title: Substitutions at NS3 Residue 155, 156, or 168 of Hepatitis C Virus Genotypes 2 to 6 Induce Complex Patterns of Protease Inhibitor Resistance.
abstract: Various protease inhibitors (PIs) currently are becoming available for treatment of hepatitis C virus (HCV). For genotype 1, substitutions at NS3 protease positions 155, 156, and 168 are the main determinants of PI resistance. For other genotypes, similar substitutions were selected during PI treatment but were not characterized systematically. To elucidate the impact of key PI resistance substitutions on genotypes 2 to 6, we engineered the substitutions R155A/E/G/H/K/Q/T, A156G/S/T/V, and D/Q168A/E/G/H/N/V into HCV recombinants expressing genotype 2 to 6 proteases. We evaluated viral fitness and sensitivity to nine PIs (telaprevir, boceprevir, simeprevir, asunaprevir, vaniprevir, faldaprevir, paritaprevir, deldeprevir, and grazoprevir) in Huh7.5 cells. We found that most variants showed decreased fitness compared to that of the original viruses. Overall, R155K, A156G/S, and D/Q168A/E/H/N/V variants showed the highest fitness; however, genotype 4 position 168 variants showed strong fitness impairment. Most variants tested were resistant to several PIs. Resistance levels varied significantly depending on the specific substitution, genotype, and PI. For telaprevir and boceprevir, specific 155 and 156, but not 168, variants proved resistant. For the remaining PIs, most genotype 2, 4, 5, and 6, but not genotype 3, variants showed various resistance levels. Overall, grazoprevir (MK-5172) had the highest efficacy against original viruses and variants. This is the first comprehensive study revealing the impact of described key PI resistance substitutions on fitness and PI resistance of HCV genotypes 2 to 6. In conclusion, the studied substitutions induced resistance to a panel of clinically relevant PIs, including the newer PIs paritaprevir, deldeprevir, and grazoprevir. We discovered complex patterns of resistance, with the impact of substitutions varying from increased sensitivity to high resistance.
authors.affiliation: Copenhagen Hepatitis C Program (CO-HEP), Department of Infectious Diseases and Clinical Research Centre, Hvidovre Hospital and Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Sanne B
authors.initials: SB
authors.lastname: Jensen
authors.affiliation: Copenhagen Hepatitis C Program (CO-HEP), Department of Infectious Diseases and Clinical Research Centre, Hvidovre Hospital and Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Stéphanie B N
authors.initials: SB
authors.lastname: Serre
authors.affiliation: Copenhagen Hepatitis C Program (CO-HEP), Department of Infectious Diseases and Clinical Research Centre, Hvidovre Hospital and Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Daryl G
authors.initials: DG
authors.lastname: Humes
authors.affiliation: Copenhagen Hepatitis C Program (CO-HEP), Department of Infectious Diseases and Clinical Research Centre, Hvidovre Hospital and Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Santseharay
authors.initials: S
authors.lastname: Ramirez
authors.affiliation: Copenhagen Hepatitis C Program (CO-HEP), Department of Infectious Diseases and Clinical Research Centre, Hvidovre Hospital and Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Yi-Ping
authors.initials: YP
authors.lastname: Li
authors.affiliation: Copenhagen Hepatitis C Program (CO-HEP), Department of Infectious Diseases and Clinical Research Centre, Hvidovre Hospital and Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Jens
authors.initials: J
authors.lastname: Bukh
authors.affiliation: Copenhagen Hepatitis C Program (CO-HEP), Department of Infectious Diseases and Clinical Research Centre, Hvidovre Hospital and Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark judith.margarete.gottwein@regionh.dk.
authors.forename: Judith M
authors.initials: JM
authors.lastname: Gottwein
mesh_terms: D000577:Amides; D019943:Amino Acid Substitution; D000998:Antiviral Agents / Q000494:pharmacology; D002219:Carbamates; D002460:Cell Line; D003521:Cyclopropanes; D024882:Drug Resistance, Viral / Q000235:genetics*; D005838:Genotype; D016174:Hepacivirus / Q000187:drug effects* / Q000235:genetics*; D006801:Humans; D009842:Oligopeptides / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011810:Quinoxalines / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013449:Sulfonamides; D017361:Viral Nonstructural Proteins / 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; D000998:Antiviral Agents; D002219:Carbamates; D003521:Cyclopropanes; D009842:Oligopeptides; D011480:Protease Inhibitors; D011810:Quinoxalines; D011994:Recombinant Proteins; 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; C486464:telaprevir