562 Bile acids circumvent gut dysbiosis-induced resistance to anti-PD-1

<h3>Background</h3> Intestinal dysbiosis is associated with resistance to immune checkpoint inhibitor (ICI) due to downregulation of the mucosal addressin cell adhesion molecule-1 (MAdCAM-1) culminating in the exodus of immunosuppressive α4β7⁺ Treg cells from the gut to distal tumors.^1–3 The downregulation of MAdCAM-1 is concomitant to perturbations of intestinal bile acids (BA) in tumor bearers suffering from gut dysbiosis.³ This result is consistent with the role of gut microbiota in BA metabolism. Thus, we addressed whether distinct BA could restore MAdCAM-1 expression and ICI efficacy during gut dysbiosis. <h3>Methods</h3> We generated two endothelial biosensor cell lines to operate screening of a large library of BA and agonists/antagonists of BA receptors, for their capacity to up-or downregulate MAdCAM-1 expression in vitro. In tumor bearing mice (MCA205 sarcoma and RET melanoma) subjected to anti-PD1 Abs, gut dysbiosis was induced by antibiotics or <i>Enterocloster clostridioformis</i> colonization and compensated by 5 different BA. Regulation of ileal and mLN MAdCAM-1 expression by gavages with BA was assessed by RT-qPCR. We evaluated α4β7⁺gut lymphocytes recirculation with a cell tracer³ using flow cytometry analyses focusing also on thymocytes and performed LC-mass spectrometry-metabolomics of various compartments (including tumor bed) post- BA administration. <h3>Results</h3> Two selective BA (glyco- and taurodeoxycholic acid (GDCA, TDCA)) could upregulate MAdCAM-1 expression in vitro and in vivo. These BA bypassed gut dysbiosis-induced resistance to ICI in two tumor models by preventing the exodus of α4β7⁺ Treg cells from the gut to the tumor. GDCA was the only BA to reprogram intratumoral metabolism towards an immunogenic pattern in mice (increasing polyamine, riboflavin and indole pathways). Surprisingly, GDCA was associated with a better response to ICI in patients, concomitantly to a recirculation of sjTREC (signal joint TCR rearrangement excision circles), a marker of thymopoiesis. In mice, GDCA induced the differentiation of intrathymic single positive CD8⁺ T cells <h3>Conclusions</h3> We successfully bypassed dysbiosis-related primary resistance to ICI using the conjugated secondary BA, GDCA that mediated its bioactivity through three modes of action; i) locally by increasing MAdCAM-1 expression, thereby preventing the exodus of Treg to the tumor, ii) distantly by reprograming the tumor metabolism, iii) on the thymus by replenishing the pool of naïve CD8⁺ T cells. Hence, we propose that GDCA could represent a selective metabolite to restore endothelial and intestinal homeostasis culminating in circumventing primary resistance to ICI. We are currently investigating the BA receptor involved in these beneficial effects. <h3>Acknowledgements</h3> ALMDLV is supported by Paris Saclay University. LZ, GK and MF are supported by SEERAVE Foundation. <h3>References</h3> Routy B, <i>et al</i>. Gut microbiome influences efficacy of PD-1-based immunotherapy against epithelial tumors. <i>Science</i> 2018;<b>359</b>:91–97. Derosa L,<i> et al</i>. Gut bacteria composition drives primary resistance to cancer immunotherapy in renal cell carcinoma patients. <i>Eur. Urol</i> 2020;<b>78</b>:195–206. Fidelle. A microbiota-modulated checkpoint directs immunosuppressive intestinal T cells into cancers | Science. https://www.science.org/doi/abs/10.1126/science.abo2296 (2023). <h3>Ethics Approval</h3> All animal experiments were performed in compliance with French and European laws and regulations. The local institutional animal ethics board and French Ministère de la Recherche approved all mouse experiments (permission numbers: 2023-024-42156). Experiments were performed in accordance with Government and institutional guidelines and regulations.