Simultaneous affinity maturation and developability enhancement using natural liability-free CDRs. Issue 1 (31st December 2022)

• Record Type: Journal Article
• Authors: Teixeira, Andre A. R.; D'Angelo, Sara; Erasmus, M. Frank; Leal-Lopes, Camila; Ferrara, Fortunato; Spector, Laura P.; Naranjo, Leslie; Molina, Esteban; Max, Tamara; DeAguero, Ashley; Perea, Katherine; Stewart, Shaun; Buonpane, Rebecca A.; Nastri, Horacio G.; Bradbury, Andrew R. M.
• Is Part Of: MAbs. Volume 14:Issue 1(2022)
• Publication Date: 2022-12-31
• Subjects: Affinity maturation -- CDR shuffling -- developability -- liability -- library construction -- off-rate -- scFv; Monoclonal antibodies -- Therapeutic use -- Periodicals; Monoclonal antibodies -- Periodicals; Antibodies, Monoclonal -- Periodicals; 616.0798
• DOI: 10.1080/19420862.2022.2115200
• Languages: English
• ISSNs: 1942-0862

Abstract

ABSTRACT: Affinity maturation is often a necessary step for the development of potent therapeutic molecules. Many different diversification strategies have been used for antibody affinity maturation, including error-prone PCR, chain shuffling, and targeted complementary-determining region (CDR) mutation. Although effective, they can negatively impact antibody stability or alter epitope recognition. Moreover, they do not address the presence of sequence liabilities, such as glycosylation, asparagine deamidation, aspartate isomerization, aggregation motifs, and others. Such liabilities, if present or inadvertently introduced, can potentially create the need for new rounds of engineering, or even abolish the value of the antibody as a therapeutic molecule. Here, we demonstrate a sequence agnostic method to improve antibody affinities, while simultaneously eliminating sequence liabilities and retaining the same epitope binding as the parental antibody. This was carried out using a defined collection of natural CDRs as the diversity source, purged of sequence liabilities, and matched to the antibody germline gene family. These CDRs were inserted into the lead molecule in one or two sites at a time (LCDR1-2, LCDR3, HCDR1-2) while retaining the HCDR3 and framework regions unchanged. The final analysis of 92 clones revealed 81 unique variants, with each of 24 tested variants having the same epitope specificity as the parental molecule. Of these, the average affinity improved by over 100 times (to 96 pM), and the best affinity improvement was 231-fold (to 32 pM). Abbreviations: CDR: complementarity determining region; FACS: fluorescence-activated cell sorting; ka : association rate; kd : dissociation rate; KD : dissociation constant; scFv: single-chain variable fragment; SPR: surface plasmon resonance