We have previously shown that p53 disruption sensitizes certain cancer cell types to cisplatin (CDDP) (Fan et al., 1995). In the present study we investigated the role of the p53 downstream effector, p21CIP1/WAF1 (p21), in this sensitization. Studies were performed in human colon cancer HCT-116 cells and murine embryonic fibroblasts (MEF) with intact versus disrupted p21 genes. For comparison, HCT-116 cells lacking p53 function were also prepared through stable transfection with the human papillomavirus type-16 E6 gene. HCT-116/E6 cells were found to be more sensitive than control transfectants to CDDP and another DNA crosslinking agent, nitrogen mustard (HN2). HCT-116 cells with disrupted p21 genes also exhibited greater CDDP and HN2-sensitivity than parental HCT-116 cells. In contrast, the clonogenic survival of HCT-116 cells exposed to ionizing radiation, adriamycin, taxol or vincristine was not affected by p53 or p21 disruption. Sensitization of HCT-116/p21-/- cells to CDDP and HN2 was not limited to the HCT-116 cell background since MEF from p21 knockout mice were also more sensitive to these DNA crosslinking agents. Investigations into a possible cause of this enhanced sensitivity revealed that HCT-116 cells lacking p53 or p21 function exhibited a reduced ability to repair cisplatin-damaged CAT-reporter plasmids transfected into the cells. In addition, we found that HCT-116/p21-/- cells were much more susceptible to HN2-induced cell cycle delay than parental cells. Our results suggest that p21 disruption preferentially sensitizes at least some cell types to DNA crosslinking agents.