A novel site on γ3 subunits important for assembly of GABAA receptors
Tu1872 NF-κβ Subunits Confer Divergent and Overlapping Functions in Apoptosis Resistance of...
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AGA Abstracts After day 12, tumor growth was measured every 3 days (x-axis) and reported as "Tumor volume" on the Y-axis. "Untreated" indicates that tumor cells were not transduced. Tu1868 PTEN Loss in the Neoplastic Stroma of Pancreatic Ductal Adenocarcinoma (PDAC) Is a Strong Predictor of Distant Metastasis Eva Karamitopoulou, Inti Zlobec, Beat Gloor, Alessandro Lugli, Aurel Perren Background: PTEN is considered a major tumor suppressor in PDAC. PTEN loss is known to promote epithelial-mesenchymal transition (EMT) thus increasing the invasive and metastatic potential of tumor cells. Our aim was to determine the role of PTEN in PDAC progression from localized to metastatic disease and to correlate PTEN alterations with other prognostic features as well as with overall and disease-free survival of the patients. Methods: Immunohis- tochemistry for PTEN was performed on a multipunch Tissue Microarray (TMA) of 120 well characterized PDACs (4 tumor cores each) with full clinico-pathological, survival time, follow-up (metastasis and recurrence) and adjuvant therapy information. Another 3 TMAs including precancerous lesions (PanINs), matched lymph node metastases and normal pan- creatic parenchyma were stained in parallel. In the PDAC-TMA, PTEN immunoreactivity was evaluated both in the tumor- and in the stromal cells. Results: We found a significant progressive loss of PTEN expression from normal pancreatic ductal epithelia (21.5%) to precancerous lesions (PanINs; 13%) and to PDAC cells (6.2%), with reappearance of PTEN in a subset of matched lymph node metastases (12.3%) (p ,0.0001). Moreover, PTEN loss in the tumor cells was associated with higher tumor grade (p=0.0338) and showed a trend towards increased T-stage (p=0.0596). PTEN loss in the neoplastic stroma was strongly associated with the presence of distant metastasis (p=0.0045) and showed a trend towards worse overall (p=0.0791) and disease-free survival (p=0.066). Sensitivity, specificity, positive and negative predictive values for loss of stromal PTEN expression and metastatic disease were 60%, 80.2%, 25% and 94.8% respectively. Conclusions: PTEN loss in PDAC seems to play an important role in the neoplastic progression and correlates to a more aggressive phenotype. Moreover, loss of PTEN in the neoplastic stroma shows a strong association with distant metastasis, suggesting that the stromal cells are actively participating in the process of pancreatic cancer dissemination. Tu1869 Targeting Pancreatic Cancer Stem Like Cells Using a Nanoparticle Modified by L-Fucose Rishu Takimoto, Makoto Yoshida, Kazuyuki Murase, Yasushi Sato, Masahiro Hirakawa, Fumito Tamura, Tsutomu Sato, Takahiro Osuga, Koji Miyanishi, Kohichi Takada, Tsuyoshi Hayashi, Masayoshi Kobune, Junji Kato Pancreatic ductal adenocarcinoma has a dismal prognosis because of its aggressiveness and the lack of effective therapies. New strategies to improve treatment and survival are therefore urgently required. Numerous fucosylated antigens in sera serve as tumor markers for cancer detection and evaluation of treatment efficacy. Increased expression of fucosyltransferases has also been reported for pancreatic cancer. These enzymes accelerate malignant transformation through fucosylation of sialylated precursors, suggesting a crucial requirement for fucose by pancreatic cancer cells. With this in mind, we developed fucose-bound nanoparticles as vehicles for delivery of anticancer drugs specifically to cancer cells. L-fucose-bound liposomes containing Cy5.5 or Cisplatin were effectively delivered into CA19-9 expressing pancreatic cancer cells. Furthermore, these nanoparticles could target CD24+/CD44+ pancreatic cancer stem like cells since CD44 is also one of the fucosylated-antigen. Excess L-fucose decreased the efficiency of Cy5.5 introduction by L-fucose-bound liposomes, suggesting L-fucose- receptor-mediated delivery. Intravenously injected L-fucose-bound liposomes carrying Cis- platin were successfully delivered to pancreatic cancer stem like cells, mediating efficient tumor growth inhibition as well as prolonging survival in mouse xenograft models. This modality represents a new strategy for pancreatic cancer stem cell-targeting therapy. Tu1870 Pifithrin-μ, a New HSP70 and Autophagy Inhibitor, Can Enhance Antitumor Effects of TRAIL on Pancreatic Cancer Hiroyuki Monma, Mamoru Harada, Nanae Harashima, Eiji Hira, Yasunari Kawabata, Yoshitsugu Tajima Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) can induce apoptosis of various types of cancers with little cytotoxicity to normal cells through the stimulation of death receptors. In this study, after confirming that heat shock protein (HSP) 70 and autophagy play protective roles in antitumor effects of TRAIL, we examined a sensitizing effect of pifithrin-μ, which is an HSP70 and autophagy inhibitor, on TRAIL-induced antitumor effects on pancreatic cancer cell lines. TRAIL decreased the cell viability and induced apoptosis in three out of four cell lines tested. Regarding TRAIL-sensitive MiaPaca-2 and Panc-1, knockdown of HSP70 or Beclin-1, a key molecule for autophagy, with RNA interference significantly enhanced TRAIL-induced antitumor effects, i.e., decreased the cell viability and colony-forming capacity. Pifithrin-μ had the potential to induce both apoptosis and cell growth arrest. In addition, pifithrin-μ seemed to inhibit I κBα degradation in TRAIL-treated cancer cells probably via inhibition of the autophagy-lysosome pathway, thereby resulting in inhibition of NF- κB activation in TRAIL-treated cancer cells. In a xenograft mouse model, combination therapy with TRAIL plus pifithrin-μ significantly inhibited MiaPaca-2 tumor growth compared to treatment with either agent alone. These results suggest that pifithrin- μ is a promising agent for use in therapies intended to enhance the antitumor effects of TRAIL on pancreatic cancer. S-868 AGA Abstracts Tu1871 Validation of MD Anderson Group B Borderline Resectable Pancreatic Cancer Tak Geun Oh, Moon Jae Chung, Seungmin Bang, Seung Woo Park, Jae Bock Chung, Si Young Song, Jinsil Seong, Ho Kyoung Hwang, Chang Moo Kang, Woo Jung Lee, Jeong Youp Park BACKGROUND: Borderline resectable pancreatic cancer (BRPC) is categorized into the group A, B, and C according to MD Anderson criteria. The group A indicates patients with tumor abutment of the visceral arteries or short-segment occlusion of the superior mesenteric vein. In the group B, patients have findings suggestive but not diagnostic of metastasis, such as carcinomatosis, paraaortic lymph node metastasis. In this study we tried to validate whether group B of BRPC could be truly categorized as the borderline resectable group, and concurrent chemoradiotherapy (CCRT) could be a treatment option. STUDY DESIGN: Among the patients who received preoperative CCRT, we defined patients with BRPC into the group A or B. Rate of operation, recurrence rate, progression-free survival, and overall survival were compared between the group A and B. RESULTS: Between November 2005 and April 2012, 53 patients with pancreatic adenocarcinoma were classified as borderline resectable. In the group A, 23 (60.5%) of 38 patients underwent pancreatectomy after CCRT, but, in the group B, only 5 (33.3%) of 15 patients underwent pancreatectomy, mainly due to the progression of suspected distant metastasis. There was significant difference in overall survival between the group A and B patients (median OS; group A : 21.2 months vs group B : 10.2 months, p=0.007). Of the patients who underwent pancreatectomy, the group B had higher recurrence rate compared to the group A (recurrence rate; group A : 11 of 23 patients (47.8%) vs group B : 5 of 5 patients (100%), p=0.033). Also, of the patients who underwent pancreatectomy, the group B showed shorter disease-free survival compared to group A (group A : 29.0 months vs group B : 8.3 months, p=0.054). CONCLUSIONS: This is the first report to validate the definition of BPRC and indication for CCRT . We suggest that the group B BRPC should be considered as systemic diseases and treated with systemic therapy first rather than CCRT. Table 1. Comparison of survival outcome between group A and B borderline resectable pancreatic cancer for all of 53 patients Table 2. Comparison of survival outcome between group A and B borderline resectable pancreatic cancer for 28 patients who had pancreatectomy Tu1872 NF-κB Subunits Confer Divergent and Overlapping Functions in Apoptosis Resistance of Pancreatic Cancer Cells Claudia Geismann, Frauke Grohmann, Susanne Sebens, Gabriele Wirths, Anita Dreher, Robert Haesler, Philip C. Rosenstiel, Günter Schneider, Sebastian Zeissig, Stefan Schreiber, Heiner Schäfer, Alexander Arlt Pancreatic ductal adenocarcinoma (PDAC) represents one of the deadliest malignancies with an overall life expectancy of six months despite palliative radio-chemotherapy. The transcription factor NF- κB has been shown to be a critical component of dysregulated transcription factor activity conferring profound apoptosis resistance. Despite extensive data on the role of the most abundant NF- κB subunit p65/RelA in PDAC apoptosis control, there is only little knowledge on the role of the other Rel subunits. In the present study, three pancreatic carcinoma cell lines (Panc1, Patu8988, MiaPaca2) were analyzed for the role of RelA, RelB and c-Rel in resistance against TRAIL, etoposide or gemcitabine induced apoptosis. Resistant Panc1 and Patu8988 cells exhibit a high basal NF- κB activity that was further strongly induced by TRAIL and moderately upregulated by etoposide or gemcitabine. In contrast, sensitive MiaPaca2 cells displayed only little basal NF- κB activity and only weak induction by TRAIL, etoposide or gemcitabine. Transfection with siRNA against the RelB or c-Rel subunits of NF- κB sensitized the TRAIL and chemotherapy resistant Panc1 and Patu8988 cells in a comparable fashion like siRNA targeting the p65/RelA subunit. Gel shift analysis revealed that, together with the p65/RelA subunit, c-Rel is part of the TRAIL, etoposide or gemcitabine inducible NF- κB complex in PDAC. In addition the basal NF- κB complex, mediating resistance against chemotherapeutic drug induced apoptosis, is com- posed of all three subunits as revealed by Shift-Western analysis. Western-Blot-analysis of known anti-apoptotic NF- κB target genes (e.g. Bcl-2 family and cIAPs) and mRNA-array based expression analysis revealed a divergent non overlapping function of the Rel NF- κB subunits in established and novel anti-apoptotic NF- κB target genes. Furthermore, c-Rel elicits a proapoptotic function in the sensitive MiaPaca2 cell line as indicated by an increase in the apoptotic response in cells transfected with c-Rel specific siRNA. In conclusion, all
Transcript of Tu1872 NF-κβ Subunits Confer Divergent and Overlapping Functions in Apoptosis Resistance of...
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sAfter day 12, tumor growth was measured every 3 days (x-axis) and reported as "Tumorvolume" on the Y-axis. "Untreated" indicates that tumor cells were not transduced.
Tu1868
PTEN Loss in the Neoplastic Stroma of Pancreatic Ductal Adenocarcinoma(PDAC) Is a Strong Predictor of Distant MetastasisEva Karamitopoulou, Inti Zlobec, Beat Gloor, Alessandro Lugli, Aurel Perren
Background: PTEN is considered a major tumor suppressor in PDAC. PTEN loss is known topromote epithelial-mesenchymal transition (EMT) thus increasing the invasive and metastaticpotential of tumor cells. Our aim was to determine the role of PTEN in PDAC progressionfrom localized to metastatic disease and to correlate PTEN alterations with other prognosticfeatures as well as with overall and disease-free survival of the patients. Methods: Immunohis-tochemistry for PTEN was performed on a multipunch Tissue Microarray (TMA) of 120well characterized PDACs (4 tumor cores each) with full clinico-pathological, survival time,follow-up (metastasis and recurrence) and adjuvant therapy information. Another 3 TMAsincluding precancerous lesions (PanINs), matched lymph node metastases and normal pan-creatic parenchyma were stained in parallel. In the PDAC-TMA, PTEN immunoreactivitywas evaluated both in the tumor- and in the stromal cells. Results: We found a significantprogressive loss of PTEN expression from normal pancreatic ductal epithelia (21.5%) toprecancerous lesions (PanINs; 13%) and to PDAC cells (6.2%), with reappearance of PTENin a subset of matched lymph node metastases (12.3%) (p ,0.0001). Moreover, PTEN lossin the tumor cells was associated with higher tumor grade (p=0.0338) and showed a trendtowards increased T-stage (p=0.0596). PTEN loss in the neoplastic stroma was stronglyassociated with the presence of distant metastasis (p=0.0045) and showed a trend towardsworse overall (p=0.0791) and disease-free survival (p=0.066). Sensitivity, specificity, positiveand negative predictive values for loss of stromal PTEN expression and metastatic diseasewere 60%, 80.2%, 25% and 94.8% respectively. Conclusions: PTEN loss in PDAC seemsto play an important role in the neoplastic progression and correlates to a more aggressivephenotype. Moreover, loss of PTEN in the neoplastic stroma shows a strong associationwith distant metastasis, suggesting that the stromal cells are actively participating in theprocess of pancreatic cancer dissemination.
Tu1869
Targeting Pancreatic Cancer Stem Like Cells Using a Nanoparticle Modifiedby L-FucoseRishu Takimoto, Makoto Yoshida, Kazuyuki Murase, Yasushi Sato, Masahiro Hirakawa,Fumito Tamura, Tsutomu Sato, Takahiro Osuga, Koji Miyanishi, Kohichi Takada,Tsuyoshi Hayashi, Masayoshi Kobune, Junji Kato
Pancreatic ductal adenocarcinoma has a dismal prognosis because of its aggressiveness andthe lack of effective therapies. New strategies to improve treatment and survival are thereforeurgently required. Numerous fucosylated antigens in sera serve as tumor markers for cancerdetection and evaluation of treatment efficacy. Increased expression of fucosyltransferases hasalso been reported for pancreatic cancer. These enzymes accelerate malignant transformationthrough fucosylation of sialylated precursors, suggesting a crucial requirement for fucoseby pancreatic cancer cells. With this in mind, we developed fucose-bound nanoparticles asvehicles for delivery of anticancer drugs specifically to cancer cells. L-fucose-bound liposomescontaining Cy5.5 or Cisplatin were effectively delivered into CA19-9 expressing pancreaticcancer cells. Furthermore, these nanoparticles could target CD24+/CD44+ pancreatic cancerstem like cells since CD44 is also one of the fucosylated-antigen. Excess L-fucose decreasedthe efficiency of Cy5.5 introduction by L-fucose-bound liposomes, suggesting L-fucose-receptor-mediated delivery. Intravenously injected L-fucose-bound liposomes carrying Cis-platin were successfully delivered to pancreatic cancer stem like cells, mediating efficienttumor growth inhibition as well as prolonging survival in mouse xenograft models. Thismodality represents a new strategy for pancreatic cancer stem cell-targeting therapy.
Tu1870
Pifithrin-μ, a New HSP70 and Autophagy Inhibitor, Can Enhance AntitumorEffects of TRAIL on Pancreatic CancerHiroyuki Monma, Mamoru Harada, Nanae Harashima, Eiji Hira, Yasunari Kawabata,Yoshitsugu Tajima
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) can induce apoptosisof various types of cancers with little cytotoxicity to normal cells through the stimulationof death receptors. In this study, after confirming that heat shock protein (HSP) 70 andautophagy play protective roles in antitumor effects of TRAIL, we examined a sensitizingeffect of pifithrin-μ, which is anHSP70 and autophagy inhibitor, on TRAIL-induced antitumoreffects on pancreatic cancer cell lines. TRAIL decreased the cell viability and induced apoptosisin three out of four cell lines tested. Regarding TRAIL-sensitive MiaPaca-2 and Panc-1,knockdown of HSP70 or Beclin-1, a key molecule for autophagy, with RNA interferencesignificantly enhanced TRAIL-induced antitumor effects, i.e., decreased the cell viability andcolony-forming capacity. Pifithrin-μ had the potential to induce both apoptosis and cellgrowth arrest. In addition, pifithrin-μ seemed to inhibit I κBα degradation in TRAIL-treatedcancer cells probably via inhibition of the autophagy-lysosome pathway, thereby resultingin inhibition of NF-κB activation in TRAIL-treated cancer cells. In a xenograft mouse model,combination therapy with TRAIL plus pifithrin-μ significantly inhibited MiaPaca-2 tumorgrowth compared to treatment with either agent alone. These results suggest that pifithrin-μ is a promising agent for use in therapies intended to enhance the antitumor effects ofTRAIL on pancreatic cancer.
S-868AGA Abstracts
Tu1871
Validation of MD Anderson Group B Borderline Resectable Pancreatic CancerTak Geun Oh, Moon Jae Chung, Seungmin Bang, Seung Woo Park, Jae Bock Chung, SiYoung Song, Jinsil Seong, Ho Kyoung Hwang, Chang Moo Kang, Woo Jung Lee, JeongYoup Park
BACKGROUND: Borderline resectable pancreatic cancer (BRPC) is categorized into the groupA, B, and C according to MD Anderson criteria. The group A indicates patients with tumorabutment of the visceral arteries or short-segment occlusion of the superior mesenteric vein.In the group B, patients have findings suggestive but not diagnostic of metastasis, such ascarcinomatosis, paraaortic lymph node metastasis. In this study we tried to validate whethergroup B of BRPC could be truly categorized as the borderline resectable group, and concurrentchemoradiotherapy (CCRT) could be a treatment option. STUDY DESIGN: Among thepatients who received preoperative CCRT, we defined patients with BRPC into the groupA or B. Rate of operation, recurrence rate, progression-free survival, and overall survivalwere compared between the group A and B. RESULTS: Between November 2005 and April2012, 53 patients with pancreatic adenocarcinoma were classified as borderline resectable.In the group A, 23 (60.5%) of 38 patients underwent pancreatectomy after CCRT, but, inthe group B, only 5 (33.3%) of 15 patients underwent pancreatectomy, mainly due to theprogression of suspected distant metastasis. There was significant difference in overall survivalbetween the group A and B patients (median OS; group A : 21.2 months vs group B : 10.2months, p=0.007). Of the patients who underwent pancreatectomy, the group B had higherrecurrence rate compared to the group A (recurrence rate; group A : 11 of 23 patients(47.8%) vs group B : 5 of 5 patients (100%), p=0.033). Also, of the patients who underwentpancreatectomy, the group B showed shorter disease-free survival compared to group A(group A : 29.0 months vs group B : 8.3 months, p=0.054). CONCLUSIONS: This is thefirst report to validate the definition of BPRC and indication for CCRT . We suggest thatthe group B BRPC should be considered as systemic diseases and treated with systemictherapy first rather than CCRT.Table 1. Comparison of survival outcome between group A and B borderline resectablepancreatic cancer for all of 53 patients
Table 2. Comparison of survival outcome between group A and B borderline resectablepancreatic cancer for 28 patients who had pancreatectomy
Tu1872
NF-κB Subunits Confer Divergent and Overlapping Functions in ApoptosisResistance of Pancreatic Cancer CellsClaudia Geismann, Frauke Grohmann, Susanne Sebens, Gabriele Wirths, Anita Dreher,Robert Haesler, Philip C. Rosenstiel, Günter Schneider, Sebastian Zeissig, Stefan Schreiber,Heiner Schäfer, Alexander Arlt
Pancreatic ductal adenocarcinoma (PDAC) represents one of the deadliest malignancieswith an overall life expectancy of six months despite palliative radio-chemotherapy. Thetranscription factor NF-κB has been shown to be a critical component of dysregulatedtranscription factor activity conferring profound apoptosis resistance. Despite extensive dataon the role of the most abundant NF-κB subunit p65/RelA in PDAC apoptosis control, thereis only little knowledge on the role of the other Rel subunits. In the present study, threepancreatic carcinoma cell lines (Panc1, Patu8988, MiaPaca2) were analyzed for the role ofRelA, RelB and c-Rel in resistance against TRAIL, etoposide or gemcitabine induced apoptosis.Resistant Panc1 and Patu8988 cells exhibit a high basal NF- κB activity that was furtherstrongly induced by TRAIL and moderately upregulated by etoposide or gemcitabine. Incontrast, sensitive MiaPaca2 cells displayed only little basal NF- κB activity and only weakinduction by TRAIL, etoposide or gemcitabine. Transfection with siRNA against the RelBor c-Rel subunits of NF-κB sensitized the TRAIL and chemotherapy resistant Panc1 andPatu8988 cells in a comparable fashion like siRNA targeting the p65/RelA subunit. Gel shiftanalysis revealed that, together with the p65/RelA subunit, c-Rel is part of the TRAIL,etoposide or gemcitabine inducible NF-κB complex in PDAC. In addition the basal NF- κBcomplex, mediating resistance against chemotherapeutic drug induced apoptosis, is com-posed of all three subunits as revealed by Shift-Western analysis. Western-Blot-analysis ofknown anti-apoptotic NF-κB target genes (e.g. Bcl-2 family and cIAPs) and mRNA-arraybased expression analysis revealed a divergent non overlapping function of the Rel NF- κBsubunits in established and novel anti-apoptotic NF- κB target genes. Furthermore, c-Relelicits a proapoptotic function in the sensitive MiaPaca2 cell line as indicated by an increasein the apoptotic response in cells transfected with c-Rel specific siRNA. In conclusion, all
three Rel-subunits are critical mediators of NF- κB dependent anti-apoptotic signalling inPDAC by subunit specific regulation of specific target genes.
Tu1873
Systemic Delivery of Fiber Redesigned Infectivity-Selective OncolyticAdenovirus for Pancreatic CancerYoshiaki Miura, Joohee Han, Julia Davydova, Masato Yamamoto
Systemic therapies for cancer, including pancreatic cancer, have provided only limited efficacyfor patients to date. Adenovirus (Ad) has frequently been used as a backbone of oncolyticviral agents. However, unlike loco-regional therapy, systemic application of cancer genetherapy mandates different level of selectivity of gene delivery. So far, insufficient cancerselectivity at the stage of infection has greatly hampered efficacy of Ad-based therapy uponsystemic administration. The controlled distribution and selective transduction of the vectorwould overcome the obstacles for systemic delivery and enable efficient systemic treatmentof cancer. We have recently developed transductionally-targeted Ads by high-throughputscreening of high-diversity (10^9-level) Ad-library. Mesothelin (MSLN) is highly over-expressed in pancreatic cancers. Ad with redesigned AB-loop targeted to MSLN (AdML-VTIN)was successfully isolated from the high throughput screening of Ad library by infectivityfor MSLN-expressing 293 cells (293-MSLN). The in vitro binding of AdML-VTIN corre-sponded to MSLN-expression of each cell line, and the suppression of MSLN-expressionwith siRNA or antibody blocked the binding to MSLN-expressing cells, which indicates thevector is targeted to mesothelin. In MSLN-positive cells (Panc-1), this virus exhibited higherbinding ability than that of Ad with 5/3 modified fiber which is known to exhibit highestinfectivity in pancreatic cancer cells. The in vivo antitumor effect was compared in Panc-1(MSLN-positive) and MiaPaCa-2 (MSLN-negative) subcutaneous xenografts. The injectionof the MSLN-retargeted oncolytic Ad showed significant antitumor effect against Panc-1tumors, and disappearance of tumors was observed in 4 out of 8 mice. Contrarily, the samevirus showed no antitumor effect in MiaPaCa-2. Viral DNA amount in the tumors correlatedto the anti-tumor effect. Upon systemic administration in the nude mice with Panc-1subcutaneous xenografts, AdML-VTIN showed significantly lower liver sequestration com-pared to the wild-type Ad5, and the copy number of this retargeted virus was significantly(1000-fold) higher in the tumor. These data indicates that this vector enables efficientsystemic delivery and treatment of MSLN-expressing pancreatic cancer xenografts. In thisstudy, our MSLN-targeted Ad vector exhibited selective infectivity to MSLN-positive pan-creatic cancer cells in vitro and in vivo. Moreover, this virus showed dramatically augmenteddelivery to the tumors and significantly reduced liver sequestration upon systemic administra-tion. This new genetically modified Ad, which is transductionally retargeted to pancreaticcancer, may embody a next generation systemic therapy for this devastating disease.
Tu1874
Earlier Age of Presentation of Pancreatic Adenocarcinoma in Individuals WithChronic PancreatitisArpan Mohanty, Nilesh H. Shah, Michelle A. Anderson, Aaron R. Sasson, Kristen J. Vogel,Simon Sherman, David C. Whitcomb, Randall Brand
BACKGROUND: Pancreatic cancer (PC) screening is reserved for the subset of the generalpopulation at high risk for PC. Patients with chronic pancreatitis (CP) are at increased riskfor PC; a recent meta-analysis reported a relative risk of 13.3 [Best Pract Res Clin Gastroent-erol. 2010 Jun;24(3)]. The exact mechanism that links the two diseases is unknown butmay be caused by a shared exposure to risk factors such as cigarette smoking and alcoholconsumption and/or facilitation of tumor initiation and promotion by inflammation. Wehypothesized that if the latter mechanism was true, the inflammatory state induced fromCP would lead to an earlier age of onset of PC development. The goal of this study was tocompare the age of onset and stage of PC in patients with a history of CP to those withoutCP. METHODS: Prospective data on 660 individuals with PC from themulti-center PancreaticCancer Collaborative Registry was reviewed for a history of CP. Identified CP cases werematched with controls based on gender, alcohol use and tobacco use. Propensity scorematching was used to match, with replacement, each case with its 3 closest controls. Tobaccouse was classified as non-smoker; ongoing use of tobacco: cessation of tobacco use , 10years prior to diagnosis of PC; and tobacco use . 10 years prior to diagnosis of PC. Alcoholusers were classified as past or current users of alcohol or non-users. When more than 3controls were an exact match (ties) for a particular case, the ties were all used and weightedequally. The average treatment effect on the treated (ATT) for age of PC diagnosis and PCstage were calculated. RESULTS: A total of 16 CP patients from the 660 PC cases wereidentified. 56% of CP patients were males. After matching, CP patients were diagnosed 4.9years (95% CI: 0.6-9.2 years) earlier than controls (p=0.025). There was no statisticaldifference in PC stage (I/II vs. III/IV) between control and CP cases. CONCLUSIONS: Ahistory of CP appears to have an independent effect on age of onset of PC with a ~5 year earlierdiagnosis, supporting the role of inflammation in PC carcinogenesis. A lack of difference inPC stage, suggests that this result is not related to more frequent abdominal imaging in theCP group. Findings from this study should be factored into any future PC surveillancerecommendations for CP patients. Further studies are warranted to determine if shared geneticsusceptibility factors involving inflammatory pathways are present in CP and PC patients.
Tu1875
Fibroblast Activation Protein (FAP) Expression of Cancer-AssociatedFibroblast Predicts Poor Prognosis in Patients With PancreaticAdenocarcinomaTomoya Kawase, Koji Yoshida, Keisuke Hino
[Background and Aim] In many solid tumors, the stroma is increasingly recognized to beimportant in promoting tumor proliferation, invasion, metastasis, and chemoresistance.Especially, several studies using mouse pancreatic ductal adenocarcinoma(PDAC)modelrevealed that fibroblast activation protein(FAP) played an important role in tumorgenesis
S-869 AGA Abstracts
by reducing the surrounding immunity. FAP, or seprase, is a 170kDa dimer surface glycopro-tein and expressed on cancer associated fibroblasts(CAF). The aim of this study was toinvestigate the relationship between FAP expression of CAF and the prognosis in patientswith PDAC. [Materials and Methods] During 2008-2012 paraffin-embedded specimens ofPDAC were collected from 50 patients under the approval by the ethics committee of theinstitution. Standard immunohistochemical detection of FAP was performed using anti-FAPprimary antibody. The expression levels of FAP were classified into 4 degrees (negative, 1+,2+, 3+) and compared with patients' clinical course. [Results] FAP of CAF was detected in92%(46/50) of specimens and its expression level was 1+ in 16, 2+ in 19 and 3+ in 11specimens. The expression level of FAP was negatively correlated with overall survival ofpatients (p,0.001). Furthermore multivariate analysis showed FAP expression level, clinicalstaging of pancreatic cancer extracted as significant factors associated with overall survival.[Conclusion] The present results indicated that FAP expression of CAF is one significantfactors associated with overall survival of patients with pancreatic ductal adenocarcinoma.
Tu1876
Feasibility of Pathway Analysis in EUS-FNA Obtained Tissue of PancreaticCancer Patients: A Step Towards Personalized Based MedicineWesley K. Utomo, Vilvapathy Narayanan, Katharina Biermann, Casper H. van Eijck,Marco J. Bruno, Maikel P. Peppelenbosch, Henri Braat
Introduction: Pancreatic cancer (PC) remains one of the most lethal common malignanciesworldwide. The five year survival of all stages combined is only 5%. Deregulation of severalcore signaling pathways have been identified in PC and distinct pathways are affectedbetween different patients. Hence, personalized based medicine is an attractive approach inthe treatment of PC. Here, we aim to study the feasibility of pathway analysis using tissueobtained by Endoscopic Ultrasound-Fine Needle Aspiration (EUS-FNA); the activation ofmammalian target of Rapamycin (mTOR) in specific by the measurement of phosphorylatedS6 (pS6), a downstream protein of the mTOR pathway. Experimental design: Immunohisto-chemical (IHC) staining of pS6 on Formalin Fixed Paraffin Embedded (FFPE) material fromPC resection specimens was performed. Forty nine slides(33 tumor regions and 36 healthyregions) were stained and scored by an expert pathologist. To study mTOR activation andinhibition by rapamycin(Rap), PC cell lines (BxPC3, SU8686) were analyzed by means ofwestern blot (cleaved caspase 3, to measure apoptosis, and pS6), cell viability assays andflow cytometry (pS6). Single cell suspensions obtained through EUS-FNA of pancreatictumors were stained for cytokeratin 8/18 (CK8/18) and pS6 expression and subsequentlyanalyzed using flow cytometry. Results: The variability of IHC stained FFPE material wascompared by means of the coefficient of variation of 63.73% in tumor regions (n=33)compared to 35.69% in healthy regions (n=36). Levels of pS6 were 6-30 fold lower after 2hours of incubation with 0.1 μM Rap. Cell viability were 43% and 38% after 3 days of 12.5μM Rap. On flow cytometry analysis of cell lines, Rap reduced the Mean FluorescenceIntensity (MFI) in CK8/18 positive cells of pS6 by 75%(BxPC3) and 80%(SU8686). Validationin fresh EUS-FNA tissue by flow cytometry analysis showed higher expression of pS6 in 2out of 6 (33%) confirmed PC. MFI reductions of 80% and 83% of pS6 in CK8/18 positivepopulations were observed after incubation with 0.1 μM Rap. Absolute numbers of CK8/18positive, pS6 positive cells also decreased by 98% and 86.4%. Conclusions: As hypothesized, agreat variation in the expression of pS6 exists between PC patients in FFPE material. Thisfinding might indicate a possible subpopulation with higher activation of the mTOR pathwayin PC patients. In PC cell lines, Rap reduced cell viability in a dose dependent manner andinhibits pS6 expression without leading to apoptosis. Validation in EUS-FNA tissue showeda marked reduction of pS6 expression after incubation with Rap. The results of this studyshow the feasibility of pathway analysis in EUS-FNA tissue which is a first step towardspersonalized based medicine in PC. Further studies in EUS-FNA tissue are being undertakento determine other activated signaling pathways in PC.
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Changes in Vascularization and Innervation Patterns in PanIN Lesions AsRevealed by Novel Three-Dimensional Imaging TechniquesYa-Yuan Fu, Jennifer M. Bailey, Janivette Alsina, Subhash Kulkarni, Shiue-Cheng Tang,Steven D. Leach, Pankaj J. Pasricha
Background & Aims: Pancreatic Intraepithelial Neoplasia (PanIN) is a pancreatic cancerprecursor lesion. Due to the opacity of pancreas, microtome-based planar microscopy isa routinely used method to observe PanIN lesions. However, this technique limits ourunderstanding of the morphology and development of PanIN and pancreatic cancer byrestricting tissue visualization in a 2-dimensional (2-D) space, rather than rendering a true3-dimensional (3-D) image of an intact developing PanIN. In order to identify and generatethe intact 3-D image of PanIN for facilitating better and early diagnosis of pancreatic cancer,we aimed to (1) apply Ulex europaeus agglutinin I (UEA-I) as a new cell surface marker tospecifically label the PanIN lesion, (2) characterize the altered vascular and neural networksby vessel painting the pancreatic tissue with wheat germ agglutinin (WGA) and immunostain-ing, and (3) combine the fluorescent staining with 3-D microscopy to observe the nativePanIN lesions at a subcellular level. Methods: We used two independent models of murinePanIN. The first involves the expression of Kras from the progenitor cells of the pancreas:Pdx-1:Cre; LsL-KrasG12D mice (9 months old). The second involves expression of Krasfrom adult acinar cells: Mist1:CreERT2;LsL-KrasG12D;mTmG mice (analyzed 6 weeks afteradministration of tamoxifen), and control mice (Mist1:CreERT2;mTmG). The mice wereperfused with fluorescent conjugated WGA to reveal the blood vessels of pancreas. Theinnervation was labeled by immunostaining (using pan-neuronal marker PGP9.5). We usedtwo independent methods to label PanIN. First, the mTmG background enables the visualiza-tion of GFP+ cells expressing KrasG12D. Second, UEA-I was used as a novel cell surfacemarker to label the PanINs. The labeled tissues were treated with the optical-clearing solutionFocusClear (Gastroenterology, 139:p1100-1105, 2010) to prepare transparent specimensfor confocal microscopy. Results: Comparisons of staining for various lectins revealed UEA-I to be a specific label for PanIN lesions. This allowed us to develop robust 3-D visualizationof PanIN lesions and their relationship to blood vessels and nerves. We discovered anincrease in vessel density in PanIN-bearing mice relative to normal pancreas. At the same
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