Identification of a novel HIV-1 inhibitor targeting Vif-dependent degradation of human APOBEC3G protein

J Biol Chem. 2015 Apr 17;290(16):10504-17. doi: 10.1074/jbc.M114.626903. Epub 2015 Feb 27.

Abstract

APOBEC3G (A3G) is a cellular cytidine deaminase that restricts HIV-1 replication by inducing G-to-A hypermutation in viral DNA and by deamination-independent mechanisms. HIV-1 Vif binds to A3G, resulting in its degradation via the 26 S proteasome. Therefore, this interaction represents a potential therapeutic target. To identify compounds that inhibit interaction between A3G and HIV-1 Vif in a high throughput format, we developed a homogeneous time-resolved fluorescence resonance energy transfer assay. A 307,520 compound library from the NIH Molecular Libraries Small Molecule Repository was screened. Secondary screens to evaluate dose-response performance and off-target effects, cell-based assays to identify compounds that attenuate Vif-dependent degradation of A3G, and assays testing antiviral activity in peripheral blood mononuclear cells and T cells were employed. One compound, N.41, showed potent antiviral activity in A3G(+) but not in A3G(-) T cells and had an IC50 as low as 8.4 μM and a TC50 of >100 μM when tested against HIV-1Ba-L replication in peripheral blood mononuclear cells. N.41 inhibited the Vif-A3G interaction and increased cellular A3G levels and incorporation of A3G into virions, thereby attenuating virus infectivity in a Vif-dependent manner. N.41 activity was also species- and Vif-dependent. Preliminary structure-activity relationship studies suggest that a hydroxyl moiety located at a phenylamino group is critical for N.41 anti-HIV activity and identified N.41 analogs with better potency (IC50 as low as 4.2 μM). These findings identify a new lead compound that attenuates HIV replication by liberating A3G from Vif regulation and increasing its innate antiviral activity.

Keywords: Antiviral Agent; Cytidine Deaminase; High Throughput Screening (HTS); Human Immunodeficiency Virus (HIV); Viral Protein; Virology.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • APOBEC-3G Deaminase
  • Anti-HIV Agents / chemistry
  • Anti-HIV Agents / pharmacology*
  • Biological Assay
  • Cell Line
  • Cytidine Deaminase / genetics*
  • Cytidine Deaminase / metabolism
  • Fluorescence Resonance Energy Transfer
  • Gene Expression Regulation
  • HEK293 Cells
  • HIV-1 / drug effects*
  • HIV-1 / genetics
  • HIV-1 / metabolism
  • Host-Pathogen Interactions
  • Humans
  • Leukocytes, Mononuclear / drug effects*
  • Leukocytes, Mononuclear / metabolism
  • Leukocytes, Mononuclear / virology
  • Primary Cell Culture
  • Proteasome Endopeptidase Complex / drug effects
  • Proteasome Endopeptidase Complex / metabolism
  • Proteolysis
  • Signal Transduction
  • Small Molecule Libraries / chemistry
  • Small Molecule Libraries / pharmacology*
  • Structure-Activity Relationship
  • T-Lymphocytes / drug effects*
  • T-Lymphocytes / metabolism
  • T-Lymphocytes / virology
  • Virus Replication / drug effects
  • vif Gene Products, Human Immunodeficiency Virus / antagonists & inhibitors
  • vif Gene Products, Human Immunodeficiency Virus / genetics*
  • vif Gene Products, Human Immunodeficiency Virus / metabolism

Substances

  • Anti-HIV Agents
  • Small Molecule Libraries
  • vif Gene Products, Human Immunodeficiency Virus
  • vif protein, Human immunodeficiency virus 1
  • Proteasome Endopeptidase Complex
  • ATP dependent 26S protease
  • APOBEC-3G Deaminase
  • APOBEC3G protein, human
  • Cytidine Deaminase