P53 Dependence of Fas Induction and Acute Apoptosis in Response to 5-fluorouracil-leucovorin in Human Colon Carcinoma Cell Lines

We examined the patterns of induction of apoptosis, Fas expression, and the influence of the status of the p53 tumor suppressor gene, in response to treatment of human colon carcinoma cell lines to 5-fluorouracil (FUra) combined with leucovorin (LV) under conditions of both DNA-directed (HT29, VRC5/c1, and RKO) and RNA-directed (HCT8 and HCT116) cytotoxicity. Acute apoptosis was induced in cell lines expressing wtp53 (RKO, HCT8, and HCT116), independent of the mechanism of FUra action. In HT29 cells that expressed mp53, apoptosis was a delayed event. Cell lines undergoing DNA-directed FUra cytotoxicity demonstrated marked accumulation of cells in S-phase (HT29 and RKO), whereas those lines undergoing RNA-directed cytotoxicity (HCT8 and HCT116) demonstrated marked cell cycle phase arrest in G2-M, both reversible by dThd. dThd partially protected HCT8 and HCT116 cells from FUra-LV-induced apoptosis but had no influence on FUra-LV-induced loss in clonogenic survival. In cells expressing wtp53, the Fas death receptor was induced in response to FUra-LV treatment. FUra-LV sensitized RKO cells to the anti-Fas monoclonal antibody CH-11 that was completely reversed by dThd, demonstrating the involvement of DNA damage in FUra-LV-induced, Fas-dependent sensitization to CH-11. In contrast, FUra-LV sensitized HCT116 cells to CH-11-induced apoptosis, which was not dThd reversible. Transduction of HT29 cells with Ad-wtp53 induced elevated Fas expression and sensitized the cells to FUra-LV-induced apoptosis. Data indicate that the presence of a wtp53 gene determines FUra-LV-induced Fas expression, the kinetics of FUra-LV-induced apoptosis and not the extent of apoptosis induced, both being independent of the mechanism of FUra action. Therefore, in colon carcinomas that express wtp53, the approach to sensitize tumors to Fas-mediated apoptosis may be further enhanced from the effect of FUra-LV in elevating Fas expression in a p53-dependent manner.