Metagenomics and culturomics reveal the dual role of gut microbiome in the development of immune-related toxicities and immune checkpoint inhibitors efficacy in cancer

Background Despite their unprecedented clinical success, immune checkpoint inhibitors (ICI) are frequently associated with immune-related adverse events (irAE). Growing evidence indicates that the occurrence of irAE may correlate with enhanced ICI efficacy, although the underlying mechanisms remain unknown. Most studies investigating the role of the gut microbiome in oncology have relied on sequencing approaches, particularly shotgun metagenomics. Although microbiome profiling revealed strong associations between specific bacterial taxa and clinical outcomes, they have limitations, including an inability to detect low-abundance or uncharacterized bacteria. To overcome these limitations, we combined shotgun metagenomics and culturomics on fecal samples collected from patients with melanoma and non-small cell lung cancer (NSCLC), at baseline and at the onset of ir-colitis. Results We first validated across three independent cohorts of 589 patients with melanoma or NSCLC treated with ICI that grade ≥ 2 irAE were associated with significantly longer overall survival and progression-free survival. Complementary analysis by shotgun metagenomics and culturomics reported a decrease in alpha diversity. Metagenomics results showed enrichment of Ruminococcus gnavus and Streptococcus vestibularis at baseline in grade ≥ 2 irAE patients, while Clostridium paraputrificum and Streptococcus spp. were isolated by culturomics from baseline stool samples from ir-colitis patients. Longitudinal analysis of paired stool samples revealed a shift in microbiome composition with enrichment of Paraclostridium bifermentans and Clostridium paraputrificum , lower lipopolysaccharide and higher flagellin concentrations at baseline compared with the time of ir-colitis. Fecal microbiota transplantation from a patient with ir-colitis into mice induced surrogate markers of colon inflammation and enhanced the anti-tumor activity of combined anti-PD-1/CTLA-4. P. bifermentans isolated from this patient sample demonstrated direct epithelial barrier disruption in epithelial Caco-2 cells, characterized by decreased ZO-1 and Occludin and increased TNF-α and IL-1β expression. Moreover, in the DSS colitis model, P. bifermentans worsened weight loss. In a separate tumor model, it amplified the anti-tumor effect of dual ICI. This beneficial effect was maintained after treatment with the < 3 kDa filtered supernatant Conclusion Altogether, our results suggest that P. bifermentans promotes subclinical colitis while increasing the efficacy of dual ICI. This provides a potential microbiome-derived link between irAE and improved anti-tumor responses.