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Objective: To explore the role of cyclophilin A (CyPA) in sensitization of lung adenocarcinoma to radiotherapy using CRISPR/Cas9 technology.Methods: A CyPA knockout human lung adenocarcinoma cell line H1975 was established by CRISPR/Cas9 technology.Groups included a control group (wildtype),CyPA knockout group,traditional Chinese medicine (TCM) extract with Fuzhengzengxiao decoction group,and TCM extract with Fuzhengzengxiao decoction + CyPA knockout group.Each group was exposed to radiation at doses of 0,2,4,6,and 8 Gy.After 24 h,MTr assays were used to determine the survival rate of lung cancer cells and calculate radiosensitivity.The qPCR was used to measure mRNA expression of DDIT3,CDKN1A,and CDC25A associated with DNA damage repair.Results: Without irradiation,Fuzhengzengxiao decoction reduced the survival rate of lung adenocarcinoma cells (P ＜.0001).After irradiation,TCM extract with Fuzhengzengxiao decoction,CyPA knockout,and TCM extract with Fuzhengzengxiao decoction + CyPA knockout groups had reduced survival rates (P ＜.0001) and radiosensitivity was increased significantly.Expression of DDIT3,CDKN1A,and CDC25A was upregulated after knockout of CyPA (P ＜.0001).Expression of DDIT3 and CDC25A was increased after irradiation in wildtype cells treated with TCM extract with Fuzhengzengxiao decoction (DDIT3,P ＜.0001;CDC25A,P =.0059).The TCM extract with Fuzhengzengxiao decoction + CyPA knockout group also had increased expression of DDIT3 and CDC25A after irradiation (P ＜.0001).Conclusion: Fuzhengzengxiao decoction significantly decreases the survival rate of lung cancer cells,and its mechanism may be related to radiosensitization by decreasing expression of CyPA and inducing G1/S cell cycle arrest.