Nuclear factor kappa B (NF-kappa B) is a transcription factor that is known to regulate apoptosis when cells are exposed to DNA-damaging agents such as ionizing radiation and cytotoxic drugs. We sought to determine if inhibition of NF-kappa B could enhance radiosensitivity in human colon cancer cells in vitro and in vivo. To inhibit NF-kappa B activation specifically, we constructed a recombinant adenovirus vector expressing a truncated form of the inhibitor protein I kappa B alpha (I kappa B alpha Delta N) that lacks the phosphorylation sites essential for activation of NF-kappa B, and transfected two human colon cancer cell lines (HT29 and HCT15) with this vector. In vitro colony-forming assays revealed that the overexpression of the stable I kappa B alpha by AxI kappa B alpha Delta N infection significantly suppressed cell growth after irradiation in both cell lines as compared to infection with a control vector, AxLacZ. Treatment with AxI kappa B alpha Delta N and irradiation successfully inhibited the growth of HT29 xenografted subcutaneous tumors in nude mice with an 83.8% volume reduction on day 38 as compared to the untreated tumors. Furthermore, it was demonstrated that apoptosis was increased by adenovirus-mediated gene transduction of I kappa B alpha Delta N in vitro and in vivo. These results indicated that inhibition of NF-kappa B could enhance radiosensitivity through an increase in radiation-induced apoptosis. We believe that radio-gene therapy using adenovirus-mediated gene transduction of I kappa B alpha Delta N could be an attractive candidate as a treatment strategy for colorectal cancer.