B190
Gamma secretase as a potential therapeutic target in pancreatic adenocarcinoma
Chris Mann1, Chris Neal1, Chris Briggs1, Mai-Kim Cheng1, William Steward1, David Berry2, Maggie Manson1
1Department of Cancer Studies and Molecular Medicine, University of Leicester, Leicester, UK, 2Department of Hepatobiliary and Pancreatic Surgery, University Hospitals of Leicester, Leicester, UK
Background
Pancreatic carcinoma has a dismal prognosis and is largely resistant to current chemotherapeutic agents. The Notch signalling pathway plays a role in carcinogenesis of several solid malignancies. The aim of this study was to determine the role of Notch in pancreatic carcinoma and investigate its potential therapeutic modulation by inhibiting gamma secretase, the enzyme responsible for pathway activation.
Method
Four pancreatic adenocarcinoma cell lines were used (ASPC-1, BxPC-3, MiaPaCa-2, PANC-1) to examine the effect of gamma secretase inhibition (GSI-I, Calbiochem, UK) on cell proliferation (Coulter counter), viability (ATP assay), apoptosis (Annexin V binding) and cell-cycle (FACS analysis).
Results
All cell lines expressed Notch pathway constituents. GSI-I inhibited activation of Notch-1, -3, and -4, as well as expression of the target gene Hes-1 in a dose-dependent manner. Treatment with 0.1-5μM also dose-dependently reduced proliferation in all cell lines, associated with G2-M cell-cycle arrest, and resulted in reduction in viability and increased apoptosis. All lines entered apoptosis 4-8 hours after treatment, with >90% of cells necrotic by 72 hours after treatment with 5μM GSI-I. At doses as low as 0.1μM, GSI-I induced apoptosis at 24 hours. Combination of GSI-I and Gemcitabine resulted in an additive reduction in cell viability and increase in apoptosis, compared to treatment with either agent alone. Knockdown of Notch 1 with siRNA significantly decreased viability in the three lines expressing this form.
Conclusion
The Notch pathway is important in the pathogenesis of pancreatic adenocarcinoma. This data demonstrates that modulation of this pathway using gamma secretase inhibitors is a promising therapeutic approach.
