title: Correlating the Impact of Well-Defined Oligosaccharide Structures on Physical Stability Profiles of IgG1-Fc Glycoforms.
abstract: As part of a series of articles in this special issue describing 4 well-defined IgG1-Fc glycoforms as a model system for biosimilarity analysis (high mannose-Fc, Man5-Fc, GlcNAc-Fc and N297Q-Fc aglycosylated), the focus of this work is comparisons of their physical properties. A trend of decreasing apparent solubility (thermodynamic activity) by polyethylene glycol precipitation (pH 4.5, 6.0) and lower conformational stability by differential scanning calorimetry (pH 4.5) was observed with reducing size of the N297-linked oligosaccharide structures. Using multiple high-throughput biophysical techniques, the physical stability of the Fc glycoproteins was then measured in 2 formulations (NaCl and sucrose) across a wide range of temperatures (10°C-90°C) and pH (4.0-7.5) conditions. The data sets were used to construct 3-index empirical phase diagrams and radar charts to visualize the regions of protein structural stability. Each glycoform showed improved stability in the sucrose (vs. salt) formulation. The HM-Fc and Man5-Fc displayed the highest relative stability, followed by GlcNAc-Fc, with N297Q-Fc being the least stable. Thus, the overall physical stability profiles of the 4 IgG1-Fc glycoforms also show a correlation with oligosaccharide structure. These data sets are used to develop a mathematical model for biosimilarity analysis (as described in a companion article by Kim et al. in this issue).
authors: Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047; Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, Kansas 66047.
authors: Apurva S
authors: AS
authors: More
authors: Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047; Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, Kansas 66047.
authors: Vishal M
authors: VM
authors: Toprani
authors: Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047.
authors: Solomon Z
authors: SZ
authors: Okbazghi
authors: Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047; Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, Kansas 66047.
authors: Jae H
authors: JH
authors: Kim
authors: Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047; Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, Kansas 66047.
authors: Sangeeta B
authors: SB
authors: Joshi
authors: Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047; Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, Kansas 66047.
authors: C Russell
authors: CR
authors: Middaugh
authors: Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047.
authors: Thomas J
authors: TJ
authors: Tolbert
authors: Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, Kansas 66047; Macromolecule and Vaccine Stabilization Center, University of Kansas, Lawrence, Kansas 66047. Electronic address: volkin@ku.edu.
authors: David B
authors: DB
authors: Volkin
mesh_terms: D004355:Drug Stability; D006023:Glycoproteins / Q000737:chemistry*; D006031:Glycosylation; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry*; D009844:Oligosaccharides / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000737:chemistry; D011487:Protein Conformation
chemical_list: D006023:Glycoproteins; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D009844:Oligosaccharides; D011092:Polyethylene Glycols
keywords: Fc; IgG; conformation; formulation; glycosylation; monoclonal antibody; spectroscopy; stability