HIV-1 maturation inhibitors (MIs) disrupt the final step in the HIV-1 protease-mediated cleavage of the Gag polyprotein, between capsid (CA) and spacer peptide 1 (SP1), leading to the production of non-infectious virus. BMS-955176 is a second generation MI with improved antiviral activity toward polymorphic Gag viruses. This study was conducted to understand the underlying mechanistic reasons for improvements to polymorphic coverage.
Antiviral activities of wild type (wt) and Gag polymorphic viruses were studied using single and multiple cycle replication formats. Gag CA/SP1 cleavage kinetics of wt and polymorphic virus-like particles (VLP) were determined by a novel LC/MS assay. A radiolabel binding assay was used to determine VLP/MI affinities and off rates. The cleavage, binding and antiviral data was used to create a integrated model for calculating rates of CA/SP1 cleavage.
In antiviral assays, early generation MIs (EGMIs) do not achieve maximal percent inhibition (100% MPI) of certain Gag polymorphs, even at saturating concentrations, resulting in low levels of breakthrough virus (partial antagonism). BMS-955176 achieves higher MPIs of polymorphs, which is correlated with lower EC50 values. Mechanistic studies show that more rapid CA/SP1 cleavage of polymorphs (2-10-fold vs. wt) is responsible for reduced viral sensitivity to EGMIs. Enzymatically, in vitro inhibition of wt CA/SP1 cleavage by the EGMI BVM is lost at longer times, while inhibition by BMS-955176 persists. The origin of antiviral and enzymatic improvements by BMS-955176 lies in its higher affinity to Gag polymorphs. Biochemical (CA/SP1 cleavage rates, MI-specific Gag affinities) and virological (MPI) data were combined to create an integrated kinetic model to calculate rate reductions of CA/SP1 cleavage by MIs. These predictions are in accord with both preclinical antiviral potency and clinical viral load reductions as a function of MI and Gag polymorph.
The improved polymorphic antiviral activity and biochemical profile of BMS-955176 over EGMIs is a consequence of its higher affinity/slower rate of dissociation from its Gag target. Modeling calculates MI-induced reductions in the rates of CA/SP1 Gag cleavage that correlate well with preclinical antiviral potencies and MI monotherapy clinical response data. These findings offer new insights into MI activity and mechanism and may prove useful to a further understanding of clinical responses to MIs.