Affinity Improvement And Humanization Of Anti-Her2 Antibody A21 Based On Computational Simulation

Chimeric antibody chA21 is an anti-Her2 engineered human-mouse chimeric antibody, which specifically inhibits proliferation of tumor cells overexpressing Her2. Compared to another clinical anti-Her2 antibody Herceptin, chA21 has several drawbacks in antigen affinity and immunogenicity. To improve clinical potential of chA21, we investigate the affinity improvement and humanization of chA21 basing on computational simulation.We improve affinity of A21 by computational designed single mutation. First, we refine the epitope of A21 by mutagenesis scanning. Then the antigen-contact sites of A21 are identified by computational prediction and site-directed mutation. Basing on these results, we investigate the interaction pattern of A21-Her2 by molecular docking and improve the affinity of A21 by computational design. According to the pattern of antibody-antigen interaction, we propose a theoretical model to explain the relationship between epitope and mechanism of A21.To eliminate the immunogenicity of A21, we study the humanization of A21 variable region by CDR-grafting. Based on the sequence and structure of A21 variable region, a humanized antibody huCC49 is selected as the template for A21 CDR-grafting, then several residues in framework region are replaced for affinity restoration. We design several humanized variants of A21, such as H1, H1-1 and H1-2. Then these humanized variants are espressed in 293T eukaryotic expression system and determined the antigen affinity by ELISA. The results suggest that the humanized variants of A21—H1, H1-1 and H1-2, maintain the affinity of A21 and have better clinical potential.