Abstract PO-055: Signaling regulation of epithelial-mesenchymal transition in the hypoxic tumor microenvironment of pancreas cancer

Abstract

Abstract Epithelial-mesenchymal transition (EMT) is a normal developmental process by which epithelial cells lose their polarity and cell-cell adhesions to become more motile and invasive mesenchymal cells. EMT is associated with the earliest stages in metastasis and promotes chemoresistance in a variety of carcinomas, including pancreatic ductal adenocarcinoma (PDAC). While specific growth factors (e.g., transforming growth factor beta) are perhaps the most well studied inducers of EMT, the low oxygen tension (hypoxia) characteristic of the PDAC tumor microenvironment has also been reported to drive EMT. The signaling pathways that promote EMT in the hypoxic PDAC tumor microenvironment may therefore represent useful drug targets for combination therapy approaches as adjuvants to augment the efficacy of chemotherapy. However, hypoxia-driven EMT has not been well characterized in PDAC, nor have the relevant hypoxia-driven signaling pathways been identified. Our lab recently developed a statistical model of the multivariate signaling regulation of growth factor-induced EMT to identify that the ERK, JNK, and NF-kappaB pathways coordinate to drive EMT in PDAC cells. In the present study, we sought to determine the roles of those pathways in promoting EMT in PDAC cells in response to hypoxia. We created an in vitro model of hypoxia-driven EMT by culturing PDAC cells in 1% oxygen and characterizing dynamic changes in the expression of hypoxia inducible factors (HIF) 1 and 2 alpha, as well as the concomitant loss of E-cadherin protein and transcripts, increased abundance of vimentin-positive cells, and increased expression of the mesenchymal transcripts VIM, SNAI1, and SNAI2. Hypoxia-driven EMT was observed in human PDAC cell lines, patient-derived xenograft cell lines, and KPC murine cell lines. To probe for the activation of specific signaling pathways, we used a combination of immunoblotting and immunofluorescence microscopy. Automated image analysis of immunofluorescence images was used to understand the relevance of specific signaling pathways for explaining the heterogeneity of EMT within populations of cells. We further identified potential kinase targets for antagonizing EMT in the hypoxic microenvironment and confirmed their roles by pharmacological inhibition and/or siRNA-mediated knockdown. We also found that specific kinase pathways are involved in the stabilization of HIF proteins in hypoxia. The relevance of these pathways in driving EMT is being further validated through analyses of a patient-derived xenograft mouse model of PDAC and a HIF1A deletion/Kras mutant autochthonous mouse model of PDAC. Citation Format: Brooke A. McGirr, Nicholas M. Seyler, Matthew J. Lazzara. Signaling regulation of epithelial-mesenchymal transition in the hypoxic tumor microenvironment of pancreas cancer [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2020 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2020;80(22 Suppl):Abstract nr PO-055.