Tissue-resident Limosilactobacillus reuteri modulates intratumoral arachidonic acid metabolism to enhance CD8+ T cell-mediated anti-PD1 response.

BACKGROUND

Tissue-resident bacteria have emerged as modulators of host responses to tumor immunotherapy. However, the immunological and metabolic mechanisms by which tissue-resident microbiota influence immune checkpoint blockade in colorectal cancer (CRC) remain incompletely defined.

METHODS

We profiled the microbial composition of colorectal tissues from CRC patients and identified differentially enriched taxa between tumors and adjacent non-tumor tissues(NTs). By focusing on tissue-resident bacteria within the colorectal tumor microenvironment, immunocompetent mouse models were used to evaluate the impact of Limosilactobacillus reuteri (enriched in NTs) on tumor growth, survival, and anti-PD1 efficacy. Single-cell RNA sequencing combined with flow cytometric analysis was applied to characterize the phenotypic features of CD8⁺ T cells following anti-PD1 treatment with or without L.reuteri. Untargeted metabolomics combined with flow cytometry was conducted to investigate the association between tissue-resident L.reuteri and intratumoral lipid metabolic remodeling.

RESULTS

L.reuteri administration significantly enhanced the efficacy of anti-PD1 therapy, reduced tumor burden (MC38: 95% CI −755.20 to −210.90 mm3, P < 0.0001; CT26: 95% CI −558.30 to −340.70 mm3, P < 0.0001), and prolonged survival (MC38: log-rank P < 0.01; CT26: log-rank P < 0.01) across multiple mouse models. Mechanistically, L.reuteri reduced CD8⁺ T-cell exhaustion and strengthened effector function. Untargeted metabolomic profiling revealed that L.reuteri remodeled intratumoral lipid metabolism, reflected by a moderate increase in arachidonic acid (AA) levels. These metabolic changes promoted CD8⁺ T-cell activation and cytotoxicity within the TME.

CONCLUSION

Our study demonstrated that tissue-resident L.reuteri enhances anti-PD1 efficacy in CRC by alleviating CD8⁺ T cell exhaustion through reprogramming intratumoral lipid metabolism, highlighting the therapeutic potential of microbiota-based immunomodulation.