title: HCV-NS3 inhibitors: determination of their kinetic parameters and mechanism.
abstract: Existing HCV protease inhibitors fall into two categories: reversible and non-covalent, such as BILN-2061, and covalent and reversible, exemplified by SCH-503034 and VX-950. In this work, the characterization of the kinetics of these three inhibitors is presented. SCH-503034 and VX-950 initially bind to the genotype 1b HCV NS3/4A protease to form a low affinity complex, with K(i) values of 5 and 5.8 microM respectively. The ability of those two compounds to form a second covalent complex (EI) results in a potency increase, with overall K(i) values of 20 and 45 nM, respectively. The increase in potency can be explained by their slow dissociation rate, forming complexes with half-lives of 2 h (VX-950) and 5 h (SCH-503034). Although BILN-2061 has been described as a fast reversible, non-covalent inhibitor, our results show a slow binding two-step mechanism. Contrary to SCH-503034 and VX-950, BILN-2061 can form a high affinity first complex with a K(i) value of 3.9 nM, and an overall K(i) of 0.14 nM. The half-life of the BILN-2061 EI complex is shorter (t(1/2) approximately 0.7 h) than that of the other two compounds. The potency of these compounds is genotype dependent, and a kinetic analysis using NS3/4A from genotype 3a indicates that the loss of potency of SCH-503034 and VX-950 relative to genotype 1 is mainly due to the slow on-rate and faster off-rate for the formation of the EI complex. In the case of BILN-2061, a better fit is obtained using a one-step model, indicating that the loss of potency is due to an increase in the off-rate of the EI complex.
authors.affiliation: Discovery Biology Department, Pfizer Global Research and Development, Sandwich, Kent CT13 9NJ, UK. Victoria.Flores@pfizer.com
authors.forename: María Victoria
authors.initials: MV
authors.lastname: Flores
authors.forename: Joanne
authors.initials: J
authors.lastname: Strawbridge
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Ciaramella
authors.forename: Romuald
authors.initials: R
authors.lastname: Corbau
mesh_terms: D019943:Amino Acid Substitution; D000998:Antiviral Agents / Q000493:pharmacokinetics / Q000494:pharmacology*; D002219:Carbamates / Q000493:pharmacokinetics / Q000494:pharmacology; D024882:Drug Resistance, Viral / Q000235:genetics; D005838:Genotype; D006207:Half-Life; D016174:Hepacivirus / Q000187:drug effects* / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D007700:Kinetics; D047028:Macrocyclic Compounds / Q000493:pharmacokinetics / Q000494:pharmacology; D008954:Models, Biological; D009154:Mutation; D009842:Oligopeptides / Q000493:pharmacokinetics / Q000494:pharmacology; D011392:Proline / Q000031:analogs & derivatives / Q000493:pharmacokinetics / Q000494:pharmacology; D011804:Quinolines / Q000493:pharmacokinetics / Q000494:pharmacology; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics; D012697:Serine Endopeptidases / Q000235:genetics; D015842:Serine Proteinase Inhibitors / Q000493:pharmacokinetics / Q000494:pharmacology*; D013844:Thiazoles / Q000493:pharmacokinetics / Q000494:pharmacology; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
chemical_list: D000998:Antiviral Agents; D002219:Carbamates; D047028:Macrocyclic Compounds; D009842:Oligopeptides; D011392:Proline; D011804:Quinolines; D011994:Recombinant Proteins; D012697:Serine Endopeptidases; D015842:Serine Proteinase Inhibitors; D013844:Thiazoles; D017361:Viral Nonstructural Proteins; C479555:BILN 2061; C084422:NS3 protein, hepatitis C virus; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C486464:telaprevir; C512204:N-(3-amino-1-(cyclobutylmethyl)-2,3-dioxopropyl)-3-(2-((((1,1-dimethylethyl)amino)carbonyl)amino)-3,3-dimethyl-1-oxobutyl)-6,6-dimethyl-3-azabicyclo(3.1.0)hexan-2-carboxamide