Multi-omics investigation for the tumor immune microenvironment and spatial heterogeneity in hepatocellular carcinoma with PVTT: MyCAFs and NID1 in immune evasion.

Abstract

e16171 Background: Hepatocellular carcinoma (HCC), accounting for 70%-90% of primary liver cancers, often involves portal vein tumor thrombus (PVTT), a significant adverse prognostic factor linked to worsened liver function and treatment challenges. This study uses multi-omics analysis to investigate the tumor immune microenvironment in HCC with PVTT, focusing on the role of cancer-associated fibroblasts (CAFs) in PVTT formation and identifying potential therapeutic targets to improve clinical outcomes. Methods: We utilized nCounter technology, single-cell RNA sequencing (scRNA-seq), digital spatial profiling (DSP), and Olink proteomics to investigate intra-tumoral spatial heterogeneity from transcriptomics, proteomics, and immunology perspectives. Key molecules promoting PVTT formation were identified via differentially expressed genes (DEGs) analysis, spatial expression and single-cell analysis. Identified Acarbose as a potential drug for treating PVTT was conducted using 3D QSAR, pharmacophore modeling, and molecular dynamics simulations. The clinical potential of Acarbose targeting NID1 in PVTT treatment was further validated through cell proliferation assays, immune-functional mouse models, immunofluorescence, and real-world clinical data analysis. Results: A comprehensive multi-omics analysis of 85 samples from 47 HCC patients revealed that myofibroblast-associated cells (myCAFs) dominate the tumor microenvironment (TME) of PVTT, shaping a tumor immune-suppressive environment. The macrophage-to-myofibroblast transition (MMT) induced myCAF formation, establishing an immune barrier in the primary tumor. A cross-analysis of DEGs from DSP data and scRNA-seq identified Nidogen1 (NID1) as a key gene driving PVTT formation. NID1 was upregulated in MMT cells and myCAFs within the PVTT TME, contributing to immunosuppression and PVTT formation. Computational virtual screening identified acarbose as a potential therapeutic agent. When combined with tislelizumab, acarbose significantly enhanced T cell-mediated tumor cytotoxicity, inhibited tumor growth, and reduced NID1 expression, demonstrating superior efficacy compared to anti-PD-1 monotherapy, both in vitro and in vivo. Real-world data showed that long-term use of acarbose in diabetic HCC patients reduced NID1 levels and prevented PVTT formation, supporting acarbose as a promising strategy to target NID1 and inhibit PVTT. Conclusions: Our findings demonstrate the critical role of myCAFs in shaping the tumor immune-suppressive environment that promotes PVTT formation. Targeting NID1, a key driver of this process, with acarbose effectively disrupts the myCAF-mediated immune barrier and synergistically enhances the efficacy of immunotherapy.