The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing

Begolo, D. et al. (2018) The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing. PLoS Pathogens, 14(9), e1007315. (doi: 10.1371/journal.ppat.1007315) (PMID:30252911) (PMCID:PMC6173450)

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Kinetoplastid parasites—trypanosomes and leishmanias—infect millions of humans and cause economically devastating diseases of livestock, and the few existing drugs have serious deficiencies. Benzoxaborole-based compounds are very promising potential novel anti-trypanosomal therapies, with candidates already in human and animal clinical trials. We investigated the mechanism of action of several benzoxaboroles, including AN7973, an early candidate for veterinary trypanosomosis. In all kinetoplastids, transcription is polycistronic. Individual mRNA 5'-ends are created by trans splicing of a short leader sequence, with coupled polyadenylation of the preceding mRNA. Treatment of Trypanosoma brucei with AN7973 inhibited trans splicing within 1h, as judged by loss of the Y-structure splicing intermediate, reduced levels of mRNA, and accumulation of peri-nuclear granules. Methylation of the spliced leader precursor RNA was not affected, but more prolonged AN7973 treatment caused an increase in S-adenosyl methionine and methylated lysine. Together, the results indicate that mRNA processing is a primary target of AN7973. Polyadenylation is required for kinetoplastid trans splicing, and the EC50 for AN7973 in T. brucei was increased three-fold by over-expression of the T. brucei cleavage and polyadenylation factor CPSF3, identifying CPSF3 as a potential molecular target. Molecular modeling results suggested that inhibition of CPSF3 by AN7973 is feasible. Our results thus chemically validate mRNA processing as a viable drug target in trypanosomes. Several other benzoxaboroles showed metabolomic and splicing effects that were similar to those of AN7973, identifying splicing inhibition as a common mode of action and suggesting that it might be linked to subsequent changes in methylated metabolites. Granule formation, splicing inhibition and resistance after CPSF3 expression did not, however, always correlate and prolonged selection of trypanosomes in AN7973 resulted in only 1.5-fold resistance. It is therefore possible that the modes of action of oxaboroles that target trypanosome mRNA processing might extend beyond CPSF3 inhibition.

Item Type:Articles
Additional Information:Work by IMV, FG and MPB (Figs 5 & 6, S1 Figs S2 & S3 Fig, Tables S5, 6 & S7) was funded by a core grant to the Wellcome Centre for Molecular Parasitology (104111/Z/14/Z) and by the Medical Research Council (MR/K008749/1). Cattle (ZB) and goat challenge studies and in vitro testing (KG) of T. congolense and T. vivax (Fig 6, S1 Table) were organised and supported by the Global Alliance for Livestock Veterinary Medicines (GALVmed) (TGR, MJW) and funded by the Bill & Melinda Gates Foundation [OPP1093639] and UK aid from the UK Government. IP and RCW (Figs S9, S11, and S12, and Table S9) thank the Klaus Tschira Foundation for support. The work by DB (the rest of the paper) was supported by European Commission FP7 Marie Curie Initial Training Network "ParaMet" (grant number 172 290080) and by core funding from the state of Baden-Württemberg.
Keywords:Research article, biology and life sciences, physical sciences, medicine and health sciences.
Glasgow Author(s) Enlighten ID:Vincent, Dr Isabel and Giordani, Dr Federica and Barrett, Professor Michael
Creator Roles:
Vincent, I. M.Investigation, Writing – original draft, Writing – review and editing
Giordani, F.Investigation, Writing – original draft, Writing – review and editing
Barrett, M. P.Conceptualization, Formal analysis, Funding acquisition, Methodology, Project administration, Supervision, Visualization, Writing – original draft, Writing – review and editing
Authors: Begolo, D., Vincent, I. M., Giordani, F., Pöhner, I., Witty, M. J., Rowan, T. G., Bengaly, Z., Gillingwater, K., Freund, Y., Wade, R. C., Barrett, M. P., and Clayton, C.
College/School:College of Medical Veterinary and Life Sciences > School of Cancer Sciences
College of Medical Veterinary and Life Sciences > School of Infection & Immunity
Journal Name:PLoS Pathogens
Publisher:Public Library of Science
ISSN (Online):1553-7374
Copyright Holders:Copyright © 2018 Begolo et al.
First Published:First published in PLoS Pathogens 14(9):e1007315
Publisher Policy:Reproduced under a Creative Commons License

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
371799The Wellcome Centre for Molecular Parasitology ( Core Support )Andrew WatersWellcome Trust (WELLCOTR)104111/Z/14/Z & AIII - PARASITOLOGY
571301ParaMet - A systematic approach to understanding parasite metabolism.Sylke MullerEuropean Commission (EC)ParaMet290080-FP7III - PARASITOLOGY