| Download | - View accepted manuscript: Production of α2,6-sialylated IgG1 in CHO cells (PDF, 904 KB)
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| DOI | Resolve DOI: http://doi.org/10.1080/19420862.2015.1029215 |
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| Author | Search for: Raymond, Céline1 ; Search for: Robotham, Anna1 ; Search for: Spearman, Maureen; Search for: Butler, Michael; Search for: Kelly, John1 ; Search for: Durocher, Yves1 |
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| Name affiliation | - National Research Council Canada. Human Health Therapeutics
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| Type | Article |
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| Journal title | mAbs |
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| ISSN | 1942-0862
1942-0870 |
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| Volume | 7 |
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| Issue | 3 |
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| Pages | 571–583; # of pages: 13 |
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| Subject | B4GALT1; CHO cells; ECL, Erythrina Cristagalli lectin; GT, β1,4-galactosyltransferase 1; HILIC, hydrophilic interaction liquid chromatography; IgG1; LC-ESI-MS, liquid chromatography coupled to electrospray ionization mass spectrometry; MAL-II, Maackia Amurensis lectin II; N-glycosylation; PEI, polyethylenimine; SIAT1; SNA, Sambucus Nigra agglutinin; ST6, α2,6-sialyltransferase 1; TZM, trastuzumab (Herceptin®); cIEF, capillary zone electrophoresis isoelectric focusing; mAbs, monoclonal antibodies; sialylation; transfection; α2,3SA, α2,3-linked sialic acid; α2,6SA, α2,6-linked sialic acid |
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| Abstract | The presence of α2,6-sialic acids on the Fc N-glycan provides anti-inflammatory properties to the IgGs through a mechanism that remains unclear. Fc-sialylated IgGs are rare in humans as well as in industrial host cell lines such as Chinese hamster ovary (CHO) cells. Facilitated access to well-characterized α2,6-sialylated IgGs would help elucidate the mechanism of this intriguing IgG's effector function. This study presents a method for the efficient Fc glycan α2,6-sialylation of a wild-type and a F243A IgG1 mutant by transient co-expression with the human α2,6-sialyltransferase 1 (ST6) and β1,4-galactosyltransferase 1 (GT) in CHO cells. Overexpression of ST6 alone only had a moderate effect on the glycoprofiles, whereas GT alone greatly enhanced Fc-galactosylation, but not sialylation. Overexpression of both GT and ST6 was necessary to obtain a glycoprofile dominated by α2,6-sialylated glycans in both antibodies. The wild-type was composed of the G2FS(6)1 glycan (38%) with remaining unsialylated glycans, while the mutant glycoprofile was essentially composed of G2FS(6)1 (25%), G2FS(3,6)2 (16%) and G2FS(6,6)2 (37%). The α2,6-linked sialic acids represented over 85% of all sialic acids in both antibodies. We discuss how the limited sialylation level in the wild-type IgG1 expressed alone or with GT results from the glycan interaction with Fc's amino acid residues or from intrinsic galactosyl- and sialyl-transferases substrate specificities. |
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| Publication date | 2015-04-15 |
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| Publisher | Taylor & Francis Group |
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| Language | English |
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| Peer reviewed | Yes |
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| NRC number | NRC-HHT-53284 |
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| NPARC number | 21275405 |
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| Export citation | Export as RIS |
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| Report a correction | Report a correction |
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Record identifier | e6369b2f-f442-4fe0-96b1-6e446ebe188f | | Record created | 2015-07-08 |
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| Record modified | 2016-05-09 |
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