title: Potent inhibitors of HIV-1 integrase display a two-step, slow-binding inhibition mechanism which is absent in a drug-resistant T66I/M154I mutant.
abstract: Two-metal binding HIV-1 integrase inhibitors (INIs) are potent inhibitors of HIV-1 in vitro and in patients. We report here for the first time the kinetics of inhibition of integrase-catalyzed strand transfer. First, the IC(50) values for each of six structurally distinct INIs decreased when a preincubation was included: S-1360 (1.3 microM vs 0.12 microM), L-731,988 (130 nM vs 9 nM), L-870,810 (130 nM vs 4 nM), raltegravir (300 nM vs 9 nM), elvitegravir (90 nM vs 6 nM), and GSK364735 (90 nM vs 6 nM). When reactions with these INIs were initiated with integrase, progress curve analyses indicated time-dependent inhibition, which could be fitted to a two-step mechanism of binding. Overall fitted K(i) values matched the IC(50) values measured with a preincubation: S-1360 (0.17 microM), L-731,988 (34 nM), L-870,810 (2.4 nM), raltegravir (10 nM), elvitegravir (4.0 nM), and GSK364735 (2.5 nM). To begin to understand the mechanism for this slow onset of inhibition and its possible impact on drug resistance, studies of resistance mutations were initiated. T66I/M154I exhibited little if any time-dependent inhibition by any of the six INIs, as measured by differences in potency upon preincubation or by progress curve analysis. These data demonstrate that slow binding is a signature of two-metal binding INIs, and that the second slow step is required for full potency. We discuss a possible structural explanation of the second slow step of inhibition and also the relationship between loss of time-dependent inhibition and drug resistance of this important new class of HIV-1 antiretroviral drugs.
authors.affiliation: Department of Virology, GlaxoSmithKline Pharmaceuticals, 5 Moore Drive, Research Triangle Park, North Carolina 27709-3398, USA. epgarvey@bellsouth.net
authors.forename: Edward P
authors.initials: EP
authors.lastname: Garvey
authors.forename: Benjamin
authors.initials: B
authors.lastname: Schwartz
authors.forename: Margaret J
authors.initials: MJ
authors.lastname: Gartland
authors.forename: Scott
authors.initials: S
authors.lastname: Lang
authors.forename: Wendy
authors.initials: W
authors.lastname: Halsey
authors.forename: Ganesh
authors.initials: G
authors.lastname: Sathe
authors.forename: H Luke
authors.initials: HL
authors.lastname: Carter
authors.forename: Kurt L
authors.initials: KL
authors.lastname: Weaver
mesh_terms: D001483:Base Sequence; D017931:DNA Primers; D024882:Drug Resistance, Viral / Q000235:genetics*; D019427:HIV Integrase / Q000187:drug effects*; D019428:HIV Integrase Inhibitors / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics; D007700:Kinetics; D009154:Mutation
chemical_list: D017931:DNA Primers; D019428:HIV Integrase Inhibitors; D019427:HIV Integrase; C514801:p31 integrase protein, Human immunodeficiency virus 1