Abstracts
1998 Digestive Disease Week

#4472

HER2/NEU OVEREXPRESSION INTERFERES WITH RETINO-BLASTOMA PROTEIN-MEDIATED CELL CYCLE REGULATION IN HUMAN PANCREATIC CARCINOMA CELLS. H. Roh, J. Pippen and J.A. Drebin, Department of Surgery, Washington University School of Medicine, St. Louis, MO.

Introduction: Overexpression of the HER2/neu oncogene is observed in 60-70% of human pancreatic carcinomas. The effects of down-regulating the expression of the HER2/neu gene product, p185, using antisense (AS) oligonucleotides, on the proliferation and cell cycle progression of the human pancreatic carcinoma cell line Panc1 were examined. Methods: Panc1 cells, which overexpress HER2/neu, were obtained from ATCC. AS oligonucleotides, as well as sense (S) and scrambled antisense (SC) controls were added to cultures at a final concentration of 1 uM in the presence of 10 ug/ml lipofectin for 4 hours. Cell cultures were then placed in fresh media. Effects on HER2/neu expression were determined by northern and western blotting. Expression of p21cip, p27kip and the phosphorylated and unphosphorylated forms of the retinoblastoma (Rb) protein were determined by western blotting. Cell proliferation was examined by BRdU-uptake ELISA. Cell cycle distribution was determined using dual channel flow cytometry in the presence of propidium iodide following pulsed BRdU exposure.

Results: HER2/neu AS treatment significantly inhibits HER2/neu mRNA and p185 protein expression in Panc1 cells; S and SC control oligonucleotides have no effect on HER2/neu expression. AS mediated down-regulation of HER2/neu expression causes upregulation of the cyclin-dependent kinase inhibitors p21cip and p27kip and a resulting decrease in Rb protein phosphorylation. Treatment of Panc1 cells with HER2/neu AS oligonucleotides results in > 80% inhibition of Panc1 cell proliferation as determined by BrDU incorporation. Control oligonucleotides have no significant effect on cell proliferation. AS oligonucleotide treatment results in a blockade of cell cycle progression with a corresponding accumulation of cells in G1:

Interestingly, these effects are independent of the p16 and p53 tumor suppressor genes, which have undergone genetic deletions in Panc1 cells.

Conclusions: HER2/neu overexpression contributes to the loss of cell cycle control in human pancreatic carcinoma cells by suppressing the expression of p21cip and p27kip with resulting effects on Rb phosphorylation. Strategies which down-regulate HER2/neu expression may be of use in the therapy of some pancreatic tumors.

Copyright 1996 - 1998, SSAT, Inc. Revised 29 June 1998.