Identification of a Small‐Molecule Inhibitor of HIV‐1 Assembly that Targets the Phosphatidylinositol (4,5)‐bisphosphate Binding Site of the HIV‐1 Matrix Protein

Abstract

The development of drug resistance remains a critical problem for current HIV-1 antiviral therapies, creating a need for new inhibitors of HIV-1 replication. We previously reported on a novel anti-HIV-1 compound, N(2)-(phenoxyacetyl)-N-[4-(1-piperidinylcarbonyl)benzyl]glycinamide (14), that binds to the highly conserved phosphatidylinositol (4,5)-bisphosphate (PI(4,5)P(2)) binding pocket of the HIV-1 matrix (MA) protein. In this study, we re-evaluate the hits from the virtual screen used to identify compound 14 and test them directly in an HIV-1 replication assay using primary human peripheral blood mononuclear cells. This study resulted in the identification of three new compounds with antiviral activity; 2-(4-{[3-(4-fluorophenyl)-1,2,4-oxadiazol-5-yl]methyl})-1-piperazinyl)-N-(4-methylphenyl)acetamide (7), 3-(2-ethoxyphenyl)-5-[[4-(4-nitrophenyl)piperazin-1-yl]methyl]-1,2,4-oxadiazole (17), and N-[4-ethoxy-3-(1-piperidinylsulfonyl)phenyl]-2-(imidazo[2,1-b][1,3]thiazol-6-yl)acetamide (18), with compound 7 being the most potent of these hits. Mechanistic studies on 7 demonstrated that it directly interacts with and functions through HIV-1 MA. In accordance with our drug target, compound 7 competes with PI(4,5)P(2) for MA binding and, as a result, diminishes the production of new virus. Mutation of residues within the PI(4,5)P(2) binding site of MA decreased the antiviral effect of compound 7. Additionally, compound 7 displays a broadly neutralizing anti-HIV activity, with IC(50) values of 7.5-15.6 μM for the group M isolates tested. Taken together, these results point towards a novel chemical probe that can be used to more closely study the biological role of MA and could, through further optimization, lead to a new class of anti-HIV-1 therapeutics.

Cellular aspartyl proteases promote the unconventional secretion of biologically active HIV-1 matrix protein p17

“…At present, the p17 protein is the target of a therapeutic vaccine aimed at boosting and better directing the immune response against the functional epitope involved in the p17/p17 receptors interactions59. At the same time, inhibitors of the p17/PI(4,5)P 2 interaction60 as well as inhibitors of γ-secretase61 are major targets for drug development. Identification of the fine mechanisms underlying p17 secretion will help to develop further therapeutic interventions for eliminating the presence of the viral toxin in the extracellular microenvironment.…”

An Optimized, Synthetic DNA Vaccine Encoding the Toxin A and Toxin B Receptor Binding Domains of Clostridium difficile Induces Protective Antibody ResponsesIn Vivo

“…The RBD regions of TcdA and TcdB were amplified from either pARBD-NQ or pBRBD-NQ using primers designed to facilitate subcloning into the ligation-independent cloning prokaryotic expression vector pETHSUL as described previously (24). TcdA and TcdB RBD proteins were overproduced in Escherichia coli and purified using the subtractive purification strategy outlined by Zentner et al (25).…”

Phospholipases: at the crossroads of the immune system and the pathogenesis of HIV-1 infection

“…Indeed, 2 different approaches that disrupt HIV-1 replication by interfering with PIP 2 -mediated viral release were described. In particular, an anti-PI-specific Ab inhibits HIV-1 infection of primary macrophages [178], whereas a small-molecule inhibitor of HIV-1 assembly, which competes with PIP 2 for binding to MA, diminishes the production of new virus exhibiting broadspectrum, anti-HIV-1 activity [179]. In vitro studies.…”