The Study On The Biological Character Of The Anti-CD20 Engineered Antibodies

The CD20 antigen is restricted to B-cell. Importantly, it is expressed highly on B-cell malignancy cells. Monoclonal antibodies have provided an alternative approach to B-cell Non-Hodgkin's lymphoma (NHL). On the base of previous study, the current study have evaluated the specifically binding activity, the avidity, the ability to lyse human target cell lines in vitro and in vivo of a novel mouse/human anti-CD20 monoclonal antibody, TGLA (IgG1κ). In addition, with computer-aided molecular modeling design method and DNA recombination technology, two engineered antibodies from anti-CD20 antibody,1-28 (IgM), were developed. Their binding activity was analyzed, and the results shown that Cγ1 is crucial for the binding activity of engineered antibodies of IgG isotype that is from IgM-type antibody.1. Characterization of a novel chimeric anti-CD20 monoclonal antibody, TGLAIn vitro studies, TGLA was found to be very similar to rituximab in terms of antigen-binding specificity and binding avidity. TGLA also bound strongly to CD20+ cells. The chimeric CD20 monoclonal antibody, TGLA could mediate complement-dependent cytotoxicity (CDC), lyse human B-lymphoid cell lines (Daudi and Raji cell lines) through antibody-dependent cell-mediated cytotoxicity (ADCC) and inhibit B-lymphoid cell growth. In vivo the chimeric antibody, TGLA, alone yielded median survival increases of up to 1.7-fold compared with saline-treated controls.2. Designs, construction and characterization of anti-CD20 engineered antibodyBased on the computer-guided homology modeling and distance geometry analysis, the variable heavy (VH) and variable light (VL) chain domains of murine anti-human CD20 mAb 1-28 (IgM), fusing VL-linker-VH with human IgG1 hinge and Fc regions (Named as LH23, VL-Linker-VH-Hγ-Cγ2-Cγ3) or the entire IgG1 heavy constant regions (LH1-3, VL-linker-VH-Cγ1-Hγ-Cγ2-Cγ3), respectively. The two plasmids were constructed and transiently transfected in 293T cells. Western blot showed both of LH23 and LH1-3 were expressed with dimmer formation. Two engineered antibodies could bind specifically to CD20 expressed on Daudi and Raji cells. These results shown that human Cγ1 domain is crucial for the binding activity of engineered antibodies of IgG isotype that is from IgM-type antibody, which were consistent with the theoretical results of computer-aided homology modeling method.