Resistance profile of the HIV-1 maturation inhibitor GSK3532795 in vitro and in a clinical study.

GSK3532795 (formerly BMS955176) is a second-generation maturation inhibitor (MI) that progressed through a Phase 2b study for treatment of HIV-1 infection. Resistance development to GSK3532795 was evaluated through in vitro methods and was correlated with information obtained in a Phase 2a proof-of-concept study in HIV-1 infected participants. Both low and high concentrations of GSK3532795 were used for selections in vitro, and reduced susceptibility to GSK3532795 mapped specifically to amino acids near the capsid/ spacer peptide 1 (SP1) junction, the cleavage of which is blocked by MIs. Two key substitutions, A364V or V362I, were selected, the latter requiring secondary substitutions to reduce susceptibility to GSK3532795. Three main types of secondary substitutions were observed, none of which reduced GSK3532795 susceptibility in isolation. The first type was in the capsid C-terminal domain and downstream SP1 region (including (Gag numbering) R286K, A326T, T332S/N, I333V and V370A/M). The second, was an R41G substitution in viral protease that occurred with V362I. The third was seen in the capsid N-terminal domain, within the cyclophilin A binding domain (V218A/M, H219Q and G221E). H219Q increased viral replication capacity and reduced susceptibility of poorly growing viruses. In the Phase 2a study, a subset of these substitutions was also observed at baseline and some were selected following GSK35323795 treatment in HIV-1-infected participants

Improving Physical Properties via CH Oxidation: Chemical and Enzymatic Approaches

Physicochemical properties constitute a key factor for the success of a drug candidate. Whereas many strategies to improve the physicochemical properties of small heterocycle-type leads exist, complex hydrocarbon skeletons are more challenging to derivatize because of the absence of functional groups. A variety of C-H oxidation methods have been explored on the betulin skeleton to improve the solubility of this very bioactive, yet poorly water-soluble, natural product. Capitalizing on the innate reactivity of the molecule, as well as the few molecular handles present on the core, allowed oxidations at different positions across the pentacyclic structure. Enzymatic oxidations afforded several orthogonal oxidations to chemical methods. Solubility measurements showed an enhancement for many of the synthesized compounds

Amination of Nitro-Substituted Heteroarenes by Nucleophilic Substitution of Hydrogen

An open-air method for the transition metal-free direct amination of nitro(hetero)arenes by anilines is disclosed. In this methodology, an aromatic C-H bond is substituted via oxidative nucleophilic aromatic substitution of hydrogen (ONSH). DFT calculations and mechanistic studies support a dianion pathway with oxidation by molecular oxygen as the rate-limiting step

Synthesis of Bicyclo[1.1.0]butanes from Iodo-Bicyclo[1.1.1]pentanes

We describe a two step process for the synthesis of substituted bicyclo[1.1.0]butanes. A photo-Hunsdiecker reaction generates iodo-bicyclo[1.1.1]pentanes under metal-free conditions at room temperature. These intermediates react with nitrogen and sulfur nucleophiles to afford substituted bicyclo[1.1.0]butane products

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

<div><p>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. <i>In vitro</i> 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 <i>in vitro</i> antiviral observations and correlate with observed <i>in vivo</i> 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.</p></div

Schematic for Cleavage of CA/SP1.

<p>A) Schematic for the processing HIV Gag at CA/SP1 and SP1/NC sites by HIV-protease. B) Detail of the cleavage region around CA/SP1 showing sites for HIV-1 protease cleavage (H1 and H2) and sites for subsequent cleavage by trypsin (T).</p

Modeling of the rate of CA/SP1 cleavage of HIV-1 WT, V370A, V362I and ΔV370 VLP in the presence of 300 nM MI.

<p>Modeled fractional rate of production of SP1 peptide from Gag VLP cleavage using model 2a at 300 nM MI, as noted in text; no MI: diamonds; BVM: squares; BMS-955176: triangles; y-axis: fraction of CA/SP1 cleavage is a surrogate for production of mature virus, as indicated in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005990#ppat.1005990.g005" target="_blank">Fig 5</a>; A) WT; B) V370A; C)V362I; D) ΔV370</p

Inhibition of HIV-1 protease mediated CA/SP1 cleavage by MIs.

<p>A) BVM, B) BMS-955176: Inhibition of CA/SP1 cleavage of WT, ΔV370 and A364V VLPs <i>in vitro</i> as monitored by LC/MS analysis (Materials and Methods); values are an average of 9 replicates; bars are SEM; MI concentrations: 3 μM.</p