Acquired Genetic Variants, not Tumor Mutation Burden, Drive Resistance to Immunotherapy in Hepatocellular Carcinoma

Introduction: While immunotherapy has emerged as a promising treatment option, no reliable predictive biomarker has been established for immunotherapy in hepatocellular carcinoma (HCC). In this study, we used genetic analyses to verify the prognostic significance of tumor mutation burden (TMB) in HCC and to search for other cancer characteristics related to prognosis. Methods: Patients with HCC who received a combined therapy of atezolizumab and bevacizumab were prospectively enrolled between July 2020 and April 2023. Circulating tumor DNA analysis was performed using next-generation sequencing before immunotherapy (baseline) and 3 weeks after initiation of immunotherapy (follow-up), from which we retrieved non-synonymous baseline, follow-up, vanished (detected only at baseline), and acquired (detected only at follow-up) variants. Gene sets related to cancer hallmarks and representative oncogenic pathways were curated. Results: Forty-two patients were enrolled in this study. Higher TMB was not correlated with better prognosis. Instead, it showed an inverse relationship, with higher baseline TMB significantly associated with shorter progression-free survival (PFS) (median 2.73 vs 9.17 months, P=0.04). Among other cancer characteristics, acquired variants in the Wnt/β-catenin (PFS: P=1.14×10-4; overall survival (OS): P=0.004) and ATP-dependent chromatin remodeling (PFS: P=0.002; OS: P=0.006) pathways were significantly correlated with worse prognosis. Conclusion: In HCC, TMB is not a reliable predictive biomarker for immunotherapy. Instead, the emergence of acquired genetic variants in the Wnt/β-catenin and chromatin remodeling pathways act as a key driver of resistance.