction and color discrimination) tests [University of Pennsylvania Smell Identification Test (UPSIT) and Farnsworth-Munsell 100 (FM-100)]. Pearson chi-square tests were performed to compare the proportion of patients with clinically impaired symptoms in both groups based on normative data. Results: Sixty-one (36%) patients had MCI. The proportion of patients having at least two motor measures impaired was higher in iRBD-MCI (63%) than iRBD-NC (35%) patients (p<0.001). Moreover, the proportion of patients with impaired performance on the FM-100 was higher in iRBD-MCI (66%) than iRBD-NC (32%) patients (p<0.001). However, the proportions were similar between the two groups for the UPSIT (iRBD-MCI=66%, iRBD-NC=58%). Conclusions: iRBD patients with MCI are more severely altered on motor and color discrimination functions.

P554 / #1267


EARLY COGNITIVE IMPAIRMENT IN PARKINSON DISEASE

Lecture Title:

B. Paradowski, J. Chojdak-Łukasiewicz Medical University, Neurology, Wroclaw, Poland

Aims: Objective: Parkinson's disease (PD) is a progressive neurodegenerative disease which for a very long time was predominantly considered as a motor disorder. However, it is frequently associated with dementia, which is the most debilitating non-motor symptom of Parkinson’s disease.The risk of developing dementia is six times higher in PD patients than in general population and increases with longer duration of disease. We assessed the results of a neuropsychological test in PD patients without recognized dementia. Methods: To explore the presenting early symptoms of dementia we assessed 64 patients with clinical diagnosis of PD (29 female) with mean age 68,3±9,8. Disease duration ranged from 0,25 to 24 years (6,2±4,6).The neuropsychological assessment was carried for all the patients and included: MMSE (Mini Mental State Examination), CDT (Clock Drawing Test), verbal fluency test (Controlled Oral Word Association Test - COWAT in both categories semantic and phonemic). The stage of the PD was assessed according to the five stage Hoehn-Yahr scale. Results: The mean results of MMSE was 27,2±2,3 point, CDT 8,0±2,0 point, COWAT s16,1±5,7 and COWAT f 12,0±4,7. A significant correlation has been demonstrated between age and mean values of MMSE scale , CTD and fluency test. Conclusions: 1.Majority of the PD patients even without recognized dementia suffer with visuospatial and semantic fluency dysfunctions. 2. The level of cognitive impairment in PD is dependent on patient age and the motor symptom severity assessed using the H-Y scale. 3.Cognitive impairment are present even with patients with short disease duration.

P555 / #1104


EARLY COGNITIVE DECLINE AFTER BILATERAL SUBTHALAMIC DEEP BRAIN STIMULATION IN PARKINSON'S DISEASE PATIENTS WITH GBA MUTATIONS

Lecture Title:

G. Mangone1, S. Bekadar2, F. Cormier-Dequaire1, K. Tahiri2, A. Welaratne3, V. Czernecki4, F. Pineau4, C. Karachi5, A. Castrioto6, F. Durif7, C. Tranchant8, D. Devos9, S. Thobois10, W. Meissner11, M.S. Navarro5, P. Cornu5, S. Lesage12, A. Brice12, M.L. Welter13, J.-C. Corvol1 1Sorbonne Université, Inserm U1127, CNRS UMR 7225, Institut du Cerveau et de la Moelle épinière, Paris, France. Assistance Publique Hôpitaux de Paris, Hôpital Pitié-Salpêtrière, Département de Neurologie, Clinical Research Center Neurosciences., Neurology, Paris, France, 2Sorbonne Université, Inserm U1127, CNRS UMR 7225, Institut du Cerveau et de la Moelle épinière, Paris, France., 75013, Paris, France, 3Assistance Publique Hôpitaux de Paris, Hôpital Pitié-Salpêtrière, Département de Neurologie, Clinical Research Center Neurosciences, Paris, France., 75013, Paris, France, 4Sorbonne Université, Inserm U1127, CNRS UMR 7225, Institut du Cerveau et de la Moelle épinière, Paris, France. Assistance Publique Hôpitaux de Paris, Hôpital Pitié-Salpêtrière, Département de Neurologie, Clinical Research Center Neurosciences, Paris, Fran, 75013, Paris, France, 5Assistance Publique des Hopitaux de Paris, Hôpital Pitié-Salpetrière, département de Neurochirurgie, Neurosurgery, Paris, France, 6Unité des Troubles du Mouvement, Département de Neurologie, CHU de Grenoble, Université de Grenoble Alpes, INSERM U1216, Département De Neurologie, Chu De Grenoble, Université De Grenoble Alpes, Grenoble, France, 7Service de Neurologie, CHU Clermont-Ferrand, Université Clermont Auvergne, Département De Neurologie, Clermont Ferrand, France, 8Hôpitaux Universitaires de Strasbourg, Neurology, Strasbourg, France, 9Univ Lille, Inserm, CHU Lille, U1172,lille Neurosciences & Cognition- Team « Degenerative & Vascular Cognitive Disorders ", Lille, France, 10Neurologie C, Hôpital Neurologique Pierre Wertheimer, Hospices Civils de Lyon, 69500 Bron, France; Univ Lyon, Université Claude Bernard Lyon 1, Faculté de Médecine Lyon Sud, Lyon, France; Institut des Sciences Cognitives Marc Jeannerod, UMR 5229, CNRS, Br, Département De Neurologie, Lyon, France, 11University Bordeaux, Institute For Neurodegenerative Diseases, Cnrs Umr5293, Bordeaux, France, 12Centre de Recherche de l'Institut du Cerveau et de la Moelleépinière, Institut National De La Santé Et De La Recherche Médicale, Paris, France, 13Sorbonne Université, Inserm U1127, CNRS UMR 7225, Institut du Cerveau et de la Moelle épinière, Paris, France. Assistance Publique Hôpitaux de Paris, Hôpital Pitié-Salpêtrière, Département de Neurologie, Clinical Research Center Neurosciences., Neurology, Paris, France

Aims: Subthalamic nucleus deep brain stimulation (STN-DBS) has demonstrated its efficacy on motor complications in advanced Parkinson’s disease (PD) but does not modify disease progression. Genetic forms of PD have been associated with different cognitive progression profiles. Methods: Patients with STN-DBS were screened for LRRK2, GBA, and PRKN mutations at the Pitié-Salpêtrière Hospital between 1997 and 2009. Patients with known monogenetic forms of PD from six other centers were also included. The Mattis Dementia Rating Scale (MDRS) was used to evaluate cognition at baseline and one-year post-surgery. The standardized Unified PD Rating Scale (UPDRS) evaluation On and Off medication/DBS was also administered. A generalized linear model adjusted for sex, ethnicity, age at onset, and disease duration was used to evaluate the effect of genetic factors on MDRS changes. Results: We analyzed 208 patients (131 males, 77 females, 54.3±8.8 years)(Figure 1) including 25 GBA, 18 LRRK2, 22 PRKN, and 143 PD patients without mutations. PRKN patients were younger and had a longer disease duration at baseline (Figure 2). A GBA mutation was the only significant genetic factor associated with MDRS change (β=-2.51, p=0.009). GBA mutation carriers had a more pronounced post-operative MDRS decline (3.2±5.1) than patients with LRRK2 (0.9±4.8), PRKN (0.5±2.7) or controls (1.4±4.4) (Figure 3). The motor response to DBS was similar between groups (Figure 4).

Conclusions: GBA mutations are associated with early cognitive decline following STN-DBS. Neuropsychological assessment and discussions on the benefit/risk ratio of DBS are particularly important for this population.

P556 / #366


STRUCTURAL IMAGING PREDICTORS OF COGNITIVE SEVERITY IN PARKINSON’S DISEASE

Lecture Title:

M. Shahid1, C. Mcdaniel1, T. Billet2, A. Ribbens2, N. Barros2, T. Montine3, K. Poston1 1Stanford University School of Medicine, Neurology & Neurological Sciences, Palo Alto, United States of America, 2Icometrix, Research & Development, Leuven, Belgium, 3Stanford University School of Medicine, Pathology, Palo Alto, United States of America

Aims: To identify differences in structural imaging biomarkers in Parkinson’s disease (PD) without (PD-Nonimpaired) and with (PD-Impaired) cognitive impairment. Methods: We applied Icobrain software to 3D-T1 and 2D-FLAIR MRI scans of 69 PD-Nonimpaired, 50 PD-Impaired, and 63 Controls. We quantified 20 biomarkers of atrophy in the whole brain, temporal cortex, frontal cortex, parietal cortex and the hippocampus. We performed between-group comparisons and univariate/multivariate correlation analyses to determine biomarker predictors of cognitive severity. Results: Using a rank-based ANCOVA correcting for age, all PD had lower midbrain-to-pons ratio (p<0.001) compared to Controls. By contrast, PD-Impaired had lower hippocampal (p<0.001), total gray matter (p<0.01), and total cortical gray matter volume (p<0.05), compared to the PD-Nonimpaired. In PD-Impaired, MoCA scores positively correlated with right hippocampal (R=0.356;p=0.012) and white matter volume (R=0.374;p=0.008), and negatively correlated with lateral ventricle volume (R=-0.368;p=0.009) and lateral ventricles-to-whole brain volume ratio (R=-0.376;p=0.008). In a multivariate backward stepwise linear regression, we iteratively removed imaging biomarkers that did not significantly predict the MoCA until the best fitting model (lowest-AIC) was obtained. In all PD, 35.8% of the MoCA variance was explained by imaging biomarkers, mostly the temporal cortex, whole brain, and FLAIR hyperintense volume. In PD-Impaired, 47% of the MoCA variance was explained by imaging biomarkers, mostly the whole brain, white matter, occipital cortex, and lateral ventral volumes. Conclusions: While PD is associated with midbrain atrophy, cognitive impairment in PD is associated with hippocampal and whole brain atrophy. Icobrain software is a promising tool for quantification of clinically meaningful imaging biomarkers.

P557 / #1079


USE OF ADDENBROOKE'S COGNITIVE EXAMINATION III (ACE-III) IN PATIENTS WITH PARKINSON'S DISEASE

Lecture Title:

N. Sousa, S. Brucki University of Sao Paulo, Neurology, Sao Paulo, Brazil

Aims: To evaluate the applicability of ACE-III in patients with PD. Methods: A total of 126 patients and 34 controls, paired by age and schooling, participated in this study. There was a higher prevalence of men than women in the clinical group. In clinical, mood and anxiety, as well as functional measures were observed higher scores for the clinical group. Regarding the severity of the disease, it was composed mainly of patients in scores I and II of Hoehn & Yahr stages. In the present study, 83 was the cut grade with the ability to discriminate dementia syndrome, showing 100% sensitivity and 0.9061 area under the curve, with a standard error of 0.0427 (95% confidence interval: 0.8223 -0.9899). Results: The results showed good diagnostic accuracy of the instrument, as well as significative correlation with standardized neuropsychological battery tests for this population. Conclusions: The data suggest that patients with PD presented difficulties related to attentional and executive measures when compared to the control group. Differences between groups were more identified when stratified by the PD group, due to demographic and clinical factors. Despite the small size of the control group, it was observed that this one presented superior performance in all indices when compared to the clinical group, mainly global scores and attention/executives subscores. Conclusions: These results reinforce the usefulness of ACE-III as an instrument for assessing dementia in PD.

P558 / #1557


CROSS-SECTIONAL ASSOCIATIONS BETWEEN COGNITION AND MOBILITY IN INDIVIDUALS WITH PARKINSON’S DISEASE MILD COGNITIVE IMPAIRMENT

Lecture Title:

N. Sousa1, R. Macedo2, S. Brucki3 1University of Sao Paulo, Neurology, Sao Paulo, Brazil, 2Hospital SARAH of Rehabilitation, Neurological Rehabilitation, Salvador, Brazil, 3University os Sao Paulo, Neurology, Sao Paulo, Brazil

Aims: To investigate the association between cognitive functions and gait performance (balance, gait speed and mobility) in patients with Parkinson’s disease-mild cognitive impairment attended to a hospital neurorehabilitation program in Brazil. Methods: The inclusion criteria were: (a) PD diagnosis, according to the criteria of the UK, Brain Bank (b) an age over 50 years, (c) Hoehn and Yahr stages I–III; (d) Beck Depression Inventory ≤16. Descriptive and inferential statistics (Pearson's correlation) were used for data analysis.The cognitive functions were evaluated through Digit Span Test, Trail Making Test (TMT A, B and B-A), Verbal Fluency Test, Addenbrooke’s Cognitive Examination III (total and domain scores).The motor function through 10-Meter Walking Test, Mini BESTest and Timed Up and Go Test. Results: A total of 107 patients (79 males, 28 females). Mean age 60.92 (8.28) years, disease duration 5.49 (4.16) years, schooling 11.73 (4.12) presenting PD-MCI, according to the MDS PD-MCI Level II diagnostic criteria was recruited for this study. The balance skills were significantly associated with global cognition (p=0.020) and verbal fluency (p=0.048), visuospatial (p=0.006), TMT B (p=0.006), TMT B-A (p=0.006). Gait speed showed associated with global cognition scores (p=0.026), memory (p= 0.002), and TMT B (p=0.017). Functional mobility showed a significant association with all cognitive tests, independent of demographic and clinical aspects. Conclusions: These findings might help early identification of cognitive deficits or motor dysfunctions in patients with PD who may benefit from rehabilitative strategies.Future prospective studies are needed to investigate the effects of cognitive training on motor performance.

P559 / #1444


MEDICATION MANAGEMENT IN PARKINSON DISEASE PATIENTS WITH MILD COGNITIVE IMPAIRMENT

Lecture Title:

B. Sumbul Sekerci1, H. Hanagasi2, B. Bilgic2, Z. Tufekcioglu2, M. Emre2 1Bezmialem Vakif University, Faculty of Pharmacy, Department Of Clinical Pharmacy, Istanbul, Turkey, 2Istanbul University, Istanbul Faculty of Medicine, Department Of Neurology, Behavioral Neurology And Movement Disorders Unit, Istanbul, Turkey

Aims: The main feature that distinguishes mild cognitive impairment (MCI) from dementia is the absence of significant functional decline due to cognitive impairment. However, in Parkinson's Disease patients with MCI (PD-MCI) the effect of cognitive impairment on complex instrumental daily activities such as medication management is not well established. Methods: Fifty-eight patients diagnosed with idiopathic Parkinson's disease were evaluated according to Level 2 Movement Disorders Society diagnostic criteria for PD-MCI. A detailed neuropsychological battery (including tests for attention and working memory, executive functions, language, visuo-spatial functions, episodic memory, and prospective memory) was applied to the patients. The raw scores of neuropsychological tests were converted into z-scores. Also, a standardized performance-based assessment (Medication Management Ability Assessment) was administered to evaluate medication management. Results: The PD-MCI group obtained significantly lower scores in medication management assessment (p <0.01) and made more mistakes with regard to prescription instructions (e.g, took more or less medication, did not use medications as instructed with regard to meal times). Medication management performance in this group was positively correlated with event-based prospective memory. Conclusions: Medication management is an efficient complex daily activity where a minimal decrease in cognitive impairment may be observed. PD patients with MCI have impaired medication management. Diagnosing MCI in PD is important so that appropriate measures can be taken to provide support and improve medication management.

P560 / #674


SELF, INSULA & DEMENTIA WITH LEWY BODIES

Lecture Title:

A. Tisserand1,2, N. Philippi1,2, A. Botzung1, F. Blanc1,2 1Hôpitaux Universitaires de Strasbourg, Geriatrics And Neurology Units, Research And Resources Memory Center (cmrr), Strasbourg, France, 2CNRS, Icube Laboratory, Umr 7357, Strasbourg, France

Aims: The objective is to review evidence of the relationship between the insular cortex, the Self, and Dementia with Lewy bodies (DLB). Indeed, modifications of the Self, such as anosognosia and other changes in personal identity are frequently reported by relatives and noticed by clinicians. Those changes could be related to insular damage, a region central to the Self, which atrophy occurs early in the course of the disease (Blanc et al., 2017). Methods: We performed researches on PubMed, using keywords and MeSH subheadings, limited to papers in English. The terms “Lewy bodies”, “Insula”, “Self”, and different aspects related to the self, such as “subjective sense of self”, “self-awareness”, “self-concept” and “autobiographical memory” were used to indicate the themes of interest. Results: The evidence suggest that insula is involved in processing various aspects of the self, notably the prerequisites allowing the emergence of self-awareness (Craig, 2009; Damasio, 2003). Moreover, changes in personal tastes have been highlighted in DLB and related to insular atrophy (Philippi et al., 2020). However, there is, to date, little work exploring the different components of the self (i.e. subjective sense of self and self-awareness; self-concept; autobiographical memory) in DLB (Pennington et al., 2020). Conclusions: Changes in Self have rarely been explored in DLB, whereas they appear clinically obvious and could be related to insular atrophy. We therefore plan to investigate the various aspects of the Self in DLB, compared to Alzheimer disease and healthy elderly population, by combining behavioral measures and multimodal neuroimaging.

P561 / #1352

Topic: Theme C: α-Synucleinopathies / C4.j. Imaging, Biomarkers, Diagnostics: Other

MULTIVARIATE PREDICTION OF COGNITIVE DECLINE IN PARKINSON’S DISEASE

Lecture Title:

J. Harvey1, R. Reijnders2, G. Shireby1, A. Duits2, S. Köhler2, B. Creese1, K. Lunnon1, E. Pishva2 1University of Exeter, Medical School, Exeter, United Kingdom, 2Maastricht University, Department Of Psychiatry And Neuropsychology, Maastricht, Netherlands

Aims: Cognitive impairment is a common non-motor symptom in Parkinson’s disease (PD), with high variability in onset, rate and trajectory of decline. We aimed to establish a powerful multivariate prediction model of cognitive trajectory in PD using baseline genomic, epigenomic and known clinical and cerebrospinal fluid (CSF) markers for cognitive function. Methods: We used the Montreal Cognitive Assessment (MoCA) and latent class mixed models to identify classes of cognitive trajectories in the Parkinson's Progression Markers Initiative (PPMI) drug-naïve de-novo PD cohort over a 7-year follow-up. For patients with genotyping data, we computed polygenic risk scores (PRS) for PD, Alzheimer’s disease and educational attainment. After splitting groups into training and test subsets, we used leave-p-out cross-validation differential expression analysis to identify robust features of DNA methylation and miRNA expression profiled in the training dataset. In order to establish an optimal predictive model of cognitive function trajectory, we tested multiple classifiers using machine-learning (ML) algorithms integrating genomic and epigenomic features with a set of clinical and CSF markers. Results: On average, the largest class (62.3%) presented with stable cognitive function, whilst remaining cases showed longitudinal cognitive decline. Integrating clinical and CSF markers with top ranked epigenomic features and PRSs in the training set, ML resulted in accurate prediction of cognitive class in the test dataset. Conclusions: Machine-learning methods showed additive predictive accuracy with integrated epigenomic and genomic features for identified cognitive trajectories in PD.

P562 / #835


SALIENCE NETWORK TOPOLOGY AND INTEGRITY ASSOCIATES WITH COGNITIVE AND AFFECTIVE IMPAIRMENT IN PARKINSON’S DISEASE.

Lecture Title:

L. Jonkman1, Y. Fathy1, H. Berendse2, M. Schoonheim1, W. Van De Berg1 1Amsterdam UMC, location VUmc, Anatomy And Neurosciences, Amsterdam, Netherlands, 2Amsterdam UMC, location VUmc, Neurology, Amsterdam, Netherlands

Aims: The salience network, consisting of the anterior insular cortex (AIC) and anterior cingulate cortex (ACC), shows extensive pathology in Parkinson disease (PD), but remains understudied in-vivo. This study quantified structural damage to this network in PD using diffusion tensor imaging (DTI) and explored clinical correlates. We hypothesize that structural disconnection of AIC sub-regions, and microstructural damage of tracts between AIC sub-regions and the ACC, associate with cognitive and affective impairment in PD. Methods: MRI of 53 PD patients and 15 controls included 3D-T1 for segmenting the AIC into ventral (vAI) and dorsal (dAI) anterior insula. DTI-based probabilistic tractography was performed with subsequent structural network construction and topological assessment of eigenvector and betweenness centrality. Microstructural tract integrity was quantified between vAI, dAI and ACC, using fractional anisotrophy (FA) and mean diffusivity (MD). Lastly, a battery of cognitive and affective tests was included and associated with centrality measures, and with FA and MD of tracts. Results: The dAI had significantly higher eigenvector centrality in PD than controls (p<0.01), and associated with higher depression scores (left dAI only, r=0.28,p<0.05). Tracts between dAI and ACC showed lower FA and higher MD in PD (p<0.05), and associated with semantic fluency, working memory capacity, executive functioning, and anxiety symptoms (range 0.002<p<0.05).

Conclusions: This study provides evidence for clinically relevant structural damage to the salience network in PD, primarily centered around the dAI. The observed alterations were associated with PD-related cognitive impairment and affective symptoms, possibly due to axonal damage to white matter tracts connected with the ACC.

P563 / #1734


SEEING THE WOODS FOR THE TREES? DEMENTIA WITH LEWY BODIES PRESENTING AS POSTERIOR CORTICAL ATROPHY

Lecture Title:

M. Kelly, S. O'Dowd Tallaght University Hospital, Neurology, Dublin, Ireland

Aims: Posterior cortical atrophy (PCA) is a clinico-radiological syndrome characterised by progressive decline in visual processing and other posterior cognitive functions, relatively intact memory and language in the early stages, and atrophy of posterior brain regions. Often considered a “visual variant” of Alzheimer’s disease, a number of other pathological substrates are recognised. DLB is the second most common neurodegenerative disease and increasingly a number of non-parkinsonism dominant phenotypes are recognised. In this case series we aim to highlight the clinical entity of "PCA-Plus (DLB)", focusing particularly on the role played by modern imaging techniques and CSF biomarkers in diagnosis. Methods: We present a series of four patients presenting with PCA and apply recent consensus criteria1 to diagnose the underlying neurodegenerative condition, utilising DAT- and PET- imaging and CSF biomarkers where appropriate. Results: 3 cases meet criteria for "Probable PCA-Plus (DLB)" and one "Possible PCA-Plus (DLB)". Conclusions: We review the current classification of PCA and highlight the importance of deep clinico-radiological phenotyping in neurodegenerative disease to guide targeted interventions and establish future trial-ready cohorts. 1 Crutch SJ et al. Alzheimers Dement. 2017 Aug; 13(8): 870–884

P564 / #1039


A SUBCORTICAL VARIANT OF DEMENTIA WITH LEWY BODIES?

Lecture Title:

N. Philippi1,2, H. Durand1, V. Noblet2, A. Botzung1, F. Blanc1,2 1University Hospital of Strasbourg, Geriatrics And Neurology Services, Research And Resources Memory Center (cmrr), Strasbourg, France, 2CNRS and University of Strasbourg, Icube Laboratory, Umr 7357, Strasbourg, France

Aims: The objective of the present work is to discuss the existence of a subcortical variant of dementia with Lewy bodies (DLB) based on two previous study. In the first study, the objective was to test the existence of a subcortical atrophy, contrasting with overall preserved neocortex in DLB patients. In the second study, we described a series of DLB patients with an initial presentation suggestive of normal pressure hydrocephalus (NPH). Methods: In the first study, we included 55 DLB patients and compared their cerebral volumes to those of 51 AD patients and to those of 17 matched healthy control subjects (CS). We examined the ratio of the TVV against the total cortical volume (Cortex), extracted using freesurfer. In the second study, we retrospectively compared the profile of 21 DLB patients fulfilling the criteria of possible NPH on initial presentation, to those of 13 patients with probable NPH. Results: In the first study, we found that DLB patients had a significantly higher TVV/Cortex ratio compared to both CS and AD patients. In the second study, we highlighted that some DLB patients have a clinical and radiological profile suggestive of NPH. DLB patients had a marked subcortical atrophy, with not significance difference in the dilatation of lateral ventricles and third ventricle compared to HPN patients, though they had a less acute callosal angle and less fourth ventricular dilatation. Conclusions: We suggest the existence of a subcortical variant of DLB, with a clinical presentation and marked subcortical atrophy suggestive of NPH.

P565 / #229


OLFACTORY DYSFUNCTION AND SEVERITY OF TREMOR - IS THERE A CONNECTION?

Lecture Title:

D.D. Pokhabov1, M. Tunik1, V. Abramov2, M. Sadovsky3, D. Pokhabov1 1Krasnoyarsk state medical university, Neurological Diseases, Krasnoyarsk, Russian Federation, 2Siberian clinical center of FMBA, Neurological Diseases, Krasnoyarsk, Russian Federation, 3Institute of computational modeling of SB RAS, Informatics And Modern Technologies, Krasnoyarsk, Russian Federation

Aims: Neurological disorders is a problem for a number of people worldwide. Parkinson's disease (PD) and essential tremor (ET) are among them. Tremor is the common symptom both for PD (rest tremor) and ET (postural-kinetic tremor). Olfactory dysfunction is stipulated to be the first manifest of PD preceding the movement disorders. It happens due to the neurodegenerative process starting in olfactory bulbs. Currently available data on the olfactory function in ET are insufficient and some of them are contradictory. Hence, our aim was to obtain tremor data of PD and ET patients and the data of their olfactory function to verify our scientific hypothesis on the inverse relation between tremor manifestation and olfactory function decay: slighter tremor is accompanied with worse smell perception, and vice versa. Methods: We had three groups of patients: PD, ET and healthy people. An examination procedure of olfactory function was based on extended olfactory Sniffin' sticks test. For tremor testing we used wireless device to monitor electrophysiological signals. We used elastic map technique to cluster and analyze all data. Results: Combination of tremor data and smell perception provides clear and apparent distinction of PD patients from ET ones. Proven inverse relation between tremor level and olfactory function decay is the most sounding result of our work. Indeed, ET patients showed better olfactory function results accompanied by severe tremor, as compared to PD patients. Conclusions: The presented results could be implemented for early differential diagnostics of PD and ET, as well as for the improvement of individual therapy for such patients.

P566 / #769


ΑSYN OLIGOMERS DETECTION BY PROXIMITY LIGATION ASSAY IN SKIN BIOPSIES IS A BIOMARKER FOR PARKINSON’S DISEASE

Lecture Title:

E. Vacchi1,2, C. Senese2, A. Kaelin-Lang1,2,3,4, G. Melli1,2,3 1Università della Svizzera Italiana (USI), Faculty Of Biomedical Sciences, Lugano, Switzerland, 2Ente Ospedaliero Cantonale (EOC), Laboratory For Biomedical Neurosciences, Lugano, Switzerland, 3Ente Ospedaliero Cantonale (EOC), Neurology Department, Lugano, Switzerland, 4University of Bern, Department Of Neurology, Bern, Switzerland

Aims: Peripheral nervous system is involved at the early phases of Parkinson disease (PD), and pathological alpha-synuclein (αSyn) has been detected in skin autonomic nerve fibers of PD patients. By proximity ligation assay (PLA), we assessed the presence of oligomeric αSyn in skin nerve fibers of PD and atypical parkinsonisms (AP). Methods: We enrolled 30 idiopathic PD, 9 Multiple system atrophy (MSA), 11 AP with tauopathies (AP-TAU) and 20 age-matched healthy controls (HC). A double 3mm punch skin biopsy was performed at two anatomical sites (cervical and ankle) and skin sections were double-stained with PLA for oligomeric αSyn and with PGP9.5, a panaxonal marker. Results: PD and MSA patients showed higher signal compared to HC and AP-Tau; in particular, PD showed higher signal especially at the cervical site while MSA at the ankle site. Semi-quantitative analysis revealed higher amount of co-localization signals in PD and MSA, and highlighted a high variability within PD group. The combination of co-localization and dotted signal allowed an accurate discrimination between PD and other groups, in particular between PD and HC (sensitivity 77%, specificity 80%) and AP-Tau (sensitivity 82%, specificity 80%). Conclusions: Our data provide evidence that αSyn oligomers detection by PLA in skin biopsy identify PD. This finding endorses the hypothesis that αSyn oligomers analysis in skin biopsies could be used as a reliable diagnostic biomarker for PD living patients.

P567 / #1121

Topic: Theme C: α-Synucleinopathies / C5.b. Genetics, Epidemiology: Disease-causing mutations

PD GENERATION DURING COVID-19: TRANSITIONING TO REMOTE RECRUITMENT FOR GENETIC COUNSELING AND TESTING

Lecture Title:

R. Alcalay1, A. Naito2, A. Hall2, K. Marder1, M. Nance3, M. Schwarzschild4, T. Simuni5, A.-M. Wills4, L. Cook6, J. Schulze6, J. Verbrugge6, C. Casaceli7, E. Mackenzie2, J. Beck2 1Columbia University Vagelos College of Physicians and Surgeons, Neurology, New York, United States of America, 2Parkinson's Foundation, Research, New York, United States of America, 3Park Nicollet Struthers Parkinson's Center, Neurology, Minneapolis, United States of America, 4Massachusetts General Hospital, Neurology, Charlestown, United States of America, 5Northwestern University Feinberg School of Medicine, Department Of Neurology, Chicago, United States of America, 6Indiana University School of Medicine, Medical And Molecular Genetics, Indianapolis, United States of America, 7University of Rochester, Clinical Trials Coordination Center, Rochester, United States of America

Aims: Social distancing and safety requirements during COVID-19 has severely affected the recruitment to clinical research. The Parkinson’s Foundation PD GENEration (NCT04057794) aims to work with clinicians to provide access to genetic testing and counseling for people with Parkinson’s disease (PD). Here, we describe the conversion of an in-person protocol to include remote recruitment, the obstacles overcome, and the accomplishments in its implementation. Methods: The pilot stage of PD GENEration includes six sites in the USA. We invite people with PD interested in obtaining information on their genetic status to participate. CLIA-certified genetic testing is performed on SNCA, PRKN, LRRK2, GBA, PINK1, PARK7 and VPS35 genes. During the recruitment visit, participants are randomized to receiving results from their local site (clinician or genetic counselors), or remotely by a genetic counseling core at Indiana University. Recruitment was halted because of COVID-19. Results: To continue, we implemented a remote recruitment arm with altered workflow. Changes are as follows: pre-screening questionnaire is completed online, consent is signed electronically, DNA is collected using a buccal swab, and MoCA and MDS-UPDRS are conducted remotely. In addition, all genetic counseling is conducted remotely. Licensure challenges due to participants located across state lines have led to local participants being randomized as before, but now out-of-state participants are directed to centralized genetic counseling. As of 09OCT2020 and six weeks from implementation, 93 have newly enrolled with 48 via telemedicine.

Conclusions: We demonstrate that, in spite of obstacles, remote recruitment for genetic testing and counseling for PD is feasible and popular. Preliminary findings from up-to-date genotyping will be presented.

P568 / #1671

Topic: Theme C: α-Synucleinopathies / C5.c. Genetics, Epidemiology: GWAS, genetic associations, susceptibility & protective genes

COMT POLYMORPHISM IS NOT ASSOCIATED WITH LEVODOPA-INDUCED DYSKINESIA DEVELOPMENT IN RUSSIAN PATIENTS WITH PARKINSON'S DISEASE

Lecture Title:

G. Akhmadeeva1, I. Khidiyatova2, I. Gilyazova2, G. Tayupova1, S. Umutbaev3, A. Baitimerov4, R. Magzhanov5 1Clinical Institute of Neurology and Rehabilitation, National Medical Holding Limited Liability Company «medstandart», Уфа, Russian Federation, 2Institute of Biochemistry and Genetics, Ufa Research Study Center, Ufa, Russian Federation, 3State budgetary healthcare institution Kuvatov Republican clinical hospital, State Budgetary Healthcare Institution Kuvatov Republican Clinical Hospital, Уфа, Russian Federation, 4Clinical Institute of Neurology and Rehabilitation, National Medical Holding Limited Liability Company «medstandart», Ufa, Russian Federation, 5Bashkir State Medical University Ministry of Healthcare of the Russian Federation, Bashkir State Medical University Ministry Of Healthcare Of The Russian Federation, Ufa, Russian Federation

Aims: We examined the influence of rs4680 polymorphism of COMT gene on levodopa-induced dyskinesia (LID) development in Parkinson’s disease (PD) patients. Methods: Our prospective 10 years clinical study included 320 PD patients. Among them there are 80 PD patients completing dyskinesia evaluation using of MDS-UPDRS scale (parts IV). PD patients were divided into PD with LID (PD-LID) and PD without LID (PD-NORM). The analysis of rs4680 polymorphism of COMT gene using was performed using Taq-man assays. Linear regression analysis and one-way ANOVA test are used. Results: There were no statistical differences in the distribution of age and sex between PD-LID patients and PD-NORM patients (Table 1). Linear regression analysis demonstrated increased LID risk for age of onset PD, duration and total daily dose of levodopa (levodopa equivalent dose, LED) (Table 1).

No differences in rs4680 genotype or allele frequencies were observed between PD-LID and PD-NORM patients (Table 2).

Table 1. Demographic characteristics of PD-LID and PD-NORM patients PD-LID PD-NORM P-value

Sex (male/female) 22/33 6/19 0,361

Age of onset PD (year) 54,98±8,79 59,42±5,7 0,039

Duration of levodopa (year) 9,03±3,95 6,88±3,14 0,021

LED (mg) 1112,03±567,88 856,3±380,07 0,001

Conclusions: Few studies examining associations between rs4680 and LID showed inconsistent results: only one study considering the impact potential confounders (the duration and dosage of dopaminergic therapy) confirmed the presence of this relation. This study suggested that rs4680 of COMT gene is not a genetic risk factor for levodopa-induced dyskinesia development. This study will be continued on expanded samples. The study was supported by RFBR grant No.19-015-00331

P569 / #1327


A META-ANALYSIS OF GWAS IN DEMENTIA WITH LEWY BODIES

Lecture Title:

E. Gibbons1, A. Rongve2, A. Ruiz3, O. Andreassen4, O.B.O. Demgene Network5, O.B.O. European Dlb Consortium6, O.B.O. Genomic Research At Fundació Ace3, O.B.O. International Dlb Genetics Consortium7, J. Bras1 1Van Andel Research Institute, Center For Neurodegenerative Science, Grand Rapids, United States of America, 2University of Bergen/European DLB Consortium, Department Of Clinical Medicine, Bergen, Norway, 3Fundació ACE, Institut Català de Neurociències Aplicades - Universitat Internacional de Catalunya, Research Center And Memory Clinic, Barcelona, Spain, 4University of Oslo, Norwegian Centre For Mental Disorders Research (norment), Institute Of Clinical Medicine, Oslo, Norway, 5DemGene Network, Demgene Network, Oslo, Norway, 6European DLB Consortium, European Dlb Consortium, Bergen, Norway, 7International DLB Genetics Consortium, International Dlb Genetics Consortium, Grand Rapids, United States of America

Aims: Dementia with Lewy bodies (DLB) is the second most common form of dementia and has phenotypic overlaps with both Alzheimer’s (AD) and Parkinson’s (PD) diseases. DLB is far from well characterized genetically, largely due to lack of familial cases and difficulty in recruiting large, reliable cohorts given the high rate of misdiagnosis. Methods: In this study, we conduct the largest meta-analysis of genome-wide association studies performed to date on 3,306 DLB cases and 8,500 controls. Results: We confirm reported associations and find supporting evidence for a previously suggestive signal on chromosome 16. In addition, we analyze this association alongside previously reported AD and PD findings to highlight genetic similarities and differences between the three diseases. Conclusions: We report evidence for potential differences in the regulatory mechanisms that distinguish DLB, as well as common trait correlations between the three diseases.

P570 / #1058


FUNCTIONAL SCREENING OF CANDIDATE GENES FOR EARLY-ONSET RECESSIVE PARKINSON’S DISEASE

Lecture Title:

J. Täger1, V. Drouet2, O. Corti2, M. Sakshi-Bedi1, S. Mazzitelli1, J.-C. Corvol3, S. Lesage2, P. Heutink1, A. Brice2 1German Center for Neurodegenerative diseases, Genome Biology Of Neurodegenerative Diseases, Tübngen, Germany, 2Centre de Recherche de l'Institut du Cerveau et de la Moelleépinière, Institut National De La Santé Et De La Recherche Médicale, Paris, France, 3Hôpital Pitié-Salpêtrière, AP-HP, Faculté de Médecine de Sorbonne Université, UMR S 1127, Inserm U 1127, and CNRS UMR 7225, and Institut du Cerveau et de la Moëlle épinière, Neurology, Paris, France

Aims: Early-onset Parkinson’s disease (EOPD) is characterized by an age of onset between 20 and 40 years but similar to late-onset PD, a substantial portion of the genetic component remains at present undefined. Based on the “French Parkinson’s Disease Genetics Study Group” network, a unique collection of autosomal recessive and EOPD families, whole exome sequencing in 77 cases and 22 relatives identified 27 genes with potential pathogenic mutations. However, most genes were not replicated in other PD cases and therefore need functional data to support their pathogenicity in EOPD. Methods: BE(2)-M17 cells cultured in an automated cell culture system were transduced with lentiviral vectors encoding shRNAs targeting individual candidate genes. For each candidate gene we aimed to use three different shRNA vectors. The knockdown efficiency was determined using RT-qPCR. By using mRNA-Seq, the effect of each target gene knockdown on disease-relevant pathways was investigated. Additionally, two microscopic assays for interrogating effects following candidate gene knockdown on early processes of mitophagy or mitochondrial morphology and a plate reader-based assay was performed to determine mitochondrial ATP production. Results: One gene was identified as hit in three assays causing a reduction of mitochondrial number and increase in mitophagy and ATP production while additional eight genes were hits in single assays. The mRNA sequencing identified alterations in mitochondrial pathways supporting an involvement of candidate genes in mitochondrial function. Conclusions: The screening of PD-relevant phenotypes successfully provided us with nine candidate genes related to EOPD which need to be confirmed in secondary screenings.

P571 / #1451

Topic: Theme C: α-Synucleinopathies / C5.d. Genetics, Epidemiology: Aging

DNA METHYLATION MITOTIC AGING: ACCELERATED PROLIFERATIVE HISTORY OF HEMATOPOIETIC CELLS IN PARKINSON’S DISEASE

Lecture Title:

K. Paul1, A. Binder2, S. Horvath3,4, Q. Yan1, I. Del Rosario1, J. Bronstein5,6, B. Ritz1 1University of California, Los Angeles, Department Of Epidemiology, Los Angeles, United States of America, 2University of Hawaii Cancer Center, Population Sciences In The Pacific Program, Honolulu, United States of America, 3University of California, Los Angeles, Department Of Human Genetics, Los Angeles, United States of America, 4University of California, Los Angeles, Department Of Biostatistics, Los Angeles, United States of America, 5University of California, Los Angeles, Department Of Neurology, Los Angeles, United States of America, 6University of California, Los Angeles, Brain Research Institute, Los Angeles, United States of America

Aims: A DNA methylation (DNAm) biomarker, named the epiTOC mitotic clock, has been developed to measure the proliferative history of blood[1]. We hypothesize that Parkinson’s disease (PD) patients exhibit an accelerated hematopoietic mitotic tick rate relative to controls, which is reflective of chronic, low-level systemic immune activation. We aim to investigate whether an acceleration of the DNAm mitotic tick rate is associated with PD susceptibility or longitudinal symptom progression. Methods: We estimated the DNAm-based mitotic age of blood in a population-based study of 569 PD patients and 228 controls using the epiTOC model and 450k DNAm data[1]. We regressed the DNAm mitotic age on chronologic age to remove the variation explained by age (termed AccelEpiTOC). We used logistic regression to estimate PD risk and controlled for blood cell composition along with standard covariates. To assess progression, we relied on 336 PD patients with repeated examinations for the UPDRS-III and MMSE (2-4 follow-up exams, mean follow-up=4.7 years (SD=2.8)), and repeated-measures linear mixed models reporting the interaction between AccelEpiTOC*time. Results: We found the PD patients exhibited a significantly more accelerated mitotic tick rate in blood than controls (AccelEpiTOC OR per SD=2.06, 95% CI=1.52, 2.79, p=2.6e-6). AccelEpiTOC was also associated with longitudinal cognitive decline over follow-up (per year, β=-0.06 (SE=0.02), p=0.01) and suggestively faster development of motor symptoms as measured by the UPDRS-III (β=0.19 (SE=0.11), p=0.08). Conclusions: The current study provides a first look and striking findings implicating accelerated hematopoietic cell mitosis, possibly reflecting imbalances in immune pathways, in PD pathogenesis and progression.

P572 / #1469

Topic: Theme C: α-Synucleinopathies / C5.e. Genetics, Epidemiology: Environmental risk factors

RELATIONSHIP BETWEEN SMOKING, ALCOHOL, CAFFEINE INTAKE AND PARKINSONISM IN A POPULATION-BASED SAMPLE AGED 75+ YEARS – PIETÀ STUDY.

Lecture Title:

M. Barbosa1, T. Vale2, E. Resende1, H. Guimarães1, J.C. Machado3, M. Almeida1, F. Cardoso4, P. Caramelli1 1Ambulatório de Neurologia Cognitiva e do Comportamento - Hospital das Clínicas da UFMG, Internal Medicine Dept, Belo Horizonte - MG, Brazil, 2Universidade Federal de Juiz de Fora - Minas Gerais, Neurologia, Juiz de Fora - MG, Brazil, 3Faculdade Ciências Médicas de Minas Gerais, Geriatric Medicine, Belo Horizonte - MG, Brazil, 4Ambulatório de Distúrbios de Movimento - Hospital das Clínicas da UFMG, Internal Medicine Dept, Belo Horizonte - MG, Brazil

Aims: Many studies have found protective effect of smoking and caffeine consumption on incidence of PD. Alcohol intake has been associated with reduced PD risk, although heavy use was related with increased risk. Objective: To investigate the association of cigarette smoking, alcohol and caffeine intake with parkinsonism (PS) and Parkinson’s disease (PD) in a population-based sample of elderly 75+ years. Methods: Cross-sectional analysis of data from individuals with PS diagnosis (n=64) and controls (n=556) from the Pietà study, a community-based investigation on brain aging, carried out in Caeté, southeast Brazil. Subjects underwent a thorough functional, clinical, psychiatric and neurological evaluation, responded to a questionnaire about previous habits of life, including consumption of caffeine, alcohol and cigarette smoking. Results: Sample was comprised mostly by women (63.9%), median of three years of schooling. Median age was significantly different for the groups: 82 for the PS vs. 80 for controls (p<0.001). Caffeine consumption was inversely associated with PD diagnosis [OR = 4.567, CI 95% 1.388-15.022; p=0.003]. Previous intake of 2+ cups/day was more frequent in controls comparing to PS individuals (p=0.03). We did not find an association between cigarette smoking and occurrence of PS or PD. For alcohol consumption, no drinking was more frequent in the control group and taking 5+ drinks during the week was associated with undetermined PS (n=14; p<0.01). Conclusions: Caffeine intake in this community-dwelling elderly population was more frequent in controls comparing to PS subjects. We found relationship between higher alcohol intake and PS. We could not find association between cigarette smoking and PS/PD.

P573 / #849


A53T -MUTATED HUMAN Α-SYNUCLEIN IS INVOLVED IN BLOOD GLUCOSE REGULATION OF WESTERN TYPE DIET-FED PARKINSON’S DISEASE MICE

Lecture Title:

M. Daurer1, S. Flunkert1, R. Rabl1, H. Roemer2, B. Hutter-Paier1 1QPS Austria GmbH, Neuropharmacology, Grambach, Austria, 2Karl-Franzens University of Graz, Institute Of Zoology, Graz, Austria

Aims: Besides genetic modifications, diabetes and obesity have been suggested to be a risk factor for the development and progression of Parkinson’s Disease (PD). To better understand the impact of diabetes and obesity caused by diet and PD, we focused on investigating the effects of a Western type diet (WTD) that contains high fat and sucrose, as well as supplemented cholesterol on a PD mouse model. Methods: To this end, genetically modified A53T mice, overexpressing A53T-mutated human α-Synuclein and non-transgenic (NTG) littermates were fed a WTD or control diet (CD) starting at 3.5 months of age. To monitor the animals’ response to the diet, the food intake, and body weight were continuously evaluated. Additionally, glucose metabolism was investigated using the glucose tolerance test after 5, 11, and 26 weeks on the diet. Results: Both, NTG and A53T mice on WTD had a higher caloric intake which resulted in an elevated weight gain compared to CD-fed control animals of both genotypes. Intriguingly, fasting blood glucose levels were increased in NTG mice but not in A53T animals on WTD. Injecting a glucose solution intraperitoneally led to higher blood glucose levels in WTD-fed A53T and NTG mice 5 and 11 weeks after starting the dietary intervention. However, after 26 weeks, only NTG mice demonstrated elevated glucose levels. Conclusions: In summary, our findings add evidence indicating an impact of A53T-mutated human α-Synuclein on glucose regulation. Further research on this topic might pave the road for new therapeutic approaches in the field of diabetes and PD.

P574 / #1479


DNA METHYLATION BIOMARKER FOR CUMULATIVE LEAD EXPOSURE IS ASSOCIATED WITH PARKINSON’S DISEASE

Lecture Title:

K. Paul1, S. Horvath2,3, I. Del Rosario1, J. Bronstein4,5, B. Ritz1 1University of California, Los Angeles, Department Of Epidemiology, Los Angeles, United States of America, 2University of California, Los Angeles, Department Of Human Genetics, Los Angeles, United States of America, 3University of California, Los Angeles, Department Of Biostatistics, Los Angeles, United States of America, 4University of California, Los Angeles, Department Of Neurology, Los Angeles, United States of America, 5University of California, Los Angeles, Brain Research Institute, Los Angeles, United States of America

Aims: Lead, a known neurotoxicant, has previously received attention in Parkinson’s disease (PD), but findings in epidemiologic studies have been equivocal. Two DNA methylation-based biomarkers for cumulative lead exposure measured in the tibia and patella have recently been developed. Using these biomarkers for the first time, we aim to evaluate the influence of cumulative lead exposure on PD risk in two, independent population-based studies with over 2600 participants. Methods: For our analysis, we used genome-wide DNA methylation data from two publicly available (GEO) PD studies, with 1528 PD patients and 1168 controls. We generated two epigenetic biomarkers for cumulative lead exposure (tibia and patella), developed in the Normative Aging Study. We modeled PD with logistic regression. Results: PD status was strongly associated with the DNAm biomarker for tibia-lead levels in both cohorts. Specifically, per unit DNAm tibia-lead increase, in the two cohorts, we estimated an OR for PD of 2.06 (95% CI=1.66, 2.56; p=5.4E-11) and of 1.60 (95% CI=1.20, 2.15; p=0.001), with a meta-OR of 1.89 (95% CI=1.59, 2.24; p=8.1E-13). The DNAm biomarker for patella-lead was not associated with PD. Conclusions: In two large population-based PD studies, we estimated cumulative, bone-lead levels applying an externally developed epigenetic biosensor, and found estimated tibia-lead levels to be strongly associated with an increased risk of PD, but not patella-lead. The weight of evidence from previous and our current study supports a positive association between tibia-measured lead and PD, representing lead exposure that extended over many decades rather than shorter-term exposures reflected in patella-lead levels.

P575 / #1255

Topic: Theme C: α-Synucleinopathies / C5.g. Genetics, Epidemiology: Other

INCREASED MENOPAUSAL AGE BASED ON A POLYGENIC SCORE DECREASES PREVALENCE OF PARKINSON’S DISEASE

Lecture Title:

C. Kusters1, K. Paul2, A. Duarte Folle2, A. Keener3,4, J. Bronstein3,5, L. Bertram6,7, J. Hansen8, S. Horvath1,9, J. Sinsheimer1,9,10, C. Lill11,12, B. Ritz2,3,13 1University of California, Los Angeles, Department Of Human Genetics, Los Angeles, United States of America, 2University of California, Los Angeles, Department Of Epidemiology, Los Angeles, United States of America, 3University of California, Los Angeles, Department Of Neurology, Los Angeles, United States of America, 4Greater Los Angeles Veterans Affairs Medical Center, Parkinson's Disease Research, Education, And Clinical Center, Los Angeles, United States of America, 5University of California, Los Angeles, Brain Research Institute, Los Angeles, United States of America, 6University of lubeck, Lubeck Interdisciplinary Platform For Genome Analytics, Institutes Of Neurogenetics & Cardiogenetics, Lubeck, Germany, 7University of Oslo, Department Of Psychology, Centre For Lifespan Changes In Brain And Cognition, Oslo, Norway, 8Danish Cancer Society, Danish Cancer Society Research Center, Copenhagen, Denmark, 9University of California, Los Angeles, Department Of Biostatistics, Los Angeles, United States of America, 10University of California, Los Angeles, Department Of Computational Medicine, David Geffen School Of Medicine, Los Angeles, United States of America, 11University of Lübeck and University Medical Center Schleswig-Holstein,, Section For Translational Surgical Oncology And Biobanking, Department Of Surgery, Lübeck, Germany, 12Imperial College, Ageing Epidemiology Research Unit, School Of Public Health, London, United Kingdom, 13University of California, Los Angeles, Department Of Environmental Health, Los Angeles, United States of America

Aims: Sex hormones may protect dopaminergic neurons, possibly preventing or delaying the onset of Parkinson’s disease (PD). Studies of PD and age of menarche or menopause as indicators of lifelong hormone exposure reported inconsistent findings, possibly due to difficult reporting accurate assessments. Here we employed polygenic scores (PGS) for age at menopause and age at menarche that depend on self-report to estimate PD risk. Methods: Relying on two population-based studies (PASIDA, Denmark, and PEG, USA) that enrolled 1,739 female subjects (805 PD patients, 934 controls) of European ancestry, we assessed associations between PD status and PGS constructed based on large external GWAS (UK biobank) on menopause and menarche age, respectively. In addition, we also employed two additional PGS, for PD and for resting heart rate, as positive and negative controls, respectively. Results: In our populations, the menopause and menarche PGS, which were constructed based on independent GWAS data, were associated with the respective age of menopause or menarche. We observed a lower risk of developing PD with each standard deviation increase of the PGS for age of menopause (OR: 0.88, 95%CI: 0.79-0.98). There was no association between the age at menarche PGS and PD. As expected, the PD risk PGS was and the resting-heart-rate PGS was not associated with PD. Conclusions: A genetically predicted later age at menopause was associated with a decreased risk of PD in women, supporting the hypothesis that sex hormones may be neuroprotective in PD.

P576 / #745

Topic: Theme C: α-Synucleinopathies / C6.a. Cell, Molecular and Systems Biology: Αlpha-synuclein

CELLULAR SENESCENCE MEDIATES THE PROPAGATION OF ALPHA-SYNUCLEIN

Lecture Title:

E.-J. Bae1, S.-J. Lee2 1Seoul National University, College of Medicine, Neuroscience Research Center, Seoul, Korea, Republic of, 2Seoul National University, Biomedical Sciences, Seoul, Korea, Republic of

Aims: In Parkinson disease, α-synuclein pathology initiates in specific brain regions and its expansion to broader areas follows. Although the mechanism of the progression of the disease has not been fully unfolded, accumulating evidence advocate propagation of α-synuclein as its root principle. In previous study, we demonstrated that the propagation of α-synuclein aggregates was a continuous process in which a cycle of events repeats: internalization of extracellular aggregates, co-aggregation between the internalized and endogenous α-synuclein, and secretion of these co-aggregates. However, the mechanism of propagation of α-synuclein remains unclear. Given that autopsy studies of the brans from aged people who had not been diagnosed with a neurological disease reported the presence of pathological protein aggregates, we hypothesized that cellular senescence could regulate the propagation of α-synuclein. Methods: Here, we established cellular senescence model in primary cortical neurons and monitored the senesence markers which showed positive correlation with the secretion of α-syuclein aggregates. Results: Aged neurons showed an alterations in lysosomal compartment and an increase in a secretory phenotypes including Senesecence-Associated Secretory Phenotype (SASP). The secretion of β-hexosaminidase, a maker of lysosomal exocytosis was increased in a age-depenent manner, which shows positive correlation with the secretion of α-synuclein. UC2288, a p21 inhibitor reduced senescence-induced secretion of α-synuclein and β-hexosaminidase. We found that the secretion of α-synuclein and β-hexosaminidase are sensitive to vacuolin-1, an inhibitor of lysosomal exocytosis. Furthermore, inhibiting lysosomal exocytosis caused nearly a total hindrance of the cell-to-cell transmission. Conclusions: These results suggest that cellular senescence accelerates the propagation of α-synuclein, thereby promoting secretion of α-synuclein via lysosomal exocytosis.

P577 / #1300

Topic: Theme C: α-Synucleinopathies / C6.a. Cell, Molecular and Systems Biology: Αlpha-synuclein

ALPHA-SYNUCLEIN AS A MARKER OF TEMPORAL LOBE ATROPHY IN AD? - PRELIMINARY REPORT

Lecture Title:

I. Winkel1, B. Paradowski2, P. Lewczuk3 1Alpha-synuclein as a marker of temporal lobe atrophy in AD? - preliminary report, Department And Clinic Of Geriatrics, Ścinawa, Poland, 2Medical University of Wrocław, Department Of Neurology, Wrocław, Poland, 3Lab for Clinical Neurochemistry, Department Of Psychitary, Erlangen, Germany

Aims: Introduction Alpha-synuclein (ASN) is a protein located mainly in the presynaptic part of nerve endings. In neurodegenerative diseases it is deposited in the form of protein aggregates which are the main component of Lewy's bodies and neurites, also in Alzheimer's disease (AD). Purpose of the study: The aim of the study was to assess the degree of atrophy of the medial structures of the temporal lobe and the level of ASN in the cerebrospinal fluid (CSF) at patients with neurochemically confirmed AD. Methods: Material and methods: The study included 80 patients with the neurochemical diagnosis of AD (including 54 women, 26 men) and 34 (including 18 women, 16 men) as a control group, without signs of dementia. The ASN level in CSF was marked using kits from MSD, Rockville, Maryland, USA. The degree of atrophy of the medial temporal lobe structures was assessed using the MTA visual scale (Medial Temporal lobe Atrophy). Results: The ASN level was statistically significantly higher in the AD group compared to the control group (p<0.001). At patients with MTA = 3 and MTA = 4, a statistically significantly lower ASN level was found compared to patients with MTA = 0. Conclusions: 1. Medial atrophy of the temporal lobe in AD correlated with a decline in ASN. 2. Testing the ASN level may be an indicator of the progressive decline and progression of the disease in AD, but it requires further research on a larger group of patients.

P578 / #1304

Topic: Theme C: α-Synucleinopathies / C6.b. Cell, Molecular and Systems Biology: LRKK2, parkin, PINK1, DJ-1 and other PD related genes

THE ROLE OF LRRK2 AND INFLAMMATION IN AN Α-SYNUCLEIN-BASED MODEL FOR PARKINSON’S DISEASE

Lecture Title:

D. Cabezudo1, V. Van Lieshout1, C. Van Den Haute1,2, A. Van Der Perren1, E. Lobbestael1, V. Baekelandt1 1KU Leuven, Department Of Neurosciences, Laboratory For Neurobiology And Gene Therapy, Leuven, Belgium, 2KU Leuven, Leuven Viral Vector Core, Leuven, Belgium

Aims: Mutations in LRRK2 such as the G2019S are the most common cause of familial Parkinson’s disease, but as suggested by its incomplete penetrance, additional triggers might be necessary for the disease to develop. Recent evidence points to an important function of LRRK2 in inflammatory processes, which leads to the hypothesis that an inflammatory trigger is needed for pathogenic LRRK2 mutations to induce PD. We aim to gain more insight in how inflammation contributes to neurodegeneration in PD and to elucidate the role of LRRK2 in these processes. Methods: We model PD in mice carrying LRRK2 wild-type or G2019S (knock-in) by overexpressing human α-synuclein in the substantia nigra using adeno-associated viral vectors (rAAV). In parallel, we study the effect of inflammatory triggers in these mutant mice. Phenotypic readouts include motor behavior using several tests (cylinder, pole test and hanging wire), neurodegeneration and neuroinflammation analyzed by immunohistochemistry and the presence of immune cells in the brain, investigated by flow cytometry. Results: We found that the presence of the LRRK2 G2019S mutation aggravates dopaminergic cell loss and increases neuroinflammation, both by overexpression of α-synuclein with an rAAV2/7 vector and by systemic inflammatory cues. Conclusions: These results suggest that mutant LRRK2 is involved in PD pathogenesis via its role in immune cells and thus might increases the susceptibility to develop PD by affecting the immune system. Therefore, we are currently investigating how inflammatory triggers accelerate the observed neurodegeneration and neuroinflammation induced by α-synuclein and to what extent pathogenic LRRK2 is involved in these processes.

P579 / #1435


PINK1-PARKIN MITOPHAGY-SPECIFIC DISEASE SIGNATURE

Lecture Title:

F. Fiesel1, Y. Ren1, J. Bredenberg1, Z. Quicksall1, Z. Wszolek2, O. Ross1, W. Springer1 1Mayo Clinic, Neuroscience, Jacksonville, United States of America, 2Mayo Clinic Florida, Neurology, Jacksonville, United States of America

Aims: Mutations in PINK1 and PARKIN are linked to early-onset Parkinson’s disease (PD). The encoded proteins, PINK1, a mitochondrial kinase, and PARKIN, an E3 Ubiquitin ligase, function together in a neuroprotective mitochondrial quality control pathway that is conserved across cell types. PINK1 and PARKIN label damaged mitochondria with phosphorylated Ubiquitin chains and thereby target them for degradation in lysosomes through autophagy (mitophagy). Several studies have shown increased mitochondrial damage and involvement of impaired mitochondrial quality control in PD and it is possible that failure of the PINK1-PARKIN pathway plays a role also for some cases of idiopathic disease. Here we set out to identify a specific disease signature of mitophagy impairment. Methods: We have used a uniquely large cohort of patients’ fibroblasts and genome-edited neuronal cells with loss-of-function mutations in PINK1 and/or PARKIN as model systems for mitophagy impairment to systematically analyze transcriptional and metabolomic changes. Results: We have already identified several significant changes in PINK1-PARKIN mutant cells that might be used as a mitophagy-specific signature. Conclusions: Once fully validated, the list of changes might help to develop biomarker panels and/or to stratify idiopathic PD patients for clinical studies.

P580 / #926


INVOLVEMENT OF LRRK2 IN THE LYSOSOMAL AND EXOSOMAL RELEASE

Lecture Title:

M. Sakurai, T. Kuwahara, T. Eguchi, T. Iwatsubo Graduate School of Medicine, The University of Tokyo, Department Of Neuropathology, Tokyo, Japan

Aims: LRRK2 is a causative gene product of Parkinson’s disease that phosphorylates a subset of Rab GTPases. We have previously shown that LRRK2 and its substrate Rab10 are relocalized onto enlarged lysosomes upon treatment with a lysosomal stressor chloroquine (CQ), and facilitate the exocytic release of lysosomal contents (Eguchi et al, PNAS 2018). We investigated the mechanism whereby LRRK2 and related proteins regulate the release of lysosomal contents under lysosomal stress conditions. Methods: We analyzed the release of lysosomal proteins and exosomes in RAW264.7 cells and primary mouse embryonic fibroblasts (MEFs) under CQ treatment. Calcium-dependent lysosomal exocytosis was analyzed by measuring the activity of b-hexosaminidase, a lysosomal hydrolase, in media. Results: LRRK2/Rab10-mediated exocytic release of cathepsins was detectable in RAW264.7 cells but not in other cells upon CQ exposure, suggesting a cell type-specific mechanism. The knockdown of LRRK2 or Rab10 suppressed the release of exosomes under CQ exposure as well. LRRK2 inhibition did not suppress the calcium-dependent lysosomal exocytosis in primary MEFs treated with ionomycin. Knockdown of autophagy-related proteins ATG5 or ATG16L1 suppressed lysosomal translocation of LRRK2 and exocytic release of cathepsins upon CQ treatment. Conclusions: We revealed the involvement of LRRK2-Rab10 in the release of lysosomal cathepsins and exosomes, by an as yet unknown mechanism that may be distinct to the classical, calcium-dependent, lysosomal exocytosis. The clarification of the relationship between the ATG proteins and the LRRK2-Rab10 pathway may be key to the understanding of the lysosomal stress-related release mechanism.

P581 / #1037

Topic: Theme C: α-Synucleinopathies / C6.f. Cell, Molecular and Systems Biology: Metabolomics, transcriptomics, lipidomics, proteomics

HUMAN-SPECIFIC TRANSCRIPTOMIC PROFILING OF VENTRAL AND DORSAL MIDBRAIN DOPAMINE NEURONS

Lecture Title:

L. Parkkinen1, J. Monzón-Sandoval2, I. Poggiolini1, R. Wade-Martins3, C. Webber2 1University of Oxford, Nuffield Department Of Clinical Neurosciences, Oxford, United Kingdom, 2Cardiff University, Uk Dementia Research Institute, Cardiff, United Kingdom, 3University of Oxford, Department Of Physiology, Anatomy And Genetics, Oxford, United Kingdom

Aims: To identify intrinsic genetic differences that account for the differential vulnerability of the neurons in substantia nigra pars compacta (SNpc) in Parkinson’s brain. Neuronal loss in the SNpc in Parkinson’s brain is not uniform as dopamine neurons from the ventral (vulnerable) tier are lost more rapidly than those of the dorsal (resistant) tier. Methods: We compared the transcriptional profiles of ~100 laser captured micro-dissected SNpc neurons from each tier obtained from seven healthy brain donors. Results: Expression levels of dopaminergic markers were similar across the tiers while markers specific to the neighbouring ventral tegmental area were virtually undetected. After accounting for unwanted sources of variation, we identified 106 differentially expressed genes (DEGs) between the tiers. The genes upregulated in the dorsal/resistant SNpc tier neurons displayed coordinated patterns of expression, their protein products had more interactions than expected by chance and they demonstrated evidence of functional convergence. No significant shared functionality was found for genes upregulated in the ventral/vulnerable SNpc tier. Surprisingly, none of the identified DEGs were among the familial genes or genome-wide associated loci. Finally, we found some DEGs in opposite tier orientation between human and analogous mouse populations Conclusions: Our results highlight functional enrichments of vesicular trafficking, ion transport/homeostasis and oxidative stress genes showing higher expression in the resistant SNpc dorsal neurons. Furthermore, the comparison of gene expression variation in human and mouse SNpc populations strongly argues for the need of human-focused Omics studies

P582 / #1059

Topic: Theme C: α-Synucleinopathies / C6.g. Cell, Molecular and Systems Biology: Epigenetics, histone modification, DNA methylation

PARKINSON’S DISEASE ALTERS GENOME-WIDE DNA METHYLATION AND DNA HYDROXYMETHYLATION IN HUMAN BRAIN

Lecture Title:

A. Bernstein, N. Kuhn, J. Kochmanski Michigan State University, Translational Neuroscience, Grand Rapids, United States of America

Aims: 5-methylcytosine (5-mC) and its oxidized form, 5-hydroxymethylcytosine (5-hmC) are distinct, stable epigenetic marks that have separate genomic distributions and regulatory functions in the brain. Existing work has shown associations between DNA methylation and Parkinson’s disease (PD), but little work has examined the potential role for DNA hydroxymethylation in the context of neurodegenerative disease. Methods: We utilized human post-mortem parietal cortex from the Banner Sun Health Research Institute Brain Bank from control (n=50) and PD (n=50) patients to identify differences in 5-mC and 5-hmC between control and diseased brains. Neuronal populations were enriched using magnetic-assisted cell sorting for the neuronal marker, NeuN. Genome-wide 5-mC and 5-hmC were measured using the Illumina EPIC array paired with bisulfite treatment and oxidative bisulfite treatment. We used our novel pipeline of mixed-effects modeling to identify cytosines that showed altered 5mC and 5hmC by disease status. Results: We identified a large number of cytosines with paired changes in 5-mC and 5-hmC that are not identified when this data was analyzed separately. Gene ontology and network analysis identified pathways important for neuronal development and maintenance, as well as synaptic function. Conclusions: We found significant effects of PD status on the balance between 5-mC and 5-hmC at the base-pair level in pathways that are important for PD pathogenesis. Our analyses suggest that there are widespread shifts in the balance between 5-mC and 5-hmC at genes relevant to disease etiology that are not captured when 5-mC and 5-hmC are analyzed as distinct datasets.

P583 / #1630

Topic: Theme C: α-Synucleinopathies / C7.a. Animal Models: Transgenic rodents

TAKING A NEW LOOK AT PARKINSON’S DISEASE: RETINAL PHENOTYPING OF A TRANSGENIC ΑLPHA-SYNUCLEIN MOUSE MODEL

Lecture Title:

L. Veys, J. Devroye, L. Cools, S. Mentens, M. Christiaens, G. Gelders, L. Moons, L. De Groef Neural Circuit Development and Regeneration Research Group, KU Leuven, Biology, Leuven, Belgium

Aims: Because many PD patients present with visual disabilities and the eye has unique properties in terms of accessibility and amenability for in vivo imaging/electrophysiology, the eye is an attractive organ for clinical research on Parkinson’s disease (PD) symptoms and potential biomarkers in the eye, and for fundamental research into PD disease mechanisms. Therefore, we characterized PD-related retinal manifestations in (Thy-1)-h[A30P]alpha-synuclein mice. Methods: (Thy-1)-h[A30P]alpha-synuclein and WT mice were studied using (immuno)histological analyses, complemented with in vivo optical coherence tomography (OCT) and electroretinograms. Results: In transgenic yet not in WT animals, immunohistochemistry revealed prominent αSYN expression in somata and neurites in the nerve fiber layer, ganglion cell layer, and inner plexiform layer; as well as in cell bodies in the inner nuclear layer. A fraction of this αSYN was phosphorylated and found in sparse cell bodies in the ganglion cell layer, yet after staining for ThioS, aggregates were not detected. OCT imaging revealed thinning of the retina from 15 months onwards in transgenic mice as compared to WT animals. Despite this retinal atrophy, we could not pinpoint specific cell types that account for this thinning, when counting dopaminergic or melanopsin+ cells. However, functional changes underscore these OCT abnormalities: electroretinograms of transgenic animals revealed oscillatory potential differences, and, at later ages, pSTR differences, as compared to WTs. Conclusions: (Thy-1)-h[A30P]alpha-synuclein mice show retinal manifestations including alpha-synuclein depositions, inner retinal atrophy, electrophysiological abnormalities. These findings mirror some retinal manifestations in PD patients, and emphasize that the retina can complement brain research in studying PD.

P584 / #138

Topic: Theme C: α-Synucleinopathies / C7.b. Animal Models: Primates, naturally occuring models

GCI-INDUCED NEURODEGENERATION AND SYNUCLEINOPATHY IN NON-HUMAN PRIMATES

Lecture Title:

B. Dehay1, M. Teil1, S. Dovero1, M.-L. Arotcarena1, M. Bourdenx1, S. Camus1, G. Porras1, M.-L. Thiolat1, N. Kruse2, B. Mollenhauer2, I. Trigo-Damas3, C. Estrada4, N. Garcia-Carrillo5, M.T. Herrero4, P. Derkinderen6, M. Vila7, J. Obeso3, E. Bezard1 1Institute of Neurodegenerative Diseases, Cnrs Umr5293-univ. Bordeaux, BORDEAUX, France, 2Paracelsus-Elena- Klinik, Kassel, University Medical Center Goettingen, Institute Of Neuropathology, Goettingen, Germany, 3HM CINAC, Hm Puerta Del Sur And Ciberned And Ceu-san Pablo University Madrid, Mostoles, Spain, 4Clinical and Experimental Neuroscience Unit, School of Medicine, Biomedical Research Institute of Murcia (IMIB), University of Murcia, Campus Mare Nostrum, Institute Of Research On Aging (iuie), School Of Medicine, University Of Murcia, Murcia, Spain, 5Centro Experimental en Investigaciones Biomedica (CEIB), University Of Murcia, Murcia, Spain, 6Inserm, U1235, Nantes University, Nantes, France, 7Neurodegenerative Diseases Research Group, Vall d’Hebron Research Institute (VHIR)-Center for Networked Biomedical Research on Neurodegenerative Diseases (CIBERNED), Department Of Biochemistry And Molecular Biology, Autonomous University Of Barcelona (uab), Barcelona, Spain

Aims: This study aimed at determining the potential to induce MSA pathology in non-human primates. Aggregation of alpha-synuclein has been implicated in several neurodegenerative diseases, termed synucleinopathies, which include Parkinson’s Disease (PD), Dementia with Lewy Bodies (DLB) and Multiple System Atrophy (MSA). These synucleinopathies are characterized by the deposit of α-synuclein aggregates in intracellular inclusions in neurons and/or glial cells. Unlike in PD and DLB, where these aggregates are located predominantly in neurons, MSA is associated with cytoplasmic inclusions of α-synuclein in oligodendrocytes. These glial cytoplasmic inclusions (GCIs) are the pathological hallmarks of MSA and are associated with neuroinflammation, demyelination and, ultimately, neurodegeneration. Methods: To this end, we inoculated MSA-derived GCI fractions into the striatum of baboon monkeys that were terminated 2 years later. Extensive histochemical and biochemical analyses were performed on the whole brain and biological fluids to evaluate pathological markers known to be affected in MSA. Results: We characterized neurodegeneration in these GCI-injected monkeys by describing the pattern of dopaminergic loss, as well as loss of oligodendrocytes, in the striatum and in the substantia nigra. We also characterized the regional distribution and variations of α-synuclein immunoreactivity in several brain structures, as well as its pathological state. Finally, we described the demyelination, neuroinflammation and occurrence of intracellular inclusion formation in this model. Conclusions: We demonstrated that injection of GCI from MSA patients into the striatum induces nigrostriatal neurodegeneration and oligodendroglial cell loss, synucleinopathy and inflammation in the brain of baboon monkeys.

P585 / #907

Topic: Theme C: α-Synucleinopathies / C7.e. Animal Models: Other

A53T ALPHA-SYNUCLEIN PROMOTES A RAPID NEURODEGENERATION AND MOTOR DEFICIT FOLLOWING AAV VECTOR MEDIATED EXPRESSION IN THE RAT SUBSTANTIA NIGRA

Lecture Title:

T. Burke1,2, L. Ryan1, J. Prenderville2,3, E. Sokolowska2, D. Kozareva1,2, N. Upton4, A. Harkin1 1Trinity College Institute of Neuroscience (TCIN), Neuropsychopharmacology Research Group, Dublin, Ireland, 2Transpharmation Ireland Ltd, Transpharmation Ireland Ltd, Dublin, Ireland, 3Trinity College Dublin School of Medicine, Department Of Physiology, Dublin, Ireland, 4Transpharmation Ltd, Transpharmation Ltd, London, United Kingdom

Aims: Hereditary forms of Parkinson’s disease can be caused by duplication, triplication and missense mutations in the alpha-synuclein encoding gene (e.g. A53T). Adeno-associated viral (AAV) vector mediated expression of alpha-synuclein in different forms provides a means for studying alpha-synuclein-induced motor deficits and pathology in animal models. Attempts with different AAV serotypes generate mixed results. This study investigates the effects of AAV serotype 5 with insert for human wildtype alpha-synuclein (AAV5-CBA-alpha-synuclein1x1013vg/ml) and AAV serotype 1/2 with insert for human A53T alpha-synuclein (AAV1/2-CMV-A53T-alpha-synuclein5x1012vg/ml) delivery to the rat substantia nigra(SN) and assesses motor behavioural deficits and dopamine neuronal cell loss. AAV empty-vector, AAV-enhanced-green-fluorescent-protein(eGFP) and phosphate buffered saline(PBS) were included as controls. Methods: Male and female rats received a unilateral stereotaxic injection into the SN. Vector-mediated expression of alpha-synuclein was assessed at multiple timepoints post-injection, and behavioural tests of motor function and immunohistochemical analysis of dopaminergic cell loss were conducted. Results: AAV5 successfully transduced and expressed alpha-synuclein in nigral dopaminergic neurons 12 weeks post-injection, with forelimb-use asymmetry and impaired skilled forelimb function evident, however there was no significant nigral dopaminergic cell loss. Dopamine cell loss was apparent in the AAV5-eGFP group, compared to both the AAV5 alpha-synuclein and PBS groups. Motor behavioural deficits became evident 3 weeks post-AAV1/2 A53T delivery with alpha-synuclein expression and dopaminergic cell loss evident in the SN 4 weeks post-vector delivery. Conclusions: In conclusion, delivery of AAV1/2 to express human A53T alpha-synuclein in the rat SN provides a rapid and progressive course of alpha-synuclein aggregation, accompanied by dopaminergic nigrostriatal degeneration and associated motor deficits.

P586 / #638


EFFECT OF 6-HYDROXYDOPAMINE AND OVARIECTOMY IN HEART RATE VARIABILITY PARAMETERS OF FEMALE RODENTS.

Lecture Title:

T. De La Rosa Macías1, D. Amado Scerni1, F. Scorza1, A.C. Takakura2, F. Da Costa Souza2, V. Sanabria Calvo1, V. Cassia Gonçalves1 1Universidade Federal de São Paulo, Neurologia-neurociencias, São Paulo, Brazil, 2Universidade de São Paulo, Farmacologia, São Paulo, Brazil

Aims: Validate the heart rate variability parameters of female Wistar after ovariectomy and striatal injection of 6-OHDA. Methods: Female Wistar rats were divided into four groups: 6-OHDA, Sham, 6-OHDA+OVX, Sham+OVX. Ovariectomy was performed in OVX groups. Two weeks after ovariectomy 6-hydroxydopamine was injected bilaterally into the striatum, total dose of 48µg. Forty days after lesion, animals had cardiac electrodes implanted and wired to an output connector fixed to the skull cap. Cardiac ECG signal was recorded and transduced by an amplification circuit and then digitized by the PowerLab v8 decive (ADInstruments). Heart rate variability parameters and R-R interval distribution were processed using LabChart software (ADInstruments). All statistical tests were performed using SPSS 21.0 (IBM). Results: Lesion of the substantia nigra pars compacta dopaminergic neurons after striatal injection of 6-OHDA was greater in the 6-OHDA groups compared to Sham (p<0.001). We also found a significant effect of ovariectomy in reducing dopaminergic lesion (p=0.047). Heart rate variability parameters were compared between groups using non-parametric tests, with no significant results. However, we did found significant effect of ovariectomy and 6-hydroxydopamine in RRi tachogram distribution (K-S p<0.001). Conclusions: 6-hydroxidopamine seems to affect cardiac function in rodents by lesioning central autonomic regions, mimicking some features of dysautonomia observed in Parkinson patients. The mechanisms behind these effects and the relation with female hormones remains elusive, and this results could help shed some light as it’s the first experience testing this using a high dose and long lesion period of 40 days.

P587 / #1730


NEUROPROTECTIVE AND ANTI-APOPTOTIC EFFECTS OF GASTRODIN IN DOPAMINERGIC NEURONS IN 6-ODHA-INDUCED PD MODEL

Lecture Title:

R. Haddadi, N. Ali-Qaradsh Hamadan university of medical sciences, Hamedan, Pharmacology, Hamadan, Iran

Aims: This study aims to evaluate the neuroprotective and anti-apoptotic effects of Gst Intra-Cerebro Ventricular (icv) injection into the SNc of 6-hydroxydopamine (6-OHDA) rats’ model of PD. Methods: In pre-treatment group, i.c.v administration of Gst (20, 40 and 80 μg/3 μl/rat) into SNc of normal Male Wistar rats was performed for five consecutive days, while in the treatment group, 6-OHDA-induced PD rat models (8 μg/2 μl/rat) were microinjected with Gst for successive 10 days. 21 days after 6-ODHA infusion, rats were evaluated for apomorphine rotation test. After that, the neuroprotective effects of Gst on the level of pro-apoptotic factors (Caspase-3, BAX), the neuroinflammation initiators (MYD88, TLR-4), and TH enzyme were analyzed by western blot assay. Results: Based on our findings, the effect of Gst in pre-treatment groups, showed a significant reduction in apomorphine-induced rotation in comparison with 6-OHDA lesion rats. Moreover, the protein levels of BAX, Caspase-3, TLR-4, and MYD-88 in both groups have demonstrated a significant decrease. Interestingly, the level of TH enzyme, as a significant mediator of the dopamine synthesis pathway, face a considerable increase. Conclusions: Conclusion: The existing drugs used to treat PD do not have the required efficacy to improve or slow down the process of neurons destruction. According to our data, Gst as a safe and effective herbal medicine was able to prevent cell death of dopaminergic neurons and reduce apoptosis. Ultimately, Gst could introduce as a useful and novel compound for improving the clinical outcome of PD.

P588 / #672


DEVELOPMENT OF EARLY STAGE PARKINSON’S DISEASE MOUSE MODEL USING LOW DOSE ROTENONE MICROEMULSION TARGETING OLFACTORY BULBS

Lecture Title:

M. Sharma, J. Kaur, S. Rakshe, A. Khairnar NIPER-A(National Institute of Pharmacology and Toxicology), Pharmacology And Toxicology, Gandhinagar, India

Aims: The major challenge in Parkinson’s disease (PD) is development of progressive mouse model to cater the need of development of disease modifying therapy. Aim of the present study was to develop the alpha-synuclein (aSyn) pathology in olfactory bulb (OB) locally using low dose rotenone microemulsion which may later progress to substantia nigra (SN). Methods: We administered rotenone mucoadhesive microemulsion (0.1mg/kg) in C57Bl/6 mice (3 months) intranasally for 9 weeks and kept the animals without rotenone administration for 16 weeks. Behavioral studies for olfactory dysfunction and motor impairment were performed every second week till 9th week and later performed only on 25th week. After behavior the animals were sacrificed and OB and striatum were isolated to perform western blot to detect TH, GFAP, aSyn and its oligomeric form. Results: We found significant olfactory dysfunction and motor impairment on 25th week after intranasal administration which was not visible till 9th week in rotenone group compare to vehicle treated animals. We found significant increase in aSyn oligomeric to monomeric level in OB and striatum in rotenone group compared to vehicle. We did not find alterations in TH level in both OB and Striatum suggesting the behavioral alterations might have occurred due to alpha-synuclein accumulation induced changes. We did not find alteration in GFAP level in both OB and striatum. Conclusions: Low dose intranasal rotenone administration is enough to induce behavioral impairment and alpha synuclein accumulation but it does not lead to development of dopaminergic neuronal loss in OB and striatum in C57Bl/6 mice.

P589 / #841


SYSTEMIC INFLAMMATION TRIGGERED BY LPS DOES NOT EXACERBATE THE TOXICITY OF ALPHA-SYNUCLEIN PREFORMED FIBRILS IN MICE

Lecture Title:

M. Soerensen1, K. Luk2, R.C.-C. Chang1,3 1The University of Hong Kong, Laboratory Of Neurodegenerative Diseases, Lks Faculty Of Medicine, Pok Fu Lam, Hong Kong, Hong Kong PRC, 2University of Pennsylvania Perelman School of Medicine, Center For The Neurodegenerative Disease Research, Philadelphia, United States of America, 3The University of Hong Kong, State Key Laboratory Of Brain And Cognitive Sciences, Pok Fu Lam, Hong Kong PRC

Aims: Parkinson’s disease is the second most common neurodegenerative disease worldwide, where many patients gradually develop Parkinson’s disease dementia. Preformed fibrils of alpha-synuclein (PFFs) can seed and transmit endogenous alpha-synuclein (aSyn) to phosphorylate and accumulate in LB-like structures, but severe effects can be observed long after injection. Our study aims to investigate whether systemic immune responses could accelerate the spread of aSyn pathology across the brain of C57BL/6 mice. Methods: PBS/PFFs (dosage: 3ug PFF) were stereotaxically injected into the medial forebrain bundle of wildtype mice, which later was challenged by one single intraperitoneal injection of PBS/LPS (dosage: 5 mg/kg). The behavior was examined 90 days after stereotaxical injection of PBS/PFF, and immunohistochemical techniques were used to detect aSyn pathology and other molecular effects. Results: aSyn PFFs per se showed a mild spread to various regions of the brain after injection into the medial forebrain bundle with a significant loss of tyrosine hydroxylase immunoreactive positive neurons in the substantia nigra ipsilateral to injection. However, no behavioral defects were observed. LPS failed to exacerbate the effect of aSyn PFFs when administered intraperitoneal. Conclusions: Our results agree with ours and other findings that aSyn PFFs injected into the brain mildly spreads across the brain, but with no apparent effect on behavior. However, non-sterilized systemic inflammation does not further speed up the spreading process.

P590 / #1832

Topic: Theme D: TDP43- and C9orf72-Related Diseases / D1. Disease Mechanisms, Pathophysiology

DIPEPTIDE PROTEIN REPEAT OLIGOMERS TOXICITY IN ALS AND FTD

Lecture Title:

N. Bhatt1, N. Puangmalai1, M. Montalbano1, S. Garcia1, M. Carretero-Murillo Pop2, U. Sengupta1, J. Rudra3, R. Kayed1 1University of Texas Medical Branch, Neurology, Galveston, United States of America, 2Baylor College of Medicine, School Of Medicine, Houston, United States of America, 3Washington University in St. Louis, Department Of Biomedical Engineering, St. Louis, United States of America

Aims: The hexanucleotide repeat expansion in C9orf72 is one of the most common causes of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The hexanucleotide expansion generates toxic dipeptide protien repeats including GP, GA, GR, PA, and PR. Given increasing evidence that soluble species, like oligomers, are the main cause of neurotoxicity in neurodegenerative conditions such as Alzheimer’s and Parkinson’s disease, we investigated the ability of DPRs to aggregate and form toxic oligomers as well as interact with amyloidogenic proteins like tau. Methods: Here, we have taken a reductionist approach by synthesizing short dipeptides repeats (GA, PR, and GR) of varying lengths and used biophysical as well as biochemical assays to characterize the aggregates in vitro and in cellular models. We also used immunobloting and immunostaining to detect DPR oligomers in C9orf72 associated ALS and FTD patients. Results: The results suggest the propensity for DPRs, specially GR and PR, to form oligomeric structures which can cause toxicity. We also tested the toxicity of the varying DPR aggregates alone and in the presence with tau protein in cellular models. Moreover, we investigated the presence of DPR oligomers in ALS and FTD human samples and their potential role in disease pathogenesis. Conclusions: Many studies have investigated the toxicity associated with DPRs however it is still unclear the role of DPRs oligomers in disease progression. Thus, the ability to detect and characterize oligomeric DPRs has great potential to further the understanding of these diseases and aid in the development of targeted therapeutics.

P591 / #1364


TDP43-HDAC6 INTERACTION AND ITS ROLE ON ALS PATHOGENESIS

Lecture Title:

M. Bordoni1, E. Scarian2,3, D. Sproviero2, V. Fantini3,4, O. Pansarasa2, C. Cereda2 1Università degli Studi di Milano, Dipartimento Di Scienze Farmacologiche E Biomolecolari (disfeb), Centro Di Eccellenza Sulle Malattie Neurodegenerative, Milan, Italy, 2IRCCS Mondino Foundation, Genomic And Post-genomic Unit, Pavia, Italy, 3University of Pavia, Department Of Brain And Behavioural Sciences, Pavia, Italy, 4Golgi-Cenci Foundation, Laboratory Of Neurobiology And Neurogenetic, Abbiategrasso, Italy

Aims: TDP43 was found to bind the mRNA of histone deacetylase 6 (HDAC6), inhibiting its RNA translation. Many studies suggested that HDAC6 participates in the regulation of the autophagic pathway. Thus, aim of this work is to evaluate the binding TDP43-HDAC6 in amyotrophic lateral sclerosis (ALS) and the effect of the inhibition of HDAC6 in cellular model. Methods: Firstly, we evaluated the binding of TDP43 to HDAC6 by RNA immunoprecipitation in peripheral blood mononuclear cells in sporadic ALS (sALS) patients and in healthy subjects. In a second time, we over-expressed TDP43 to investigate the effect on our cellular model, analysing the level of HDAC6 protein. Finally, we studied autophagy by both western blot and immunofluorescence after HDAC6 silencing. Results: We found that TDP43 binds HDAC6 mRNA in sALS patients, suggesting that the increase of TDP43 in cytoplasmic compartment leads to a decreased RNA translation of HDAC6 protein. Moreover, we found that an over-expression of TDP43 leads to the decrease of HDAC6 protein level. These data confirmed the regulating role of cytoplasmic TDP43 in HDAC6 translation. Finally, we found that a decreased level of HDAC6 leads to a stop of autophagy pathway, which is typical of ALS patients’ cells. Conclusions: Our work provides new insight in the pathogenesis of ALS, in particular by investigating a pivotal role of TDP43 in autophagy impairment.

P592 / #247


TRANSCRIPTOME CHARACTERIZATION OF THE FRONTAL CORTEX IN FRONTOTEMPORAL LOBAR DEGENERATION

Lecture Title:

O. Dols-Icardo, S. Sirisi-Dolcet, V. Montal, B. Sabariego, L. Cervera-Carles, L. Molina-Porcel, L. Muñoz, R. Blesa, J. Fortea, A. Lleó, J. Clarimon Memory Unit, Sant Pau Biomedical Research Institute, Hospital de la Santa Creu i Sant Pau, Neurology, Barcelona, Spain

Aims: Our aim was to identify alterations in the transcriptomic signature and cell subpopulations occurring in the frontal cortex of patients with frontotemporal lobar degeneration (FTLD). Methods: We performed total RNA and small RNA sequencing in the frontal cortex (Brodmann area 46) of patients neuropathologically diagnosed with FTLD-TDP (without mutations in genes causing FTLD (n = 10)), FTLD-TDP associated with the C9orf72 repeat expansion (n = 11), FTLD-tau (n = 13, six of them carrying the p.P301L mutation in the MAPT gene) and 7 controls without neuropathological alterations in the same brain region. Using bioinformatics tools, alterations in the expression of genes, isoforms and gene co-expression networks were evaluated. Furthermore, changes in cell proportions were identified through cell-type deconvolution using human brain derived single-cell RNA sequencing data as a reference. Results: Our results demonstrate significant changes in gene and isoform expression, which are markedly related to synaptic dysfunction in all FTLD groups. The gene co-expression modules identified supported these results. Furthermore we found a specific signature of transcriptome alterations in each FTLD group. Finally, changes in the proportion of specific microglial and astrocyte subpopulations were demonstrated. Conclusions: Our data reveal a significant downregulation of synaptic-related transcriptome changes in FTLD. In addition, a specific signature of transcriptome alterations characterizes each FTLD subgroup and might be used as surrogate biomarkers of the underlying FTLD neuropathological subtype during life or as novel therapeutic targets. Finally, we identified alterations in particular microglial and astrocyte subpopulations that were common in all FTLD subtypes.

P593 / #1526


HNRNP K MISLOCALISATION IN FRONTOTEMPORAL LOBAR DEGENERATION LEADS TO CRYPTIC EXON INCLUSION.

Lecture Title:

A. Gatt1, A. Bampton1, S. Cappelli2, J. Humphrey3,4,5,6, E. Buratti2, T. Lashley1, P. Fratta7 1Institute of Neurology, University College London, Queen Square Brain Bank, Department Of Neurodegenerative Disease, London, United Kingdom, 2International Centre for Genetic Engineering and Biotechnology (ICGEB), Molecular Pathology Group, Trieste, Italy, 3Friedman Brain Institute, Icahn School of Medicine at Mount Sinai, Nash Family Department Of Neuroscience, New York City, United States of America, 4Icahn Institute for Data Science and Genomic Technology, Icahn School of Medicine at Mount Sinai, Department Of Genetics And Genomic Sciences, New York City, United States of America, 5Icahn School of Medicine at Mount Sinai, Estelle And Daniel Maggin Department Of Neurology, New York City, United States of America, 6Icahn School of Medicine at Mount Sinai, Ronald M. Loeb Center For Alzheimer’s Disease, New York City, United States of America, 7Queen Square Institute of Neurology, University College London, Department Of Neuromuscular Diseases, London, United Kingdom

Aims: HnRNP K is a heterogeneous nuclear ribonucleoprotein (hnRNP) that shares structural and functional similarities with TDP-43. Like other hnRNPs, hnRNP K plays an important role in pre-mRNA splicing in the nucleus and can also shuttle to the cytoplasm. We find that hnRNP K is predominantly cytoplasmic in Frontotemporal Lobar Degeneration (FTLD). The loss of nuclear TDP-43 in disease states is directly linked to splicing misregulation and aberrant RNA processing, leading to cryptic exon inclusion in gene targets and the generation of truncated proteins. We aimed to investigate changes in RNA processing and presence of cryptic events upon hnRNP K mislocalisation in FTLD. Methods: Bioinformatic analysis of a neuronal cell line with hnRNP K knockdown identified a set of cryptic events. We have validated these events using PCR amplification and gel electrophoresis in both SH-SY5Y cells with hnRNP K knockdown and post-mortem human brain tissue. Results: The loss of nuclear hnRNP K results in the inclusion of cryptic exons in multiple gene targets in both SH-SHY5Y cells with hnRNP K knockdown and FTLD brain regions exhibiting hnRNP K nuclear loss. Conclusions: The mislocalisation of hnRNP K from the nucleus to the cytoplasm in FTLD is associated with the inclusion of cryptic exons in multiple gene targets. This suggests that similarly to TDP-43, hnRNP K plays a crucial role in RNA processing and splicing regulation which when impaired could lead to transcriptional dysregulation and disease pathogenesis.

P594 / #473

Topic: Theme D: TDP43- and C9orf72-Related Diseases / D2. Therapeutic Targets, Mechanisms for Treatment

SCO-SPONDIN DERIVED PEPTIDE, A PROMISING INNOVATIVE DRUG-CANDIDATE TO PROLONG SURVIVAL IN A MOUSE MODEL OF AMYOTROPHIC LATERAL SCLEROSIS

Lecture Title:

L. Beckett1, N. Delétage2, J. Korpikoski1, S. Lemarchant2, J. Le Douce2, Y. Godfrin2,3, M. Voutilainen1 1Institute of Biotechnology, Regenerative Neuroscience, Helsinki, Finland, 2Axoltis Pharma, Preclinical R&d, Lyon, France, 3Godfrin Life Sciences, -, Caluire-et-Cuire, France

Aims: NX peptide is an innovative peptide with pleiotropic functions derived from the subcommissural organ (SCO)-spondin, a large glycoprotein that strongly contributes to neuronal survival during development. The aim of this study was to evaluate the therapeutic effect of NX peptide in the SOD1G93A

mouse model of amyotrophic lateral sclerosis (ALS). Methods: Symptomatic ALS SOD1G93A mice were treated with either vehicle or different doses of NX peptide (2.5, 5 or 10 mg/kg) from 90-days old until disease end-stage. Clinical status and body weight were analyzed twice a week and motor behavioral tests were performed weekly (rotarod, grip strength) or every other week (multiple static rods). Results: Treatment with NX peptide at 10 mg/kg significantly extended the lifespan of SOD1G93A mice; the median survival of vehicle-treated SOD1G93A mice was of 144 ± 10 days (n=11) whereas treatment with NX peptide increased the lifespan of SOD1G93A mice to 154 ± 6 days (n=10). Beneficial effects were also observed in behavioral assays. Conclusions: NX represents a promising disease-modifying drug-candidate in the context of ALS. Indeed, it prolongs the survival of ALS mice and improves motor performance in the SOD1G93A mouse model. We will now further investigate the cellular and molecular mechanisms underlying the beneficial effects of NX peptide on ALS disease progression/survival. A phase 1 clinical trial in healthy volunteers started to evaluate the tolerability of NX.

P595 / #783

Topic: Theme D: TDP43- and C9orf72-Related Diseases / D4. Imaging, Biomarkers, Diagnostics

MATHEMATICAL MODELLING REVEALS THE CORRELATES OF COGNITIVE IMPAIRMENT ACROSS THE FTLD SPECTRUM

Lecture Title:

C. Cividini1, F. Agosta1, S. Basaia2, E.G. Spinelli1, V. Castelnovo1, E. Canu2, N. Riva3, G. Magnani4, F. Caso4, M. Filippi5 1IRCCS San Raffaele Scientific Institute and Vita-Salute San Raffaele University, Division Of Neuroscience, Milano, Italy, 2IRCCS San Raffaele Scientific Institute, Division Of Neuroscience, Milano, Italy, 3IRCCS San Raffaele Scientific Institute, Unit Of Neurorehabilitation, Milano, Italy, 4IRCCS San Raffaele Scientific Institute, Unit Of Neurology, Milano, Italy, 5IRCCS San Raffaele Scientific Institute; Vita-Salute San Raffaele University, Division Of Neuroscience; Unit Of Neurology, Milano, Italy

Aims: The aim was to apply mathematical modelling to unravel MRI connectomic signatures of cognitive/behavioural impairment in amyotrophic lateral sclerosis (ALS), lying on a continuum with the behavioral variant of frontotemporal dementia (bvFTD). Methods: Eighty-three ALS, 35 bvFTD and 61 controls underwent clinical/cognitive evaluations and MRI scan. ALS patients were classified in 54 ALS pure-motor (ALSpm), 21 ALS with cognitive/behavioural impairment (ALSci/bi) and 8 ALS with FTD (ALS-FTD). The structural value of the brain connections was considered and normalized relative to controls. Statistical distribution analysis was performed identifying structural patterns between bvFTD and ALSpm and to assess ALS-like or a bvFTD-like patterns in ALSci/bi and ALS-FTD. Results: Compared to ALSpm, bvFTD showed greater structural involvement of the connections within and between frontal, temporal and parietal lobes. Conversely, ALSpm patients showed greater involvement of the connections between sensorimotor and basal ganglia areas, compared with bvFTD. ALSci/bi and ALS-FTD showed an ALSpm-like pattern within sensorimotor and basal ganglia areas. Within the frontotemporoparietal networks ALSci/bi and ALS-FTD behaved differently: ALSci/bi showed a bvFTD-like pattern only within the parietotemporal connections, while ALS-FTD showed a bvFTD-like pattern also within frontal and between frontal and sensorimotor areas. Conclusions: A widespread structural damage characterized bvFTD, particularly in the connections within frontotemporal areas and between frontal and other lobes, while a more focal structural damage within sensorimotor-basal ganglia areas characterized ALSpm. ALSci/bi and ALS-FTD showed an ALS-like pattern with the exception of a greater damage within the frontotemporoparietal network, signature of the occurrence of cognitive impairment. Funding:The Italian Ministry of Health(GR-2011-02351217;GR-2013-02357415;RF-2011-02351193) and AriSLA(ConnectALS).

P596 / #902


BRAIN MRI SIGNATURES OF ATROPHY IN GENETIC FRONTOTEMPORAL LOBAR DEGENERATION

Lecture Title:

E.G. Spinelli1, A. Ghirelli1, N. Riva2, S. Basaia3, C. Cividini1, G. Magnani4, F. Caso4, P. Caroppo5, S. Prioni5, L. Tremolizzo6, I. Appollonio6, V. Silani7, P. Carrera8, M. Filippi1, F. Agosta1 1IRCCS San Raffaele Scientific Institute and Vita-Salute San Raffaele University, Division Of Neuroscience, Milano, Italy, 2IRCCS San Raffaele Scientific Institute, Unit Of Neurorehabilitation, Milano, Italy, 3IRCCS San Raffaele Scientific Institute, Division Of Neuroscience, Milano, Italy, 4IRCCS San Raffaele Scientific Institute, Unit Of Neurology, Milano, Italy, 5Fondazione IRCCS Istituto Neurologico Carlo Besta, Unit Of Neurology 5 - Neuropathology, Milan, Italy, 6"San Gerardo" Hospital and University of Milano-Bicocca, Unit Of Neurology, Monza, Italy, 7IRCCS Istituto Auxologico Italiano, Department Of Neurology And Laboratory Of Neuroscience, Milan, Italy, 8IRCCS San Raffaele Scientific Institute, Laboratory Of Clinical Molecular Biology, Milan, Italy

Aims: To assess cortical, subcortical and cerebellar grey matter (GM) atrophy using magnetic resonance imaging (MRI) in patients with disorders of the frontotemporal lobar degenerations (FTLD) spectrum with known genetic mutations. Methods: Sixty-six patients carrying FTLD-related mutations were enrolled, including 44 with pure motor neuron disease (MND) and 22 with frontotemporal dementia (FTD). Sixty-one patients with sporadic FTLD (sFTLD) matched for age, sex and disease severity with genetic FTLD (gFTLD) were also included, as well as 52 healthy controls (HC). A whole-brain voxel-based morphometry (VBM) analysis was performed. GM volumes of subcortical and cerebellar structures were also obtained. Results: Compared with HC, GM volume loss on VBM was greater and more diffuse in genetic FTD, followed by sporadic FTD and genetic MND cases, whereas sporadic MND (sMND) showed focal motor cortical atrophy. Patients carrying GRN and C9ORF72 mutations showed the most widespread cortical volume loss. Greater atrophy of the parietal cortices and thalami was found, globally, in gFTLD patients compared with sFTLD and, particularly, in C9-MND compared with sMND. When assessing deep GM structures, gFTLD patients showed significant volume loss compared with sFTLD in the caudate nuclei and thalami, in particular comparing C9-MND with sMND. At the cerebellar level, greater atrophy of the right lobule VIIb could discriminate gFTLD from sFTLD patients. Conclusions: Measures of deep GM and cerebellar involvement might be useful markers of gFTLD, particularly C9ORF72-related disorders, regardless of the clinical presentation within the FTLD spectrum. Supported by: Italian Ministry of Health (RF-2011-02351193; GR-2011-02351217) and European Research Council (StG-2016_714388_NeuroTRACK).

P597 / #688


DEVELOPMENT OF A REAGENT FOR ASSAYING TDP-43 UTILIZING IMMUNOMAGNETIC REDUCTION

Lecture Title:

C.S.Y. Yang1, H.-C. Liu2, M.-J. Chiu3, T.-F. Chen3, C.-H. Lin3, H.-H. Chen2 1MagQu Co., Ltd., President Office, New Taipei City, Taiwan, 2MagQu Co., Ltd., R&d Division, New Taipei City, Taiwan, 3National Taiwan University Hospital, Department Of Neurology, Taipei, Taiwan

Aims: TDP-43 is a promising biomarker for frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). However, studies on TDP-43 in human biofluid are rare. In this work, the authors utilized an ultrasensitive technology, called immunomagnetic reduction (IMR), to develop the reagent for assaying TDP-43. Methods: The preclinical performance characteristics of the TDP-43 reagent, such as the standard curve, detection limits, assay linearity, dilution recovery range, assay reproducibility, spike recovery, reagent stability, and interference tests, were explored according to the CLSI guidelines. Plasma samples from normal controls (NC, n = 27) and from patients with frontotemporal dementia (FTD, n = 9), Alzheimer’s disease (AD, n = 34) and Parkinson’s disease (PD, n = 10) were collected for TDP-43 assays using the IMR TDP-43 reagent. Results: The low-detection limit of assaying TDP-43 was 0.68 fg/ml, and the upper-detection limit was 100 pg/ml. There was no significant interference effect when assaying TDP-43 mixed with hemoglobin, bilirubin, intralipid, albumin, etc. The FTD patients had significantly higher levels of plasma TDP-43 (0.419±0.193 fg/ml) compared to the NC subjects (0.163±0.097 fg/ml), AD patients (0.165±0.082 fg/ml) and PD patients (0.069±0.068 fg/ml). Through analysis of the ROC curve, the cut-off value of plasma TDP-43 for discriminating FTD from the other patient groups was 0.237 fg/ml, which resulted a clinical sensitivity of 0.889 and a specificity of 0.831. Conclusions: These results demonstrate the feasibility of assaying plasma TDP-43 to specifically identify FTD.

P598 / #1276

Topic: Theme E: Vascular Diseases / E1. Disease Mechanisms, Pathophysiology

DIET-INDUCED HIGH CARDIOVASCULAR RISK IMPAIRS BRAIN HIPPOCAMPAL METABOLISM AND LONG-TERM SPATIAL MEMORY IN MIDDLE-AGED MALE RATS

Lecture Title:

I.N. Alves1,2,3, D. Mena1,2,3, D.F. Silva1,3, D.E. Vaz1,4, E. Candeias1, C.R. Oliveira1,5, P.F. Monteiro6,7, P.I. Moreira1,3,8, A.I. Duarte1,3,9 1CNC - Center for Neuroscience and Cell Biology, Rua Larga, Faculty of Medicine (Pólo 1, 1st Floor), University Of Coimbra, Coimbra, Portugal, 2University of Coimbra, N/a, Coimbra, Portugal, 3CiBB - Center for Innovative Biomedicine and Biotechnology, University Of Coimbra, Coimbra, Portugal, 4University of Coimbra, Department Of Life Sciences, Coimbra, Portugal, 5Faculty of Medicine, University of Coimbra, Laboratory Of Biochemistry, Coimbra, Portugal, 6Centro Hospitalar e Universitário de Coimbra (CHUC, E.P.E.), Clinical Cardiology Research Unit, Cardiology Department, Coimbra, Portugal, 7Faculty of Medicine, University of Coimbra, N/a, Coimbra, Portugal, 8Faculty of Medicine, University of Coimbra, Laboratory Of Physiology, Coimbra, Portugal, 9Institute for Interdisciplinary Research (IIIUC), University Of Coimbra, Coimbra, Portugal

Aims: High cardiovascular risk (HCR) comprises aging-/lifestyle-associated diseases (e.g. hypertension, dyslipidemia, diabetes) that raise the risk for cognitive impairment/Alzheimer’s disease (AD). e hypothesized that HCR modulates brain metabolism and cognitive performance at midlife, predisposing to AD. We aimed to evaluate the impact of HCR on hippocampal metabolism, cognition and AD-like pathology in middle-aged male rats fed on salted diet vs. the well-known spontaneously-hypertensive (SHR) rats. Methods: We analyzed peripheral features of HCR, spatial memory and hippocampal levels of cAMP, cholesterol and phospho-Tau protein. Results: Blood pressure, insulin levels and HOMA-IR were higher in salted diet-fed rats, while body weight, glycemia, HbA1C and cholesterol were similar to SHR rats. This suggests that salted diet induces HCR in middle-aged rats. This was accompanied by an impairment of their long-term, hippocampal-related spatial memory, lower hippocampal cAMP, cholesterol and phospho-Tau levels than in SHR rats. Conclusions: Diet-induced HCR impairs brain metabolism and long-term spatial memory in middle-aged rats. However, this issue deserved further clarification. Funding sources: European Regional Development Fund (ERDF), through the Centro 2020 Regional Operational Programme (Projects CENTRO-01-0145-FEDER-000012 (HealthyAging2020), POCI-010145-FEDER-016577); by COMPETE 2020 - Operational Programme for Competitiveness and Internationalization; by Portuguese funds via FCT – Fundação para a Ciência e a Tecnologia (Projects P2020-PTDC/NEU-NMC/0412/2014, PTDC/SAU-TOX/117481/2010, POCI-01-0145-FEDER-007440; UIDB/NEU/04539/2020); by Santander-Totta & Faculty of Medicine, University of Coimbra (PEPITA 2018); by European Social Fund (Post-Doctoral Researcher contract DL57/2016 #SFRH/BPD/84473/2012 to AI Duarte; the MSc Researcher Fellowships CENTRO-01-0145-FEDER-000012#1 and #2 (HealthyAging2020) to IN Alves and D Mena). *IN Alves and D Mena contributed equally to this study.

P599 / #230


BAVACHIN INDUCES APOPTOSIS THROUGH INVOLVEMENT OF ROS/MFN2/AKT PATHWAY AND NF-ΚB: IMPORTANCE OF HERBAL MEDICINE FOR THE TREATMENT OF CANCEROUS DISORDERS.

Lecture Title:

D. Patel Sam Higginbottom University of Agriculture, Technology and Sciences, Department Of Pharmaceutical Sciences, Payagraj, India

Aims: Psoralea corylifolia has been used for the treatment of various form of skin disorders. Bavachin is also called phytoestrogen found to be present the medicinal plants in the pure form and have been isolated and purified from Psoralea corylifolia. Methods: Here in the present investigation, numerous important scientific work have been collected from literature work and analyzed to know the anti-cancer potential of Bavachin. Detailed pharmacological activities of Bavachin have been analyzed through data analysis of various scientific works from literature work. Importance of various molecular pathways and the involvement of ROS/Mfn2/Akt pathway, NF-κB and STAT3 have been also investigated through data analysis of different scientific works of literature work to get the molecular mechanism. Results: Effect of bavachin on human hepatocellular carcinoma (HepG2) cells have been investigated in various scientific research work and found to inhibit cell proliferation and trigger endoplasmic reticulum (ER) stress due to inhibition of the phosphorylation of protein kinase B (Akt) and cell apoptosis by Mfn2-Akt pathway. In another scientific investigation, effect of bavachin in multiple myeloma (MM) cell lines have been investigated and found to decrease viability of MM cell lines after bavachin treatment. It inhibited the activation of nuclear factor kappa B (NF-κB) and signal transducer and activator of transcription 3 (STAT3). Further scientific research work analysis also revealed that bavachin induced apoptosis by the activation of caspase-3 and caspase-9, implicating the involvement of the mitochondrial pathway. Conclusions: Data analysis of literature revealed the biological importance of ROS/Mfn2/Akt pathway in the anticancer mechanism of bavachin.

P600 / #237


BIOLOGICAL IMPORTANCE OF HYPERIN AS VALUABLE DIETARY FLAVONOID FOR THE TREATMENT OF HUMAN DISORDERS: THERAPEUTIC ROLE OF DIFFERENT ENZYMES THROUGH SCIENTIFIC DATA ANALYSIS

Lecture Title:

D. Patel, K. Patel Sam Higginbottom University of Agriculture, Technology and Sciences, Department Of Pharmaceutical Sciences, Payagraj, India

Aims: Plant and their derived product including various forms of marketed Herbal products have been used in the various system of medicine for the treatment of human disorders. Plant have numerous phytochemical which represents broad range of pharmacological activities and promising biological potential in the modern system of medicine. Hyperin is a flavonoidal class plant chemical found to be present in the Hypericum perforatum and Drosera rotundifolia. Methods: Present study aim is to collect and analyzed all the available scientific information of hyperin for the treatment of various forms of human disorders. Present study includes detailed literature data analysis of hyperin for their pharmacological activities and molecular mechanistic study through different scientific research works. All the collected literature data’s were analyzed statistically to get significant results. Literature database analysis was performed in the present investigation to predict the molecular mechanism of hyperin. Interaction of hyperin with various enzymes was also investigated through literature database analysis in the present investigation in order to know their biological potential. Results: Analysis of the collected scientific research data’s of hyperin from various literature sources revealed the biological importance of hyperin in the medicine. Literature data analysis signified their free radical scavenging potential and antioxidant properties. Literature data analysis revealed the protective effects of hyperin against various form of cytotoxicity. Data analysis revealed the biological importance of TNFα, ALT and AST in the various form of human disorders. Conclusions: Present study signified the potential health benefits of hyperin for the treatment of human disorders and related complication.

P601 / #1512


DETERMINING MECHANISMS BY WHICH VARIATIONS IN THE MTHFR GENE CAUSE CEREBROVASCULAR DAMAGE

Lecture Title:

A. Reagan1, M. Sasner2, A. Haber3, G. Carter1,4, K. Christensen5, R. Rozen5, G. Howell1 1The Jackson Laboratory, Research, Bar Harbor, United States of America, 2The Jackson Laboratory, Alzheimer's Disease Center, Maine, United States of America, 3The Jackson Laboratory, Research, Farmington, United States of America, 4The Jackson Laboratory, Model-ad, Bar Harbor, United States of America, 5McGill University, Human Genetics, Montreal, Canada

Aims: Alzheimer’s disease (AD) often presents with a cerebrovascular component that contributes to cognitive decline. The C677T variant in methylenetetrahydrofolate reductase (MTHFRC677T) has been associated with AD and vascular dementia. MTHFR codes for an enzyme involved in folate and homocysteine metabolism, and the MTHFRC677T polymorphism is associated with vascular inflammation. Although MTHFR deficiency has been modeled using mice with a Mthfr null allele, the mechanism(s) by which the MTHFRC677T variant increases risk for vascular damage are not understood. Methods: To determine the precise contribution(s) of the MTHFRC677T variant to vascular dysfunction, we utilized CRISPR to create a novel knock in (KI) mouse carrying the MTHFRC677T allele on the C57BL/6J background. We assessed plasma metabolites in young and middle-aged mice carrying one or two copies of the risk allele and performed bulk RNA sequencing and methyl sequencing on brain tissue. Liver and brain enzyme function and cerebrovascular features were assessed in young cohorts. Results: MthfrC677T mice have reduced liver and brain enzymatic function and increased blood homocysteine levels, which corresponds to human data. Mice with two risk alleles show significantly reduced genome-wide methylation in the brain at both young and middle ages, as well as differential expression of more than 300 genes at a young age. Activity data from a larger study in progress suggests that aged female mice carrying the Mthfr risk variant run significantly more than their WT littermates (n=6). Conclusions: A novel mouse strain has been created to determine the precise mechanisms by which the MTHFRC677T polymorphism increases risk for cerebrovascular dysfunction.

P602 / #539

Topic: Theme E: Vascular Diseases / E2. Therapeutic Targets, Mechanisms for Treatment

IN VITRO EFFECTS ON CONTRACTILE ACTIVITY OF SMALL BRAIN VESSELS ISOLATED FROM RATS OF ALCESEFOLISIDE AND MAURITIANIN, ISOLATED FROM ASTRAGALUS MONSPESSULANUS SPP. MONSPESSULANUS

Lecture Title:

B. Angelov Medical University of Sofia - Pharmaceutical Faculty, Department Of Pharmacology, Pharmacotherapy And Toxicology, Sofia, Bulgaria

Aims: Alzheimer’s disease and Parkinson’s diseases are the two primary and most common neurodegenerative diseases in elderly individuals. Reduction in the regional cerebral blood flow (rCBF) and neuronal loss in the brain can be observed in patients with Alzheimer’s disease and Parkinson’s disease. The biggest factors that compromise the rCBF are atherosclerosis, diabetes, and obesity. Be altering the blood flow to the brain we may suggest that we can affect the pathophysiological mechanisms that cause these diseases. There is evidence that Flavonoids exhibit well-expressed endothelial-independent vasodilatation. The aim of our study is to investigate the effects of the Flavonoids - Mauritianine and Alcesefoliside isolated from the aerial part of Astragalus Monspessulanus spp. Monspessulanus on the contractility of a.basilaris isolated from rats. A. Basilaris and the two vertebral arteries supply blood to the circle of Willis which plays an important role in the cerebral blood circulation. Methods: The substances were isolated from the aerial part of Astragalus monspessulanus subsp. monspessulanus. For further reference: Krasteva I, Bratkov V, Bucar F, et al. Flavoalkaloids and flavonoids from Astragalus monspessulanus. 2015;78(11):2565-71. For our investigation of the contractility of the blood vessels, we used wired myography (model 410A, JP Trading, Denmark). Results: The examined substances lead to insignificant changes in the vascular tone which cannot be explained and do not correspond to the available data on the effect of total flavonoids on the cardiovascular system. Conclusions: The examined substances lead to insignificant changes in the vascular tone.

P603 / #1277


EFFECT OF PERIPHERAL ADMINISTRATION OF EVOLOCUMAB ON CEREBRAL CORTEX OF MIDDLE-AGED RATS AT HIGH CARDIOVASCULAR RISK

Lecture Title:

D.F. Silva1,2, D. Mena1,2,3, I.N. Alves1,2,3, D.E. Vaz1,4, P.F. Monteiro5,6, P.I. Moreira1,2,7, A.I. Duarte1,2,8 1CNC - Center for Neuroscience and Cell Biology, Rua Larga, Faculty of Medicine (Pólo 1, 1st Floor), University Of Coimbra, Coimbra, Portugal, 2CiBB - Center for Innovative Biomedicine and Biotechnology, University Of Coimbra, Coimbra, Portugal, 3University of Coimbra, N/a, Coimbra, Portugal, 4University of Coimbra, Department Of Life Sciences, Coimbra, Portugal, 5Centro Hospitalar e Universitário de Coimbra (CHUC, E.P.E.), Clinical Cardiology Research Unit, Cardiology Department, Coimbra, Portugal, 6Faculty of Medicine, University of Coimbra, N/a, Coimbra, Portugal, 7Faculty of Medicine, University of Coimbra, Laboratory Of Physiology, Coimbra, Portugal, 8Institute for Interdisciplinary Research (IIIUC), University Of Coimbra, Coimbra, Portugal

Aims: Diets with high fat and salt content are increasingly associated with an exacerbation of cardiovascular risk, cardiovascular disease, brain/cognitive dysfunction and, ultimately, Alzheimer’s disease (AD). Thus, besides its efficient use in the treatment of dyslipidemia, the lipid-lowering PCSK9 inhibitor Evolocumab may be also effective against high cardiovascular risk (HCR)-related brain/cognitive changes. We hypothesized that Evolocumab protects the brain of middle-aged rats at HCR. We analyzed the effects of peripherally-administered Evolocumab on markers for HCR, AD and neurodegeneration/death in diet-induced HCR rats. Methods: We used blood and brain lysates, together with colorimetry/fluorimetry approaches to assess the above-mentioned parameters. Results: High-fat salted diet induced hypercholesterolemia, hypertriglyceridemia, hyperinsulinemia and diabetes in middle-aged rats, suggesting that they were at HCR. The Evolocumab-mediated reduction their blood cholesterol was accompanied by a slight reduction in brain phospho-Tau protein levels, and in the activities of caspases-1 and -12 in HCR rats. Conclusions: The peripheral administration of Evolocumab may constitute a neuroprotective strategy against HCR-induced brain/cognitive dysfunction and AD. However, this issue needs further clarification. European Regional Development Fund (ERDF), through the Centro 2020 Regional Operational Programme (Projects CENTRO-01-0145-FEDER-000012 (HealthyAging2020), POCI-010145-FEDER-016577); by COMPETE 2020 - Operational Programme for Competitiveness and Internationalization; by Portuguese funds via FCT – Fundação para a Ciência e a Tecnologia (Projects P2020-PTDC/NEU-NMC/0412/2014, PTDC/SAU-TOX/117481/2010, POCI-01-0145-FEDER-007440; UIDB/NEU/04539/2020); by Santander-Totta & Faculty of Medicine, University of Coimbra (PEPITA 2018); by European Social Fund (Post-Doctoral Researcher contract DL57/2016 #SFRH/BPD/84473/2012 to AI Duarte; the MSc Researcher Fellowships CENTRO-01-0145-FEDER-000012#1 and #2 (HealthyAging2020) to IN Alves and D Mena). DF Silva, D Mena, IN Alves contributed equally.

P604 / #973


THE EPIGENETIC MODULATOR APABETALONE DOWNREGULATES BRAIN ENDOTHELIAL ACTIVATION AND MONOCYTE ADHESION

Lecture Title:

S. Wasiak1, L. Fu1, E. Daze1, D. Gilham1, B. Rakai1, S. Stotz1, L. Tsujikawa1, C. Sarsons1, D. Studer2, K. Rinker2, R. Jahagirdar1, N. Wong1, M. Sweeney3, J. Johansson3, E. Kulikowski1 1Resverlogix Corp, R&d, Calgary, Canada, 2University of Calgary, Cellular And Molecular Bioengineering Research Lab, Calgary, Canada, 3Resverlogix Corp, Clinical Development, San Francisco, United States of America

Aims: Increased immune activity at the blood brain barrier (BBB) can exacerbate neuroinflammation. Bromodomain and extraterminal (BET) proteins are histone acetylation readers that activate cytokine-dependent transcription in monocytes and endothelial cells. Targeting BETs with epigenetic therapies may reduce immune-brain interactions during neuroinflammation. Methods: hCMEC/D3 human brain microvascular endothelial cells (HBMVECs), grown on plastic or suspended filters served as a BBB model. Monolayers treated with TNFα+IFNγ received apabetalone, BET degrading compound MZ-1, or solvent (DMSO) for 4-24h. Cytokines were measured in the apical and basolateral supernatants via multiplex immunoassays. Gene expression was measured by PCR. Surface adhesion proteins were assessed by FACS in primary HBMVECs and THP-1 monocytes. THP-1 adhesion to primary HBMVECs was measured in laminar flow conditions. Results: In cytokine-stimulated filter-grown hCMEC/D3 monolayers, apabetalone bilaterally reduced protein secretion of proinflammatory cytokines including MCP-3, fractalkine, GM-CSF, MCP-1, IL-6, IL-8, IP-10 and RANTES (-40% to -90%). Transcription of these genes was BET-dependent, as apabetalone or knockdown with MZ-1 reduced their expression. In cytokine-stimulated primary HBMVECs, apabetalone reduced surface abundance of VCAM-1 (-80%) and E-selectin (-50%), proteins involved in monocyte adhesion. Consequently, apabetalone countered THP-1 adhesion to cytokine-activated HBMVECs in laminar flow assays. Apabetalone-treated THP-1 had lower surface abundance of chemokine receptors CCR1, CCR2 and CX3CR1 (-60 to -80%), which is expected to lower interactions between monocytes and endothelial cells. Conclusions: Apabetalone decreases immune activation of the neuroendothelium, potentially reducing neuroinflammation. This may explain beneficial effects of apabetalone on cognition recently demonstrated in diabetic coronary artery disease patients (phase 3 clinical trial BETonMACE).

P605 / #308


BIOLOGICAL IMPORTANCE OF EUPAFOLIN AGAINST INFLAMMATORY DISORDERS AND OTHER COMPLICATIONS: THERAPEUTIC BENEFIT AND MEDICINAL IMPORTANCE

Lecture Title:

D. Patel, K. Patel Sam Higginbottom University of Agriculture, Technology and Sciences, Department Of Pharmaceutical Sciences, Payagraj, India

Aims: Plants have been used by the human being for the treatment of various health complications including various types of neurological disorders. Plant belongs to the different family are well known for their rich source of pure plant chemical called phytoconstituents. Eupafolin is a 6- methoxy-5,7,3′,4′-tetrahydroxyflavone class phytochemical found to be present in the Phyla nodiflora and Gaillardia aristata. Methods: Eupafolin have been well known for their antioxidant and anti-inflammatory activity in the medicine. Here in the present investigation numerous scientific literature databases have been searched to collect needed information of eupafolin for the treatment of human disorders. Biological importances of eupafolin in the medicine for their therapeutic potential have been investigated through literature data analysis of various scientific research works. Detailed pharmacological potential of eupafolin have been investigated through literature data analysis. Biological importance of various biomolecules such as IL-1α, IL-1β and TNF in the human disorders has been investigated through literature data analysis. Results: Literature data analysis revealed the biological importance and therapeutic benefit of eupafolin in the treatment of inflammatory disorders through scientific data analysis of current research works. Further biological importance of eupafolin in the medicine is mainly due to their inhibitory potential on NO and down-regulation of various types of inflammatory mediators including cytokines. Literature data analysis of current research work revealed the biological importance of IL-1α, IL-1β and TNF in the medicine. Conclusions: Literature data analysis revealed the therapeutic benefit and biological importance of eupafolin for the treatment of human disorders.

P606 / #1385


PLASMA BRAIN-DERIVED NEUROTROPHIC FACTOR IN ON-PUMP CORONARY SURGERY PATIENTS WITH AND WITHOUT A SHORT COURSE OF PHYSICAL PREHABILITATION

Lecture Title:

O. Trubnikova1, E. Moskin1, I. Tarasova1, Y. Argunova2, O. Barbarash3 1Research Institute of Complex Issues of Cardiovascular Diseases, Laboratory Of Neurovascular Pathology, Kemerovo, Russian Federation, 2Research Institute of Complex Issues of Cardiovascular Diseases, Laboratory Rehabilitation, Kemerovo, Russian Federation, 3Research Institute of Complex Issues of Cardiovascular Diseases, Institute Directorate, Kemerovo, Russian Federation

Aims: To evaluate the effect of physical preoperative training for the plasma level of brain-derived neurotrophic factor (BDNF) in patients undergoing elected on-pump coronary artery bypass grafting (CABG). Methods: The study included 95 male CABG-candidates, which were divided into 2 groups: with (n=45) and without training (n=50). The preoperative physical training consisted of course of intensive training on a treadmill during 5-7 days. The plasma BDNF level was assessed in the peripheral blood at baseline, immediately after the training course and at the 7th day after CABG using a single-phase ELISA (R&D Systems, Inc., USA). Statistical processing was carried out using the STATISTICA 8.0. Results: It was shown an increase of the plasma BDNF level in patients immediately after a course of aerobic training as compared to the baseline values (23.3 [15.9; 30.1] and 31.8 [29.7; 42.8] ng/ml, (p <0.0001)). The patients without a training course had no significant BDNF changes (25.7 [14.4; 30.0] and 26.1 [25.2; 32.9], p = 0.7)). The intergroup differences were significant at this time period (p = 0.006). At the 7th day after CABG, the higher BDNF level was also observed only in the patients with aerobic training as compared to the patients without training 26.8 [20.4; 30.2] and 25.6 [20.6; 26.9] ng/ml (p = 0.035). Conclusions: High plasma BDNF was observed in the patients with a short course of aerobic physical training in perioperative period. It was proposed that this protective mechanism is a consequence of physical preoperative training for development of cerebrovascular complications after on-pump CABG.

P607 / #375

Topic: Theme E: Vascular Diseases / E4. Imaging, Biomarkers, Diagnostics

PARKINSONISM AND OTHER MOVEMENT DISORDERS ASSOCIATED WITH INTRACRANIAL AND EXTRACRANIAL ARTERY DISEASES

Lecture Title:

S.J. Chung, N. Choi, K.W. Park, S. Jo, H.-S. Ryu Asan Medical Center, Neurology, Seoul, Korea, Republic of

Aims: Intracranial and extracranial artery diseases may cause movement disorders such as parkinsonism, chorea, dystonia, myoclonus, and tremor. We aimed to investigate the clinical characteristics and prognosis of parkinsonism and other movement disorders caused by intracranial and extracranial artery diseases. Methods: We conducted a retrospective, cross-sectional, and multicenter study. We analyzed clinical and imaging data of the patients with parkinsonism and other movement disorders caused by intracranial and extracranial artery diseases. All data were collected from 14 University hospitals in South Korea. Results: Total 71 patients were enrolled in this study. Our patients presented with chorea (n=46, 64.8%), dystonia (n=10, 14.1%), limb-shaking movements (n=8, 11.3%), myoclonus (n=7, 9.9%), tremor (n=4, 5.6%), and parkinsonism (n=1, 1.4%). Four patients (5.6%) had two or more abnormal movements such as chorea with myoclonus or chorea with dystonia. On neuroimaging, 30 patients had the focal stenosis or occlusion in the middle cerebral artery opposite to the abnormal movements, 9 patients had posterior cerebral artery disease, 13 patients had intracranial internal carotid artery (ICA) disease, 17 patients had extracranial ICA disease, and 17 patients had moyamoya disease. Twenty patients were improved spontaneously and 37 patients were improved with medical or surgical treatments. Fourteen patients showed that movement disorders (9 chorea, 4 dystonia, 1 tremor and 1 limb-shaking movement disorder) persisted after appropriate treatments. Conclusions: Hemichorea was the most frequent movement disorders caused by intracranial and extracranial artery diseases, which may be persistent after treatments. This study may provide useful information about parkinsonism and other movement disorders associated with vascular etiologies.

P608 / #283


BREVICAN AND NEUROCAN AS POTENTIAL CSF BIOMARKERS FOR DIFFERENTIATION BETWEEN VASCULAR DEMENTIA AND ALZHEIMER’S DISEASE

Lecture Title:

K. Minta1, G. Brinkmalm1,2, E. Portelius1,2, P. Johansson3,4, J. Svensson4,5, P. Kettunen1, A. Wallin1, H. Zetterberg1,2,6,7, K. Blennow1,2, U. Andreasson1,2 1Institute of Neuroscience and Physiology, the Sahlgrenska Academy at the University of Gothenburg, Department Of Psychiatry And Neurochemistry, Mölndal, Sweden, 2Sahlgrenska University Hospital, Clinical Neurochemistry Laboratory, Mölndal, Sweden, 3Lund University, Department Of Clinical Sciences, Helsingborg, Sweden, 4Institute of Medicine, the Sahlgrenska Academy at the University of Gothenburg, Department Of Internal Medicine And Clinical Nutrition, Gothenburg, Sweden, 5Skaraborg Central Hospital, Department Of Endocrinology, Skövde, Sweden, 6UCL Institute of Neurology, Department Of Neurodegenerative Disease, London, United Kingdom, 7UCL, Uk Dementia Research Institute, London, United Kingdom

Aims: Brevican and neurocan, CNS-specific extracellular matrix proteoglycans, are cleaved and degraded by various metalloproteinases. However, their fragmentation profile is largely undetermined in cerebrospinal fluid (CSF). The aim of the study was to quantify tryptic peptides derived from brevican and neurocan proteins in human CSF of patients with Alzheimer’s disease (AD), vascular dementia (VaD) and healthy controls. Methods: The first cohort included 75 participants (25 patients with AD, 7 with mild cognitive impairment (MCI) diagnosed with AD upon follow-up, 10 patients with VaD or MCI diagnosed with VaD upon follow-up, and 33 healthy controls and cognitively stable MCI patients). The second cohort included 31 participants (5 AD patients, 14 subcortical small vessel disease (SSVD) patients and 12 healthy controls). Twenty tryptic peptides derived from brevican (n=9) and neurocan (n=11) proteins were quantified using parallel reaction monitoring mass spectrometry. Results: In the first cohort, nearly all CSF concentrations of brevican and neurocan peptides were significantly decreased in VaD compared with AD (AUC=0.83-0.93, p≤0.05) as well as with controls (AUC=0.79-0.87, p≤0.05). In the second cohort, the CSF concentrations of two brevican peptides were significantly decreased in SSVD compared with AD (AUC=0.86-0.91, p≤0.05) and with controls (AUC=0.80-0.82, p≤0.05), although other brevican and neurocan peptides showed an apparent trend to be decreased in SSVD compared with AD group (AUC=0.64-80, p>0.05). No brevican or neurocan peptides differed between AD and controls. Conclusions: Brevican and neurocan peptides are potential differential diagnostic biomarkers for VaD, with ability to discriminate VaD from AD.

P609 / #1025


NEUROVASCULAR UNCOUPLING OF CEREBRAL BLOOD FLOW AND GLUCOSE METABOLISM IN APOE4, TREM2, AND APOE4.TREM2 MICE

Lecture Title:

P. Territo1, P. Lin1, S. Persohn1, A. Bedwell1, K. Eldridge1, R. Speedy1, K. Kotredes2, R. Pandey3, A. Oblak1, M. Sasner2, G. Carter4, B. Lamb1, G. Howell2 1Indiana University School of Medicine, Stark Neuroscience Research Institute, Indianapolis, United States of America, 2Jackson Laboratory, Model-ad, Bangor, United States of America, 3THE JACKSON LABAROTARY FOR GENOMIC MEDICINE, Carter Lab, FARMINGTON, United States of America, 4The Jackson Laboratory, Model-ad, Bar Harbor, United States of America

Aims: Objectives: Alzheimer’s disease (AD) is the most common cause of dementia in the United States, with approximately 95% of patients exhibiting sporadic Late-Onset AD (LOAD). Apolipoprotein E4 (APOE4), the strongest genetic risk factor of LOAD, increases AD risk by 3-12 fold depending on copy number, and recnet work on the triggering receptor expressed on myeloid cell 2 (TREM2) shows the R47H variant increases AD risk by 2-3 fold. Therefore, developing a noninvasive phenotype would advance our understanding. Methods: Using PET/MRI, we established the perfusion-metabolism profiles across 27 brain regions in both sexes by using 64Cu-PTSM and 18F-FDG in the APOE4 (APOEE4/E4), TREM2 (TREM2R47H), and double (APOEE4/E4.TREM2R47H) knockin (KI) mice, and compared these to blood chemistry and nanoString transcriptomic analysis. Results: Analysis comparing 12mo to 4mo time point revealed that male APOE4 mice and both sexes of TREM2 had hypoperfusion and metabolism, while female APOE4 mice showed an uncoupled hyper-perfusion and hypo-metabolism phenotype, and was correlated to human AD pathology. In double KI mice, perfusion and metabolism showed a mixed phenotype which was region dependent. Cross-sectional analysis of KI compared to C57BL/6J mice at 12mo showed reduced glucose metabolism. Male APOE4, TREM2, and double mice showed hypo-perfusion and metabolism, while female double mice showed metabolic uncoupling compared to C57BL/6J. RNAseq and immunohistology was peformed, and key genes involved in the regulation of cerebral perfusion, glucose transportation, and metabolism were altered. Conclusions: These data indicate perfusion-metabolism strategy may be able to identify AD-related patterns that are observed in the transcriptomic profiling.

P610 / #1253

Topic: Theme E: Vascular Diseases / E5. Genetics, Epidemiology

POST STROKE DEPRESSION: PREVALENCE AND ASSOCIATED FACTORS IN A TUNISIAN COHORT

Lecture Title:

E. Jarrar, M. Belhi, M. Ben Halima, A. Mili, N. Amara, R. Douma, S. Naija, A. Hassine, S. Ben Amor Sahloul Hospital, Neurology Departement, Sousse, Tunisia

Aims: To investigate the prevalence of depression and its associated factors among patients suffered from stroke, in a Tunisian hospital. Methods: This cross-sectional study was carried out among patients suffering from acute stroke in neurology department in Sahloul Hospital over a period of one year. Participants were administered questionnaires to profile their socio-demographic and clinical characteristics. Subsequently, they were assessed with the Montreal Cognitive Assessment (MoCA), Beck Depression Inventory (IDB) and NIHSS at one week and at 3 months. Results: One hundred forty two patients were included. The mean age was 65 years. The sex ratio was 1.68. PSD prevalence was 57% initially and 51.1% at 3 months. We found significant correlations between PSD at one week and HTA (p=0.003) and stroke severity assessed by NIHSS (p=0.012). However PSD at 3 months were significantly associated with sex (p=0.004), marital status (p=0.04), physical disability (p=0.001) and cognitive impairment (P=0.021). Conclusions: Findings in this study support the need to focus on moods disorders post stroke. Early rehabilitation and psychological interventions targeting people with not modifiable related factors are needed to improve well-being.

P611 / #820

Topic: Theme E: Vascular Diseases / E7. Animal Models

MUMEFURAL AMELIORATES COGNITIVE IMPAIRMENT IN CHRONIC CEREBRAL HYPOPERFUSION VIA REGULATING THE SEPTOHIPPOCAMPAL CHOLINERGIC SYSTEM AND NEUROINFLAMMATION

Lecture Title:

J. Bang, M.-S. Kim, W.K. Jeon Korea Institute of Oriental Medicine, Herbal Medicine Research Division, Daejeon, Korea, Republic of

Aims: Chronic cerebral hypoperfusion (CCH) causes cognitive impairment and neurogenic inflammation by reducing blood flow. We previously showed that Fructus mume (F. mume) improves cognitiveimpairmentandinhibitsneuroinflammationinaCCHratmodel. Oneofthecomponentsof F.mume, Mumefural (MF), is known to improve blood flow and inhibit platelet aggregation. Whether MF affects cerebral and cognitive function remains unclear. We investigated the effects of MF on cognitive impairment and neurological function-related protein expression in the rat CCH model, established by bilateral common carotid arterial occlusion (BCCAo). Methods: Three weeks after BCCAo, MF (20, 40, or 80 mg/kg) was orally administrated once a day for 42 days. Using Morris water maze assessment, MF treatment significantly improved cognitive impairment. Results: MF treatment also inhibited cholinergic system dysfunction, attenuated choline acetyltransferase-positive cholinergic neuron loss, andregulatedcholinergicsystem-relatedproteinexpressionsinthebasalforebrainandhippocampus. MF also inhibited myelin basic protein degradation and increased the hippocampal expression of synapticmarkersandcognition-relatedproteins. Moreover,MFreducedneuroinflammation,inhibited gliosis, and attenuated the activation of P2X7 receptor, TLR4/MyD88, NLRP3, and NF-κB. Conclusions: This study indicatesthatMFamelioratescognitiveimpairmentinBCCAoratsbyenhancingneurologicalfunction and inhibiting neuroinflammation.

P613 / #258

Topic: Theme F: Prion Diseases / F4. Imaging, Biomarkers, Diagnostics

UNDERSTANDING THE ROLE OF SALIVARY EXOSOMES IN NEURODEGENERATIVE DISORDERS

Lecture Title:

S. Rastogi1, K. Rani1, P. Vishwakarma1, F. Nikolajeff2, S. Kumar1 1ALL INDIA INSTITUTE OF MEDICAL SCIENCES, Department Of Biophysics, DELHI, India, 2LULEA UNIVERSITY OF TECHNOLOGY, Department Of Health Sciences, LULEA, Sweden

Aims: 1.Isolation of exosomes from saliva of AD, CI (cognitively impaired), PD patients and healthy controls (HC) as well morphological characterisation and quantification 2.To correlate the expression level of hallmark proteins of AD and PD with disease severity staging 3.Development of an early screening methodology to check progression of Alzheimer’s and Parkinson’s disease using NTA Methods: Exosomes were isolated from saliva samples by chemical precipitation method. The morphological characterisation by transmission electron microscopy followed by quantification through NTA. The exosomes were validated by automated western-blot analyser using antibodies against CD63 (exosomal surface marker), L1CAM (neuronal-origin exosome). The expression level of hallmark proteins of AD and PD was achieved after performing western-blotting against amyloid-beta (Aβ) oligomer, (Aβ) monomer, phospho-tau in case of AD and against alpha-synuclein in case of PD. Results: We observed a significant difference in salivary exosomes concentration in CI and AD patients (p=0.0023) as well in PD (p= < 0.0001) when compared to the healthy control (HC). We validated our findings with western-blotting to check the expression level of oligomer amyloid-beta, phosphorylated tau protein, and alpha-synuclein from salivary exosomes. The Aβ oligomer/fibril abundance (p=0.0291), p-tau (p=0.0325) and Aβ protein abundance (p=0.0198) was significantly high in AD and CI as well the α-syn abundance (p=<0.0004) was significantly high in PD patients in comparison to HC. Conclusions: The concentration of exosomes obtained in NTA correlates well with pre-determined disease severity staging and level of hallmark proteins in case of PD and AD. It has a potential to be developed as an early screening methodology.

P614 / #1213

Topic: Theme F: Prion Diseases / F6. Cell, Molecular and Systems Biology

PROTEIN MISFOLDING CYCLIC AMPLIFICATION OF SPORADIC CREUTZFELDT-JAKOB DISEASE PRIONS IS STRONGLY SEED AND SUBSTRATE DEPENDENT

Lecture Title:

M. Bélondrade1, S. Nicot1, C. Mayran1, L. Bruyere-Ostells1, M. Di Bari2, E. Levavasseur3, S. Lehmann4, S. Haïk3, R. Nonno2, D. Bougard1 1Pathogenesis and Control of Chronic Infections UMR1058, Etablissement Français Du Sang, Montpellier, France, 2Istituto Superiore di Sanita, Department Of Food Safety, Nutrition And Veterinary Public Health, Roma, Italy, 3Institut du cerveau, INSERM U1127, CNRS UMR7225, Sorbonne Universités, Paris, France, 4IRMB, INM, Université de Montpellier, INSERM, CHU de Montpellier, Laboratoire De Biochimie-protéomique Clinique, Montpellier, France

Aims: Unlike variant Creutzfeldt-Jakob disease prions, sporadic Creutzfeldt-Jakob disease prions have been shown to be difficult to amplify in vitro by protein misfolding cyclic amplification (PMCA). In this study, we aimed to examine the ability of PMCA to amplify sCJD prions using three different PrP substrates. Methods: Brain samples from patients with sCJD of each representative subtype (MM1/MV1, MM2, MV2, VV1 and VV2) were serially tenfold diluted from 10-4 to 10-9 (w/v) and submitted to 4 PMCA rounds using normal brain homogenates originating either from human PrP transgenic mice (TgMet and TgVal) or from bank voles. Protease resistant prion proteins (PrPres) were detected by western blot after proteinase K digestion. Results: Bank vole substrate allowed the propagation of all sCJD subtypes with preservation of the initial molecular PrPres type. In contrast, PMCA in human PrP substrates led to a PrPres molecular shift during heterologous (M/V129) PMCA reactions. Combining PMCA efficiency with PrPres WB profiles obtained in the different substrates confirmed the classification of 4 distinct major sCJD prion strains (M1, M2, V1, and V2). The level of sensitivity required to detect VV2 sCJD prions in cerebrospinal fluid was achieved. Conclusions: Our results based on seeding activity properties of sCJD prions demonstrate that the efficiency of PMCA replication depends on both the sCJD prion strain considered and the nature of the PMCA substrate. Our findings highlight the potential of PMCA to support sensitive sCJD prions amplification, which might be amenable to their detection in human CSF samples.

P615 / #1132

Topic: Theme G: Huntington's and Other Neurodegenerative Diseases / G1. Disease Mechanisms, Pathophysiology

'MITOCHONDRIAL DYSFUNCTION': A PRIMARY EVENT IN GLUTAMATE NEUROTOXICITY.

Lecture Title:

B. Adebisi Osun State University Nigeria, Anatomy, Osogbo, Nigeria

Aims: The review aimed at establishing excitotoxic neuronal death, associated with neurodegenerative diseases and hypoxic insults, results from excessive exposure of excitatory neurotransmitters, like Glutamate. Hypothesis have also been tested that mitochondria are the primary targets in excitotoxicity. Methods: Researches of Alejandro and colleagues in 1996, were reviewed, where they activated the N-Methyl-D-Aspartate, NMDA, subtype of glutamate receptors for 20 minutes. The intracellular calcium ions were measured, alongside the mitochondria membrane potential, Delta Psi, using confocal imaging on cultural rats hippocampal neurons. Cyclosporin A was also used to block mitochondria permeability transition pore to see if there will be allowance of recovery of Delta Psi, in order to prevent neuronal cell death. Results: Massive influx of calcium ions, arising from overstimulation of NMDA subtypes of glutamate receptors, were observed. Sustained activation for 20 minutes elicited a reversible elevation of calcium ions. Longer activation for about 50 minutes initiated an irreversible influx of calcium ions. The susceptibility to NMDA-induced calcium ion overload is increased, when 20 minutes stimuli are applied to neurons pretreated with electron transport chain inhibitors, thereby implicating Mitochondria in calcium ions homeostasis, during excitotoxic challenges. Furthermore, Mitochondria Membrane potential, Delta Psi, exhibited prominent and persistent depolarization in response to NMDA, which closely parallels the incidence of neuronal death and that blockade of mitochondria permeability transition pore by cyclosporin allowed complete recovery of Delta Psi and prevented cell death. Conclusions: Understanding the underlying mechanisms of neuronal cell death brought by excitotoxicity could be instrumental in intercepting the cascade of events leading to Neurodegeneration.

P616 / #1314


UNRAVELING GLYCOLIC PATHWAYS ON AMYOTROPHIC LATERAL SCLEROSIS PATIENTS PRESENTING MUTANT SOD1 OR UNKNOWN MUTATIONS

Lecture Title:

F.G. Silva1, T. Cunha-Oliveira1, D. Mena1, V.A. Sardão1, A.I. Duarte1,2, P.J. Oliveira1 1CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Mitoxt - Mitochondrial Toxicology And Experimental Therapeutics, Cantanhede, Portugal, 2Institute for Interdisciplinary Research (IIIUC), University Of Coimbra, Coimbra, Portugal

Aims: Energy impairment and mitochondria dysfunction play a pivotal role in the neurodegeneration related with amyotrophic lateral sclerosis (ALS), especially in the mutant SOD1 (mutSOD1) subtype of patients. We hypothesized that different alterations occur in metabolic pathways of lymphoblasts from ALS patients with mutSOD1 and unknown (undSOD1) mutations. Hence, we aimed to evaluate different glycolytic alterations on patients with mutSOD1 and unknown mutations. Methods: Lymphoblasts from one ALS patient with mutSOD1 and one with undSOD1, as well as a sex-/age-matched control were obtained from Coriell. Oxygen consumption (OCR) and extracellular-acidification (ECAR) rates were measured by using the Seahorse XFe96 Extracellular Flux-Analyzer. Phenotypic metabolic microarrays were performed using the Biolog MitoPlate S-1 system. Hexokinase and lactate dehydrogenase activities, as well as lactate levels were measured by colorimetric assays. Statistical comparisons were performed by Kruskal-Wallis and Dunn's multiple comparisons test, with P-value<0.05 as significant. Results: undSOD1 lymphoblasts showed higher mitochondrial oxidation associated to glucose-6-PO4 as a substrate (p<0.05) and higher values of glycolytic ATP-production rate (p<0.05), compared to control, suggesting the activation of glycolysis. In concordance, the increase in hexokinase activity (p<0.01) and lactate levels (p<0.05) observed in undSOD1 lymphoblasts, compared to the control, reinforces this activation. Conversely, mutSOD1 lymphoblasts had lower mitochondrial ATP-production rate in comparison to undSOD1 lymphoblasts (p<0.05), suggesting an impairment of mitochondrial function. Conclusions: Conclusion: Glycolysis seems to be activated in lymphoblasts with undSOD1, while lymphoblasts with mutSOD1 present a distinct mitochondrial profile characterized by an impairment on mitochondrial function. Funding: POCI-01-0145-FEDER-029391, PTDC/MED-FAR/29391/2017, PTDC/BTM-SAL/29297/2017-POCI-01-0145-FEDER-029297, UIDB/04539/2020.

P617 / #1714


FAII SKIN MECHANORECEPTORS INDICATED AS A CANDIDATE TO EXPLAIN DEFICITS IN GRIP FORCE MODULATION IN PARKINSON DISEASE.

Lecture Title:

V. Gusbin1, B. Dachy2, I. Neu1, T. Pages1, J. Mcintyre3,4, A. Bengoetxea1 1Université Libre de Bruxelles, Faculté Des Sciences De La Motricité, Bruxelles, Belgium, 2CHU Brugmann, Neurology, Bruxelles, Belgium, 3TECNALIA, Basque Research And Technology Alliance (brta), Donostia-San Sebastián, Spain, 4Ikerbasque Research Foundation, ., Bilbao, Spain

Aims: Purpose: Basal ganglia are intrinsically involved in the planning and the modulation of the grip force. Grip force is tuned by hand and finger muscle long-latency reflexes (LLR). PD patients show increased grip force, and increased amplitude and latency for LLR (spindle and cutaneous origin), showing modifications at peripheral and central levels of precision grip neural circuits. In this study we try to identify which skin mechanoreceptor could be impaired. Methods: Method: Ten parkinsonian patients at stage between 1.5 and 3 on the Hoehn and Yahr scale, and between 10% and 35% on the motor examination section of the UPDRS (56 points reported as a percentage), participated in this study. We recorded muscular activity of the first dorsal interosseous of the hand during 3 grip tasks: 1) static, 2) weighted and 3) pulp anesthetized by cooling skin temperature below 25°C. Grip and load force were measured with two three-dimensional force sensors from Optoforce ©, electrical stimulations were sent by Digitimer © and EMG was recorded by Spike2©. Results: Compared to healthy age-matched subjects, electrical perception threshold was significantly increased as was the latency for LLR under cooling anesthesia. Concerning grip force, the grip force/load force ratio increased for a lighter weight (300gr) and with a anesthetized finger pulp in patients compared to healthy controls. Conclusions: Conclusion: Our results suggests that the well-known overshoot phenomenon in PD grip force could be the result of a deficiency of FAII skin mechanoreceptors.

P618 / #853


MICRORNA BIOGENESIS DEFICITS IN HUNTINGTON’S DISEASE

Lecture Title:

S. Petry1, R. Keraudren1, I. St-Amour1, S. Hebert1,2 1Centre de recherche du CHU de Quebec-UL, Neurosciences, Quebec, Canada, 2Laval University, Psychiatry And Neurosciences, Quebec, Canada

Aims: Early transcriptional deregulation is considered to be one of the key events involved in Huntington’s disease (HD) pathogenesis. Many studies have identified microRNA (miRNA) alterations in HD brain and peripheral system. Furthermore, miRNA-deficient mice develop HD-related symptoms. Interestingly, we and others have shown that huntingtin (HTT) could participate in modulating miRNA maturation, strengthening the link between defective miRNA biogenesis and neurodegenerative disorders. We now aim to better understand the role of miRNA biogenesis in HTT-mediated neurodegeneration. Methods: Using Western blot and qRT-PCR, we analyzed major components of the miRNA biogenesis pathway in a cohort of human HD brain (cortex and putamen, N=80). This was combined with the analysis of miRNA maturation of selected miRNA species (miR-10b, miR-132, miR-212, miR-127, miR-128, miR-196). We finally investigated the effects of specifically deleting the miR-132/212 cluster in adult HD mice. Results: We observed significant changes in miRNA biogenesis members and miRNA maturation in human HD brain, some of which occurred prior to overt neurodegeneration (e.g., miR-132). Similar biogenesis defects were observed in HD mouse and cell models. Deletion of the miR-132/212 cluster, which is strongly downregulated early in disease course, affected mouse behaviour and HD-related gene networks in mice. Conclusions: Our data are consistent with a predominant role for miRNA biogenesis alterations as underlying mechanism of miRNA alterations, transcriptional dysregulation, and clinical onset during HD progression. Further investigations are required to delineate the functional relationship between HTT and miRNA biogenesis; more so given the growing interest in gene silencing therapies requiring the endogenous miRNA/RNAi pathway.

P619 / #1036


SPONTANEOUS NEURODEGENERATION IN AGING C57BL/6 MICE AS MEASURED BY MORRIS-WATER-MAZE AND NOVEL OBJECT RECOGNITION

Lecture Title:

J.O. Hendrickx1, S. De Moudt1, E. Calus2, D. Van Dam2,3, G. De Meyer1 1Laboratory of Physiopharmacology, Faculty Of Pharmaceutical, Biomedical And Veterinary Sciences, University Of Antwerp, Wilrijk, Belgium, 2Laboratory of Neurochemistry and Behaviour, Institute Born-Bunge, Department Of Biomedical Sciences, University Of Antwerp, Wilrijk, Belgium, 3University of Groningen and University Medical Center Groningen, Department Of Neurology And Alzheimer Research Center, Groningen, Netherlands

Aims: Over past decades, the majority of neurodegenerative studies focused on β-amyloid diseases, tauopathies or α-synucleinopathies, while spontaneous neurodegenerative aging was often overlooked. In this study, we aimed for the behavioural phenotyping of aging C57BL/6 mice. Methods: C57BL/6 mice were studied at 2 (n=14), 4 (n=11), 6 (n=25), 9 (n=9) and 12 (n=21) months of age. Memory and spatial learning were assessed by a Morris Water Maze (MWM) and a Novel Object Recognition (NOR) test. Data are represented as mean ± SEM. Results: MWM measurements revealed signs of age-related memory loss in C57BL/6 animals from the age of 6 months onward (Table 1). The NOR assessment strengthened the latter finding by decreasing discrimination indexes (DI) and recognition indexes (RI) (Table 2). Table 1: Values of MWM parameters shown per age category over the total range of eight trials with *p<0.05**p<0.01,**p<0.001,****p<0.0001 compared to 2 months measurements. Data are presented as mean ± SEM.

2 months 4 months 6 months 9 months 12 months

Total path length (m) 95±8 112±8 127±5** 133±9* 129±8**

Escape latency (s) 478±35 582±48 755±39*** 773±57*** 831±42****

Swim speed (m/s) 1.42±0.03 1.40±0.06 1.22±0.03** 1.18±0.05*** 1.08±0.02****

Table 2: Measurements of DI and RI per age category with *p<0.05,**p<0.01,****p<0.0001. Data are represented as mean ± SEM.

2 months 4 months 6 months 9 months 12 months

DI (A.U.) 0.3±0.1 0.2±0.1 0.1±0.1* -0.1±0.1** -0.2±0.1****

RI (%) 65±3 58±3 53±3* 43±3** 40±3****

Conclusions: Clear signs of spontaneous cognitive decline were observed in aging C57BL/6 mice.

P620 / #696


CHANGES IN COGNITIVE FUNCTION DURING AGING

Lecture Title:

A. Sidenkova Ural State Medical University, Psychiatry, Yekaterinburg, Russian Federation

Aims: Currently, the number of cases of pathological aging of the central nervous system, represented by a violation of cognitive functions, is increasing. 148 people involved. Their age is 27 -74 years. We applied the screening neuropsychological method. . The dynamic heterogeneity of the cognitive profile during aging was revealed. The deterioration in the performance of the graphomotor test was the most age-specific. In older study participants, a decrease in the visual gnosis test correlated with a decrease in non-verbal intelligence. Methods: neuropsychological, statistical Results: A significant difference in the performance of the graphomotor test between the subjects of the age subgroup of 27-40 years and the subgroup of 41-50 years was statistically confirmed. The test was carried out using a marker that left a mark on the paper, the parameter of the total severity of regulatory errors introduced into the processing, in the form of the sum of penalty points normalized with respect to the number of series of patterns performed by each test subject. Conclusions: 1.Performance indicators of neuropsychological subtests of the right-handed group under study decrease unevenly as they age 2. In the older participants of the study, the evaluation of the strategy of the new figure correlated with a slight decrease in predominantly nonverbal and to a lesser extent with verbal Executive functions, with a slow pace of neurocognitive decisions of the highest order and a lower speed of information processing.

P621 / #344

Topic: Theme G: Huntington's and Other Neurodegenerative Diseases / G2. Therapeutic Targets, Mechanisms for Treatment

7,8-DIHYDROXYFLAVONE ATTENUATES COGNITIVE DEFICIT IN ICV-STZ RAT MODEL OF ALZHEIMER’S DISEASE REVERSING OXIDATIVE STRESS, MITOCHONDRIAL DYSFUNCTION, AND INSULIN RESISTANCE

Lecture Title:

A. Akhtar UIPS, Panjab University, Division Of Pharmacology, Chandigarh, India

Aims: Sporadic Alzheimer’s disease (SAD) is the most commonly prevalent dementia and progressive neurodegenerative disease. A polyphenolic flavonoid, 7,8-dihydroxyflavone (7,8-DHF) was used to reverse the cognitive deficit in a rat SAD model by reversing oxidative imbalance, mitochondrial enzyme dysfunction, and insulin resistance. Methods: For the SAD model, streptozotocin (STZ-3 mg/kg) was injected intracerebroventricularly (ICV) in male Wistar rats to induce cognitive dysfunction. Cognitive functions were evaluated by Morris water maze (MWM) and novel object recognition (NOR) tests, while locomotor activity was determined in actophtometer. 7, 8-DHF was given orally in doses of 5 mg/kg, 10 mg/kg, and 20 mg/kg, and reference standard drug rivastigmine in a dose of 2 mg/kg. Antioxidant enzymes, mitochondrial enzyme complexes were determined biochemically, insulin-degrading enzyme (IDE) and p-tau by ELISA, and histopathology by H&E staining. Results: 7,8-DHF attenuated cognitive deficit induced by ICV-STZ in MWM and NOR. Moreover, in the cortex and hippocampus regions of the brain, levels of reduced glutathione, catalase, superoxide dismutase, and mitochondrial complex enzymes and increased lipid peroxidation, protein carbonylation, and nitrite levels were subsequently reversed by 7,8-DHF and rivastigmine. IDE and p-tau protein were found to be altered. Histopathological examination revealed halted neurodegeneration. Conclusions: Conclusively, 7,8-DHF was found to be neuroprotective in the ICV-STZ rat model of SAD by ameliorating oxidative stress, mitochondrial dysfunction, and insulin resistance, thereby improving cognitive functions evident with the behavioral results. Hence, these results can further have clinical relevance in human AD.

P622 / #1215


MAINTENANCE OF NCX1 AND NCX2 ACTIVATION IN A MOUSE MODEL OF FAMILIAR ALS PREVENTS MISFOLDED SOD1 ACCUMULATION AND AMELIORATES MOTOR BEHAVIOR AND PROLONGS SURVIVAL RATE

Lecture Title:

S. Anzilotti, V. Valsecchi, G. Pignataro Federico II university of Naples, Neuroscience, Naples, Italy

Aims: Objectives: Imbalance in cellular ionic homeostasis is a hallmark of several neurodegenerative diseases including ALS. Sodium-calcium exchanger(NCX) is a membrane antiporter that, operating in a bidirectional way, couples the exchange of Ca2+ and Na+ ions in neurons and glial cells, thus controlling the intracellular homeostasis of these ions. Accordingly, the identification of new compounds capable of increasing NCX activity may be a suitable strategy to limit this devastating disorder. The aim of the present study is to evaluate the possible neuroprotective effects of NCX activator, Neurounina, on motor neurons degeneration in a mice model of familiar ALS. Methods: We performed real time PCR, western blotting, microfluorimetry and confocal microscopy experiments on spinal cord of SOD1 G93A and wild type mice Results: ALS mice showed a reduction in the expression and activity of NCX1 and NCX2 consistent with disease progression, therefore we aimed to investigate their role in ALS pathophysiology. Notably, we demonstrated that, in SOD1G93A mice, the prolonged NCX1 and NCX2 pharmacological activation:(1)prevented the reduction in NCX activity observed in spinal cord;(2)preserved motor neurons survival in G93A mice;(3)prevented the spinal cord accumulation of misfolded SOD1;(4)reduced astroglia and microglia activation in the spinal cord;(5)improved the lifespan and mitigated motor symptoms of ALS mice. Conclusions: The present study highlights the significant role of NCX in the pathogenesis of this neurodegenerative disorder and it might represent an important step in defining a new pharmacological approach for ALS

P623 / #886


DEVELOPMENT OF A THERAPY FOR ALZHEIMER’S DISEASE BASED ON COUNTERACTING THE PATHOLOGICAL EFFECTS OF APOE4

Lecture Title:

A. Bar-El1, H. Ben Moshe Ninio1, D.M. Michaelson2 1tel aviv university, Neurobiology, tel aviv, Israel, 2Tel Aviv Universtiy, Neurology, Tel Aviv, Israel

Aims: The overall objective of this study was to develop a low molecular weight and brain permeable druggable ABCA1 activator that can reverse the hypo-lipidation of apoE4 and the associated brain pathological effects. Methods: High-Throughput Screening (HTS) was first performed utilizing a cellular cholesterol efflux assay. Potential hits were then subjected to various other assays in order to select the most promising candidates. An orthogonal assay (i.e. digestion of senile plaques) was used in order to assess the extent to which the hits can reverse an efflux-independent apoE4 driven pathology, which was followed by cytotoxicity measurements. Target recognition experiments were then preformed utilizing microscale thermophoresis assay for assessing the extent to which the hits bind to ABCA1. Subsequent in vivo experiments were performed in which representative compounds were injected I.C.V. Results: Analysis of the orthogonal plaque digestion, direct interaction with ABCA1 utilizing target recognition experiments (MST) and the lack of toxicity led to determining a lead candidate. The lead candidate was injected into apoE4 TR mice and revealed reversal of the hypo-lipidation of apoE4 and a parallel increase in the level of this molecule, serves as a proof of concept to be beneficial in counteracting apoE4 pathological effects in-vivo. Conclusions: These findings provide a proof of principal that low molecular weight druggable ABCA1 agonists can reverse the hypo-lipidation and pathological effects of apoe4 and thus have important therapeutic potential.

P624 / #1274


DUAL ADMINISTRATION OF LIRAGLUTIDE AND GHRELIN IMPROVES BRAIN MITOCHONDRIAL ENERGY METABOLISM IN THE R6/2 MOUSE MODEL FOR HUNTINGTON´S DISEASE

Lecture Title:

D. Mena1,2,3, I.N. Alves1,2,3, M. Sjögren4, D.F. Silva1,3, C.R. Oliveira1,3,5, P.I. Moreira1,3,6, M. Björkqvist4, A.I. Duarte1,3,4,7 1CNC - Center for Neuroscience and Cell Biology, Rua Larga, Faculty of Medicine (Pólo 1, 1st Floor), University Of Coimbra, Coimbra, Portugal, 2University of Coimbra, N/a, Coimbra, Portugal, 3CiBB - Center for Innovative Biomedicine and Biotechnology, University Of Coimbra, Coimbra, Portugal, 4Brain Disease Biomarker Unit, Department of Experimental Medical Sciences, Wallenberg Neuroscience Center, Lund University, N/a, Lund, Sweden, 5Faculty of Medicine, University of Coimbra, Laboratory Of Biochemistry,, Coimbra, Portugal, 6Faculty of Medicine, University of Coimbra, Laboratory Of Physiology, Coimbra, Portugal, 7University of Coimbra, Institute For Interdisciplinary Research (iiiuc), Coimbra, Portugal

Aims: Huntington's disease (HD) is an incurable neurodegenerative disorder, characterized by motor/cognitive deficits, mitochondrial dysfunction, diabetes and cachexia. We hypothesized that co-administration of the anti-type 2 diabetes drug liraglutide with the orexigenic hormone ghrelin attenuates brain mitochondrial dysmetabolism in HD. We aimed to evaluate the effect of the peripheral treatment with liraglutide and ghrelin on brain mitochondrial metabolism in early symptomatic R6/2 HD mice. Methods: We evaluated the effect of a 2-week co-injection of liraglutide and ghrelin on brain cortical lactate levels, mitochondrial respiratory chain complexes’ activities (by colorimetry) and adenine nucleotide levels (by HPLC). Results: Liraglutide plus ghrelin stimulated brain mitochondrial respiratory chain complex IV, and reduced lactate and AMP levels in R6/2 mice brain, possibly improving their energy status. Conclusions: Liraglutide plus ghrelin ameliorate brain mitochondrial metabolism upon HD. However, this issue deserves further clarification. Funding sources: European Regional Development Fund (ERDF), through the Centro 2020 Regional Operational Programme (Projects CENTRO-01-0145-FEDER-000012 (HealthyAging2020), POCI-010145-FEDER-016577); by COMPETE 2020 - Operational Programme for Competitiveness and Internationalization; by a Seed Fund from European Huntington’s Disease Network (EHDN); by the Swedish Research Council; by Neuro Sweden; by Kocks Foundation; by Portuguese funds via FCT – Fundação para a Ciência e a Tecnologia (Projects P2020-PTDC/NEU-NMC/0412/2014, PTDC/SAU-TOX/117481/2010, POCI-01-0145-FEDER-007440 and UIDB/NEU/04539/2020); by Santander-Totta & Faculty of Medicine, University of Coimbra (PEPITA 2018); by European Social Fund (Post-Doctoral Researcher contract DL57/2016 #SFRH/BPD/84473/2012 to AI Duarte; MSc Researcher Fellowships CENTRO-01-0145-FEDER-000012#1 and #2 (HealthyAging2020) to D Mena and IN Alves).

P625 / #478


SCO-SPONDIN DERIVED PEPTIDE PROTECTS RAT AND HUMAN NEURONS AGAINST GLUTAMATE-INDUCED EXCITOTOXICITY

Lecture Title:

S. Lemarchant1, N. Delétage1, J. Le Douce1, Y. Godfrin1,2 1Axoltis Pharma, Preclinical R&d, Lyon, France, 2Godfrin Life Sciences, -, Caluire-et-Cuire, France

Aims: The subcommissural organ (SCO)-spondin is a brain-specific glycoprotein that strongly contributes to neuronal survival during development. The SCO becomes atrophic in adults, thereby losing protective functions of damaged nervous cells. Using rat and human neuronal cultures, we have evaluated the neuroprotective effect of NX peptide, derived from the SCO-spondin, against glutamate toxicity. Methods: Primary neurons were exposed to glutamate (glu) and treated with NX peptide (100, 250, 500 µg/mL). Neuronal survival and neurite network were assessed using immunohistochemistry or biochemistry. The mechanism of action was investigated by exposing neurons to inhibitors targeting receptors and intracellular mediators that trigger apoptosis, neuronal survival, or neurite growth. Results: NX promoted neuronal survival and prevented neurite network retraction on rat cortical and hippocampal neurons. NX triggered neuroprotection via β1-integrin and Notch receptors, activation of the PI3K/mTOR pathway and modulation of the apoptotic cascade. NX neuroprotective effect was confirmed in human cortical neurons: reduction of lactate dehydrogenase release (-52% in NX/glu-treated neurons compared to glu-treated neurons) and recovery of normal basal level of apoptotic cells (14, 28 and 13% for vehicle-, glu- and NX/glu-treated neurons, respectively). Conclusions: Collectively, these results show that NX peptide represents a promising translational drug-candidate to reduce glutamate-induced neuronal death. Proof of concepts in CNS animal models are under investigation to evaluate NX neuroprotective action. A phase 1 clinical trial in healthy volunteers is ongoing to evaluate the tolerability of NX peptide.

P626 / #730


CANNABINOIDS' NEUROPROTECTIVE EFFECT AS AN ALTERNATIVE TREATMENT FOR PARKINSON'S DISEASE: A SYSTEMATIC REVIEW

Lecture Title:

A.C. Ortiz1, A. Gavioli2, S.R. Ortiz2 1Faculdade de Medicina do ABC, Centro Universitário Saúde ABC, Medicine, Santo André, Brazil, 2São Judas Tadeu University, Postgraduate Program In Aging Sciences, São Paulo, Brazil

Aims: Parkinson's disease (PD) is a neurodegenerative disease caused by the deterioration of substantia nigra pars compacta’s dopaminergic pathways, resulting in a decrease in stimulation of the motor cortex and consequent symptoms linked to the reduction of automatic movements. It is known that several substances have protective effects against PD, among them cannabinoids (CB), obtained from Cannabis sativa, with emphasis on Cannabidiol (CBD) and Δ-9-tetrahydrocanbinol (Δ9-THC). The aim of this study was to assess previous data describing the protective role of cannabinoids in PD and their mechanisms of action, focused on improving symptoms and quality of life of individuals with the disease. Methods: Searches in PubMed and PMC were performed and articles published within the last 5 years that described protective effects of cannabinoids in individuals with PD, addressing their mechanism of action, were included. Results: Until now, 179 articles that establish a link between cannabinoids and PD were found. Their initial analysis revealed significant neuroprotective effect, but the mechanisms of action of CB in the central nervous system, which contribute to the regression of PD, still vary and there is no consensus on which theory is the most acceptable. Conclusions: The literature that reports the effects of PD cannabinoids is not conclusive, but studies show that there is a connection between the activation of cannabinoid pathways, Parkinson's disease and BDNF production that can be explored to obtain favorable clinical outcomes. Future studies are necessary to determine which of these substances’ mechanism of action actually guarantees their protective effect against neurodegeneration.

P627 / #884


COMPARISON OF TWO DIFFERENT IN VITRO MODELS OF NEUROINFLAMMATION: CYTOKINE RELEASE AND INFLAMMASOME ACTIVATION IN ORGANOTYPIC SLICES AND BV2 CELLS

Lecture Title:

I. Schilcher, T. Loeffler, S. Flunkert, B. Hutter-Paier QPS Austria GmbH, Neuropharmacology, Grambach, Austria

Aims: TREM2 variants as genetic risk factors for AD as well as current research on the inflammasome activation are giving rise for the importance of glial cells in AD. In the current study, two different LPS-stimulated in vitro systems of neuroinflammation were assessed in terms of cytokine release and expression of the inflammasome component NLRP3. The response to LPS treatment of organotypic brain slices, a mixed-model with intact cell-cell interactions, was compared to the response of the immortalized murine microglia cell line BV2. Methods: Organotypic hippocampal brain slices were prepared from early postnatal C57Bl6 mouse pups. To stimulate inflammation, slices were incubated with LPS at numerous concentrations and cytokine and sTREM2 release into the supernatant was measured by Mesoscale Discovery (MSD) immunosorbent assay. NLRP3 expression was examined with protein simple WES technology. Dexamethasone or MCC950 served as reference items to counteract the inflammatory response. A comparable treatment paradigm was used for microglial BV2 cells. Results: LPS stimulation differently changed cytokine and sTREM2 release into the supernatant as well as NLRP3 expression depending on concentration and duration within slices and cells. The reference items were able to downregulate the inflammatory response to various degrees. Conclusions: Disease-relevant inflammatory pathways are activated by LPS stimulation in both systems and this process can be reversed by reference compounds. While BV2 cells can serve as high-through-put screening tool, organotypic brain slices closely resemble the in vivo situation offering several advantages for early screenings, like maintenance of the interplay of different cell types in the postnatal brain.

P628 / #431


EFFECT OF SKIN ALLERGENS TARGETING NRF2 TRANSCRIPTION FACTOR ON ALZHEIMER’S DISEASE HALMARKS

Lecture Title:

A. Silva1, M. Pereira2, M. Carrascal3, G. Brites4, B. Neves5, P. Moreira1, R. Resende1, M. Silva4, A. Santos4, C. Pereira2, M. Cruz4 1University of Coimbra, Center For Neuroscience And Cell Biology And Institute For Biomedical Imaging And Life Sciences, Coimbra, Portugal, 2University of Coimbra, Medicine Faculty, Coimbra, Portugal, 3Tecnimede Group, Tecnimede Investigation Department, Sintra, Portugal, 4University of Coimbra, Pharmacy Faculty, Coimbra, Portugal, 5University of Aveiro, Department Of Medical Sciences And Institute For Biomedicine (ibimed), Aveiro, Portugal

Aims: Since 1) experimental evidences highlight nuclear factor (erythroid-derived 2)-like 2 (Nrf2) as a molecular target in Alzheimer’s disease (AD); 2) electrophilic cysteine-reactive skin allergens activate Nrf2; and 3) the skin allergen dimethyl fumarate (DMF) have demonstrated neuroprotective activity in in vivo models of neurodegenerative diseases, the main objective of this work was to evaluate the putative therapeutic role of skin allergens in AD. Methods: We studied the effect of the cysteine-reactive allergens 1,4-phenylenediamine (PPD) and methyl heptine carbonate (MHC) on 1) neuronal redox balance, mitochondrial polarization and calcium homeostasis using wild-type N2a neuronal cells (N2a-wt) and N2a cells overexpressing human wild-type amyloid precursor protein (APP) (N2a-APPwt) by RT-PCR, western blot (WB) and using fluorescent probes; and 2) on neuroinflammation, using microglia BV-2 cells exposed to Lipopolysaccharides (LPS), by measuring gene and protein levels of pro-inflammatory mediators by RT-PCR and WB, respectively. DMF was used as a positive control and phthalic anhydride (PA, mainly lysine-reactive), was used as a negative control. Results: DMF, PPD and MHC increased Hmox1 gene and HMOX1 protein levels in N2a-APPwt cells suggesting Nrf2-dependent antioxidant activity. MHC, but also PA, rescued N2a-APPwt mitochondrial membrane potential and calcium levels, in a Nrf2-independent pathway. All the chemicals showed anti-inflammatory activity by decreasing iNOS protein levels in microglia. Conclusions: This work highlights the potential neuroprotective and anti-inflammatory role of the selected skin allergens in in vitro models of AD, and supports further studies envisaging the validation of the results using in vivo AD models.

P629 / #1225


RTP801 MEDIATES SYNAPTIC PLASTICITY VIA EXTRACELLULAR VESICLES

Lecture Title:

J. Solana Balaguer1,2,3, G. Campoy Campos1,2,3, L. Pérez Sisqués1,2,3, N. Martín Flores1,2,3,4, A. Coll Manzano1,2,3,5, M. Masana Nadal1,2,3,5, J. Alberch Vie1,2,3,5, C. Malagelada Grau1,2,3 1University of Barcelona, Department Of Biomedicine, Barcelona, Spain, 2Institut de Neurociències, School Of Medicine And Health Sciences Of The University Of Barcelona, Barcelona, Spain, 3Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Department Of Biomedicine, Barcelona, Spain, 4University College London, Department Of Cell And Developmental Biology, London, United Kingdom, 5IDIBAPS-Instituto de Investigaciones Biomédicas August Pi i Sunyer, School Of Medicine And Health Sciences Of The University Of Barcelona, Barcelona, Spain

Aims: Neurodegenerative diseases such as Parkinson’s disease (PD), Alzheimer’s disease (AD) and Huntington’s disease (HD) induce an impairment in synaptic plasticity which eventually leads to cognitive symptoms in patients. Extracellular vesicles, which are involved in intercellular communication, have been also described to have an important role in synaptic plasticity, as they are carriers of bioactive miRNAs, proteins and lipids involved in synaptic processes which can influence firing rate in the recipient neurons. RTP801/REDD1 is a pro-apoptotic protein which levels are elevated in compromised neuronal populations from patients with neurodegenerative disorders. The aim of this study is to examine the potential role of neuronal EVs in synaptic plasticity and to investigate whether RTP801 is involved. Methods: Extracellular vesicles were isolated by ultracentrifugation from Sprague Dawley or WT/RTP801 KO C57BL6/129 cortical neurons at DIV14. Sister neuronal cultures were treated with EVs for 24h. Samples were subjected to immunohistochemistry, Western blotting and calcium imaging analysis. Results: We found that neuronal EVs per se enhance the consolidation of glutamatergic synapses in recipient neurons. Indeed, RTP801 seems to modulate this effect by affecting the production and the release of exosomes and their proteomic content. Conclusions: All these data propose neuronal EVs as potential neuroprotective agents and RTP801 as a therapeutic target to prevent synaptic plasticity impairment and neurodegeneration.

P631 / #379

Topic: Theme G: Huntington's and Other Neurodegenerative Diseases / G3. Drug Development, Clinical Trials

NEURO-PROTECTIVE ROLE OF PIPERAMIDE DERIVATIVE (D4) ON THE NEURODEGENERATION UPSTREAM PATHWAYS OF LATE-ONSET PROTEINOPATHIES

Lecture Title:

S. Shahbazi, T. Zakerali nencki institute of experimental biology, Neurobiology, Warsaw, Poland

Aims: - Design and development natural product based druggable molecule for neuroinflammatory leading neurodegeneration diseases - Evaluation of the bioactivity and pharmacological properties of synthesized druggable molecule D4 Methods: D4 ((2E,4E)-5-(benzo[d][1,3]dioxol-5-yl)-N-(4-(hydroxymethyl)phenyl)penta-2,4-dienamide)) was designed and its pharmacological properties were predicted. The bioactivity of piperine, D4 along with Aspirin was predicted using Glide 5.5 module at biologically relevant PH, with the Extra Precision (XP) mode targeting IKK-β, NF-kB (in 3 subunits, IKB, P50, and P65), COX-1, and COX-2 with PDB IDs, 4KIK, 1NFI (IKB, P50, and P65), 1DIY and 1CVU&3LN1 (COX-2). D4 was synthesized and its impact was tested on 2 types of inflammation-induced human brain cell-lines, microglia, and astrocyte using LPS 100ng/ml. cells were treated with 250 ng of D4 and 9500 ng/ml of Aspirin for 12 and 24h treatment according to the practical EC50 value of the known drug molecules obtained from FDA database. The efficacy of D4 was tested using various molecular biology techniques such as MTT assay, western blot, ELISA, qrtPCR, and Immunocytochemistry. Results: The result indicated that D4, with ideal pharmacoproperties for orally administration, significantly reduced the level of inflammatory cytokines in both proteome and genome level by inhibitory function on NF-kB translocation pathways in comparison with Aspirin as the well-known nonselective NSAID. D4 treatment increased the level of anti-inflammatory cytokines such as IL-10 and PPAR-α which help to alleviate inflammation and healing. Conclusions: D4 with its attractive pharmacological properties and amazing efficacy can be considered as a promising lead molecule for further clinical tests and pharmacological drug investigation.

P632 / #406

Topic: Theme G: Huntington's and Other Neurodegenerative Diseases / G4. Imaging, Biomarkers, Diagnostics

FUNCTIONAL CHANGES IN NONFLUENT VARIANT OF PRIMARY PROGRESSIVE APHASIA

Lecture Title:

D. Akhmadullina1, S. Fomenko1, Y. Shpilyukova1, D. Grishina2, E. Berdnikovich3, R. Konovalov4, E. Fedotova1 1Research Center for Neurology, Neurogenetics Department, Moscow, Russian Federation, 2I.M.Sechenov Fist Moscow State medical University, Neurology Department, Moscow, Russian Federation, 3Research Center for Neurology, Outpatient Neurology Department, Moscow, Russian Federation, 4Research Center for Neurology, Department Of Radiology, Moscow, Russian Federation

Aims: Non-fluent variant of primary progressive aphasia (nfvPPA) is a clinical syndrome of language decline characterized by agrammatism and/or speech apraxia and local atrophy involving mainly inferior frontal gyrus (IFG) and supplemental motor area (SMA). Functional changes may be more widespread than detectable structural alterations, and resting state functional magnetic resonance imaging (rs-fMRI) could provide an alternative approach to study this condition. Our aim was to explore patterns of functional changes in nfvPPA. Methods: We performed rs-fMRI in 9 patients with nfvPPA and 9 age-matched healthy controls (HC). Functional connectivity, fractional amplitude of low frequency fluctuations (fALLF) and local connectivity (LC) were analyzed. ROI-to-ROI connectivity (RRC) analysis was conducted in CONN toolbox. All statistical tests were computed considering age and gender as nuisance covariates. Results: Intrinsic connectivity, fALFF and LC were reduced in left IFG, posterior middle temporal gyrus, supramarginal gyrus and somatosensory regions in nfvPPA group compared to HC (p<0,05, FDR corrected). RRC analysis revealed not only expected reduced connectivity between IFG and posterior superior temporal gyrus but also among salience network regions which is more common in behavioral variant frontotemporal dementia. Moreover, according to RRC nfvPPA was characterized by a marked disrupted connectivity between left anterior insula and anterior cingulate cortex, SMA, putamen and caudate that are known to be involved in semantic analysis and production of speech. Conclusions: Obtained results show that nfvPPA is characterized by a widespread disruption of connectivity which affects not only IFG and SMA but also other regions that are responsible for speech and behavior.

P633 / #1502


CLINICAL CASE: FTIR ANALYSIS OF A PATIENT WITH ALZHEIMER'S DISEASE

Lecture Title:

A.G. Cárdenas-Pérez1, A.L. Vega Rodríguez2, J.S. Flores-Robles2, A.D. Bertadillo-Jilote2, D.G. García-Gutiérrez2, J. Campos-Guillén2, M.D.P. Figueroa-Corona3, M.A. Meraz-Ríos3, N.F. Santos-Sánchez4, B. Hernández-Carlos4, R. Salas-Coronado4, K.I. Lira-De León2 1Universidad Autónoma de Querétaro, Specialty In Clinical Biochemistry, Faculty Of Chemistry, QUERETARO, Mexico, 2Universidad Autónoma de Querétaro, Specialty In Clinical Biochemistry And Master In Diagnostic Clinical Chemistry, Faculty Of Chemistry, QUERETARO, Mexico, 3CINVESTAV-IPN, Molecular Biomedicine, Ciudad de Mexico, Mexico, 4Universidad Tecnológica de la Mixteca, Agroindustries Institute, Oaxaca, Mexico

Aims: The aim of this study was related the biochemical and anthropometric parameters and the history of a patient diagnosed with Alzheimer Disease (AD) with the results obtained by Fourier transform infrared spectroscopy (FTIR). Methods: Anthropometric measurements were taken from a patient previously diagnosed with AD, and a blood sample was obtained to collect serum and subsequently carry out biochemical measurements of cholesterol, triglycerides, cHDL, cLDL and glucose. To obtain spectra, the samples were pulverized and subjected to lyophilization. Subsequently, the samples are taken at random to obtain their spectrum in three replications. Results: An 87-year-old woman with AD had biochemical parameters of; cholesterol 172 mg/dL, triglycerides 119 mg/dL, cHDL 34.8 mg/dL, cLDL 113.4 mg/dL and glucose 84 mg/dL, anthropometric with abdominal circumference of 87 cm and blood pressure of 110/60. In the FTIR spectrum a separation of the data was observed in the regions 469 to 399 cm-1, 897 to 828 cm-1 and 1182 to 1113 cm-1. Conclusions: Most of the variations were found in the region between 900 and 400 cm-1. These vibrations are related to alterations at the level of sugars that make up nucleic acids in patients with AD, which can serve as a canvas to focus subsequent studies on in-depth analysis and validation of said region.

P634 / #981


A NEUROIMAGING STUDY OF THE EFFECTS OF EARLY VS LATE ANTI-INFLAMMATORY TREATMENT IN THE TGF344-AD ALZHEIMER’S DISEASE RAT MODEL

Lecture Title:

C. Fowler1, D. Madularu2, G. Devenyi3,4, J. Breitner5,6, J. Near3,6 1McGill University, Biological And Biomedical Engineering, Montreal, Canada, 2Northeastern University, Centre For Translational Neuroimaging, Boston, United States of America, 3Douglas Hospital Research Institute, Cerebral Imaging Centre, Verdun, Canada, 4McGill University, Psychiatry, Quebec, Canada, 5Douglas Hospital Research Institute, Perry Pavillion - Human Neuroscience Division, Verdun, Canada, 6McGill University, Psychiatry, Montreal, Canada

Aims: Alzheimer’s disease (AD) is a progressive neurodegenerative disorder with no effective treatments or known biomarkers for definitive diagnosis. Observable changes in brain pathology are now known to appear up to decades prior to clinical symptoms, substantiating the need for early detection and early intervention. To this end, Magnetic Resonance Spectroscopy offers a unique window into the brain; this non-invasive imaging technique permits the characterization of brain neurochemistry and is translatable from animal models to human AD subjects. The aim of this study was to characterize longitudinal changes in neurochemistry in the TgF344-AD rat model of AD, during normal disease progression and under treatment conditions, to determine the utility of MRS as an outcome measure for detecting disease modification. Methods: MRS spectra were acquired from the hippocampus at 4, 10, and 16-months of age in treated and untreated TgF344-AD rats, and untreated wildtypes (WT); n=~20/group for 80% power to detect a 3% effect size. Treatment consisted of either early (4-10 months) or late (10-16 months) treatment with Naproxen, a common non-steroidal anti-inflammatory drug. A linear mixed effects model was used to assess age by group interactions, while controlling for sex. Results: Early but not late chronic Naproxen treatment appears to mitigate disease-dependent changes in myo-inositol and taurine. Treatment effects were not seen for other disease-dependent changes, including decreased N-acetylaspartate and aspartate/glutamate, and increased total choline.

Conclusions: Preliminary results are consistent with the hypothesis that early intervention, rather than after the appearance of symptoms, is crucial for effective disease modification.

P635 / #1228


EVALUATION OF SALIVARY HEME OXYGENASE-1 AS A POTENTIAL BIOMARKER OF PARKINSON DISEASE AND OTHER NEURODEGENERATIVE CONDITIONS

Lecture Title:

J. Galindez1, L. Juwara2, M. Cressatti1, M. Gornitsky3, A. Velly3, H. Schipper1 1McGill University/Lady Davis Institute, Department Of Neurology And Neurosurgery, Montreal, Canada, 2McGill University/Lady Davis Institute, Department Of Quantitative Life Sciences, Montreal, Canada, 3Jewish General Hospital, Department Of Dentistry, Montreal, Canada

Aims: Parkinson’s disease (PD) is the second most common neurodegenerative disease that affects 2% of the population over 65 years of age. PD affects over 6 million people worldwide. No diagnostic biological tests are currently available and diagnosis can only be confirmed at autopsy. In an earlier study, protein levels of heme oxygenase-1 (HO-1), an inducible stress response protein important for heme catabolism and implicated in PD pathology, was reported to be elevated in PD subjects’ saliva. We aimed to evaluate salivary HO-1 protein levels of PD subjects and assess its potential as a biomarker of PD. Methods: Three hundred and seven subjects comprised of 75 PD patients, 162 non-neurological controls, 37 degenerative neurological controls, and 33 non-degenerative neurological controls were recruited. Levels of salivary HO-1 and total protein were measured using enzyme-linked immunosorbent assay and bicinchoninic acid assay, respectively. Analyses were adjusted by age, sex, total protein and relevant comorbidities. Results: We report significantly elevated salivary total protein and HO-1 concentration in PD subjects compared to non-neurological and non-degenerative neurological controls. Receiver operating characteristic curve analyses using HO-1 in combination with covariates showed an area under the curve (AUC) of 86% and 88%, respectively. Additionally, salivary HO-1 was significantly increased in patients with neurodegenerative conditions (n = 112) relative to those without (n = 195), exhibiting an AUC of 86.0%. Conclusions: Salivary HO-1 in combination with covariates may serve as a marker to distinguish patients with neurodegenerative conditions from the rest of the population.

P636 / #1252


NUTRIENTS IN THE PREVENTION OF ALZHEIMER'S DISEASE

Lecture Title:

E. Jarrar1, S. Naija1, M. Mhiri1, H. Hamdouni2, R. Douma1, N. Amara1, M. Ben Halima1, A. Hassine1, A. Omezzine2, S. Ben Amor1, A. Bouselema2 1Sahloul Hospital, Neurology Departement, Sousse, Tunisia, 2Sahloul Hospital, Biochemistry Department, sousse, Tunisia

Aims: To assess dietary patterns and vitamins levels in people with Alzheimer's disease (AD) as compared to healthy controls. Methods: This is a cross-sectional case-control study conducted in the Department of Neurology at the university hospital of Sahloul-Sousse-Tunisia. We included volunteered patients followed for AD diagnosed in accordance to the NINCDS-ADRDA criteria and 313 healthy controls. All subjects underwent structured clinical neurological evaluation, cognitive tests, biological assessments and brain imaging. Dietary patterns were collected with a food-frequency questionnaire. Results: One hundred thirty-seven patients and 313 healthy controls were included. The mean age at onset was 71 ± 9.7 years. The sex ratio of AD patients was 1.07 and 1.04 in healthy controls. The median MMSE was 14 ± 6. High blood pressure, diabetes and high cholesterol were more frequently found in AD patients with a statistically significant difference (p <0.001). Consumption of spices (OR=0.33), curcuma (OR=0.15), Dry fruits (OR=0.4), fig (OR=0.24), olive oil (OR=0.25), blue fish (OR=0.31), seafood (OR=0.25) and dark chocolate (OR=0.29) was lower in patients compared to controls with a statistically significant difference. Hypocalcemia, Vitamin B12 and Vitamin D Deficiencies were significantly more frequently associated with AD. Conclusions: Our study may provide a useful basis of existence of dietary specific patterns for AD in Tunisians. However, more research is needed before any of these factors can be considered a proven strategy to prevent Alzheimer's disease.

P637 / #666


PET CLASSIFICATION FOR MILD COGNITIVE IMPAIRMENT (MCI) AND ALZHEIMER’S DISEASE (AD) AGAINST HEALTHY ONE USING CNN FEATURE EXTRACTION

Lecture Title:

B. Khagi1, G. Kwon2 1Chosun university, Information And Communication Engineering, Seosuk-Dong, Dong-Gu, Korea, Republic of, 2Chosun university, Information And Communication Engineering, Gwangju, Korea, Republic of

Aims: To classify the PET images into three classes AD, MCI and HC (healthy controls), which eventually helps to identify early dementia condition. Methods: A 3D CNN based model was implemented to generate the features from each PET image, and then the features were extracted from the final fully connected layer of the CNN, which were directly fed into other classifier performing feature reduction technique like PCA. This will help to ease the classification and reduce the feature dimension, making it useful for final classification. Training was performed in around 1700 PETs and testing in 750 PETs.

Results: are tabulated below as the ground truth is shown along with classification accuracy. Conclusions: We successfully classified PET image with CNN+PCA +KNN, to get good results. The application of PCA helped to improve the classification accuracy.

P638 / #462


ALZHEIMER’S DISEASE DIAGNOSIS USING HIPPOCAMPAL AND AMYGDALA SUBFIELD OF STRUCTURAL MRI, RESTING STATE FMRI AND CSF

Lecture Title:

U. Khatri, G. Kwon Chosun university, Information And Communication Engineering, Gwangju, Korea, Republic of

Aims: Alzheimer’s disease (AD) is a convoluted neurodegenerative disease engaging variety of pathogenic factors, and disease etiology detection has been a major interest of researchers. Neuroimaging is a key and crucial means to analyze the disease. It is the prime current scientific research trend for combining other modality and neuroimaging data to explorer deep inside the potential insight of AD with the complementarities among multi-model biomarkers. Machine learning scheme possess huge potentiality and have attained some achievement in this research field. A few literatures have introduce some solution to the outcome of multi-modal data analysis, however the overall diagnostic framework for multi-modal fusion and analysis of fusion results has thus far been neglected. The full possibilities of multimodal biomarkers measures thus yet to be explorer optimally. Methods: We propose to combine a voxel-wise discriminative framework applied to multi-measure resting state fMRI, hippocampal sub-field, amygdala nuclei volume obtained from Structural MRI and CSF biomarkers using joint mutual information based features selection and MKL-SVM approach to enhanced diagnostic classification of AD. Results: For 37 clinically diagnosed AD patients, 94 MCI and 51 cognitively healthy controls. Obtained results validate that the fusion of distinct biomarkers perform well with Accuracies 93.19%, sensitivity 96.04% and specificity 91.15% for AD vs HC, Accuracies 87.70%, sensitivity 94.73% and specificity 85.33% for MCIs vs MCIc classifications respectively. Conclusions: The results indicated that the machine learning technique could effectively analyze multi-modal data fusion, giving new vision and perspectives for the analysis of Alzheimer’s disease.

P639 / #1513


CLINICAL CASE: PATIENT WITH ALZHEIMER'S DISEASE AND THEIR RISK FOR METABOLIC SYNDROME

Lecture Title:

A.L. Vega Rodríguez1, J.A. Romero-Muñoz2, A.G. Cárdenas-Pérez2, J.S. Flores-Robles1, A.D. Bertadillo-Jilote1, D.G. García-Gutiérrez1, M.D.P. Figueroa-Corona3, M.A. Meraz-Ríos3, N.F. Santos-Sánchez4, B. Hernández-Carlos4, R. Salas-Coronado4, K.I. Lira-De León1 1Universidad Autónoma de Querétaro, Specialty In Clinical Biochemistry And Master In Diagnostic Clinical Chemistry, Faculty Of Chemistry, QUERETARO, Mexico, 2Universidad Autónoma de Querétaro, Specialty In Clinical Biochemistry, Faculty Of Chemistry, QUERETARO, Mexico, 3CINVESTAV-IPN, Molecular Biomedicine, Ciudad de Mexico, Mexico, 4Universidad Tecnológica de la Mixteca, Agroindustries Institute, Oaxaca, Mexico

Aims: The aim of this study was analyzed the biochemical and anthropometric parameters for Metabolic Syndrome (MS) risk and the history of a patient diagnosed with Alzheimer Disease (AD). Methods: In a patient previously diagnosed with AD, the anthropometric measurements were taken and a blood sample was obtained to collect serum and subsequently carry out biochemical measurements of cholesterol, triglycerides, cHDL, cLDL and glucose. Using the criteria for diagnosis of MS according to the Harmonization of Metabolic Syndrome, the risk for MS was evaluated. Results: An 88-year-old woman with AD had biochemical parameters of cholesterol 217 mg/dL, triglycerides 154 mg/dL, cHDL 30 mg/dL, cLDL 156.2 mg/dL and glucose 72 mg/dL, anthropometric with abdominal circumference of 78 cm and blood pressure of 100/70. In the previous results, we can observe that the levels of triglycerides and cHDL were high and low respectively. Conclusions: Although MS was not observed in the results obtained, two parameters like triglycerides and cHDL were altered, and the main reason could be because the patient was controlled for diabetes, which resulted with normal valuesof the glucose parameter. The opportune diagnosis of these pathologies, allows diminishing possible complications, which allow offering a better quality of life to the patient, with an adequate and accessible treatment.

P640 / #1430


SELECTION THE TARGET FOR REHABILITATION INTERVENTION USING VIDEO ANALYSIS OF MOVEMENTS IN PARKINSON'S DISEASE.

Lecture Title:

E. Zubrickaya1, E.Y. Mozheiko2, S.B. Ismailova2, S. Prokopenko2 1Voino-Yasenetskiy Krasnoyarsk State Medical University of the Ministry of Health, Department Of Physical And Rehabilitation Medicine, Krasnoyarsk, Russian Federation, 2Professor V. F. Voino-Yasenetsky Krasnoyarsk State Medical University, Department Of Nervous Diseases With A Course Of Postgraduate Education, Krasnoyarsk, Russian Federation

Aims: Identification of the leading mechanism of gait disorders as a target of rehabilitation in a patient with PD using three-dimensional video analysis of movements. Methods: A study was carried out of a patient with a diagnosis of PD stage 3.5 according to Hoehn and Yahr, with postural and gait disorders using the method of three-dimensional video analysis of movements (VAM) on the Vicon Motion Capture Systems hardware and software complex. Results: Based on the examination, changes in the tempo-rhythm parameters of gait in a patient with PD in comparison with a healthy person were revealed: acceleration of the pace of walking with shortening the length of single and double strides, a decrease in the time of double support, acceleration of the moment of leg separation and a decrease in walking speed. Analysis of the angular characteristics of the step also revealed a decrease in the amplitude of flexion-extension in the hip, knee, and ankle joint with the greatest manifestation on the side with clinically more pronounced parkinsonism, insufficient knee flexion and hip extension, excessive dorsal flexion of the foot with insufficient plantar flexion. Conclusions: The identification of the individual characteristics of locomotion in a patient with PD makes it possible to form a plan of a targeted personalized recovery program. The method of three-dimensional VAM can be a diagnostic tool that is able to objectively assess the existing movement disorders. The method allows to achieve better results in the rehabilitation of movement disorders and improve the quality of life in patients with PD.

P641 / #1008

Topic: Theme G: Huntington's and Other Neurodegenerative Diseases / G5. Genetics, Epidemiology

GENOME-WIDE ASSOCIATION STUDY OF IMAGING IN ALZHEIMER'S DISEASE

Lecture Title:

J. Del-Aguila, F. Farias, E. Dhungel, C. Cruchaga Washington University in St. Louis, Department Of Psychiatry, St. Louis, United States of America

Aims: Deposition of amyloid plaques in the brain is one of the two main pathological hallmarks of Alzheimer’s disease (AD). Standardized Uptake Value (SUV) is a nuclear medicine term, used in positron emission tomography (PET) that selectively detects in vivo amyloid deposition in the brain for AD. In this project, a genome-wide association study (GWAS) in patients with AD was performed to investigate their genetic association with amyloid plaques. Methods: We collected in vivo amyloid deposition in the brains of 4467 subjects from three collaborative centers, the Alzheimer ’s disease Neuroimaging Initiative (ADNI), the Knight Alzheimer’s Disease Research Center (Knight-ADRC) and the Anti-Amyloid Treatment in Asymptomatic Alzheimer's Disease (A4). Then we performed a meta-analysis of genome-wide association studies to identify novel genetic loci for this endophenotype. Quality control analysis was performed following standard GWAS protocols Results: The APOE region showed the most significant association where several SNPs surpassed the genome-wide significant threshold, with APOE*4 being most significant (p = 5.82e-128; β = 0.18). Interestingly, after conditioning on APOE*4, two suggestive associated loci were found in chromosome 7 and chromosome 16. Conclusions: We have identified novel signals in non-APOE regions that affect amyloid deposition in the brain. Replication in another dataset is necessary in order to validate these findings.

P642 / #1013


A LONGITUDINAL GENOME-WIDE ASSOCIATION STUDY OF PROGRESSION IN ALZHEIMER'S DISEASE

Lecture Title:

J. Del-Aguila1, E. Dhungel2, F. Farias3, C. Cruchaga2 1Washington University in St. Louis, Department Of Psychiatry, St. Louis, United States of America, 2Washington University School of Medicine, Psychiatry, St Louis, United States of America, 3Washington University in St. Louis, Psychiatry, St. Louis, United States of America

Aims: The rate of disease progression is an important factor to consider regarding Alzheimer’s disease (AD). Even with the multiple cognitive assessment tools, it is contest to predict the disease stages due to the wide variability among patients in their rate of progression to AD. We performed a longitudinal genome-wide association study to investigate the presence of a genetic liability to faster progression in patients with AD Methods: We combined a sample of 3497 patients with AD from three different centers, the Alzheimer’s Disease Neuroimaging Initiative (ADNI), the Knight Alzheimer’s Disease Research Center (Knight-ADRC) and the National Alzheimer's Coordinating Center (NACC) that have been assessed at multiple timepoints (> 3) for change in CDR Sum of Boxes (CDR-SB) for this analysis. A linear mixed-model repeated measure framework was used to account for correlation between repeated measures in the same patient. Results: The sample was 49.3% male, with mean age at recruitment being 74.9 years. Mean CDR-SB baseline was 3.02 at first assessment and 11.54 at last assessment, and mean number of assessments was 3.16. After Bonferroni correction for multiple testing, we found seven loci to be significantly associated with progression. It is importance to notice that apolipoprotein E (ApoE) genotype was not associated with progression. Conclusions: After multiple testing correction, seven loci were significantly associated with progression in Alzheimer’s Disease. ApoE genotype was not found to affect the rate progression. Among these loci located in chromosome 1, 2, 6, 8, 10, 11 and 15, we think that most of them were spurious associations.

P643 / #1376


MITOCHONDRIAL DNA HAPLOGROUPS AND THREE INDEPENDENT POLYMORPHISMS HAVE NO ASSOCIATION WITH THE RISK OF PARKINSON’S DISEASE IN EAST INDIAN POPULATION.

Lecture Title:

T. Saha1, S. Roy1, R. Chakraborty1, A. Biswas2, K. Ray3, J. Ray2, M. Sengupta1 1University of calcutta, Genetics, KOLKATA, India, 2University of calcutta, Neuroscience, KOLKATA, India, 3ATGC diagnostics pvt ltd, Genetics, KOLKATA, India

Aims: Parkinson’s disease (PD) is a multifaceted illness affecting ~0.3% of the world population. The complex genetic bases of PD have not been fully elucidated. Several studies suggest mitochondrial DNA variants to be associated with PD. Here, we have explored the possibility of genetic association between mitochondrial haplogroups as well as 3 independent SNPs with PD in a representative East Indian cohort. Methods: The Asian mtDNA haplogroups: M, N, R, B, D, M7, and 3 other SNPs: 4336 T/C, 9055 G/A, 13708 G/A were genotyped in 100 sporadic PD patients and 100 matched controls via conventional PCR-RFLP-sequencing approach. Results: The distribution of mtDNA haplogroups as well as 3 single polymorphisms did not show any significant differences (p value > 0.05) between patients and controls. Conclusions: This is the first of its kind of study from Indian subcontinent that suggests no association of selected mitochondrial DNA variations with PD.

P644 / #693


ASSOCIATION BETWEEN GAIT SPEED AND COGNITIVE IMPAIRMENT AMONG COMMUNITY-DWELLING OLDER POPULATION: THE JAPAN PROSPECTIVE STUDIES COLLABORATION FOR AGING AND DEMENTIA (JPSC-AD)

Lecture Title:

K. Suzuki1, H. Niimura1, R. Shikimoto1, H. Kida1, Y. Miyasaka1, M. Shibata2, T. Honda3, T. Ohara4, J. Hata3, T. Ninomiya3, M. Mimura1 1Keio University School of Medicine, Department Of Neuropsychiatry, Tokyo, Japan, 2Kyushu University, Center For Cohort Studies, Graduate School Of Medical Sciences, Fukuoka, Japan, 3Kyushu University, Department Of Epidemiology And Public Health, Graduate School Of Medical Sciences, Fukuoka, Japan, 4Kyushu University, Department Of Neuropsychiatry, Graduate School Of Medical Science, Fukuoka, Japan

Aims: Previous studies have shown the slower gait speed is associated with cognitive impairment. However, few studies have shown reproducibility of this result using consistent methods. Thus external validity was limited. This study aimed to investigate the association between gait speed and cognitive impairment among community-dwelling elderly in Japan. Methods: A total of 9,002 community-dwelling participants aged 65 years or older, who have available the data of gait speed and cognitive function, were enrolled in selected eight regions of Japan. Participants were grouped into either normal cognitive function, cognitive impairment (mild cognitive impairment or dementia). Gait speed was divided into quintiles. As statistical analysis, we investigated the age-and sex-adjusted prevalence of cognitive impairment according to the gait speed levels. We also performed a logistic regression analysis to assess the association between gait speed and cognitive impairment after adjusting for confounders (e.g. age, sex, education, research region). Results: Among the study participants, 2,901 (32.2%, mean age 79.9 [standard deviation 7.8]) had cognitive impairment. Age- and sex-adjusted prevalence of cognitive impairment was 14.4%, 15.7%, 20.4%, 24.3%, and 31.2% in each quintile, from the fastest quintile. Lower gait speed levels were significantly associated with a higher likelihood of cognitive impairment after adjusting for confounders: the multivariable-adjusted odds ratios (95% confidence intervals) were, 1.39 (1.13-1.71) for quintile 3, 1.58 (1.29-1.94) for quintile 4, and 1.98 (1.60-2.44) for quintile 5 against quartile 1. Conclusions: The present findings suggest that subjects with slower gait speed are more likely to have cognitive impairment among community-dwelling Japanese elderly.

P645 / #1388


TONGUE BITING DYSTONIA AND FACIAL GRIMACING AS AN INITIAL MANIFESTATION OF CHOREA-ACANTHOCYTOSIS

Lecture Title:

W.T. Yoon Kangbuk Samsung Hospital, Sungkyunkwan Univerisity School of Medicine, Neurology, SEOUL, Korea, Republic of

Aims: Chorea-acanthocytosis is a rare hereditary disease caused by a mutation in a gene VPS13A that directs structural proteins in red blood cells. Chorea-acanthocytosis is a form of neuroacanthocytosis and is characterized clinically by a Huntington disease-like phenotype with progressive neurological symptoms including movement disorders, psychiatric manifestations and cognitive disturbance. It often shows itself as a mixed movement disorders, in which chorea, tics, dystonia and even parkinsonism may appear as s symptoms. Our aim is to report a case of unusual manifestation of chorea-acanthocytosis which showed frequent tongue biting dystonia and facial grimacing initially. Methods: Single case report. Results: A 43-year-old man presented to us with excessively frequent tongue biting and facial grimacing. He had 12-year history of diabetes and 5-year history of epilepsy on medication. In neurologic examination, he showed tongue biting dystonia and facial grimacing as well as chewing like oromandibular dyskinesia, mild dysarthria and bilateral cerebellar ataxia. His father had early-onset unspecified dementia. Brain magnetic resonance imaging (MRI) showed mild caudate and cerebellar atrophy. Dopamine transporter positron emission tomography-computed tomography and electroencephalogram showed no abnormality. Further genetic testing revealed a mutation of VPS13A gene confirming he had chorea-acanthocytosis. After he was prescribed aripiprazole and clonazepam, most movement symptoms improved. Conclusions: For now, we report for the first time a rare manifestation of chorea-acanthocytosis initially presenting tongue biting dystonia and facial grimacing. As like this case, if there are tongue biting dystonia and facial grimacing, it should be considered diagnosis and genetic test of chorea-acanthocytosis.

P646 / #860

Topic: Theme G: Huntington's and Other Neurodegenerative Diseases / G6. Cell, Molecular and Systems Biology

DIFFERENTIAL ALTERATIONS OF MOTOR BEHAVIOR AND ITS INTERNEURONAL ACTIVITY CORRELATES DURING DISEASE PROGRESSION IN A HUNTINGTON’S DISEASE MOUSE MODEL

Lecture Title:

S. Blumenstock1,2, K. Voelkl1,2, N. Del Grosso1, R. Klein2, I. Dudanova1 1Max Planck Institute of Neurobiology, Molecular Neurodegeneration Group, Martinsried, Germany, 2Max Planck Institute of Neurobiology, Department Of Molecules – Signaling – Development, Martinsried, Germany

Aims: Dysfunction of the cortico-striatal circuit is central in the development of Huntington’s disease (HD). Previous studies have shown an increase in cortical excitatory neuron activity, possibly related to a loss of inhibitory control through interneurons. However, the involvement of cortical microcircuits in the development of HD and their representation of disease-related behavior still remain open questions. Methods: We used chronic in vivo 2-photon calcium imaging of genetically targeted parvalbumin (PV), somatostatin (SST) and vasointestinal peptide (VIP) interneurons in the primary motor cortex of R6/2 HD mice and controls. Simultaneously, three-dimensional pose estimation of motor behavior was performed along presymptomatic and symptomatic phases of disease progression. Results: Unbiased machine learning analysis shows abnormalities in body posture, gait and forepaw movements in head-fixed R6/2 mice. Motor behavior is differentially encoded by interneuron subtypes in healthy mice, with PV and VIP activity positively and SST activity negatively correlating with locomotion, whereas grooming is strongly represented by SST activity. In R6/2 mice, interneuron population activity is differentially altered, with PV cells showing a decrease and SST cells an increase in activity. Moreover, neural representations of motor behavior are disturbed and can be observed before the onset of overt disease symptoms. Histologically, we find a partial loss of immunopositivity for SST and VIP, but not PV. Conclusions: Our study provides unprecedented detail on cell-type specific cortical function changes that occur during the progression of HD. Understanding how cortical microcircuit activity shapes motor behavior in health and disease is a crucial step towards developing effective and targeted therapy.

P647 / #1383


CHARACTERIZATION OF MENINGEAL FIBROBLASTS FROM HUMAN BRAIN DONORS

Lecture Title:

V. Fantini1,2, A. Davin1, C. Pandini3,4, T.E. Poloni5,6, V. Medici5, M. Bordoni7, O. Pansarasa4, A. Guaita1,6, C. Cereda4 1Golgi-Cenci Foundation, Laboratory Of Neurobiology And Neurogenetic, Abbiategrasso, Italy, 2University of Pavia, Department Of Brain And Behavioural Sciences, Pavia, Italy, 3University of Pavia, Department Of Biology And Biotechnology, Pavia, Italy, 4IRCCS Mondino Foundation, Genomic And Post-genomic Unit, Pavia, Italy, 5Golgi-Cenci Foundation, Department Of Neurology And Neuropathology, Abbiategrasso, Italy, 6ASP Golgi-Redaelli Geriatric Hospital, Department Of Rehabilitation, Abbiategrasso, Italy, 7Università degli Studi di Milano, Dipartimento Di Scienze Farmacologiche E Biomolecolari (disfeb), Centro Di Eccellenza Sulle Malattie Neurodegenerative, Milan, Italy

Aims: Meninges surround and protect the central nervous systems. In adulthood, meninges maintain the homeostasis of the CNS releasing trophic factors, and during brain injury, resident neural stem cells (NSCs) are differentiated to overcome the damage. These NSCs form a stem cell niche, and it has been observed the presence of Nestin+, which characterized neural stem cells, also in the adult brain without injury. The aim of this work was to study and characterize meningeal fibroblasts (MFs) and the self-renewal potential with the presence of NSCs, which partially overcome the neurodegeneration during aging. Methods: We selected 6 brain donors and we isolated MFs and skin fibroblasts (SFs), as control. We evaluated cell mobility, cell alignment, and growth rate. We measured different protein markers fibronectin, serpinh1, beta-III-tubulin, and nestin by immunofluorescence and differentially expressed genes through whole transcriptome analysis. Results: MFs and SFs show a clear difference in terms of cell mobility; MFs started to grow out from the tissue more quickly than SFs. SFs show a great alignment at 100% confluency. The expression of fibronectin, serpinh1 and beta-III-tubulin is the same in Ms and SFs, while the nestin is more evident in MFs. RNA-Seq analysis showed 1145 deregulated RNAs, with the involvement of extracellular matrix component and focal adhesion, confirming the differences observed at the morphological level. Conclusions: MFs and SFs show a different signature and the presence of nestin in Ms and the different deregulated signaling pathways, allow us to hypothesize that MFs could be used as a new source of neurons.

P648 / #877


NOVEL PROTEOSTASIS REPORTER MOUSE REVEALS DIFFERENT EFFECTS OF CYTOPLASMIC AND NUCLEAR AGGREGATES ON PROTEIN QUALITY CONTROL IN NEURONS

Lecture Title:

I. Dudanova1, S. Blumenstock1,2, E.K. Schulz-Trieglaff1,2, A.-L. Bolender1,2, K. Voelkl1,2, P. Lapios1,2, F.U. Hartl3, M. Hipp3,4, R. Klein2 1Max Planck Institute of Neurobiology, Molecular Neurodegeneration Group, Martinsried, Germany, 2Max Planck Institute of Neurobiology, Department Of Molecules – Signaling – Development, Martinsried, Germany, 3Max Planck Institute of Biochemistry, Department Of Cellular Biochemistry, Martinsried, Germany, 4University Medical Center Groningen, Department Of Biomedical Sciences Of Cells And Systems, Groningen, Netherlands

Aims: Cellular protein quality control machinery is important for preventing protein misfolding and aggregation, and decline in protein homeostasis (proteostasis) is believed to play a crucial role in age-related neurodegenerative disorders. However, how proteostasis capacity of neurons changes in different diseases is not yet sufficiently understood, and progress in this area has been hampered by the lack of tools to monitor proteostasis in mammalian models. The objective of our study was to develop and validate reporter mice for in vivo analysis of neuronal proteostasis. Methods: We generated transgenic mice expressing EGFP-fused firefly luciferase (Fluc). Fluc is a conformationally unstable protein that requires chaperones for proper folding and can therefore be used as a proteostasis sensor. Defects of proteostasis manifest by a decrease in luciferase activity and by appearance of intracellular Fluc-EGFP foci due to reduced solubility of the sensor. Results: Using Fluc-EGFP reporter line, we found a marked proteostasis impairment in rTg4510 taupathy mice, but not in R6/2 and HD94 Huntington’s disease mouse lines. Mechanistic investigations in primary neuronal cultures demonstrated that cytoplasmic, but not nuclear, aggregates cause defects of cellular proteostasis. Conclusions: The effects of protein aggregates on the protein quality control system are cellular compartment-specific and differ between neurodegenerative diseases. The new Fluc-EGFP reporter mice represent a useful tool for detailed studies of proteostasis in various neurodegeneration models.

P649 / #1761


SUBSTRATE-INDUCED CLUSTERING ACTIVATES TRIM-AWAY OF PROTEINS

Lecture Title:

A. Mukadam1, J. Zeng2, D. Clift2, L. James2, W. Mcewan1 1University of Cambridge, Uk Dementia Research Institute, Cambridge, United Kingdom, 2University of Cambridge, Mrc Laboratory Of Molecular Biology, Cambridge, United Kingdom

Aims: Trim-Away is a powerful new technology that acutely and rapidly degrades proteins using intracellular antibodies and the E3 RING ligase and cytosolic antibody receptor TRIM21. How TRIM21 is catalytically-activated upon substrate engagement during either its normal immune function or when re-purposed for targeted protein degradation is unknown. We aimed to determine the mechanism of TRIM21 activation. We further investigated whether TRIM21 and antibodies can be used for the selective degradation of host proteins implicated in neurodegeneration as a research tool and as a potential therapeutic strategy. Methods: We introduced antibodies and constructs expressing mRNA to cells through electroporation, permitting the selective degradation of endogenous proteins. Results: Using antibodies against an artificial substrate containing a repeated epitope sequence we show that it takes a threshold number of antibodies to be bound to the substrate in question to activate degradation via TRIM21. We demonstrate that clustering activates ubiquitin ligase activity on adjacent TRIM21 dimers. We exploit this mechanism to demonstrate that expanded poly-glutamine tracts can be specifically degraded at the protein level, while leaving wild-type length tracts unaffected. Conclusions: Our results demonstrate that, like other antibody receptors, TRIM21 is activated by ligation of polyvalent immune complexes. This mechanism provides a novel strategy for the selective depletion of cytoplasmic proteins that may have broad research and therapeutic implications.

P650 / #1402


SPATIALLY RESOLVED TRANSCRIPTOMICS IN THE APPSWE [TG2576] MOUSE MODEL OF ALZHEIMER’S DISEASE

Lecture Title:

C. Uytingco, J. Chew, A. Hartnett, N. Weisenfeld, S. Williams, S. Ziraldo, F. Meschi, K. Miller, Y. Yin 10x Genomics, Research And Development, Pleasanton, United States of America

Aims: Identifying individual cells and their genetic makeup using RNA-seq is critical for understanding their roles in how the central nervous system (CNS) functions in both healthy and diseased states. Here we demonstrate characterization of spatial transcriptomic patterns in the APPSWE [Tg2576] mouse model of familial Alzheimer’s Disease using the Visium Spatial Gene Expression Solution. Methods: Serial sections of fresh-frozen tissue samples were placed onto Visium Spatial Gene Expression slides. The slides consist of ~5,000 spot arrays with uniquely barcoded probes allowing for the capture of native mRNA. Captured molecules were sequenced on an Illumina NovaSeq. Using the Space Ranger v1.1 analysis pipeline and the Loupe Browser v4.1 desktop software, the RNA-seq data were merged with H&E and immunostained tissue images to align reads, perform clustering, and gene expression analysis. Results: We demonstrated an improved version of the spotted oligo array technology that increases tissue coverage and spatial resolution with reduced spot size, and increased spot number and packing density. We captured spatial patterns of gene expression and mapped the information back to H&E and immunostained images with regional annotations. Correlation analysis demonstrated differential spatial gene expression between the control and APPSWE [Tg2576] mouse model including the Pmch gene within the hypothalamic region. Conclusions: Together, this confluence of imaging and sequencing is a valuable tool for understanding the relationships between CNS cell types in healthy and diseased samples with an unbiased anatomical and gene expression-driven method.

P651 / #1107


THE ROLE OF NITRIC OXIDE AND CYTOKINES IN THE DEVELOPMENT OF GLAUCOMA PATHOGENESIS

Lecture Title:

H. Zanginyan1, G. Ghazaryan1, L. Hovsepyan2 1Institute of Molecular biology NAS of Armenia, Laboratory Of Molecular Membranology, Yerevan, Armenia, 2Institute of Molecular biology, Laboratory Of Molecular Membranology, Head of department Molecular membranology, Armenia

Aims: Primary open-angle glaucoma (POAG) is a chronic neurodegenerative disease with progressive death of retinal ganglion cells and optic nerve axon. It is accompanied by the loss of visual sensitivity fields. Studies have shown that neuropathy in glaucoma involves immunological factors. Purpose of the work: to study the production of nitric oxide and study the features of TNF-α pg / ml, IL-6, IL-8 in patients with POAG. Methods: 35 patients with POAG were examined. The age of the surveyed ranged from 35 to 75 years and averaged 55.6 years. The control group consisted of 20 ophthalmologically healthy people of comparable age. Blood serum served as the material for the study. The concentration of cytokines in the peripheral blood was determined by the ELISA method. Nitric oxide was determined by photometry at a wavelength of 546 nm. Results: As shown by the research results, patients have an increased level of nitric oxide, TNF-α, (44.0 ± 3.8 pg / ml versus 10.8 ± 2 pg / ml in healthy people,), IL-6 and IL-8 .The synthesis of NO synthase is induced by proinflammatory cytokines; macrophages activated by cytokines and bacterial endotoxins increase NO synthesis. The formation of cytokines is an important element in maintaining the homeostasis of the body. However, if there is an overproduction of cytokines, damage to the optic nerve is possible. Conclusions: Изменения концентрации цитокинов важны роль в механизме действия зрительного нерва при глаукоме, нормализации метаболизма оксида азота может быть новым способом лечения и ранней оценки степени тяжести патологических процессов.

P652 / #1297

Topic: Theme G: Huntington's and Other Neurodegenerative Diseases / G7. Animal Models

OCTODON DEGUS: A NEW RODENT MODEL TO STUDY PARKINSON’S DISEASE

Lecture Title:

L. Cuenca-Bermejo1,2,3, E. Pizzichini4, P. Gallo-Soljancic1,2, C. Sánchez-Rodrigo3, A.-M. González-Cuello1, E. Fernández-Villalba1,2,3, M.T. Herrero1,2,3 1Institute for Aging Research, University Of Murcia, Murcia, Spain, 2Biomedical Research Institute of Murcia, Clinical & Experimental Neuroscience (nice), Murcia, Spain, 3University of Murcia, Human Anatomy And Psychobiology, Murcia, Spain, 4Department of Biology and biotechnology “Charles Darwin” (BBCD), Sapienza, University Of Rome, Roma, Italy

Aims: The diurnal rodent Octodon degus (degus) has been claimed an interesting natural model for neuroscience and other human age-related disorders, since this animal spontaneously develops Alzheimer’s disease-like pathology, cognitive decline, diabetes, retinal degeneration or atherosclerosis. The main goal of this work was to test if the O. degus is sensitive to MPTP-induced neurotoxicity. Methods: Ten adult male degus were divided into control and MPTP groups. Animals from the MPTP group received two intraperitoneal injections of MPTP per week (10 mg/kg/injection, 5 weeks), until they reached a cumulative dose of 100 mg/kg. Motor activity was evaluated by open-field test and cognition by the Barnes Maze test. Dopaminergic degeneration was studied in both the ventral mesencephalon and the striatum. Neuroinflammation was analyzed in the Substantia Nigra pars compacta (SNpc), the Ventral Tegmental Area (VTA), the striatum and in the dorsal hippocampus. Results: As the cumulative dose of MPTP increased, the motor condition was significantly reduced in the parkinsonized degus compared with the control group, together with cognitive alterations. Postmortem analyses revealed a significant decrease in the number of TH+ neurons in the mesencephalon of MPTP animals (both SNpc and VTA) compared with the controls, although the dopaminergic terminals in the striatum were not significantly affected. Immunohistochemical expression of astroglial and microglial markers was significantly increased in both SNpc and striatum of parkinsonized animals, together with a significant microgliosis at the hippocampal level. Conclusions: These results show that the O. degus is susceptible to MPTP intoxication and, therefore, could be a suitable model for experimental Parkinsonism.

P653 / #855


OCD AND NEURODEGENERATION IN SPRED2-DEFICIENT MICE IS ASSOCIATED WITH BRAIN VENTRICLE ENLARGEMENT AND ULTRASONIC VOCALIZATION CHANGES

Lecture Title:

D. Hepbasli1, M. Ullrich1,2, K. Schuh1 1Intitute of Physiology, Ag Schuh, Wurzburg, Germany, 2Institute for rare Diseases, University Hospital Wuerzburg, Wurzburg, Germany

Aims: Deficiency of SPRED2, a protein ubiquitously expressed in brain and a potent suppressor of Ras/ERK/MAPK cascades, causes OCD in combination with neurodegeneration in mice. Prompted by increased brain ventricles and augmented levels of Tau and P-Tau in brains of SPRED2-KOs, the objective was to verify that OCD in SPRED2-KOs is accompanied by neurodegeneration. Neurodegeneration is often associated with language problems or underdevelopment. Therefore, we examined the communication of mice throughout their life. To determine differences in ultrasonic vocalizations (USV), we analyzed call rate, call subtype profile, and acoustic parameters (duration, bandwidth, and mean peak frequency) in young and old SPRED2-KOs and analyzed the calls with the deep-learning Software DeepSqueak. Methods: After perfusion of the brains with PFA, they were cut into 100µm slices using a Vibratom. 3D reconstruction and quantification of brain and ventricle volumes was done with Fiji and Imaris. Speech development and USV usage was analyzed by two self-trained Faster-RCNN neural networks in Deepsqueak. Results: Ventricle system size quantification in brains of young and old WT and SPRED2-KOs revealed a significant volume increase of the ventricle system of aged KOs. USV recording of male mice while interacting with females, revealed significant differences between WT and KO, both age- and genotype-dependent. Conclusions: Our findings confirmed neurodegeneration in SPRED2-KOs by an increase of the ventricle system volume and provided the first classification of WT vs. SPRED2-KO USVs. This indicates that SPRED2 is involved in the pathogenies of stress- and anxiety-related disorders like OCD but also of neurodegenerative disorders.

P654 / #264


LOSS OF RETINAL GANGLION CELLS IS ACCOMPANIED BY COMPLEMENT RESPONSE IN A NEW COMBINATION GLAUCOMA MOUSE MODEL

Lecture Title:

S. Reinehr1, J. Wulf1, A. Hensel1, R. Fuchshofer2, H.B. Dick1, S. Joachim1 1Ruhr-University Bochum, Experimental Eye Research Institute, Bochum, Germany, 2University Regensburg, Institute Of Human Anatomy And Embryology, Regensburg, Germany

Aims: Besides an elevated intraocular pressure (IOP) as a main risk factor, an involvement of the immune system is discussed in glaucoma. However, pathomechanisms are currently only investigated in models based on one pathogenic factor. Hence, we combined an IOP-dependent and an IOP-independent glaucoma model to study the underlying complex mechanisms more precisely. Methods: 6-week-old CTGF mice, which develop an IOP-dependent glaucoma damage, were used (CTGF). The corresponding wildtype animals served as controls (WT). To investigate immunological alterations, CTGF (Combination) and WT mice (ONA) were immunized with an optic nerve antigen homogenate. Six weeks after immunization, retinal ganglion cells and complement system components were examined via immunohistology (n=10/group) and quantitative real time PCR (RT-qPCR; n=4/group). Results: A loss of retinal ganglion cells was noted in ONA mice compared to WT (p=0.01). An even stronger RGC loss was observed in Combination animals in comparison to WT (p=0.009) and ONA mice (p=0.02). C3 mRNA levels, a component of the terminal complement system pathway, was significantly upregulated in ONA (p<0.001) and Combination (p<0.001) mice compared to WT animals. In addition, mRNA levels of all C1q subunits (C1qa, C1qb, and C1qc) were significantly upregulated in ONA (all: p<0.05) as well as in Combination mice (all: p<0.05), showing an activation of the classical pathway. Conclusions: The new combined glaucoma model reveals an authentic depiction of the multifactorial pathogenesis in glaucoma. Furthermore, the results emphasize the contribution of the complement system. Hence, complement inhibition might be a beneficial therapy for glaucoma patients in the future.

P655 / #1330


NEURONAL DELETION OF NDR1/2 KINASES IMPAIRS AUTOPHAGY AND LEADS TO NEURODEGENERATION

Lecture Title:

F. Rosianu, S. Mihaylov, A. Martiniuc, N. Eder, S. Claxton, H. Flynn, B. Snijders, S. Tooze The Francis Crick Institute, Kinases And Brain Development Lab, London, United Kingdom

Aims: In neurons constitutive autophagy plays a vital role in ensuring a constant turnover of organelles and proteins, while a failure to carry out autophagy results in neurodegeneration. NDR kinases are evolutionarily conserved serine/threonine kinases that play roles in membrane trafficking, cell polarisation, neuronal differentiation and autophagy. Two highly similar isoforms, NDR1 and NDR2, are found in mammals. We aim to study the roles of NDR1/2 in mammalian neurons in vivo. Methods: Using a conditional knockout mouse model, we deleted both NDR isoforms in excitatory neurons in the cortex and hippocampus. Results: Dual loss of NDR1/2 causes excessive membrane protrusions and Atg9 mislocalisation from Golgi to peripheral compartments as early as postnatal day 20. At later ages, we observed progressive accumulation of p62-positive aggregates and ubiquitinated proteins and a reduction in the number of LC3-positive autophagosomes, consistent with impaired autophagy. These alterations result in degeneration of upper cortical layers in parallel with astrocytic and microglial activation, hallmarks of neurodegeneration. Quantitative mass spectrometry analysis of NDR1/2 knock-out mice revealed major alterations in endocytosis proteins and confirmed previously-reported NDR1 substrates implicated in membrane trafficking (Ultanir et. al., Neuron, 2012). In cultured neurons infected with NDR1/2 shRNA transferrin uptake is significantly reduced and the surface levels of transferrin receptor and Atg9 are increased, indicating defects in membrane recycling. Retrograde Atg9 trafficking in axons is also significantly impaired. Conclusions: We conclude that NDR1/2 are required for membrane and Atg9 recycling, as well as efficient autophagy, and deletion of these kinases leads to neurodegeneration.

P656 / #1658

Topic: Theme H: Demyelinating Diseases / H6. Cell, Molecular and Systems Biology

NERVE GROWTH FACTOR NEUTRALIZATION PROMOTES OLIGODENDROGENESIS BY INCREASING MIR-219 LEVELS.

Lecture Title:

R. Scardigli1, R. Brandi2, C. Giorgi3, I. Arisi4, F. La Regina5, F. Malerba6, S. Amadio7, C. Volontè7, M. D'Onofrio2, A. Cattaneo8 1CNR, Institute Of Translational Pharmacology, Rome, Italy, 2European Brain Research Institute (EBRI), Genomics Facility, Roma, Italy, 3CNR, Institute Of Molecular Biology And Pathology, Rome, Italy, 4European Brain Research Institute (EBRI), Bioinformatics Facility, Rome, Italy, 5European Brain Research Institute (EBRI), Animal Care, Rome, Italy, 6European Brain Research Institute (EBRI), Ngf Lab, Rome, Italy, 7Fondazione Santa Lucia, Cellular Neurobiology, Rome, Italy, 8Scuola Normale Superiore (SNS), Laboratorio Di Biologia Bio@sns, Pisa, Italy

Aims: Alterations of the crosstalk between the nervous and immune systems may underlie the inflammatory and neurodegenerative changes characterizing Multiple Sclerosis (MS). Among the molecules that actively interconnect these two systems is the Nerve Growth Factor (NGF), involved in both regulating inflammation and in brain homeostasis. Additionally, NGF is known to modulate oligodendrogenesis and reduce myelination, further suggesting an involvement of NGF in MS ethiology. Despite that, the role of NGF in oligodendrocytes ontogenesis and myelination is still poorly defined, as are the underlying mechanisms involved. To better understand the molecular pathway by which NGF modulates myelination, we investigated NGF downstream targets relevant to oligodendrogenesis, focusing our attention on microRNAs. Methods: We used AD11 mice, in which postnatal expression of the anti-NGF antibody αD11 leads to NGF neutralization. We performed a microRNA profiling of AD11 brain samples and qRT-PCR analyses, as well as oligodendrocytes in vitro differentiation from AD11 neurospheres. Results: NGF deprivation leads to an increase of miR-219 levels and a downregulation of its predicted targets. AD11 neurospheres give rise to more oligodendrocytes and this process is dependent on miR-219, as shown by decoy-mediated inhibition of this microRNA. Moreover, treatment of AD11 neurospheres with NGF inhibits miR-219 upregulation and consequently oligodendrocytes differentiation, while aD11 treatment of WT progenitors and rat primary OPC increases miR-219 expression and the number of mature oligodendrocytes. Conclusions: Overall, this study indicates that NGF inhibits oligodendrogenesis and myelination by downregulating miR-219 levels, suggesting a novel molecular circuitry that can be exploited for the discovery of new effectors for remyelination.

P657 / #685

Topic: Theme H: Demyelinating Diseases / H7. Animal Models

EVALUATION OF THE NEUROPROTECTIVE EFFECT OF NEWLY SYNTHESIZED 4-AMINOPYRIDINE DERIVATIVES ON CUPRIZONE-INDUCED DEMYELINATION OF MICE BRAIN

Lecture Title:

I. Kostadinova1, B. Landzhov2, N. Danchev1, L. Vezenkov3 1Medical University of Sofia, Faculty of Pharmacy, Department Of Pharmacology, Pharmacotherapy And Toxicology, Sofia, Bulgaria, 2Medical University of Sofia, Faculty of Medicine, Department Of Anatomy, Histology And Embryology, Sofia, Bulgaria, 3University of Chemical Technology and Metallurgy, Department Of Organic Chemistry, Sofia, Bulgaria

Aims: Immunohistochemical evaluation of the protective effect of long-term administration of newly synthesized 4-aminopyridine derivatives (4-AP) on cuprizone-induced demyelination in mice. Methods: The model of demyelination was performed by the administration of neurotoxic agent cuprizone to mice. Cuprizone model of multiple sclerosis permits detection of changes in the brain of mice with different methology including ultrastructural analysis by electron microscope, biochemical aasay for myelin specific components, histochemical and immunohistochemical evaluation of changes in different structures in the brains of mice. Our modifications of the method were related to the mice strain, age and the route of cuprizone administration. The derivatives of 4-AP were administered orally to the mice for 6 weeks. Immunohistochemical studies were conducted with gluthation-S transferase isoform pi (GST-π) antibody in order to assess the degree of demyelination. Results: The results show that cuprizone, administered with the drinking water (0.2% concentration) for a period of 6 weeks leads to the induction of detectable demyelination. The results from the immunohistochemical assay indicate that some of the tested compunds antagonize the cuprizone-induced demyelination to a different degree. Conclusions: The results of our investigations proved that 16 week old albino mice, line H were susceptible to cuprizone-induced demyelination. According immunohistochemical evaluation the investigated compounds antagonize the cuprizone-induced demyelination to a different degree.

P658 / #731

Topic: Theme I: Lysosomal Storage Diseases / I1. Disease Mechanisms, Pathophysiology

GBA P.E326K PARKINSON’S DISEASE AND DEMENTIA WITH LEWY BODY MOUSE MODEL

Lecture Title:

L. Clark Columbia University Medical Center, Pathology And Cell Biology, New York, United States of America

Aims: Biallelic mutations in the Glucocerebrosidase (GBA) gene cause Gaucher’s disease (GD), one of the most common lysosomal storage disorders. We and others have shown that specific mutations in GBA, in the heterozygous state, are a risk factor for Parkinson’s disease (PD) and Dementia with Lewy bodies (DLB). Among the GBA variants associated with PD and DLB, the p.E326K allele is the most common risk factor. To further our understanding of the mechanistic link between this specific GBA variant and the development of PD and DLB, we are developing a GBA p.E326K mouse model. Methods: A point mutation knockin mouse model, Gba p.E344K (aka GBA p.E326K), was generated by genome editing using CRISPR-Cas9 technology. Behavioral phenotyping will be performed to assess motor and cognitive function. Neuropathological evaluation of brains will include assessment of Lewy bodies (a-Synuclein) and Glucosylceramide. Results: A total of 30 F0 mice were produced by microinjection and 25 positive Gba p.E344K F1 mice were identified by PCR and sequencing. Characterization of the mouse model is currently ongoing and behavioral and neuropathological data will be presented. Conclusions: We are developing a mouse model of GBA p.E326K associated PD and DLB. Currently, therapies being developed to treat GBA associated PD include glucosylceramide synthase inhibitors and the molecular chaperone, ambroxol hydrochloride. Development of a mouse model for the most common GBA variant associated with PD, the GBA p.E326K variant, and determining the disease mechanism will allow a personalized medicine approach and open up new avenues for therapeutic development. Funding: National Institutes of Health NS113038 (Clark).

P659 / #941

Topic: Theme I: Lysosomal Storage Diseases / I1. Disease Mechanisms, Pathophysiology

DYRK1A CHANGES AND MODIFICATION OF THE ENDO-LYSOSOMAL COMPARTMENT IN THE LOCUS COERULEUS OF POST-MORTEM HUMAN BRAINS IN NEURODEGENERATIVE DISEASE

Lecture Title:

M. Fructuoso1, A. Mohammad1, J.-M. Delabar1, S. Boluda1,2,3, L. Stimmer2, Y. Vermeiren4,5, P. De Deyn4,5, C. Duyckaerts1,2, M.-C. Potier1,2 1Paris Brain Institute, Team Alzheimer’s And Prion Diseases, Paris, France, 2Institut du cerveau, INSERM U1127, CNRS UMR7225, Sorbonne Universités, Paris, France, 3Laboratoire de neuropathologie Raymond Escourolle, Sorbonne Université, Hôpital de la Pitié-Salpêtrière, Département De Neuropathologie - Pôle Des Maladies Du Système Nerveux, Paris, France, 4Institute Born-Bunge, University of Antwerp, Department Of Biomedical Sciences,laboratory Of Neurochemistry And Behaviour, Wilrijk, Belgium, 5University of Groningen and University Medical Center Groningen, Department Of Neurology And Alzheimer Research Center, Groningen, Netherlands

Aims: The locus coeruleus (LC) is the main source of noradrenaline in the brain, and its neurodegeneration is an early event in Alzheimer’s disease (AD), Parkinson’s disease (PD), and Down syndrome (DS). DYRK1A, a kinase encoded by a gene mapping to chromosome 21, is overexpressed in DS and phosphorylates proteins from signaling pathways relevant to AD pathogenesis, from the endolysosomal pathway, and induces alterations in noradrenergic profiles. Our study addresses the relationship between levels of DYRK1A, LC neurodegeneration, and endolysosomal alterations in AD, PD and DS. Methods: We measured the levels of DYRK1A in frozen LC, hippocampus and prefrontal cortex of AD, PD, DS, and healthy age-matched controls (Ctr) using a customized Meso Scale Discovery (MSD) assay. Paraffin fixed LC sections of the same individuals were used for the analysis of the density of pathological markers (Aβ, P-TAU, α-synuclein), the endosomes (Rab5), and lysosomes (Cathepsin B) in tyrosine hydroxylase positive cells using immunolabeling. Results: PD and AD LC presented the most severe noradrenergic cell loss. DS individuals with dementia had the most prominent tau pathology. As compared to Ctr, AD cases showed enlarged endosomes whereas PD cases had smaller lysosomes. As expected, DYRK1A levels in DS brains were increased. Additionally, according to previous data in plasma, our preliminary results showed non-significant lowering of DYRK1A levels in dementia-affected brains, especially in prefrontal cortex of PD cases. Conclusions: We validate the use of human homogenates for DYRK1A dosage, and propose a role of DYRK1A in dementia. In addition, we conclude that endolysosomal alterations may be disorder-specific.

P660 / #1552

Topic: Theme I: Lysosomal Storage Diseases / I4. Imaging, Biomarkers, Diagnostics

TRANSCRANIAL SONOGRAPHY (TCS) – POSITRON EMISSION TOMOGRAPHY (PET) RELATIONSHIP IN PATIENTS WITH GBA1 MUTATIONS

Lecture Title:

G. Lopez1, D. Eisenberg2, M. Gregory2, K. Berman2, E. Sidransky1 1National Institutes of Health, National Human Genome Research Institute, Bethesda, United States of America, 2National Institutes of Health, National Institute Of Mental Health, Bethesda, United States of America

Aims: Mutations in GBA1, the gene coding for the lysosomal enzyme glucocerebrosidase, are the most common genetic risk factor for parkinsonism. We employed both ultrasonography and molecular neuroimaging in individuals carrying GBA1 mutations with and without Parkinson disease and examined the relationship between maximal substantia nigra echogenicity area with [18F]-FDOPA specific uptake (Ki). Methods: Thirty-four patients with GBA1 mutations (7 with PD), underwent both [18F]-FDOPA PET neuroimaging and TCS nigral echogenicity measurement conducted independently by two different examiners during the same patient visit. Maximal area of echogenicity was obtained bilaterally, as well as [18F]-FDOPA specific uptake (Ki) analyzed by voxel-wise interrogation. Results: Individuals with PD were found to have a larger echogenic area than those without PD, consistent with current ultrasonography literature in these patients. There was a robust inverse relationship between TCS measurements and 18[F]-FDOPA specific uptake (Ki) across individuals with PD. The greater echogenicity in the subjects with PD paralleled the group differences observed with 18[F]-FDOPA PET, in which significantly reduced dopamine synthesis was exclusively seen in subjects with clinical PD. However, in at- risk subjects without PD, the TCS results did not correlate with Ki on PET, suggesting that neither modality may be predictive. Conclusions: In our cohort, there is a close correspondence between nigral echogenicity measurements and striatal presynaptic dopamine synthesis capacity in subjects with GBA1 associated PD but not in carriers without PD. Continued evaluation may shed light on the utility of TCS as a predictive tool in the prodromal phase of the disease.

P661 / #615

Topic: Theme J: COVID-19: Impact on Brain Neurodegenerative Diseases / J1e. Impact of Covid-19 on clinical trials: Telemedicine, Digital Health and E-Trials

ALZHEIMER’S DISEASE CLINICAL TRIAL ADAPTATIONS TO COVID-19

Lecture Title:

C. Martinez, J. Stanek, B. Swearingen T3D Therapeutics, Inc., Clinical Development, Research Triangle Park, United States of America

Aims: To provide a new research framework for conducting hybrid remote/onsite clinical outcomes assessments in AD clinical trials to mitigate operational compromises arising from COVID-19 demands. We present the PIONEER study – T3D959-202 as an example of implementing this new framework for remote adaptations to AD clinical trials. Methods: The risk of contracting the SARS-CoV-2 virus in health care settings impacts clinical sites, monitors, subjects, and caregivers. Concerns for subject and site adherence to protocol, data reliability and consistency, and trial integrity have caused disruptions to clinical trials. We utilize ongoing adaptations to the PIONEER study – T3D959-202 to address clinical research concerns. Results: Adaptations of the PIONEER study provide a framework for hybrid remote/onsite clinical outcomes assessments with caregiver assistance. Continuous eSource and remote monitoring protect data quality against interruptions to onsite monitoring. Training of home-health nurses, cognitive and clinical raters, and subjects and caregivers on contingent at-home procedures protects data integrity should a subject or caregiver be unable or unwilling to come into the clinic. Modification of the statistical analysis plan factors in the potential loss of subjects. Conclusions: The hybrid remote/onsite research framework in the PIONEER study implements modifications that overcome obstacles to trial conduct presented by the COVID-19 pandemic. Application of this framework provide: (1) support alternatives to onsite visits; (2) facilitates technology use to allow for seamless adaptation to at-home visits; (3) maintains the integrity of study data; and (4) plans for modifications to statistical analysis due to new impediments to subject retention.

P662 / #1258

Topic: Theme K: Psychiatric Symptoms in Neurodegenerative Diseases / K1. Disease Mechanisms, Pathophysiology

INVESTIGATING INFLAMMATORY RESPONSES IN A CORTICOSTERONE-INDUCED MODEL OF DEPRESSION

Lecture Title:

S.L. Dunnett, R. Wang, R.C.-C. Chang Laboratory of Neurodegenerative Diseases, School of Biomedical Sciences, LKS Faculty of Medicine, The University Of Hong Kong, Pokfulam, Hong Kong PRC

Aims: In this project, we aim to use a corticosterone-induced model to investigate the inflammatory responses that occur in depression, which may further increase the risk of developing neurodegenerative diseases in the future. Methods: Corticosterone was administered daily to Sprague-Dawley (SD) rats for 14, 21 or 28 days. Afterwards, a series of behavioral tests were performed to assess their anxiety, memory and depressive behaviors. Different brain regions and tissues were collected to conduct immunohistochemical analysis and qPCR. Results: Following 21 days of corticosterone administration, subjects displayed decreased adrenal and bodyweight, indicating successful absorption of corticosterone into the system. From the Open Field test, we observed the development of anxiety-like behaviors. Additionally, in the 21-day time point, we observed the development of systemic inflammation from qPCR analysis, but no significant neuroinflammation. Conclusions: In these experiments, we utilized a corticosterone-induced model of depression to investigate the possible inflammatory links between depression and neurodegenerative diseases. 21 days of corticosterone treatment was sufficient to lead to the development of anxiety-like symptoms, as well as leading to the development of systemic inflammation, yet no neuroinflammation. This is the first time we have observed this phenomenon, which deserves further investigation. Future work will be conducted to see how the inflammatory response changes over different durations of corticosterone treatment, and the impact this has on the brain. Acknowledgement: SLD is awarded by Hong Kong PhD Fellowship.

P663 / #928


RISK OF DEVELOPING EPILEPSY IN ALZHEIMER'S DISEASE PATIENTS

Lecture Title:

J.H. Lee National Health Insurance Service Ilsan Hospital, Neurology, Goyang-si, Korea, Republic of

Aims: Previous studies have reported conflicting results about the prevalence of seizures in AD, and few epidemiological studies in the Asia have been found. We examine the demographic, clinical characteristics and the incidence for seizures with AD patients compared to non-AD patients in a prospective, longitudinal, community-based cohort with long follow-up. Methods: The data was collected from National Health Insurance Service-National Elderly cohort(NHIS-elderly) Database to define patients with AD from 2004-2006 based on having KCD code G30, F00. We performed a 1:5 case-control matched analysis using propensity score matching, which were based on age, sex and household income. We conducted Cox proportional hazards regression analysis to estimate the risk of epilepsy in AD. Results: In the cohort study, the patients with AD have higher chances for epilepsy than those without AD The Hazard ratios for epilepsy with AD are 2.773(95% CI: 2.515-3.057). The study also shows that the gender male, comorbidities such as hypertension, hyperlipidemia, diabetes, and chronic kidney disease increases the risk of developing epilepsy. Patients have 1.527 (95% CI: 1.375–1.695) times higher mortality rate when epilepsy occurred in the AD group than in the control group Conclusions: Out data show that AD significantly has higher risk for the development for epilepsy.

P664 / #386


DEPRESSION AND NIGRAL NEURON DENSITY IN LEWY SPECTRUM DISEASES

Lecture Title:

L. Saari1, L. Heiskanen1, M. Gardberg2, V. Kaasinen1 1Turku University Hospital, Neurocenter, Turku, Finland, 2Turku University Hospital and Institute of Biomedicine, University of Turku, Department Of Pathology, Laboratory Division, Turku, Finland

Aims: Parkinson’s disease (PD) and other Lewy body spectrum diseases (LBDs) are associated with a specific risk for clinical depression. Previous studies have suggested that lower numbers of mesencephalic dopamine neurons are associated with depressive symptoms in elderly individuals without LBD. In this clinicopathological study, we sought out to investigate whether LBD patients with comorbid depression have lower densities of dopaminergic neurons in the substantia nigra pars compacta (SNc) compared to patients without depression. Methods: Nigral tyrosine-hydroxylase (TH)-positive neuron densities were measured postmortem from midbrain sections. The area of SNc was evaluated with Fast Luxol Blue staining and neurons were semi-automatically counted. Seventy-three LBD patients were included to the study (43 patients with a clinical phenotype of PD and 28 patients with dementia with Lewy bodies). Twelve patients had clinical co-morbid depression antemortem extending to late stage of LBD. Results: SNc neuron density (n/mm²) was significantly lower in patients with depression than in patients without depression (4.34 vs 6.60, p<0.001). There were no differences in other variables between groups, such as motor disease severity, antiparkinsonian medications or disease duration. Conclusions: The results of this study indicate that clinical depression in LBD is associated with lower densities of TH-positive neurons in the SNc. The results thus point to a relevant role of mesencephalic dopamine neurons in the modulation of mood in LBD patients.

P665 / #1143


MACHINE LEARNING APPROACHES TO POST-MORTEM PHENOTYPE BRAIN DONORS: A NEW WAY TO STUDY MOLECULAR CHANGES IN THE BRAIN

Lecture Title:

J. Vogelgsang1, S. Berretta1, T. Klengel2, T. Mccoy3 1McLean Hospital, Harvard Medical School, Dpartment Of Basic Neuroscience, Translational Neuroscience Laboratory, Belmont, United States of America, 2McLean Hospital, Harvard Medical School, Dpt. Of Basic Neuroscience, Translational Molecular Genomics Laboratory, Belmont, United States of America, 3MGH, Dept. Of Psychiatry, Center For Quantitative Health, Harvard Medical School, Boston, United States of America

Aims: Clinical observation, medical history and a psychopathological examination describing different pathological behaviour or symptoms are the pillars of psychiatric diagnosis and treatment planning. In contrast to other medical disciplines, supportive technical diagnostics tools, such as laboratory measurements or imaging are less well established to facilitate preventive, diagnostic, and therapeutical efficacy monitoring approaches. This study aims to combine post-mortem psychiatric dimensional phenotyping using machine learning approaches and neuropathological changes. Methods: Donated brains from the Harvard Brain and Tissue Resource Center (HBTRC) underwent a neuropathological examination including Braak & Braak staging, semiquantitative grading of amyloid plaques, neuronal cell loss, and microvascular lesions. Clinical records as provided by the caregivers were digitalized and RDoCs were quantified using patter recognition techniques. RDoCs were correlated with neuropathological changes in the corresponding brains. Results: Cognition highly correlates with amyloid plaques in frontal and parietal lobe but not in the temporal or occipital lobe. No correlations between RDoCs and B&B stages were observed. Conclusions: Post-mortem AI based analysis of electronic health records is a novel and promising approach to include multidimensional, clinical orientated and symptom-based evaluation of corresponding donors. This approach enables novel strategies to study a broader span of psychiatric changes in post-mortem human brain tissue.

P666 / #1245

Topic: Theme K: Psychiatric Symptoms in Neurodegenerative Diseases / K2. Therapeutic Targets, Mechanisms for Treatment

COGNITIVE-BEHAVIOURAL GROUP THERAPY FOR THE TREATMENT OF SOCIAL ANXIETY IN SCHIZOPHRENIA AND ITS IMPACT ON QUALITY OF LIFE: A META-ANALYTIC SYNTHESIS

Lecture Title:

M. Azharuddin1, M. Sharma2 1School of Pharmaceutical Education and Research, Jamia Hamdard, Pharmaceutical Medicine, Division Of Pharmacology, New Delhi, India, 2School of Pharmaceutical Education and Research, Jamia Hamdard, Pharmacology, New Delhi, India

Aims: There is clinical uncertainty over the effectiveness of CBGT for the treatment of social anxiety in schizophrenia. This study estimated the effectiveness of CBGT interventions in improving social anxiety symptoms, general anxiety, distress, depression, symptoms of schizophrenia, and quality of life. Methods: A systematic search on MEDLINE and Cochrane Central Register of Controlled Trials, was performed with pairing relevant keywords to identify the articles for the last 10 years. Eligible studies evaluating the efficacy of CBGT versus control group for social anxiety in schizophrenia. The Outcome measures were social anxiety, general psychopathology, and quality of life. A random-effects model was used to calculate the pooled mean difference (MD) with 95% confidence interval (CI). Meta-analysis was performed using RevMan 5.3 software. Results: Studies were included with a total of 49 participants, the majority was male. Mean age of patients was 39.6 years. Results from the meta-analysis showed significant reductions in symptoms of social anxiety; Social Interaction Anxiety Scale (SIAS) were observed after treatment [MD -7.48; 95% CI -14.46, -0.50, P=0.04]. Significant improvement was also observed in general psychopathology; Calgary Depression Scale for Schizophrenia (CDSS) [MD -5.09; 95% CI -6.73, -3.45, P <0.001]. There was no significant heterogeneity amongst the studies [I2=47%; P=0.07]. There was no significant improvement observed in the quality of life [MD 4.64; 95% CI -1.16, 10.45, P= 0.12] after treatment. Conclusions: The current study suggests that CGBT for social anxiety disorder is effective in schizophrenia. Furthermore, evidence from well-controlled RCTs and real-world studies with long-term follow-up required to confirm the present findings.

P667 / #663


IMPROVEMENT OF IMPULSE CONTROL DISORDER IN PARKINSON'S DISEASE PATIENTS TREATED WITH SUBTHALAMIC NUCLEUS STIMULATION

Lecture Title:

E.J. Choi1, C.S. Lee2, D.G. Lee3 1Ulsan University Hospital, Ulsan University College of Medicine, Department Of Neurology, Ulsan, Korea, Republic of, 2ASAN MEDICAL CENTER, Department Of Neurology, Seoul, Korea, Republic of, 3Faculty of Medicine, The University of British Columbia, Vancouver, British Columbia, Canada, Neurology, Vancouver, Canada

Aims: To find effects of subthalamic deep brain stimulation on impulse control disorder in patients with Parkinson's disease Methods: Subjects: Among 69 patients with PD who received STN-DBS between March 2013 and July 2015 in Asan Medical Center, eleven from fifteen patients with ICD were selected based on Questionnaire for ICD in PD (QUIP) scores (4 or higher) before DBS. Evaluation: UPDRS III, QUIP, Addenbrooke's cognitive examination (ACE-R) before and after DBS surgery. PET: [18F]FDG and [18F]FP-CIT PET were performed for all subjects. Regional metabolic rates were obtained from the ratio using the gray matter (GM) as reference. MRI: 3.0T MRI(pre-DBS MR 3.0T) was performed for all subjects. Data analysis: UPDRS III, QUIP and ACE-R scores were compared between pre-DBS and post-DBS stages. Results: The eleven patients with ICD had a mean age 40.4±9.2 (mean±SD), and the mean age of PD onset was 46.6±10.7. The mean interval between the onset of Parkinsonism and the onset of ICD was 4.91 ±2.77 years. The mean duration from ICD symptoms to get STN DBS operation was 3.18 ±2.99. After the STN DBS operation, the amount of L-dopa necessary was 668±314mg/day that might be decreased to a half of the amount needed before the operation. (p<0.005) Also, the QUIP score of the patients was 5.3±2.0 before STN DBS, and 1.5±1.8 after STN-DBS (p<0.005). Conclusions: Our data showed that ICD is more common in young patients with PD, and that STN-DBS reduced LEDD markedly, to a half of pre-DBS LEDD, and provided evidence that STN-DBS improved ICD.

P668 / #305


MEDICINAL IMPORTANCE AND VISNAGIN FOR THE TREATMENT OF NEUROINFLAMMATORY DISORDERS: PHYSIOLOGICAL FUNCTIONS IN THE OXIDATIVE STRESS THROUGH SCIENTIFIC DATA ANALYSIS

Lecture Title:

D. Patel Sam Higginbottom University of Agriculture, Technology and Sciences, Department Of Pharmaceutical Sciences, Payagraj, India

Aims: Visnagin is the main active phytoconstituents of Ammi visnaga belongs to the furanochromone class chemical which have physiological functions against oxidative stress and neurodegenerative disorders. Methods: In order to know the medicinal importance and of visnagin for the treatment of neurodegenerative disorders, in the present investigation biological potential of visnagin against oxidative stress and neurodegenerative disorders have been investigated through scientific data analysis of various literature works. Importance of visnagin for the treatment of neuroinflammatory disorders have been investigated in the present investigation through literature data analysis of various scientific research works. However pharmacological activities of visnagin for their effectiveness against oxidative stress induced neurodegenerative disorders have been also investigated in the present investigations through literature data analysis. Literature data analysis of the numerous scientific research works of visnagin have been also carried out to know the biological importance of and health beneficial potential of visnagin in the medicine. Results: Scientific data analysis of various scientific research work of the literature revealed the biological importance of visnagin in the medicine and other allied health sectors. Literature data analysis in the present investigation revealed the biological importance of visnagin in the neuroinflammation and oxidative stress. Literature data analysis revealed the biological importance of visnagin on cell injury. Molecular study data analysis signified the therapeutic importance of visnagin in the medicine for the treatment of oxidative stress induced neurodegenerative disorders. Conclusions: Literature data analysis of various research works signified the biological potential of visnagin in the oxidative stress induced neurodegenerative disorders.

P669 / #1088

Topic: Theme K: Psychiatric Symptoms in Neurodegenerative Diseases / K3. Drug Development, Clinical Trials

MEASUREMENT OF FACIAL, VOCAL, AND LIMB TREMOR USING MACHINE LEARNING CORRELATES WITH IN-PERSON CLINICAL SCORES

Lecture Title:

A. Abbas, V. Yadav, D. Desai, L. Zhang, V. Koesmahargyo, A. Paley, I. Galatzer-Levy AiCure, Research And Development, New York, United States of America

Aims: Measurement of tremor induced by disorders such as Parkinson’s Disease and Essential Tremor can be subjective and have poor reliability. Traditional in-person assessments are burdensome for both patients and clinicians. We demonstrate the accuracy of machine learning methods that can objectively and remotely quantify facial, vocal, and hand tremor, along with their validity to track treatment response in patients. Methods: 222 individuals (106 Female) with Essential Tremor enrolled in a randomized controlled trial were assessed by trained raters for tremor severity using the The Essential Tremor Rating Assessment Scale (TETRAS), including individual assessment of facial, vocal, and limb tremor. Videos of these assessments (n = 490) were used to train and test machine learning-based measurements of facial tremor, vocal tremor, and hand tremor. Results: Measurement of facial tremor correlated strongly with the TETRAS (p = 0.03) and tracked alongside the TETRAS over the 4 timepoints of the study [F(3,30) = 0.31; p= 0.81]. Measurement of vocal tremor correlated with the TETRAS (p = 0.02). Importantly, the vocal tremor score showed a significant response to treatment [F(3,45) = 4.33; p = 0.009] when the traditional score did not [F (3,45) = 1.82; p = 0.16]. Finally, measurement of hand tremor correlated strongly with the TETRAS total (p = 0.01) and tracked alongside the TETRAS over the course of the study [F(3,39) = 2.02; p = 0.13]. Conclusions: Machine learning-based measurement of tremor allows for accurate and objective assessment of disease severity and treatment response in individuals with tremor disorders. Digital measures can be used in clinical applications requiring low-burden assessments.

P670 / #309


COGNITIVE PROFILE IN VERY EARLY STAGE PARKINSON DISEASE

Lecture Title:

A. Evlice1, M. Erdem2, D.M. Demirkiran3 1ADANA CİTY HOSPİTAL, Neurology, adana, Turkey, 2ÇUKUROVA UNİVERSİTY, Neurology, adana, Turkey, 3CUKUROVA UNİVERSİTY, Neurology, adana, Turkey

Aims: Parkinson’s disease (PD) is a neurodegenerative disorders characterized by both motor and nonmotor symptoms mainly due to striatal dopamine deficiency. Dementia is an important nonmotor symptom of PD. The purpose of this study is to determine the cognitive profile in very early stages of PD and to investigate its relation to the clinical features of the disease. Methods: Thirty PD patients with a disease duration of ≤5 years and 20 healthy controls were included in this study. MMSE, digit span (DS), clock drawing (CD), verbal and visual memory tests were administered. Correlation of neurocognitive tests with age, education, duration of disease, initial symptom, treatment groups, Unified Parkinson’s Disease Rating Scale motor scores (UPDRS) and Hoehn & Yahr Staging (H&Y) scores were evaluated. All analyses were performed using SPSS 20 statistical software package programme. Results: There were 30 PD patients (11 female) and 20 healthy controls (10 female). Duration of disease was 29.42 ± 16.88 (3-60) months. There was a significant difference only in CD between PD and healthy controls. Patients with H&Y stage 1 (unilateral involvement) of PD were better than H&Y stage 2 (bilateral involvement) on backward DS, CD, and visual memory tests. Verbal memory negatively correlated with both age and UPDRS motor scores. DS tests showed negative correlation with treatment groups, they were worse in L-dopa treated patients. Conclusions: This study shows that CD, DS, verbal and visual memory tests were affected even in very early PD, in relation to age and severity (bilateral involvement) of disease.

P671 / #661


ROLE OF NATURAL HERBS TO AVERT PARKINSONS ASSOCIATED DEFICITS: ASSOCIATION WITH AGE, DIABETES, BEHAVIOR AND DOPAMINE IN BRAIN

Lecture Title:

S.A.M. Khan, A. Khan, H. Rafiq THE AGA KHAN UNIVERSUTY, Biological And Biomedical Sciences, KARACHI, Pakistan

Aims: Background: Parkinson's disease (PD) is a neurodegenerative disease, characterized by tremors, stiffness and difficulties in walking, balance, and coordination with dopamine deficiency in the brain. Dopamine is released by substantia nigra and is responsible for transmitting messages that plan and control body movements. Few FDA-approved drugs are available for treatment but possess unwanted side effects. Therefore, finding cost-effective herbal remedies with minimal side effects and better efficacy is necessary. For this reason, methanolic extracts from natural herbs saffron (S) and chamomile (C) are considered in this study for treatment. Methods: Fifty aged, diabetic white albino Wistar rats of 2 years, weighing 300 grams, were randomly divided into five groups (n=5), healthy control groups (HC), disease controls (DC), and test groups receiving S (10 mg/kg), C (30 mg/kg), S-C (10mg/kg-30mg/kg) respectively for two weeks. Behavioral tests for locomotion, motor impairment, exploration, and memory were evaluated through field tests (OFT) and new object recognition tests (NOR) and brain biochemistry comprised of DA and MAO analysis. Results: it was observed that brain biochemistry locomotion, movement disabilities and memory in DC decreased significantly in the OFT, NORT, while a significant improvement was observed in all test groups and HC with increased brain dopamine and MAO inhibition especially in the combination group. Conclusions: Natural herbs saffron and chamomile with inhibitory MAO activities and DA modulation can avoid age-related brain dysfunctions causing movement disorders, and despair that are hall marks of PD. Clinical trial with same herbs on human subjects is currently underway for treatment.

P672 / #862

Topic: Theme K: Psychiatric Symptoms in Neurodegenerative Diseases / K4. Imaging, Biomarkers, Diagnostics

REPEATED, SELF-ADMINISTERED, WIRELESS DRY EEG IN THE HOME RELIABLY MEASURES THE NEUROPHYSIOLOGICAL BASIS OF COGNITIVE PROCESSES IN OLDER ADULTS

Lecture Title:

F. Barbey1,2, F. Farina1, A. Buick3, L. Rueda Delgado2, J. Dyer3, M.N. Islam2, B. Murphy2, H. Nolan2, R. Whelan1 1Trinity College Dublin, Psychology, Dublin, Ireland, 2BrainWaveBank Ltd, Brainwavebank, Dublin, Ireland, 3BrainWaveBank Ltd, Brainwavebank, Belfast, United Kingdom

Aims: Neurodegeneration progresses slowly, making early detection and on-going tracking a challenge. Electroencephalography (EEG) is a non-invasive measure of brain activity but its burdensomeness has prevented large scale use. In contrast, wireless mobile dry EEG technology could provide an easy-to-use, scalable, objective, and alternative way of measuring of brain function suitable for large-scale screening and treatment tracking. Methods: We report data from a 6-week long study (n=52 healthy participants, mean age: 67.9 years, std:5.0; 26 female) using a wireless dry EEG headset (www.brainwavebank.com). Participants completed gamified visual oddball and Flanker tasks, during at-home unsupervised EEG. EEG signals were automatically pre-processed and poor quality sessions removed. We quantified signal variability by computing standard deviation of evoked potentials across multiple session aggregates, which were computed by randomly selecting, without replacement, N sessions from each participant’s data. Aggregated sessions were then averaged, and the process repeated 250 times. Results: In total, 1,530 EEG sessions were collected. On average, participants recorded 29.3 sessions (of 30 sessions requested, std: 7.6). Signal variability decreased nonlinearly when increasing the numbers of sessions aggregated.

Conclusions: This proof-of-concept study demonstrated that it is possible to reliably collect clinically relevant domain-specific markers of brain function and cognitive performance – remotely and without researcher supervision – via wireless dry-sensor EEG. Older adults demonstrated a very high adherence to the system – near-daily use. This finding has strong implications for AD research as it suggests that larger scale brain electrophysiology studies evaluating cognitive function are possible for the first time.

P673 / #561


CORRELATION OF FRONTAL ATROPHY AND CSF TAU LEVELS WITH NEUROPSYCHIATRIC SYMPTOMS IN PATIENTS WITH COGNITIVE IMPAIRMENT: A MEMORY CLINIC EXPERIENCE

Lecture Title:

M. Cotta Ramusino1, G. Perini1, M. Capelli1, M. Picascia2, D. Franciotta3, L. Farina4, A. Costa1 1IRCCS Mondino Foundation, Department Of Brain And Behavior, Pavia, Italy, 2IRCCS Mondino Foundation, Laboratory Of Neuropsychology, Irccs Mondino Foundation, Pavia, Italy, Pavia, Italy, 3IRCCS Mondino Foundation, Laboratory Of Neuroimmunology, Irccs Mondino Foundation, Pavia, Italy, Pavia, Italy, 4IRCCS Mondino Foundation, Neuroradiology Unit, Irccs Mondino Foundation, Italy, Pavia, Italy

Aims: Behavioral and psychological symptoms of dementia (BPSD) are a distressful condition. We aimed to investigate the BPSD distribution in subjects with cognitive impairment, and the potential correlations between BPSD and neurodegeneration in terms of cerebrospinal fluid (CSF) tau and brain atrophy. Methods: One-hundred patients with mild cognitive impairment (MCI) or dementia (Alzheimer’s disease; Lewy-body disease, LBD; frontotemporal dementia; vascular dementia) underwent a complete diagnostic workup, including 3T-MRI and/or CT and CSF. Cortical atrophy was assessed with MTA, PA and GCA-F scales. BPSD were rated using Neuropsychiatric Inventory (NPI), and BPSD clusters were defined according to the European Alzheimer Disease Consortium. Results: Delusions, hallucinations and psychosis cluster were differently distributed among the diagnostic groups (p<0.05, p<0.001, and p<0.05), with LBD patients showing higher scores for hallucinations (vs MCI, p<0.001, and AD, p<0.05) and psychosis cluster (vs MCI, p<0.05). In primary dementias, we found a negative trend between NPI total score and tau levels (p=0.08), while a positive relationship was observed in MCI (p=0.60). Higher GCA-F scores were associated to delusions and apathy (p<0.05, on both hemispheres) and to hallucinations (left: p<0.01, right: p<0.05). GCA-F scores were positively correlated with delusions and psychosis cluster (right: p<0.05), and agitation/aggression (left: p<0.05). Conversely, nighttime disturbances were positively correlated with both GCA-F and MTA scores (left: p<0.01; right: p<0.05). Conclusions: Our results suggest that psychotic symptoms are significantly more represented in LBD patients and that CSF tau and frontal atrophy could be useful indicators of the occurrence and severity of BPSD in clinical practice.

P674 / #288


EARLY DETECTION OF PSYCHOSIS SYMPTOMS IN PARKINSON’S DISEASE BASED ON NEUROPSYCHOLOGICAL TESTS AND DEMOGRAPHICS: A MACHINE LEARNING APPROACH

Lecture Title:

A. Fernandez-Quilez1,2, T. Tjersland3, A. Podhraski3, K. Oppedal1,4, M.C. Gonzalez2,5 1Stavanger University Hospital, Department Of Radiology, Stavanger Medical Imaging Laboratory (smil), Stavanger, Spain, 2University of Stavanger, Department Of Quality And Health Technology, Faculty Of Health Sciences, Stavanger, Norway, 3University of Stavanger, Faculty Of Science And Technology, Stavanger, Norway, 4Stavanger University Hospital, Centre For Age-related Medicine (sesam), Stavanger, Norway, 5Stavanger University Hospital, The Norwegian Centre For Movement Disorders, Stavanger, Norway

Aims: Early detection of psychosis symptoms in Parkinson’s Disease (PD) patients based on neuropsychological tests and demographic data and to determine the most important tests and features for it. Methods: 416 subjects diagnosed with PD and part of the Parkinson’s Progression Markers Initiative (PPMI) were used to evaluate machine learning (ML) techniques such as logistic regression (LR), support vector machines (SVM), decision trees (DT) and random forests (RF). Given the heterogeneous nature of the PPMI data different preprocessing techniques were evaluated, such as mode and mean imputation to deal with missing values. Furthermore, due to the small population (n=416) and large number of features dimensionality reduction techniques such as principal component analysis (PCA) were employed. PCA was compared to using the total score of each neuropsychological test instead of each individual item contributing to it. Finally, the longitudinal nature of the PPMI data was exploited by deriving features that contained longitudinal information useful for the task at hand. Results: We obtained an area under the curve (ROC-AUC) of 0.792 with SVM, derived longitudinal features and by making use of the total score of each neuropsychological test as a dimensionality reduction technique. Inference of the results showed that duration of PD and Montreal Cognitive Assessment were among the most important features to detect early psychosis symptoms. Conclusions: This study shows the plausibility of the application of machine learning techniques to the early detection of psychosis symptoms in PD patients and to identify those patients at risk for more rapidly progressive disease.

P675 / #410


PSYCHOLOGICAL RESILIENCE ENHANCES THE ORBITOFRONTAL NETWORK IN THE ELDERLY WITH MILD COGNITIVE IMPAIRMENT

Lecture Title:

C. Pae Bucheon St Mary Hospital Catholic University of Korea, Psychiatry, Bucheon, Korea, Republic of

Aims: It has been suggested that maintaining the efficient organization of the brain's functional connectivity (FC) supports neuroflexibility under neurogenerative stress. This study examined psychological resilience-related FC in 112 older adults with mild cognitive impairment (MCI). Methods: Using a resting-state functional magnetic resonance imaging (fMRI) approach, we investigated reorganization of the orbitofrontal gyrus (OFG)/amygdala (AMG)/hippocampus (HP)/parahippocampal gyrus (PHG) FC according to the different levels of resilience scale. Results: Compared with the low resilient group, the high resilient group had greater connectivity strengths between the left inferior OFG and right superior OFG (P < 0.05, Bonferroni corrected), between the right inferior OFG and left PHG (P < 0.05, Bonferroni corrected), and between the right middle OFG and left PHG (false discovery rate < 0.05). Conclusions: Psychological resilience may be associated with enhancement of the orbitofrontal network in the elderly with MCI.

P676 / #732


RISK FACTORS FOR THE DEVELOPMENT OF DEMENTIA WITH LEWY BODIES

Lecture Title:

D. Tsuang1, S. Payne2, J. Shofer2, A. Shutes-David2, G. Li3 1VA Puget Sound, Geriatric Research Education And Clinical Center, Seattle, United States of America, 2VA Puget Sound, Grecc, Seattle, United States of America, 3University of Washington, Psychiatry And Behavioral Sciences, Seattle, United States of America

Aims: This study sought to evaluate the potential risk factors for the development of probable dementia with Lewy bodies (pDLB) versus probable AD (pAD) in NACC participants with baseline mild cognitive impairment (MCI). Methods: Participants in the NACC who were diagnosed with naMCI or amnestic MCI (aMCI) were followed longitudinally until the development of pDLB (n=95) or pAD;( n=1537). To identify predictors of pDLB (versus pAD), we performed logistic regression with baseline MCI subtype as primary predictor along with a set of characeteristics such as gender, age at pDLB/pAD, UPDRS, NPI, neuropsychological battery and APOE E4. Results: Although most participants (i.e., 78% of participants with naMCI and 96% of participants with aMCI) developed pAD, participants who developed pDLB were significantly more likely than participants who developed pAD have baseline nMCI (35% vs. 8%), to be male (81% vs. 48%), experience a younger age of onset (77 vs. 80 years,; p<.003 for all comparisons). These associations were maintained in a multivariate logistic regression model such that the likelihood of pDLB declined after age 80. Development of pDLB was also associated with higher UPDRS scores, lower executive functioning, and night behavioral disturbances (p<.0001) at baseline.

OR (95% CI) p

Male vs. female 4.6 (2.7, 7.9) <.0001

naMCI vs. aMCI 7.5 (4.5, 12) <.0001

Age at diagnosis, 85 vs. 75 0.61 (0.42, 0.88) .024

Conclusions: naMCI, Gender, age, and baseline symptoms may serve as risk factors for the development of pDLB, particularly when considered together. However, most participants with MCI who present to memory disorder centers eventually develop pAD.

P677 / #978


A PATIENT WITH PROGRESSIVE NON-FLUENT APHASIA WITH LONG-TERM PSYCHIATRIC SYMPTOMS

Lecture Title:

Ö. Türkoğlu, B.D. Ulug, Y. Ayhan Hacettepe University Faculty of Medicine, Psychiatry, Ankara, Turkey

Aims: To report a patient whose been followed in psychiatric clinic with late-onset long lasting recurrent depression with a final diagnosis of primary progressive aphasia non-fluent variant. Methods: We will present long term symptoms,signs,detailed neuropsychological profile,treatments,MRI findings of the patient. Results: A 64-year-old right handed woman presented with depressive complaints 15 years prior.She was followed up in a psychiatric clinic with recurrent depressive episodes and prescribed with different antidepressants including paroxetine, duloxetine and intermittent clonazepam for her anxiety symptoms. Seven years after the onset of depressive symptoms,she started to complain about her cognition.Her chief cognitive complaint was trouble remembering words. These complaints became apparent, interfering with daily functioning within 4 years.Her first neuropsychological evaluation revealed moderate impairment in executive functions,complex attention. Her speech became effortful and non-fluent which was interrupted by lengthy pauses within and between utterances,with paraphasic errors.Neuropsychological examination a year later revealed severe impairment in executive functions,visuospatial skills,complex attention; moderate impairment in attention; mild impairment in memory. Brain MRI displayed mild atrophy more prominent on left fronto-insular cortex with no hippocampal atrophy which led to the diagnosis of PPA non-fluent variant.Family history was significant for her deceased grandfather having Alzheimer’s dementia diagnosis. Conclusions: Neuropsychiatric syndromes such as depression,agitation,apathy may accompany PPAs. However data on the neuropsychiatric profile of presymptomatic sporadic PNFA patients are relatively scarce. In our case,depression might precede the language symptoms as well as it might present as a comorbid condition. It is important for physicians to reconsider their diagnosis when cognitive symptoms ensue during the course of psychiatric illness.

P678 / #1334

Topic: Theme K: Psychiatric Symptoms in Neurodegenerative Diseases / K5. Genetics, Epidemiology

APATHY OCCURS IN MORE THAN HALF OF NON-DEMENTED PARKINSON’S DISEASE PATIENTS

Lecture Title:

S. De Waele1,2, P. Cras1,3, D. Crosiers1,4 1Institute Born-Bunge, University Of Antwerp, Edegem, Belgium, 2University Hospital of Antwerp, Neurology, Edegem, Belgium, 3University Hospital Antwerp, Department Of Neurology, Edegem, Belgium, 4University Hospital of Antwerp, Department Of Neurology, Edegem, Belgium

Aims: The objective of our study was to determine the prevalence of apathy in a cohort representative of the real-life Parkinson’s disease (PD) population. Methods: We recruited PD patients through our outpatient clinic. We included patients of all ages, disease duration and Hoehn-and-Yahr stages. Patients suffering from cognitive decline and/or depression were excluded. Patients were asked to complete the Lille Apathy Rating Scale (LARS). Presence of depression was verified using the Beck Depression Inventory, possible cognitive decline was screened for using the Montreal Cognitive Assessment Scale. Results: To date we enrolled 44 PD patients in our study. 75 percent of patients were male. Mean age was 68.84 (±10.91) years. We found that 56.8 percent of patients were apathetic. Of all 44 patients, 25 percent were mildly apathetic (LARS scores 21 to -17), 18.2 percent were moderately apathetic (-16 to -10) and 13.6 percent were severely apathetic (-9 and above). Conclusions: In previous studies, apathy was reported to occur in around 20 to 25 percent of non-demented PD patients. Other studies employed different methods ranging from one item questionnaires to specific apathy scales. However, when employing a much more sensitive scale such as the LARS, apathy was identified in nearly 60 percent of our study sample. Actual prevalence may be three times higher than previously reported. Apathy is significantly associated with worse motor manifestations and increased caregiver burden. Its high prevalence calls for increased screening in everyday clinical practice to identify those affected and to guide future therapeutic interventions.

P679 / #1535

Topic: Theme K: Psychiatric Symptoms in Neurodegenerative Diseases / K5. Genetics, Epidemiology

ALLELIC VARIANT -521C>T (RS1800955) OF PROMOTER REGION OF GENE DRD4 AS A GENETIC PREDICTOR OF DEVELOPING ANTIPSYCHOTIC-INDUCED EXTRAPYRAMIDAL SYNDROME

Lecture Title:

E. Vaiman, N. Shnayder, R. Nasyrova V. M. Bekhterev National Medical Research Center of Psychiatry and Neurology, Department Of Personalized Psychiatry And Neurology, St Petersburg, Russian Federation

Aims: Study of association of antipsychotic-induced extrapyramidal syndrome (EPS) with variant -521CT (rs1800955) of promoter region of gene DRD4 encoding dopamine receptor D4. Methods: We tested 65 patients (53 men - 81.54%, 12 women - 18.46%) with a diagnosis of F20 to determine the severity of EPS on the background of antipsychotic (AP) monotherapy on the scales BARS, SAS, ESRS. The molecular genetic study of carriage of SNV rs1800995 of DRD4 gene promoter was carried out. The first group of observations -22 patients (33.8%), who received typical AP. The second group of observations - 43 patients (66.2%), who received atypical AP. Testing was carried out in dynamics: 1 visit - before AP therapy; 2 visit - 8 weeks after the start of therapy. Results: In the analysis of the data, including calculation of odds ratios, there were no statistically significant associations between carriage of allelic variants of the studied SNV rs1800955 of DRD4 gene promoter and the probability of development of AP-induced EPS. Conclusions: The results of the pilot pharmacogenetic study indicate that the carrier of allelic variants -521CT (rs1800955) of promoter region of gene DRD4 encoding dopamine receptor D4 is not associated with the development of AP-induced EPS in the study population, which may be due to the peculiarities of gene drift and sample size.

P680 / #336

Topic: Theme K: Psychiatric Symptoms in Neurodegenerative Diseases / K7. Animal Models

HUC-MSCS AMELIORATED CUMS-INDUCED DEPRESSION BY MODULATING COMPLEMENT C3 SIGNALING-MEDIATED MICROGLIAL POLARIZATION DURING ASTROCYTE-MICROGLIA CROSSTALK

Lecture Title:

J. Li, M. Wang the First Hospital of Hebei Medical University, Neurology, Shijiazhuang, China

Aims: Background: Major depressive disorder (MDD) has been shown to be related to immune inflammation and the complement system. Previous studies have suggested that human umbilical cord mesenchymal stem cells (hUC-MSCs) play an important role in inflammatory diseases. Methods: hUC-MSCs were administered into chronic unpredictable mild stress model (CUMS) mice through the tail vein once a week for 4 weeks. After the administration of hUC-MSCs, the depression-like and anxiety-like phenotypes, neuronal histopathology, synaptic-related protein expression and inflammatory index of the mice were assessed. Microglial M1/M2 polarization and the expression of C3a in astrocytes and C3aR in microglia was detected by immunofluorescence co-localization. Then, CUMS mice were injected with a C3aR antagonist, and the expression of C3a and C3aR and microglial polarization were observed. Results: Based on the sucrose preference and tail suspension tests, hUC-MSCs ameliorated the depression-like behaviors of CUMS mice. Additionally, the anxiety-like behaviors of CUMS mice in the open-field and plus-maze tests were improved after the administration of hUC-MSCs. hUC-MSCs altered microglia polarization by alleviating complement C3a-C3aR signaling activation, which decreased pro-inflammatory factor levels and increased anti-inflammatory factor levels, alleviating neuronal damage and synaptic deficits. Conclusions: hUC-MSCs have therapeutic effects on anxiety-like and depressive-like phenotypes caused by CUMS. They can alter the polarization of microglia by inhibiting C3a-C3aR signaling to reduce neuroinflammation.

P681 / #538


AGGRESSION AS A VERY EARLY MARKER OF ALZHEIMER'S DISEASE: A NEW MOUSE BEHAVIORAL TEST

Lecture Title:

I. Popova1, A. Osypov2, K. Mukhina3 1Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences, Laboratory Of System Organization Of Neurons, Pushchino, Russian Federation, 2Institute of Higher Nervous Activity and Neurophysiology, Russian Academy of Sciences, Laboratory Of Cellular Neurobiology Of Learning, Moscow, Russian Federation, 3Moscow State University, Faculty Of Biotechnology, Moscow, Russian Federation

Aims: Behavioral tests for revealing early neuropsychiatric symptom of AD as aggressiveness are poorly developed on animal models. We have developed a new "Fights" behavioral test for mice with AD model to detect aggressive interaction within a group of mice previously socially isolated. Methods: Mice underwent a neurosurgery for a guide cannula implantation and a social isolation for a period of one month in order to level the social hierarchy. After this period, animals received single i.c.v. injections of 1μl vehicle (control group) or 1μl of Aβ1-42 (Aβ group). To test for aggressiveness, all mice from each experimental group were placed together in a round chamber (diameter 50 cm) for 1 hour. The latent time of the first attack, the number of episodes of fights, their average duration, the number of participants and attackers were analyzed. Experiments were carried out at 18:00–21:00. The RealTimer (Open Science, Russia) were used for video registration and subsequent analysis. Results: A new “Fights» test were performed following 1 week of i.c.v. drug administration. The test revealed a significant increase in aggression in Aβ mice. The most significant changes affected the level of motivation of aggression – the latent period of the first attacks, the number of fights, the total time of fights and number of initiators. Conclusions: Experiments have shown that the "Fights" test can reliably reveal an increase of the level of aggression in mice with Aβ-model of Alzheimer's disease on the very early stage of pathology (1 week after Aβ1-42 injection).

P682 / #579


EFFECTS OF ENVIRONMENTAL ENRICHMENT ON MOOD AND HIPPOCAMPAL NEUROPLASTICITY IN THE YAC128 MOUSE MODEL OF HUNTINGTON'S DISEASE

Lecture Title:

P. S. Brocardo, E. Plácido, P. Gomes Welter, A.L. Severo Rodrigues Federal University of Santa Catarina, Neuroscience Graduate Program, Florianopolis, Brazil

Aims: Huntington’s disease (HD) is a genetic neurodegenerative disorder characterized by motor, neuropsychiatric, and cognitive deficits. The objective of this study was to determine if environmental enrichment (EE) exposure has positive effects on depressive-like behavior and hippocampal neuroplasticity in the YAC128 HD mice. Methods: Male and female wild-type (WT) and YAC128 mice (n= 8–10) were exposed to EE for 2 months (2-4 months) or to standard housing conditions. At the end of this period, depressive-like behavior was assessed using the tail suspension test and the splash test. Twenty-four-hour after behavioral testing animals were perfused with 0.9% NaCl followed by 4% paraformaldehyde. Serial brain coronal sections were obtained on a vibratome at 30 μm thickness. Cell proliferation in the dentate gyrus was assessed with the cell cycle markers Ki-67 and PCNA, and neuronal differentiation with the Doublecortin (DCX). The total number of immunoreactive cells was counted and statistical differences were analyzed with two-way ANOVA. Further, dendritic arborization was evaluated by Sholl analysis with DCX labeled neurons and statistical differences were assessed by repeated-measures ANOVA. Results: Exposure to EE for 2 months prevented the occurrence of depressive-like behavior in YAC128 mice. EE increased cell proliferation and dendritic arborization in WT mice. Neuronal differentiation was also increased by EE exposure in WT and YAC128 mice when compared to their control groups housed in standard housing conditions. Conclusions: Our results indicate that exposure to EE is able to modulate mood and hippocampal neuroplasticity in WT and YAC128 mice.

P683 / #1141

Topic: Theme L: Patient Care and Support / L1.b. Dementia and Cognitive Dysfunction: Mobile applications, social networks

DEMENTIA SCREENING USING AN APPLICATION PROGRAM WITH SIMPLE QUESTIONS AND SYMPTOMS

Lecture Title:

Y. Daté1,2, H. Tabuchi3, M. Mimura3, J. Nakahara2, D. Ito2 1Keio University School of MedicineFaculty of Nursing and Medical Care, Graduate School Of Health Management, Fujisawa City, Kanagawa, Japan, 2Keio University School of Medicine, Neurology, Shinjuku City, Tokyo, Japan, 3Keio University School of Medicine, Neuropsychiatry, Shinjuku City, Japan

Aims: As the population of patients with cognitive decline grows, physicians and caregivers need brief screening tools. Methods: “Attended With” (AW) and “Head-Turning Sign” (HTS) factors and participants’ replies to following questions were recorded: “Do you feel you’re in trouble in your daily life ?”, [no consciousness (C-) or consciousness+ (C+)], “Could you tell me about your daily pleasure or pastime?” [no pleasure (P-) or pleasure + (P+)], “What are notable current/recent news/topics?” [no news (N-) or news+ (N+)]. We enrolled 162 consecutive cases. Results: The sensitivity and specificity of each question were calculated, and the population attributable risk percent % (PAR%) of (AW and HTS+), (C- and P-), (C- and N-), (P- and N-), respectively, were calculated. As we have already reported, AW had high sensitivity, 87.4, 95.8% (CN vs aMCI + AD, CN + aMCI vs AD) but the sensitivity of HTS was only 46.4, 57.7%, and HTS showed high specificity, 100.0, 71.8%. C- had high sensitivity, 80.6, 87.5%, whereas P- and N- had high specificity, both 83.9% in CN vs aMCI + AD, 88.1% and 75.9% in CN + aMCI vs AD, respectively. In combination analysis, the combination of (C- and N-) is as powerful as (AW and HTS+) in screening AD. Conclusions: Then we have opened a website (https://neucop-q.site/) with which a general physician or a caregiver can easily fill in the symptoms and patients’ replies to three questions and then have the sensitivity and specificity concerning whether the patient in front of him or her has any cognitive impairment.

P684 / #1086

Topic: Theme L: Patient Care and Support / L1.b. Dementia and Cognitive Dysfunction: Mobile applications, social networks

CLINICAL APPLICATION OF THE EXPERIMENTAL ADL TEST FOR PEOPLE WITH COGNITIVE IMPAIRMENT USING IOT TECHNOLOGY

Lecture Title:

H.-W. Lee1, Y.-H. Lim1, Y. Baek1, S.J. Kang2 1Kyungpook National University Chilgok Hospital, Department Of Neurology, Daegu, Korea, Republic of, 2College of IT Engineering, Kyungpook National University, School Of Electronics Engineering, Daegu, Korea, Republic of

Aims: We employed a hospital-based Internet of Things (IoT) platform to validate the role of real-time activities of daily living (ADL) measurement as a digital biomarker for cognitive impairment in an hospital setting. Methods: All participants were assessed with structured intensive neuropsychological tests. The results of ADL tasks were categorized into success or fail. The total number of successful task and the average success proportion of each group was calculated. Time to complete the total tasks and total number of prompting were also measured. Results: The patients with dementia, with MCI, and cognitively normal older adults performed ADL tasks in a hospital setting. Significant differences in the average success rate of the tasks were found among dementia, MCI, and normal, respectively. Dementia group showed the lowest success proportion (49.3%) compared with MCI group (78.3%) and normal group (97.4%). Correlation between classical ADL scales and the number of completed ADL tasks was statistically significant. In particular, instrumental ADL (I-ADL) had stronger relationship with the number of completed ADL tasks than Barthel’s ADL (B-ADL). Dementia group required more time and coaching to accomplish the tasks when compared to MCI and normal groups. Conclusions: This study demonstrated that there is a clear relationship between the performance of experimental ADL tasks and the severity of cognitive impairment. The evaluation of ADLs involving the IoTs platform in an ecological setting allows accurate assessment and quantification of the patient’s functional level. Given the significant association with I-ADL, this evaluation may prove useful in detecting and diagnosing early stage of dementia.

P685 / #1600

Topic: Theme L: Patient Care and Support / L1.c. Dementia and Cognitive Dysfunction: Cognitive training

IMPROVING COGNITIVE PERFORMANCE OF OLDER ADULTS WITH MILD COGNITIVE IMPAIRMENT USING A SERIOUS GAME PLATFORM

Lecture Title:

H.-T. Jung1, J.-F. Daneault2, B. Ready3, B. Louis4, H. Lee5, B. Kim5, S. Lee1 1University of Massachusetts Amherst, College Of Information And Computer Sciences, Amherst, United States of America, 2Rutgers University, Department Of Rehabilitation And Movement Sciences, Newark, United States of America, 3University of Massachusetts Amherst, Department Of Psychological And Brain Sciences, Amherst, United States of America, 4Université de Montréal, Montreal Heart Institute & Department Of Medicine, Montreal, Canada, 5Woorisoft, Clinic, Daegu, Korea, Republic of

Aims: Mild Cognitive Impairment (MCI) is characterized by cognitive decline greater than expected for normal aging. Recently, serious games have become a potential tool to train and improve cognitive function among older adults with MCI. Serious games are video games designed for education and enhancing patients' health-related conditions. Existing games improve cognition in persons with MCI but do not offer a user-friendly interface that supports self-administration, hampering widespread implementation in homes and clinical settings. In this work, we demonstrate that older adults with MCI can self-administer Neuro-World, a serious game platform for cognitive training with a user-friendly touchscreen interface, and this leads to improved cognitive function. Methods: We conducted a pilot cohort study in eight older adults with MCI. Subjects self-administered cognitive training using Neuro-World for 30 minutes per day, twice a week, for six weeks. Cognitive function was assessed using all sub-scores and the total score of the Mini-Mental State Examination (MMSE) and Digit Forward/Backward Span (DFS and DBS, respectively). Additionally, the Quality of Life in Alzheimer's Disease (QoL-AD) was completed to identify any potential impacts of the intervention on QoL. Results: Participants showed statistically significant improvement in their overall cognitive function (1.38 ± 0.52 improvement in total MMSE), working memory (1.13 ± 0.99 improvement in DFS), and QoL (3.38 ± 3.11 improvement in QoL-AD). Conclusions: Neuro-World has great potential to be self-administered by older adults with MCI to improve their cognitive abilities and QoL, which warrants a large-scale randomized controlled trial.

P686 / #801

Topic: Theme L: Patient Care and Support / L1.e. Dementia and Cognitive Dysfunction: Support devices & monitoring

IMPROVING MEDICATION ADHERENCE IN ISOLATED ALZHEIMER’S DISEASE PATIENTS USING “AUTOMATED TELEPHONE REMINDER”

Lecture Title:

J.Y. Song, J.-W. Jang, J.H. Jhoo, G.H. Byeon, Y. Kim Kangwon National University Hospital, Neurology, Chuncheon, Korea, Republic of

Aims: Medication adherence is essential for effective treatment. Alzheimer's disease (AD) patients have difficulty taking medication because of their memory impairments. We aimed to investigate if there is an effect of automated telephone reminder service on improving medication adherence and reduction of the decline of cognitive function in isolated AD patients. Methods: This study was a single-center, randomized clinical trial conducted from March 2019 to February 2020 at the Hongcheon Dementia Center in Korea. We enrolled AD patients who lived alone or lived with a cognitively impaired spouse. We provided an automated telephone reminder service for taking medication to the intervention group for six months. The control group was provided with general guidelines for taking the medication every month. The participants underwent neuropsychological assessment at the beginning and the end of the study. Statistical significance was tested using Wilcoxon rank sum test. Results: Thirty participants were randomly allocated and 29 participants were analyzed. The mean age was 79.6 (SD 6.0) years, 79.3% were female. There was no significant difference in medication adherence between the two groups. However, a subgroup analysis with whom showed the answering rate above 70% showed a significant difference in the medication adherence between the two groups (intervention: 94.6 % (SD 3.8); control: 88.2 % (SD 9.0), p=0.0478). There was no significant difference in the change of cognitive function between the two groups. Conclusions: If compliance is good, telephone reminders might be effective in improving medication adherence. It is necessary to develop a reminder tool that can improve compliance.

P687 / #878

Topic: Theme L: Patient Care and Support / L1.f. Dementia and Cognitive Dysfunction: Quality of life

A NEW CAR-FREE LIFE: THE NEED FOR DEMENTIA-PATIENTS TO RETURN THEIR DRIVER'S LICENSES

Lecture Title:

M. Kanai, Y. Hirata, Y. Shimazaki, S. Yokota, E. Morita, M. Osaki, B. Mihara Mihara Memorial Hospital, Neurology, Isesaki, Japan

Aims: In Japan, people aged 75 or above who wish to have their driver’s license renewed will have to submit a medical certificate to determine whether they full in the first category during a preliminary inspection. An investigation of the situation is required if a subject belonging to the first category require medical approval. Methods: The subjects were 70 patients who visited our center for dementia-related diseases between June 2019 and May 2020, and needed to renew their driver's license. The patients were subject to neuropsychological tests, electroencephalography, and brain imaging (magnetic resonance imaging and single-photon emission computed tomography). Results: Dementia was diagnosed in 32 individuals, Alzheimer’s disease in 22, vascular dementia in 5, mixed dementia in 2, frontotemporal lobar degeneration in 2, and dementia with Lewy bodies in 1. Thirty patients had mild cognitive impairment (MCI), of whom seven revisited 6 months later, and two progressed to Alzheimer’s disease. Some patients diagnosed with dementia responded positively to the recommendation of voluntarily returning their driver’s licenses. However, some patients wished to retain their driver’s licenses despite the diagnosis of dementia. Conclusions: It is necessary to take measures to provide support to people with dementia, so that they do not experience a deterioration in their quality of life after they stop driving. Particularly, patients diagnosed with MCI need early consideration, and they should return their driver’s licenses. It is important for such patients to start preparing for a car-free life before the onset of cognitive decline.

P688 / #471


EGO INTEGRITY, GEROTRANSCENDENCE, AND RELATED FACTORS IN ELDERLY INDIVIDUALS: A POPULATION-BASED CROSS-SECTIONAL STUDY CONDUCTED IN TOKYO, JAPAN

Lecture Title:

H. Kida1, H. Niimura1, R. Shikimoto1, K. Suzuki1, Y. Miyasaka1, M. Takayama2, M. Mimura1 1Keio University School of Medicine, Department Of Neuropsychiatry, Tokyo, Japan, 2Keio University, Faculty Of Science And Technology, Yokohama, Japan

Aims: Japan is facing several issues pertaining to having a super-aged society. Elderly individuals require psychological support to cope with aging. E.H. and J.M. Erikson suggested that ego integrity and gerotranscendence are developmental stages of old and oldest-old age (Erikson’s 8th and 9th stages). This study aimed to understand how elderly individuals can acquire these two stages. Further, it aimed to identify related factors that could help them achieve psychological well-being. Methods: Questionnaire surveys and face-to-face interviews were conducted with older adults (aged 65 to 84 years) living in Tokyo. We assessed their developmental stages (ego integrity and gerotranscendence) using scales that assess Erikson’s 8th and 9th stage of development. Additionally, we assessed their quality of life (EQ5D), activities of daily living (ADL) (Barthel Index), instrumental ADL (TMIG-IC), sleep quality (PSQI), physical activity (GPAQ), grip strength, walking speed, depressive symptoms (GDS), and cognitive function (MMSE). Multiple regression analyses were performed to examine the relationships among variables after controlling for age, sex, and education. Results: The final sample comprised 1,099 older adults [596 females; mean (SD) age: 72.9 (5.2) years]. The 8th-stage score was negatively correlated with higher ADL and positively correlated with grip strength. The 9th-stage score was negatively correlated with higher ADL. Conclusions: This study identified factors associated with the developmental stages of elderly individuals. The negative relationship between Erikson’s two stages and ADL may suggest that positive acceptance of one’s physical deterioration and achievement of the two developmental stages may improve elderly individuals’ psychological well-being.

P689 / #899


EFFECT OF PLASMA EXCHANGE WITH ALBUMIN REPLACEMENT IN ALZHEIMER’S TREATMENT ON PATIENT-REPORTED HEALTH-RELATED QUALITY OF LIFE (HRQOL)

Lecture Title:

R. Lipton1, M. Boada2, O. López3, W. Stewart4, L. Podger5, M.C. Runken6, A. Davis6, L. Núñez7, A. Páez7, D. Serrano8 1Albert Einstein Colloge of Medicine, Department Of Neurology, New York, United States of America, 2Institut Català de Neurociències Aplicades - Universitat Internacional de Catalunya, Research Center And Memory Clinic, Barcelona, Spain, 3University of Pittsburgh School of Medicine, Department Of Neurology And Psychiatry, Pittsburgh, United States of America, 4Medcurio, N/a, Oakland, United States of America, 5Pharmerit (an OPEN Health company), Patient Centered Outcomes, London, United Kingdom, 6Grifols SSNA, Medical Affairs, Research Triangle Park, United States of America, 7Grifols, Alzheimer's Research Group, Sant Cugat del Vallès, Barcelona, Spain, 8Pharmerit (an OPEN Health company), N/a, Chapel Hill, United States of America

Aims: Alzheimer’s Management By Albumin Replacement (AMBAR) is a multicenter, randomized, blinded and placebo-controlled, Phase 2b/3 clinical trial, previously published elsewhere (Boada et al., Alzheimer’s Dement 2020). HRQoL was a secondary endpoint, measured by Quality of Life–Alzheimer’s Disease (QoL-AD). As a prelude to cost-effectiveness analyses, the impact of albumin replacement on HRQoL of people with Alzheimer’s Disease (AD) was assessed. Methods: The QoL-AD analysis population was the full analysis set (FAS). Change from baseline to final visit in QoL-AD total scores for placebo and pooled treatment arms was assessed using ANCOVA. Baseline QoL-AD, age, and Mini Mental State Examination (MMSE) were included as covariates. For this analysis, patients were classified as performing in a mild-spectrum AD subgroup according to both Clinical Dementia Rating Scale–Sum of Boxes (CDR-SB) and MMSE thresholds. Results: The FAS included 63 participants in the placebo arm and 160 in the pooled treatment arm; mild-spectrum subgroup included 27 in the placebo arm and 78 in the pooled treatment arm. Within FAS, change from baseline marginal mean was 0.25 for placebo and 1.5 for treatment (difference: 1.26, p=0.0361). Within the mild-spectrum subgroup, change from baseline marginal mean was -0.57 for placebo and 1.75 for pooled treatment (difference: 2.3, p=0.0136); treatment arm stratified proportions achieving MID were 39% and 20% for pooled treatment and placebo, respectively. Conclusions: In comparison with placebo, plasma exchange followed by albumin replacement therapy is associated with improvements in QoL-AD scores for the FAS, with numerically larger improvements for the mild-spectrum subset.

P690 / #673


DYSPHAGIA IN ALZHEIMER'S DISEASE: A SYSTEMATIC REVIEW

Lecture Title:

A. Mira1,2, R. Gonçalves2, I. Tello Rodrigues1,3 1Escola Superior de Saúde do Alcoitão, Speech And Language Therapy, Alcabideche, Portugal, 2Centro Hospitalar e Universitário do Algarve - Centro de Medicina de Reabilitação do Sul - Algarve - Portugal, Speech And Language Therapy, São Brás de Alportel, Portugal, 3Center for Innovative Care and Health Technology (CiTechCare) – IPL, Speech And Language Therapy, Leiria, Portugal

Aims: Dysphagia has been described in recent literature as one of the most relevant co-morbilities of Alzheimer's disease. However, few studies on the specific characteristics and progression of dysphagia exist. The aim of this systematic review was to identify the specific characteristics, progression, and prevalence of dysphagia in Alzheimer's disease. Methods: The search was conducted by two independent researchers in March2020 of published literature from the last 10 years in the PubMed (Medline), EBSCO, ScienceDirect, and BASE databases. A critical appraisal and an evidence level analysis using Joanna Briggs Institute Critical Appraisal and the Effective Public Health Practice Project's (EPHPP) "Quality Assessment Tool for Quantitative Studies" tools were conducted. Results: From the search, appraisal, and analysis of the results, 26 studies were eligible for full review. The extracted data suggest that cortical changes occur in the neural swallowing network long before clinical symptoms appear. Dysphagia progresses, as does Alzheimer's disease, in a continuum and, its severity depends of individual variability. There are a few studies regarding therapeutic approaches for minimizing symptoms and complications. Dysphagia is unequivocally linked to Alzheimer's disease co-morbidities since severe dysphagia leads to malnutrition, dehydration, pneumonia, increases in falls or ulcers, cognitive and behavioral decline, and even death. Regarding prevalence, the researchers found no studies were published in the last 10 years. Conclusions: Dysphagia is a complex and important co-morbility in Alzheimer's disease and has an impact on quality of life and access to and costs of healthcare.

P691 / #865

Topic: Theme L: Patient Care and Support / L1.g. Dementia and Cognitive Dysfunction: Functional foods

DIETARY HABITS INFLUENCE THE SHORT AND LONG-TERM FRAILTY RISK. RESULTS FROM THE INVECE.AB COHORT STUDY.

Lecture Title:

M.C. Mimmi1, A. Davin2, A. Ceretti2,3, O. Pansarasa1, T.E. Poloni3,4, C. Cereda1, A. Guaita2,3 1IRCCS Mondino Foundation, Genomic And Post-genomic Center, Pavia, Italy, 2Golgi-Cenci Foundation, Laboratory Of Neurobiology And Neurogenetic, Abbiategrasso, Italy, 3Golgi-Cenci Foundation, Department Of Neurology And Neuropathology, Abbiategrasso, Italy, 4ASP Golgi-Redaelli Geriatric Hospital, Department Of Rehabilitation, Abbiategrasso, Italy

Aims: Frailty is a geriatric syndrome which manifests as the reduction of functional reserves. It exposes the individual to an increased risk of adverse events, disability and mortality. This work focuses on diet as an adjustable factor that can affect the incidence of frailty. Methods: The subjects were selected from the "InveCe.Ab" study, which in 2010 recruited 1321 people born between 1935-39. The cohort was re-evaluated in 2014 and in 2018. Frailty was measured via a cumulative "Frailty Index" (FI), identifying three classes of subjects: Fit, PreFrail and Frail. Explanatory dietary variables captured the current and past intake of fresh fruit, fresh vegetables and fish. A Habitual Mediterranean Diet score (HMedDiet) was obtained by combining the consumption of those three classes of food. Results: Frailty incidence resulted inversely associated with the HMedDiet score, indicating an especially strong response to the consumption of fruit and vegetables. The latter was significantly lower in Frail subjects than in the rest of the population. Our data demonstrate that each unit increment of HMedDiet score consistently reduces the four-year risk of developing frailty (OR 0.68; 95%CI: 0.52-0.88) The same effect is confirmed also in a longer term perspective (eight years) (OR 0.80; 95%CI: 0.66-0.98). Conclusions: Our data show that maintaining a diet with frequent intake of fruit, vegetables and fish is associated with a lower occurrence of frailty in an elderly population. The adherence to a healthy nutrition style, started at least in the sixties reduces the risk of developing frailty in the late seventies.

P692 / #524

Topic: Theme L: Patient Care and Support / L1.h. Dementia and Cognitive Dysfunction: Behavioral & psychiatric symptoms

BASELINE FINDINGS OF THE FILIPINO MULTICOMPONENT INTERVENTION TO MAINTAIN COGNITIVE PERFORMANCE AMONG HIGH-RISK POPULATIONS (FINOMAIN) STUDY

Lecture Title:

M.C. Del Moral1, J. Dominguez1, R.M. Del Moral2 1St. Luke's Medical Center, Institute For Neurosciences, Quezon City, Philippines, 2University of the Philippines Manila, College Of Pharmacy, Manila, Philippines

Aims: (1) To discuss a multicomponent intervention targeting Filipino populations at high-risk for cognitive decline; (2) To compare FINOMAIN with other existing multicomponent interventions for the prevention of dementia in the elderly; (3) To communicate the results of an on-going multicomponent intervention in a community setting. Methods: FINOMAIN is an on-going community study which integrates nutrition counselling, standard vascular care, and physical exercise in the form of a ballroom dance protocol called INDAK (Improving Neurocognition through Dance and Kinesthetics). The multicomponent intervention will last for 48 weeks. Nutrition counselling and standard vascular care are given every third month, and INDAK is given twice a week for 60 minutes each session. FINOMAIN is a two-arm, single-blind, cluster randomized controlled trial of a multicomponent intervention to prevent cognitive decline among at-risk elderly. A total sample of 71 participants (intervention=38, control=33) from 8 community clusters in Quezon City, Metro Manila joined the study. Results: The comprehensive assessment included (1) cognitive measures (including CDR, ADAS-Cog, AD 8, MoCA-Philippines, MMSE-Philippines, Verbal Fluency, Digit Symbol Substitution Test, Trail Making Test A and B), (2) behavioural and functional measures (including NPI, GDS, EQVAS Health-Related QoL, Lawton's IADL, DAD, Time Up and Go, BBS), (3) anthropometric measures, and (4) medical and laboratory measures, including blood amyloid (MDS-AD). The intervention and control groups are comparable in all baseline measures (p's>0.05). Compliance to INDAK protocol was 97.4% (>/=80%). Conclusions: FINOMAIN is a much-needed study to confirm the efficacy of multicomponent interventions in preventing cognitive decline among at-risk elderly in a community setting.

P693 / #1305

Topic: Theme L: Patient Care and Support / L1.j. Dementia and Cognitive Dysfunction: Other

A COMPARISON STUDY BETWEEN LIPID-BASED NANOPARTICLES AND POLYMERIC-BASED NANOPARTICLES IN NOSE-TO-BRAIN DELIVERY OF MELOXICAM AS ANTI ALZHEIMER’S DISEASE

Lecture Title:

H. Akel, I. Csoka Faculty of Pharmacy - University of Szeged, Institute Of Pharmaceutical Technology And Regulatory Affairs, szeged - Csongrad, Hungary

Aims: Nanoparticles were classified as preferable nose-to-brain carriers due to the desirable properties they show, especially in neurodegenerative disorders. The presented work aimed to prepare SLNs and PBNs with the help of the QbD concept to do an in vitro comparison between them both. Methods: QbD approach was followed to prepare the optimized formulations by Double-emulsion solvent evaporation. The EE& DL, Morphological study, and in-vitro permeation and release tests have been done to compare the two nanocarriers, then the mucoadhesion test was performed on the optimal candidate investigating the effect of chitosan coating. Results: SLNs showed higher EE& DL than PBNs 72.71&2.23, 62.29&2.41, respectively. Encapsulation of meloxicam in both nanoformulations demonstrated better in-vitro release and permeation behavior in comparison with the plain drug with the superiority of SLNs. SLNs showed good mucoadhesiveness properties enhanced by chitosan coating. Conclusions: Formulation of meloxicam, a model of anti-AD medicines, in SLNs for the nose to brain delivery, showed promote release and permeation properties over PBNs, with good mucoadhesion characteristics enhanced by chitosan coating.

P694 / #1056


ADVANTAGE OF THE ELECTRONIC MEDICAL RECORD IN ALZHEIMER'S DISEASE AND RELATED DISORDERS: DIVERSITY STUDIES

Lecture Title:

S. Clos, T. Chabrashvili SUNY Upstate Medical University College of Medicine, Neurology And Neuroscience, Syracuse, United States of America

Aims: The electronic medical record (EMR) is an excellent resource to evaluate trends in patient data. A broad array of neurodegenerative disorders are seen amongst the diverse population of Central New York (CNY). We focused on Alzheimer’s disease and related disorders in this study. Our goal is to evaluate trends and disparities in neurodegenerative diagnoses amongst patients in the area. Methods: The EMR EPIC platform was used to create a cohort of patients using keywords Alzheimer’s disease (AD), vascular dementia (VaD), and mild cognitive impairment (MCI). Microsoft Excel was used to compile data and generate graphs. Results: In our cohort of 8,401 patients, 4,768 (56.8%) had AD, 2,149 (25.6%) VaD, and 2,815 (33.5%) MCI. Among AD, 89.2% were Caucasian, 5.7% African American, 4.5% unknown, 0.5% American Indian, 0.6% Asian, and <0.1% Native Hawaiian/Other Pacific Islander. Among VaD, 87.0% were Caucasian, 8.5% African American, 4.2% unknown, 0.7% American Indian, 0.5% Asian, and 0% Native Hawaiian/Other Pacific Islander. For MCI, 85.7% were Caucasian, 9.4% African American, 4.6% unknown, 0.5% American Indian, 0.7% Asian, and 0% Native Hawaiian/Other Pacific Islander. The CNY population represents 84.2% Caucasian, 7.6% African American, 1.4% other race, 2.9% 2+ races, 0.6% American Indian, 3.4% Asian, and <0.1% Native Hawaiian/Other Pacific Islander. Conclusions: EMRs can assist studies of neurodegenerative disorders in diverse patient populations to improve healthcare and to address health equity research as part of the portfolio in the focus area.

P695 / #637


VISUAL AND OCULAR DETERMINANTS OF CENTRAL VISUAL PATHWAY INTEGRITY AND VISUAL ASSOCIATION AREAS

Lecture Title:

D. Garzone1,2, M. Mauschitz1,2, R. Finger2, M. Reuter3,4,5, M. Breteler1,6, A. Aziz1,7 1German Center for Neurodegenerative Diseases (DZNE), Population Health Sciences, Bonn, Germany, 2University of Bonn , Faculty of Medicine, Department Of Ophthalmology, Bonn, Germany, 3Harvard Medical School, Department Of Radiology, Boston, United States of America, 4German center for neurodegenerative diseases (DZNE), Image Analysis, Bonn, Germany, 5Massachusetts General Hospital, A.a. Martinos Center For Biomedical Imaging, Boston, United States of America, 6Faculty of Medicine, University of Bonn, Institute For Medical Biometry, Informatics And Epidemiology (imbie), Bonn, Germany, 7Faculty of Medicine, University of Bonn, Department Of Neurology, Bonn, Germany

Aims: Decreased visual acuity (VA) can induce trans-neuronal degeneration (TND) along the entire visual pathway (VP) and is a risk factor for cognitive impairment. However, the relation between VA and higher order (sub)cortical visual integration centers is less well known. We therefore aimed to investigate the interrelations among VA, retinal layer structure and volumes of structures involved in central visual processing. Methods: We used baseline data from participants of the Rhineland Study who had both optic coherence tomography (OCT) and MRI data available, excluding participants with a known brain disease (N=2596). VA and retinal layer thickness were assessed using autorefraction and spectral domain-OCT, respectively. Volumes and cortical thickness were derived from 3 Tesla MRI T1-weighted brain scans using FreeSurfer. Multivariate regression and structured equation models were used to assess the associations between VA, retinal structure and brain MRI measurements. Results: VA was significantly associated with retinal thickness as well as hippocampal volume, but not with volumes or thickness of visual association areas. The relation between retinal thickness and volumes/thickness of visual association areas was mediated by the pericalcarine cortex (for cortical thickness, indirect effect=0.29, 95%CI [0.12-0.48], total effect 0.62, 95% CI [0.29-0.97], with the effect size significantly increasing at lower levels of VA (moderated mediation index 0.19, 95% CI [0.04-0.44]). Conclusions: We found that VA is associated with the structural integrity of the retina and other central regions, suggesting that TND due to decreased VA may partly account for the previously reported association between visual impairment and cognitive decline.

P696 / #1644


HEALTH SYSTEM PREPAREDNESS FOR ALZHEIMER’S DISEASE TREATMENT: VETERANS HEALTH ADMINISTRATION

Lecture Title:

D. Miller1,2, Q. Shao2, M. Palnati2, S. Mcdannold2, Q. Zhang3, A. Tahami3, W. Xia4,5, N. Palacios1, L. Moo5 1University of Massachusetts at Lowell, Center For Population Health, Lowell, United States of America, 2Bedford VA Healthcare System, Center For Healthcare Organization And Implementation Research, Bedford, United States of America, 3Easai Inc., Neurology Business Group, Woodcliff Lake, United States of America, 4Boston University School of Medicine, Pharmacology And Experimental Therapeutics, Boston, United States of America, 5Bedford VA Healthcare System, Grecc, Bedford, United States of America

Aims: Disease-modifying treatments (DMT) in Alzheimer disease (AD), a debilitating neurodegenerative disorder, are extensively investigated. We aimed to evaluate healthcare system capacity and readiness in the Department of Veterans Affairs (VA) for AD DMT when approved in US. Methods: Electronic health records from VA outpatient, inpatient, and extended care from 2019 were analyzed to identify patients with clinical diagnosis of mild cognitive impairment (MCI), AD, or AD related dementia (ADRD) based on ICD-10-CM codes, and to determine the distribution of dementia diagnoses, specialist follow-up, and use of biomarker testing leading up to the diagnosis. Results: We identified a total of 230,092 veterans with MCI (90,835), AD (35,197), or ADRD (135,532) with most receiving a diagnosis of dementia not-otherwise-specified (NOS) (101,766). We found that 43.9% of individuals visited a specialist in the year (34.0% neurologist, 13.5% geriatrician, 52.1% geriatric psychiatrist). Only 1.2% received a brain Positron Emission Tomography (PET) scan, although 94.7% of them subsequently visited a specialist. There was no clinical cerebrospinal fluid (CSF) testing for β-amyloid or tau proteins. Conclusions: Less than a half of veterans who received clinical diagnosis of MCI, AD, or ADRD in 2019 had a specialist visit during the year. PET imaging and CSF testing were virtually unused among these patients to confirm the status of AD biomarkers. The apparent overuse of NOS dementia coding may be due, in part, to lack of certainty in AD diagnosis. Substantial healthcare resources are needed to advance decision making for AD DMT.

P697 / #1210


A COMPARATIVE ANALYSIS OF ALZHEIMER’S DISEASE PRESENTATION IN HAWAII'S ASIAN, CAUCASIAN, AND NATIVE HAWAIIAN POPULATIONS

Lecture Title:

Z. Miyamoto1,2, E.L. Luo2,3, C. Llantero2,4, C. Yee2,5, C. Nakamura2,6, P. Borman7,8, J. Viereck7,8, K. Liow7,8 1University of Hawaiʻi at Mānoa, Department Of Molecular Biosciences And Bioengineering, Honolulu, United States of America, 2Hawaii Pacific Neuroscience, Clinical Research, Honolulu, United States of America, 3University of Hawaiʻi at Mānoa, School Of Life Sciences, Honolulu, United States of America, 4University of Notre Dame, College Of Science, Notre Dame, United States of America, 5Boston University, Department Of Psychological And Brain Sciences, Boston, United States of America, 6Chaminade University, School Of Nursing, Honolulu, United States of America, 7Hawaii Pacific Neuroscience, Alzheimer’s Research Unit & Memory Disorders Center, Honolulu, United States of America, 8John A. Burns School of Medicine, University of Hawaii, Medicine, Honolulu, United States of America

Aims: This study investigated how Alzheimer’s disease (AD) presents differently, in terms of diagnostic characteristics, severity of cognitive impairment, and intensity of behavioral disturbance, across different racial populations on Oahu, Hawai’i. Methods: Data was extracted from Hawai’i Pacific Neuroscience’s late onset AD patient records compiled over the past decade. Racial affiliation, diagnostic age and body mass index (BMI), Mini Mental State Examination (MMSE) score, Geriatric Depression Scale (GDS) reportation, and usage of psychotropic medication were noted. GDS analysis only included patients with an MMSE score of 15 or higher for valid reports. Results: Average diagnosis age in all groups fell within a year of each other, with the youngest group being Native Hawaiians, at 79.8 years. Native Hawaiians averaged a diagnosis BMI in the overweight range, with 25.93. Asians posted the lowest BMI at 23.54. A significant difference (p<0.05) in average MMSE score was identified, with Caucasians at 22.4, Asians at 21.1, and Native Hawaiians at 18.2. Proportion of patients on behavioral medication within each race was highest in Native Hawaiians, at 47% (n = 17), and lowest in Asians, at 27% (n = 38). GDS analysis suggested no significant difference in depressive severity, but was limited due to MMSE cutoffs. Conclusions: These findings suggest that Native Hawaiians present with greater cognitive impairment severity and behavioral disturbance intensity than Asians or Caucasians diagnosed around the same age. BMI and socioeconomic status could factor into the observed disparity. Outreach would benefit Native Hawaiians in slowing the disease progression.

P698 / #1257


NEUROCOGNITIVE KEYPAD ASSESSMENT TOOL (NKAT) PILOT STUDY

Lecture Title:

A. Ng1, A. Sritharan2, A. Thananjeyan2, M. Thomas3, S. Garga4, S. Marathe5, J. Stocker6 1Fiona Stanley Hospital, -, Perth, Australia, 2Dubbo Base Hospital, -, Dubbo, Australia, 3Western Sydney University, -, Sydney, Australia, 4Westmead Hospital, -, Sydney, Australia, 5Royal Prince Alfred Hospital, -, Sydney, Australia, 6St George Hospital, -, Sydney, Australia

Aims: Traditional pen-and-paper cognitive assessments are time-consuming and maybe frustrating for individuals with motor symptoms. It also requires trained clinicians to meet the patient in a face-to-face setting which would also introduce an assessor bias. The NKAT aims to rapidly identify and accurately measure a quantifiable cognitive aggregate in individuals with early cognitive impairment. Methods: NKAT measures the duration the user takes to input numbers for three tasks. The tool logs the number of errors made and the time spent on each task. The initial input is then used to correct for individual keystroke pace. Stable inpatients with no prior diagnosis of neurological or cognitive issues were recruited. Trained assessors completed an initial screening of the patients with the 4AT to rule out delirium. The AD8 was then used to screen for any cognitive complaints; DASS21 for depression, anxiety and stress levels; and the 3MS and MoCA for baseline cognitive function. The NKAT was then completed independently by the patient on a touchscreen or tablet. Results: 34 in-patients were recruited (29 normals, 5 reported cognitive deficits). There was no difference between the groups for gender, ADL, iADL, depression, anxiety, stress levels, MMSE or MoCA. Individuals with cognitive complaints were older (51.90±7.86vs37.90±9.55,p=0.004), had more cognitive complaints (1.60±0.89vs0,p=0.011) and took longer for the corrected NKAT repeat shuffle task (17.80±7.40vs7.42±4.24seconds,p=0.026). Conclusions: The pilot study on inpatients within a hospital environment demonstrates its utility in distinguishing individuals with early cognitive deficits. Monitoring keystroke speed can also correct for individuals with tremors which may affect traditional task taking abilities.

P699 / #502


A PILOT STUDY TO DISTINGUISH BETWEEN HEALTHY PARTICIPANT AND PATIENT WITH ALZHEIMER’S DISEASE AND PARKINSON’S DISEASE BASED ON VOCAL ANALYSIS: FOLLOW-UP REPORT

Lecture Title:

Y. Omiya1, T. Takano1, M. Higuchi2, M. Nakamura2, S. Shinohara2, S. Mitsuyoshi2, N. Sanjo3, H. Terashi4, S. Tokuno2 1PST Inc., Research And Development, Yokohama, Japan, 2The University of Tokyo, Bioengineering, Tokyo, Japan, 3Tokyo Medical and Dental University, Neuropathology, Tokyo, Japan, 4Tokyo Medical University, Neurology, Tokyo, Japan

Aims: To extend healthy life expectancy, early detection and preventive treatment of disease are important. To estimate health conditions using voice is effective for diagnosis support. In this paper, we examined a pilot study to distinguish between healthy (HE) participant and patient with Alzheimer’s disease (AD) and Parkinson’s disease (PD) based on vocal analysis. Compared to our previous report at the ADPD2020 conference, we applied to another data and improved algorithm to alleviate impact of recording condition differences. Methods: We collected voices from HE (n = 23), AD (n = 22), and PD (n = 20) participants in the consulting rooms of two hospitals. The participants uttered by repeating “pa-ta-ka” for about 5 seconds. Then, we considered influenced by motor functional decline on voice by disease and calculated six features related to intensity from each recorded voice. Here, to reduce the impact of recording condition differences, we adapted the normalization to voice and using the median value as a threshold for detection of the peak position. Based on those six features, we examined the possibility of distinguishing between HE, AD, and PD participants by the Tukey-Kramer method. Results: In case the Tukey-Kramer method was conducted in three groups of HE, AD, and PD, significant differences (t<0.05) at least one or more combinations were observed in five of six features. Conclusions: Consequently, the features relative to voice intensity has the potential to work well in classifying participants with HE, AD, and PD, suggesting the utility of the classification algorithm for detecting diseases based on speech.

P700 / #351


USE OF MULTISENSORY INTEGRATION TASKS IN NEUROCOGNITIVE TRAINING WITH OLDER ADULTS

Lecture Title:

J. Pinto1, A. Dores1,2, B. Peixoto3,4, B.B. Vieira De Melo5, F. Barbosa1, A. Geraldo1 1Laboratory of Neuropsychophysiology, Faculty Of Psychology And Education Sciences, University Of Porto, Porto, Portugal, 2School of Health, Polytechnic Institute Of Porto, Porto, Portugal, 3CESPU, University Institute of Health Sciences, Department Of Social And Behaviour Sciences, Paredes, Portugal, 4NeuroGen, Center For Health Technology And Services Research (cintesis), Porto, Portugal, 5Psychosocial Rehabilitation Laboratory, Center for Rehabilitation Research, School Of Health Of The Polytechnic Of Porto, Porto, Portugal

Aims: Objectives: A growing body of literature has investigated how to maximize the efficacy of neurocognitive training in older adults, namely by improving the ecological validity of the interventions. Multisensory Integration (MI) has the potential to improve the ecological validity of neurocognitive interventions due to the need of adaptation to the naturalistic environment and the relationship between neurocognitive and sensory functioning. The authors aimed to determine the characteristics of MI tasks, their usefulness as functional and cognitive assessment, and as an intervention in older adults with and without neurocognitive impairment. Methods: The authors conducted a literature search using quality assessment of narrative reviews criteria, following previous suggestions by Baethge and colleagues (2019) and Green and colleagues (2016). Results: Results included tasks of detection, discrimination, sensory illusion, temporal judgment, and sensory conflict. Overall, results were inconsistent regarding the enhancement of MI with age, but pointed that older adults may have an expanded window of integration, needing more time to integrate two stimuli. The association of neurocognitive impairment and spatial aspects of gait through the visual-somatosensory integration was also reported in older adults with mild cognitive impairment and dementia. Conclusions: This review highlights the potential advantages of combining MI with neurocognitive training in the rehabilitation of older adults with and without neurocognitive impairment, according to the neurocognitive model of Luria and provides recommendations to guide future studies.

P701 / #1741


SUCCESSFUL IMPLEMENTATION OF A NATION-WIDE PROGRAMME FOR DEMENTIA PREVENTION IN LUXEMBOURG: PROGRAMME DEMENCE PREVENTION (PDP)

Lecture Title:

V. Schröder1,2, A. Kaysen1, J. Fritz3, L. Pauly1,2, A. Skrozic1,2, D. Mcintyre3, S. Köhler4, A. Leist5, T. Hartmann6,7, E. Kalbe8, R. Dodel9, R. Krüger1,2,3 1University of Luxembourg, Luxembourg Centre For Systems Biomedicine, Belvaux, Luxembourg, 2Centre Hospitalier de Luxembourg, Department Of Neurology, Luxembourg, Luxembourg, 3Luxembourg Institute of Health, Transversal Translational Medicine, Strassen, Luxembourg, 4Maastricht University, Alzheimer Centrum Limburg, School For Mental Health And Neuroscience, Maastricht, Netherlands, 5University of Luxembourg, Institute For Research On Socio-economic Inequality, Esch-sur-Alzette, Luxembourg, 6Universitat Des Saarlandes, Neurology, Homburg, Germany, 7Saarland University, Deutsches Institut Für Demenz Prävention (didp), Medical Faculty, Homburg, Germany, 8Medical Faculty and University Clinic Cologne, Department Of Medical Psychology | Neuropsychology And Gender Studies, Köln, Germany, 9University Duisburg-Essen, Chair Of Geriatric Medicine, Essen, Germany

Aims: Implement the first nation-wide integrated care programme for at risk individuals to prevent dementia by the means of personalised lifestyle interventions. Methods: Participants with subjective cognitive decline and mild cognitive impairment are referred to the programme by their treating physician and undergo an extensive cognitive evaluation and dementia risk factor assessment. Based on these results, individualised lifestyle interventions (such as cognitive training, dietary advice, physical/social activities) are offered via different national partners. In order to evaluate the impact on their cognitive status and the presence of risk factors, participants are followed-up annually. Results: We established a national prevention network and successfully reached our target population by means of efficient communication strategies, stakeholder engagement and outreach events. Furthermore, the network fostered interdisciplinary communication between healthcare providers, which was supported by promising results of a survey conducted among the referring doctors. The collection of a large amount of data regarding cognitive status, risk factors and adherence to suggested lifestyle interventions allows gaining insights into the effectiveness of personalised lifestyle interventions in reducing dementia risk factors. To ensure continuity of the programme despite the adversities caused by the COVID-19 pandemic and to further raise awareness for dementia prevention, the implementation of eHealth technologies (e.g. smartphone application), is planned. Conclusions: We provide evidence for the feasibility of the implementation of a nation-wide dementia prevention programme offering personalised lifestyle interventions, which is easily transferrable to other countries. Future results from this programme may allow for integrating innovative prevention interventions into the regular healthcare system.

P702 / #640


PRESENTATION OF THREE FRENCH SCREENING TOOL TO DETECT THE LEXICO-SEMANTIC BREAKDOWN IN MILD COGNITIVE IMPAIRMENT AND ALZHEIMER’S DISEASE

Lecture Title:

I. Simoes Loureiro1, M. Taverne1, I. Rouleau2, L. Lefebvre1, S. Joubert3, M.-J. Chasles4, E. Delage5, J. Cole4 1UMONS, Cognitive Psychology And Neuropsychogy, Mons, Belgium, 2Département de psychologie, Université du Québec à Montréal, & CRIUGM, Centre De Recherche Du Centre Hospitalier De L'université De Montréal (crchum), Montreal, Canada, 3Université de Montréal & CRIUGM, Département De Psychologie, Montreal, Canada, 4Université du Québec à Montréal, & CRIUGM, Département De Psychologie, Montreal, Canada, 5Université de Montréal & CRIUGM, Département De Psychologie Et Criugm, Montreal, Canada

Aims: Lexico-semantic difficulties are amongst the first symptoms of Alzheimer’s disease (AD). Lexical impairment manifests itself through word retrieval difficulties while semantic disorder impairs general knowledge. This profile already occurs in mild cognitive impairment (MCI), constituting a predictive feature of conversion in AD. The aim of this study is to present three tools to rapidly screen lexical and semantic disorder. Methods: 97 participants allocated in 3 groups contributed to this Belgian-Quebec study: one group of MCI (N=35, MOCA (French-Canadian) =24.18+/-2.3 ; MMSE (Belgian) (N=7) =26.71+/-1.38), one group of AD (N=13 Belgian, MMSE=24.7+/-2.52) and one control group (N=49, MOCA (French-Canadian)=27.79+-1.9 ; MMSE (Belgian) (N=15)=29.07+/-.79). Three tools were administered : 1) the mini Semantic Knowledge Questionnaire (SKQ) composed of 12 multiple choice questions interrogating semantic properties of objects ; 2) The Short Test of naming for Alzheimer’s Disease (STN-AD) comprising 11 pictures in black and white to name and 3) the mini-Montreal Semantic Memory Protocol (mini-MSMP) comprising 26 questions interrogating functional or categorical features of objects. Results: The ANOVA indicated that all the three tools (mini-SKQ : F(2 ;94)=8.440 ; p<.001 ; STN-AD : F(2 ;94)=17.760 ; p<.001; mini-MSMP: F(2 ;94)=7.273 ; p=.001) allowed for differentiating our groups. Post hoc tests showed that MCI and AD performed poorer than the control group for all three tools (p<.05). Conclusions: The mini-SQK, the STN-AD and the mini-MSMP are three original tools used to evaluate the early lexical and semantic alteration. Their rapid administration made them instrument of choice for an early detection of MCI and AD.

P703 / #1251


USEING OF PSYCHOEDUCATIONAL PROGRAMS IN THE STRUCTURE OF MEDICAL AND PSYCHOLOGICAL SUPPORT OF FAMILIES WITH ALZHEIMER'S DISEASE PATIENTS.

Lecture Title:

I. Strelnikova Kharkiv National Medical University, Psychiatry, Narcology, Medical Psychology And Social Work, Kharkive, Ukraine

Aims: The goal. Investigation of the psychological state of the families of patients with Alzheimer's disease and the use of psychoeducational programs in the structure of medical and psychological support of the family membersr of these patients. Methods: . A comprehensive clinical and psychopathological examination of 64 families of patients with Alzheimer's disease. The main group - 42 families who took part in the program of medical and psychological support with the use of psychoeducation. The control group - 22 families who did not receive medical and psychological support. Used the Spielberger-Khanin anxiety scale, K. Thomas' method of "Determining ways to resolve conflicts" in the adaptation of NV Grishina (according to D. Ya. Raigorodsky, 2002), the questionnaire of nervous and mental stress (TA Nemchin, 1984), anxiety and depression scale M. Hamilton, 1967, methods of studying styles of stress-coping behavior "Coping Strategy" (adapted by TA Kryukova, 2002), lectures of psychoeducation. Results: . The main group had a positive effect with the reduction of anxiety and depressive disorders (59.3% of families), reducing frustration to the nominal disease (73.8%), family conflict (69, 7%), harmonization of family relations (54.76%). Among the control group, 31.81% of families were diagnosed with increased manifestations of psychopathological symptoms, in 59.09% of families the clinical picture of anxiety and depressive disorders remained without significant changes, in 40.90% of families the deterioration of family relationships was admited. Conclusions: . The obtained results allowed to substantiate the importency of medical and psychological support of families with Alzheimer's disease patients and the use of psycho-educational programs.

P704 / #1636


MODELING THE ASSOCIATION OF SELECTED MEDICATIONS ON THE OCCURRENCE OF POSSIBLE ALZHEIMER’S DISEASE: ATORVASTATIN AND LISINOPRIL OUTPERFORM SIMVASTATIN

Lecture Title:

Y. Wang1, M. Li1,2, L. Kazis2,3, W. Xia4,5 1Bentley University, Mathematical Sciences, Waltham, United States of America, 2Bedford VA Healthcare System, Center For Healthcare Organization And Implementation Research, Bedford, United States of America, 3Boston University School of Public Health, Health Law, Policy And Management, Boston, United States of America, 4Boston University School of Medicine, Pharmacology And Experimental Therapeutics, Boston, United States of America, 5Bedford VA Healthcare System, Geriatric Research Education Clinical Center, Bedford, United States of America

Aims: Associations of long term use of angiotensin converting enzyme inhibitor (ACEI) and statin medications are explored on the risk of occurrence of Alzheimer’s disease (AD). Methods: From an existing Department of Veterans Affairs (VA) corporate data warehouse (VA-CDW), we extracted 73,605 dementia with possible AD patients and 660,314 non-AD patients from 1999 to 2018, based on ≥2 ICD-9/10-CM codes from separate clinical visits, and analyzed 97,163 patients prescribed the target medications of Lisinopril, Atorvastatin, or Simvastatin using a non-randomized cohort design. We utilized logistic regression and Cox proportional hazard models with or without propensity score weighting to calculate the odds ratio (OR) and hazard ratios (HR) of taking individual medications on the occurrence of dementia with possible AD. Results: Compared to those medicated with Simvastatin, Atorvastatin was associated with a lower OR (0.23, 95% confidence intervals (CI) (0.21, 0.25)) and HR (0.73, (0.66, 0.81)); propensity score adjusted (PSA) OR (0.39) and PSA HR (0.70) on occurrence of AD. Lisinopril significantly reduced the OR (0.71, (0.68, 0.75)) and HR (0.91, (0.87, 0.96)); PSA OR (0.74) and PSA HR (0.93) on occurrence of AD. A significantly lower risk of developing AD was found among patients taking Atorvastatin for <2 years. Lisinopril, compared to Simvastatin, demonstrated a significant reduction on the occurrence of AD among those who initiated medication younger than 65-years of age and had taken the medication for < 5 years. Conclusions: Atorvastatin and Lisinopril outperform the commonly used Simvastatin, associated with lower occurrence of possible AD onset.

P705 / #567

Topic: Theme L: Patient Care and Support / L2.b. Movement Disorders: Mobile applications, Telemedicine, Digital Health, E-trials & social networks

AN ADAPTIVE STEP DETECTION ALGORITHM FOR WAIST-WORN WEARABLE DEVICES: A FEASIBILITY STUDY IN OLDER ADULTS

Lecture Title:

L. Peraza1, K. Kinnunen1, R. Joules1, R. Wolz1,2 1IXICO Plc, R&d, London, United Kingdom, 2Imperial College London, Department Of Computing, London, United Kingdom

Aims: Accurate step detection from wearable accelerometers is a key analysis requirement in clinical gait studies. Moreover, the large amount of data recorded prohibits the use of manual labelling, calling for reliable algorithms that automate or semi-automate the labelling process. In this investigation, we designed and propose a step detection algorithm based on adaptive thresholds and tested its performance with accelerometry data acquired from an older-adult group. Methods: We reanalysed a publicly available database recorded from an orthogeriatric population diagnosed with osteoporosis (N=17, mean age=75.5) [Keppler 2019, PLoS One]. Steps were labelled by video annotation and participants performed four types of parcours: Hallway and lobby walks (back and forth), Figure 1. Our proposed algorithm detects peaks from a heavily processed signal and adapts the step-detection threshold according to the amplitude to account for participant differences. Algorithm’s initial parameters were tuned using accelerometry data from one healthy participant (age=38) and the performance independently tested with the orthogeriatric accelerometry database. Results: The total number of steps detected showed a high agreement with the ground truth (mean absolute error = 0.030, SD=0.043, Figure 1). Step detection precision was >0.976 and sensitivity was >0.964 for the four parcours (Figure 2).

Figure 1

Figure 2

Conclusions: Our proposed step detection algorithm showed to be highly accurate detecting steps from older adults. Gait analysis in older adults is particularly challenging due to the possible presence of movement disorders such as gait shuffling. More research work will be necessary to test our algorithm in patients diagnosed with gait disorders.

P706 / #1207

Topic: Theme L: Patient Care and Support / L2.b. Movement Disorders: Mobile applications, Telemedicine, Digital Health, E-trials & social networks

ADDED VALUE OF WEARABLE SENSORS FOR FALL RISK ASSESSMENT IN AN OLDER ADULT GROUP

Lecture Title:

L. Peraza1, K. Kinnunen1, R. Joules1, R. Wolz1,2 1IXICO Plc, Clinical Science, London, United Kingdom, 2Imperial College London, Department Of Computing, London, United Kingdom

Aims: Falls represent the leading cause of death-related injuries in older adults. Free-living activity recorded from wearables may offer a method to characterise people at risk. In this investigation we re-analyse a publicly available database to evaluate the added value of wearables in identifying fall risk. Methods: Recruited healthy older adults were categorised as fallers if self-reported two or more falls (N=31) and as controls if only one or none (N=37) over a year period [Weiss2013, Neurorehabil-Neural Repair]. Participants had cognitive and in-clinic gait assessments and showed no evidence of gait or cognitive disorders. Participants were given a wearable sensor positioned near L5 and wore it for three days. From the accelerometry data, gait bouts were identified using an in-house step-detection algorithm, divided in 20-second segments, and 74 features extracted per segment (Figure 1). Using LOO cross-validation with random forests, we classified fallers vs non-fallers by including 1) free-living features, 2) in-clinic gait/cognitive scores, and 3) in-clinic and free-living measures combined.

Results: The free-living features resulted in a 63% accuracy (50% sensitivity, 81% specificity). The in-clinic scores showed an accuracy of 72% (61% sensitivity, 81% specificity). When combining both feature sources, accuracy was 75% (61% sensitivity, 86% specificity). Prominent features are shown in Figure 2.

Conclusions: Free-living accelerometry helped to slightly improve fall prediction over clinical scores. These findings are in line with previous work published on the cohort analysed. Future work will focus on whether wearable assessments can provide additional benefits in Parkinson’s disease patients as opposed to the healthy participants studied here.

P707 / #506

Topic: Theme L: Patient Care and Support / L2.f. Movement Disorders: Quality of life

FAMILY SUPPORT AND LEGAL LIABILITY WHEN THE DEMENTIA PATIENTS CAUSE AN ACCIDENT IN JAPAN

Lecture Title:

R. Hatanaka SHOBI University, Faculty Of Policy Science, Saitama, Japan

Aims: In 2007, 91 years old man with dementia walked into railway negligently ,and died by train accident. Japan Railway(JR) sued his 83 years old wife and his son for paying damage making up transportation cost. His son lives apart from their house and son’s wife moves near to take care of them.His 83 years old wife is 1 level graded under nursing insurance . This presentation tries to focus on the topic about legal liability of family members having dementia patient in Japan. For example, whether the family has legal liability when dementia patient becomes assailant of traffic accidents was well discussed in current Japan. Methods: Method is case analysis. Researcher follow some legal cases about gurdians were asked civil liability. Results: This case showed us the issues“Does family have supervisory responsibility?” and“If they have, did they do their responsibility?” Final decision of this case was judged in March 2016 .Supreme court denied family’ s liability to pay dameges. Because it is difficult to expect his wife take care of him actually in the situation as his wife was frail and his son lived far from them . On the other hand, this case showed if family members can take care of him actually, they may have legal liability possibly. Conclusions: Comparing other past case in Japan, family tended to have legal liability when same case had been occurred. In current situation, Japanese courts have changed their attitude for family liability in aging society.

P708 / #407


PREDISTIM: A FRENCH MULTI-CENTRIC, MULTI-MODAL LONGITUDINAL PROSPECTIVE COHORT TO PREDICT STN DBS RESPONSE IN PARKINSON’S DISEASE ON QUALITY OF LIFE TO REFINE DBS INDICATION

Lecture Title:

A.-S. Rolland1, C. Moreau1, S. Thobois2, A. Eusebio3, E. Hainque4, T. Rouaud5, J.-L. Houeto6, S. Drapier7, D. Guehl8, D. Maltete9, C. Tranchant10, C. Giordana11, P. Krystkowiak12, L. Hopes13, C. Hubsch14, B. Jarraya15, F. Durif16, O. Rascol17, J.-C. Corvol4, D. Devos1 1Univ Lille, Inserm, CHU Lille, U1172,lille Neurosciences & Cognition- Team « Degenerative & Vascular Cognitive Disorders ", Lille, France, 2Hospices civils de Lyon, Neurology, Bron, France, 3Aix Marseille Universite, AP-HM, Hopital de la Timone, Neurology, Marseille, France, 4Hôpital Pitié-Salpêtrière, AP-HP, Faculté de Médecine de Sorbonne Université, UMR S 1127, Inserm U 1127, and CNRS UMR 7225, and Institut du Cerveau et de la Moëlle épinière, Neurology, Paris, France, 5• Clinique Neurologique, Hôpital Guillaume et René Laennec,, Neurology, Nantes cedex, France, 6Centre Expert Parkinson, CIC-INSERM 1402, CHU Poitiers, Neurlogy, Poitiers, France, 7CHU Pont Chaillou, Neurology, Rennes cedex, France, 8Institut des Maladies Neurodégénératives cliniques, CHU de Bordeaux, Explorations Fonctionnelles Du Système Nerveux, bordeaux, France, 9Rouen University Hospital and University of Rouen, France; INSERM U1239, Laboratory of Neuronal and Neuroendocrine Differentiation and Communication, Neurology, Mont-saint-aignan, France, 10Hôpitaux Universitaires de Strasbourg, Neurology, Strasbourg, France, 11• CHU Nice, Centre Expert Parkinson, Neurology, Nice, France, 12Amiens University Hospital, Neurosurgery, Amiens, France, 13Nancy University hospital, Neurology, Nancy, France, 14Fondation Ophtalmologique A de Rothschild, Neurology, Paris, France, 15Foch Hospital, Versailles Saint-Quentin-en-Yvelines University, Neurology, Suresnes, France, 16Université Clermont Auvergne, EA7280, Clermont-Ferrand University Hospital, Neurology, Clermont-ferrand, France, 17UMR 1214, Inserm/ups – Tonic, Toulouse Neuroimaging Center, Chu Purpan, Toulouse, France

Aims: Determine predictive factors of quality of life, 1 and 5y after Subthalamic Nucleus (STN) Deep Brain Stimulation (DBS) to refine DBS indication with multimodal parameters (clinical, radiological, biological, genetic). Methods: 17 french centers from the NS-Park network implemented a web-based solution to prospectively collect clinical information from PD patients’ candidates to DBS. Protocol includes 4 visits: inclusion(V0), time of surgery(Vc), 1y post-surgery(V1) and 5y post-surgery(V5). Clinical data include demography, risk factors, medical/surgical history, general impact, motor/non-motors symptoms, adverse event, stimulation parameters, treatments. In parallel, biological samples/MRIs were performed to create a biobank/brain-imaging bank. Inclusions ended in June 2019 and follow-up will continue up to 2024. Results: PREDISTIM cohort includes 647 advanced PD patients. Motor/non-motor data are collected into an eCRF. A central biobank ensures quality/conformity of all biological samples (DNA/plasma/serum/CSF). A central brain-imaging bank ensure the quality of all MRIs sequences. Statistical approaches with joint latent class analyses, linear mix models and machine learning will be used. Conclusions: PREDISTIM is the largest prospective multi-centric cohort of PD patients at the stage of severe motor fluctuations. It will allow 1) to define weighted/multimodal predictors of STN DBS response on quality of life at 5 years 2) to refine new DBS inclusion criteria with a decisional tree including new parameters and 3) to offer to a large database to assess surrogate biomarkers of disease progression from advanced stage to late stage with dementia and severe axial disorders in order to stratify PD population for future trials on advanced disorders contributing to the precise medicine.

P709 / #422


THE BURDEN OF PROGRESSIVE SUPRANUCLEAR PALSY ON PATIENTS AND CAREGIVERS BY PSP PHENOTYPE: A CROSS-SECTIONAL STUDY

Lecture Title:

A. Scowcroft1, A. Klein2, T. Gasalla3, A. Avbersek4, J. Wright5, J. Mellor5, A. Thompson6, D. Pillas1 1UCB Pharma, Global Real World Evidence, Slough, United Kingdom, 2UCB Pharma, Tau And Psp, Brussels, Belgium, 3UCB Pharma, Neurodegeneration Mission, Raleigh, United States of America, 4UCB Pharma, Tau Mission, Braine-l’Alleud, Belgium, 5Adelphi Real World, Rare Diseases, Bollington, United Kingdom, 6UCB Pharma, Early Evidence Strategy, Brussels, Belgium

Aims: Progressive supranuclear palsy (PSP) is a rare, relentlessly progressive, ultimately fatal neurodegenerative brain disease. This study aimed to assess the burden of PSP on patients, caregivers and healthcare systems by PSP phenotype according to the 2017 Movement Disorder Society diagnostic criteria. Methods: Between July and November 2018, data from the Adelphi PSP Disease Specific Programme, a cross-sectional study of neurologists and people living with PSP in the USA, France, Germany, Italy, Spain and the UK, were collected and reviewed. Results: Data from 892 people living with PSP were evaluated, mean age was 70.2 years; 58% were male. PSP phenotype was reported for 242 (27%) of patients (PSP-Richardson’s syndrome: n=96, PSP-Parkinsonism: n=88, PSP-progressive gait freezing: n=28, PSP-corticobasal syndrome: n=12, PSP-predominant frontal presentation: n=9, PSP-speech/language disorder: n=9) and 650 patients had no reported phenotype. Most patients reported cognitive, motor, behavioural, and ocular symptoms; 47–100% of patients (across phenotypes) had moderate-to-severe disease at diagnosis. Post-diagnosis, the majority of patients (55–100%, across phenotypes) were provided with a visual and/or mobility aid, and/or required home modification to facilitate their needs. Patients required multiple types of healthcare professional for disease management (mean 3.0–4.4, across phenotypes), and the majority reported receiving care from at least one caregiver (mean 1.3–1.8, across phenotypes). Conclusions: There is a high burden on patients, caregivers, and healthcare systems across all PSP phenotypes. Although phenotypes manifest different symptoms and are associated with different diagnostic pathways, once diagnosed with PSP, patients typically receive similar care.

P710 / #1792


IMPACTS OF CLASSICAL MUSIC AND DANCING ON COGNITIVE FUNCTIONS IN PARKINSON'S DISEASE

Lecture Title:

S. Shokhimardonov, E. Nuriddinova, S. Kuzieva, S. Khudjanov Tashkent Medical Academy, Neurology, Tashkent, Uzbekistan

Aims: To study changes in the cognitive plan among patients with Parkinson's disease after conducting dance classes and listening to classical music based on rehabilitation therapy under the TMA program, evaluate the dynamics of changes in such cognitive functions, memory, speech, attention, etc. according to MMSE and MoCa rating scales. Methods: Patients were divided into two groups. The first group (n = 27) underwent therapy with rehabilitation therapy with TMA and the control group (n = 26) who received standardized treatment without this program. The lesson was held once a week for 1 hour for 12 weeks at the base of rehabilitation of neurological patients at the TMA clinic. Patients had an early stage of PD without cognitive impairment (Addenbrook score: DG = 92.3 ± 2.7, CG = 90.8 ± 3.9). Both groups took cognitive function tests on the first day of admission, as well as on days 21 and 48. Results: Comparison of the results showed that the cognitive functions of patients such as verbal training (= 28%), delay (= 28%), recall delay (= 20%), memory capacity for verbal material (= 12%) improved in relation to the control group. (p <0.05) Conclusions: Rehabilitation with the help of dance lessons and classical music have a clear benefit from psychological symptoms, preservation and optimization of cognitive function. Long-term studies are necessary to confirm the persistence of these effects.

P711 / #858


ARE ERRORS IN ADMINISTRATION OF TIME CRITICAL MEDICATION ASSOCIATED WITH INCREASED LENGTH OF STAY IN PARKINSON’S INPATIENTS?

Lecture Title:

A. Smith University of Exeter Medical School, Knowledge Spa, Truro, United Kingdom

Aims: The National Institute of Health and Care Excellence (NICE) advises doses of Parkinson’s medication to be given within 30 minutes either side of scheduled dose time to avoid harmful side effects. This research compares Royal Cornwall Hospital Trust’s (RCHT) performance against NICE guidelines and determines if doses received outside guidelines are associated with increased length of hospital stay (LoS). Methods:

A retrospective review of electronic health records between the range 01/06/19 – 31/08/19 was performed. Of 169 admissions screened, 57 were included in statistical analysis. Admissions were categorised as follows: Group 1 (n = 21) – patients with 0-1 doses received outside guidelines Group 2 (n = 36) – patients with 2+ doses received outside guidelines Statistical analysis was performed using Kolmogorov-Smirnov (KS) and Mann Whitney U (MWU) tests.

Results:

Doses analysed = 746 Doses within guideline = 407 (54.6%) Doses outside guideline = 339 (45.4%) Group 1 Mean LoS = 1.17 days Group 2 Mean LoS = 5.95 days KS test statistic (0.59) exceeded the critical value (0.18) indicating significant difference between distributions of LoS. MWU test showed significant difference in mean LoS between groups (P = < 0.001). Conclusions: 45.4% of Parkinson’s patients in RCHT were placed at risk of developing harmful side effects. Patients who received 2+ doses outside guidelines stayed 4.78 days longer, reducing patient wellbeing and increasing susceptibility to hospital-acquired infections. Future research should analyse reasons behind dose errors to inform policy change/educational strategies. Future research should adjust LoS for confounding factors to confirm if significant difference persists.

P712 / #1574

Topic: Theme L: Patient Care and Support / L2.g. Movement Disorders: Behavioral & psychiatric symptoms

THE RELATIONSHIP BETWEEN DYSARTHRIA AND COGNITION IN IDIOPATHIC PARKINSON’S DISEASE

Lecture Title:

K. Brown, K. Spencer University of Washington, Speech And Hearing Sciences, Seattle, United States of America

Aims: The aim of this study is to examine the relationship between speech impairment and global cognitive functioning in individuals with idiopathic Parkinson’s disease (PD). Converging evidence suggests an association between the presence of dysarthria and cognitive decline and that this relationship may be mediated by primary motor phenotype (i.e., tremor-dominant and non-tremor dominant). Methods: In Experiment 1 (pre-COVID19 restrictions), 9 participants with PD completed a test battery including Unified Parkinson’s Disease Rating Scale (UPDRS) ratings, a speech sample, and the Parkinson’s Disease Cognitive Rating Scale.Ten experienced listeners will rate naturalness and understandability of speech on a Visual Analog Scale as an initial determination of the relationship between speech and cognition. In Experiment 2 (during COVID19 restrictions), 192 individuals with PD were examined from the Parkinson’s Progression Marker Initiative (PPMI)—a longitudinal, controlled database of de novo PD. Speech ratings and primary motor presentation in early disease will be used as predictive factors of later cognitive decline and progression to dementia. Speech and primary motor profile were rated using the UPDRS; cognitive categorization was determined by PPMI investigators. Results: Preliminary analyses suggest an emerging relationship between speech and cognitive decline in PD that is not attributed to overall severity and may be influenced by PD motor subtype. Conclusions: Initial findings support the premise that a breakdown in shared neural channels results in changes to speech and cognitive performance, and that this relationship may differ between tremor-dominant and non-tremor phenotypes. Therapeutic implications will be discussed.

P713 / #236

Topic: Theme L: Patient Care and Support / L2.g. Movement Disorders: Behavioral & psychiatric symptoms

CORRELATION OF HOEHN AND YAHR STAGES AND MOCA-INA SCORE IN PARKINSON’S DISEASE PATIENTS IN MOHAMMAD HOESIN GENERAL HOSPITAL PALEMBANG

Lecture Title:

Y. Harun1, S. Marisdina2, A.K. Kurniawan2, S. Irfanuddin3 1MOHAMMAD HOESIN GENERAL HOSPITAL PALEMBANG, Neurology, Palembang, Indonesia, 2Dr. Mohammad Hoesin General Hospital, Neurology, Palembang, Indonesia, 3Medical Faculty of SriwijayaUniversity,Palembang, Physiology, Palembang, Indonesia

Aims: Objectives: To determine the correlation between Hoehn and Yahr stages as well as sociodemographic factors and MoCA-INA score Parkinson's Disease. Methods: Method: This study is a cross-sectional correlation test using primary and secondary data. A total of 38 subjects were obtained from 1 January 2019 - 31 January 2020 in the outpatient installation of Mohammad Hoesin General Hospital Palembang. Sociodemographic data, history of hypertension, history of type 2 diabetes mellitus, onset of motoric symptoms, and MoCA-INA score were taken for analysis Results: The majority of subjects were male (65.8%), 61-70 years old (42.1%), with high school education (47.4%). 17 patients (44.7%) had hypertension, and 7 patients (18.4%) had a history of type II DM. The mean age of onset for motoric symtomps was 59.94 ± 10.95 years. There was no correlation between Hoehn and Yahr stages and MoCA-INA score (r = -0,012; p = 0.942), as well as visuospatial / executive functions, naming, attention, language, abstraction, delayed recall, and orientation (r<0,4; p>0,05) in Parkinson's Disease patient. Weak correlation occurs between age and cognitive function (r = -0.357; p = 0.028), but there was no correlation between educational level, history of hypertension and diabetes mellitus type II, onset of motoric symptoms and MoCA-INA score (r<0,4; p>0,05) in Parkinson’s Disease patients. Conclusions: Conclusion: Hoehn and Yahr stages do not affect MoCA-INA score in Parkinson's Disease patient.

P714 / #310

Topic: Theme L: Patient Care and Support / L2.g. Movement Disorders: Fall prevention & patient protection

INVESTIGATION FOR WHEELCHAIR DEPENDENCE OF DEMENTIA PATIENTS BY THE GAIT ANALYSIS PERFORMED WITH MOFF BAND®

Lecture Title:

Y. Chiba1,2, A. Kumamoto1, N. Noguchi1, A. Yoshimi1,2, A. Suda1,2, A. Kase1 1Yokohama Maioka Hospital, Psychiatry, Yokohama-shi, Japan, 2Yokohama City University Graduate school of Medicine, Psychiatry, yokohama-shi, Japan

Aims: Patients with dementia suffer from motor dysfunction in the mid-later course of the disease. Falls should be prevented or treated safely by walking mobility aids such as a wheelchair. The gait analysis is helpful for evaluating the risk of falls, but it is difficult for patients with dementia to set many sensors in their bodies. Moff band®, Moff Co. Ltd, Tokyo, Japan, is an easy-to-use and commercially-available wearable smart device detecting 3-dimensional physical movements. Methods: We performed the Moff band's gait analysis for 34 patients with dementia who came to our hospital for cognitive evaluation or treatment for behavioral and psychological symptoms. We collected their demographic data, cognitive function, CT scan findings, medications, gait analysis parameters evaluated by Moff band, and followed them for six months. We compared their data between dementia patients with and without wheelchair dependence by t-test or Fisher exact test. We performed a false discovery rate correction by Benjamini & Hochberg method for multiple comparisons, and performed the logistic regression analysis for wheelchair dependence by the related gait analysis parameters. Results: Eleven patients got wheelchair dependence during six months. The score of the clinical dementia rating scale was significantly higher and the angle of hip extensor was significantly lower in the dementia patients with wheelchair dependence than those without. The odds ratio for wheelchair dependence of an angle of hip extensor was 1.32 (95% CI: 1.09~1.61). Conclusions: The angle of hip extensor measured by Moff band might indicate the necessity of a wheelchair for patients with dementia.

P715 / #808

Topic: Theme L: Patient Care and Support / L2.h. Movement Disorders: Other

PSYCHOMETRIC PROPERTIES OF THE CROATIAN VERSION OF THE NON-MOTOR SYMPTOMS SCALE FOR PARKINSON’S DISEASE

Lecture Title:

Z. Uzarevic1, A. Soldo Koruga2,3, I. Kampic2, Z. Popijac2, S. Butkovic Soldo2,3 1Faculty of Education, University Of Osijek, Osijek, Croatia, 2Clinic for Neurology, Clinical Hospital Centre Osijek, Osijek, Croatia, 3Faculty of Medicine, University Of Osijek, Osijek, Croatia

Aims: The aim was to evaluate the psychometric properties of the Croatian version of the Non-Motor Symptoms Scale (NMSS), a tool to assess non-motor symptoms (NMSs) in Parkinson’s disease (PD). Methods: A cross cultural adaptation of the NMSS into Croatian and a psychometric analysis of the translated version of the NMSS was carried out in PD patients. NMSS includes nine domains: cardiovascular, sleep/fatigue, mood/cognition, perceptual problems/hallucinations, attention/memory, gastrointestinal tract, urinary, sexual function and miscellaneous. The quality of data was descriptively analysed and Chronbach’s alpha coefficient assessed internal consistency. Pearson correlation were performed on the nine domains. The level of significance was set to p<0.05. Results: Seventy-six PD patients were assessed (mean age 68.4±9.6 years; 45% women; mean length of disease 8.7±4.6 years). Mean NMSS score was 110.68±58.76, where the more common NMSs in PD patients were related to the following domains: mood/cognition (22.86%), sleep/fatigue (19.74%), sexual function (13.65%) and urinary (11.58%). Chronbach’s alpha for the NMSS total score was 0.92 (range for domains: 0.48-0.96). Total NMSS score was highly correlated with each of the nine domains (Pearson correlation range: 0.45-0.82). Conclusions: The Croatian version of NMSS can be considered a comprehensive and helpful measure for NMSs in Croatian-speaking PD patients.

P716 / #1080


UTILIZATION OF THE ELECTRONIC MEDICAL RECORD: PARKINSONIAN COHORT

Lecture Title:

S. Clos, T. Chabrashvili SUNY Upstate Medical University College of Medicine, Neurology And Neuroscience, Syracuse, United States of America

Aims: The electronic medical record (EMR) is a useful tool in evaluating trends to improve diagnosis and treatment. Methods: The EMR EPIC platform was used to provide a cohort using keywords Parkinson’s disease (PD), parkinsonism, dementia with Lewy bodies (DLB), multiple system atrophy (MSA), corticobasal syndrome (CBS), and progressive supranuclear palsy (PSP). Microsoft Excel was utilized to compile data. Results: In our cohort of 3,235 patients, 2,905 (89.8%) had PD, 278 (8.6%) parkinsonism, 304 (9.4%) DLB, 45 (1.4%) MSA, 16 (0.5%) CBS, 71 (2.2%) PSP, and 187 (5.8%) PD with DLB. Average age of onset was 76.36 (PD), 57.57 (parkinsonism), 80.53 (DLB), 65.49 (MSA), 72.46 (CBS), and 76.48 (PSP). Male predominance occurred in PD (60.7%), parkinsonism (58.8%), and DLB (60.9%). Female predominance occurred in MSA (55.6%), CBS (53.8%), and PSP (52.2%). Marital status was reflected by 57.4% of PD, 43.1% parkinsonism, 60.9% DLB, 44.4% MSA, 76.9% CBS, and 75.4% PSP patients. Depression and anxiety were detected in 29.4% and 19.0% of PD, 54.7% and 34.7% of parkinsonism, 43.1% and 28.4% of DLB, 40.0% and 35.6% of MSA, 46.2% and 61.5% of CBS, and 42.0% and 18.8% of PSP patients, respectively. Psychosis and sleep disorders were detected in 14.4% and 21.0% of PD, 40.9% and 27.4% of parkinsonism, 48.5% and 22.4% of DLB, 35.6% and 22.2% of MSA, 15.4% and 53.9% of CBS, and 14.5% and 17.4% of PSP patients, respectively. Conclusions: The cohort selected from the EMR provides an adequately representative parkinsonian population. This data can be used as a platform for future studies.

P717 / #1124


ALZHEIMER'S DISEASE MORTALITY TRENDS IN TURKEY, 2009-2018

Lecture Title:

N. Doğan, İ. Doğan Afyonkarahisar Health Sciences University, Biostatistics And Medical Informatics, Afyonkarahisar, Turkey

Aims: Background: With the increase in the elderly population in the world, prevalence and mortality rates of Alzheimer's disease (AD), which is the most common cause of dementia, have been observed. This ecological study aims to analyze the temporal mortality trends of AD inTurkey. Methods: Method: Data on the Turkish population and the number of deaths due to AD were obtained from the Türkish Statistical Institute database (2009-2018). The age-standardized rates were calculated using the direct method, according to the proposed World standard population. The trend in AD mortality rates was tested for sex and age using joinpoint regression analysis. Results: Almost 96,000 AD deaths occurred in Turkey from 2009 to 2018. In general, the model with a single joinpoint was found to be the best significant model. Standardized AD mortality rates have increased by 13.3% each year from 2009 to 2015. Although it has decreased by 0.5% annually from 2015 to the end of the period, this decrease is not statistically significant. When evaluated by gender, age-standardized Alzheimer's mortality rates from 2009 to 2015 showed a significant increase of approximately 13% per year (both sex). Until 2015, significant increases were observed in all age groups in both women and men. Although there was a downward trend after 2015, none of these trends are significant. Conclusions: Conclusion: The findings of the studies may be important for the development of the health system related to AD. Further efforts are required to reduce mortality from AD in the Turkish population.

P718 / #633


STUDY ON THE WEIGHT LOSS IN PARKINSON'S DISEASE

Lecture Title:

H. Kujirai1, Y. Kanda2, K. Ishigaki3, E. Akazawa3, M. Kitazaki2, Y. Miyamoto2, M. Shibasawa4, T. Ishikawa4, Y. Hasegawa4, A. Fujita5, Y. Matsushima6, Y. Ono7, S. Orimo1 1Kanto Central Hospital, Neurology, Tokyo, Japan, 2Kanto Central Hospital, Nursing Department, Tokyo, Japan, 3Kanto Central Hospital, Nutrition Management Department, Tokyo, Japan, 4Kanto Central Hospital, Department Of Rehabilitation, Tokyo, Japan, 5Kanto Central Hospital, Pharmaceutical Department, Tokyo, Japan, 6Kanto Central Hospital, Clinical Laboratory, Tokyo, Japan, 7Tokyo Institute of Technology, School of Computing, Laboratory Of Human Communication And Co-creation System, Kanagawa, Japan

Aims: Around 50% of people with Parkinson's disease (PD) show weight loss, however the exact mechanism of weight loss remains to be clarified. In this study, we conducted study to clarify the relationship between weight loss and clinical characteristics together with laboratory findings in PD. Methods: Seventy-nine PD people (48 women, age 70.6 years, disease duration 7.6 years) were enrolled. Various indicators to evaluate PD symptoms, blood data, and body components were examined. We divided into “lean group: LG” with BMI of less than 18.5 and “non-lean group: NLG” with BMI of 18.5 or more. We compared various data between two groups. Results: The LG was 23 (29.1%) (21 women). There were no differences in age, disease duration, MDS-UPDRS III, PDQ39, and levodopa equivalent dose between two groups, but there were significantly more women and higher Hoehn-Yahr stage (LG 2.6/NLG 1.98) in the LG. For body components (LG/NLG), body water (25.3/32.1)L, protein (6.6/8.5)kg, mineral (2.4/3.0)kg, body fat (5.4/13.3)kg, muscle mass (32.3/41.1)kg were significantly decreased in the LG. The decrease in body fat (46%) was particularly large. Blood data (LG/NLG) showed no differences in blood glucose, total protrein, total cholesterol, and zinc. Albumin (4.1/4.25), transthyretin (22.1/27.3)mg/dL, triglyceride (78.5/106.6)mg/dL, cholinesterase (257.5/306.3)mg/dL, and Hb (12.5/13.4)g/dL were significantly decreased in the LG. Conclusions: In PD patients with around H-Y stage 2, 29.1% were thin, and most of them were women. In thin PD patients, albumin, transthyretin, triglyceride, cholinesterase, and Hb were decreased. All body components were decreased, and the decrease in fat amount was particularly remarkable.

P719 / #490


A NEW PARKINSON’S DISEASE TRIAL NETWORK: LESSONS LEARNED FROM THE GLOBAL ALZHEIMER’S PLATFORM FOUNDATION® TRIAL NETWORK

Lecture Title:

K. Smith1, G. Goldfeder1, J. Bork1, L. Zisko1, R. Mohs1, Z. Mari2, H. Fernandez3 1Global Alzheimer's Platform Foundation, Clinical Operations/ Trial Recruitment And Study Solutions, Washington D.C., United States of America, 2Cleveland Clinic Lou Ruvo Center, Parkinson Disease And Movement Disorders, Las Vegas, United States of America, 3Cleveland Clinic Lerner College of Medicine, Cleveland Clinic Center For Neurological Restoration, Cleveland, United States of America

Aims: To design a novel site network that addresses operational challenges in Parkinson’s disease (PD) clinical trials. Methods: The Global Alzheimer’s Platform Foundation® (GAP) is designed to improve the efficiency and speed of Alzheimer’s disease (AD) clinical trials. The network of 70+ academic and private sites (GAP-Net) has made measurable progress in accelerating study start-up (SSU) and positively impacting recruitment. GAP recently conducted a needs analysis to understand unique requirements of executing PD trials that have not already been addressed by other organizations, determine the transferability of GAP’s current programs to a PD network, and design a network with programs that address unmet needs. Results: No PD-specific SSU and recruitment services were identified in existing organizations. Existing organizations are primarily academic sites. GAP created a novel PD network that complements existing organizations. The collaborative network of high-performing academic and private sites will implement SSU and PD-specific recruitment tactics. All PD sites have agreed to a single IRB. GAP leveraged existing AD programs for the PD network including accelerated SSU, in-person and remote recruitment and prescreening strategies, site optimization program, and transportation services. New PD-specific programs being developed: Recruitment steering committees (community stakeholders guiding enrollment). Outreach activities with community partners (physical / occupational therapists, movement disorder exercise classes). Materials promoting awareness of the benefits of clinical trials. Conclusions: GAP’s PD network is designed to provide PD-specific solutions to SSU and recruitment and potentially reduce the time and cost of PD clinical trials. Network formation is underway with several potential trials identified.

P720 / #1002

Topic: Pre-Conference Symposia: A. Common Features of Neurodegenerative Diseases Flash Presentation: senescence

TAU TRANSGENIC MICE AND NONHUMAN PRIMATE MODELS SUPPORT MOVING SENOLYTIC THERAPY TO CLINICAL TRIALS

Lecture Title:

G. Gillispie1, V. Garbarino2, A. Ruggiero3, K. Kavanagh3, M. Orr1 1Wake Forest School of Medicine, Department Of Internal Medicine Section Of Gerontology And Geriatric Medicine, Winston-Salem, United States of America, 2UT Health San Antonio, Gerontology, San Antonio, United States of America, 3Wake Forest School of Medicine, Department Of Pathology, Winston-Salem, United States of America

Aims: Senescent cells accumulate with aging throughout the body and contribute to pathology and dysfunction. We previously determined that tau protein aggregation triggers the cellular senescence stress response in the brain, and removing senescent cells improved brain structure/function in tau transgenic mice (rTg4510). The objective of this study was to compare different drug treatments in rTg4510 mice and investigate pharmacology and target engagement in nonhuman primates (NHPs) to inform clinical trial design. Methods: WT and rTg4510 mice: We performed a direct comparison between dasatinib plus quercetin (D+Q), which demonstrated efficacy in our previous studies, and fisetin, a senolytic with a potentially lower side effect profile. Outcome measures included behavioral analyses and postmortem evaluation of target engagement in the brain. NHPs: Using the most promising drug candidate (D+Q), we tested pharmacodynamics and target engagement in old aged NHPs. Results: Mice: We observed an interaction of drug, sex and genotype on outcome measures. In totality, D+Q provided the greatest benefit on neuropathology and behavior with the lowest side effect profile. NHPs: We detected both drugs in NHP biofluids after a single oral dose; both were rapidly cleared and demonstrated unique t1/2. A decrease in senescent cell burden demonstrated target engagement. Conclusions: In the four years since presenting our initial findings for tau-induced senescence at AD/PD 2017, these additional studies further support translating senolytic therapy to humans. A phase 2 clinical trial of D+Q in older adults with mild cognitive impairment/early Alzheimer’s disease is planned for 2021.

P721 / #986

Topic: Pre-Conference Symposia: A. Common Features of Neurodegenerative Diseases Flash Presentation: senescence

OXIDATIVE STRESS REDUCES PRONGF TRANSPORT IN BASAL FOREBRAIN CHOLINERGIC NEURONS

Lecture Title:

A. Shekari1, C. Wu2, M. Fahnestock3 1McMaster University, Neuroscience Graduate Program, Hamilton, Canada, 2University of California at San Diego, Neurosciences, La Jolla, United States of America, 3McMaster University, Psychiatry & Behavioural Neurosciences, Hamilton, Canada

Aims: Profound and early basal forebrain cholinergic neuron (BFCN) degeneration is a hallmark of Alzheimer’s disease (AD). BFCNs depend for their survival and function on nerve growth factor (NGF) which is retrogradely transported from BFCN target tissue. We have shown that the retrograde transport of proNGF, the only detectable form of NGF in the brain, is reduced with age in cultured BFCNs and coincides with the loss of proNGF receptor TrkA. TrkA levels are partially regulated by protein tyrosine phosphatase (PTP)1B, an enzyme whose activity is reduced by oxidation of an active site cysteine residue. We sought to determine whether mechanisms related to oxidative stress, a longstanding hypothesized contributor to AD, account for these reductions. Methods: E18 rat BFCNs were cultured in microfluidic chambers. Oxidative stress was induced by culturing BFCNs in antioxidant-poor medium, or TCS401, a pharmacological PTP1B inhibitor, was added to the somal compartment. TrkA levels were determined by immunocytochemistry. Quantum dot-labeled proNGF was added to the axonal compartment, and proNGF accumulation was quantified in the somal compartment. Results: Elevation of intracellular oxidative stress via antioxidant deprivation significantly reduced TrkA immunoreactivity and proNGF retrograde transport. TCS401 treatment also significantly reduced proNGF retrograde transport and TrkA levels and reduced axonal uptake of proNGF-KKE, a mutant form of proNGF that only binds TrkA. Conclusions: Our results suggest that increased oxidative stress, which occurs in aging and AD, reduces TrkA levels and proNGF transport through a PTP1B-dependent mechanism. Impaired proNGF retrograde transport due to TrkA loss via aberrant PTP1B oxidation may contribute to BFCN degeneration in AD.

P722 / #1363

Topic: Pre-Conference Symposia: B. Common Features of Neurodegenerative Diseases Flash Presentation: resilience

PATHOLOGICAL BRAIN CHANGES THAT, BEYOND PLAQUES AND TANGLES, BEST PREDICT COGNITIVE DECLINE IN ALZHEIMER’S DISEASE

Lecture Title:

A.C. Amaral Massachusetts General Hospital, Neurology, Charlestown, United States of America

Aims: The presence of classic Alzheimer's (AD) pathology (plaques and tangles) does not always accurately predict clinical outcome at the individual level. Understanding the underlying alterations that lead to impaired cognition in the presence of plaques and tangles and the mechanisms that may provide protection against cognitive decline is key to develop cognitive neuroprotective therapies. Our objective is to characterize pathological brain changes that, in the presence of AD pathology, best predict neuronal/synaptic derangement and impaired cognition. Methods: We conducted quantitative histopathological and biochemical assessments of the entorhinal cortex (EC) in subjects with equivalent loads of plaques and tangles at postmortem and whose antemortem cognition was either normal (resilient) or impaired (AD). Results: We previously reported that, despite similar burdens of plaques and tangles, AD brains exhibited significant neuronal loss, reduction in cortical thickness, and abnormal accumulation of soluble p-Tau in synapses compared to resilient brains. Our current results further show that AD brains exhibited higher burdens of TDP-43 aggregates, enhanced glial cell activation, increased deposits of fibrin(ogen) and expression of complement proteins that are part of the innate immune system compared to resilient brains. These neuroinflammatory changes had a robust negative association with loss of neurons and synaptic integrity. Conclusions: These data provide further insight into pathways and molecular targets potentially involved in human brain resilience to classic AD pathology. Our results suggest that neuroinflammation may have a larger role than previously recognized in the anatomical disruption of neurons and synapses, and the subsequent cognitive decline that characterize AD.

P723 / #1367

Topic: Pre-Conference Symposia: C. Common Features of Neurodegenerative Diseases Flash Presentation: neurovascular

RELATIONSHIP OF CHOROIDAL THICKNESS TO CLINICAL AND IMAGING METRICS IN PARKINSON’S DISEASE: A PILOT STUDY WITH OPTICAL COHERENCE TOMOGRAPHY

Lecture Title:

G. Brown1, J. Nguyen1, D. Brown2, M. Lewis1, M. Camacci3, S. Kim3, G. Du1, L. Kong4, X. Huang1, E. Bowie3 1Pennsylvania State University, Neurology, Hershey, United States of America, 2Lewis Katz School of Medicine, Department Of Ophthalmology, Philadelphia, United States of America, 3Pennsylvania State University, Ophthamology, Hershey, United States of America, 4Pennsylvania State University, Biostatistics, Hershey, United States of America

Aims: Visual changes are common in Parkinson’s disease (PD) with recent studies suggesting retinal involvement. This study investigates the relationship between neurovascular alterations in the eye and other PD-related processes. Methods: Ten PD and 12 control (HC) participants (2 eyes each) underwent optical coherence tomography (OCT) evaluation. Retinal nerve fiber (RNFL), ganglion cell/inner plexiform (GCIP), and choroidal (vascular) thickness were measured in 4 subregions [superior (S), temporal (T), inferior (I), and nasal (N)] and at 3 foveal distances (1, 1.5, and 3 mm). Retinal and choroidal thickness measurements were compared between PD and controls and associated with clinical [motor, olfaction, visuospatial function] and Substantia Nigra pars compacta (SNpc) MRI metrics. Results: Compared to Controls, choroidal thickness was increased in PD, adjusting for age, sex, and axial length. We identified no changes in RNFL or GCIP. Choroidal thickness negatively correlated with UPDRS-III in the temporal and nasal quadrant at 1mm and nasal quadrant at 3mm, and positively correlated with a figure-copy task on average, and superior 1, 1.5 and 3mm locations and temporal 1 and 1.5mm areas. Choroidal thickness negatively correlated with MRI in SNpc in all regions except superior and nasal quadrants at 3mm. Conclusions: We found patient’s with PD to experience alterations in choroidal thickness that relates to clinical and SNpc MRI metrics of disease severity. Further studies are warranted to better understand the clinical and scientific relevance and potential use of early monitoring of choroidal changes in PD.

P724 / #746


MENINGEAL LYMPHATIC DRAINAGE MODULATES ANTI-AΒ IMMUNOTHERAPY IN ALZHEIMER’S DISEASE

Lecture Title:

S. Da Mesquita1, Z. Papadopoulos2, T. Dykstra2, C. Cruchaga3,4, O. Harari3,4, D. Holtzman4,5, J. Kipnis2,4 1Mayo Clinic, Department Of Neuroscience, Jacksonville, United States of America, 2Washington University, Department Of Pathology And Immunology, Division Of Immunobiology, St. Louis, United States of America, 3Washington University in St. Louis, Department Of Psychiatry, St. Louis, United States of America, 4Washington University School of Medicine, Hope Center For Neurological Disorders, St. Louis, United States of America, 5Washington University, Neurology, St. Louis, United States of America

Aims: The brain-draining meningeal lymphatic vasculature modulates brain fluid flow, learning and memory in aged mice and brain amyloid beta (Aβ) clearance in mouse models of Alzheimer’s disease (AD). While defective meningeal lymphatic drainage exacerbated brain and meningeal Aβ pathology, it is unknown if or how altered brain drainage mediated by meningeal lymphatics affects the brain blood vasculature, microglial function or anti-Aβ immunotherapy in AD. Methods: Meningeal lymphatic function was evaluated in 5xFAD mice at different ages. Brain blood endothelial cell and microglial transcriptomes were assessed by single cell RNA sequencing and the changes in Aβ plaque clearance by monoclonal antibodies were measured upon prolonged manipulation of meningeal lymphatic drainage in adult or aged AD transgenic mice. eQTL analysis was performed to test for a potential association between SNPs that are highly expressed in lymphatic endothelial cells and increase the risk for AD and gene expression variation in microglia isolated from the AD brain. Results: Herein, we show that modulating meningeal lymphatic function in AD transgenic mice affects the clearance of Aβ by monoclonal antibodies, a feature that is closely associated with transcriptomic changes in brain blood endothelial cells and microglia. Furthermore, SNPs that are highly expressed in lymphatic vasculature and associated with increased risk for AD are also markers of genetic variance in microglia isolated from the AD brain. Conclusions: Altogether, our data emphasizes the notion that manipulating brain drainage by meningeal lymphatic vasculature could affect brain cell function and provide an important adjuvant to monoclonal antibody-based immunotherapies for the treatment of AD.

P725 / #603


FMNL2 INTERACTS WITH CEREBROVASCULAR RISK FACTORS TO ALTER ALZHEIMER’S DISEASE RISK

Lecture Title:

R. Mayeux1, N. Raghavan2, S. Sariva2, A. Lee2, Y. Gao2, D. Reyes-Dumeyer2, P. De Jager2, D. Bennett3, V. Menon2, R. Lantigua2, W. Kukull4, A. Brickman2, J. Manly2, J. Gutierrez1, B. Vardarajan2, G. Tosto2 1Columbia University, Neurology, New York, United States of America, 2Columbia University, Taub Institute, New York, United States of America, 3Rush University, Rush Alzheimer’s Disease Center, Rush University Medical Center, Chicago, United States of America, 4University of Washington, Epidemiology, Seattle, United States of America

Aims: Late-onset Alzheimer’s disease affects more than 6 million Americans, and is frequently co-morbid with cerebrovascular disease. Neuropathological data indicate that brain tissue from up to 70% of patients with Alzheimer’s disease have the hallmark neuritic plaques and neurofibrillary tangles with varying degrees of cerebrovascular disease. Given the high frequency of this comorbidity, a number of studies have assessed the relationship between cerebrovascular risk factors such as hypertension, being overweight, diabetes and coronary heart disease and Alzheimer’s disease. Methods: Participants age 65 years or older from five multi-ethnic cohorts (N=14,669) were included in genome-wide association meta-analyses for AD including an interaction factor for a cardiovascular risk factor score created from body mass index, hypertension, heart disease, and diabetes. Significant gene level results were substantiated using neuropathological and gene expression data. Results: At the gene-level, FMNL2 interacted with the cardiovascular risk factor score to significantly modify Alzheimer's disease risk (p= 7.7x10-7). A SNP within FRMD4B, rs1498837, was nominally significant (p=7.95x10-7). Increased FMNL2 expression was significantly associated with brain infarcts and Alzheimer's disease. Conclusions: FMNL2 is highly expressed in the brain and has been associated with ischemic stroke and failures in endosomal trafficking, a major pathway in AD pathology. The results highlight an interaction between FMNL2 and cardiovascular risk factors on susceptibility to Alzheimer's disease.

P726 / #1200


META-ANALYSIS OF GENOME-WIDE DNA METHYLATION IDENTIFIES SHARED ASSOCIATIONS ACROSS NEURODEGENERATIVE DISORDERS

Lecture Title:

M. Nabais Et Al1,2 1University of Queensland, Institute For Molecular Biosciences, Brisbane, Australia, 2University of Exeter Medical School, Complex Disease Epigenomics Group, Exeter, United Kingdom

Aims: People with neurodegenerative disorders can show diverse clinical syndromes, genetic heterogeneity, and distinct brain pathological changes, but studies report overlap between these features. DNA methylation provides a way to explore this overlap and heterogeneity as it is determined by the combined effects of genetic variation and the environment. In this study we aimed to identify shared blood DNA methylation differences between 471-1,522 controls and people with Alzheimer’s disease (N = 161), amyotrophic lateral sclerosis (N = 3,032) and Parkinson’s disease (N = 1,132). Methods: We used a mixed-linear model method (MOMENT) that has been shown to account for the effect of (un)known confounders, to test for association of each DNA methylation site with each disorder. Results: While only three probes were found to be genome-wide significant in each MOMENT association analyses of amyotrophic lateral sclerosis and Parkinson’s disease (and none with Alzheimer’s disease), a fixed-effects meta-analysis of the three disorders results in 12 genome-wide significant differentially methylated positions. Predicted immune cell-type proportions were disrupted across all disorders. Protein inflammatory markers were correlated with profile sum-scores derived from disease-associated immune cell-type proportions in a healthy ageing cohort. In contrast, they were not correlated with MOMENT DNA methylation-derived profile sum-scores, calculated using effect sizes of the 12 differentially methylated positions as weights. Conclusions: We have identified shared differentially methylated positions in whole blood between neurodegenerative disorders that point to shared pathogenic mechanisms. These shared differentially methylated positions may reflect causes or consequences of disease, but they are unlikely to reflect cell-type proportion differences.

P727 / #1410


IDENTIFYING FACTORS AND PREDICTIVE MODEL FOR HOSPITALIZATION OUTCOMES IN GERIATRIC PATIENTS WITH NEURODEGERATIVE DISEASES

Lecture Title:

L. Wang1, X. Wang1, C. Ezeana1, M. Puppula1, Y. He1, Z. Yin1, E. Lai2, S. Wong1 1Houston Methodist Research Institute, Smab, Houston, United States of America, 2Houston Methodist Hospital, Houston Methodist Stanley H. Appel Department Of Neurology, Houston, United States of America

Aims: The risk of being affected by neurodegenerative diseases which include Alzheimer’s and Parkinson’s increases dramatically with age, and this group of geriatric patients will use up more healthcare costs and be in hospitals more often than other geriatric patients. It is important to understand all significantly associated factors so as to lessen the hospital ordeal and achieve good hospitalization outcomes. Objectives. To identify factors associated with hospitalization outcomes for geriatric patients with neurodegeneration. To combine these factors to develop an effective machine learning-based model for predicting patient outcomes as early as the 2nd day of admission. Methods: We retrieved clinical records of 18,374 geriatric neurodegeneration encounters at Houston Methodist’s 8-hospital system over ten years (January 2010 - December 2019), including over 150 risk factors categorized into three patient risk groups: pre-hospitalization risk factors, in-hospital risk factors, and comorbidities during hospitalization. Hospitalization outcomes, based on length of stay, discharge disposition and complications, were categorized into good or poor outcomes. Identification of factors with significant difference between these two groups was performed using ANOVA test. Using these significant risk factors, we trained a deep learning classification model to predict patient hospitalization outcomes. Results: We narrowed down to 20 significant factors using ANOVA. Our classification model performance based on AUC was 85% and outperformed prevalent baseline models, such as logistic regression, in identifying patients with good outcomes. Conclusions: We are implementing the deep learning predictive model in the clinics to aid physicians and caregivers at Houston Methodist in resource reallocation to improve patient outcomes.

Book of abstracts · Topic: Theme A: β-Amyloid Diseases / A1.a. Disease Mechanisms,...

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