Abstract 2891: Gastrin signaling and Par-4 in pancreatic cancer
Authors: Kristin K. Fino, Rosalyn B. Irby, Jill P. Smith
Journal: Cancer Research (2011)

Abstract Background: The gastrointestinal peptide, gastrin, has been identified as a growth factor in pancreatic cancer. Gastrin stimulates growth of pancreatic cancer through the cholecystokinin (CCK)-2 receptor. Prostate apoptosis response-4 (Par-4) is a tumor-suppressor protein that induces apoptosis in cancer cells but not in normal cells. Par-4 is down-regulated in a variety of cancers, including pancreatic where it is correlated with lower rate of survival. The anti-apoptotic activity of Par-4 is reduced when active Akt1 binds and phosphorylates it. In colon cancer cells, Par-4 also binds to NFκB and repressed NFκB gene transcription. Gastrin stimulation mediated through the CCK-2R regulates NFκB and Akt activity. Hypothesis: Stimulation of human pancreatic cancer cells with gastrin affects Par-4 activity which may be mediated through Akt. Methods: PANC-1 cells were serum starved for 16-hours, pretreated with a CCK-2R specific inhibitor (YM022) or vehicle for 1hr, and then stimulated with 10-8 M gastrin-17 for 20min. Cells were immediately lysed and the whole cell lysate was subjected to immunoprecipitation with magnetic Dynabeads bound to antibodies specific for Par-4 or NFκB(p65). Immunoprecipitation samples were probed for Par-4, and NFκB(p65), and cell lysates were probed for Par-4, NFκB(p65), Akt, phospho-p65 (Ser536), and phospho-Akt (Ser473). PANC-1 cells were transfected with siRNA specific to the CCK-2R or appropriate scrambled control delivered by Lipofectamine 2000. Protein was isolated 72 hours after transfection. Results: Pretreatment of PANC-1 cells with YM022 before gastrin stimulation blocked phosphorylation of Akt while total levels of Akt remained unchanged. siRNA knock-down of CCK-2R also resulted in a decrease of Akt activation. Phosphorylation of NFκB(p65) was increased in cells pretreated with YM022 compared to cells treated with gastrin alone; total levels of NFκB(p65) remained unchanged. Binding of Par-4 to NFκB(p65) was enhanced in cells treated with both gastrin and YM022, compared to cells treated with gastrin alone. Total levels of Par-4 remained unchanged. Conclusion: For the first time we provide evidence supporting that gastrin stimulation of pancreatic cancer cells alters activity of Par-4. Gastrin stimulation decreases binding of Par-4 to NFκB(p65) in PANC-1 cells while pretreatment with YM022 blocks this decrease in binding. The change in Par-4 activity may be mediated through Akt because YM022 and gastrin stimulation alters their activity and they are known binding partners of Par-4. Since Par-4 sensitizes cells to apoptosis induction as exemplified by sensitization of colon cancer cells to the chemotherapeutic agent, 5-fluorouracil, when Par-4 is delivered in vivo, understanding of the mechanism by which Par-4 is down-regulated is important in developing therapies to take advantage of Par-4 induced sensitization to apoptosis. Supported by: R01 CA117926 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2891. doi:10.1158/1538-7445.AM2011-2891