Mechanistic Studies and Modeling Reveal the Origin of Differential Inhibition of Gag Polymorphic Viruses by HIV-1 Maturation Inhibitors.

Zeyu Lin,Ira B Dicker,Nicholas A Meanwell,Dieter Drexler,Hong Yang,Alicia Regueiro-Ren,Joseph Cantone,Tricia Protack,Mark Krystal,Mark Cockett,Hao Lu,Zheng Liu,Beata Nowicka-Sans,Max Lataillade,Tian Yuan,Ronald Swanstrom

doi:10.1371/journal.ppat.1005990

Abstract

HIV-1 maturation inhibitors (MIs) disrupt the final step in the HIV-1 protease-mediated cleavage of the Gag polyprotein between capsid p24 capsid (CA) and spacer peptide 1 (SP1), leading to the production of infectious virus. BMS-955176 is a second generation MI with improved antiviral activity toward polymorphic Gag variants compared to a first generation MI bevirimat (BVM). The underlying mechanistic reasons for the differences in polymorphic coverage were studied using antiviral assays, an LC/MS assay that quantitatively characterizes CA/SP1 cleavage kinetics of virus like particles (VLPs) and a radiolabel binding assay to determine VLP/MI affinities and dissociation kinetics. Antiviral assay data indicates that BVM does not achieve 100% inhibition of certain polymorphs, even at saturating concentrations. This results in the breakthrough of infectious virus (partial antagonism) regardless of BVM concentration. Reduced maximal percent inhibition (MPI) values for BVM correlated with elevated EC50 values, while rates of HIV-1 protease cleavage at CA/SP1 correlated inversely with the ability of BVM to inhibit HIV-1 Gag polymorphic viruses: genotypes with more rapid CA/SP1 cleavage kinetics were less sensitive to BVM. In vitro inhibition of wild type VLP CA/SP1 cleavage by BVM was not maintained at longer cleavage times. BMS-955176 exhibited greatly improved MPI against polymorphic Gag viruses, binds to Gag polymorphs with higher affinity/longer dissociation half-lives and exhibits greater time-independent inhibition of CA/SP1 cleavage compared to BVM. Virological (MPI) and biochemical (CA/SP1 cleavage rates, MI-specific Gag affinities) data were used to create an integrated semi-quantitative model that quantifies CA/SP1 cleavage rates as a function of both MI and Gag polymorph. The model outputs are in accord with in vitro antiviral observations and correlate with observed in vivo MI efficacies. Overall, these findings may be useful to further understand antiviral profiles and clinical responses of MIs at a basic level, potentially facilitating further improvements to MI potency and coverage.

Highlights

There are more than 1.2 million individuals in the United States (CDC data)[1] and more than 35 million worldwide infected with HIV, with 39 million people already having died from the disease and 2.3 million new cases reported in 2013.[2]
These data were integrated into a semi-quantitative kinetic model whose outputs are in accord with in vitro antiviral observations and correlate with observed in vivo maturation inhibitors (MIs) efficacies and the results of recent crystal and cryo-electron tomography structures
HIV-1 maturation inhibitors (MIs) are a class of agents that may be effective in the treatment of HIV-1.[9,10,11,12] MIs disrupt the final step in the HIV-1 protease-mediated cleavage of the HIV-1 Gag polyprotein between capsid (CA) and spacer peptide 1 (SP1), a step which is responsible for a major conformational rearrangement of viral proteins within the virion that leads to the production of infectious virions.[13,14,15]

Summary

Introduction

There are more than 1.2 million individuals (age 13 years older) in the United States (CDC data)[1] and more than 35 million worldwide infected with HIV, with 39 million people already having died from the disease and 2.3 million new cases reported in 2013.[2]. BMS-955176 (GSK3532795) was developed as a second generation MI that possesses antiviral activity against viruses containing BVM-resistant Gag polymorphisms.[9, 19, 23, 33,34,35,36,37,38,39,40] It is currently in Phase 2b clinical trials.[41,42,43] an understanding of the mechanism for how BMS-955176 achieves this improved antiviral coverage has not been described Such an understanding at the mechanistic level is of intrinsic interest, potentially providing further insights into the maturation process itself, and the biology and biochemistry of HIV-1 infection. Of clinical importance, such understanding may be of value to help guide the development of newer MIs with further improvements to MI activity, genotypic coverage and spectrum

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