Multicenter Quality Control of Hepatitis C Virus Protease Inhibitor Resistance Genotyping

Abstract

ccHepatitis C virus (HCV) protease inhibitor resistance-associated substitutions are selected during triple-therapy breakthrough. This multicenter quality control study evaluated the expertise of 23 French laboratories in HCV protease inhibitor resistance genotyping. A panel of 12 well-defined blinded samples comprising two wild-type HCV strains, nine transcripts from synthetic NS3 mutant samples or from clinical strains, and one HCV RNA-negative sample was provided to the participating laboratories. The results showed that any laboratory with expertise in sequencing techniques should be able to provide reliable HCV protease inhibitor resistance genotyping. Only a 0.7% error rate was reported for the amino acid sites studied. The accuracy of substitution identification ranged from 75% to 100%, depending on the laboratory. Incorrect results were mainly related to the methodology used. The results could be improved by changing the primers and modifying the process in order to avoid cross-contamination. This study underlines the value of quality control programs for viral resistance genotyping, which is required prior to launching observational collaborative multicenter studies on HCV resistance to direct-acting antiviral agents. H epatitis C virus (HCV) resistance-associated variants have been shown to emerge rapidly with the use of direct-acting antiviral (DAA) agents with a low-to-moderate barrier to resistance when used as monotherapies (1). Despite significant viral load reductions during administration of the two most advanced protease inhibitors, telaprevir and boceprevir, viral breakthroughs often occur as a result of the outgrowth of variants bearing a number of well-identified amino acid substitutions that confer resistance to these agents (2, 3). The very early selection of resistant viral populations in patients receiving protease inhibitors is in keeping with the preexistence of such variants in most HCVinfected patients before the initiation of treatment. These variants most often preexist in very low proportions and are detectable only by means of next-generation sequencing (NGS) (4-7), whereas they can sometimes be present as major viral populations detectable by means of population sequencing (8-11). In a previous multicenter study, we described the natural genetic variability of the NS3 protease and the presence of variants resistant to protease inhibitors in patients infected with the HCV genotype 1 to 5 strains that are circulating in France in the absence of specific antiviral pressure (12). Although genotyping of HCV resistance to NS3 protease inhibitors has not found a clear indication in clinical practice in the context of the triple combination of pegylated alpha interferon, ribavirin, and either telaprevir or boceprevir, such genotyping is mandatory in clinical trials and cohort studies where these molecules are used, and it might find utility in the coming era of interferon-free regimens. In particular, the impact of such

Naturally Occurring Resistance-Associated Variants of Hepatitis C Virus Protease Inhibitors in Poor Responders to Pegylated Interferon-Ribavirin

“…Each of the 22 participating laboratories performed its own PCR amplification and sequence analysis of HCV NS3/4A using either the ANRS protocol for genotype 1 (n ϭ 13 laboratories) (11), the pangenotypic NS3 amplification described by Besse et al (n ϭ 5 laboratories) (7), or their own laboratory method (n ϭ 4 laboratories). All centers but one participated in the ANRS NS3 quality control, which was recently reported (16).…”

Therapy with boceprevir or telaprevir in HIV/hepatitis C virus co-infected patients to treat recurrence of hepatitis C virus infection after liver transplantation

“…In the event of virological failure, resistance to TVR or BOC was proven as previously described [14]. In the event of virological response, the viral load was monitored each month until 6 months after treatment (SVR 24).…”