The role of Notch3 and related pathways in pancreatic cancer
Introduction: Pancreatic cancer is a malignancy with a very poor prognosis, and therefore insights into the derangement in cell signalling pathways which result in cancer development may provide important information in understanding the disease process, and in developing future therapeutic strategies. The aim of this study was to examine the expression and potential importance of several intracellular signalling proteins. Particular attention was given to Notch3. Methods: An immunohistochemical study was performed on 23 samples of human pancreatic adenocarcinorna and 12 samples of normal pancreas, to assess expression of Notch3, cyclin D1, pAkt, STAT3 and pSTAT3, β-catenin, and pin1. The degree of staining, and intracellular locations of the proteins were assessed. Immunohistochemical data were correlated with clinical parameters, including tumour resectability, size, presence of metastases, and plasma levels of CA19-9. In vitro studies were also performed on two pancreatic cell lines. Basal levels of protein expression were assessed by western blotting. The effect of Notch inhibition by γ-secretase inhibitors and by transfecting cells with Notch3 siRNA was also examined. Cell survival studies were performed using the Annexin V apoptosis assay. Results: Pin1 was not overexpressed in pancreatic adenocarcinoma. However, Notch3 was significantly overexpressed in the cytoplasm of 73.9% of tumours. Nuclear expression, never observed in normal ductal tissue, was noted in 43.5% of cases. No tumour expressing nuclear Notch3 was surgically resectable, and there was a significant correlation between nuclear Notch3 staining and increasing levels of CA19-9. There were also significant correlations between expression and intracellular location of Notch3 and each of STAT3, pSTAT3, and pAkt, but not cyclin D1. Notch3 was expressed in untreated ASPC1 cells, but not in PANC1 cells. Its levels could be reduced using either γ-secretase inhibitors or siRNA. Notch inhibition also resulted in a fall in levels of pAkt, but initial experiments did not demonstrate an effect upon cell survival. Conclusion: The presence of Notch3 in tumour nuclei is likely to represent functional activation of the protein, and is clearly linked to more aggressive tumours. In the future, targeting the Notch pathway may prove to be of therapeutic benefit.