Ripk2 promotes CD8+ T cell inactivation and hepatocellular carcinoma progression through Myb/Cxcl9 and Pax5/Adpgk signaling pathways

Abstract Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide, characterized by complex pathogenesis and immune escape mechanisms. The tumor immune microenvironment plays an increasingly recognized role in tumorigenesis, progression, and treatment. Receptor-interacting serine/threonine-protein kinase 2 (Ripk2) plays a pivotal role in inflammatory responses and immune regulation. This study aimed to investigate the role of Ripk2 in the HCC immune microenvironment, particularly its impact on macrophage function, using a macrophage Ripk2 knockout mouse model (Ripk2 CKO ), single-cell sequencing analysis (scRNA-seq) and flow cytometry (FCM). We found that inhibiting or deleting Ripk2 altered the intratumoral microbiota within macrophages, significantly increasing the abundance of Streptomyces collinus ( Sc ). This alteration not only prompted a metabolic shift in macrophages but also significantly upregulated the transcription factor Myb expression, promoting Cxcl9 secretion, thereby enhancing CD8 + T cell infiltration, activity, and proliferation. Furthermore, Ripk2 was found to promote lactate production via the Pax5/Adpgk pathway, leading to CD8 + T cell dysfunction and forming an immunosuppressive feedback loop in HCC. Combining a Ripk2 inhibitor (GSK583) with a PD-1/PD-L1 inhibitor (BMS202) reduced the hepatotoxicity of BMS202 monotherapy and improved therapeutic efficacy. In conclusion, this study provides new insights into the role of Ripk2 in the HCC immune microenvironment and lays an experimental foundation for the development of Ripk2-targeted immunotherapy strategies.