The induction of tumor cell death by anticancer therapy results from a genetic program of autonomous cell death termed apoptosis. Because the p53 tumor suppressor gene is a critical component for induction of apoptosis in response to DNA damage, its inactivation in cancers may be responsible for their resistance to genotoxic anticancer agents. The cellular response to DNA damage involves a cell-cycle arrest at both the G1/S and G2/M transitions; these checkpoints maintain viability by preventing the replication or segregation of damaged DNA. The arrest at the G1 checkpoint is mediated by p53-dependent induction of p21WAF1/CIP1, whereas the G2 arrest involves inactivation of p34cdc2 kinase. Following DNA damage, p53-deficient cells fail to arrest at G1 and accumulate at the G2/M transition. We demonstrate that abrogation of G2 arrest by caffeine-mediated activation of p34cdc2 kinase results in the selective sensitization of p53-deficient primary and tumor cells to irradiation-induced apoptosis. These data suggest that pharmacologic activation of p34cdc2 kinase may be a useful therapeutic strategy for circumventing the resistance of p53-deficient cancers to genotoxic anticancer agents.