GLRX3, a novel secretory biomarker of pancreatic cancer based on pancreatic cancer stem cell characteristics

GLRX3, a novel secretory biomarker of pancreatic cancer based on pancreatic cancer stem cell characteristics

Abstracts / Pancreatology 13 (2013) S1–S80 contributing to T-helper cell mediated inflammation and early b-cell dysfunction. Keywords: Stellate cells,...

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Abstracts / Pancreatology 13 (2013) S1–S80

contributing to T-helper cell mediated inflammation and early b-cell dysfunction. Keywords: Stellate cells, Pancreatic islets, Beta cell dysfunction, Chronic pancreatitis

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and biological roles of this important family of epigenomic oncogenes in PDAC. Keywords: EZH2, Epigenetics, Pancreatic cancer

[O 05]. GLRX3, a novel secretory biomarker of pancreatic cancer based on pancreatic cancer stem cell characteristics Sun a Kim, Soo Been Park, Si Young Song Internal Medicine, Yonsei University College of Medicine, Seoul, South Korea

[O 04]. Studies on polycomb identify new EZH2-type epigenetic oncogene for pancreatic cancer Gwen Lomberk, Raul Urrutia Laboratory of Epigenetics and Chromatin Dynamics, GI Research Unit, Department of Medicine, Mayo Clinic, Rochester, MN, United States Background/aim: Histone methyltransferase EZH2, one of the best characterized epigenomic oncogenes, forms an obligate Polycomb repressive complex with SUZ12 and EED. Polycomb-mediated repression influences gene expression across the entire gamut of biological processes. Deregulation of EZH2 expression has been implicated in human PDAC. However, to date, most EZH2-mediated function has been ascribed to a single protein product of the EZH2 locus. Methods: PCR and western blot were used to detect the transcripts and protein products, respectively. Immunoprecipitation and H3-K27me3 ELISA were used to characterize the functional complex. Localization of proteins was observed by immunofluorescence with specific antibodies. EZH2a or EZH2b-carrying adenovirus was used to infect cells prior to RNA extraction for microarray analysis. Results: We report that the EZH2 locus undergoes alternative splicing to yield at least two structurally and functionally distinct EZH2 methyltransferases. The longest protein is the conventional enzyme, EZH2a, while EZH2b, characterized here, represents a novel isoform. We find that EZH2b is overexpressed in PDAC, localizes to the nucleus, complexes with EED/SUZ12, deposits the H3-K27me3 mark, and mediates gene silencing. Increased levels of EZH2b also induce cell proliferation, demonstrating that this protein is functional in the regulation of processes previously attributed to EZH2a. Through genome-wide expression profiling, we demonstrate that EZH2b governs a pattern of gene repression that is often ontologically redundant from that of EZH2a, but also divergent for a wide variety of target genes. Lastly, enhanced expression of EZH2b, as observed in the human situation, enhances the growth of PDAC xenografts. Conclusions: These results characterize EZH2b as novel pancreatic oncogene. These data support the notion that the regulation of EZH2-mediated gene silencing, with an expanded repertoire of EZH2 writers, is more complex than previously anticipated and should guide future studies aimed at understanding the biochemical

Background/aim: Pancreatic cancer is one of the most lethal diseases, which is difficult to diagnose and resistant to conventional treatment such as chemotherapy and radiotherapy. Cancer stem cells are involved in carcinogenesis, cancer progression and recurrence. We tried to find secretory biomarkers associated with pancreatic cancer stem cells using proteomic analysis. Methods: Pancreatic cancer stem like cells were enriched using sphere culture method. Proteomic analysis was done with secreted protein in culture medium of sphere and adherent cells. Identified proteins were confirmed with western blot, immunohistochemical staining and ELISA. shRNA for GLRX3 was used to study the function of GLRX3 in pancreatic cancer. Results: Proteomic analysis showed that total of 55 protein spots from sphere cells were increased compared to adherent cells. Among them, 41 spots were identified using MALDI-TOF. Proteins known to be associated with cancer stem cells such as HSP90AB1, ALDH, and vimentin were expressed in sphere cells. The overexpression of 5 proteins including GLRX3 was confirmed by western blot in sphere cells and patient serum. Immunohistochemical staining in TMA of human pancreatic cancer tissue revealed that GLRX3 was detected in 15 (57.7%) out of 25 pancreatic cancer tissues regardless of cancer stages. To examine GLRX3 involvement in pancreatic carcinogenesis, we used shRNA to generate GLRX3knockdown cells. Compared to the mock, knockdown of GLRX3 in human pancreatic cancer cell lines decreased in vitro proliferation, clonogenecity, and sphere formation. GLRX3 knockdown reverses EMT in pancreatic cancer cells. ELISA showed patients with unresectable pancreatic cancer and high blood GLRX3 levels had poor survival compared to those with low blood GLRX3 levels (155 days vs 463 days, pvalue <0.000). Conclusions: This study shows that secretory protein GLRX3 may be a useful prognostic marker in pancreatic cancer. More researches are needed to find their roles in cancer stem cells and clinical implication. Keywords: GLRX3, Pancreatic cancer, Cancer stem cell, Marker

[O 06]. Prognostic impact of tumor infiltrating regulatory T cell (Foxp3D) to activated cytotoxic T lymphocyte (granzyme BD) ratio in resected left-sided pancreatic cancer Ho Kyoung Hwang, Hyoung-il Kim, Chang Moo Kang, Kyung Sik Kim, Dong Sup Yoon, Woo Jung Lee Surgery, Yonsei University College of Medicine, Seoul, South Korea Background/aim: Tumor infiltrating activated cytotoxic T lymphocyte (Granzyme Bþ) has been reported to have an anti-tumor effect; on the other hand, Regulatory T cell (Foxp3þ) has a suppressive role in antitumor immune response. The aim of this study was to investigate the correlation between the balance of different types of tumor infiltrating lymphocytes (TILs) and prognosis in resected left-sided pancreatic cancer.