Limited studies have indicated that some chemotherapy agents activate the transcription factor nuclear factor-κB (NF-κB), and that this leads to suppression of the apoptotic potential of the chemotherapy. In contrast, it was reported recently that stable inhibition of NF-κB in four different cancer cell lines did not lead to augmentation of the chemotherapy-induced apoptosis. In this study, we have focused on colorectal cancer, which is known to be highly resistant to genotoxic chemotherapy and gamma irradiation. We show that the topoisomerase I inhibitor 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin(CPT-11) activates NF-κB in most colorectal cancer cell lines. We then examine a therapeutic strategy that uses adenovirus-mediated transfer of the super-repressor IκBα to inhibit NF-κB activation as an adjuvant approach to promote chemosensitivity in colorectal tumor cells to treatment with CPT-11. These data demonstrate that the protection from apoptosis induced in response to CPT-11 treatment is effectively inhibited by the transient inhibition of NF-κB in a variety of human colon cancer cell lines and in a tumor xenograft model, resulting in a significantly enhanced tumoricidal response to CPT-11 via increased induction of apoptosis. These findings indicate that the activation of NF-κB by chemotherapy is an important underlying mechanism of inducible chemoresistance.