Activation of the transcription factor NF-κB has been shown to protect cells from tumor necrosis factor-alpha, chemotherapy, and radiation-induced apoptosis. NF-κB-dependent cIAP expression is a major antiapoptotic mechanism for that. NF-κB activation and cIAP expression in A549 lung cancer cells which is relatively resistant to radiation-induced cell death were investigated for the mechanism of radioresistance.

We used A549 lung cancer cells and Clinac 1800C linear accelerator for radiation. Cell viability test was done by MTT assay. NF-κB activation was tested by luciferase reporter gene assay, Western blot for IκBalpha degradation, and electromobility shift assay. For blocking NF-κB, MG132 and transfection of IκBα-superrepressor plasmid construct were used. cIAP expression was analyzed by RT-PCR and cIAP2 promoter activity was performed using luciferase assay system.

MTT assay showed that cytotoxicity even 48 hr after radiation in A549 cells were less than 20%. Luciferas assay demonstrated weak NF-κB activation of 1.6±0.2 fold compared to PMA-induced 3.4±0.9 fold. Radiation-induced IκBalpha degradation was observed in Western blot and NF-κB DNA binding was confirmed by EMSA. However, blocking NF-κB using MG132 and IκBalpha-superrepressor transfection did not show any sensitizing effect for radiation-induced cell death. The result of RT-PCR for cIAP1 & 2 expression was negative induction while TNF-α showed strong expression for cIAP1 & 2. The cIAP2 promoter activity also did not show any change compared to positive control with TNF-α.

We conclude that activation of NF-κB does not determine the intrinsic radiosensitivity of cancer cells, at least for the cell lines tested in this study.