Circulating Cell-Free DNA Profiling Predicts the Therapeutic Outcome in Advanced Hepatocellular Carcinoma Patients Treated with Combination Immunotherapy

SIMPLE SUMMARY: Atezolizumab/bevacizumab (Atezo/Bev) combination immunotherapy has become a front-line therapy for unresectable hepatocellular carcinoma (u-HCC), but some patients are initially nonresponders. We investigated the potential of cell-free DNA (cfDNA)/circulating tumor DNA (ctDNA) as biomarkers for predicting the therapeutic outcome of u-HCC patients treated with anti-programmed cell death1-ligand1 (PD-L1)/vascular endothelial growth factor (VEGF) therapy. Patients with high levels of cfDNA showed a significantly lower overall response rate and shorter progression-free survival and overall survival (OS) than those with low levels of cfDNA. Ultradeep sequencing of cfDNA showed that the telomerase reverse transcriptase (TERT) promoter, tumor protein 53 (TP53) and catenin beta 1 (CTNNB1) were the most frequently mutated genes in ctDNA. Lastly, a TERT ctDNA mutation and a high alpha-fetoprotein (AFP) level were independent predictors of shorter OS in u-HCC patients treated with Atezo/Bev therapy and could stratify their prognoses. Collectively, cfDNA/ctDNA profiling may be useful to predict therapeutic outcome in u-HCC patients treated with Atezo/Bev therapy. ABSTRACT: Combination immunotherapy with anti-programmed cell death1-ligand1 (PD-L1) and anti-vascular endothelial growth factor (VEGF) antibodies has become the standard treatment for patients with unresectable HCC (u-HCC). However, limited patients obtain clinical benefits. Cell-free DNA (cfDNA) in peripheral blood contains circulating tumor DNA (ctDNA) that reflects molecular abnormalities in tumor tissue. We investigated the potential of cfDNA/ctDNA as biomarkers for predicting the therapeutic outcome in u-HCC patients treated with anti-PD-L1/VEGF therapy. We enrolled a multicenter cohort of 85 HCC patients treated with atezolizumab and bevacizumab (Atezo/Bev) between 2020 and 2021. Pretreatment plasma was collected, and cfDNA levels were quantified. Ultradeep sequencing of cfDNA was performed with a custom-made panel for detecting mutations in 25 HCC-related cancer genes. We evaluated the association of cfDNA/ctDNA profiles and clinical outcomes. Patients with high plasma cfDNA levels showed a significantly lower response rate and shorter progression-free survival and overall survival (OS) than those with low cfDNA levels. ctDNA detected in 55% of HCC patients included the telomerase reverse transcriptase (TERT) promoter in 31% of these patients, tumor protein 53 (TP53) in 21%, catenin beta 1 (CTNNB1) in 13% and phosphatase and tensin homolog (PTEN) in 7%. The presence or absence of ctDNA did not predict the efficacy of Atezo/Bev therapy. Twenty-six patients with a TERT mutation had significantly shorter OS than those without. The presence of a TERT mutation and alpha-fetoprotein (AFP) ≥ 400 ng/mL were independent predictors of poor OS according to multivariate Cox proportional hazard analysis and could be used to stratify patients treated with Atezo/Bev therapy based on prognosis. In conclusion, pretreatment cfDNA/ctDNA profiling may be useful for predicting the therapeutic outcome in u-HCC patients treated with anti-PD-L1/VEGF therapy.