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Synthesis, Screening and Characterization of Novel Potent Arp2/3 Inhibitory Compounds Analogous to CK-666

Branched actin networks polymerized by the Actin-related protein 2 and 3 (Arp2/3) complex play key roles in force generation and membrane remodeling. These networks are particularly important for cell migration, where they drive membrane protrusions of lamellipodia. Several Arp2/3 inhibitory compoun...

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Autores principales: Fokin, Artem I., Chuprov-Netochin, Roman N., Malyshev, Alexander S., Romero, Stéphane, Semenova, Marina N., Konyushkin, Leonid D., Leonov, Sergey V., Semenov, Victor V., Gautreau, Alexis M.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9189929/
https://www.ncbi.nlm.nih.gov/pubmed/35707404
http://dx.doi.org/10.3389/fphar.2022.896994
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author Fokin, Artem I.
Chuprov-Netochin, Roman N.
Malyshev, Alexander S.
Romero, Stéphane
Semenova, Marina N.
Konyushkin, Leonid D.
Leonov, Sergey V.
Semenov, Victor V.
Gautreau, Alexis M.
author_facet Fokin, Artem I.
Chuprov-Netochin, Roman N.
Malyshev, Alexander S.
Romero, Stéphane
Semenova, Marina N.
Konyushkin, Leonid D.
Leonov, Sergey V.
Semenov, Victor V.
Gautreau, Alexis M.
author_sort Fokin, Artem I.
collection PubMed
description Branched actin networks polymerized by the Actin-related protein 2 and 3 (Arp2/3) complex play key roles in force generation and membrane remodeling. These networks are particularly important for cell migration, where they drive membrane protrusions of lamellipodia. Several Arp2/3 inhibitory compounds have been identified. Among them, the most widely used is CK-666 (2-Fluoro-N-[2-(2-methyl-1H-indol-3-yl)ethyl]-benzamide), whose mode of action is to prevent Arp2/3 from reaching its active conformation. Here 74 compounds structurally related to CK-666 were screened using a variety of assays. The primary screen involved EdU (5-ethynyl-2′-deoxyuridine) incorporation in untransformed MCF10A cells. The resulting nine positive hits were all blocking lamellipodial protrusions and cell migration in B16-F1 melanoma cells in secondary screens, showing that cell cycle progression can be a useful read-out of Arp2/3 activity. Selected compounds were also characterized on sea urchin embryos, where Arp2/3 inhibition yields specific phenotypes such as the lack of triradiate spicules and inhibition of archenteron elongation. Several compounds were filtered out due to their toxicity in cell cultures or on sea urchin development. Two CK-666 analogs, 59 (N-{2-[5-(Benzyloxy)-2-methyl-1H-indol-3-yl] ethyl}-3-bromobenzamide) and 69 (2,4-Dichloro-N-[2-(7-chloro-2-methyl-1H-indol-3-yl) ethyl]-5-[(dimethylamino) sulfonyl] benzamide), were active in all assays and significantly more efficient in vivo than CK-666. These best hits with increased in vivo potency were, however, slightly less efficient in vitro than CK-666 in the classical pyrene-actin assay. Induced-fit docking of selected compounds and their possible metabolites revealed interaction with Arp2/3 that suppresses Arp2/3 activation. The data obtained in our screening validated the applicability of original assays for Arp2/3 activity. Several previously unexplored CK-666 structural analogs were found to suppress Arp2/3 activation, and two of them were identified as Arp2/3 inhibitors with improved in vivo efficiency.
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spelling pubmed-91899292022-06-14 Synthesis, Screening and Characterization of Novel Potent Arp2/3 Inhibitory Compounds Analogous to CK-666 Fokin, Artem I. Chuprov-Netochin, Roman N. Malyshev, Alexander S. Romero, Stéphane Semenova, Marina N. Konyushkin, Leonid D. Leonov, Sergey V. Semenov, Victor V. Gautreau, Alexis M. Front Pharmacol Pharmacology Branched actin networks polymerized by the Actin-related protein 2 and 3 (Arp2/3) complex play key roles in force generation and membrane remodeling. These networks are particularly important for cell migration, where they drive membrane protrusions of lamellipodia. Several Arp2/3 inhibitory compounds have been identified. Among them, the most widely used is CK-666 (2-Fluoro-N-[2-(2-methyl-1H-indol-3-yl)ethyl]-benzamide), whose mode of action is to prevent Arp2/3 from reaching its active conformation. Here 74 compounds structurally related to CK-666 were screened using a variety of assays. The primary screen involved EdU (5-ethynyl-2′-deoxyuridine) incorporation in untransformed MCF10A cells. The resulting nine positive hits were all blocking lamellipodial protrusions and cell migration in B16-F1 melanoma cells in secondary screens, showing that cell cycle progression can be a useful read-out of Arp2/3 activity. Selected compounds were also characterized on sea urchin embryos, where Arp2/3 inhibition yields specific phenotypes such as the lack of triradiate spicules and inhibition of archenteron elongation. Several compounds were filtered out due to their toxicity in cell cultures or on sea urchin development. Two CK-666 analogs, 59 (N-{2-[5-(Benzyloxy)-2-methyl-1H-indol-3-yl] ethyl}-3-bromobenzamide) and 69 (2,4-Dichloro-N-[2-(7-chloro-2-methyl-1H-indol-3-yl) ethyl]-5-[(dimethylamino) sulfonyl] benzamide), were active in all assays and significantly more efficient in vivo than CK-666. These best hits with increased in vivo potency were, however, slightly less efficient in vitro than CK-666 in the classical pyrene-actin assay. Induced-fit docking of selected compounds and their possible metabolites revealed interaction with Arp2/3 that suppresses Arp2/3 activation. The data obtained in our screening validated the applicability of original assays for Arp2/3 activity. Several previously unexplored CK-666 structural analogs were found to suppress Arp2/3 activation, and two of them were identified as Arp2/3 inhibitors with improved in vivo efficiency. Frontiers Media S.A. 2022-05-30 /pmc/articles/PMC9189929/ /pubmed/35707404 http://dx.doi.org/10.3389/fphar.2022.896994 Text en Copyright © 2022 Fokin, Chuprov-Netochin, Malyshev, Romero, Semenova, Konyushkin, Leonov, Semenov and Gautreau. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Pharmacology
Fokin, Artem I.
Chuprov-Netochin, Roman N.
Malyshev, Alexander S.
Romero, Stéphane
Semenova, Marina N.
Konyushkin, Leonid D.
Leonov, Sergey V.
Semenov, Victor V.
Gautreau, Alexis M.
Synthesis, Screening and Characterization of Novel Potent Arp2/3 Inhibitory Compounds Analogous to CK-666
title Synthesis, Screening and Characterization of Novel Potent Arp2/3 Inhibitory Compounds Analogous to CK-666
title_full Synthesis, Screening and Characterization of Novel Potent Arp2/3 Inhibitory Compounds Analogous to CK-666
title_fullStr Synthesis, Screening and Characterization of Novel Potent Arp2/3 Inhibitory Compounds Analogous to CK-666
title_full_unstemmed Synthesis, Screening and Characterization of Novel Potent Arp2/3 Inhibitory Compounds Analogous to CK-666
title_short Synthesis, Screening and Characterization of Novel Potent Arp2/3 Inhibitory Compounds Analogous to CK-666
title_sort synthesis, screening and characterization of novel potent arp2/3 inhibitory compounds analogous to ck-666
topic Pharmacology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9189929/
https://www.ncbi.nlm.nih.gov/pubmed/35707404
http://dx.doi.org/10.3389/fphar.2022.896994
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