Assay strategies for the discovery and validation of therapeutics targeting Brugia pahangi Hsp90

Taldone, T., Gillan, V., Sun, W., Rodina, A., Patel, P., Maitland, K., O'Neill, K., Chiosis, G. and Devaney, E. (2010) Assay strategies for the discovery and validation of therapeutics targeting Brugia pahangi Hsp90. PLoS Neglected Tropical Diseases, 4(6), e714. (doi:10.1371/journal.pntd.0000714)

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Abstract

The chemotherapy of lymphatic filariasis relies upon drugs such as diethylcarbamazine and ivermectin that largely target the microfilarial stages of the parasite, necessitating continued treatment over the long reproductive life span of the adult worm. The identification of compounds that target adult worms has been a long-term goal of WHO. Here we describe a fluorescence polarization assay for the identification of compounds that target Hsp90 in adult filarial worms. The assay was originally developed to identify inhibitors of Hsp90 in tumor cells, and relies upon the ability of small molecules to inhibit the binding of fluorescently labelled geldanamycin to Hsp90. We demonstrate that the assay works well with soluble extracts of Brugia, while extracts of the free-living nematode C. elegans fail to bind the probe, in agreement with data from other experiments. The assay was validated using known inhibitors of Hsp90 that compete with geldanamycin for binding to Hsp90, including members of the synthetic purine-scaffold series of compounds. The efficacy of some of these compounds against adult worms was confirmed in vitro. Moreover, the assay is sufficiently sensitive to differentiate between binding of purine-scaffold compounds to human and Brugia Hsp90. The assay is suitable for high-throughput screening and provides the first example of a format with the potential to identify novel inhibitors of Hsp90 in filarial worms and in other parasitic species where Hsp90 may be a target.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Devaney, Professor Eileen and O'Neill, Ms Kerry and Maitland, Ms Kirsty and Gillan, Dr Victoria
Authors: Taldone, T., Gillan, V., Sun, W., Rodina, A., Patel, P., Maitland, K., O'Neill, K., Chiosis, G., and Devaney, E.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
Journal Name:PLoS Neglected Tropical Diseases
Publisher:Public Library of Science
ISSN:1935-2727
ISSN (Online):1935-2735
Published Online:15 June 2010
Copyright Holders:Copyright © 2010 The Authors
First Published:First published in PLoS Neglected Tropical Diseases 4(6):e714
Publisher Policy:Reproduced in accordance with the copyright policy of the publisher

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
441591A comparative analysis of Hsp90 its possible role in drug resistance in nematode parasitesEileen DevaneyBiotechnology and Biological Sciences Research Council (BBSRC)BB/E013473/1Infection Immunity and Inflammation Life Sciences