A Human-Derived Monoclonal Antibody Targeting Extracellular Connexin Domain Selectively Modulates Hemichannel Function
Connexin hemichannels, which are plasma membrane hexameric channels (connexons) composed of connexin protein protomers, have been implicated in a host of physiological processes and pathological conditions. A number of single point pathological mutations impart a “leaky” character to the affected he...
Main Authors: | , , , , , , , , , , , , |
---|---|
Format: | Article |
Language: | English |
Published: |
Frontiers Media S.A.
2019-06-01
|
Series: | Frontiers in Physiology |
Subjects: | |
Online Access: | https://www.frontiersin.org/article/10.3389/fphys.2019.00392/full |
id |
doaj-b3ac6b95d41f4cd7a0dd04a751006fbd |
---|---|
record_format |
Article |
spelling |
doaj-b3ac6b95d41f4cd7a0dd04a751006fbd2020-11-24T21:30:38ZengFrontiers Media S.A.Frontiers in Physiology1664-042X2019-06-011010.3389/fphys.2019.00392447350A Human-Derived Monoclonal Antibody Targeting Extracellular Connexin Domain Selectively Modulates Hemichannel FunctionGaia Ziraldo0Gaia Ziraldo1Damiano Buratto2Yuanyuan Kuang3Liang Xu4Andrea Carrer5Andrea Carrer6Chiara Nardin7Chiara Nardin8Francesco Chiani9Anna Maria Salvatore10Gaetano Paludetti11Gaetano Paludetti12Richard A. Lerner13Guang Yang14Francesco Zonta15Fabio Mammano16Fabio Mammano17Fabio Mammano18CNR Institute of Cell Biology and Neurobiology, Monterotondo, ItalyInstitute of Otolaryngology, Università Cattolica del Sacro Cuore, Rome, ItalyShanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, Shanghai, ChinaShanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, Shanghai, ChinaShanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, Shanghai, ChinaCNR Institute of Cell Biology and Neurobiology, Monterotondo, ItalyDepartment of Physics and Astronomy “G. Galilei”, University of Padova, Padua, ItalyCNR Institute of Cell Biology and Neurobiology, Monterotondo, ItalyDepartment of Physics and Astronomy “G. Galilei”, University of Padova, Padua, ItalyCNR Institute of Cell Biology and Neurobiology, Monterotondo, ItalyCNR Institute of Cell Biology and Neurobiology, Monterotondo, ItalyInstitute of Otolaryngology, Università Cattolica del Sacro Cuore, Rome, ItalyFondazione Policlinico Universitario A. Gemelli IRCCS, Rome, ItalyShanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, Shanghai, ChinaShanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, Shanghai, ChinaShanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, Shanghai, ChinaCNR Institute of Cell Biology and Neurobiology, Monterotondo, ItalyShanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, Shanghai, ChinaDepartment of Physics and Astronomy “G. Galilei”, University of Padova, Padua, ItalyConnexin hemichannels, which are plasma membrane hexameric channels (connexons) composed of connexin protein protomers, have been implicated in a host of physiological processes and pathological conditions. A number of single point pathological mutations impart a “leaky” character to the affected hemichannels, i.e., make them more active or hyperactive, suggesting that normal physiological condition could be recovered using selective hemichannel inhibitors. Recently, a human-derived monoclonal antibody named abEC1.1 has been shown to inhibit both wild type and hyperactive hemichannels composed of human (h) connexin 26 (hCx26) subunits. The aims of this work were (1) to characterize further the ability of abEC1.1 to selectively modulate connexin hemichannel function and (2) to assess its in vitro stability in view of future translational applications. In silico analysis of abEC1.1 interaction with the hCx26 hemichannel identified critically important extracellular domain amino acids that are conserved in connexin 30 (hCx30) and connexin 32 (hCx32). Patch clamp experiments performed in HeLa DH cells confirmed the inhibition efficiency of abEC1.1 was comparable for hCx26, hCx30 and hCx32 hemichannels. Of note, even a single amino acid difference in the putative binding region reduced drastically the inhibitory effects of the antibody on all the other tested hemichannels, namely hCx30.2/31.3, hCx30.3, hCx31, hCx31.1, hCx37, hCx43 and hCx45. Plasma membrane channels composed of pannexin 1 were not affected by abEC1.1. Finally, size exclusion chromatography assays showed the antibody does not aggregate appreciably in vitro. Altogether, these results indicate abEC1.1 is a promising tool for further translational studies.https://www.frontiersin.org/article/10.3389/fphys.2019.00392/fullconnexin hemichannelsrare diseasesphage display librariestherapeutic monoclonal antibodiesmolecular dynamicspatch clamp |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Gaia Ziraldo Gaia Ziraldo Damiano Buratto Yuanyuan Kuang Liang Xu Andrea Carrer Andrea Carrer Chiara Nardin Chiara Nardin Francesco Chiani Anna Maria Salvatore Gaetano Paludetti Gaetano Paludetti Richard A. Lerner Guang Yang Francesco Zonta Fabio Mammano Fabio Mammano Fabio Mammano |
spellingShingle |
Gaia Ziraldo Gaia Ziraldo Damiano Buratto Yuanyuan Kuang Liang Xu Andrea Carrer Andrea Carrer Chiara Nardin Chiara Nardin Francesco Chiani Anna Maria Salvatore Gaetano Paludetti Gaetano Paludetti Richard A. Lerner Guang Yang Francesco Zonta Fabio Mammano Fabio Mammano Fabio Mammano A Human-Derived Monoclonal Antibody Targeting Extracellular Connexin Domain Selectively Modulates Hemichannel Function Frontiers in Physiology connexin hemichannels rare diseases phage display libraries therapeutic monoclonal antibodies molecular dynamics patch clamp |
author_facet |
Gaia Ziraldo Gaia Ziraldo Damiano Buratto Yuanyuan Kuang Liang Xu Andrea Carrer Andrea Carrer Chiara Nardin Chiara Nardin Francesco Chiani Anna Maria Salvatore Gaetano Paludetti Gaetano Paludetti Richard A. Lerner Guang Yang Francesco Zonta Fabio Mammano Fabio Mammano Fabio Mammano |
author_sort |
Gaia Ziraldo |
title |
A Human-Derived Monoclonal Antibody Targeting Extracellular Connexin Domain Selectively Modulates Hemichannel Function |
title_short |
A Human-Derived Monoclonal Antibody Targeting Extracellular Connexin Domain Selectively Modulates Hemichannel Function |
title_full |
A Human-Derived Monoclonal Antibody Targeting Extracellular Connexin Domain Selectively Modulates Hemichannel Function |
title_fullStr |
A Human-Derived Monoclonal Antibody Targeting Extracellular Connexin Domain Selectively Modulates Hemichannel Function |
title_full_unstemmed |
A Human-Derived Monoclonal Antibody Targeting Extracellular Connexin Domain Selectively Modulates Hemichannel Function |
title_sort |
human-derived monoclonal antibody targeting extracellular connexin domain selectively modulates hemichannel function |
publisher |
Frontiers Media S.A. |
series |
Frontiers in Physiology |
issn |
1664-042X |
publishDate |
2019-06-01 |
description |
Connexin hemichannels, which are plasma membrane hexameric channels (connexons) composed of connexin protein protomers, have been implicated in a host of physiological processes and pathological conditions. A number of single point pathological mutations impart a “leaky” character to the affected hemichannels, i.e., make them more active or hyperactive, suggesting that normal physiological condition could be recovered using selective hemichannel inhibitors. Recently, a human-derived monoclonal antibody named abEC1.1 has been shown to inhibit both wild type and hyperactive hemichannels composed of human (h) connexin 26 (hCx26) subunits. The aims of this work were (1) to characterize further the ability of abEC1.1 to selectively modulate connexin hemichannel function and (2) to assess its in vitro stability in view of future translational applications. In silico analysis of abEC1.1 interaction with the hCx26 hemichannel identified critically important extracellular domain amino acids that are conserved in connexin 30 (hCx30) and connexin 32 (hCx32). Patch clamp experiments performed in HeLa DH cells confirmed the inhibition efficiency of abEC1.1 was comparable for hCx26, hCx30 and hCx32 hemichannels. Of note, even a single amino acid difference in the putative binding region reduced drastically the inhibitory effects of the antibody on all the other tested hemichannels, namely hCx30.2/31.3, hCx30.3, hCx31, hCx31.1, hCx37, hCx43 and hCx45. Plasma membrane channels composed of pannexin 1 were not affected by abEC1.1. Finally, size exclusion chromatography assays showed the antibody does not aggregate appreciably in vitro. Altogether, these results indicate abEC1.1 is a promising tool for further translational studies. |
topic |
connexin hemichannels rare diseases phage display libraries therapeutic monoclonal antibodies molecular dynamics patch clamp |
url |
https://www.frontiersin.org/article/10.3389/fphys.2019.00392/full |
work_keys_str_mv |
AT gaiaziraldo ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT gaiaziraldo ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT damianoburatto ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT yuanyuankuang ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT liangxu ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT andreacarrer ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT andreacarrer ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT chiaranardin ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT chiaranardin ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT francescochiani ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT annamariasalvatore ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT gaetanopaludetti ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT gaetanopaludetti ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT richardalerner ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT guangyang ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT francescozonta ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT fabiomammano ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT fabiomammano ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT fabiomammano ahumanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT gaiaziraldo humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT gaiaziraldo humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT damianoburatto humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT yuanyuankuang humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT liangxu humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT andreacarrer humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT andreacarrer humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT chiaranardin humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT chiaranardin humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT francescochiani humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT annamariasalvatore humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT gaetanopaludetti humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT gaetanopaludetti humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT richardalerner humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT guangyang humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT francescozonta humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT fabiomammano humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT fabiomammano humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction AT fabiomammano humanderivedmonoclonalantibodytargetingextracellularconnexindomainselectivelymodulateshemichannelfunction |
_version_ |
1725962503583694848 |