期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2015
卷号：112
期号：50
页码：15354-15359
DOI：10.1073/pnas.1510944112
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceImmunized animals are a key source of monoclonal antibodies used to treat human diseases. Before clinical use, animal antibodies are typically "humanized" by laborious and suboptimal methods that transfer their full target binding loops (a.k.a. CDRs) into human frameworks. We report an optimal method, where the CDRs from species such as rodents and chickens can be adapted to fit human frameworks in which we have clinical and manufacturing confidence. The Augmented Binary Substitution (ABS) process exploits the fundamental plasticity of antibody CDRs to ultrahumanize antibodies from key species in a single pass. ABS results in a final antibody that is much closer to human germ line in the frameworks and CDRs, minimizing immunogenicity risks in man and maximizing the therapeutic potential of the antibody. Although humanized antibodies have been highly successful in the clinic, all current humanization techniques have potential limitations, such as: reliance on rodent hosts, immunogenicity due to high non-germ-line amino acid content, v-domain destabilization, expression and formulation issues. This study presents a technology that generates stable, soluble, ultrahumanized antibodies via single-step complementarity-determining region (CDR) germ-lining. For three antibodies from three separate key immune host species, binary substitution CDR cassettes were inserted into preferred human frameworks to form libraries in which only the parental or human germ-line destination residue was encoded at each position. The CDR-H3 in each case was also augmented with 1 {+/-} 1 random substitution per clone. Each library was then screened for clones with restored antigen binding capacity. Lead ultrahumanized clones demonstrated high stability, with affinity and specificity equivalent to, or better than, the parental IgG. Critically, this was mainly achieved on germ-line frameworks by simultaneously subtracting up to 19 redundant non-germ-line residues in the CDRs. This process significantly lowered non-germ-line sequence content, minimized immunogenicity risk in the final molecules and provided a heat map for the essential non-germ-line CDR residue content of each antibody. The ABS technology therefore fully optimizes the clinical potential of antibodies from rodents and alternative immune hosts, rendering them indistinguishable from fully human in a simple, single-pass process.
