We aimed to investigate HBx genetic elements correlated with hepatitis B virus (HBV) -related hepatocellular carcinoma (HCC) and their impact on (a) HBV replicative efficiency, (b) HBx binding to ...circular covalently closed DNA (cccDNA), (c) apoptosis and cell-cycle progression, and (d) HBx structural stability.
This study included 123 individuals chronically infected with HBV: 27 with HCC (77.9% (21/27) genotype D; 22.1% (6/27) genotype A) and 96 without HCC (75% (72/96) genotype D; 25.0% (24/96) genotype A). HepG2 cells were transfected by wild-type or mutated linear HBV genome to assess pre-genomic RNA (pgRNA) and core-associated HBV-DNA levels, HBx-binding onto cccDNA by chromatin immunoprecipitation-based quantitative assay, and rate of apoptosis and cell-cycle progression by cytofluorimetry.
F30V was the only HBx mutation correlated with HCC (18.5% (5/27) in HCC patients versus 1.0% (1/96) in non-HCC patients, p 0.002); a result confirmed by multivariate analysis. In vitro, F30V determined a 40% and 60% reduction in pgRNA and core-associated HBV-DNA compared with wild-type (p <0.05), in parallel with a significant decrease of HBx binding to cccDNA and decreased HBx stability. F30V also decreased the percentage of apoptotic cells compared with wild-type (14.8 ± 6.8% versus 19.1 ± 10.1%, p <0.01, without affecting cell-cycle progression) and increased the probability of HBx-Ser-31 being phosphorylated by PI3K-Akt kinase (known to promote anti-apoptotic activity).
F30V was closely correlated with HBV-induced HCC in vivo, reduced HBV replicative efficiency by affecting HBx-binding to cccDNA and increased anti-apoptotic HBx activity in vitro. This suggests that F30V (although hampering HBV's replicative capacity) may promote hepatocyte survival, so potentially allowing persistent production of viral progeny and initiating HBV-driven hepatocarcinogenesis. Investigation of viral genetic markers associated with HCC is crucial to identify those patients at higher risk of HCC, who hence deserve intensive liver monitoring and/or early anti-HBV therapy.
First-generation protease-inhibitors (PIs) have suboptimal efficacy in GT-1 patients with advanced liver disease, and patients experiencing treatment failure may require urgent retreatment. Our ...objective was to analyse the real-life efficacy of interferon (IFN)-free retreatment after PI-failure, and the role of genotypic-resistance-testing (GRT) in guiding retreatment choice.
In this multi-centre observational study, patients retreated with IFN-free regimens after first-generation PI-failure (telaprevir-boceprevir-simeprevir) were included. Sustained-virological-response (SVR) was evaluated at week 12 of follow-up. GRT was performed by population-sequencing.
After PI-failure, 121 patients (cirrhotic=86.8%) were retreated following three different strategies: A) with ‘GRT-guided’ regimens (N=18); B) with ‘AASLD/EASL recommended, not GRT-guided’ regimens (N=72); C) with ‘not recommended, not GRT-guided’ regimens (N=31). Overall SVR rate was 91%, but all 18 patients treated with ‘GRT-guided’ regimens reached SVR (100%), despite heterogeneity in treatment duration, use of PI and ribavirin, versus 68/72 patients (94.4%) receiving ‘AASLD/EASL recommended, not GRT-guided’ regimens. SVR was strongly reduced (77.4%) among the 31 patients who received a ‘not recommended, not GRT-guided regimen’ (p <0.01). Among 37 patients retreated with a PI, SVR rate was 89.2% (33/37). Four GT-1a cirrhotic patients failed an option (C) simeprevir-containing treatment; three out of four had a baseline R155K NS3-RAS. All seven patients treated with paritaprevir-containing regimens reached SVR, regardless of treatment duration and performance of a baseline-GRT.
Retreatment of PI-experienced patients can induce maximal SVR rates in real life. Baseline-GRT could help to optimize retreatment strategy, allowing PIs to be reconsidered when chosen after a RASs evaluation.
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