Abstract
The Fc-fusion protein atacicept is currently under clinical investigation for its biotherapeutic application in autoimmune diseases owing to its ability to bind the two cytokines B-Lymphocyte Stimulator (BLyS) and A PRoliferation-Inducing Ligand (APRIL). Like typical recombinant IgG-based therapeutics, atacicept is a glycoprotein whose glycosylation-related heterogeneity arises from the glycosylation-site localization, site-specific occupation and structural diversity of the attached glycans. Here, we present a first comprehensive site-specific N- and O-glycosylation characterization of atacicept using mass spectrometry-based workflows. First, N- and O-glycosylation sites and their corresponding glycoforms were identified. Second, a relative quantitation of the N-glycosylation site microheterogeneity was achieved by glycopeptide analysis, which was further supported by analysis of the released N-glycans. We confirmed the presence of one N-glycosylation site, carrying 47 glycoforms covering 34 different compositions, next to two hinge region O-glycosylation sites with core 1-type glycans. The relative O-glycan distribution was analyzed based on the de-N-glycosylated intact protein species. Overall, N- and O-glycosylation were consistent between two individual production batches.
Original language | English |
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Pages (from-to) | 1053-1063 |
Number of pages | 11 |
Journal | mAbs |
Volume | 11 |
Issue number | 6 |
Early online date | 26 Jul 2019 |
DOIs | |
Publication status | Published - 18 Aug 2019 |
Funding
This project was supported by the European Union (Seventh Framework Programme HighGlycan project, grant number: 278535)
Funders | Funder number |
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Seventh Framework Programme | 278535 |
European Commission | |
Seventh Framework Programme |
Keywords
- atacicept
- Fc-fusion protein
- glycosylation
- N-glycans
- N-glycopeptides
- O-glycopeptides