Thomson, R.I., Gardner, R.A., Strohfeldt, K. et al. (4 more authors) (2017) Analysis of Three Epoetin Alpha Products by LC and LC-MS Indicates Differences in Glycosylation Critical Quality Attributes, Including Sialic Acid Content. ANALYTICAL CHEMISTRY, 89 (12). pp. 6455-6462. ISSN 0003-2700
Abstract
Erythropoietin (EPO) is one of the main therapeutics used to treat anemic patients, greatly improving their quality of life. In this study, biosimilars Binocrit and a development product, called here CIGB-EPO, were compared to the originator product, Eprex. All three are epoetin alpha products, reputed to have similar glycosylation profiles. The quality, safety, and efficacy of this biotherapeutic depend on the following glycosylation critical quality attributes (GCQAs): sialylation, N-glycolyl-neuraminic acid (Neu5Gc) content, branching, N-acetyl-lactosamine (LacNAc) extensions, and O-acetylation pattern. Reverse-phase ultra-high-pressure liquid chromatography (RP-UHPLC) analysis of acid-released, 1,2-diamino-4,5-methylenedioxybenzene (DMB) labeled sialic acid derivatives and hydrophilic interaction liquid chromatography (HILIC) in combination with mass spectrometry (HILIC-UHPLC-MS) of procainamide (PROC) labeled N-glycans were the analytical tools used. An automated method for enzymatic release and PROC labeling was applied for the first time to the erythropoiesis stimulating agent (ESA) products, which facilitated novel, in-depth characterization, and allowed identification of precise structural features including the location of O-acetyl groups on sialic acid (SA) moieties. Samples were digested by a sialate-O-acetylesterase (NanS) to confirm the presence of O-acetyl groups. It was found that Eprex contained the greatest relative abundance of O-acetylated derivatives, Binocrit expressed the least Neu5Gc, and CIGB-EPO showed the greatest variety of high-mannose-phosphate structures. The sialylation and LacNAc extension patterns of the three ESAs were similar, with a maximum of four N-acetyl-neuraminic acid (Neu5Ac) moieties detected per glycan. Such differences in SA derivatization, particularly O-acetylation, could have consequences for the quality and safety of a biotherapeutic, as well as its efficacy.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | Copyright © 2017 American Chemical Society ACS Editors' Choice - This is an open access article published under an ACS AuthorChoice License, which permits copying and redistribution of the article or any adaptations for non-commercial purposes (http://pubs.acs.org/page/policy/authorchoice_termsofuse.html) |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Medicine, Dentistry and Health (Sheffield) > School of Clinical Dentistry (Sheffield) |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 19 Jul 2017 11:04 |
Last Modified: | 19 Jul 2017 11:04 |
Published Version: | https://doi.org/10.1021/acs.analchem.7b00353 |
Status: | Published |
Publisher: | American Chemical Society |
Refereed: | Yes |
Identification Number: | 10.1021/acs.analchem.7b00353 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:119112 |