Effect And Mechanism Of Tumor Suppressor CMTM7 On Radiosensitivity Of Non-small Cell Lung Cancer

Objective:Non-small cell lung cancer has become the leading cause of cancer death.Radiation therapy is an indispensable treatment method for patients with early non-surgical and locally advanced non-small cell lung cancer.Enhancing the radiosensitivity of lung cancer is very important to improve the efficacy of radiotherapy.The purpose of this study was to investigate the effect of tumor suppressor CMTM7 on the sensitivity of non-small cell lung cancer to radiotherapy,and to provide new ideas for improving the radiotherapy effect of patients with non-small cell lung cancer.Methods:CRISPR / Cas9 technology was used to interfere with CMTM7 expression by knocking out CMTM7 in non-small cell lung cancer cells A549.RT-PCR and Western Blot were used to verify the CMTM7 knockout.6MV-X-rays of 0Gy,2Gy,4Gy,6Gy,and 8Gy were used to irradiate wild-type A549-WT cells and CMTM7 knock-out A549-B2 cells,and clone formation experiment was used to detect the effect of CMTM7 knockout on radiosensitivity of lung cancer cells.The effect of CMTM7 on the apoptosis of non-small cell lung cancer cells after 0Gy and 6Gy irradiation was detected by Annexin V / PI double staining method and flow cytometry.Propidium iodide(PI)staining and flow cytometry were used to detect the effects of CMTM7 on the cell cycle of non-small cell lung cancer after 0Gy and 6Gy irradiation.Western Blot was used to detect the effects of CMTM7 on the expression of AKT signaling pathway and γ-H2 AX protein,a DNA damage marker,after 0Gy and 6Gy irradiation.A549-WT and A549-B2 transplanted nude mice models were irradiated with 0Gy and 10 Gy,and the changes in body weight and tumor volume of nude mice were recorded,respectively.Results:RT-PCR and Western Blot confirmed that CMTM7 expression was high in A549-WT cells but no CMTM7 expression in A549-B2 cells,that is,CRISPR / Cas9 technology successfully knocked out CMTM7 in wild type A549 cells.Under different doses of X-ray irradiation,A549-B2 cells had stronger colony-forming ability and faster growth rate than A549-WT cells,namely,CMTM7 knockout significantly promoted cell proliferation under radiotherapy conditions.There was almost no difference in apoptosis between the A549-WT group and the A549-B2 group after irradiation,showing that CMTM7 had no significant effect on the apoptosis of non-small cell lung cancer cells induced by radiotherapy.CMTM7 knockout promoted radiation-induced G2 phase arrest.After knocking out CMTM7,γ-H2 AX protein expression was significantly down-regulated at the indicated time pionts,suggesting that CMTM7 knockout decreased radiation-induced DNA damage and promoted the process of DNA damage repair.Knockout of CMTM7 promotes AKT phosphorylation.The tumor volume of the CMTM7 knockout group was significantly larger than that of the wild group,and there was no significant difference in body weight between the mice in each group.Conclusions:This study demonstrates that in non-small cell lung cancer,CMTM7 knockout promotes radiotherapy-induced DNA damage repair and reduces radiosensitivity in vitro and in vivo,suggesting that CMTM7 is involved in the regulation of the sensitivity of non-small cell lung cancer to radiotherapy...