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  • Preparation and Characterization of Manganese (II) Complexes with Potential

    Therapeutic Application against α-Synuclein Aggregation

    Kate Byrne

    BSc Medicinal Chemistry & Pharmaceutical Science

    Year 3 Project

  • Overview

    • Aim of the Project

    • Parkinson’s Disease and Etiology

    • α-Synuclein and Lewy Bodies

    • Project Rationale and Objectives

    • Synthesis of the compounds

    • Characterisation of the compounds

    • Conclusion

  • Aim

    The aim of this project was to synthesize, purify and characterise three manganese(II) complexes as part of a structure-activity relationship (SAR) study of a potential

    therapeutic system for Parkinson’s disease

  • Parkinson’s Disease Parkinson’s disease, (PD) is a progressive neurological

    disease which is a result of the loss of dopamine producing brain cells

    Parkinson’s disease is named after Dr. James Parkinson who first reported symptoms of the disease in 1817 when

    it was referred to as “Paralysis Agitans”

    Etiology Idiopathic

    Figure 1: Dr. James Parkinson

  • Statistics

    • Parkinson’s disease affects 6.3 million people worldwide.

    • Over 9,000 of those affected by the disease are Irish.

    • The disease typically develops at the age of 65 however people can develop “early-onset” parkinson’s before reaching 50 years.

    • Current life expectancy: 81 years

    • Projected Life expectancy 2030: 90+ years

  • α-Synuclein α-Synuclein is a protein that is abundant in the human brain.

    In the early stages of Parkinson’s disease misfolded α-synuclein

    proteins are converted to pathological oligomers and higher

    order aggregates that form fibrils leading to the formation of

    Lewy Bodies.

    Figure 2: α-Synuclein aggregation stages

    Emil Paleek, et al., Changes in interfacial properties of α-synuclein preceding its aggregation, Analyst, 2008, 133, 76.

  • Lewy bodies are abnormal α-Synuclein protein aggregates that develop in nerve cells in regions of the brain that are

    involved in motor control.

    The presence of the Lewy Body protein aggregates in the

    brain is a pathological hallmark of Parkinson’s Disease.1

    Inhibiting α-Synuclein aggregation offers a therapeutic target for Parkinson’s Disease and evidence in the literature

    suggests a role for inorganic medicinal chemistry2

    1. He-Jin Lee, et al., Extracellular α-synuclein a novel and crucial factor in Lewy body diseases, Nature Reviews Neurology, 2014, 10, 92-98. 2. D.J. Haynes, S. Lim, P.S. Donnelly, Metal complexes designed to bind to amyloid-β for diagnosis and treatment of Alzheimer’s disease, Chem. Soc. Rev., 2014, 43, 6701-6715.

    Lewy bodies and Parkinson’s Disease

  • Lead Compound

    The complex salt, MD1 has been shown to be an excellent lead compound for the prevention of α-Synuclein aggregation.

    MD1:

    [Mn2(oda)(phen)4(H2O)2][Mn2(oda)(phen)4(oda)2]4H2O

    Where; odaH2 = octanedioic acid; phen = 1,10-phenanthroline

    Figure 4: Structures of MD1 and Octanedioic Acid

    http://www.google.ie/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&ved=0CAcQjRxqFQoTCNiXvbOj_sgCFUH-DgodfFYP4w&url=http://www.sigmaaldrich.com/catalog/product/aldrich/s5200&psig=AFQjCNE609-ohZOlrMCkToSsECmA8iKG-A&ust=1446984477245297

  • • Comprised of bi-nuclear Cationic and Anionic components (Complex salt).1

    • Bridging and terminal octanedioate ligands, deprotonated octanedioic acid

    • Highly water soluble.

    • Exhibits antioxidant capability.2

    • Reduces α-Synuclein aggregation in fungal (saccharomyces cerevisiae) and mammalian (HEK293) cellular models.2

    1. M. McCann, M. Devereux, et al. Synthesis and structure of the Mn2 (II,II) complex salt [Mn2(oda)(phen)4(H2O)2] [Mn2(oda)2(phen)4] (odaH2 = octanedioic acid): a catalyst for H2O2 disproportionation., J. Chem. Soc., Chem. Commun., 1994, 2643. 2. T. Ribeiro, M. McCann, M. Devereux, M. Pereira, et al., Evaluation of antioxidant activity of a Mn2+complex salt and its potential therapeutic use against alpha—synuclein aggregation. Manuscript in preparation. (Target Journal: Nature Communications)

  • Research question

    What are the structural aspects of MD1 that are important for its

    α-Synuclein aggregation inhibitory properties?

  • Objective of the Structure-Activity Relationship Study

    To investigate analogues of MD1 to determine how variations in structure influence the α-Synuclein aggregation inhibitory potential of this class of

    manganese(II) complex.

    Note: The analogues were varied in terms of the number of Manganese centres in the complex as well as the charge

    on the complex

  • Key Objectives

    To synthesise and characterise three known analogues of MD1 with variations in structure achieved by replacing the octanedioate ligands with aliphatic

    dicarboxylic acid ligands of varying chain length

    [Mn(hxda)(phen)2(H2O)].7H2O

    1

    [Mn(pda)(phen)]2

    [Mn2(bda)2(phen)2(H2O)].2H2O3

    Hexanedioic acid (hxdaH2)

    Pentanedioic acid (pdaH2)

    Butanedioic acid (bdaH2)

    1. M. McCann, M. Devereux, et al., Manganese(II) complexes of hexanedioic and heptanedioic acids: X-ray crystal structure of [Mn(hxda)(phen)2(H2O)].7H2O and [Mn(phen)2(H2O)][Mn(hpda)(phen)2(H2O)](hpda) .12.5H2O, Polyhedron, 1997, 16, 2741.

    2. Martin Curran, PhD Thesis, DIT/NUIM 1996.

    3. M. McCann, M. Devereux, et al., Synthesis, X-ray crystal structure and catalytic activities of manganese(II) butanedioic acid complexes [Mn(bda)(phen)2(H2O)4].2H2O and {[Mn(bda)(bipy2(H2O)2].H2O }n, Polyhedron, 1997, 16, 2547.

    Octanedioic acid (odaH2)

    http://www.google.ie/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&ved=0CAcQjRxqFQoTCPbF8Iyi_sgCFQagDgodI_EHsw&url=http://wellnessadvocate.com/?uid=7602&psig=AFQjCNGrKwcgSCyczD-QZMAfkxJA9NS8-g&ust=1446984133921066http://www.google.ie/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&ved=0CAcQjRxqFQoTCJKw6tWj_sgCFUeNDwodfxENbg&url=http://www.sigmaaldrich.com/catalog/product/sigma/a26357&psig=AFQjCNHdRmFP7ngNfYLnVzdZ306YvH1JMw&ust=1446984553695267https://www.google.ie/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&ved=0CAcQjRxqFQoTCLW3vJOn_sgCFUKfDgodcO4Exg&url=https://answers.yahoo.com/question/index?qid=20111029072246AA1nULk&psig=AFQjCNHjvQWi61Hr9N7EydxL7Xb3A3qzZw&ust=1446985445384287http://www.google.ie/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&ved=0CAcQjRxqFQoTCNiXvbOj_sgCFUH-DgodfFYP4w&url=http://www.sigmaaldrich.com/catalog/product/aldrich/s5200&psig=AFQjCNE609-ohZOlrMCkToSsECmA8iKG-A&ust=1446984477245297

  • Structure of the Analogue Complexes

    [Mn(hxda)(phen)2(H2O)].7H2O : Mononuclear (1 manganese centre), 2 phens per manganese, neutral charge

    [Mn(pda)(phen)]: Polymeric, 1 phen per manganese, neutral

    charge.

    [Mn2(bda)2(phen)2(H2O)].2H2O: Binuclear (2 manganese centres), 1 phen per

    manganese, neutral charge

  • Synthetic Scheme Stage One: Generation of the manganese(II) dicarboxylate

    precursors

    Where; R = -(CH2)4 - or -(CH2)3 -or -(CH2)2 -

  • Synthetic Scheme Stage Two: Reaction of the precursor complexes with 1,10-

    phenanthroline to produce the analogues of MD1

    {Mn(OOC-(CH2)n-COO)}x + 4 phen → [Mn(hxda)(phen)2(H2O)].7H2O (where n = 4) [Mn(pda)(phen)] (where n = 3) [Mn2(bda)2(phen)2(H2O)].2H2O (where n = 2)

    EtOH

    Reflux

  • Characterisation

    The three precursor complexes and the three analogue complexes were characterised using the following

    techniques:

    • Infra-Red (IR) Spectroscopy

    • Magnetic Susceptibility Analysis

    • Inductively-Coupled Mass Spectrometry (ICP-MS)

  • Infrared Spectroscopy An IR spectrum was recorded of:

    • The Ligand

    • The Precursor Complex

    • The Final Complex

    Each of the spectra was then overlayed

  • Hexanedioic Acid

  • [Mn(hxda)].H2O

  • [Mn(hxda)(phen)2(H2O)].7H2O

  • Hexanedioic Acid Spectra Overlay

  • Infrared Spectroscopy

    Precursor Complex and Analogue Complex Similarities

    Characteristic Carbonyl Peak

    • Asymmetric Stretch 1590-1547cm-1

    • Symmetric Stretch 1400-1420cm-1

  • Determination of [Mn] Using ICP

    y = 66202x + 5669.9

    R² = 0.9999

    0

    100000

    200000

    300000

    400000

    500000

    600000

    -1 0 1 2 3 4 5 6 7 8 9

    Intensity

    Concentration (ppm)

    ICP Standards

  • Mn2+ Analysis Name of Complex Theoretical

    Concentration

    (ppm)

    Experimental

    Concentration

    (ppm)

    [Mn(bda)].2H2O 5.3 4.647

    [Mn(pda)].H2O 5.36 4.001

    [Mn(hxda)].H2O 5 3.468

    Drug 1,

    [Mn2(bda)2(phen)2(H2O)].2

    H2O

    5.4 9.5

    4.75 x 2

    Drug 2, [Mn(pda)(phen)] 5.45 4.325

    Drug 3,

    [Mn(hxda)(phen)2(H2O)].7

    H2O

    4.68 1.468

  • Magnetic Susceptibility Balance

    µ𝑠 .𝑜. = 𝑛(𝑛 + 2)

    Where;

    N = number of unpaired electrons, n=5

    µ𝑠 .𝑜. = 5(5 + 2) = 35 = 5.92

    µs.o. = 5.92B.M.

  • Gram Magnetic Susceptibility Calculation

    Χg =

    c ∗ l ∗ (R − R0)

    109 ∗ m

    C= calibration constant, 1.05

    L= sample length, cm

    R= reading taken of sample

    R0 = reading taken with no sample

    M= sample mass (g)

  • 𝛸𝑀 = 𝛸𝑔 ∗ 𝑀

    Where;

    𝛸𝑀 = Molar Magetic Susceptibility

    𝛸𝑔= Gram Magnetic Susceptibility (cm3 g-1)

    M= Molar Mass (g mol-1)

    Molar Magnetic Susceptibility

  • Effective Magnetic Moment

    µEFF = 2.828 𝑇𝑋𝑀

    µEFF = effective magnetic moment, B.M.

    T= Temperature (K), 292K

    XM = Molar magnetic Susceptibility, cm3 mol-1

  • Name of Substance Theoretical Value

    B.M.

    Experimental

    Value B.M.

    [Mn(bda)].2H2O 5.92 5.87

    [Mn(pda)].H2O 5.92 6.16

    [Mn(hxda)].H2O 5.92 3.09

    Drug 1,

    [Mn2(bda)2(phen)2(H2O)].

    2H2O

    5.92 8.91

    4.455 x 2

    Drug 2, [Mn(pda)(phen)] 5.92 6.36

    Drug 3,

    [Mn(hxda)(phen)2(H2O)].

    7H2O

    5.92 8.62

  • Conclusion

    1. The complex salt [Mn2(oda)(phen)4(H2O)2] [Mn2(oda)(phen)4(oda)2]4H2O (MD1) is an excellent lead candidate for the prevention of α-Synuclein aggregation.

    2. Three known analogues of MD1 {[Mn(hxda)(phen)2(H2O)].7H2O; [Mn(pda)(phen)]; and [Mn2(bda)2(phen)2(H2O)].2H2O along with their precursor manganese(II) dicarboxylate complexes have been synthesised using methods previously published

    3. All six complexes have been characterised using a range of analytical techniques.

    4. Further work is required to purify the complexes in preparation for their use in a structure-activity relationship study.

  • Acknowledgements

    I would like to thank my supervisors Professor Michael Devereux and Dr. Patricia Ennis for their help and support

    when carrying out this project.

    I would also like to thank the School of Chemical & Pharmaceutical Sciences for allowing me to use their

    equipment.

  • Thank You