The Hepatitis C Virus Replicon Presents a Higher Barrier to Resistance to Nucleoside Analogs than to Nonnucleoside Polymerase or Protease Inhibitors

Abstract

Specific inhibitors of hepatitis C virus (HCV) replication that target the NS3/4A protease (e.g., VX-950) or the NS5B polymerase (e.g., R1479/R1626, PSI-6130/R7128, NM107/NM283, and HCV-796) have advanced into clinical development. Treatment of patients with VX-950 or HCV-796 rapidly selected for drug-resistant variants after a 14-day monotherapy treatment period. However, no viral resistance was identified after monotherapy with R1626 (prodrug of R1479) or NM283 (prodrug of NM107) after 14 days of monotherapy. Based upon the rapid selection of resistance to the protease and nonnucleoside inhibitors during clinical trials and the lack of selection of resistance to the nucleoside inhibitors, we used the replicon system to determine whether nucleoside inhibitors demonstrate a higher genetic barrier to resistance than protease and nonnucleoside inhibitors. Treatment of replicon cells with nucleoside inhibitors at 10 and 15 times the 50% effective concentration resulted in clearance of the replicon, while treatment with a nonnucleoside or protease inhibitor selected resistant colonies. In combination, the presence of a nucleoside inhibitor reduced the frequency of colonies resistant to the other classes of inhibitors. These results indicate that the HCV replicon presents a higher barrier to the selection of resistance to nucleoside inhibitors than to nonnucleoside or protease inhibitors. Furthermore, the combination of a nonnucleoside or protease inhibitor with a nucleoside polymerase inhibitor could have a clear clinical benefit through the delay of resistance emergence.Hepatitis C virus (HCV) is a positive-strand RNA virus that is a member of the Hepacivirus genus within the Flaviridae family. There are an estimated 170 million individuals chronically infected with HCV worldwide, which amounts to almost 3% of the global population (1). In the United States, an estimated 20,000 new HCV infections occurred in 2005, adding to the approximately 4 million individuals previously infected with HCV (2, 38). Liver cirrhosis, as a result of HCV infection, is currently the leading reason justifying liver transplantation; however, reinfection occurs immediately posttransplantation and can result in graft loss (39).The current treatment of pegylated alpha interferon in combination with ribavirin results in a sustained viral response in approximately 50% of HCV patients infected with genotype (GT) 1 virus, the most prevalent GT worldwide. Therefore, a specific HCV antiviral therapy is highly desirable. Viral proteases and viral polymerases have been validated as clinically effective targets for a number of different viruses, including human immunodeficiency virus, hepatitis B virus, and herpesviruses (6,7,14,15). Two potential drug targets encoded by HCV are the NS3/4A serine protease and the NS5B RNA-dependent RNA polymerase (5). Several anti-HCV compounds that inhibit the activity of either the NS3/4A protease or the NS5B RNA-dependent RNA polymerase have resulted in decreased viral loads when administered to HCV-infected pat...

Hepatitis C Virus Nucleotide Inhibitors PSI-352938 and PSI-353661 Exhibit a Novel Mechanism of Resistance Requiring Multiple Mutations within Replicon RNA

“…Mutations that confer resistance to the NS3 protease, NS5A, and NS5B nonnucleoside inhibitors have been readily selected both in vitro and in vivo (7,16,22,24). In contrast, a higher barrier of resistance exists for NS5B nucleoside analogs (17). To date, only two mutations within HCV NS5B have been found to be associated with decreased susceptibility to nucleoside/-tide analogs: S96T, which confers resistance to 4Ј-azidocytidine (R1479) (13), and S282T, which confers resistance to 2Ј-Cmethyl-modified nucleoside/-tides (e.g., IDX184 and INX-08189) (18,29; J. F. McCarville et al, presented at the 5th International Workshop on Hepatitis C-Resistance and New Compounds, Boston, MA, 24 to 25 June 2010) and 2Ј-F-2Ј-Cmethyl pyrimidine nucleoside/-tide analogs (e.g., RG7128 and PSI-7977) (1,26).…”

Genotype and Subtype Profiling of PSI-7977 as a Nucleotide Inhibitor of Hepatitis C Virus

“…Two classes of inhibitors directed at the activity of NS5B are currently in clinical development: nucleoside/ tide analogs, which target the active site, and nonnucleoside inhibitors (NNIs), which target other regions of the polymerase (39). Compared to NNIs, nucleoside/tide inhibitors have broader genotype coverage and a higher barrier to viral resistance (9,24,28). Previously, we reported the anti-HCV activity of PSI-7851, a phosphoramidate nucleotide prodrug of 2=-F-2=-C-methyluridine monophosphate (18).…”

Use of Illumina Deep Sequencing Technology To Differentiate Hepatitis C Virus Variants

“…Substitutions that generated boceprevir resistance included those detected in a patient treated with telaprevir, plus V170A/T and V55A substitutions (13,41). Of the nonnucleoside inhibitors of RdRp displaying inhibitory activities against the RdRp enzyme at NS5B, few have been reported in the in vivo resistance data (27). This is because studies of antiviral efficacy are generally limited to 3 to 5 days.…”