Tumor-suppressing multi-enterobacteria enhance the anti-PD-1/PD-L1 efficacy in microsatellite stable colorectal cancer.

Gut microbiome plays a pivotal role in modulating immunotherapy responses in colorectal cancer (CRC) treatment. While individual enterobacteria have been identified as enhancers of anti-PD-1/anti-PD-L1 therapy, the synergistic effects of multiple probiotic strains remain insufficiently explored. In this study, we investigated the therapeutic potential of Tumor-Suppressing Multi-Enterobacteria (TSME), a consortium of nine beneficial intestinal probiotic strains, in enhancing anti-PD-1/anti-PD-L1 therapy for microsatellite stable (MSS) CRC. Using a tumor-bearing mouse and employing techniques including flow cytometry, immunohistochemistry, ELISA, and genomic sequencing, we found that TSME significantly improved the efficacy of immune checkpoint inhibitors (ICIs) by optimizing tumor immune and microbe microenvironment. Specifically, the addition of TSME increased CD8+ T cell infiltration and reshaped cytokine profiles, including reducing pro-inflammatory cytokines (IL-17, IL-1β, IL-6, and TNF-α) while elevating anti-inflammatory factors (IFN-γ). Moreover, TSME significantly up-regulated key immune pathways, including TNF signaling, cytokine-cytokine receptor interaction, and JAK-STAT signaling. In addition, TSME restructured the gut microbiome, increasing the abundance of beneficial bacteria such as Akkermansia and Alistipes. These findings highlight the synergistic effect of the multi-strain probiotics in enhancing ICI efficacy. Well-formulated probiotic consortia offer a promising strategy for enhancing immunotherapy outcomes in MSS CRC and advancing broader implementation of microbiome-assisted precision oncology.