Format

Send to

Choose Destination
Angew Chem Int Ed Engl. 2016 Feb 12;55(7):2361-7. doi: 10.1002/anie.201508723. Epub 2016 Jan 12.

Optimal Synthetic Glycosylation of a Therapeutic Antibody.

Author information

1
Department of Chemistry, University of Oxford, Chemistry Research Laboratory, Mansfield Road, Oxford, OX1 3TA, UK.
2
Department of Pharmacology, University of Oxford, Oxford, OX1 3QT, UK.
3
Department of Chemistry, University of Oxford, Chemistry Research Laboratory, Mansfield Road, Oxford, OX1 3TA, UK. carol.robinson@chem.ox.ac.uk.
4
Department of Chemistry, University of Oxford, Chemistry Research Laboratory, Mansfield Road, Oxford, OX1 3TA, UK. ben.davis@chem.ox.ac.uk.

Abstract

Glycosylation patterns in antibodies critically determine biological and physical properties but their precise control is a significant challenge in biology and biotechnology. We describe herein the optimization of an endoglycosidase-catalyzed glycosylation of the best-selling biotherapeutic Herceptin, an anti-HER2 antibody. Precise MS analysis of the intact four-chain Ab heteromultimer reveals nonspecific, non-enzymatic reactions (glycation), which are not detected under standard denaturing conditions. This competing reaction, which has hitherto been underestimated as a source of side products, can now be minimized. Optimization allowed access to the purest natural form of Herceptin to date (≥90 %). Moreover, through the use of a small library of sugars containing non-natural functional groups, Ab variants containing defined numbers of selectively addressable chemical tags (reaction handles at Sia C1) in specific positions (for attachment of cargo molecules or "glycorandomization") were readily generated.

KEYWORDS:

antibodies; endoglycosidases; glycoengineering; glycosylation; native mass spectrometry

PMID:
26756880
PMCID:
PMC4973692
DOI:
10.1002/anie.201508723
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Wiley Icon for PubMed Central
Loading ...
Support Center