The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.

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...

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Autores principales:	Daniela Begolo, Isabel M Vincent, Federica Giordani, Ina Pöhner, Michael J Witty, Timothy G Rowan, Zakaria Bengaly, Kirsten Gillingwater, Yvonne Freund, Rebecca C Wade, Michael P Barrett, Christine Clayton
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2018
Materias:	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5
Acceso en línea:	https://doaj.org/article/e1674f3ac3f24f8baaf483e779b5f349
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spelling	oai:doaj.org-article:e1674f3ac3f24f8baaf483e779b5f3492021-12-02T19:59:38ZThe trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.1553-73661553-737410.1371/journal.ppat.1007315https://doaj.org/article/e1674f3ac3f24f8baaf483e779b5f3492018-09-01T00:00:00Zhttps://doi.org/10.1371/journal.ppat.1007315https://doaj.org/toc/1553-7366https://doaj.org/toc/1553-7374Kinetoplastid 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.Daniela BegoloIsabel M VincentFederica GiordaniIna PöhnerMichael J WittyTimothy G RowanZakaria BengalyKirsten GillingwaterYvonne FreundRebecca C WadeMichael P BarrettChristine ClaytonPublic Library of Science (PLoS)articleImmunologic diseases. AllergyRC581-607Biology (General)QH301-705.5ENPLoS Pathogens, Vol 14, Iss 9, p e1007315 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5
spellingShingle	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5 Daniela Begolo Isabel M Vincent Federica Giordani Ina Pöhner Michael J Witty Timothy G Rowan Zakaria Bengaly Kirsten Gillingwater Yvonne Freund Rebecca C Wade Michael P Barrett Christine Clayton The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.
description	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.
format	article
author	Daniela Begolo Isabel M Vincent Federica Giordani Ina Pöhner Michael J Witty Timothy G Rowan Zakaria Bengaly Kirsten Gillingwater Yvonne Freund Rebecca C Wade Michael P Barrett Christine Clayton
author_facet	Daniela Begolo Isabel M Vincent Federica Giordani Ina Pöhner Michael J Witty Timothy G Rowan Zakaria Bengaly Kirsten Gillingwater Yvonne Freund Rebecca C Wade Michael P Barrett Christine Clayton
author_sort	Daniela Begolo
title	The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.
title_short	The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.
title_full	The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.
title_fullStr	The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.
title_full_unstemmed	The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.
title_sort	trypanocidal benzoxaborole an7973 inhibits trypanosome mrna processing.
publisher	Public Library of Science (PLoS)
publishDate	2018
url	https://doaj.org/article/e1674f3ac3f24f8baaf483e779b5f349
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The trypanocidal benzoxaborole AN7973 inhibits trypanosome mRNA processing.

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