Single-cell landscape identifies the immunophenotypes and microenvironments of HBV-positive and HBV-negative liver cancer

Background: HBV infection leads to HCC and affects immunotherapy. We are exploring the tumor ecosystem in HCC to help gain a deeper understanding and design more effective immunotherapy strategies for patients with HCC with or without HBV infection. Methods: Single-cell RNA sequencing series were integrated as a discovery cohort to interrogate the tumor microenvironment of HBV-positive (HBV+) HCC and HBV-negative (HBV−) HCC. We further dissect the intratumoral immune status of HBV+ HCC and HBV− HCC. An independent cohort, including samples treated with immune checkpoint blockade therapy, was used to validate the major finding and investigate the effect of HBV infection on response to immunotherapy. Results: The interrogation of tumor microenvironment indicated that regulatory T cells, exhausted CD8⁺ T cells, and M1-like Macrophage_MMP9 were enriched in HBV+ HCC, while mucosa-associated invariant T cells were enriched in HBV− HCC. All subclusters of T cells showed high expression of immune checkpoint genes in HBV+ HCC. Regulatory T cells enriched in HBV+ HCC also showed more robust immunosuppressive properties, which was confirmed by cross talk between immune cell subsets. The ability of antigen presentation with major histocompatibility complex-II was downregulated in HBV+ HCC and this phenomenon can be reversed by immunotherapy. Two types of HCC also present different responses to immunotherapy. Conclusions: There is a more immunosuppressive and exhausted tumor microenvironment in HBV+ HCC than in HBV− HCC. This in-depth immunophenotyping strategy is critical to understanding the impact of HBV and the HCC immune microenvironment and helping develop more effective treatments in patients with HCC.