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Biophysical Characterization and Stability of Modified IgG1 Antibodies with Different Hexamerization Propensities.
- Source :
-
Journal of Pharmaceutical Sciences . Jun2022, Vol. 111 Issue 6, p1587-1598. 12p. - Publication Year :
- 2022
-
Abstract
- The hexamerization of natural, human IgG antibodies after cell surface antigen binding can induce activation of the classical complement pathway. Mutations stimulating Fc domain-mediated hexamerization can potentiate complement activation and induce the clustering of cell surface receptors, a finding that was applied to different clinically investigated antibody therapeutics. Here, we biophysically characterized how increased self-association of IgG1 antibody variants with different hexamerization propensity may impact their developability, rather than functional properties. Self-Interaction Chromatography, Dynamic Light Scattering and PEG-induced precipitation showed that IgG variant self-association at neutral pH increased in the order wild type (WT) < E430G < E345K < E345R < E430G-E345R-S440Y, consistent with functional activity. Self-association was strongly pH-dependent, and single point mutants were fully monomeric at pH 5. Differential Scanning Calorimetry and Fluorimetry showed that mutation E430G decreased conformational stability. Interestingly, heat-induced unfolding facilitated by mutation E430G was reversible at 60°C, while a solvent-exposed hydrophobic mutation caused irreversible aggregation. Remarkably, neither increased dynamic self-association propensity at neutral pH nor decreased conformational stability substantially affected the stability of concentrated variants E430G or E345K during storage for two years at 2-8°C. We discuss how these findings may inform the design and development of IgG-based therapeutics. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 00223549
- Volume :
- 111
- Issue :
- 6
- Database :
- Academic Search Index
- Journal :
- Journal of Pharmaceutical Sciences
- Publication Type :
- Academic Journal
- Accession number :
- 156911045
- Full Text :
- https://doi.org/10.1016/j.xphs.2022.02.016