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Discovery of E2730, a novel selective uncompetitive GAT1 inhibitor, as a candidate for anti‐seizure medication
OBJECTIVE: As of 2022, 36 anti‐seizure medications (ASMs) have been licensed for the treatment of epilepsy, however, adverse effects (AEs) are commonly reported. Therefore, ASMs with a wide margin between therapeutic effects and AEs are preferred over ASMs that are associated with a narrow margin be...
Autores principales: | , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley and Sons Inc.
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10472371/ https://www.ncbi.nlm.nih.gov/pubmed/37052238 http://dx.doi.org/10.1002/epi4.12741 |
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author | Fukushima, Kazuyuki Higashiyama, Hiroyuki Kazuta, Yuji Hashimoto, Keisuke Watanabe, Naoto Furuya, Yoshiaki Ito, Yoshimasa Wu, Ting Kosasa, Takashi Talos, Delia M. Song, Yeri Roberts, Nicholas S. Jensen, Frances E. Hanada, Takahisa Ido, Katsutoshi |
author_facet | Fukushima, Kazuyuki Higashiyama, Hiroyuki Kazuta, Yuji Hashimoto, Keisuke Watanabe, Naoto Furuya, Yoshiaki Ito, Yoshimasa Wu, Ting Kosasa, Takashi Talos, Delia M. Song, Yeri Roberts, Nicholas S. Jensen, Frances E. Hanada, Takahisa Ido, Katsutoshi |
author_sort | Fukushima, Kazuyuki |
collection | PubMed |
description | OBJECTIVE: As of 2022, 36 anti‐seizure medications (ASMs) have been licensed for the treatment of epilepsy, however, adverse effects (AEs) are commonly reported. Therefore, ASMs with a wide margin between therapeutic effects and AEs are preferred over ASMs that are associated with a narrow margin between efficacy and risk of AEs. E2730 was discovered using in vivo phenotypic screening and characterized as an uncompetitive, yet selective, inhibitor of γ‐aminobutyric acid (GABA) transporter 1 (GAT1). Here, we describe the preclinical characteristics of E2730. METHODS: Anti‐seizure effects of E2730 were evaluated in several animal models of epilepsy: corneal kindling, 6 Hz–44 mA psychomotor seizure, amygdala kindling, Fragile X syndrome, and Dravet syndrome models. Effects of E2730 on motor coordination were assessed in accelerating rotarod tests. The mechanism of action of E2730 was explored by [(3)H]E2730 binding assay. The GAT1‐selectivity over other GABA transporters was examined by GABA uptake assay of GAT1, GAT2, GAT3, or betaine/GABA transporter 1 (BGT‐1) stably expressing HEK293 cells. To further investigate the mechanism for E2730‐mediated inhibition of GAT1, in vivo microdialysis and in vitro GABA uptake assays were conducted under conditions of different GABA concentrations. RESULTS: E2730 showed anti‐seizure effects in the assessed animal models with an approximately >20‐fold margin between efficacy and motor incoordination. [(3)H]E2730 binding on brain synaptosomal membrane was abolished in GAT1‐deficient mice, and E2730 selectively inhibited GAT1‐mediated GABA uptake over other GABA transporters. In addition, results of GABA uptake assays showed that E2730‐mediated inhibition of GAT1 positively correlated to the level of ambient GABA in vitro. E2730 also increased extracellular GABA concentration in hyperactivated conditions but not under basal levels in vivo. SIGNIFICANCE: E2730 is a novel, selective, uncompetitive GAT1 inhibitor, which acts selectively under the condition of increasing synaptic activity, contributing to a wide margin between therapeutic effect and motor incoordination. |
format | Online Article Text |
id | pubmed-10472371 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | John Wiley and Sons Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-104723712023-09-02 Discovery of E2730, a novel selective uncompetitive GAT1 inhibitor, as a candidate for anti‐seizure medication Fukushima, Kazuyuki Higashiyama, Hiroyuki Kazuta, Yuji Hashimoto, Keisuke Watanabe, Naoto Furuya, Yoshiaki Ito, Yoshimasa Wu, Ting Kosasa, Takashi Talos, Delia M. Song, Yeri Roberts, Nicholas S. Jensen, Frances E. Hanada, Takahisa Ido, Katsutoshi Epilepsia Open Original Articles OBJECTIVE: As of 2022, 36 anti‐seizure medications (ASMs) have been licensed for the treatment of epilepsy, however, adverse effects (AEs) are commonly reported. Therefore, ASMs with a wide margin between therapeutic effects and AEs are preferred over ASMs that are associated with a narrow margin between efficacy and risk of AEs. E2730 was discovered using in vivo phenotypic screening and characterized as an uncompetitive, yet selective, inhibitor of γ‐aminobutyric acid (GABA) transporter 1 (GAT1). Here, we describe the preclinical characteristics of E2730. METHODS: Anti‐seizure effects of E2730 were evaluated in several animal models of epilepsy: corneal kindling, 6 Hz–44 mA psychomotor seizure, amygdala kindling, Fragile X syndrome, and Dravet syndrome models. Effects of E2730 on motor coordination were assessed in accelerating rotarod tests. The mechanism of action of E2730 was explored by [(3)H]E2730 binding assay. The GAT1‐selectivity over other GABA transporters was examined by GABA uptake assay of GAT1, GAT2, GAT3, or betaine/GABA transporter 1 (BGT‐1) stably expressing HEK293 cells. To further investigate the mechanism for E2730‐mediated inhibition of GAT1, in vivo microdialysis and in vitro GABA uptake assays were conducted under conditions of different GABA concentrations. RESULTS: E2730 showed anti‐seizure effects in the assessed animal models with an approximately >20‐fold margin between efficacy and motor incoordination. [(3)H]E2730 binding on brain synaptosomal membrane was abolished in GAT1‐deficient mice, and E2730 selectively inhibited GAT1‐mediated GABA uptake over other GABA transporters. In addition, results of GABA uptake assays showed that E2730‐mediated inhibition of GAT1 positively correlated to the level of ambient GABA in vitro. E2730 also increased extracellular GABA concentration in hyperactivated conditions but not under basal levels in vivo. SIGNIFICANCE: E2730 is a novel, selective, uncompetitive GAT1 inhibitor, which acts selectively under the condition of increasing synaptic activity, contributing to a wide margin between therapeutic effect and motor incoordination. John Wiley and Sons Inc. 2023-05-18 /pmc/articles/PMC10472371/ /pubmed/37052238 http://dx.doi.org/10.1002/epi4.12741 Text en © 2023 The Authors. Epilepsia Open published by Wiley Periodicals LLC on behalf of International League Against Epilepsy. https://creativecommons.org/licenses/by/4.0/This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Original Articles Fukushima, Kazuyuki Higashiyama, Hiroyuki Kazuta, Yuji Hashimoto, Keisuke Watanabe, Naoto Furuya, Yoshiaki Ito, Yoshimasa Wu, Ting Kosasa, Takashi Talos, Delia M. Song, Yeri Roberts, Nicholas S. Jensen, Frances E. Hanada, Takahisa Ido, Katsutoshi Discovery of E2730, a novel selective uncompetitive GAT1 inhibitor, as a candidate for anti‐seizure medication |
title | Discovery of E2730, a novel selective uncompetitive GAT1 inhibitor, as a candidate for anti‐seizure medication |
title_full | Discovery of E2730, a novel selective uncompetitive GAT1 inhibitor, as a candidate for anti‐seizure medication |
title_fullStr | Discovery of E2730, a novel selective uncompetitive GAT1 inhibitor, as a candidate for anti‐seizure medication |
title_full_unstemmed | Discovery of E2730, a novel selective uncompetitive GAT1 inhibitor, as a candidate for anti‐seizure medication |
title_short | Discovery of E2730, a novel selective uncompetitive GAT1 inhibitor, as a candidate for anti‐seizure medication |
title_sort | discovery of e2730, a novel selective uncompetitive gat1 inhibitor, as a candidate for anti‐seizure medication |
topic | Original Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10472371/ https://www.ncbi.nlm.nih.gov/pubmed/37052238 http://dx.doi.org/10.1002/epi4.12741 |
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