Obatoclax Inhibits Alphavirus Membrane Fusion by Neutralizing the Acidic Environment of Endocytic Compartments

Antimicrob Agents Chemother. 2017 Feb 23;61(3):e02227-16. doi: 10.1128/AAC.02227-16. Print 2017 Mar.

Abstract

As new pathogenic viruses continue to emerge, it is paramount to have intervention strategies that target a common denominator in these pathogens. The fusion of viral and cellular membranes during viral entry is one such process that is used by many pathogenic viruses, including chikungunya virus, West Nile virus, and influenza virus. Obatoclax, a small-molecule antagonist of the Bcl-2 family of proteins, was previously determined to have activity against influenza A virus and also Sindbis virus. Here, we report it to be active against alphaviruses, like chikungunya virus (50% effective concentration [EC50] = 0.03 μM) and Semliki Forest virus (SFV; EC50 = 0.11 μM). Obatoclax inhibited viral entry processes in an SFV temperature-sensitive mutant entry assay. A neutral red retention assay revealed that obatoclax induces the rapid neutralization of the acidic environment of endolysosomal vesicles and thereby most likely inhibits viral fusion. Characterization of escape mutants revealed that the L369I mutation in the SFV E1 fusion protein was sufficient to confer partial resistance against obatoclax. Other inhibitors that target the Bcl-2 family of antiapoptotic proteins inhibited neither viral entry nor endolysosomal acidification, suggesting that the antiviral mechanism of obatoclax does not depend on its anticancer targets. Obatoclax inhibited the growth of flaviviruses, like Zika virus, West Nile virus, and yellow fever virus, which require low pH for fusion, but not that of pH-independent picornaviruses, like coxsackievirus A9, echovirus 6, and echovirus 7. In conclusion, obatoclax is a novel inhibitor of endosomal acidification that prevents viral fusion and that could be pursued as a potential broad-spectrum antiviral candidate.

Keywords: Zika virus; antiviral; chikungunya virus; envelope protein; resistant mutant; virus entry.

MeSH terms

  • Animals
  • Antiviral Agents / pharmacology*
  • Cell Line
  • Cell Membrane / drug effects
  • Cell Membrane / virology
  • Chikungunya virus / drug effects*
  • Chikungunya virus / genetics
  • Chikungunya virus / growth & development
  • Cricetinae
  • Drug Resistance, Viral / genetics
  • Endosomes / drug effects*
  • Endosomes / metabolism
  • Epithelial Cells / drug effects
  • Epithelial Cells / virology
  • Gene Expression
  • Hepatocytes / drug effects
  • Hepatocytes / virology
  • Humans
  • Hydrogen-Ion Concentration / drug effects
  • Indoles
  • Lysosomes / drug effects*
  • Lysosomes / metabolism
  • Membrane Fusion / drug effects*
  • Membrane Glycoproteins / genetics
  • Membrane Glycoproteins / metabolism
  • Mutation
  • Neutral Red / metabolism
  • Proto-Oncogene Proteins c-bcl-2 / genetics
  • Proto-Oncogene Proteins c-bcl-2 / metabolism
  • Pyrroles / pharmacology*
  • Semliki forest virus / drug effects*
  • Semliki forest virus / genetics
  • Semliki forest virus / growth & development
  • Viral Envelope Proteins / genetics
  • Viral Envelope Proteins / metabolism
  • Virus Internalization / drug effects
  • Virus Replication / drug effects
  • West Nile virus / drug effects
  • West Nile virus / genetics
  • West Nile virus / growth & development
  • Yellow fever virus / drug effects
  • Yellow fever virus / genetics
  • Yellow fever virus / growth & development
  • Zika Virus / drug effects
  • Zika Virus / genetics
  • Zika Virus / growth & development

Substances

  • Antiviral Agents
  • BCL2 protein, human
  • E1 glycoprotein, Semliki forest virus
  • Indoles
  • Membrane Glycoproteins
  • Proto-Oncogene Proteins c-bcl-2
  • Pyrroles
  • Viral Envelope Proteins
  • Neutral Red
  • obatoclax