A Chemical Chaperone Restores Connexin 26 Mutant Activity

Publikation: Beitrag in FachzeitschriftArtikelForschungPeer-Review

Autoren

  • Dahua Wang
  • Hongling Wang
  • Lu Fan
  • Tobias Ludwig
  • Andre Wegner
  • Frank Stahl
  • Jennifer Harre
  • Athanasia Warnecke
  • Carsten Zeilinger

Externe Organisationen

  • Medizinische Hochschule Hannover (MHH)
  • Technische Universität Braunschweig
Forschungs-netzwerk anzeigen

Details

OriginalspracheEnglisch
Seiten (von - bis)997–1005
Seitenumfang9
FachzeitschriftACS Pharmacology & Translational Science
Jahrgang6
Ausgabenummer7
Frühes Online-Datum1 Juni 2023
PublikationsstatusVeröffentlicht - 14 Juli 2023

Abstract

Mutations in connexin 26 (Cx26) cause hearing disorders of a varying degree. Herein, to identify compounds capable of restoring the function of mutated Cx26, a novel miniaturized microarray-based screening system was developed to perform an optical assay of Cx26 functionality. These molecules were identified through a viability assay using HeLa cells expressing wild-type (WT) Cx26, which exhibited sensitivity toward the HSP90 inhibitor radicicol in the submicromolar concentration range. Open Cx26 hemichannels are assumed to mediate the passage of molecules up to 1000 Da in size. Thus, by releasing radicicol, WT Cx26 active hemichannels in HeLa cells contribute to a higher survival rate and lower cell viability when Cx26 is mutated. HeLa cells expressing Cx26 mutations exhibited reduced viability in the presence of radicicol, such as the mutants F161S or R184P. Next, molecules exhibiting chemical chaperoning activity, suspected of restoring channel function, were assessed regarding whether they induced superior sensitivity toward radicicol and increased HeLa cell viability. Through a viability assay and microarray-based flux assay that uses Lucifer yellow in HeLa cells, compounds 3 and 8 were identified to restore mutant functionality. Furthermore, thermophoresis experiments revealed that only 3 (VRT-534) exhibited dose-responsive binding to recombinant WT Cx26 and mutant Cx26K188N with half maximal effective concentration values of 19 and ~5 µM, respectively. The findings of this study reveal that repurposing compounds already being used to treat other diseases, such as cystic fibrosis, in combination with functional bioassays and binding tests can help identify novel potential candidates that can be used to treat hearing disorders.

ASJC Scopus Sachgebiete

Zitieren

A Chemical Chaperone Restores Connexin 26 Mutant Activity. / Wang, Dahua; Wang, Hongling; Fan, Lu et al.
in: ACS Pharmacology & Translational Science, Jahrgang 6, Nr. 7, 14.07.2023, S. 997–1005.

Publikation: Beitrag in FachzeitschriftArtikelForschungPeer-Review

Wang, D, Wang, H, Fan, L, Ludwig, T, Wegner, A, Stahl, F, Harre, J, Warnecke, A & Zeilinger, C 2023, 'A Chemical Chaperone Restores Connexin 26 Mutant Activity', ACS Pharmacology & Translational Science, Jg. 6, Nr. 7, S. 997–1005. https://doi.org/10.1021/acsptsci.3c00056
Wang, D., Wang, H., Fan, L., Ludwig, T., Wegner, A., Stahl, F., Harre, J., Warnecke, A., & Zeilinger, C. (2023). A Chemical Chaperone Restores Connexin 26 Mutant Activity. ACS Pharmacology & Translational Science, 6(7), 997–1005. https://doi.org/10.1021/acsptsci.3c00056
Wang D, Wang H, Fan L, Ludwig T, Wegner A, Stahl F et al. A Chemical Chaperone Restores Connexin 26 Mutant Activity. ACS Pharmacology & Translational Science. 2023 Jul 14;6(7):997–1005. Epub 2023 Jun 1. doi: 10.1021/acsptsci.3c00056
Wang, Dahua ; Wang, Hongling ; Fan, Lu et al. / A Chemical Chaperone Restores Connexin 26 Mutant Activity. in: ACS Pharmacology & Translational Science. 2023 ; Jahrgang 6, Nr. 7. S. 997–1005.
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title = "A Chemical Chaperone Restores Connexin 26 Mutant Activity",
abstract = "Mutations in connexin 26 (Cx26) cause hearing disorders of a varying degree. Herein, to identify compounds capable of restoring the function of mutated Cx26, a novel miniaturized microarray-based screening system was developed to perform an optical assay of Cx26 functionality. These molecules were identified through a viability assay using HeLa cells expressing wild-type (WT) Cx26, which exhibited sensitivity toward the HSP90 inhibitor radicicol in the submicromolar concentration range. Open Cx26 hemichannels are assumed to mediate the passage of molecules up to 1000 Da in size. Thus, by releasing radicicol, WT Cx26 active hemichannels in HeLa cells contribute to a higher survival rate and lower cell viability when Cx26 is mutated. HeLa cells expressing Cx26 mutations exhibited reduced viability in the presence of radicicol, such as the mutants F161S or R184P. Next, molecules exhibiting chemical chaperoning activity, suspected of restoring channel function, were assessed regarding whether they induced superior sensitivity toward radicicol and increased HeLa cell viability. Through a viability assay and microarray-based flux assay that uses Lucifer yellow in HeLa cells, compounds 3 and 8 were identified to restore mutant functionality. Furthermore, thermophoresis experiments revealed that only 3 (VRT-534) exhibited dose-responsive binding to recombinant WT Cx26 and mutant Cx26K188N with half maximal effective concentration values of 19 and ~5 µM, respectively. The findings of this study reveal that repurposing compounds already being used to treat other diseases, such as cystic fibrosis, in combination with functional bioassays and binding tests can help identify novel potential candidates that can be used to treat hearing disorders.",
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T1 - A Chemical Chaperone Restores Connexin 26 Mutant Activity

AU - Wang, Dahua

AU - Wang, Hongling

AU - Fan, Lu

AU - Ludwig, Tobias

AU - Wegner, Andre

AU - Stahl, Frank

AU - Harre, Jennifer

AU - Warnecke, Athanasia

AU - Zeilinger, Carsten

N1 - Parts of the project were funded by DFG Cytolabs ZE 338/16-1 and Cluster of Excellence Hearing4All (EXC 2177/1).

PY - 2023/7/14

Y1 - 2023/7/14

N2 - Mutations in connexin 26 (Cx26) cause hearing disorders of a varying degree. Herein, to identify compounds capable of restoring the function of mutated Cx26, a novel miniaturized microarray-based screening system was developed to perform an optical assay of Cx26 functionality. These molecules were identified through a viability assay using HeLa cells expressing wild-type (WT) Cx26, which exhibited sensitivity toward the HSP90 inhibitor radicicol in the submicromolar concentration range. Open Cx26 hemichannels are assumed to mediate the passage of molecules up to 1000 Da in size. Thus, by releasing radicicol, WT Cx26 active hemichannels in HeLa cells contribute to a higher survival rate and lower cell viability when Cx26 is mutated. HeLa cells expressing Cx26 mutations exhibited reduced viability in the presence of radicicol, such as the mutants F161S or R184P. Next, molecules exhibiting chemical chaperoning activity, suspected of restoring channel function, were assessed regarding whether they induced superior sensitivity toward radicicol and increased HeLa cell viability. Through a viability assay and microarray-based flux assay that uses Lucifer yellow in HeLa cells, compounds 3 and 8 were identified to restore mutant functionality. Furthermore, thermophoresis experiments revealed that only 3 (VRT-534) exhibited dose-responsive binding to recombinant WT Cx26 and mutant Cx26K188N with half maximal effective concentration values of 19 and ~5 µM, respectively. The findings of this study reveal that repurposing compounds already being used to treat other diseases, such as cystic fibrosis, in combination with functional bioassays and binding tests can help identify novel potential candidates that can be used to treat hearing disorders.

AB - Mutations in connexin 26 (Cx26) cause hearing disorders of a varying degree. Herein, to identify compounds capable of restoring the function of mutated Cx26, a novel miniaturized microarray-based screening system was developed to perform an optical assay of Cx26 functionality. These molecules were identified through a viability assay using HeLa cells expressing wild-type (WT) Cx26, which exhibited sensitivity toward the HSP90 inhibitor radicicol in the submicromolar concentration range. Open Cx26 hemichannels are assumed to mediate the passage of molecules up to 1000 Da in size. Thus, by releasing radicicol, WT Cx26 active hemichannels in HeLa cells contribute to a higher survival rate and lower cell viability when Cx26 is mutated. HeLa cells expressing Cx26 mutations exhibited reduced viability in the presence of radicicol, such as the mutants F161S or R184P. Next, molecules exhibiting chemical chaperoning activity, suspected of restoring channel function, were assessed regarding whether they induced superior sensitivity toward radicicol and increased HeLa cell viability. Through a viability assay and microarray-based flux assay that uses Lucifer yellow in HeLa cells, compounds 3 and 8 were identified to restore mutant functionality. Furthermore, thermophoresis experiments revealed that only 3 (VRT-534) exhibited dose-responsive binding to recombinant WT Cx26 and mutant Cx26K188N with half maximal effective concentration values of 19 and ~5 µM, respectively. The findings of this study reveal that repurposing compounds already being used to treat other diseases, such as cystic fibrosis, in combination with functional bioassays and binding tests can help identify novel potential candidates that can be used to treat hearing disorders.

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JF - ACS Pharmacology & Translational Science

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ER -

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