Clinical and veterinary trypanocidal benzoxaboroles target CPSF3

Richard Wall, Eva Rico Vidal, Iva Lukac, Fabio Zuccotto, Sara Elg, Ian Gilbert, Yvonne R. Freund, Michael R. K. Alley, Mark Field, Susan Wyllie, David Horn (Lead / Corresponding author)

Research output: Contribution to journalArticle

17 Citations (Scopus)
265 Downloads (Pure)

Abstract

African trypanosomes cause lethal and neglected tropical diseases, known as sleeping sickness in humans and nagana in animals. Current therapies are limited, but fortunately, promising therapies are in advanced clinical and veterinary development, including acoziborole (AN5568 or SCYX-7158) and AN11736, respectively. These benzoxaboroles will likely be key to the World Health Organization's target of disease control by 2030. Their mode of action was previously unknown. We have developed a highcoverage overexpression library and use it here to explore drug mode of action in Trypanosoma brucei. Initially, an inhibitor with a known target was used to select for drug resistance and to test massive parallel library screening and genome-wide mapping; this effectively identified the known target and validated the approach. Subsequently, the overexpression screening approach was used to identify the target of the benzoxaboroles, Cleavage and Polyadenylation Specificity Factor 3 (CPSF3, Tb927.4.1340). We validated the CPSF3 endonuclease as the target, using independent overexpression strains. Knockdown provided genetic validation of CPSF3 as essential, and GFP tagging confirmed the expected nuclear localization. Molecular docking and CRISPRCas9-based editing demonstrated how acoziborole can specifically block the active site and mRNA processing by parasite, but not host CPSF3. Thus, our findings provide both genetic and chemical validation for CPSF3 as an important drug target in trypanosomes and reveal inhibition of mRNA maturation as the mode of action of the trypanocidal benzoxaboroles. Understanding the mechanism of action of benzoxaborole-based therapies can assist development of improved therapies, as well as the prediction and monitoring of resistance, if or when it arises.

Original languageEnglish
Pages (from-to)9616-9621
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume115
Issue number38
Early online date5 Sep 2018
DOIs
Publication statusPublished - 18 Sep 2018

Keywords

  • CPSF73
  • Drug discovery
  • Genetic screening
  • N-myristoyltransferase
  • Ysh1
  • Trypanosoma brucei brucei/drug effects
  • Trypanocidal Agents/pharmacology
  • Humans
  • Boron Compounds/pharmacology
  • Drug Resistance/drug effects
  • Gene Knockdown Techniques
  • Cleavage And Polyadenylation Specificity Factor/antagonists & inhibitors
  • Cell Nucleus/genetics
  • High-Throughput Screening Assays/methods
  • Valine/analogs & derivatives
  • Gene Library
  • Benzamides/pharmacology
  • RNA, Messenger/metabolism
  • RNA, Protozoan/metabolism
  • Animals
  • CRISPR-Cas Systems
  • RNA Processing, Post-Transcriptional/drug effects
  • Protozoan Proteins/antagonists & inhibitors
  • Molecular Docking Simulation
  • Trypanosomiasis, African/prevention & control

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