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Cited by 73Viral Infectious Diseases and Gene Expression in InsectsGlycosylation and Glycoproteins ResearchProtein purification and stability

Chinese hamster ovary (CHO) host cell engineering to increase sialylation of recombinant therapeutic proteins by modulating sialyltransferase expression

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Lin, Nan, Mascarenhas, Joaquina, Sealover, Natalie R., George, Henry J., Brooks, Jeanne, Kayser, Kevin J., Gau, Brian, Yasa, Isil +2 more·2015·Biotechnology Progress
DOI 10.1002/btpr.2038PMC PMC4492121
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Abstract

N‐Glycans of human proteins possess both α2,6‐ and α2,3‐linked terminal sialic acid (SA). Recombinant glycoproteins produced in Chinese hamster overy (CHO) only have α2,3‐linkage due to the absence of α2,6‐sialyltransferase (St6gal1) expression. The Chinese hamster ST6GAL1 was successfully overexpressed using a plasmid expression vector in three recombinant immunoglobulin G (IgG)‐producing CHO cell lines. The stably transfected cell lines were enriched for ST6GAL1 overexpression using FITC‐Sambucus nigra (SNA) lectin that preferentially binds α2,6‐linked SA. The presence of α2,6‐linked SA was confirmed using a novel LTQ Linear Ion Trap Mass Spectrometry (LTQ MS) method including MSn fragmentation in the enriched ST6GAL1 Clone 27. Furthermore, the total SA (mol/mol) in IgG produced by the enriched ST6GAL1 Clone 27 increased by 2‐fold compared to the control. For host cell engineering, the CHOZN® GS host cell line was transfected and enriched for ST6GAL1 overexpression. Single‐cell clones were derived from the enriched population and selected based on FITC‐SNA staining and St6gal1 expression. Two clones (“ST6GAL1 OE Clone 31 and 32”) were confirmed for the presence of α2,6‐linked SA in total host cell protein extracts. ST6GAL1 OE Clone 32 was subsequently used to express SAFC human IgG1. The recombinant IgG expressed in this host cell line was confirmed to have α2,6‐linked SA and increased total SA content. In conclusion, overexpression of St6gal1 is sufficient to produce recombinant proteins with increased sialylation and more human‐like glycoprofiles without combinatorial engineering of other sialylation pathway genes. This work represents our ongoing effort of glycoengineering in CHO host cell lines for the development of “bio‐better” protein therapeutics and cell culture vaccine production. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:334–346, 2015

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  • The ST6Gal1 gene of the Chinese hamster was also recently used in stable expression of different CHO cell lines producing recombinant IgGs.

    “As genome-editing strategies, the ST6Gal1 gene of the Chinese hamster was also recently used in stable expression of different CHO cell lines producing recombinant IgGs (Lin et al., 2015).”

    From Therapeutic glycoprotein production in mammalian cells (2017) · cites this paper as Lin et al., 2015

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Cited by

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Authors
Lin, N., Mascarenhas, J., Sealover, N. R., George, H. J., Brooks, J., Kayser, K. J., Gau, B., Yasa, I., Azadi, P., & Archer‐Hartmann, S.
Affiliations
  • Sigma-Aldrich (United States) (US)
  • University of Georgia (US)
Corresponding author
Lin, Nan
ORCID
Venue
Biotechnology Progress
Publisher
Wiley
Published
2015-03
Volume
31
Issue
2
Pages
334-346
ISSN
  • 1520-6033 (ISSN-L)
  • 8756-7938
DOI
10.1002/btpr.2038
PMC
PMC4492121
OpenAlex
W2123900742
Cited by
73
References
3
Citations by year
2026: 8 · 2025: 5 · 2024: 1 · 2023: 4 · 2022: 4 · 2021: 10 · 2020: 9 · 2019: 8 · 2018: 8 · 2017: 7 · 2016: 5 · 2015: 4
FWCI
3.47
Type
journal article
Language
en
Open access
Licence
onlinelibrary.wiley.com/termsAndConditions#vor (vor)
Locations
  • Biotechnology Progress (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
Funders
National Institute of General Medical Sciences (P41GM103490)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Viral Infectious Diseases and Gene Expression in Insects
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Glycosylation and Glycoproteins Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein purification and stability
Keywords
sialylation, α2,6-linked sialic acid, glycoengineering, N-glycan profiling
MeSH terms
Animals, Cricetulus, Cricetinae, Polysaccharides, Recombinant Proteins, Sialyltransferases, CHO Cells, N-Acetylneuraminic Acid, Cell Engineering, Metabolic Engineering

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