TY - JOUR
T1 - A multiorganism pipeline for antiseizure drug discovery
T2 - Identification of chlorothymol as a novel γ-aminobutyric acidergic anticonvulsant
AU - Jones, Alistair
AU - Barker-Haliski, Melissa
AU - Ilie, Andrei S.
AU - Herd, Murray B.
AU - Baxendale, Sarah
AU - Holdsworth, Celia J.
AU - Ashton, John Paul
AU - Placzek, Marysia
AU - Jayasekera, Bodiabaduge A. P.
AU - Cowie, Christopher J. A.
AU - Lambert, Jeremy J.
AU - Trevelyan, Andrew J.
AU - White, H. Steve
AU - Marson, Anthony G.
AU - Cunliffe, Vincent T.
AU - Sills, Graeme J.
AU - Morgan, Alan
N1 - Funding Information:
This work was supported by a Medical Research Council (MRC) DiMeN PhD studentship for A.J. awarded to A.M./G.J.S./V.T.C./A.G.M.; by an MRC pump-priming translational research award G0802527 to M.P.; and by a grant from Tenovus Tayside to J.J.L. A.S.I. was supported by National Institute for Health Research UK via an Academic Clinical Fellowship in Neurology. M.B.-H. was supported by the Institution of Translational Health Sciences (KL2 TR002317). We thank Professor David Sullivan of the Johns Hopkins School of Medicine (Baltimore, MD) for supplying the Johns Hopkins Clinical Compound Collection Library. Worm strains used in this work were provided by the Caenorhabditis Genetics Center, which is funded by the National Institutes of Health National Center for Research Resources; and by the National Bioresource Project for the Experimental Animal ?Nematode C. elegans? from the laboratory of Dr Shohei Mitani (Tokyo Women's Medical University, Tokyo, Japan). We thank Jeff Barclay (University of Liverpool) for advice on C elegans work, and Jack Paveley (University of Sheffield) for help with data analysis.
Publisher Copyright:
© 2020 The Authors. Epilepsia published by Wiley Periodicals LLC on behalf of International League Against Epilepsy
Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.
PY - 2020/10
Y1 - 2020/10
N2 - Objective: Current medicines are ineffective in approximately one-third of people with epilepsy. Therefore, new antiseizure drugs are urgently needed to address this problem of pharmacoresistance. However, traditional rodent seizure and epilepsy models are poorly suited to high-throughput compound screening. Furthermore, testing in a single species increases the chance that therapeutic compounds act on molecular targets that may not be conserved in humans. To address these issues, we developed a pipeline approach using four different organisms.Methods: We sequentially employed compound library screening in the zebrafish, Danio rerio, chemical genetics in the worm, Caenorhabditis elegans, electrophysiological analysis in mouse and human brain slices, and preclinical validation in mouse seizure models to identify novel antiseizure drugs and their molecular mechanism of action.Results: Initially, a library of 1690 compounds was screened in an acute pentylenetetrazol seizure model using D rerio. From this screen, the compound chlorothymol was identified as an effective anticonvulsant not only in fish, but also in worms. A subsequent genetic screen in C elegans revealed the molecular target of chlorothymol to be LGC-37, a worm γ-aminobutyric acid type A (GABAA) receptor subunit. This GABAergic effect was confirmed using in vitro brain slice preparations from both mice and humans, as chlorothymol was shown to enhance tonic and phasic inhibition and this action was reversed by the GABAA receptor antagonist, bicuculline. Finally, chlorothymol exhibited in vivo anticonvulsant efficacy in several mouse seizure assays, including the 6-Hz 44-mA model of pharmacoresistant seizures.Significance: These findings establish a multiorganism approach that can identify compounds with evolutionarily conserved molecular targets and translational potential, and so may be useful in drug discovery for epilepsy and possibly other conditions.
AB - Objective: Current medicines are ineffective in approximately one-third of people with epilepsy. Therefore, new antiseizure drugs are urgently needed to address this problem of pharmacoresistance. However, traditional rodent seizure and epilepsy models are poorly suited to high-throughput compound screening. Furthermore, testing in a single species increases the chance that therapeutic compounds act on molecular targets that may not be conserved in humans. To address these issues, we developed a pipeline approach using four different organisms.Methods: We sequentially employed compound library screening in the zebrafish, Danio rerio, chemical genetics in the worm, Caenorhabditis elegans, electrophysiological analysis in mouse and human brain slices, and preclinical validation in mouse seizure models to identify novel antiseizure drugs and their molecular mechanism of action.Results: Initially, a library of 1690 compounds was screened in an acute pentylenetetrazol seizure model using D rerio. From this screen, the compound chlorothymol was identified as an effective anticonvulsant not only in fish, but also in worms. A subsequent genetic screen in C elegans revealed the molecular target of chlorothymol to be LGC-37, a worm γ-aminobutyric acid type A (GABAA) receptor subunit. This GABAergic effect was confirmed using in vitro brain slice preparations from both mice and humans, as chlorothymol was shown to enhance tonic and phasic inhibition and this action was reversed by the GABAA receptor antagonist, bicuculline. Finally, chlorothymol exhibited in vivo anticonvulsant efficacy in several mouse seizure assays, including the 6-Hz 44-mA model of pharmacoresistant seizures.Significance: These findings establish a multiorganism approach that can identify compounds with evolutionarily conserved molecular targets and translational potential, and so may be useful in drug discovery for epilepsy and possibly other conditions.
KW - drug discovery
KW - epilepsy
KW - GABA
KW - nematode
KW - zebrafish
UR - http://www.scopus.com/inward/record.url?scp=85089366916&partnerID=8YFLogxK
U2 - 10.1111/epi.16644
DO - 10.1111/epi.16644
M3 - Article
C2 - 32797628
AN - SCOPUS:85089366916
SN - 0013-9580
VL - 61
SP - 2106
EP - 2118
JO - Epilepsia
JF - Epilepsia
IS - 10
ER -