Rapid high yield production of different glycoforms of Ebola virus monoclonal antibody.

<h4>Background</h4>Fc-glycosylation of monoclonal antibodies (mAbs) has profound implications on the Fc-mediated effector functions. Alteration of this glycosylation may affect the efficiency of an antibody. However, difficulties in the production of mAbs with homogeneous N-glycosylation...

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Autores principales:	Alexandra Castilho, Natasha Bohorova, Josephine Grass, Ognian Bohorov, Larry Zeitlin, Kevin Whaley, Friedrich Altmann, Herta Steinkellner
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2011
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/c6fd415b0f934f4bb541f430c2038645
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spelling	oai:doaj.org-article:c6fd415b0f934f4bb541f430c20386452021-11-18T07:35:54ZRapid high yield production of different glycoforms of Ebola virus monoclonal antibody.1932-620310.1371/journal.pone.0026040https://doaj.org/article/c6fd415b0f934f4bb541f430c20386452011-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/22039433/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203<h4>Background</h4>Fc-glycosylation of monoclonal antibodies (mAbs) has profound implications on the Fc-mediated effector functions. Alteration of this glycosylation may affect the efficiency of an antibody. However, difficulties in the production of mAbs with homogeneous N-glycosylation profiles in sufficient amounts hamper investigations of the potential biological impact of different glycan residues.<h4>Methodology/principal findings</h4>Here we set out to evaluate a transient plant viral based production system for the rapid generation of different glycoforms of a monoclonal antibody. Ebola virus mAb h-13F6 was generated using magnICON expression system in Nicotiana benthamiana, a plant species developed for commercial scale production of therapeutic proteins. h-13F6 was co-expressed with a series of modified mammalian enzymes involved in the processing of complex N-glycans. Using wild type (WT) plants and the glycosylation mutant ΔXTFT that synthesizes human like biantennary N-glycans with terminal N-acetylglucosamine on each branch (GnGn structures) as expression hosts we demonstrate the generation of h-13F6 complex N-glycans with (i) bisected structures, (ii) core α1,6 fucosylation and (iii) β1,4 galactosylated oligosaccharides. In addition we emphasize the significance of precise sub Golgi localization of enzymes for engineering of IgG Fc-glycosylation.<h4>Conclusion</h4>The method described here allows the efficient generation of a series of different human-like glycoforms at large homogeneity of virtually any antibody within one week after cDNA delivery to plants. This accelerates follow up functional studies and thus may contribute to study the biological role of N-glycan residues on Fcs and maximizing the clinical efficacy of therapeutic antibodies.Alexandra CastilhoNatasha BohorovaJosephine GrassOgnian BohorovLarry ZeitlinKevin WhaleyFriedrich AltmannHerta SteinkellnerPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 6, Iss 10, p e26040 (2011)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Alexandra Castilho Natasha Bohorova Josephine Grass Ognian Bohorov Larry Zeitlin Kevin Whaley Friedrich Altmann Herta Steinkellner Rapid high yield production of different glycoforms of Ebola virus monoclonal antibody.
description	<h4>Background</h4>Fc-glycosylation of monoclonal antibodies (mAbs) has profound implications on the Fc-mediated effector functions. Alteration of this glycosylation may affect the efficiency of an antibody. However, difficulties in the production of mAbs with homogeneous N-glycosylation profiles in sufficient amounts hamper investigations of the potential biological impact of different glycan residues.<h4>Methodology/principal findings</h4>Here we set out to evaluate a transient plant viral based production system for the rapid generation of different glycoforms of a monoclonal antibody. Ebola virus mAb h-13F6 was generated using magnICON expression system in Nicotiana benthamiana, a plant species developed for commercial scale production of therapeutic proteins. h-13F6 was co-expressed with a series of modified mammalian enzymes involved in the processing of complex N-glycans. Using wild type (WT) plants and the glycosylation mutant ΔXTFT that synthesizes human like biantennary N-glycans with terminal N-acetylglucosamine on each branch (GnGn structures) as expression hosts we demonstrate the generation of h-13F6 complex N-glycans with (i) bisected structures, (ii) core α1,6 fucosylation and (iii) β1,4 galactosylated oligosaccharides. In addition we emphasize the significance of precise sub Golgi localization of enzymes for engineering of IgG Fc-glycosylation.<h4>Conclusion</h4>The method described here allows the efficient generation of a series of different human-like glycoforms at large homogeneity of virtually any antibody within one week after cDNA delivery to plants. This accelerates follow up functional studies and thus may contribute to study the biological role of N-glycan residues on Fcs and maximizing the clinical efficacy of therapeutic antibodies.
format	article
author	Alexandra Castilho Natasha Bohorova Josephine Grass Ognian Bohorov Larry Zeitlin Kevin Whaley Friedrich Altmann Herta Steinkellner
author_facet	Alexandra Castilho Natasha Bohorova Josephine Grass Ognian Bohorov Larry Zeitlin Kevin Whaley Friedrich Altmann Herta Steinkellner
author_sort	Alexandra Castilho
title	Rapid high yield production of different glycoforms of Ebola virus monoclonal antibody.
title_short	Rapid high yield production of different glycoforms of Ebola virus monoclonal antibody.
title_full	Rapid high yield production of different glycoforms of Ebola virus monoclonal antibody.
title_fullStr	Rapid high yield production of different glycoforms of Ebola virus monoclonal antibody.
title_full_unstemmed	Rapid high yield production of different glycoforms of Ebola virus monoclonal antibody.
title_sort	rapid high yield production of different glycoforms of ebola virus monoclonal antibody.
publisher	Public Library of Science (PLoS)
publishDate	2011
url	https://doaj.org/article/c6fd415b0f934f4bb541f430c2038645
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Rapid high yield production of different glycoforms of Ebola virus monoclonal antibody.

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