Abstract 7422: Investigation of acquired resistance to atezolizumab-bevacizumab in hepatocellular carcinoma: Paired tumor microenvironment analysis in a mouse model and human subjects

Abstract Background Immunotherapy combinations are the standard first-line treatment for advanced hepatocellular carcinoma (HCC), yet most responders eventually develop resistance. No standard second-line therapy exists post-progression, and elucidating mechanisms of acquired resistance is essential to guide future strategies. Methods Hep53.4 murine HCC cells were subcutaneously implanted into C57BL/6 mice and treated with anti-programmed death-ligand 1 (PD-L1) monoclonal antibody (mAb) (clone 6E11, 5 mg/kg) and anti-vascular endothelial growth factor (VEGF) mAb (clone B20-4.1.1, 5 mg/kg). Tumors were harvested at baseline, responding, and resistant phase (n=6 per group) for analysis using NanoString nCounter® Pan-Cancer IO 360™, bulk RNA sequencing (RNA-seq), and immunohistochemistry (IHC) staining. Paired human tumor samples (n=4) at pre- and post-progression on atezolizumab-bevacizumab were analyzed with RNAseq. Immune cell fractions were estimated using CIBERSORT. Results NanoString analysis of different response phases - baseline, responding, and resistant phase of murine Hep53.4 HCC tumors treated with anti-PD-L1 and anti-VEGF mAb revealed distinct tumor-immune microenvironment (TME) changes. Total tumor-infiltrating lymphocytes and cytotoxic T cells increased in responding tumors but declined in resistant ones, whereas exhausted and regulatory T cells showed the opposite trend. No significant changes were observed in B cells or dendritic cells. Notably, mast cells decreased in responding tumors but increased significantly in resistant ones. RNA-seq with CIBERSORT and IHC validated immune dynamics, including mast cell fluctuations, in murine tumors. Paired human tumors also showed mast cell enrichment at progression versus baseline. Conclusion TME profiling of a murine HCC model treated with anti-PD-L1/VEGF mAbs demonstrated a dynamic shift in immune contexture, mirroring the immune response-resistance phenomenon. Mast cells decreased in response but increased upon resistance—a trend confirmed in human samples. Further investigation of the role of mast cells in acquired resistance is warranted. Citation Format: Tsung-Hao Liu, Li-Chun Lu, Yu-Yun Shao, Shu-Han Yang, Chi-Yuan Yao, Chia-Lang Hsu, Ying-Chun Shen, Ann-Lii A. Cheng, Chih-Hung Hsu. Investigation of acquired resistance to atezolizumab-bevacizumab in hepatocellular carcinoma: Paired tumor microenvironment analysis in a mouse model and human subjects [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 7422.