Abstract 5309: Single-cell-based analysis of tumor microenvironment heterogeneity in human MASLD/MASH-associated HCC: Classifications by inter-cellular network variations

Abstract Immune checkpoint inhibitor (ICI) therapy has demonstrated limited efficacy in MASLD/MASH-associated hepatocellular carcinoma (HCC), suggesting distinct anti-tumor immune responses within the tumor microenvironment (TME) of this cancer type compared to other cancers. Understanding the mechanisms driving MASLD/MASH-HCC progression and identifying novel therapeutic targets are therefore critical. While ICI therapy alone may not be sufficient, combining it with drugs targeting key cancer progression pathways may hold promising enhancement of anti-tumor efficacy in this context. In this study, we sought to elucidate the tumor-promoting pathways in MASLD/MASH-associated HCC by characterizing the comprehensive transcriptomic landscape of cancer cells and stromal cells within the tumor microenvironment (TME). We conducted single-cell RNA sequencing on the samples from 31 HCC patients and 8 healthy donors. Using this data, we classified the samples based on the cellular composition of the TME and performed interactome analysis with CellChat version 2. This analysis stratified the samples into five distinct groups: Group 1, TME with stem cell-like cancer cells and immune cell infiltration; Group 2, TME with Wnt signaling-activated cancer cells regardless of immune cell infiltration; Group 3, TME containing cancer cells not classified in the categories mentioned above with immune cell infiltration; Group 4, TME containing cancer cells not classified in the categories mentioned above without immune cell infiltration; and Group 5, TME with normal hepatocytes, as observed in healthy donor livers. Detailed analysis highlights several notable features of the tumor stroma. For example, “senescent” cancer-associated fibroblasts (CAFs) exhibiting a senescence-associated secretory phenotype (SASP) are frequently observed in the TMEs of Groups 1-3. Interactome analysis revealed a VEGFA-VEGFR2 interaction between endothelial cells and various other cell types, consistent with the hypervascularity characteristic of HCCs and supporting the clinical efficacy of VEGF inhibitors. Furthermore, group-specific interactome analysis uncovered distinct novel interactions, indicating molecular network variations among TME groups. We will present spatial distributions of each cell type in TME and TME-dependent inter-cellular interactions. We discuss their potential as biomarkers and therapeutic targets for MASLD/MASH-HCC treatment. Citation Format: Naoko Ohtani, Yoshiki Nonaka, Kanae Echizen, Yi Cheng, Takahiro Kodama. Single-cell-based analysis of tumor microenvironment heterogeneity in human MASLD/MASH-associated HCC: Classifications by inter-cellular network variations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5309.