We examined the effect of specific inhibitors of DNA polymerases alpha and delta, and beta, on cisplatin (DDP) cytotoxicity in DDP-sensitive and -resistant human 2008 ovarian carcinoma cells. Under conditions of continuous exposure to drug combinations, neither aphidicolin glycinate (AG) nor dideoxythymidine enhanced the cytotoxicity of DDP in either cell line as determined by clonogenic survival assays. However, when clonogenic survival was determined following short-term drug exposure, AG exhibited strong synergism with DDP in the DDP-resistant, but not the DDP-sensitive cells, as indicated by median effect analysis of the data. DNA polymerase alpha mRNA levels were the same in both cell lines under basal conditions. DDP-sensitive cells, but not DDP-resistant cells, were able to increase their expression of DNA polymerase alpha in response to DDP exposure. Levels of mRNA for DNA polymerase beta and for the human DNA repair gene ERCC-1 were not elevated in resistant cells, either under basal conditions or 18 hr after a 1 hr exposure to IC20 concentrations of DDP. In another human ovarian carcinoma cell line, A2780, AG and DDP were synergistic in both DDP-sensitive and -resistant variants in short-term exposure. We conclude that DNA polymerases alpha and/or delta play a role in the DDP sensitivity of human ovarian carcinoma cells.