We investigated whether radiation influences the susceptibility of non-small cell lung cancer (NSCLC) cells to NK cell mediated cytotoxicity. We found radiation treatment increased expression of programmed cell death ligand 1 (PD-L1), but decreased NK group 2, member D (NKG2D) ligand expressions in A549 and H157 NSCLC cells. Both types of changes would have protected tumor cells from the cytotoxic action of NK cells. Consistently, we detected similar alteration in these molecules in radioresistant A549R26-1 and H157R24-1 subline cells. Higher PD-L1 level was also observed in tumors of A549R26-1 cell-derived xenografts than tumors of parental A549 (A549P) cell-derived xenografts. Accordingly, we found radioresistant cells were more resistant to the cytotoxic action of NK cells than parental cells, and such resistance was decreased when neutralizing antibody (Ab) of PD-L1 was added to the radioresistant cell/NK cell co-cultures. In mechanism studies, we found that IL-6- MEK/Erk signaling contributed most significantly to the up-regulation of PD-L1/down-regulation of NKG2D ligands in radioresistant cells. The addition of the MEK/Erk inhibitor increased the susceptibility of A549R26-1 and H157R24-1 cells to NK-cell cytotoxicity while no significant effect was observed in parental cells. Moreover, we detected enhanced NK-cell cytotoxicity to radioresistant cells when PD-L1 Ab and MEK/Erk inhibitor were added together to co-cultures of tumor/NK cells compared to when PD-L1 Ab was used alone. We suggest that combined use of PD-L1 Ab and MEK/Erk inhibitor may offer better therapeutic benefits than PD-L1 Ab alone to treat NSCLC patients who are receiving radiotherapy or who are at the radioresistant stage.