Combining random mutagenesis, structure-guided design and next-generation sequencing to mitigate polyreactivity of an anti-IL-21R antibody
Open Access
- 1 January 2021
- journal article
- research article
- Published by Taylor & Francis Ltd in mAbs
- Vol. 13 (1), 1883239
- https://doi.org/10.1080/19420862.2021.1883239
Abstract
Despite substantial technological advances in antibody library and display platform development, the number of approved biotherapeutics from displayed libraries remains limited. In vivo, 20–50% of peripheral B cells undergo a process of receptor editing, which modifies the variable and junctional regions of light chains to delete auto-reactive clones. However, in vitro antibody evolution relies primarily on interaction with antigen, with no in-built checkpoints to ensure that the selected antibodies have not acquired additional specificities or biophysical liabilities during the optimization process. We had previously observed an enrichment of positive charge in the complementarity-determining regions of an anti-IL-21 R antibody during affinity optimization, which correlated with more potent IL-21 neutralization, but poor in vivo pharmacokinetics (PK). There is an emerging body of data that has correlated antibody nonspecificity with poor PK in vivo, and established a series of screening assays that are predictive of this behavior. In this study we revisit the challenge of developing an anti-IL-21 R antibody that can effectively compete with IL-21 for its highly negatively charged paratope while maintaining favorable biophysical properties. In vitro deselection methods that included an excess of negatively charged membrane preparations, or deoxyribonucleic acid, during phage selection of optimization libraries were unsuccessful in avoiding enrichment of highly charged, nonspecific antibody variants. However, a combination of structure-guided rational library design, next-generation sequencing of library outputs and application of linear regression models resulted in the identification of an antibody that maintained high affinity for IL-21 R and exhibited a desirable stability and biophysical profile.Keywords
This publication has 39 references indexed in Scilit:
- Antitumor activity, pharmacokinetics, tumor-homing effect, and hepatotoxicity of a species cross-reactive c-Met antibodyBiochemical and Biophysical Research Communications, 2017
- Reduction of Nonspecificity Motifs in Synthetic Antibody LibrariesJournal of Molecular Biology, 2017
- In silico selection of therapeutic antibodies for development: Viscosity, clearance, and chemical stabilityProceedings of the National Academy of Sciences of the United States of America, 2014
- T-cell dependent immunogenicity of protein therapeutics: Preclinical assessment and mitigationClinical Immunology, 2013
- CDR-restricted engineering of native human scFvs creates highly stable and soluble bifunctional antibodies for subcutaneous deliverymAbs, 2013
- A Fully Human, Allosteric Monoclonal Antibody That Activates the Insulin Receptor and Improves Glycemic ControlDiabetes, 2012
- A simple high-throughput purification method for hit identification in protein screeningJournal of Immunological Methods, 2008
- Neutralizing human monoclonal antibody against H5N1 influenza HA selected from a Fab-phage display libraryVirology Journal, 2008
- The Intrinsic Contributions of Tyrosine, Serine, Glycine and Arginine to the Affinity and Specificity of AntibodiesJournal of Molecular Biology, 2008
- Predicting Changes in the Stability of Proteins and Protein Complexes: A Study of More Than 1000 MutationsJournal of Molecular Biology, 2002