title: Unraveling the structural basis of grazoprevir potency against clinically relevant substitutions in hepatitis C virus NS3/4A protease from genotype 1a.
abstract: Grazoprevir is a potent pan-genotype and macrocyclic inhibitor of hepatitis C virus (HCV) NS3/4A protease and was developed for treating chronic HCV infection. In HCV genotype (GT) 1a, grazoprevir maintains potent activity against a majority of NS3 resistance-associated amino acid substitutions, including the highly prevalent and naturally occurring Q80K polymorphism that impacts simeprevir, another NS3/4A protease inhibitor. The basis for an unexpected difference in the clinical impact of some NS3 substitutions was investigated. Phenotypic analysis of resistance-associated substitutions identified in NS3 from GT1a-infected patients who failed therapy with grazoprevir (in combination with elbasvir, an inhibitor of HCV NS5A protein) showed that positions 56, 156, and 168 in NS3 were most impactful because they diminished protein-inhibitor interactions. Although an amino acid substitution from aspartic acid to alanine at position 168 (D168A) reduced the potency of grazoprevir, its combination with R155K unexpectedly nullified this effect. Molecular dynamics and free-energy surface studies indicated that Asp-168 is important in anchoring Arg-155 for ligand binding but is not critical for Lys-155 because of the inherent flexibility of its side chain. Moreover, modeling studies supported a strong direct cation-heterocycle interaction between the Lys-155 side chain of the double substitution, R155K/D168A, and the lone pair on the quinoxaline in grazoprevir. This unique interaction provides a structural basis for grazoprevir's higher potency than simeprevir, an inhibitor to which the double substitution confers a significant reduction in potency. Our findings are consistent with the detection of R155K/D168A in NS3 from virologic failures treated with simeprevir but not grazoprevir.
authors.affiliation: From the Departments of Chemistry, Modeling and Informatics and.
authors.forename: Zhuyan
authors.initials: Z
authors.lastname: Guo
authors.affiliation: Infectious Diseases, Merck Research Laboratories, Kenilworth, New Jersey 07033.
authors.forename: Stuart
authors.initials: S
authors.lastname: Black
authors.affiliation: From the Departments of Chemistry, Modeling and Informatics and.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Hu
authors.affiliation: Infectious Diseases, Merck Research Laboratories, Kenilworth, New Jersey 07033.
authors.forename: Patricia
authors.initials: P
authors.lastname: McMonagle
authors.affiliation: Infectious Diseases, Merck Research Laboratories, Kenilworth, New Jersey 07033.
authors.forename: Paul
authors.initials: P
authors.lastname: Ingravallo
authors.affiliation: Infectious Diseases, Merck Research Laboratories, Kenilworth, New Jersey 07033.
authors.forename: Robert
authors.initials: R
authors.lastname: Chase
authors.affiliation: Infectious Diseases, Merck Research Laboratories, Kenilworth, New Jersey 07033.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Curry
authors.affiliation: Infectious Diseases, Merck Research Laboratories, Kenilworth, New Jersey 07033 Ernest_asanteappiah@merck.com.
authors.forename: Ernest
authors.initials: E
authors.lastname: Asante-Appiah
mesh_terms: D000577:Amides; D019943:Amino Acid Substitution; D002219:Carbamates; D045744:Cell Line, Tumor; D003521:Cyclopropanes; D016174:Hepacivirus / Q000201:enzymology* / Q000235:genetics; D006526:Hepatitis C / Q000188:drug therapy / Q000201:enzymology / Q000235:genetics; D006801:Humans; D056004:Molecular Dynamics Simulation; D020125:Mutation, Missense; D011810:Quinoxalines / Q000737:chemistry* / Q000627:therapeutic use; D000069616:Simeprevir / Q000737:chemistry / Q000627:therapeutic use; D013449:Sulfonamides; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; D047029:Lactams, Macrocyclic
chemical_list: D000577:Amides; D002219:Carbamates; D003521:Cyclopropanes; D011810:Quinoxalines; D000069616:Simeprevir; D013449:Sulfonamides; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C578009:grazoprevir; D047029:Lactams, Macrocyclic
keywords: crystal structure; drug resistance; hepatitis C virus (HCV); molecular dynamics; protease inhibitor