Design, synthesis and evaluation of a potent substrate analog inhibitor identified by scanning Ala/Phe mutagenesis, mimicking substrate co-evolution, against multidrug-resistant HIV-1 protease

Abstract

Multidrug-resistant (MDR) clinical isolate-769, human immunodeficiency virus type-1 (HIV-1) protease (PDB ID: 1TW7), was shown to exhibit wide-open flaps and an expanded active site cavity, causing loss of contacts with protease inhibitors. In the current study, the expanded active site cavity of MDR769 HIV-1 protease was screened with a series of peptide-inhibitors that were designed to mimic the natural substrate cleavage site, capsid/p2. Scanning Ala/Phe chemical mutagenesis approach was incorporated into the design of the peptide series to mimic the substrate co-evolution. Among the peptides synthesized and evaluated, a lead peptide (6a) with potent activity (IC50: 4.4 nM) was identified against the MDR769 HIV-1 protease. Isothermal titration calorimetric analysis showed favorable binding profile for 6a against both wild type and MDR769 HIV-1 protease variants. Nuclear magnetic resonance spectrum of 15N-labeled MDR769 HIV-1 protease in complex with 6a showed perturbations in chemical shifts, indicating the peptide-induced conformational changes in protease. Modeling analysis revealed multiple contacts between 6a and MDR769 HIV-1 protease. The lead peptide-inhibitor, 6a, with high potency and good binding profile can be used as the basis for developing potent small molecule inhibitors against MDR variants of HIV.

P1 and P1′ para-fluoro phenyl groups show enhanced binding and favorable predicted pharmacological properties: Structure-based virtual screening of extended lopinavir analogs against multi-drug resistant HIV-1 protease

“…Mutations from longer side chain residues to shorter side chain residues cause an overall expansion of the active site cavity, which in turn causes weaker binding of the PI due to loss of contacts [10, 15]. Recently it has been shown that a scanning Ala/Phe chemical mutagenesis approach can be effectively used to probe the expanded active site cavity of MDR769 HIV-1 protease with peptides [16]. Based on these structural analyses, it was hypothesized that by substituting the P1 and P1' phenyl groups of LPV with extended analogs of phenyl group one can restore the lost contacts in the expanded active site cavity.…”

A multi-drug resistant HIV-1 protease is resistant to the dimerization inhibitory activity of TLF-PafF

“…Due to the expanded active site cavity, PIs show loss of contacts [11, 12] resulting in loss of potency. Although peptide-inhibitors were recently shown to be active against MDR769 HIV-1 protease [13], bioavailability of such inhibitors could be a concern. Extended lopinavir analogs were recently shown to possess better predicted binding affinities against MDR769 HIV-1 protease [14] and are yet to be evaluated.…”