Preclinical Characterization of the Antiviral Activity of SCH 900518 (Narlaprevir), a Novel Mechanism-Based Inhibitor of Hepatitis C Virus NS3 Protease

Small-molecule hepatitis C virus (HCV) NS3 protease inhibitors such as boceprevir (SCH 503034) have been shown to have antiviral activity when they are used as monotherapy and in combination with pegylated alpha interferon and ribavirin in clinical trials. Improvements in inhibitor potency and pharmacokinetic properties offer opportunities to increase drug exposure and to further increase the sustained virological response. Exploration of the structure-activity relationships of ketoamide inhibitors related to boceprevir has led to the discovery of SCH 900518, a novel ketoamide protease inhibitor which forms a reversible covalent bond with the active-site serine. It has an overall inhibition constant (K*_i) of 7 nM and a dissociation half-life of 1 to 2 h. SCH 900518 inhibited replicon RNA at a 90% effective concentration (EC₉₀) of 40 nM. In biochemical assays, SCH 900518 was active against proteases of genotypes 1 to 3. A 2-week treatment with 5× EC₉₀ of the inhibitor reduced the replicon RNA level by 3 log units. Selection of replicon cells with SCH 900518 resulted in the outgrowth of several resistant mutants (with the T54A/S and A156S/T/V mutations). Cross-resistance studies demonstrated that the majority of mutations for resistance to boceprevir and telaprevir caused similar fold losses of activity against all three inhibitors; however, SCH 900518 retained more activity against these mutants due to its higher intrinsic potency. Combination treatment with alpha interferon enhanced the inhibition of replicon RNA and suppressed the emergence of resistant replicon colonies, supporting the use of SCH 900518-pegylated alpha interferon combination therapy in the clinic. In summary, the results of the preclinical characterization of the antiviral activity of SCH 900518 support its evaluation in clinical studies.Hepatitis C virus (HCV) is a major cause of chronic hepatitis worldwide. Despite recent advances, the eradication of chronic HCV infection remains challenging. Approximately 50% of patients infected with the most prevalent form of the virus (genotype 1) fail to respond to treatment with the current standard of care of pegylated alpha interferon in combination with ribavirin (4, 13). Thus, there is a great unmet medical need for the development of new agents with activity against HCV to improve the sustained virological response (SVR). The HCV genome is translated as a single polyprotein precursor which is processed by cellular and viral proteases into structural proteins (the C, E1, and E2 proteins), followed by nonstructural (NS) proteins (the p7, NS2, NS3, NS4A, NS4B, NS5A, and NS5B proteins). The NS3 protein is multifunctional, with the N-terminal region (amino acids 1 to 181) encoding a serine protease and the remaining polypeptide encoding a helicase. The NS3 protease noncovalently binds to the cofactor NS4A, which contributes to the formation and stability of the active site and helps to anchor the NS3/NS4A complex to the membrane. The NS3 protease is responsible for the processing of the HCV polyprotein at the NS3-NS4A, NS4A-NS4B, NS4B-NS5A, and NS5A-NS5B junctions (for a review, see reference 19).The NS3 protease is essential for viral replication (6) and represents an important target for antiviral therapy. The clinical efficacies of NS3 protease inhibitors were first demonstrated with compounds targeting the enzyme active site in monotherapy and in combination with pegylated interferon: BILN 2061, a macrocyclic tripeptide in a short monotherapy trial (8), and boceprevir (SCH 503034) and telaprevir (VX-950), both of which are ketoamide derivatives (21, 22, 27, 28). Mutations conferring resistance to these small-molecule inhibitors have been identified in the NS3 protease domain by selection of replicon cells in the presence of compounds; many of the mutations selected in vitro have also been detected in patients during clinical trials of boceprevir and telaprevir (9, 11, 21, 25, 27, 28). Although encouraging clinical results have been achieved with these first-generation protease inhibitors, additional improvements in inhibitor potency and pharmacokinetic properties offers opportunities to increase drug coverage and to further increase the SVR.The combination of medicinal chemistry and structure-based design has led to the synthesis of a new compound, SCH 900518 (narlaprevir; Fig. ​Fig.1)1) (1), with improved antiviral activity in the replicon system compared with the activities of boceprevir (12) and telaprevir (16, 17). The characterization of its inhibition mechanism and in vitro resistance profile is presented in this report.Open in a separate windowFIG. 1.Structure of SCH 900518. P1 to P4 refer to different side chains.