HBX-driven tumor-intrinsic MIF contributes to CD8⁺ T cell exhaustion in HBV-related hepatocellular carcinoma

HBV-related hepatocellular carcinoma (HCC) is characterized by an immunosuppressive tumor microenvironment and limited responsiveness to immunotherapy. However, tumor-intrinsic mechanisms linking HBV-associated oncogenic programs to CD8⁺ T-cell dysfunction remain incompletely defined. We integrated single-cell RNA sequencing datasets from HBV-positive and HBV-negative HCC samples to characterize immune remodeling and tumor–immune communication. Malignant hepatocytes were identified using inferCNV and decomposed into transcriptional meta-programs by consensus non-negative matrix factorization. Ligand–receptor interactions were inferred using CellChat and validated using independent single-cell datasets, spatial transcriptomics, multiplex immunofluorescence, HBX overexpression, MIF knockdown, and an immunocompetent orthotopic mouse model. HBV-positive tumors showed enrichment of exhausted CD8⁺ T cells and increased intercellular communication complexity. MIF signaling was identified as an enriched tumor-to-CD8⁺ T-cell communication pathway, potentially mediated through CD74-associated receptor complexes. Malignant cells from HBV-positive HCC selectively upregulated MIF, which was associated with HBV-enriched viral-response and stress-adaptive transcriptional programs. Spatial transcriptomics and tissue-level imaging confirmed elevated tumor-intrinsic MIF expression in HBV-positive tumors. Functionally, HBX enhanced MIF expression and secretion, promoted tumor growth, and was associated with increased accumulation of exhaustion marker–positive CD8⁺ T cells. MIF silencing attenuated tumor progression and reduced exhaustion-associated phenotypes in the context of HBX overexpression in vivo. In primary human CD8⁺ T-cell co-culture assays, MIF silencing partially restored CD8⁺ T-cell effector activity, as reflected by increased GZMB and IFN-γ secretion. These findings support a tumor-intrinsic HBX–MIF axis that may contribute to CD8⁺ T-cell exhaustion in HBV-related HCC and suggest MIF-mediated tumor–immune communication as a potential therapeutic target.