Role of Cathepsin B gene single nucleotide polymorphisms in tropical chronic pancreatitis in Indian population

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Conclusion: Our study confirmed that IL-6, IL-8, IL-10, and sTNFr measured on admission, and CRP and pancreatic elastase measured on third day of admission represent valuable prognostic factors in the determination of severity in patients with AP.

in patients with SUVmax < 6.0 and 26.3% and 13.5% in patients with SUVmax
6.0 (p<0.001). Cox proportional model revealed that SUVmax
6.0 (odds ratio (OR): 2.52, p¼0.001), lymph node metastases (OR: 2.41, p¼0.023), and poor differentiation (OR: 1.71, p¼0.027) were significantly correlated with poor survival. Conclusions: SUVmax >6.0 was a significant predictor of postoperative early recurrence and subsequent poor survival after resection of pancreatic adenocarcinoma.

P228. Neoadjuvant chemoradiotherapy with S-1 in patients with borderline resectable pancreatic cancer with involvement of the major artery M. Suenaga 1, T. Fujii 1, S. Yamada 1, N. Okumura 1, D. Kobayashi 1, C. Tanaka 1, G. Nakayama 1, H. Sugimoto 1, M. Koike 1, S. Nomoto 1, M. Fujiwara 1, S. Takeda 1, A. Nakao 2, Y. Kodera 1. 1 Department of Gastroenterological Surgery (Surgery II), Nagoya University Graduate School of Medicine, Nagoya, Japan 2 Department of Surgery, Nagoya Central Hospital, Nagoya, Japan Background: The aim of this study was to investigate the efficacy and safety of neoadjuvant chemoradiotherapy with S-1 (oral fluoropyrimidine) followed by surgery for the treatment of borderline resectable pancreatic cancer that involved the major visceral artery. Patients and methods: Ten patients with pancreatic cancers that abutted the superior mesenteric artery in five and the common hepatic artery in five were treated with neoadjuvant chemoradiation at a single institution. Radiation therapy was delivered at a total dose of 50 Gy in 25 fractions. S-1 was administered orally at a dose of 80 mg/m(2)/day for 14 consecutive days followed by a 7-day rest period during radiation therapy. After radiotherapy and 2 courses of S-1, restaging was done to evaluate secondary resectability. Results: Of the ten patients, nine underwent a full course of neoadjuvant therapy and chemotherapy terminated in one patient because of grade 3 leukopenia. Partial response was achieved in three patients and stable disease in seven. Seven (70%) underwent surgical resection, and all had margin-negative (R0) resections. Pathological examination revealed that more than 50% of tumor cells had disappeared in all cases. Conclusions: Neoadjuvant chemoradiation with S-1 was feasible and promising therapy for borderline resectable pancreatic cancer that involves the major artery.

P229. The FDG-PET predicts postoperative early recurrence and poor prognosis after resection of pancreatic adenocarcinoma T. Sugiura, T. Yamamoto, Y. Okamura, T. Mizuno, H. Kanemoto, K. Uesaka. Division of Hapato-Biliary-Pancreatic Surgery, Shizuoka Cancer Center Hospital, Shizuoka, Japan Background: 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) has been acknowledged for its utility in staging or recurrence diagnosis of pancreatic adenocarcinoma. However, the clinical utility of the FDG-PET as a prognostic predictor of the pancreatic adenocarcinoma has not been fully evaluated. Methods: Between October 2002 and May 2010, a total of 114 patients received FDG-PET examination within 30 day before surgery. The maximum standard uptake value (SUVmax) was calculated for each primary lesion. A receiver operating characteristics (ROC) curve was constructed to estimate the optimal cutoff value of SUVmax. To determine which outcome of interests were appropriately demonstrated, ROC curve analyses were conducted on the six outcomes; 6-, 12-, and 24-months disease-free survival (DFS), 6-, 12-, and 24-months overall survival (OS). Multivariate analysis was conducted to evaluate the prognostic factor. Results: The ROC curves revealed that the SUVmax predicted the 6month DFS the most optimally (AUC: 0.757) with a cutoff value of 6.0. Of 63 patients with SUVmax
6.0, 34 (54%) had recurrence within 6 months. In contrast, only 3 of 51 (6%) with SUVmax <6.0 developed an early recurrence (p<0.001). The 3- and 5-year DFS rates were 44.7% and 44.7% in patients with the SUVmax < 6.0 and 14.0% and 14.0% in patients with SUVmax
6.0 (p<0.001). The 3- and 5-year OS rates were 55.2% and 51.5%

P230. Descriptive statistics for the endoscopic pancreatic function test: Defining normal S.L. Suleiman, V. Kadiyala, S.J. Burton, L.S. Lee, P.A. Banks, D.L. Conwell. Center for Pancreatic Disease, Brigham and Women's Hospital, Boston, MA, USA Background: The traditional secretin-stimulated Dreiling tube pancreas function test evaluates duct cell function and is the surrogate gold standard for diagnosis of chronic pancreatitis. The endoscopic pancreatic function test (ePFT) is being more widely adopted; however, the need for sedation may affect duct cell function and confound ePFT results. Aims: 1. Describe the ePFT electrolyte profile of healthy subjects. 2. Compare peak bicarbonate concentration ([HCO3]) in ePFT with sedation to that in the non-sedated Dreiling tube method. Methods: Healthy subjects (HS) with no personal or family history of pancreatic disease, no comorbities, non-smoker and alcohol intake <2 drinks/day were recruited. Subjects underwent the previously published 60-minute ePFT procedure with conscious sedation. Pancreatic fluid [Na], [Cl], [K] and [HCO3] were analyzed. Descriptive statistics, correlation and ttest were performed (SPSS Statistics v20). Results: 10 HS (5 male) of mean age 28.56 and mean BMI 244 underwent ePFT with secretin (14.8 3.6mg; 10.9-21.6mg), midazolam (7.61.9mg; 5-10mg) and fentanyl (19556.3mg; 125-300mg). [Na] and [K] were relatively stable, while [HCO3] and [Cl] exhibited an inverse relationship over the 1 hour procedure (Pearson's r¼-0.93, p¼0.022). The mean peak [HCO3] was 100.910.2 mEq/L, range 84-120mEq/L and IQR 94.5-105mEq/L. The 2SD lower limit of normal peak [HCO3] was 80.5 mEq/ L. Mean nadir [Cl] was 70.95 mEq/L, range 63-81mEq/L, median 71.5mEq/ L and IQR 68.8-72.8mEq/L. The ePFT mean peak [HCO3] was not statistically different (p¼0.14) to that of the non-sedated Dreiling tube test (968 mEq/L, 2SD lower limit of normal 80mEq/L; Somogyi 2003). Conclusion: 1. The ePFT mean peak [HCO3] in our HS cohort agrees with published normal values and the 80mEq/L diagnostic cutoff determined by the non-sedated Dreiling tube method. 2. Standard doses of conscious sedation do not appear to impact duct cell secretory function.

P231. Role of Cathepsin B gene single nucleotide polymorphisms in tropical chronic pancreatitis in Indian population S. Sundaresan 1, 2, S. Yuvaraj 1, R. Dawra 2, A.K. Dutta 1, A.J. Joseph 1, A.K. Saluja 2, A. Chacko 1. 1

Molecular Medicine Unit, Division of Gastrointestinal Sciences, Christian Medical College, Vellore 632 004, Tamilnadu, India 2 Basic & Translational Research, Division of Surgery, University of Minnesota, Minneapolis, MN, USA Background and aims: Tropical Calcific Pancreatitis (TCP) is another type of chronic pancreatitis unique to developing tropic countries India, Sri lanka and in other Asian countries with unknown origin. Majority of TCP patients are associated with SPINK mutation (N34S). The mechanism responsible for the intrapancreatic activation of trypsinogen to trypsin

Abstracts / Pancreatology 13 (2013) e1–e94

leading to pancreatitis is still controversial. Cathepsin B (CTSB) is known to activate cationic trypsinogen. Hence we aimed to evaluate the single nucleotide polymorphism in CTSB gene of TCP patients in Indian population. Subjects and method: We sequenced the coding and flanking intronic region of CTSB gene in 113 patients and 130 controls. The Polymerase Chain Reaction (PCR) product was purified and then sequenced by applied biosystems capillary automated sequencer (Applied Biosystem, Boston). Appropriate statistical analyses were performed and data was analysed by using SPSS for Windows Version 20. Results: We found six novel polymorphisms in the sequenced portion of the CTSB gene, which is associated with TCP. In Intron 2 mutant allele C12T has higher frequency in controls vs TCP patients (p¼0.004). In Exon 3 no significant differences in both allele and genotype frequency between patients vs control was observed, whereas in Intron 3 G265T heterozygous and mutant form increased in TCP by factor of 10.8. T66C mutation in both heterozygote and homozygote increased the disease odds to 1.9. In Exon 4 the mutant allele at T774C was 13.5% in TCP patients but none in the control (p¼0.0001). However in Intron 4 the polymorphism G834C showed significant difference between patients vs controls. Conclusion: This data indicates the lack of association of Leu26Val and other polymorphisms in CTSB gene in the eastern and south Indian population.

P232. Bile acid induced injury to human acinar cell does not involve Gpbar1 activation S. Sundaresan, H. Zhu, S. Kumar, R. Dawra, A. Saluja. Division of Basic and Translational Research, Department of Surgery, University of Minnesota, Minneapolis MN, USA Biliary pancreatitis accounts for majority of the cases of pancreatitis, but mechanism of the disease is not clear. Based upon the experimental animal models, it is believed that bile acids exposure due to influx of bile in pancreatic duct can cause injury to pancreatic acinar cells through a specific bile acid receptor (Gpbar1or TGR5). Because of paucity of human pancreas for experimental studies, there is no information about the mechanism of bile acids induced injury in human pancreatic acinar cells. Aim of this study was to understand the mechanism of bile acids induced injury in human pancreatic acinar cells. Methods: Human acinar cells prepared by collagenase digestion during islet isolation from healthy pancreases were used for this study. Injury in response to bile acid exposure was evaluated by trypsin activity measurement after exposure to bile acids or supramaximal concentrations of carbamyl choline chloride. Bile acid receptor presence in human acinar cells was studied by immunoprecepitation and western blotting. Receptor mediated effect was blocked by pretreatment with a specific antibody or use of a phospholipase C inhibitor. Results: Western blot analysis of the protein extract from human pancreatic acinar cells revealed presence of recently reported bile acid receptor (Gpbar1). Sodium taurolithocholate-3- sulfate (TLCS) only at high concentration (500mM) caused trypsin activation, while there was no effect at low concentration (50mM). Pretreatment of human acinar cells with a specific antibody for Gpbar1 didn't alter carbamyl choline induced trypsin activation but reduced TLCS (500mM) induced trypsin activation. Pretreatment of human acinar cells with phospholipase C inhibitor U73122 completely inhibited carbamyl choline elicited trypsin activation but reduced sodium taurolithocholate induced trypsin activation. Preincubation of human acinar cells with a low concentration of TLCS (50mM), followed by carbamyl choline (0.1mM) significantly enhanced trypsin activation as compared to carbamyl choline alone and this effect was attenuated by pretreatment with Gpbar1antibody. Conclusion: Bile acid receptor (Gpbar1) is present on human pancreatic acinar cells. Stimulation of this receptor using TLCS (50mM) doesn't cause trypsin activation but enhances response of carbamyl choline. Blocking this receptor using either a specific antibody or pretreatment with a phospholipase C inhibitor reduced TLCS (500mM) induced trypsin activation. Blocking of the receptor using antibody also attenuated the

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enhanced trypsin activation observed by subsequent carbamyl choline stimulation. In human acinar cells stimulation of Gpbar1 alone doesn't cause injury.

P233. Regulation of pluripotency markers by DCLK1 in pancreatic cancer S.M. Sureban 1, 2, R. May 1, 2, D. Qu 1, N. Ali 1, C.W. Houchen 1, 2. 1 Department of Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA 2 Veterans Affairs Medical Center, Oklahoma City, OK, USA Background: It has been shown that OCT4, SOX2, c-Myc, LIN28, Nanog and KLF4 are required for embryonic stem cell self-renewal and pluripotency and are upregulated in some aggressive cancers and cancer stem cells (CSCs). miR-145 is downregulated in various cancers and has been shown to repress OCT4, SOX2, and KLF4, thereby repressing pluripotency and tumorigenesis. DCLK1, a putative pancreatic stem cell marker is upregulated in pancreatic cancer and siRNA mediated knockdown of DCLK1 resulted in the inhibition of several oncogenes, including c-Myc, KRAS and Notch1 via miRNA related mechanisms. In this study, we aim to determine whether DCLK1 regulates factors controlling pluripotency via miR-145. We have utilized Nanoparticle (NP) technology to deliver DCLK1 specific siRNA (NP-siDCLK1). Methods: Total RNA isolated from human pancreatic tumor xenografts (AsPC-1), following treatment with NP-siDCLK1, was subjected to realtime RT-PCR for mRNA (DCLK1, OCT4, SOX2, LIN28, Nanog and KLF4) and miRNA (pri-miR-145 and pri-miR-143/145 cluster). Additionally, AsPC-1 cells were transfected with a plasmid encoding the firefly luciferase gene with a miR145 binding site at the 3’UTR to measure miR-145 expression. Results: NP-siDCLK1 treatment resulted in AsPC-1 tumor xenograft growth arrest, downregulation (w50%) of DCLK1, OCT4, SOX2, LIN28, Nanog and KLF4 mRNA with a related >2-fold increase in pri-miR-145 and pri-miR-143/145 cluster miRNA. This increase was confirmed by a corresponding reduction in miR-145 specific luciferase activity in AsPC-1 cells treated with NP-siDCLK1. Conclusions: These data taken together demonstrate a potential regulatory role for DCLK1 in the expression of pluripotency factors in pancreatic cancers via miR-145 miRNA. Therefore DCLK1 may represent a novel target for anti-cancer stem cell based strategies for pancreatic cancer and perhaps other solid tumors.

P234. The long-term survivors after curative resection for the pancreatic cancer should be paid attention to the remnant pancreatic cancer S. Suzuki 1, T. Hatori 1, A. Kimijima 1, N. Oshima 1, T. Furukawa 2, Y. Kuboki 3, K. Shimizu 3, K. Shiratori 3, M. Yamamoto 1. 1 Department of Gastroenterological Surgery, Tokyo Women's Medical University, Tokyo, Japan 2 Institute for Integrated Medical Sciences, Tokyo Women's Medical University, Tokyo, Japan 3 Department of Gastroenterology, Tokyo Women's Medical University, Tokyo, Japan Background: The remnant pancreatic cancer sometimes appears in the long-term survivors after curative resection for the invasive ductal adenocarcinoma of the pancreas (PDAC). The aim of this study was to evaluate the management of the remnant pancreatic cancer in the long-term survivors of the PDAC. Methods: 146 patients survived more than three years after curative resection (R0) for the PDAC between 1980 and 2008. A group of 16 patients who were treated for metachronous remnant pancreatic cancer were reviewed retrospectively. Results: At the first surgery, the tumor was located in the pancreatic head with 9 patients and in the pancreatic body with 7 patients. PPPD was performed in 5 patients, PD in 3 patients, DPPHR in one and DP in 7. A well