RORc expressing immune cells negatively regulate tertiary lymphoid structure formation and support their pro-tumorigenic functions

• Published in: Journal of hepatology 2024-12
• Main Authors: Cinnamon, Einat, Stein, Ilan, Zino, Elvira, Rabinovich, Stav, Shovman, Yehuda, Schlesinger, Yehuda, Salame, Tomer-Meir, Reich-Zeliger, Shlomit, Albrecht, Thomas, Roessler, Stephanie, Schirmacher, Peter, Lotem, Michal, Ben-Neriah, Yinon, Parnas, Oren, Pikarsky, Eli
• Format: Article
• Language: English
• Subjects: B cells; CD4 cells; CD8 cells; cell depletion; cholangiocarcinoma; CyTOF; liver cancer; RORc; single cell RNA sequencing; Tertiary lymphoid structure
• ISSN: 0168-8278; 1600-0641; 1600-0641
• DOI: 10.1016/j.jhep.2024.12.015

RORc-expressing immune cells play important roles in inflammation, autoimmune disease and cancer. They are required for lymphoid organogenesis and have been implicated in tertiary lymphoid structure (TLS) formation. TLSs are formed in many cancer types and have been correlated with better prognosis and response to immunotherapy. In liver cancer, some TLSs are pro-tumorigenic as they harbor tumor progenitor cells and support their growth. The processes involved in TLS development and acquisition of pro- or anti-tumorigenic roles are largely unknown. This study aims to explore the role of RORc-expressing cells in TLS development in the context of inflammation-associated liver cancer. IKKβ(EE)Hep mice, exhibiting chronic liver inflammation, TLS formation and liver cancer, were crossed with RORc knockout mice to explore RORc’s effect on TLS and tumor formation. TLS phenotypes were analyzed using transcriptional, proteomic, and immunohistochemical techniques. CD4, CD8, and B cell depletions were used to assess their contribution to liver TLS and tumor formation. RORc-expressing cells are detected within TLSs of both human patients and mice developing intrahepatic cholangiocarcinoma. In mice, these cells negatively regulate TLS formation, as excess TLSs form in their absence. CD4 cells are essential for liver TLS formation, while B cells are required for TLS formation specifically in the absence of RORc-expressing cells. Importantly, in chronically inflamed livers lacking RORc-expressing cells, TLSs become anti-tumorigenic, reducing tumor load. Anti-tumorigenic TLSs revealed enrichment of exhausted CD8 cells with effector functions, germinal center B cells and plasma cells. B cells are key in limiting tumor development, possibly via tumor-directed antibodies. RORc-expressing cells negatively regulate B cell responses and facilitate the pro-tumorigenic functions of hepatic TLSs. [Display omitted] •RORc+ cells in human and mouse livers with iCCA are primarily located within TLSs.•RORc+ cells suppress TLS formation in liver cancer. CD4 cells are essential for TLS development, while B cells are needed for TLS formation in the absence of RORc+ cells.•In chronically inflamed livers lacking RORc+ cells, TLSs become anti-tumorigenic.•Anti-tumorigenic TLSs are enriched in CD8 cells, germinal center B cells, and plasma cells.•B cells limit iCCA growth, possibly through tumor-directed antibodies.