IDDF2026-ABS-0401 A multi-cohort single-cell transcriptome atlas identifies ERO1A-expressing angiogenesis-associated macrophages as the dominant pro-tumorigenic subset driving hepatocellular carcinoma progression

Background Hepatocellular carcinoma (HCC) is a leading cause of cancer mortality worldwide, with poor outcomes in advanced disease driven by an immunosuppressive tumor microenvironment. Tumor-associated macrophages (TAMs) represent the predominant immune population in this setting, yet their functional heterogeneity and prognostic relevance remain poorly defined. We aimed to generate a unified single-cell atlas of TAMs from multiple clinical cohorts to delineate functionally distinct subsets, uncover their origins and interactions, and identify clinically actionable biomarkers and therapeutic targets. Methods Single-cell RNA sequencing data from five independent GEO cohorts (GSE202642, GSE189903, GSE151530, GSE166635, GSE242889) were integrated, encompassing 46 HCC tumors and 13 adjacent non-tumor liver samples (256,236 high-quality cells after quality control). Analysis was performed using Seurat for clustering and integration, Monocle for pseudotime trajectories, GSEA/GSVA for pathway enrichment, CellChat for intercellular communication, and TCGA-LIHC bulk RNA-seq for survival and clinical correlation analyses (univariate Cox regression). Results Eight transcriptionally distinct macrophage subsets were identified based on functional gene modules (IDDF2026-ABS-0401 Figure 1). Monocyte-derived angiogenesis-associated macrophages (Angio-Mac), selectively marked by high SPP1 expression, demonstrated the most striking expansion in tumor versus adjacent tissue (3.65-fold increase) and progressive enrichment from early (BCLC-A) to intermediate (BCLC-B) stages (IDDF2026-ABS-0401 Figure 2). These cells exhibited a pro-tumorigenic profile characterized by upregulated mTORC1 signaling, hypoxia, glycolysis, M2-like polarization, and reduced antigen-presentation capacity (IDDF2026-ABS-0401 Figure 3). Unbiased survival analysis in TCGA-LIHC identified ERO1A as the top Angio-Mac-specific gene associated with advanced disease and poor overall survival (IDDF2026-ABS-0401 Figure 4). Cell–cell communication analysis further showed intense outgoing angiogenic signals (SPP1, VEGFA) from Angio-Mac and strong tumor-specific SPP1–CD44 interactions with T/NK cells (IDDF2026-ABS-0401 Figure 5). Intermediate monocytes were identified as the potential precursors that differentiate into Angio-Mac within the HCC microenvironment (IDDF2026-ABS-0401 Figure 6). Conclusions ERO1A+ Angio-Mac represents the predominant immunosuppressive and pro-angiogenic TAM subset in HCC. This study establishes ERO1A and SPP1 as clinically actionable prognostic biomarkers and prioritizes Angio-Mac – along with its circulating intermediate monocyte precursors – as high-value targets for novel diagnostic monitoring and therapeutic strategies in advanced HCC.