BACKGROUND AND OBJECTIVE DNA-dependent protein kinase catalytic subunit (DNA-PKcs) plays an important role in repairing irradiation-induced DNA double-strand break (DSB), and affects the radiosensitivity of tissue cells. This study was to detect the expression of DNA-PKcs in different non-small cell lung cancer (NSCLC) cell lines and evaluate its correlation to radiosensitivity. METHODS The content and activity of DNA-PKcs in five NSCLC cell lines A549, H1299, L78, PGCL3 and H460 were measured by Western blot and the DNA-PK activity assay. Cell survival was analyzed using clonogenic formation assay. RESULTS The radiosensitivities of five NSCLC cell lines were different. The values of survival fraction at 2 Gy (SF2) were 0.74 in A549 cells, 0.25 in H1299 cells, 0.21 in H460 cells, 0.48 in PGCL3 cells, and 0.58 in L78 cells. The protein levels of DNA-PKcs were 3.26+/-0.98 in A549 cells, 0.51+/-0.07 in L78 cells, 0.51+/-0.11 in H1299 cells, 0.86+/-0.23 in H460 cells, and 2.60+/-0.76 in PGCL3 cells. The activity values of DNA-PKcs were 8.30+/-1.03 in A549 cells, 2.45+/-0.52 in H1299 cells, 0.11+/-0.02 in H460 cells, 4.13+/-0.87 in PGCL3 cells, and 0.42+/-0.07 in L78 cells. In adenocarcinoma and large cell carcinoma cell lines, SF2 were correlated to DNA-PKcs content (P<0.05, r=0.95) and activity (P=0.03, r=0.98). CONCLUSION DNA-PKcs is an important factor to predict the radiosensitivity in adenocarcinoma and large cell lung cancer cell lines.