Non-small cell lung cancer (NSCLC) is known to carry heavy mutation load. Besides smoking, cytidine deaminase APOBEC3B plays a key role in the mutation process of NSCLC. APOBEC3B is also reported to be upregulated and predicts bad prognosis in NSCLC. However, targeting APOBEC3B high NSCLC is still a big challenge. Here we show that APOBEC3B upregulation is significantly associated with immune gene expression, and APOBEC3B expression positively correlates with known immunotherapy response biomarkers, including: PD-L1 expression and T-cell infiltration in NSCLC. Importantly, APOBEC mutational signature is specifically enriched in NSCLC patients with durable clinical benefit after immunotherapy and APOBEC mutation count can be better than total mutation in predicting immunotherapy response. In together, this work provides evidence that APOBEC3B upregulation and APOBEC mutation count can be used as novel predictive markers in guiding NSCLC checkpoint blockade immunotherapy.