Background: In recent years, the advent of immunotherapy had provided a new approach for non-small cell lung cancer (NSCLC) patients. However, only a small subset of patients can benefit from immune checkpoint inhibitors (ICIs) and the majority of patients still show poorly durable responses. Although some biomarker for predicting clinical efficacy of immunotherapy has been reported, the more proper predictive biomarker is required to screen dominant populations for ICI efficacy. Methods: A total of three publicly available datasets of NSCLC patients treated with immunotherapy were collected for analysis. The discovery contained 57 NSCLC patients sequenced by WES from Miao et al. and another cohort of 75 patients with clinical data and WES data download from Hellman et al. studies. We collected two MSK-IMPACT cohorts including 240 and 350 NSCLC immunotherapy patients used as validation set from cBioPortal(www.cbioportal.org). Results: We detected 79 mutation genes (mutation frequency > 5% cases) associated with a better progressive free survival PFS (HR < 1, P value < 0.05) by univariate Cox regression analysis in the discovery cohort. The survival analysis results indicated that patients who carried co-occurring mutations of GRIN2A, FAT1, FLT1, ASXL2, PTPRS ,and DDR pathway showed a longer PFS (p = 0.004, HR = 0.37, 95% Cl 0.235-0.644). Notably, comparing with the non-co-mutation group, the higher tumor mutation burden (TMB)(P < 0.01) and more durable clinical benefit(P < 0.05) have been detected in the co-mutations group. Next, we validated our findings using two independent validation cohorts who received ICI therapies from MSKCC, the results showed that PFS was significantly prolonged in the co-mutation group (p = 0.036, HR = 0.67, 95%CI 0.47-0.97), and with no significant difference in overall survival. Additionally, the TMB was significantly higher in the co-mutation group than that harbored non-co-mutation in two validation cohorts(p < 0.01). Conclusions: In this study, we found that co-occurring mutations in five genes and DDR pathways can predict a better immunotherapy efficacy of NSCLC. Therefore, the co-mutation signature may have great potential as a biomarker for guiding immunotherapy of NSCLC.