Abstract A52: Hypoxia inducible factor-1 alpha (HIF-1α) promotes migration and invasion of pancreatic cancer regulated by CD133 under hypoxia.

Abstract

Background: Pancreatic cancer is a highly lethal disease and it is little sensitive to any current therapy. Over 17,000 patients died of pancreatic cancer, which is the fifth leading cause of cancer-related death in Japan. Multimodal treatments have completely failed to eradicate pancreatic cancer. A potential reason for the failure of the conventional therapeutic approach for pancreatic cancer might be explained by cancer stem cell (CSC) theory. CD133 (prominin-1) is a 5-transmembrane glycoprotein, which is well known as one of CSC markers in pancreatic cancer. We previously reported that CD133 expression in pancreatic cancer is correlated with poor prognosis and lymph node metastasis[1]. However, the function of CD133 is still obscure. On the other hand, pancreatic cancer is surrounded with desmoplastic morphology, which is under the low-oxygen condition because of its insufficient blood supply. In the present study, we hypothesized that CD133 plays a key role in invasion and metastasis for pancreatic cancer associated with HIF1-α expression. Materials and Methods: 1) We investigated the relationship between HIF-1α or CD133 expression in primary tumor tissues and survival for pancreatic cancer patients. 2) To clarify the mechanism of these relationships, we established a highly migratory cell line, Capan1M9, which is a subpopulation derived from Capan-1 cells by migration assay system. Over 90% of Capan1M9 cells express CD133[2]. Consequently, we established Capan1M9-GFP-shCD133 cells (CD133 knockdown) by lentiviral transduction method. Using these cell lines, other methods such as migration and invasion assays, wound healing assay, immunofluorescence staining, Western blot, and real-time quantitative RT-PCR were performed. Results: 1) Immunohistochemical study showed that HIF-1α expression positively correlated with poor prognosis and microvessel density in pancreatic cancer samples. 2) CD133+ cell population in Capan-1 showed more resistant to hypoxic condition (1% O2 and 0.1% O2) than CD133cell population by FACS analyses. This result indicated that CD133 contributed to the resistance of cancer cells in hypoxia. 3) We compared CD133+ cells (Capan-M9) and CD133-cells (shRNA-CD133-M9) under 1% O2 hypoxia. In hypoxia, CD133+ cells showed higher expression of HIF-1α than CD133cells. HIF-1α accumulation was observed in both of the nuclei of CD133+ and CD133cells under hypoxia. 4) HRE response was also showed high level in Luc-CD133+ cells than Luc-CD133-cells by luciferase assay in hypoxia. 5) Migration ability of CD133+ cells was higher than that of CD133-cells by wound healing assay and migration assay in hypoxic conditions. 6) Epithelial-mesenchymal transition (EMT)-related molecules such as Slug, N-cadherin and fibronectin were higher in Capan1-M9 (migratory subclone cells) than in Capan-1 (parental cells). Conclusion: These results indicated that HIF-1α stimulates migration and invasion of pancreatic cancer associating with EMT and CD133 expression under hypoxia.