40.19 Effect of HO-3867, a novel curcumin analog, on cholangiocarcinoma

Posted on January 18, 2018

H. Kazik1, S. Kunnimalaiyaan1, T. Gamblin1, M. Kunnimalaiyaan1  1Medical College Of Wisconsin,Surgical Oncology/Surgery,Milwaukee, WI, USA

Introduction: Cholangiocarcinoma (CCA) is rare, but lethal bile duct cancer which remains difficult to diagnose and treat. Currently, systemic therapies and surgery remain the viable solutions. However, CCA responds poorly to these therapies and only a few patients are candidates for potentially curative surgical resection. Therefore, an urgent need for the identification of molecularly targeted compound(s) remains. HO-3867, a curcumin analog, has shown efficacy as an inhibitor of signal transducer and activator of transcription 3 (STAT3) in ovarian cancer cell proliferation but the effects of HO-3867 in other cancer types including CCA is unknown. Therefore, the objective of this study is to test the hypothesis that HO-3867 will effectively inhibit CCA cellular proliferation.

Methods: Effect of HO-3867 (0-10 µM) on two human CCA cell lines, CCLP-1 and CC-SW-1 cellular proliferation was measured through MTT assay, colony forming ability, and real time confluency assay. Cell lysates were analyzed via western blot to determine the effect of HO-3867 on STAT3 phosphorylation as well as the levels of pro and anti-apoptotic proteins.

Results:Treatment with HO-3867 significantly reduced cellular proliferation, colony formation, and cell confluency in a dose- and time-dependent fashion in both cell lines. Following treatment with HO-3867, induction of apoptosis was evident in CCLP-1 through an increase in pro-apoptotic marker (cleaved poly ADP ribose polymerase) and a decrease in anti-apoptotic marker (cyclin D1 and survivin). HO-3867 treatment reduced the levels of phosphorylation of STAT3.

Conclusion:HO-3867 significantly reduced the growth of both CCLP-1 and CC-SW-1. To our knowledge, this is the first study on the effect of HO-3867 in CCA cell lines in vitro and provides rationale for further preclinical analysis.

 

40.17 Human Biliary Tract Cancer Contains High Levels of S1P

Posted on January 18, 2018

Y. Hirose1, M. Nagahashi1, K. Yuza1, K. Miura1, J. Sakata1, T. Kobayashi1, H. Ichikawa1, Y. Shimada1, H. Kameyama1, K. Takabe2,3, T. Wakai1  1Niigata University Graduate School Of Medical And Dental Sciences,Division Of Digestive And General Surgery,Niigata, NIIGATA, Japan 2Roswell Park Cancer Institute,Breast Surgery, Department Of Surgical Oncology,Buffalo, NY, USA 3University At Buffalo Jacobs School Of Medicine And Biomedical Sciences, The State University Of New York,Department Of Surgery,Buffalo, NY, USA

Introduction: Biliary tract cancer, including gallbladder cancer, intra- and extrahepatic bile duct cancer, is one of the most lethal diseases among gastrointestinal cancer. Previous studies have suggested that chronic inflammation is involved in the disease process of biliary tract cancer. Sphingosine-1-phosphate (S1P) has emerged as a pleiotropic sphingolipid mediator that regulates many cellular functions associated with inflammation and cancer. Indeed, it has been previously suggested that S1P plays important roles in bile duct cancer progression in experimental model. However, the roles of S1P in human biliary tract cancer has yet to be clarified. The aim of this study is to determine the levels of sphingolipids including S1P and their metabolites in biliary tract cancer tissues and normal biliary tract tissues, and to clarify the difference in the levels of each sphingolipid between the two tissues.

Methods: We examined biliary tract cancer tissues (gallbladder cancer, N=5; intrahepatic bile duct cancer, N=2; and extrahepatic bile duct cancer, N=8) and normal biliary tract tissues (normal gallbladder mucosa, N=5; normal extrahepatic bile duct mucosa, N=12) in patients with biliary tract cancer. Sphingolipids including S1P and their metabolites of dihydro-S1P (DHS1P), sphingosine (Sph), and dihydro-Sph (DHSph) were quantified by liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). The levels of each sphingolipid were compared between cancer tissues and normal tissues using the Mann-Whitney U test. All tests were two-sided and P<0.05 were considered statistically significant.

Results:In the comparison between all biliary tract cancer tissues (N=15) and all normal biliary tract tissues (N=17), the levels of S1P, DHS1P, Sph, and DHSph were significantly higher in cancer tissues than normal tissues (S1P, P=0.004; DHS1P, P=0.030; Sph, P=0.011; DHSph, P<0.001). In the comparison between intra- and extrahepatic bile duct cancer tissues (N=10) and normal bile duct mucosa (N=12), the levels of S1P, DHS1P, Sph, and DHSph were significantly higher in cancer tissues than normal tissues (S1P, P=0.004; DHS1P, P=0.030; Sph, P=0.011; DHSph, P<0.001). In the comparison between gallbladder cancer tissues (N=5) and normal gallbladder mucosa (N=5), the levels of DHSph were significantly higher in cancer tissues than normal tissues (P=0.016), but there were no significant difference in the levels of S1P, DHS1P and Sph.

Conclusion:To our knowledge, this is the first study to reveal the levels of sphingolipids including S1P in human biliary tract cancer patients by mass spectrometry. The high levels of sphingolipids in the cancer tissue may indicate the important roles of S1P in disease process of biliary tract cancer in human patients.

 

40.16 Possible Role of HOX genes in Pancreatic Ductal Adenocarcinoma Survival

Posted on January 18, 2018

K. Takabe1, E. Katsuta1, L. Yan2  1Roswell Park Cancer Institute,Breast Surgery, Department Of Surgical Oncology,Buffalo, NY, USA 2Roswell Park Cancer Institute,Department Of Biostatistics And Bioinformatics,Buffalo, NY, USA

Introduction:  Pancreatic adenocarcinoma (PDAC) is one of the most aggressive cancers with severe prognosis in general; however, we sometimes encounter exceptional long term survivors.  Although there have been many study to explore the genes that are responsible for these differences in prognosis of PDAC, it remains unknown. The HOX genes are a family of homeodomain-containing transcription factors that determine cellular identity during development. This study aimed to investigate the clinical relevance of HOX genes on the PDAC patient survival. 

Methods:  RNA-sequencing and clinical data were obtained from the Cancer Genome Atlas (TCGA) through cBioportal. Gene expression was compared between patients who survived more than 5 years (good prognosis group) and patients who died within 3 years of diagnosis (poor prognosis group). Patients whose cause of death was not pancreatic cancer in bad prognosis group were excluded for the analysis. The cut-off value of gene expression for survival analysis was determined by automated scanning and selecting the threshold yielding the lowest p-value for the each gene.

Results: Among pancreatic cancer TCGA cohort, 94 PDAC patients were confirmed of their date of death or lived longer than 5 years. 4 and 82 patients were classified as good and poor prognosis group, respectively. 1 and 3 patients were diagnosed as Stage IIA and IIB in good prognosis group, 1, 5, 9, 62, 2 and 3 patients were diagnosed as Stage IA, IB, IIA, IIB, III and IV in poor prognosis group, respectively. 120 genes were extracted as differentially expressed gene with adjusted p<0.05. Among 120 genes, 8 genes were HOX genes, including HOXA9 (logFC=7.18, adj. p<0.001), HOXA7 (logFC=3.26, adj. p<0.001), HOXA4 (logFC=2.18, adj. p<0.001), HOXA6 (logFC=5.38, adj. p<0.001), HOXA5 (logFC=3.85, p<0.001), HOXA10 (logFC=4.19, adj. p=0.001), HOXA2 (logFC=1.75, adj. p=0.036) and HOXA13 (logFC=2.87, adj. p=0.039). All of them were upregulated in good prognosis group. Then we compared the survival based upon HOX expression in whole 154 PDAC in TCGA cohort. High expression of HOXA2 (median OS: 19.8 month vs 16.8 month, p=0.005, median DFS: 16.9 month and 9.6 month, p=0.001), HOXA4 (median OS: 20.6 month vs 12.5 month, p<0.001, median DFS: 17.1 month and 9.5 month, p<0.001), and HOXA5 (median OS: 19.9 month vs 12.5 month, p<0.001, median DFS: 17.1 month and 9.5 month, p=0.001) showed significantly better both OS and DFS. 

Conclusion: This is the first report that high expression of HOX genes associate with exceptional long term survivor in PDAC. Further studies are warranted to investigate the mechanism how these gene expressions contribute to patient survival.

40.15 γH2AX Expression Discriminates Between Benign and Malignant Biliary Tract Biopsy Specimens

Posted on January 18, 2018

Y. Hirose1, M. Nagahashi1, H. Ishikawa1, K. Yuza1, K. Miura1, H. Ichikawa1, Y. Shimada1, H. Kameyama1, J. Sakata1, T. Kobayashi1, T. Wakai1  1Niigata University Graduate School Of Medical And Dental Sciences,Division Of Digestive And General Surgery,Niigata, NIIGATA, Japan

Introduction: Preoperative discrimination between benign and malignant biliary structures based on small biopsy specimens is difficult. 5.2 to 24.5 percent of biliary structures resected as malignant proved to be benign after histological examination of the surgical specimens. Recent studies have demonstrated that γH2AX, a sensitive marker for DNA double-strand breaks (DSBs), are detected at higher levels in cancer than in precancerous or normal lesions. However, γH2AX expression in biopsy specimens of biliary structures have not been examined. The aim of this study was to determine the usefulness of γH2AX expression in biopsy specimens to discriminate between benign and malignant tissues.

Methods: We examined biopsy specimens of six extrahepatic bile duct cancer tissues, seven inflammatory tissues of the bile duct, and five normal bile duct tissues. The diagnoses of these samples were confirmed by postoperative histological examination or clinical follow-up. Immunohistochemical examination of γH2AX was performed for each biopsy specimen. The nuclear staining pattern was classified into two patterns: focal staining pattern, characterized by focus formation in the nucleus; and diffuse staining pattern, characterized by peripheral– or pan-nuclear staining. The labeling index (LI) of γH2AX-positive cells was calculated for each staining pattern. Continuous variables were compared using the Mann-Whitney U test or the Kruskal-Wallis test. All tests were two-tailed, and P-values less than 0.05 were considered statistically significant.

Results:Kruskal-Wallis test revealed that the LI of γH2AX-positive cells with focal staining was significantly different between cancerous, inflamed, and normal biopsy tissues . Moreover, Dunn's multiple comparison test revealed that there was a significant difference of LI of γH2AX-positive cells with focal staining between cancer tissues and normal tissues (P < 0.01). On the other hand, there was no significant difference in the LI of γH2AX-positive cells with diffuse staining among the three different types of tissues (Kruskal-Wallis test; P > 0.05).

Conclusion:To our knowledge, this is the first study to examine the LI of γH2AX-positive cells in human biliary biopsy samples. The LI of γH2AX-positive cells with focal staining might be a useful biomarker to discriminate between benign and malignant biliary tissues.

 

40.14 High Levels of Sphingolipids in Human Pancreatic Cancer

Posted on January 18, 2018

K. Yuza1, M. Nagahashi1, Y. Hirose1, M. Nakajima1, K. Miura1, H. Ichikawa1, Y. Shimada1, J. Sakata1, H. Kameyama1, T. Kobayashi1, K. Takabe2,3, T. Wakai1  1Niigata University Graduate School Of Medical And Dental Sciences,Division Of Digestive And General Surgery,Niigata City, NIIGATA, Japan 2Roswell Park Cancer Institute,Breast Surgery, Department Of Surgical Oncology,Buffalo, NY, USA 3University At Buffalo Jacobs School Of Medicine And Biomedical Sciences, The State University Of New York,Department Of Surgery,Buffalo, NY, USA

Introduction:
Pancreatic cancer is one of the most lethal diseases and it often spreads quickly before it causes any symptoms. Elucidation of the underlying mechanisms how pancreatic cancer progresses and metastasizes is the key to improve outcome. Sphingosine-1-phosphate (S1P), a bioactive lipid mediator plays critical roles in cancer progression. S1P is involved in numerous cellular functions such as cell proliferation, migration, survival, angiogenesis and lymph angiogenesis, all of which are related to cancer progression and metastasis. Although it is expected that S1P plays an important role in pancreatic cancer progression based on the previous findings of experimental models, its role in human pancreatic cancer has never been revealed. We hypothesized that sphingolipids including S1P are produced higher in pancreatic cancer compared with normal pancreas tissue.

Methods:
Tumor and non-cancerous pancreas tissue were obtained from 10 patients with pancreatic cancer. Both tumor and non-cancerous pancreas tissue were collected from the same resected specimen, and non-cancerous tissue was collected from normal pancreas as far away as possible from the cancer site. Sphingolipids including S1P and their metabolites were measured by liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). The levels of each sphingolipid were compared between tumor and non-cancerous pancreas tissue by using the Wilcoxon matched-pairs signed rank test. All of the tests were two-sided and P values < 0.05 were considered to be statistically significant.

Results:
Levels of sphingosine, dihydro-sphingosine, S1P, and dihydro-S1P in the pancreatic cancer and normal pancreas tissue were all successfully determined. The levels of all these sphingolipids were universally higher in the cancer tissue than in the normal pancreas tissue (P<0.001 for sphingosine, dihydro-sphingosine, and S1P; P<0.05 for dihydro-S1P). We also determined the levels of each species of ceramide (C14:0, C16:0, C18:1, C18:0, C20:0, C22:0, C24:1, C24:0, C26:1 and C26:0) in the pancreatic cancer and normal pancreas tissue. We found that C14:0 alone was significantly higher in the cancer tissue than in the normal pancreas tissue.

Conclusion:
Levels of sphingolipids in cancer tissue are generally higher than normal pancreas tissue in patients with pancreatic cancer. The high levels of S1P and its metabolites in cancerous tissues implicate the important role of S1P in pancreatic cancer.
 

40.11 High Expression of DMT1 Indicated Better Prognosis in Non-B Non-C Hepatocellular Carcinoma

Posted on January 18, 2018

T. Hoki1, E. Katsuta2, L. Yan3, K. Takabe2,4, F. Ito1,4,5  1Roswell Park Cancer Institute,Center For Immunotherapy,Buffalo, NY, USA 2Roswell Park Cancer Institute,Breast Surgery, Department Of Surgical Oncology,Buffalo, NY, USA 3Roswell Park Cancer Institute,Department Of Biostatistics And Bioinformatics,Buffalo, NY, USA 4State University Of New York At Buffalo,Department Of Surgery, University At Buffalo Jacobs School Of Medicine And Biomedical Sciences,Buffalo, NY, USA 5Roswell Park Cancer Institute,Department Of Surgical Oncology,Buffalo, NY, USA

Introduction:

Hepatocellular carcinoma (HCC) is the sixth most common malignancy with poor prognosis worldwide. HCC commonly develops in patients with underlying chronic liver disease. Higher iron accumulation is present in chronic liver disease and is known to be a risk factor in the development of HCC. The divalent metal-ion transporter-1 (DMT1) is a primary importer of non-heme iron, and is ubiquitously expressed throughout the body, with highest expression in the proximal duodenum, which is the main site of iron uptake. We previously reported that mal-regulation of iron metabolism caused by increased DMT1 expression in the duodenum induced hepatocarcinogenesis. Increased expression of DMT1 in tumorous tissue has also been shown to be associated with carcinogenesis and progression in colorectal and esophageal adenocarcinoma. However, the role of DMT1 in liver of HCC patients remains unknown.

Methods:
Clinical and RNA-seq data were obtained from the Cancer Genome Atlas (TCGA). Gene expression level was compared among each AJCC Stage, and each viral infectious status. Patients were divided into two groups based on DMT1 expression level for survival analysis using Kaplan-Meier method followed by Log-rank test. The cut-off value was determined by automated scanning and selecting the threshold yielding the lowest p-value.

Results:
Of 442 HCC patients in the TCGA cohort, tumor RNA-seq data were obtained from 342 patients with overall survival (OS) and etiology data. The prevalence of HBV, HCV, dual HBV-HCV, and non-B non-C were 97(28.4%), 47(13.7%), 6(1.8%), and 192(56.1%), respectively. Clinical stages were available for 327 patients, and the patients number of stage I, II, III, and IV were 165(50.5%), 75(22.9%), 82(25.1%), and 5(1.5%), respectively. The median observation period was 19.3 months (range, 0–120.73m). There was no significant difference in the expression level of DMT1 among each clinical stage (stage I/II/III/IV) and each viral infection status (HBV/HCV/dual HBV-HCV/non-B non-C). To investigate the impact of DMT1 expression on patients’ prognosis, OS was compared between high and low expression groups in the whole cohort as well as in different viral infectious status. Interestingly, high expression of DMT1 showed better survival in non-B non-C patients (5-year OS rate: 65.2% vs 32.7%, p=0.038), HBV patients (5-year OS rate: 78.3% vs 61.9%, p=0.006), and HBV and/or HCV-infected patients (5-year OS rate: 65.7% vs 36.5%, p=0.001) compared to low expression group. Whereas there was no significant difference in OS between high and low expression groups in HCV patients (p=0.193) and the whole cohort (p=0.069).

Conclusion:
HCC with increased expression of DMT1 has better OS for HBV, HBV and/or HCV, and non-B non-C patients. These findings may imply that DMT1 in HCC tumors plays different role among different viral infectious status patients, and play roles to suppress tumor progression other than iron transportation.

40.08 Glutathione sp and Metabolic Signatures of NAFLD/NASH in the Rodent.

Posted on January 18, 2018

M. Schade1, J. Sanabria1, R. Aguilar1, M. Andryka1, A. Mallick1, J. Fannin1, J. Sanabria1,2  1Marshall University Schoool Of Medicine,Department Of Surgery,Huntington, WV, USA 2Case Western Reserve University School Of Medicine,Surgery, Nutrition And Preventive Medicine,Cleveland, OH, USA

Introduction:
Non Alcoholic Liver Disease (NAFLD) and its complications, NASH, ESLD and HCC have become the second most common cause for liver transplantation in the West. Although major advances have been done in the understanding of fatty liver disease, the metabolic disturbances that occur in the normal cell from NAFLD to NASH remains to be determined.

Methods:
C57Bl 6J mice (female: n=7/male: n=7 per each time point) were exposed to normal chow (control), high fat diet + fructose (HFD, Western diet) and methyl choline deficient diet (MCD). Liver and plasma were collected at weeks 7, 12, 16, 20, 24 and 48. Body weight, total body water, lean mass and fat compartment were determined by MRI spectroscopy. The proportion of cells undergoing mitosis or cell arrest were determined against liver apoptotic index and collagen deposition by standard stains using morphometric techniques. Quantitative protein expression of genes involved in cell metabolism or cell senescence (p53, p21, p16, mTOR1, STAT33, SIRT7, FOX01, Grb2) were determined by Western Blots.   In addition, Na/K-exchange pump regulation was determined by SRC expression.  While liver oxidative stress was determined by glutathione sp, hepatic mitochondrial ß -lipid oxidation function was determined by octanoate/butyrate ratio. Metabolites (n=852) were measured in treated plasma by LC/MS-MS. Principal component analyses (PCA) was conducted to detect metabolite differences among groups.  

Results:
The total body weight corrected for aging increased significantly in the HFD due to an increase in the fat compartment with similar lean mass and total body water (HFD vs normal chaw, p<005 by ANOVA).  The animals in the MCD group had a significant decrease in their body weight due to decreased fat, lean and total body water (MCD vs normal chaw, p<005 by ANOVA). Nonetheless, both diets induced NAFL/NASH. An increased proportion of cells in senescence was observed with decreased mitotic index and increased apoptotic activity (HFD and MCD vs normal chaw, p<0.05 by ANOVA). Morphological changes correlated with gene expression. Significant decreased in both mitochondrial ß-lipid oxidation function and Na/K-exchange pump activity after censored for aging (HFD and MCD vs normal chaw, p<0.05, by ANOVA). Glutathione ratio reduced/oxidized was significantly reduced animals with fatty liver (HFD and MCD vs normal chaw, p<0.05, ANOVA). Metabolic signatures of NAFLD and NASH after censored for aging on cell aging demonstrated disturbances in the lipid and carbohydrate metabolism.

Conclusion:

HFD and MCD generates NAFLD/NASH with decreased cellular redox status and mitochondrial ß-lipid oxidation, manifested by metabolic changes that are seen in accelerated cell senescence with characteristic plasma metabolic signatures. Described metabolic changes are of interest as therapeutic targets for arrest or reversibility of disease progression.   

 

40.07 Glutathione sp and Metabolic Signatures of Physiologic Liver Senescence in the Rodent.

Posted on January 18, 2018

M. Schade1, J. A. Sanabria1, R. Aguilar1, M. Andryka1, A. Mallick1, J. Fannin1, J. Sanabria1,2  1Marshall University Schoool Of Medicine,Department Of Surgery,Huntington, WV, USA 2Case Western Reserve University School Of Medicine,Surgery, Nutrition And Preventive Medicine,Cleveland, OH, USA

Introduction:
Human life expectancy has increased to over 80years in the last decade and more than half of the Western population will be over the age of 50 in 2020. Surgical procedures are expected to increase in number and complexity in the older. The understanding of the aging process and its metabolic implications on cell energy requirements is of paramount importance. Nevertheless, biological markers of cell stress and/or healing reservoir on the aging liver remains unclear.

Methods:
C57Bl 6J mice (female: n=7/male: n=7 per each time point) and db/db (diabetic, female=7) were exposed to normal chow. Livers and plasma were collected at different stages of animal growth and aging (weeks 7, 12, 16, 20, 24 and 48). Body weight, total body water, lean mass and fat compartment were determined by MRI spectroscopy. Cardiac function was followed by echocardiography. The proportion of cells on mitosis or in senescence was determined against liver apoptotic index and collagen deposition by standard stains using morphometric techniques. Quantitative protein expression of genes involved in cell metabolism or cell senescence (p53, p21, p16, mTOR1, STAT33, SIRT7, FOX01, Grb2) were determined by Western Blots.   In addition, Na/K-exchange pump regulation was determined by SRC expression.  While liver oxidative stress was determined by glutathione sp, hepatic mitochondrial ß -lipid oxidation function was determined by octanoate/butyrate ratio. Metabolites (n=852) were measured in treated plasma by LC/MS-MS. Principal component analyses (PCA) and partial least square discriminant analysis (PLS-DA) were conducted to detect metabolite differences among groups.

Results:
The total body weight increases with aging manly due to an increase in the fat compartment with decreased lean mass and total body water (W7vsW24vsW48, p<005 by ANOVA).  LVEF decreased and increased LV wall with age. An increased proportion of cells in senescence was observed with decreased mitotic index and increased apoptotic activity (W7vsW16vsW24vsW48, p<0.05 by ANOVA). Morphological changes correlated with expected gene expression and significant decreased in both mitochondrial ß-lipid oxidation function and Na/K-exchange pump activity (p<0.05, by ANOVA). Glutathione ratio reduced/oxidized was reduced with aging (p<0.05, ANOVA). These findings were more accentuated in the diabetic mice. Metabolic prints on cell aging showed disturbances of not only lipid but carbohydrate metabolism.

Conclusion:
Body aging under normal diet is characterized by an increase fat compartment with decreased lean mass and total body water. Changes that correlate with a decreased in liver mitochondrial function, Na/K-exchange pump activity and cell redox status, conferring metabolic signatures of liver aging. These changes must be taking into account on judging liver response to injury, liver disease development and progress and early detection of liver malignancy.
 

40.06 Prevention of early liver metastasis after pancreatectomy by NF-kB inhibitor

Posted on January 18, 2018

N. SAITO1, T. Uwagawa1, H. Shiba1, R. Hamura1, N. Takada1, H. Sugano1, T. Horiuchi1, Y. Shirai1, T. Ohashi1, K. Yanaga1, K. Yanaga1  1Jikei University School Of Medicine,Department Of Surgery,Minato-ku, TOKYO, Japan

Introduction:  Liver metastasis is a significant clinical problem early after resection of pancreatic cancer. Kocherization during early phase of the operation for pancreatic cancer prohibits the use of no-touch isolation, when tumor cells may dislodge, implant and settle in the liver through the portal vein. Matrix metalloproteinase (MMP)-2/9 degrades basement membrane and makes the tumor more susceptible to infiltration. FUT 175, a serine protease inhibitor, is known to inhibit MMP-2/9 by suppressing the NF-kB signal. Therefore, we hypothesized that preoperative treatment by FUT 175 may prevent hepatic metastasis and prolong postoperative survival of patients with post-resectional pancreatic cancer. 

Methods:  In vitro, we used a mouse pancreatic cancer cell line (PAN02). Between control and FUT 175 treatment groups, cell viability was examined by MTT assay, and the levels of cytoplasm MMP-2/9 were evaluated by Western blotting. Furthermore, activation levels of MMP-2/9 in supernatant of cultured cells were investigated by gelatin zymography. In vivo, 1´106 PAN02 cells were injected into the spleen of the mouse (C57BL6) to produce liver metastases. In treatment group, PAN02 cells were treated with FUT 175, as compared to vehicle alone in control group. The volume of liver metastasis was assessed by MRI once a week. After sacrifice, metastatic tumors were evaluated by various assays. Also, survival rates were compared.  

Results: FUT 175 did not suppress the viability of PAN02 cells for the first 24 h of treatment (p>0.05). However, the levels of cytoplasmic MMP-2/9 were down-regulated by FUT 175 after 24 h of treatment. In vivo, for control group, all animals died within 7 weeks after injection (n=10), for which MRI revealed rapid growth of metastatic tumors. On the contrary, the tumor growth rates of treatment group were very slow, and 4 of 10 animals (40%) were alive for over 7 weeks. 

Conclusion: Because of suppression of the level of MMP-2/9 in vitro and prolonged survival of liver metastasis of pancreatic cancer in mice, FUT 175 may prevent liver recurrence after pancreatectomy.

 

40.05 Sphingosine Kinase Type 1 and Type 2 Works Differently in Pancreatic Cancer

Posted on January 18, 2018

M. Nakajima1, K. Yuza1, J. Tsuchida1, Y. Hirose1, K. Miura1, H. Ichikawa1, Y. Shimada1, T. Kobayashi1, J. Sakata1, H. Kameyama1, M. Abe2, K. Sakimura2, T. Wakai1, M. Nagahashi1  1Niigata University Graduate School Of Medical And Dental Sciences,Division Of Digestive And General Surgery,Niigata City, NIIGATA, Japan 2Brain Research Institute, Niigata University,Department Of Cellular Neurobiology,Niigata, NIIGATA, Japan

Introduction: Pancreatic cancer is one of the most lethal diseases known, and it is important to develop new therapeutic agents. Sphingosine-1-phosphate (S1P) is a pleiotropic lipid mediator that regulates cell survival, migration, angiogenesis and lymphangiogenesis, which are all factors involved in cancer progression. S1P, which functions intra- and extracellularly, is generated inside the cell by two sphingosine kinases (SphK1 and SphK2). We have reported that SphK1 plays an important role in S1P secretion (J Biol Chem 2010) and cancer progression (Cancer Res 2012, J Surg Res 2016), and that SphK2 has a unique role in regulating cellular functions in the liver (Hepatology 2015). Little is known, however, about the role of SphK1 and SphK2 in pancreatic cancer progression. The aim of this study is to investigate the role of SphK1 and SphK2 in pancreatic cancer progression using SphK-knockout (KO) cells generated by CRISPR/Cas9 technology.

Methods: We generated Pan02 murine pancreatic cancer cell lines with a CRISPR/Cas9 mediated targeted deletion of the SphK1 or SphK2 gene. To investigate the role of SphK1 or SphK2 in cellular proliferation, we assessed cell growth by a spectrophotometric technique using the water-soluble tetrazolium salt, WST-8. Cell migration was measured by an in vitro scratch assay. In the animal experiments, we assessed the prognosis of C57BL/6 mice injected with the SphK1 KO or SphK2 KO Pan02 cells described above intraperitoneally.

Results: SphK2 KO Pan02 cells were significantly less proliferative than WT cells. Unexpectedly, SphK1 KO cells were significantly more proliferative than WT cells. The in vitro scratch assay indicated that SphK2 KO cells were less migratory than WT cells, and that SphK1 KO cells had greater migratory ability than WT cells. Furthermore, the animal experience showed that mice injected with SphK1 KO cells had shorter prognosis than those injected with SphK1 WT cells, while mice injected with SphK2 KO cells had longer prognosis than those injected with SphK2 WT cells. These results indicate that SphK2, rather than SphK1, may have important roles in proliferation and migratory behavior in pancreatic cancer cell lines and pancreatic cancer progression. On the other hand, SphK1 KO cells treated with gemcitabine had more survival rate than WT cells and SphK2 KO cells treated with gemcitabine had less survival rate than WT cells. These results indicate that SphK1 may have important roles in resistance against chemotherapy.

Conclusion: Our findings indicate that S1P produced by SphK1 and SphK2 may have different functions in pancreatic cancer cell. Targeting both SphK1 and SphK2 signaling pathways may be a potential strategy for pancreatic cancer treatment.

 

40.04 Diagnosis of Pancreatic Adenocarcinoma via Protein Signatures from Fine Needle Aspirations

Posted on January 18, 2018

M. H. Gerber1, D. Delitto1, B. DiVita1, S. Han1, R. M. Thomas1, J. Trevino1, S. J. Hughes1  1University Of Florida,Department Of Surgery,Gainesville, FL, USA

Introduction:  Cytological analysis of fine needle aspiration (FNA) of pancreatic lesions fails to confirm pancreatic ductal adenocarcinoma (PDAC) in up to one third of patients. We discovered that a 4-analyte protein signature from tumor lysates distinguishes PDAC from other lesions including chronic pancreatitis, thus raising the notion protein signatures could prove superior to FNA cytological analysis in the diagnosis of PDAC.  Here, we aim to translate this observation to the FNA biopsy platform.

Methods:  At time of operation, a “virtual” FNA biopsy of various pancreatic pathologies was obtained using a 19-gauge needle with six passes through the intestinal wall into the region of interest. Biopsy samples were placed in various quantities of lysis buffer and protein concentrations of each sample determined. IL-6 concentrations (one of the informative analytes in the diagnostic protein signature) were determined for each FNA biopsy sample, with and without spiking of “contaminant” plasma.

Results: Samples were collected from 27 consecutive patients undergoing pancreatectomy (PDAC n = 18; normal n = 3; pancreatitis n = 6). Protein concentrations from FNA biopsy samples diluted in 150 ul of lysis buffer ranged from 1.8 –  260.8 mg/ml with a median of 17.1 mg/ml (Interquartile range (IQR): 11.5 – 32.6 mg/ml). IL-6 levels in the FNA samples ranged from 185 – 2941 pg/ml with a median of 486 pg/ml (IQR: 247 – 740 pg/ml). Plasma concentrations of IL-6 from the same patients ranged from 59 – 312 pg/ml with a median of 111 pg/ml (IQR:  65 – 203 pg/ml).  When patient FNA samples were compared to plasma samples, FNA samples had IL-6 concentrations 3.1 to 14.9 times higher than the matched patient plasma concentration.

Conclusion: Pancreatic FNA biopsy produces adequate quantities of protein for multiple replications on a variety of protein assay platforms. Normalization of cytokine concentrations to total protein may be subject to blood/plasma contamination inherent to the FNA procedure; thus alternative normalization to a resident, soluble protein may be necessary.  Soluble protein analysis of pancreatic FNA biopsy samples remains a realistic supplement to cytology in the diagnosis of PDAC.
 

40.03 IL-23 Plays an Important Role in Short-Term Survival after Pancreatic Adenocarcinoma Resection

Posted on January 18, 2018

B. A. Krasnick1, S. M. Husain2, Y. Bi1, P. V. Dickson2,3, J. Deneve2,3, D. Shibata2,3, R. Fields1, W. G. Hawkins1, E. S. Glazer2,3  1Washington University,Surgery,St. Louis, MO, USA 2University Of Tennessee Health Science Center,Surgery,Memphis, TN, USA 3University Of Tennessee West Cancer Center,Surgery,Memphis, TN, USA

Introduction:  Pancreatic ductal adenocarcinoma (PDAC) carries the highest case fatality rate of any cancer and will likely become the second leading cause of death by 2020.  TGF-ß has a paradoxical relationship with survival in PDAC patients where it is a tumor suppressor in early stage PDAC and a tumor promotor in late stage PDAC. While TGF-ß is known to drive inflammation in the tumor microenvironment (TME), the role of interleukins in the pancreatic TME is not well understood.  We hypothesized that IL23, a pro-inflammatory interleukin associated with suppression of cytotoxic T cells, is associated with survival in PDAC.

Methods:  24 long-term survivors (>30 months) and 24 short-term survivors (<12 months) with resected PDAC were identified. Tumor sections were taken from formalin fixed, paraffin embedded blocks and protein expression of IL-23 and TGF-ß were independently investigated with immunohistochemistry utilizing quantitative analysis with CellProfiler image analysis software. Immunohistochemistry expression of IL23 or TGF-ß was determined to be high (highest quartile), low (lowest quartile), or median (within the interquartile range) based on 5 representative images of each tumor section. Comparisons with clinical outcomes were investigated with Student’s t-test, ANOVA, or multivariate regression.

Results: There was no significant difference in the average age (66 ± 12 years), gender (44% male), or clinical stage between the two groups. Patients with low TGF-ß protein expression were more likely to be in the short-term survival group (OR=2.2, P=0.018). Tumors from short-term survivors were significantly more likely to have low IL23 expression (OR=0.48, P=0.019).  In long-term survivors, neither TGF-ß nor IL-23 protein expression was associated with survival (P>0.05). There was no difference in IL23 expression in low or median TGF-ß expressing tumors (P>0.5), however, in high TGF-ß expressing tumors, long-term survivors were associated with 20% higher IL23 protein expression (P=0.008). Multivariate analysis demonstrated that long-term survival was linearly associated with increasing IL23 expression (OR=3, P=0.001) but not TGF-ß expression (P=0.07). Overall, we also found a statistical association between IL23 expression and TGF-ß expression (P<0.001) and that long-term survival was associated with a higher ratio of IL23 / TGF-ß expression (P=0.04).

Conclusion: We found that IL23 tumor expression is associated with survival after PDAC resection in short-term survivors and statistically related to TGF-ß expression for all patients.  While other groups have shown that TGF-ß is associated with survival in PDAC, we demonstrated that IL23 may be a critical component to understanding the relationship between TGF-ß expression and survival in patients with survival less than 12 months.

 

40.02 HO-1 Polymorphism and Acute Necrotic Pancreatitis Through V-cam and E-selectin Expression

Posted on January 18, 2018

A. K. Gulla1,2, A. Gulbinas3, G. Barauskas3, Z. Dambrauskas3  1Georgetown University Medical Center,Department Of Surgery,Washington, DC, USA 2Vilnius University Hospital, Santaros Clinics,Department Of Surgery,Vilnius, SANTARISKIU 2, Lithuania 3Lithuanian University Of Health Sciences, Kaunas Clinics,Department Of Surgery,Kaunas, , Lithuania

Introduction:  Acute pancreatitis is a severe and frequently a life-threatening disease, which can lead to pancreatic necrosis, acute lung injury, SIRS and MODS. The inducible enzyme heme oxygenase-1 (HO-1) is an anti-oxidative, anti-inflammatory, and cytoprotective enzyme that is induced in response to cellular stress. The HO-1 promoter contains (GT)n dinucleotide repeats and is highly polymorphic in the population. In this study, we hypothesized that the number of GT repeats in HO-1 promoter can influence the occurrence of acute necrotic pancreatitis through v-cam and e-selectin expression due to its protective function. Patients with acute pancreatitis are more likely to have long repeats than controls.

Methods: Acute pancreatitis patients (n=135) and age- and sex-matched healthy controls (n=33) were studied. Peripheral blood samples from pancreatitis patients were collected on admission. Genomic DNA was extracted from the blood samples of patient and control groups. The HO-1 promoter region with the GT repeats was PCR amplified with fluorescent tagged primers and levels of cytokines were measured.

The PCR products were analyzed by ABI 3130 genetic analyzer and the exact size of the PCR products was determined by GeneMapper software.  A short allele was defined as containing 27 GT repeats or fewer, whereas a long allele was more than 27 repeats.

Results: The subjects were categorized into 3 groups based on the genotype

Results: one short and one long alleles (S/L), two short alleles (S/S) and two long alleles (L/L). The presence of S/L was similar between the patient group (41.2%) and the controls (39.4%). Interestingly, 46.6% of patients were carriers of two long repeats (L/L) while 11.1% v-cam and 11.1 % e-selectins levels (p<0.05) vs 24.2% of control subjects, whereas 12.2% of patients were carriers of two short repeats vs 36.4% of control population. 

Conclusion: Our data demonstrate a strong bias toward longer alleles and higher levels of v-cam and e-selectins among patients with acute pancreatitis. Thus, polymorphism of the GT repeats in the HO-1 promoter region may be a risk factor for developing acute pancreatitis. Further studies are now underway to analyze the pancreatic levels of HO-1 protein in acute pancreatitis patients and controls and to determine whether the presence of the short alleles facilitate HO-1 upregulation and consequently promote its protective anti-inflammatory function in acute pancreatitis.

40.01 Modeling of Early Pancreatic Neoplasia Transcriptional Regulation in Mice

Posted on January 18, 2018

J. K. Thompson1, H. Crawford3,4, M. Pasca Di Magliano1,2, F. Bednar1  1University Of Michigan,Surgery,Ann Arbor, MI, USA 2University Of Michigan,Cell And Developmental Biology,Ann Arbor, MI, USA 3University Of Michigan,Molecular And Integrative Physiology,Ann Arbor, MI, USA 4University Of Michigan,Internal Medicine,Ann Arbor, MI, USA

Introduction: KRAS is the primary oncogenic driver in pancreatic ductal adenocarcinoma (PDA). Pancreatic acinar cells are most susceptible to transformation by Kras in mouse models of PDA. The earliest stage of transformation consists of conversion of the acinar cells to a duct-like progenitor phenotype in a process called acinar-ductal metaplasia (ADM). Networks of developmental transcription factors (TFs) are involved in fate specification and maintenance of acinar cells. We hypothesize that oncogenic Kras alters these networks to establish the neoplastic cell state within the pancreas. In this work, we aimed to validate a genetically engineered mouse model that will allow us to analyze changes in gene regulatory networks driven by oncogenic Kras in early pancreatic neoplasia.

Methods: Mice with a pancreatic acinar cell-specific, tamoxifen inducible Cre recombinase (Ela-CreER) were bred with strains containing the oncogenic Kras G12D allele and the fluorescent protein tdTomato in the Rosa26 locus. To activate the oncogenic Kras and the tdTomato lineage tracer, Ela-CreER; Kras G12D/+; R26 tdTomato mice and littermates lacking oncogenic Kras, were treated with daily gavages of tamoxifen (4mg/day) for five consecutive days. Mice were sacked one week after the initial tamoxifen gavage. We analyzed the expression of tdTomato by fluorescence microscopy and fluorescence-activated cell sorting (FACS). Total cell RNA was isolated from the sorted cells using the RNeasy Micro Kit (QIAGEN) and complementary DNA (cDNA) was synthesized with the High Capacity cDNA Reverse Transcription Kit (Applied Biosystems). We utilized a panel of 26 pancreatic developmental TFs and three acinar and duct markers (amylase, elastase, keratin 19 – CK19) with specific quantitative PCR (qPCR) TaqMan probes to characterize the isolated cells. Differences in TF expression between the wildtype and oncogenic Kras-containing mice were analyzed with a Kruskal-Wallis rank test. Statistical significance was set at p<0.05.

Results: Tamoxifen gavage induced high level of tdTomato expression in Ela-CreER mice. Fluorescence microscopy confirmed that the tdTomato+ cells were also amylase positive. RT-qPCR analysis after FACS sorting confirmed high amylase, high elastase, and low CK19 expression in the tdTomato+ cells. We also consistently found measurable levels of 23/26 (88%) of pancreatic developmental TFs in the isolated acinar cells. Preliminary analysis did not reveal significant differences in TF levels in oncogenic Kras-expressing pancreata versus wildtype controls.

Conclusion: We established a genetically engineered mouse model of early pancreatic neoplasia, which allows for specific isolation of acinar cells and their progeny. Early analysis of the model suggests that one-week activation of oncogenic Kras does not yet lead to significant developmental TF expression changes in the adult pancreas.

 

35.03 Impact of Hospital Volume on Outcomes of Laparoscopic versus Open Hepatectomy for Liver Cancer

Posted on January 18, 2018

S. W. De Geus1, G. G. Kasumova1, T. E. Sachs1, O. Akintorin1, S. Ng1, D. McAneny1, J. F. Tseng1  1Boston University,Surgery,Boston, MA, USA

Introduction:  Previous investigators have suggested that laparoscopic liver resection may be superior to an open operation based on studies at high-volume centers; however, the applicability of these findings remains unclear. This study investigates whether hospital volume is a factor in determining the short- and long-term outcomes of laparoscopic versus open hepatectomy for liver cancer.

Methods:  The National Cancer Database (NCDB) was queried for patients who underwent open or laparoscopic hepatectomy, without transplantation, for liver cancer 2010-2013. Institutions were defined as being either low-volume hospitals (LVH, ≤ 11 operations/year) or high-volume hospitals (HVH, >11 operations/year). For entire cohort and within each category, positive margin rate, 30-day mortality, readmissions, prolonged hospital stay (hospital stay ≥ 14 days), and overall survival were compared between patients who had laparoscopic and open resections, using multivariate logistic regression and Kaplan-Meier methods. 

Results: 2,867 patients underwent hepatectomy for liver cancer. Overall, 612 (21.4%) of resections were performed laparoscopically. After adjustment for covariates, resections for liver cancers at a HVH were significantly associated with lower positive-margin rates (HVH vs. LVH: 8.3% vs. 11.0%; adjusted odd ratio [AOR], 0.744; p=0.0413) and 30-day mortality (HVH vs. LVH: 3.5% vs. 6.2%; AOR, 0.646; p=0.0375). However, no significant differences were observed among the HVHs and LVHs regarding readmissions (HVH vs. LVH: 4.6% vs. 4.8%; AOR, 1.039; p=0.8482), prolonged hospital stay (HVH vs. LVH: 9.2% vs. 8.8%; AOR, 1.065; p=0.6648), or overall survival (HVH: log-rank p=0.1405; LVH: log-rank p=0.2322). Multivariate regressions showed in both HVH and LVH, laparoscopic resections were not significantly associated with positive margins (HVH: AOR, 1.246; p=0.4176; LVH: AOR, 0.991; p=0.9627), 30-day mortality (HVH: AOR, 0.755; p=0.5456; LVC: AOR, 1.037; p=0.8808), readmission (HVH: AOR, 0.834; p=0.6297; LVH: AOR, 0.698; p=0.2302) prolonged hospital stay (HVH: AOR, 0.626; p=0.1172; LVH: AOR, 0.886; p=0.5766), or overall survival (HVH: log-rank p=0.1405; LVH: log-rank p=0.2322) when compared to open.

Conclusion: Although outcomes after major operations are influenced by various factors beyond hospital volume alone, the results of this study suggest that patients with liver cancer are at higher risk of having positive resection margins and 30-day mortality if they are treated at LVH instead of HVH.  However, for both high- and low-volume hospitals, laparoscopic resections of liver cancer were associated with surgical and oncologic outcomes that were similar to those for open operations. Although residual selection bias regarding MIS vs open approach must be acknowledged, our data suggest that laparoscopic liver resection when feasible is a reasonable approach across hospital volume strata.

 

35.01 Triple-drug Therapy to Prevent Pancreas Fistula for the Patients with a High Drain Amylase Level.

Posted on January 18, 2018

T. Adachi1, S. Ono1, T. Adachi1, M. Yamashita1, T. Hara1, A. Soyama1, M. Hidaka1, K. Kanetaka1, M. Takatsuki1, S. Eguchi1  1Nagasaki University,Department Of Surgery,Nagasaki, , Japan

Introduction: A high drain amylase level is a well known predictive marker for the development of a pancreatic fistula (PF) after a pancreaticoduodenectomy (PD). Any sign of PF following a PD warrants immediate preventive measures to avoid the development of PF. We aimed to determine the efficacy of a triple-drug therapy (TDT) regimen using gabexate mesilate, octreotide, and carbapenem antibiotics to prevent PF in patients showing a high drain amylase level on postoperative day (POD1) after PD.

Methods: We enrolled 183 patients who had undergone a PD from the year 2007. Patients were divided into two groups based on the study period. The former period group (2007~2011, n = 81) included patients in whom no particular treatment had been administered even if their drain amylase level on POD1 was high ( ≥  10,000 IU/L). The latter period group (n = 102) included patients having a high drain amylase level on POD1, who had received TDT [gabexate mesilate 600 mg/day continuous intravenous (civ.), octreotide 150 µg/day civ., and carbapenem 1.0 g/day IV] along with a fasting status in one week. Any other postoperative management including the day of drain removal (POD5) was same in this study period. The primary endpoint was the incidence of PF [beyond grade B defined by the International Study Group of Pancreatic Fistula (ISGPF criteria)]. 

Results: Incidence of PF in all enrolled patients was 10.9%. Incidence of high drain amylase level ( ≥  10,000 IU/L) on POD1 in the former group was 11.1% and in the latter group was 17.6%. Incidence of PF in patients whose drain amylase level was not high ( ?  10,000 IU/L) was equivalent (8.2 vs. 4.8%, p = 0.36), between the two groups; however incidence of PF in patients with a high drain amylase level in the latter group was effectively prevented by administration of TDT showing results that were statistically significant (88.9 vs. 11.1%, p ? 0.001).

Conclusion: TDT is an effective treatment strategy to prevent PF even in patients with a high drain amylase level after a PD procedure.
 

34.10 Non-invasive Fibrosis Marker Impacts the Mortality after Hepatectomy for Hepatoma among US Veterans

Posted on January 18, 2018

F. B. Maegawa1,2, L. Shehorn3, J. B. Kettelle1,2, T. S. Riall2  1Southern Arizona VA Health Care System,Department Of Surgery,Tucson, AZ, USA 2University Of Arizona,Department Of Surgery,Tucson, AZ, USA 3Southern Arizona VA Health Care System,Department Of Nursing,Tucson, AZ, USA

Introduction:
The clinical role of non-invasive fibrosis markers (NIFM) on the mortality of patients undergoing hepatectomy for hepatocellular carcinoma (HCC) is not well established. We investigate the long-term impact of NIFM on mortality after hepatectomy for HCC. 

Methods:
This analysis utilized the Department of Veterans Affairs Corporate Data Warehouse database between 2000-2012. The severity of hepatic fibrosis was determined by the AST-platelet ratio index (APRI) and the Fibrosis-4 score (FIB-4). Kaplan-Meier survival and Cox proportional hazard regression methods were utilized for analysis. 

Results:
Mean age, MELD score, and BMI were 65.6 (SD: ± 9.4) years, 9 (SD: ± 3.1) and 28 (SD: ± 4.9) kg/m2, respectively. Most the patients were white (64.5%), followed by black (27.6%). The most common operation was partial lobectomy (56.5%) followed by right hepatectomy (28.7%). Out of 475 veterans who underwent hepatectomy for HCC, 26.3% had significant fibrosis utilizing APRI (index >1) and 29.2% utilizing FIB-4 (score > 3.25). The long-term survival among veterans with APRI > 1 was significantly worse compared to those with a normal index. Kaplan-Meier survival analysis revealed a median survival of 2.76 vs 4.38 years, respectively (Log-Rank: p< 0.0018). In contrast, the FIB-4 score was not associated with worse survival. Median survival among veterans with FIB-4 > 3.25 compared to those with a normal score was 3.28 vs 4.22 years, respectively (Log-Rank: p = 0.144). Unadjusted Cox proportional hazard regression showed that APRI >1 is associated with increased mortality (HR: 1.45; 95% CI 1.14 – 1.84). After adjusting for age, race, BMI and MELD score, APRI remained associated with increased mortality (HR: 1.36, 95% CI: 1.02 – 1.82). FIB-4 was not associated with increased mortality in both unadjusted and adjusted analysis (HR: 1.19; 95% CI: 0.94 – 1.50 and HR:1.29; 95% CI: 0.96 – 1.72, respectively).

Conclusion:
APRI can be used as a preoperative tool to predict long-term mortality after hepatectomy, refining the selection criteria for liver resection for HCC. These results suggest patients with APRI > 1 are likely to benefit from other curative therapies, such as transplantation.
 

33.06 Surgeon Annual and Cumulative Volume Variably Predict Outcomes of Complex Hepatobiliary Procedures

Posted on January 18, 2018

M. M. Symer1, L. Gade3, A. Sedrakyan2, H. Yeo1,2  1Weill Cornell Medical College,Surgery,New York, NY, USA 2Weill Cornell Medical College,Healthcare Policy,New York, NY, USA 3NewYork-Presbyterian / Queens,Surgery,New York, NY, USA

Introduction: There is a strong volume-outcome relationship in pancreatectomy, but whether the same relationship exists for other complex hepatopancreatobiliary (HPB) procedures is not known. The role of surgeon experience is clearly important, but whether it should be defined by cumulative volume or a more contemporaneous measure like annual volume is unclear. We compared the outcomes of surgeons across the spectrum of experience to better define the volume-outcome relationship in complex HPB surgery. 

Methods: We identified all patients undergoing major elective HPB operations in New York State from 2000 to 2014 using the Statewide Planning and Research Cooperative Database. Major resections such as liver lobectomy, proximal pancreatectomy, as well as bile duct resection and complex repair were included, while wedge resections, distal pancreatectomy, and percutaneous or endoscopic procedures were excluded. In-hospital mortality and perioperative outcomes were compared across four categories of surgeons based on high or low annual and high or low cumulative operative volume. Median volume was used as the cut-point for high vs. low categories.

Results:13,236 operations performed by 893 surgeons were included in the study. Median cumulative volume was 89 operations, and median annual volume was 21 operations. Similar numbers of procedures were performed by low cumulative/low annual (LCLA) volume surgeons and high cumulative/high annual (HCHA) volume surgeons (6106 vs. 6176 operations). HCHA surgeons treated slightly older patients than LCLA surgeons (63.0y vs. 61.1y, p<0.01). HCHA surgeons also treated fewer Medicaid (5.6% vs. 10.0%, p<0.01) or Black patients (5.2% vs. 10.2%, p<0.01). HCHA surgeons performed many more minimally invasive procedures (15.2% of HCHA operations vs. 5.7% of LCLA operations, p<0.01). Mortality was lowest for HCHA and highest for LCLA surgeons (1.6% vs. 3.7%, p<0.01). Adjusted odds of in-hospital mortality were lower only for those patients undergoing surgery by HCHA volume surgeons (OR 0.47 95%CI 0.32-0.67), but not HCLA volume surgeons (OR 0.58 95%CI 0.28-1.20), or LCHA volume surgeons (OR 0.82 95%CI 0.44-1.53). 30d major events (e.g. stroke, shock), reoperation, and readmission were not affected by cumulative or annual experience. 

Conclusion:In this large New York State based study of complex, elective HPB operations only surgeons with high cumulative and high annual volume have improved in-hospital mortality. In isolation, neither high cumulative volume, nor high annual volume alone were associated with improved outcomes. Racial and socioeconomic disparities in access to high-volume care persist. Interventions to regionalize complex surgical care should account for these distinctions.

 

24.09 Pioglitazone Reduces Hepatocarcinogenesis in a Rodent Model of Cirrhosis

Posted on January 18, 2018

S. Li1, S. Ghosal1, G. Arora1, D. J. Erstad1, M. Lanuti2, K. K. Tanabe1, B. Fuchs1  1Massachusetts General Hospital,Surgical Oncology,Boston, MA, USA 2Massachusetts General Hospital,Thoracic Surgery,Boston, MA, USA

Introduction:  Advanced hepatocellular carcinoma (HCC) is a leading cause of mortality worldwide with limited treatment options. There is a readily identifiable cohort of cirrhosis patients at risk and they are ideal candidates for chemoprevention. Anti-hyperglycemic agents have garnered interest for their chemo-preventive effects. Pioglitazone, a selective PPAR-y agonist, has been shown to reduce non-alcoholic steatohepatitis (NASH), but its role as an anti-fibrotic and chemo-preventive agent has yet to be elucidated. The hypothesis of this study is that Pioglitazone reduces cirrhosis and subsequent HCC development in rats with diethylnitrosamine (DEN)-induced cirrhosis.

Methods:  Male Wistar received DEN 50mg/kg by intraperitoneal injection. DEN injury reliably recapitulates human HCC development with induction of hepatic fibrosis at 8 weeks, cirrhosis at 12 weeks, and HCC by 18 weeks. DEN-injured rats were randomized to receive oral gavage of pioglitazone at 3mg/kg/day (n=9) or vehicle control (n=9). Initiation of pioglitazone coincided with the development of liver fibrosis at 8 weeks. All animals were sacrificed at 18 weeks.

Results: As expected, repeated injections of DEN in rats resulted in progressive fibrosis, cirrhosis, followed by HCC formation. Treatment with pioglitazone resulted in a 56% reduction of surface nodules relative to treatment with vehicle (7.4±4.9 vs. 17±7; p<0.005). Pioglitazone treatment resulted primarily in a reduction of nodules<8mm compared to vehicle (6.3±1.5 vs. 15.14±2.5; p<0.001). Liver sections were stained by picrosirius red to assess fibrosis. Pioglitazone significantly reduced collagen deposition in DEN-injured rats (collagen proportional area = 3.2±1.8% vs. 9.2±2%; p<0.035). This histology was confirmed by gene expression analysis with reductions in COL1A1, α-SMA, TGF-β, and TIMP1. Pioglitazone treatment resulted in an upregulation of Adiponectin, which has been shown to antagonize carcinogenesis. Pioglitazone treatment also increased AMPK signaling, a well-recognized target for anti-tumor drug discovery as well as a down regulation of the mitogenic MAPK pathway. Daily pioglitazone dosing signifcantly reduced the gene expression of progenitor cell activation including CD44, RAGE, and DLK1. 

Conclusion: Overall our data supports the hypothesis that the anti-diabetic agent pioglitazone may be repurposed as a drug to reduce fibrosis and prevent HCC. This could be beneficiary in patient management given the low cost as well as minimal side effects.

24.05 Pancreas-Specific Arid1a Deletion Induces Intraductal Papillary Mucinous Neoplasms in Mice

Posted on January 18, 2018

I. Nassour1,2, X. Sun1, S. Zhang1, X. Luo1, L. H. Nguyen1, L. Li1, L. Peng3, J. Shen4, H. Zhu1, S. Wang1,2  1University Of Texas Southwestern Medical Center,Children’s Research Institute,Dallas, TX, USA 2University Of Texas Southwestern Medical Center,Division Of Surgical Oncology,Dallas, TX, USA 3University Of Texas Southwestern Medical Center,Department Of Pathology,Dallas, TX, USA 4Stanford University,Department Of Pathology,Palo Alto, CA, USA

Introduction: Intraductal papillary mucinous neoplasms (IPMN) are precursors to pancreatic ductal adenocarcinoma (PDAC). While activating KRAS mutations are the most common alterations found in PDAC and IPMN, ARID1A, which is a component of the SWI/SNF chromatin remodeling complex, is also commonly mutated. However, the functional effects of ARID1A mutations in pancreas tumorigenesis are not known. Understanding molecular mechanisms that drive IPMN progression may lead to the development of novel therapies that prevent IPMN transformation into PDAC.

Methods: We generated transgenic mice that had pancreas-specific activating Kras mutations and Arid1a deletion (KrasG12D; Ptf1a-Cre; Arid1af/f, or “KCA” mice). We identified 35 human IPMN resection samples, performed immunohistochemistry staining for ARID1A, and graded the expression intensity. We knocked down ARID1A and MYC in human pancreatic ductal epithelial (HPDE) cells with siRNA. siScramble was used as the negative control. To measure protein synthesis, we used an assay based on the incorporation of modified puromycin molecules (O-propargyl-puromycin (OPP)) into nascent peptides. OPP was then fluorescently labelled and quantified by fluorescence-activated cell sorting. Fluorescence level was proportional to the extent of cellular protein synthesis.

Results: Mice with only pancreas-specific activating Kras mutations (KrasG12D; Ptf1a-Cre) had significant pancreatitis and pancreatic intraepithelial neoplasia, which is another type of PDAC precursor, consistent with previous reports. In contrast, KCA mice developed macroscopic mucinous pancreatic cysts at 100% penetrance. Histologic and biochemical assessment showed that these cysts most resembled gastric subtype IPMN. Evaluation of human IPMN samples revealed that gastric subtype IPMN frequently had little to no ARID1A expression while other subtypes had high expression, confirming the fidelity of our model (Fig. A). RNA-seq from Arid1a null pancreas showed significant upregulation of gene networks involved with MYC activity and protein translation. Knocking down ARID1A (siARID1A) in HPDE cells induced increased MYC expression (Fig. B) and protein synthesis (Fig. C). Concurrent knockdown of MYC (siMYC) and ARID1A abrogated the increase in protein synthesis (Fig. D).

Conclusions: Pancreas-specific ARID1A deletion resulted in the formation of gastric subtype IPMN and increased protein synthesis that was mediated through elevated MYC activity (Fig. E). These data suggest that translation is a potential therapeutic target to block the formation and progression of gastric subtype IPMN.

 

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