Abstract A78: PAR-2 activation with ATP facilitation induces migration but not proliferation of pancreatic cancer cells.

Abstract

Introduction: Protease-activated receptor 2 (PAR-2) is a G protein-coupled receptor that functions as a cell-surface sensor for coagulation factors, as well as other proteases associated with the tumor microenvironment. The activation of PAR-2 has been shown to induce proliferation of many cancer cells whereas it also may induce migration. The role of PAR-2 in pancreatic cancer remains largely unexplored however. Aim: The aim of the present study is to examine whether PAR-2 activation induces proliferation and/or migration of pancreatic cancer cells and to elucidate the underlying mechanism. Materials and Methods: BxPC-3 and Capan-2 pancreatic cancer cell lines were stimulated with PAR-2 agonist peptide or trypsin after which proliferation was assessed by BrdU assays and migration was assessed by trans-well and scratch assays. To identify the underlying signal transduction pathways, the phosphorylation status of Erk1/2 and Src was determined by Western blot analysis. Moreover, specific inhibitors of the different signal transduction pathways were tested in the migration experiments. Results: PAR-2 activation does not induce proliferation of both pancreatic cancer cell lines tested. Interestingly, cell stimulations with PAR-2 agonist peptide and trypsin did induce migration of the pancreatic cancer cells in scratch assays but not in a trans-well setting. Addition of inhibitors of PKC (Chelerythrine), MEK (U0126), Rac (NSC23766) and Src (pp1) all inhibit PAR-2 dependent migration whereas these inhibitors do not inhibit FCS driven migration. In line with these findings, stimulation of BxPC3 and Capan-2 cells does induce phosphorylation of ERK1/2 and Src. Preliminary data suggest that the seeming discrepancy between scratch assays and trans-well experiments may be due to increased metabolic activity of cells in close proximity of the scratch. It is well-known that upon scratching the monolayer of cells, ATP and Zn2+ will be released for metabolic activity. By addition of ATP inhibitor, activating of PAR-2 no longer drives cell migration in neither of the cell line. Ongoing experiments should however elucidate further of the interplay between PAR-2 and ATP with respect to cancer cell migration. Conclusion: We show that activation of PAR2 has no effect on proliferation of pancreatic cancer cells but significantly induces migration in scratch assays. We suggest activation of the PKC/Src/Rac/MEK pathway to drive PAR-2 induced pancreatic cancer cell migration. Overall our data suggest that activated PAR-2, with the facilitation of ATP may be important for pancreatic cancer metastasis in an ERK-dependent manner. Citation Format: Kun Shi, Karla C.S. Queiroz, C. Arnold Spek. PAR-2 activation with ATP facilitation induces migration but not proliferation of pancreatic cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Progress and Challenges; Jun 18-21, 2012; Lake Tahoe, NV. Philadelphia (PA): AACR; Cancer Res 2012;72(12 Suppl):Abstract nr A78.