AIM: To study the efficacy and molecular mechanism of PX-478 in enhancing radiotherapy effect of lung cancer. METHODS: H460, A549 cells were divided into blank group, radiation group and radiation combined PX-478 group. In addition to the blank group, the radiation group and the PX-478 group were given 2Gy X-ray irradiation to establish the radiation model, and the radiation combined with the PX-478 group was given 20 μmol/L PX-478 intervention after modeling, and cultured for 24 h. Inverted microscope was used to observe cell growth and cell number, CCK-8 method was used to detect cell viability, cloning was used to observe cell proliferation, flow cytometry was used to detect cell apoptosis, and Western blot was used to detect HIF-1α, GLUT1, HK2, PFK1, PKM2, LDHA protein expression. RESULTS: Compared with blank group, the number of H460,A549 cells in radiation group decreased, cell viability and proliferation ability decreased, cell apoptosis rate increased, HIF-1α, GLUT1, HK2, PFK1, PKM2, LDHA protein expression increased (P<0.01). Compared with the radiation group, the number of H460, A549 cells in the radiation combined PX-478 group was significantly decreased, the cell viability and proliferation ability were significantly weakened, the apoptosis rate was significantly increased, and the protein expressions of HIF-1α, GLUT1, HK2, PFK1, PKM2 and LDHA were significantly decreased (P<0.01).CONCLUSION: PX-478 can regulate the HIF-1α-mediated glycolysis in A549,H460 cells after radiation, regulate the energy metabolism, increase the apoptosis of tumor cells, and improve the effect of radiotherapy.