Nuclear transcription factor-kappaB (NF-kappaB) is a transcriptional complex that is rapidly activated in the course of an immediate early response of cells after exposure to different stresses including ionizing radiation (IR). To overcome the limitation of radiation therapy for thyroid cancers, we studied the response of the NF-kappaB cascade to IR in cultured normal human thyroid cells and various thyroid cancer cell lines. Exposure to IR resulted in a dose-dependent increase of DNA-binding activity of p65 and p50 subunits in all types of thyroid cells. Specific inhibitors of NF-kappaB or phosphorylation deficient mutant inhibitory protein IkappaBalpha reduced thyroid cancer cell survival after exposure to IR and enhanced IR-induced cell death in a model undifferentiated thyroid cancer cell line. Tumors harboring mutant IkappaBalpha implanted into nude mice exhibited delayed growth rate and increased radiosensitivity. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling and annexinV-propidium iodide staining revealed the increase of radiation-induced apoptosis in the cells with inhibited NF-kappaB signaling. Our results indicate that radiosensitivity of transformed thyroid cells is due in part to elevated basal activity and rapid induction of the active form of NF-kappaB. We therefore suggest that inhibition of NF-kappaB could be an effective modality for radiation therapy of advanced human thyroid cancers.