Development of purification processes for fully human bispecific antibodies based upon modification of protein A binding avidity. Issue 4 (18th May 2016)

• Record Type: Journal Article
• Authors: Tustian, Andrew D.; Endicott, Christine; Adams, Benjamin; Mattila, John; Bak, Hanne
• Is Part Of: MAbs. Volume 8:Issue 4(2016)
• Page Start: 828
• Page End: 838
• Publication Date: 2016-05-18
• Subjects: Affinity chromatography -- bispecific antibody -- downstream process -- elution pH -- heterodimeric antibody -- protein A chromatography -- protein purification; Monoclonal antibodies -- Therapeutic use -- Periodicals; Monoclonal antibodies -- Periodicals; Antibodies, Monoclonal -- Periodicals; 616.0798
• DOI: 10.1080/19420862.2016.1160192
• Languages: English
• ISSNs: 1942-0862

Abstract

ABSTRACT: There is strong interest in the design of bispecific monoclonal antibodies (bsAbs) that can simultaneously bind 2 distinct targets or epitopes to achieve novel mechanisms of action and efficacy. Multiple bispecific formats have been proposed and are currently under development. Regeneron's bispecific technology is based upon a standard fully human IgG antibody in order to minimize immunogenicity and improve the pharmacokinetic profile. A single common light chain and 2 distinct heavy chains combine to form the bispecific molecule. One of the heavy chains contains a chimeric Fc sequence form (called Fc*) that ablates binding to Protein A via the constant region. As a result of co-expression of the 2 heavy chains and the common light chain, 3 products are created, 2 of which are homodimeric for the heavy chains and one that is the desired heterodimeric bispecific product. The Fc* sequence allows selective purification of the FcFc* bispecific product on commercially available affinity columns, due to intermediate binding affinity for Protein A compared to the high avidity FcFc heavy chain homodimer, or the weakly binding Fc*Fc* homodimer. This platform requires the use of Protein A chromatography in both a capture and polishing modality. Several challenges, including variable region Protein A binding, resin selection, selective elution optimization, and impacts upon subsequent non-affinity downstream unit operations, were addressed to create a robust and selective manufacturing process.