A new approach to produce IgG4-like bispecific antibodies

• Published in: Scientific reports Vol. 11; no. 1; p. 18630
• Main Authors: Zhao, Caizhi, Zhang, Wei, Gong, Guihua, Xie, Liping, Wang, Ming-Wei, Hu, Youjia
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 20.09.2021; Nature Publishing Group; Nature Portfolio
• Subjects: 631/250; 631/61; Antibodies; Bispecific antibodies; Humanities and Social Sciences; Immunogenicity; Immunoglobulin G; Light chains; Lymphocytes T; multidisciplinary; PD-1 protein; Pembrolizumab; Science; Science (multidisciplinary); T cell receptors; Western blotting
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-021-97393-2

While achieving rapid developments in recent years, bispecific antibodies are still difficult to design and manufacture, due to mispair of both heavy and light chains. Here we report a novel technology to make bispecific molecules. The knob-into-hole method was used to pair two distinct heavy chains as a heterodimer. IgG 4 S228P CH1-CL interface was then partially replaced by T-cell receptor α/β constant domain to increase the efficiency of cognate heavy and light chain pairing. Following expression and purification, the bispecific antibody interface exchange was confirmed by Western blotting and LC–MS/MS. To ensure its validity, we combined a monovalent bispecific antibody against PD-1 (sequence from Pembrolizumab) and LAG3 (sequence from Relatlimab). The results showed that the molecule could be assembled correctly at a ratio of 95% in cells. In vitro functional assay demonstrated that the purified bispecific antibody exhibits an enhanced agonist activity compared to that of the parental antibodies. Low immunogenicity was predicted by an open-access software and ADA test.