CD95-related apoptotic machinery is functional in pancreatic cancer cells

The CD95 (FAS, Apo-1) system is a major death pathway in normal and tumor c ells. Recent evidence indicates that pancreatic cancer cells express CD95R and CD95L but are insensitive to CD95-mediated apoptosis. Here we show that treatment of human pancreatic cancer cells with RNA synthesis inhibitor ac tinomycin D (ActD) converted the phenotype of cancer cells from CD95 resist ant to CD95 sensitive. Flow cytometric analysis demonstrated that all pancr eatic cancer cell lines studied responded with cell surface CD95R and CD95L upregulation to bleomycin treatment, and PANC1 (mt p53) cells demonstrated a dose-dependent response to interferon gamma and bleomycin treatment with CD95R and CD95L upregulation. However, only bleomycin sensitized PANC1 cel ls to CD95-mediated apoptosis. Taxol sensitized PANC1 and HPAC cells to CD9 5-mediated apoptosis without surface upregulation of CD95R. These data sugg est that pancreatic cancer cells possess a p53-independent mechanism of CD9 5R and CD95L surface upregulation and that surface expression of CD95R is n ot predictive of apoptotic function. Protein extracts of HPAC and PANC1 cel ls treated for 24 hours with a combination of ActD/agonist anti-CD95 antibo dies demonstrated significantly higher Acetyl-Asp-Glu-Val-Asp-ase (DEVDase) cleavage activity (caspase 3-like activity) than extracts from cells treat ed with ActD only. In the present study, we also investigated the time kine tics of DEVDase (caspase 3-like) activation in PANC1 (mt p53) and HPAC (wt p53) pancreatic cancer cell lines. We found that DEVDase activity in PANC1 cells responds to ActD and ActD/anti-CD95 antibodies earlier than in HPAC c ells; however, at 24 hours HPAC cells demonstrated much stronger activation . Cytosolic protein extracts from untreated cells did not influence caspase 3-like activity when added to extracts from the ActD/anti-CD95 antibody-tr eated cells. Collectively, these data suggest that pancreatic cancer cells have functional CD95-related apoptotic machinery with preserved apoptotic s ignal transduction, CD95R upregulation, and caspase activation. However, th is system is blocked by some unknown protein(s) that is either located in t he organelle fraction of the cell and/or requires an intact cell for manife station of its activity.