Distinct gut virome profiles are associated with response to anti-PD-1 therapy in non-small cell lung cancer

The gut microbiota is a key modulator of immune checkpoint inhibitor (ICI) efficacy, yet the contribution of the gut virome remains poorly defined, particularly in advanced non–small cell lung cancer (NSCLC). Here, we characterized the gut virome and explored its potential role in shaping response to PD-1 blockade. We performed metagenomic virome profiling of fecal samples from 338 advanced NSCLC patients treated with PD-1 inhibitors and evaluated model generalizability in an independent external cohort (n = 30). Viral diversity, taxonomic composition, and functional potential were analyzed. Virus–bacteria co-occurrence networks were constructed, and random forest classifiers were developed to predict treatment response. Viral Shannon diversity decreased progressively with poorer clinical response, and β-diversity analyses revealed distinct virome community structures between responders (R) and non-responders (NR). Differential abundance analysis identified 194 NR-enriched vOTUs, predominantly assigned to Peduoviridae and Inoviridae, and 594 R-enriched vOTUs, mainly from Herelleviridae and Microviridae. Host prediction indicated that NR-enriched vOTUs frequently targeted bacterial genera such as Clostridium_M, Bacteroides, and Escherichia, whereas R-enriched vOTUs targeted beneficial genera such as Faecalibacterium and Roseburia. Network analyses further revealed response-specific virus–bacteria interaction modules. Functional profiling showed that NR-enriched vOTUs were associated with metabolic functions, including K01689 (eno; enolase). A virus-only random forest model outperformed a bacterium-only model in predicting response (area under the curve [AUC] = 0.768 vs. 0.664) and maintained superior performance in the external cohort (AUC = 0.742). In addition, Akkermansia muciniphila positivity was associated with a higher-diversity, responder-favorable virome configuration. The gut virome undergoes marked remodeling during anti–PD-1 therapy in advanced NSCLC and displays distinct taxonomic, ecological, and functional signatures associated with clinical outcome. These findings support the gut virome as a strong predictor of ICI response and highlight its potential as both a biomarker and a therapeutic target.