<i>Lactobacillus gallinarum</i>-derived metabolites boost anti-PD1 efficacy in colorectal cancer by inhibiting regulatory T cells through modulating IDO1/Kyn/AHR axis.

Abstract

Gut microbiota is a key player in dictating immunotherapy response. We aimed to explore the immunomodulatory effect of probiotic <i>Lactobacillus gallinarum</i> and its role in improving anti-programmed cell death protein 1 (PD1) efficacy against colorectal cancer (CRC). The effects of <i>L. gallinarum</i> in anti-PD1 response were assessed in syngeneic mouse models and azoxymethane/dextran sulfate sodium-induced CRC model. The change of immune landscape was identified by multicolour flow cytometry and validated by immunohistochemistry staining and in vitro functional assays. Liquid chromatography-mass spectrometry was performed to identify the functional metabolites. <i>L. gallinarum</i> significantly improved anti-PD1 efficacy in two syngeneic mouse models with different microsatellite instability (MSI) statuses (MSI-high for MC38, MSI-low for CT26). Such effect was confirmed in CRC tumourigenesis model. <i>L. gallinarum</i> synergised with anti-PD1 therapy by reducing Foxp3⁺ CD25⁺ regulatory T cell (Treg) intratumoural infiltration, and enhancing effector function of CD8⁺ T cells. <i>L. gallinarum</i>-derived indole-3-carboxylic acid (ICA) was identified as the functional metabolite. Mechanistically, ICA inhibited indoleamine 2,3-dioxygenase (IDO1) expression, therefore suppressing kynurenine (Kyn) production in tumours. ICA also competed with Kyn for binding site on aryl hydrocarbon receptor (AHR) and antagonised Kyn binding on CD4⁺ T cells, thereby inhibiting Treg differentiation in vitro. ICA phenocopied <i>L. gallinarum</i> effect and significantly improved anti-PD1 efficacy in vivo, which could be reversed by Kyn supplementation. <i>L. gallinarum</i>-derived ICA improved anti-PD1 efficacy in CRC through suppressing CD4+Treg differentiation and enhancing CD8+T cell function by modulating the IDO1/Kyn/AHR axis. <i>L. gallinarum</i> is a potential adjuvant to augment anti-PD1 efficacy against CRC.