Understanding the Increased Aggregation Propensity of a Light-Exposed IgG1 Monoclonal Antibody Using Hydrogen Exchange Mass Spectrometry, Biophysical Characterization, and Structural Analysis.

Autor: Bommana R; Department of Pharmaceutical Chemistry, University of Kansas, 2095 Constant Avenue, Lawrence, Kansas 66047., Chai Q; Eli Lilly Biotechnology Center, San Diego, California 92121., Schöneich C; Department of Pharmaceutical Chemistry, University of Kansas, 2095 Constant Avenue, Lawrence, Kansas 66047., Weiss WF 4th; Biopharmaceutical Research and Development, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285., Majumdar R; Biopharmaceutical Research and Development, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285. Electronic address: majumdar_ranajoy@lilly.com.
Jazyk: angličtina
Zdroj: Journal of pharmaceutical sciences [J Pharm Sci] 2018 Jun; Vol. 107 (6), pp. 1498-1511. Date of Electronic Publication: 2018 Jan 31.
DOI: 10.1016/j.xphs.2018.01.017
Abstrakt: This work compares the conformational stability, backbone flexibility, and aggregation propensity of monomer and dimer fractions of an IgG1 monoclonal antibody (mAb) generated on UVA light exposure for up to 72 h collected by preparative size-exclusion chromatography, compared with unstressed control. UVA light exposure induced covalent aggregation, and fragmentation as measured by size-exclusion chromatography, sodium dodecyl sulfate polyacrylamide gel electrophoresis, and extensive oxidation of specific methionine residues (Met 257, Met 433, and Met 109) in both size fractions identified by reverse phase chromatography coupled to mass spectrometry. Compared with unstressed mAb, both the monomer and dimer fractionated from 72 h UVA light-exposed mAb had decreased thermal melting temperatures (T m1 ) by 1.4°C as measured by differential scanning calorimetry, minor changes in tertiary structure as measured by near-UV CD, increased monomer loss, and aggregation on accelerated storage at 35°C. Hydrogen/deuterium exchange mass spectrometry identified local segments with increased flexibility in C H 2 and C H 3 domains of both size fractions, and decreased flexibility in few segments of F ab and C H 1 domains in the dimer fraction. Segment 247-256 in heavy chain, an established aggregation hotspot in IgG1 mAbs had large increase in flexibility in both size fractions compared with unstressed mAb.
(Copyright © 2018 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
Externí odkaz: