Dissecting inter-focal heterogeneity and convergent immunosuppressive ecosystems in multifocal hepatocellular carcinoma through single-cell and spatial multi-omics

Multifocal hepatocellular carcinoma (HCC) exhibits inter-focal heterogeneity and a highly immunosuppressive microenvironment, limiting therapeutic efficacy. We performed integrated single-cell and spatial multi-omics profiling across tumor foci and adjacent tissues to delineate inter-focal heterogeneity and identify convergent immunoregulatory programs and therapeutic vulnerabilities. We identified malignant hepatocytes with focus-specific copy number alterations and functional diversity, and highlighted TMED2 as a key regulator associated with hepatocarcinogenesis and prognosis. Tumor ecosystem remodeling revealed immunosuppressive niches enriched in regulatory T cells (Tregs), SPP1+/SPRED1+ tumor-associated macrophages (TAMs), THY1+ cancer-associated fibroblasts (CAFs), and COL15A1+ vascular endothelial cells (VECs), accompanied by depletion of cytotoxic CD8+ T cells, MARCO+ TAMs, and CRHBP+ lymphatic endothelial cells, with marked inter-focal variability. Mechanistically, a TMED2-ADAR axis in malignant hepatocytes enhances COMPLEMENT (C3/C5) and PROS signaling via A-to-I RNA editing, constituting a convergent immunoregulatory program across tumor foci. This axis promotes SPP1+/SPRED1+ TAM polarization, establishing an immunosuppressive myeloid niche, which, together with Tregs, drives CD8+ T cell exhaustion via PD-L1/CD80/CD86 and MHC-I signaling. In parallel, expansion of THY1+ CAFs and COL15A1+ VECs contributes to spatially heterogeneous stromal remodeling and angiogenesis. Collectively, this atlas reveals inter-focal heterogeneity and convergent immunosuppressive ecosystems driven by tumor-immune-stromal interactions, providing mechanistic insights and therapeutic targets for multifocal HCC. Graphic abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/719788v1_ufig1.gif" ALT="Figure 1"> View larger version (76K): org.highwire.dtl.DTLVardef@1ee2a21org.highwire.dtl.DTLVardef@1100dbborg.highwire.dtl.DTLVardef@f576b0org.highwire.dtl.DTLVardef@143299d_HPS_FORMAT_FIGEXP M_FIG C_FIG