Colorectal tumors frequently contain activating mutations in KRAS. ReovirusT3D is an oncolytic virus that preferentially kills tumor cells with an activated Ras pathway. Here we have assessed the contribution of endogenous mutant KRAS in human colorectal cancer cell lines to ReovirusT3D replication and to tumor cell oncolysis. In addition, treatment combinations involving ReovirusT3D, oxaliplatin, and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) were tested for their efficacy in tumor cell killing. The mutation status of KRAS did not predict the sensitivity of a panel of human colorectal cancer cell lines to ReovirusT3D. Virus replication was observed in all cell lines tested regardless of KRAS status and was not affected by deletion of endogenous mutant KRAS(D13). However, deletion of KRAS(D13) or p53 did reduce apoptosis induction by ReovirusT3D whereas deletion of beta-catenin(DeltaS45) had no effect. Likewise, KRAS(D13)- or p53-deficient cells display reduced sensitivity to oxaliplatin but not to death receptor activation by TRAIL. Finally, the treatment of colorectal cancer cells with ReovirusT3D combined with either oxaliplatin or TRAIL resulted in a nonsynergistic increase in tumor cell killing. We conclude that oncolysis of human tumor cells by ReovirusT3D is not determined by the extent of virus replication but by their sensitivity to apoptosis induction. Oncogenic KRAS(D13) increases tumor cell sensitivity to activation of the cell-intrinsic apoptosis pathway without affecting ReovirusT3D replication.