ez-ADiCon: A novel glyco-remodeling based strategy that enables preparation of homogenous antibody-drug conjugates via one-step enzymatic transglycosylation with payload-preloaded bi-antennary glycan complexes

• Published in: Bioorganic & medicinal chemistry letters Vol. 80; p. 129117
• Main Authors: Chon, Hyongi, Kanamori, Satoshi, Hibino, Kazuhiro, Nagahara, Takashi, Suzuki, Tomohiko, Ohara, Keiichiro, Narumi, Hideki
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 15.01.2023
• Subjects: Antibodies, Monoclonal - chemistry; Antineoplastic Agents - chemistry; Bi-antennary glycan; Enzymatic transglycosylation; Homogenous ADCs; Hydrophobic and Hydrophilic Interactions; Immunoconjugates - chemistry; Polysaccharides - chemistry; Site-specific; Site-specific; Homogenous ADCs; Bi-antennary glycan; Enzymatic transglycosylation
• ISSN: 0960-894X; 1464-3405
• DOI: 10.1016/j.bmcl.2022.129117

[Display omitted] The conserved N-linked glycan at the Fc domain of recombinant monoclonal antibodies is an attractive target for site-specific payload conjugation for preparation of homogenous antibody-drug conjugates (ADCs). Here, we report a novel ADC constructing strategy, named “ez-ADiCon”, that is achieved by one-step enzymatic transglycosylation of a payload-preloaded bi-antennary glycan oxazoline onto a deglycosylated antibody. In this method, a mixture of different glycoforms of the Fc-glycan is replaced with a pre-defined payload-linked glycan. Since two payloads are linked on each donor glycan substrate, efficient conjugation results in a highly homogenous ADC with mostly-four drug molecules per antibody, facilitating hydrophobic interaction chromatography analysis and purification. We validated this conjugation strategy using Monomethyl auristatin E (MMAE) and an anti-Human epidermal growth factor receptor 2 (anti-Her2) antibody as the model ADC components and demonstrated its target-specific in vitro cytotoxicity. Our novel conjugation strategy, ez-ADiCon, provides a new approach for the preparation of next generation ADCs.