Inhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells

Abstract Sialylation of recombinant therapeutic glycoproteins modulates their pharmacokinetic properties by affecting their in vivo half-life. N-glycan branching on glycoproteins increases the number of potential attachment sites for sialic acid. Here, we introduce a new approach for increasing the...

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Autores principales:	Chung-Geun Lee, Myung Jin Oh, Seung-Yeol Park, Hyun Joo An, Jung Hoe Kim
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/4f90bdf6a4af41ef87c8b171f0e83055
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spelling	oai:doaj.org-article:4f90bdf6a4af41ef87c8b171f0e830552021-12-02T12:33:00ZInhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells10.1038/s41598-018-25580-92045-2322https://doaj.org/article/4f90bdf6a4af41ef87c8b171f0e830552018-05-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-25580-9https://doaj.org/toc/2045-2322Abstract Sialylation of recombinant therapeutic glycoproteins modulates their pharmacokinetic properties by affecting their in vivo half-life. N-glycan branching on glycoproteins increases the number of potential attachment sites for sialic acid. Here, we introduce a new approach for increasing the sialylation of recombinant human erythropoietin (rhEPO) produced in CHO cells by modulating poly-N-acetyllactosamine (poly-LacNAc) biosynthesis. We did not observe an increase in rhEPO sialylation, however, until the feedback inhibition by intracellular cytidine monophosphate-N-acetylneuraminic acid (CMP-Neu5Ac), which is a limiting factor for sialylation, was released. Thus, we found that a combined approach inhibiting poly-LacNAc biosynthesis and releasing CMP-Neu5Ac feedback inhibition produces the most significant increase in rhEPO sialylation in metabolically engineered CHO cells. Furthermore, a detailed analysis of the resulting N-glycan structures using LC/MS revealed increased tri- and tetra- sialylated N-glycan structures accompanied by a reduction of di-sialylated N-glycan structures. These results validate our new approach for glycosylation engineering, and we expect this approach will be useful in future efforts to enhance the efficacy of other therapeutic glycoproteins.Chung-Geun LeeMyung Jin OhSeung-Yeol ParkHyun Joo AnJung Hoe KimNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-9 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Chung-Geun Lee Myung Jin Oh Seung-Yeol Park Hyun Joo An Jung Hoe Kim Inhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells
description	Abstract Sialylation of recombinant therapeutic glycoproteins modulates their pharmacokinetic properties by affecting their in vivo half-life. N-glycan branching on glycoproteins increases the number of potential attachment sites for sialic acid. Here, we introduce a new approach for increasing the sialylation of recombinant human erythropoietin (rhEPO) produced in CHO cells by modulating poly-N-acetyllactosamine (poly-LacNAc) biosynthesis. We did not observe an increase in rhEPO sialylation, however, until the feedback inhibition by intracellular cytidine monophosphate-N-acetylneuraminic acid (CMP-Neu5Ac), which is a limiting factor for sialylation, was released. Thus, we found that a combined approach inhibiting poly-LacNAc biosynthesis and releasing CMP-Neu5Ac feedback inhibition produces the most significant increase in rhEPO sialylation in metabolically engineered CHO cells. Furthermore, a detailed analysis of the resulting N-glycan structures using LC/MS revealed increased tri- and tetra- sialylated N-glycan structures accompanied by a reduction of di-sialylated N-glycan structures. These results validate our new approach for glycosylation engineering, and we expect this approach will be useful in future efforts to enhance the efficacy of other therapeutic glycoproteins.
format	article
author	Chung-Geun Lee Myung Jin Oh Seung-Yeol Park Hyun Joo An Jung Hoe Kim
author_facet	Chung-Geun Lee Myung Jin Oh Seung-Yeol Park Hyun Joo An Jung Hoe Kim
author_sort	Chung-Geun Lee
title	Inhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells
title_short	Inhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells
title_full	Inhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells
title_fullStr	Inhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells
title_full_unstemmed	Inhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells
title_sort	inhibition of poly-lacnac biosynthesis with release of cmp-neu5ac feedback inhibition increases the sialylation of recombinant epo produced in cho cells
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/4f90bdf6a4af41ef87c8b171f0e83055
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Inhibition of poly-LacNAc biosynthesis with release of CMP-Neu5Ac feedback inhibition increases the sialylation of recombinant EPO produced in CHO cells

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