2216 Differential expression of CYR61, CTGF and NOV in pancreatic cancer and the clinical relevance

S404 a monotherapy or in combination with chemotherapeutic agents. To date, most of studies have focused on the cytopathic effect of reovirus and the mechanisms of reovirus-induced cancer cell death. To achieve sufficient anti-tumor effect, it is crucial to prevent metastasis in addition to control tumor growth. Here, we address the role of reovirus as an inhibitor of migration and invasion ability of gastric cancer cells. Materials and Methods: In this study, we employed four gastric cancer cell lines including MKN1, MKN7, MKN45, and AGS, and accessed the efficacy of reovirus on gastric cancer cells using MTS and apoptosis assays. To investigate the effect of reovirus on cell migration and invasion, we performed wound healing assay and transwell migration/invasion assay. We also investigate the cytotoxic effect and cell motility inhibitory effect of anti-cancer agents including CPT-11 and paclitaxel. We examined the expression of reovirus protein and filamentous actin (F-actin), which plays an important role in cell motility, in gastric cancer cells infected with reovirus using immunofluorescence staining. Results: The cytotoxic effect of reovirus was almost same as those of anticancer drugs including CPT-11 and paclitaxel that are used for clinical treatment of gastric cancer. Reovirus showed the inhibition of cell migration and invasion in MKN1, MKN7, and AGS. Caspase-3 activity assay revealed that these effects were achieved without initiation of apoptotic cell death. The number of reovirus protein positive cells was higher in cell lines that showed a significant decrease in cell migration and invasion ability after treatment with reovirus. The expression level of F-actin and formation of stress fiber significantly decreased in cells expressing reovirus protein. In contrast, chemotherapeutic agents did not show any effect on cell motility and formation of stress fiber. Conclusion: We concluded that reovirus induces not only cytopathic effect, but also reduction of cell migration and invasion in gastric cancer cells via inhibition of actin polymerization. Our study suggests that reovirus may have a utility for malignancies as an inhibitor of metastasis. No conflict of interest. 2214 POSTER KIAA1324, a novel tumor suppressor in gastric cancer through inhibition of GRP78 oncoprotein J.M. Kang1 , S. Park1 , B. Lee1 , J. Kim1 , J. Park1 , S.T. Kim1 , S. Kim2 , H. Kim1 , S.J. Kim1 . 1 CHA University, CHA Cancer Institute, Seongnam-si Kyunggi-do, South Korea; 2 University of Tsukuba, International Institute for Integrative Sleep Medicine WPI-IIIS, Tsukuba, Japan Background: Inactivation of tumor suppressor genes by genetic alteration or epigenetic suppression leads to gastric carcinogenesis. Therefore, study of the genes that are suppressed in cancer is essential for understanding gastric carcinogenesis and developing gastric cancer therapy. Here, we investigated the role of KIAA1324 which is downregulated in gastric cancer. Material and Methods: We analyzed genetic and transcriptomic alterations of KIAA1324 in gastric cancer using transcriptome sequencing data and public database. The clinical impact of KIAA1324 in gastric cancer was assessed through tissue microarray analysis. Proliferation, invasiveness, drug resistance, and in vivo tumorigenic ability of gastric cancer cell lines expressing exogenous KIAA1324 were evaluated by cell viability, transwell invasion and xenograft assays. Protein interaction analysis was also conducted to identify KIAA1324 binding partners. Results: We observed KIAA1324 downregulation in 78% of gastric cancer patients. Tissue microarray analysis showed that low KIAA1324 levels were associated with poor prognosis in gastric cancer patients. In the xenograft model, KIAA1324 significantly reduced tumor formation of gastric cancer cells. KIAA1324 also suppressed proliferation, invasion, and drug resistance and induced apoptosis in gastric cancer cells. Through protein interaction analysis, we found glucose-regulated protein 78 kDa (GRP78) as a KIAA1324 binding partner. KIAA1324 blocked the anti-apoptotic activity of GRP78 by inhibiting the interaction between GRP78 and caspase 7 in the endoplasmic reticulum. Conclusions: Our study reveals a novel role of KIAA1324 as a gastric tumor suppressor via inhibition of GRP78 oncoprotein activity. Our data provide new insight into the diagnosis and treatment of gastric cancer. This work was supported by the Basic Science Research Program (2014R1A6A3A01058338 to J.M.K.) and the Bio-Synergy Research Project (NRF-2012M3A9C4048735 to S.-J.K.) of Ministry of Science, ICT and Future Planning through the National Research Foundation and a KHIDI grant (HI14C2640), Korea. No conflict of interest.

Abstracts 2215 POSTER Differential expression of CNN family members CYR61, CTGF and NOV in gastric cancer and association with disease progression X. Gao1 , K. Ji2 , Y. Jia1 , J. Li2 , J. Ji1 , W. Jiang2 , L. Ye2 . 1 Peking University Cancer Hospital and Institute, Gastrointestinal Surgery, Beijing, China; 2 Cardiff University school of Medicine, CCMRC/ICG, Cardiff, United Kingdom Background: CCN is an acronym of CYsteine-Rich protein 61 (CYR61), Connective Tissue Growth Factor (CTGF) and Nephroblastoma OVerexpressed (NOV) comprising another three members i.e. Wnt1-inducible signalling pathway protein (WISP)-1, -2 and -3. Aberrations of certain CCN members including CYR61, CTGF, WISP-1 and -3 have been reported in gastric cancer. The present study aimed to examine the clinical relevance of NOV together with CYR61, CTGF in gastric cancer by analysing their transcripts levels. Materials and Methods: Gastric tumour samples collected from 324 patients with gastric cancer during the operation together with background tissues adjacent to the tumours as paired control were stored at −80ºC. The tissue collection was approved by local ethic committee at Beijing Cancer Hospital and also with written consent from each patient. CYR61, CTGF and NOV expression in gastric tumour specimens was determined using real time quantitative PCR and the results were statistically analysed against patient clinical and pathological data using SPSS software. Results: NOV mRNA levels in gastric cancer tissues were significantly elevated when compared with their paired adjacent non-cancerous tissues (p = 0.014). Higher transcript levels of NOV were positively correlated with distant metastases (p = 0.035). Local advanced tumours with invasive expanding (T3 and T4) expressed higher levels of NOV p = 0.0036 compared with tumours at earlier stages (T1 and T2). Meanwhile, CYR61 mRNA levels in gastric tumours were also significantly increased compared with their adjacent corresponding non-cancerous tissues (p = 0.0019). Kaplan–Meier survival curves revealed that patients with CYR61 low transcript levels had longer overall survival (OS) (P = 0.018) and diseasefree survival (DFS) (P = 0.015). CTGF mRNA levels in gastric cancer tissues were significantly elevated when compared with their paired adjacent non-cancerous tissues (p = 0.014). High transcript levels of CTGF was positively correlated with tumour invasion (T1+T2 vs T3+T4, P = 0.005). Although transcript levels of CTGF and NOV exhibited no correlation with survival when we analysed them individually, combined CYR61, CTGF and NOV did show that patients with low transcript levels of all three genes had poorer OS (P = 0.027) and DFS (P = 0.021) compared with patients with higher expression of all three genes. Conclusions: NOV expression is increased in gastric cancer which is associated with local invasion and distant metastases. Similar expression patterns have been observed for the expression of CYR61 and CTGF in the current cohort. The elevated expression of these three genes in gastric cancer is associated with shorter survival of patients, in particular CYR61. Further investigations will highlight their predictive and therapeutic potential in gastric cancer. No conflict of interest. 2216 POSTER Differential expression of CYR61, CTGF and NOV in pancreatic cancer and the clinical relevance K. Ji1 , J. Li2 , X. Gao3 , C.Y. Hao4 , J. Ji3 , W. Jiang1 , L. Ye1 . 1 Cardiff University School of Medicine, CCMRC/ICG, Cardiff, United Kingdom; 2 Beijing Friendship Hospital, Capital Medical University, General Surgery, Beijing, China; 3 Peking University Cancer Hospital and Institute, Department of Gastrointestinal Surgery, Beijing, China; 4 Peking University Cancer Hospital and Institute, Department of Hepato-PancreatoBiliary Surgery, Beijing, China Background: CCN is an acronym of CYsteine-Rich protein 61 (CYR61), Connective Tissue Growth Factor (CTGF) and Nephroblastoma OVerexpressed (NOV) comprising another three members i.e. Wnt1-inducible signalling pathway protein (WISP)-1, -2 and -3. Clinical studies have revealed differential roles of certain CCN family members in cancers which can be cancer specific or dependent on local microenvironment. CTGF has been evident to promote growth of pancreatic cancer cells under hypoxia. The role played by CYR61 and NOV in pancreatic cancer remains unclear. The present study is aimed to determine the expression of CYR61, CTGF and NOV in pancreatic cancer and the corresponding clinical implications. Materials and Methods: Pancreatic tumour samples (n = 230) together with paired background pancreatic tissues were collected at Peking University Cancer Hospital immediately following the surgery with written consent from the patients. Protocol and procedures of the tissue collection

Abstracts were approved by Peking University Cancer Hospital Research Ethics Committee. RNA was extracted and converted into cDNA using a reverse transcription kit. Transcripts of CYR61, CTGF and NOV were then determined using real time PCR respectively. The association with clinical outcomes and other clinic-pathologic parameters was analysed using SPSS. Results: An increased expression of both CYR61 and CTGF was evident in pancreatic cancer tissues compared with their expression in normal background tissues respectively (P < 0.05). However, the local advanced tumours (T3 and T4) expressed lower levels of CTGF transcripts compared with tumours at earlier stages (T1 and T2), p = 0.036. Similarly, an association exits between increased expression of NOV and the local invasion though no difference was seen for NOV expression in the tumours compared with the background tissues. There is no association of CYR61 expression with histological and pathological features observed in the current cohort. The expression of the three genes is not relevant to the survival of patients when each was analysed individually. However, tumours expressed higher levels of three CCNs (CYR61, CTGF and NOV) had longer overall survival (P = 0.034). Conclusions: CYR61 and CTFG are up-regulated in pancreatic cancer. CTGF and NOV may contribute to the local invasive growth of pancreatic tumours. However, patients with tumours highly expressing of these three genes exhibited longer survival together with molecular and cellular mechanism require further clarification and investigation. No conflict of interest. 2217 POSTER The role of cyclin D1 and p21(WAF1/CIP1) in gastric cancer V. Michalaki1 , G. Frangulidis1 , A. Kondi-Pafgiti2 , J. Papakonstandinou3 , A. Vezakis4 , C. Papadimitriou4 . 1 University of Athens Areteion Hospital, 2Nd Department Of Surgery, Athens, Greece; 2 University of Athens Areteion Hospital, Histopathology Department, Athens, Greece; 3 University of Athens Areteion Hospital, 2Nd Department Of Surgery, Athens, Greece; 4 University of Athens Areteion Hospital, 2Nd Department Of Surgery, Athens, Greece Background: The deregulation of cyclin, cyclin-dependent kinases (CDKs) and their inhibitors could have a crucial role in the development of diverse human cancers. Alterations in cell cycle regulators and subsequent deregulation of the cell cycle are frequently involved in tumorigenesis and/or tumor progression. The aim of our study was to detect the abnormal expression of cyclin D1 and p21(WAF1/CIP1)in gastric carcinoma and investigate its clinicopathologic significance. Methods: Proteins of cyclin D1 were detected by immunohistochemistry in 80 cases of advanced gastric carcinoma, and 30 cases of benign gastric diseases (chronic gastritis, atrophic gastritis, gastric metaplasia, and gastric dysplasia). From each patient formaldehyde–fixed paraffin sections were stained and examined by immunohistochemistry using monoclonal antibodies.All tumor cells with distinct nuclear staining were considered positive. Results: Sixty-five patients were male and forty-five female. Normal gastric epithelium showed consistently positive immunostain for p21WAF1/CIP1. Loss of p21WAF1/CIP1 expression was noted in 65% of intestinal type adenocarcinoma and in 90% of diffuse type adenocarcinoma. Overexpression of cyclin D1 was detected in 90% of advanced gastric carcinomas. Among the various clinicopathological findings, overexpression of cyclin D1 was associated with lymph-node metastasis (P = 0.003) and recurrence (P = 0.044). Loss of p21WAF1/CIP1 expression was more frequent in diffuse type cancers (P = 0.005) and was correlated with recurrence (P = 0.002) and death (P = 0.001). Conclusions: These findings suggest that overexpression of cyclin D1 is a frequent finding in gastric cancer and immunohistochemical analysis for cell cycle regulators, might be a useful prognostic indicator in gastric cancer. No conflict of interest. 2218 POSTER Genetic and clinicopathologic analysis of gastric cancer in Colombia J.J. Suarez Olaya1 , R. Prieto2 , M. Bohorquez2 , M.M. Echeverry2 , ´ Colombia; L.G. Carvajal3 . 1 Universidad del Tolima, Tolima, Ibague, 2 ´ Colombia; 3 University of California University of Tolima, Tolima, Ibague, Davis, California, Davis, USA Background: The susceptibility to develop gastric carcinoma (GC) has been associated with the genetic characteristics of the individuals and some risk factors such as alcoholism, smoking, diet and infections. This disease ranks third in lethality in the world (723.073 cases, 8,8%); in Colombia, it is the leading cause of death from cancer in men (3.051

S405 cases, 16,2%) and the third in women (1.930 cases, 10,1%). Its etiology and histopathology have not been completely understood. Materials and Methods: The association of polymorphisms of CYP1A1, CYP2E1, CYP26B1, CDH1, PSCA and MUC1 genes was studied in 160 Colombian patients diagnosed with GC. These results were compared with 164 healthy individuals with no family history of the disease and 139 individuals of the ethnic groups Nasa-Paez and Pijaos, in order to establish their risk of developing the disease. Risk analysis for carriers of susceptible genotypes was performed using Plink 1.07. Results: Greater frequency of the disease was found in men than in Women (2:1 ratio) and in people over 50 years old, coinciding with previous studies. Interestingly, a higher percentage (21,9%) of younger individuals affected by the disease (average 39,5 years old, ranging from 17−49 years old) was observed. These results are above previous reports for the South American population (2.4−8%). The distribution of the GC cases included 45,6% Intestinal, 37,6% diffuse (higher than previous reports of 5%-15%) and 16,8% mixed GC. Stages TNM III and IV were diagnosed in 50,6% of patients. Comparative analysis of genetic markers between cases and control groups showed a relationship between the presence of the mutant PSCA gene and patients with diffuse and mixed GC. However, mutations of MUC1 and CYP2E1 genes seem to have a protective effect against the risk of developing intestinal GC. Interestingly, in the indigenous populations, mutations of MUC1, CYP26B1 and PSCA genes seem to increase the susceptibility to develop intestinal, diffuse and mixed GC. However, polymorphism in CYP1A1 y and CYP2E1 seem to be associated with a protective effect from malignant gastric lesions in those indigenous populations. Moreover, analysis of the CDH1 gene, showed no association with the susceptibility to develop the disease in the groups studied. In contrast, alcohol consumption, smoking and low socioeconomic status showed association with this pathology. Conclusions: The results of this study indicate a possible association between the polymorphism of some genes and the susceptibility to intestinal, diffuse and mixed GC. This association may partly explain the lower incidence of CG in native populations. No conflict of interest. 2219 POSTER Intratumoral injection of the oncolytic immunotherapeutic Pexa-Vec (JX-594) in liver tumors and hepatocellular carcinoma: Recommendations for clinical practice R. Lencioni1 , C.W. Kim2 , S.C. Rose3 , C.J. Breitbach4 , J.M. Burke5 , T. Hickman6 , D.H. Kirn7 , N. Stojkowitz8 , M. Lusky9 , M. Homerin10 . 1 Pisa University School of Medicine, Division of Diagnostic Imaging and Intervention, Pisa, Italy; 2 Pusan National University School of Medicine, Interventional Radiology, Busan, South Korea; 3 UCSD Medical Center, Interventional Oncology section, Hillcrest, USA; 4 Sillajen Inc., Clinical and Translational Research, San Francisco, USA; 5 Sillajen Inc., Clinical Research, San Francisco, USA; 6 Jennerex Biotherapeutics, Clinical Operations, San Francisco, USA; 7 Jennerex Biotherapeutics, Clinical Research, San Francisco, USA; 8 Transgene S.A., Clinical Operations, Illkirch-Graffenstaden, France; 9 Transgene S.A., Business Development, Illkirch-Graffenstaden, France; 10 Transgene S.A., Medical Affairs, Illkirch-Graffenstaden, France Background: Pexastimogene devacirepvec (Pexa-Vec, JX-594) is a vaccinia virus engineered to preferentially replicate in and lyse cancer cells, and to express the therapeutic transgene granulocyte macrophage colonystimulating factor (GM-CSF) to stimulate anti-tumor immunity. Pexa-Vec properties also include systemic exposure after intratumoral (IT) injection and antivascular effects against tumors. Experience from previous clinical trials with IT Pexa-Vec is summarized. Material and Methods: The feasibility, safety and efficacy of percutaneous IT injections of Pexa-Vec, using a needle with multiple side-holes or a dedicated multi-pronged needle (Quadra-Fuse® , Rex-Medical, PA, USA), was investigated in 4 trials involving 155 patients: a dose-escalation phase I study in patients with primary or metastatic liver tumors (n = 14) and 3 phase 2 studies in hepatocellular carcinoma (HCC): a randomized dose-finding phase 2 study (n = 30), a randomized Best Supportive Care-controlled phase 2 study in sorafenib-experienced patients (n = 129) and a single-arm study combining Pexa-Vec and sorafenib (n = 25). The goal of the injection procedure was to ensure a controlled, homogenous, 3-dimensional distribution of Pexa-Vec suspension into tumor tissue. Results: The Maximal Tolerated Dose for IT Pexa-Vec was identified to be 109 plaque forming units (pfu) and three IT 109 pfu injections was determined to be the effective regimen for further development in HCC. Most patients experienced transient, mild to moderate fever