%0 Journal Article
%A Peng, Jiangnan
%A Patil, Sharadrao M.
%A Keire, David A.
%A Chen, Kang
%D 2018
%T Chemical Structure and Composition of Major Glycans
Covalently Linked to Therapeutic Monoclonal Antibodies by Middle-Down
Nuclear Magnetic Resonance
%U https://acs.figshare.com/articles/journal_contribution/Chemical_Structure_and_Composition_of_Major_Glycans_Covalently_Linked_to_Therapeutic_Monoclonal_Antibodies_by_Middle-Down_Nuclear_Magnetic_Resonance/7012928
%R 10.1021/acs.analchem.8b02637.s001
%2 https://ndownloader.figshare.com/files/12868949
%K mAb quality attributes
%K Major Glycans Covalently
%K glycan anomeric peaks
%K mAb drug efficacy
%K glycan structure determination
%K middle-down NMR method
%K MS
%K glycosidic linkage position
%K chemical structure
%K chemical structure similarity
%K chemical information content
%K mAb drugs
%K mAb glycosylation characterization
%X Glycosylation
of monoclonal antibodies (mAbs) is a critical quality
attribute that can impact mAb drug efficacy and safety. The mAb glycans
are inherently heterogeneous in chemical structure and composition
of monosaccharides. The established fluorescence or mass-spectrometry
(MS) detection methods for glycosylation evaluation may require multiple
steps of glycan cleavage or extensive digestion of the mAb, chemical
labeling of the glycans, column separation and report the chemical
identity of glycans indirectly through retention time and molecular
weight values. In demonstrating chemical structure similarity and
comparability among mAb drugs, orthogonal analytical methods for measuring
glycan chemistry are needed to ensure the quality of drug products.
Here, a “middle-down” NMR method is developed as a proof-of-concept
approach to measure the domain-specific glycosylation of marketed
mAb drugs without cleavage of the glycan moieties. Complete glycan 1H/13C chemical shift assignments were obtained
at 13C natural abundance from commercial standard glycans
that allowed unambiguous determination of the chemical structure,
glycosidic linkage position, and anomeric configuration of each monosaccharide
in the major N-glycan scaffolds found in mAb molecules.
The analysis of glycan anomeric peaks in two-dimensional (2D) 1H–13C NMR spectra yielded metrics for clinically
important mAb quality attributes (i.e., galactosylation (Gal%) and
fucosylation (Fuc%)), consistent with literature results using a standard
glycan-mapping method. Therefore, the middle-down NMR method provided
a facile orthogonal measurement for mAb glycosylation characterization
with improved chemical information content on glycan structure determination
and quantification, compared to standard approaches.
%I ACS Publications