Screening of known gut microbiome-derived metabolites in anti-tumor immunity of colorectal cancer
Xinan Zhang, Zixuan Li, Hui Hua, Hao Song, Sangni Qian, Xin Huang, Guiqin Ye, Yingyu Huang, Qin Wang, Boan Zheng, Xiaodong Xu, Yiran Li, Yanli Zhu, Liping Yu, Yeting Hong, Hang Yuan, Jianbin ZhangBackground
Most colorectal cancers (CRCs) patients with microsatellite stability (MSS) have minimal benefit from immune checkpoint inhibitor monotherapy, and even after PD-1/PD-L1 blockade, T cell function remains inadequately restored, highlighting the importance of PD-1 downstream signaling events. SHP-2 is a critical effector of PD-1 downstream signaling and plays a key role in establishing the immunosuppressive tumor microenvironment in CRC. Gut microbial metabolites are emerging as regulators of anti-tumor immunity, but their role in modulating the PD-1/SHP-2 interaction remains unknown.
Methods
We established a split-luciferase complementation screening system based on the PD-1/SHP-2 interaction and systematically screened a library of human gut microbial metabolites (n=480). The effect of candidate metabolites on T cell function was assessed in vitro using flow cytometry, confocal microscopy, and cell viability assays. Using a mouse colon cancer model, we further investigated the role of taurocholic acid (TCA) and glycodeoxycholic acid (GDCA) in the tumorigenesis and immunotherapy of CRC. Finally, the clinical relevance of metabolite-producing bacterium was examined in tumor specimens from CRC patients (n=53) using fluorescence in situ hybridization and real-time PCR assays.
Results
We identified the conjugated bile acids TCA and GDCA as enhancers of the PD-1/SHP-2 interaction. Mechanistically, TCA and GDCA stabilized the PD-1/SHP-2 complex through an allosteric mechanism by promoting PD-1 phosphorylation. Functionally, two metabolites suppressed CD8 + T cell activation, proliferation, and cytotoxicity in a PD-1/SHP-2-dependent manner, thereby accelerating tumor progression in vivo . Conversely, pharmacological depletion of TCA and GDCA using the bile acid sequestrant cholestyramine (CHO) effectively reversed their immunosuppressive effect and synergized with anti-PD-1 antibody therapy to inhibit the tumorigenesis of CRC. Clinical sample analysis revealed that the abundance of Clostridium scindens , the primary bacterial source of TCA and GDCA, was inversely correlated with CD8 + T cell infiltration in tumor tissues from CRC patients.
Conclusions
This study reveals a “Bile acid-associated metabolites-Immune checkpoint” regulatory axis that drives immunosuppression in CRC. The identification of TCA and GDCA as endogenous enhancers of PD-1 signaling provides a mechanistic rationale for targeting these metabolites to overcome immunotherapy resistance.