Background and purpose: Checkpoint kinase 2 (CHK2) is activated by DNA damage and can contribute to p53 stabilization, modulating growth arrest and/or apoptosis. We investigated the contribution of CHK2 to oxaliplatin-mediated toxicity in a colorectal cancer model. Experimental approach: We evaluated the ability of CHK2 small molecule inhibitors to potentiate oxaliplatin-induced toxicity. The role of CHK2 in oxaliplatin-induced apoptosis was investigated in HCT116 cells that were wild-type (WT) or KO for CHK2. Small molecule inhibitors of CHK2 were used in combination studies with oxaliplatin in this cell model. Key results: In oxaliplatin-treated CHK2 KO cells, accelerated apoptosis was accompanied by attenuated p53 stabilization and p21 WAF-1 up-regulation correlating with increased Bax expression, cytochrome c release and elevated caspase activity. The higher levels of apoptosis in CHK2 KO cells were restored to control (WT) levels when CHK2 was re-introduced. This 'uncoupling' of p53 stabilization and Bax up-regulation in CHK2 KO cells suggested oxaliplatin-induced apoptosis was due to a p53-independent response. Combination studies revealed that CHK2 inhibitor II or debromohymenialdisine antagonized the responses to oxaliplatin. This inhibitory effect correlated with decreases in apoptosis, p53 stabilization and DNA inter-strand cross-link formation, and was dependent on the presence (but not activity) of CHK2. Conclusions and implications: Combinations of CHK2 inhibitors with oxaliplatin should further sensitize cells to oxaliplatin treatment. However, these inhibitors produced an antagonistic effect on the response to oxaliplatin, which was reversed on the re-introduction of CHK2. These observations may have implications for the use of oxaliplatin in colorectal cancer therapy in combination with therapies targeting CHK2. © 2010 The Authors.