Viral-Track integrated single-cell RNA-sequencing reveals HBV lymphotropism and immunosuppressive microenvironment in HBV-associated hepatocellular carcinoma.

The tumor microenvironment (TME) is a crucial mediator of tumor progression and treatment response. Here, we compare the immune microenvironments of HBV and non-HBV hepatocellular carcinoma (HCC) and investigate the reason for the persistence of HBV infection in the liver. We combine the Viral-Track method with scRNA sequencing and profile the transcriptomes of 70,056 cells from HBV and non-HBV-HCC patients. In addition to hepatocytes and macrophages, HBV transcripts were also detected in T and B cells using the Viral-Track method, confirming the lymphotropic nature of HBV in scRNA-sequencing data for the first time, to the best of our knowledge. HBV-HCC tumors have reduced levels of NK cells, macrophages, DCs, and increased malignant hepatocytes compared with those in non-HBV HCC. Notably, we report the enrichment of metallothioneins (MTs), particularly MT1G, in HBV-related HCC TAMs, which is associated with a worse prognosis. HBV-tumor-infiltrated CD8⁺ T cells exhibit a dysfunctional cytotoxic phenotype, characterized by upregulated MDK and CTLA4 expression and reduced IFN-γ production, unlike the non-HBV-HCC. Additionally, HBV-HCC exhibits immunosuppressive ligand-receptor interactions, whereas non-HBV-HCC exhibits antitumor ligand-receptor interactions. Our deeper understanding of the HBV-HCC ecosystem using Viral-Track integrated scRNA sequencing provides insights into immune evasion mechanisms and HBV lymphotropism associated with viral persistence.