Background: Nuclear factor E2-related factor-2 (NFE2L2) gene encodes a transcription factor which is a member of basic leucine zipper (bZIP) proteins family. Overexpression of NFE2L2 lead to cell proliferation and promoted tumor metastasis. Previous report indicated that NFE2L2 mutation (NFE2L2-MT) was an independent poor prognostic factor in esophageal squamous cell carcinoma (ESCC). However, the correlation between NFE2L2 mutation and pan-cancer types of TMB, MSI, and PD-L1 expression is unclear. Methods: TMB analysis was performed in 3,716 Chinese pan-cancer patients who underwent NGS sequencing using a 539 gene panel. The TMB calculation included synonymous and nonsynonymous mutations and InDels. MSI analysis was performed in 3,110 patients. MSI-H was defined as above 10% positive of the 195 tested microsatellites sites. The PD-L1 expression analysis was performed in 3,415 patients with immunohistochemistry staining (IHC) by antibody SP263. PD-L1 positive was defined as greater than or equal to 1%. The statistical correlation was investigated using Chi-square analysis. TMB value was compared using Wilcoxon Rank Sum test. We used TCGA public database to verify the result. Results: The mutation frequency of NFE2L2 mutation was 2.66% (99/3716). The TOP 5 cancer types were liver cancer 3.53% (14/397), lung cancer 2.97% (42/1416), colorectal cancer 2.02% (7/347), gastric cancer 1.36% (3/221), soft tissue sarcoma 0.53% (1/189). NFE2L2-MT had a significant correlation with higher TMB (p = 2.2e-16), compared with NFE2L2 wild-type (NFE2L2-WT). Among 3110 samples with MSI status, the MSI-H percentage of NFE2L2-MT and NFE2L2-WT were 8.60% (8/93) and 1.33% (40/3017), respectively (p = 1.29e-7). In 3,415 patients with PD-L1 protein expression information, the PD-L1 positive percentage of NFE2L2-MT and NFE2L2-WT were 51.52% (51/99) and 61.9% (1,268/2,048), respectively. NFE2L2-WT has higher PD-L1 positive percentage than NFE2L2-MT (p = 0.01). NFE2L2-MT was significantly correlated with higher TMB and MSI when we used TCGA data to verify, p<0.0001. However, the survival analysis of 1661 MSKCC immunotherapy cohort showed that the median OS of NFE2L2-MT vs NFE2L2-WT was 21 months vs 18 months (p=0.858), but the difference was not significant. Conclusions: NFE2L2 mutation has a very significant correlation with higher TMB and MSI, but not related to PD-L1 expression. However, whether NFE2L2-MT is related to the efficacy of immunotherapy was still unclear and more clinical data were needed.