Impact of Preexisting Hepatitis C Virus Genotype 6 NS3, NS5A, and NS5B Polymorphisms on theIn VitroPotency of Direct-Acting Antiviral Agents

Abstract

HCV genotype 6 (GT-6) is found predominantly in East and Southeast Asia. Clinical studies have focused on patients infected with hepatitis C virus (HCV) GT-6a, where high sustained virologic response (SVR) rates to direct-acting antivirals (DAAs) have been achieved. However, GT-6 is highly diverse, with 29 reported subtypes. We explored the diversity of GT-6 polymorphisms at residues associated with DAA resistance, their impact on DAA in vitro potency when evaluated in a GT-6a consensus replicon, and their association with specific GT-6 subtypes. GT-6 sequences from 25 patient-derived samples and 105 sequences from the U.S. HCV database were compared, and substitutions at resistance-associated residue positions were phenotyped against different DAAs. Preexisting resistance-associated substitutions (RASs) to NS3 protease (A156V and D168E) and NS5B nucleotide (L159F and S282C) inhibitors were rare (Ͻ4%). Preexisting RASs to NS5A inhibitors were common, especially at L28 (A/F/G/M/T/V) and R30 (E/N/S). In vitro susceptibilities of NS5A-L28A and -L28T were dramatically reduced against all tested NS5A drugs (90% effective concentration [EC 90 ] range, 119 to 2,032 nM) compared with susceptibilities against a GT-6a consensus replicon (EC 90 range, 0.1 to 19 nM). These L28 RASs preexisted in combination with R30S (EC 90 [L28A-R30S] of Ն720 nM or EC 90 [L28T-R30S] of Ն128 nM against tested DAAs) or as L28T-L31I (EC 90 [tested DAAs] of Ͼ5,000 nM) and were detected in evaluated GT-6b and -6f sequences. NS5A-L28A-R30A, observed in GT-6r, did not replicate. In conclusion, HCV GT-6b, GT-6f, and GT-6r sequences harbored highly resistant RASs to all evaluated NS5A drugs. Therefore, monitoring SVR in patients infected with these GT-6 subtypes treated with NS5A drug-containing regimens is suggested to confirm any association between noted NS5A polymorphisms and treatment failure.

Genetic diversity of genotype 6 HCV infections in France: epidemiology and consequences for treatment strategy

“…Analysis of GT6 NS3 from different subtypes identified a rare D168E polymorphism not described as modifying anti-NS3 susceptibility and no such polymorphism was found on our patient sequences. Our findings and reports from the literature are thus quite reassuring in term of second generation anti-NS3 efficacy on all GT6 subtypes, with no evidence for polymorphism significantly modifying DAA potency 11 .…”

“…Indeed, McPhee et al report in vitro high level of resistance to velpatasvir in replicons carrying both NS5A L28T/R30S RAS, a common motif found exclusively in subtype 6f and also found in one of our patients 11 . We believe awareness should be promoted when treating uncommon subtypes that may not be as susceptible to the so-called "pan-genotypic" current combinations, even though current real-world efficacy data are very reassuring.…”

“…Interestingly, 3 GT6e cirrhotic patients included in the Hepather cohort and treated with sofosbuvir-ledipasvir, achieved SVR. McPhee et al also describe a double variant NS5A L28A/R30S with a more than 10,000 fold reduced sensitivity to ledipasvir 11 . This motif was detected in one GT6f patient who had a sustained virological response to 3 months of SOF-LED + ribavirin combination treatment.…”

“…Indeed, most data on DAA efficacy to date have been derived from clinical trials conducted in Western countries, where GT6-HCV is rare and only few data sets exist for countries such as Cambodia and Vietnam with higher GT6 prevalence. Few in vitro studies have clearly identified significant difference in DAA potency according to GT6 subtypes or even selection of resistance [9][10][11][12] . Importantly, while their data validate DAA activity on subtype 6a prototypes, efficacy on less frequent subtypes is missing despite great polymorphism on residues known to influence DAA efficacy 13 .…”

Genetic diversity of genotype 6 HCV infections in France: Epidemiology and consequences for treatment strategy

“…Indeed, most data on DAA efficacy to date have been derived from clinical trials conducted in Western countries, where GT6‐HCV is rare and only few data sets exist for countries such as Cambodia and Vietnam with higher GT6 prevalence. Few in vitro studies have clearly identified significant difference in DAA potency according to GT6 subtypes or even selection of resistance . Importantly, while their data validate DAA activity on subtype 6a prototypes, efficacy on less frequent subtypes is missing despite great polymorphism on residues known to influence DAA efficacy .…”