| Title | Engineering and characterization of a humanized antibody targeting TNF-alpha and RANKL |
| Authors | Wang, Jun Du, Yuxuan Qian, Hongyan Yu, Haitao Li, Shentao Zhang, Xulong Li, Zhanguo Yuan, Huihui Zhao, Wenming |
| Affiliation | Capital Med Univ, Sch Basic Med Sci, Dept Immunol, Beijing 100069, Peoples R China. Inner Mongolia Med Univ, Sch Basic Med Sci, Dept Pathogeny Biol & Immunol, Hohhot 010110, Inner Mongolia, Peoples R China. Xiamen Univ, Affiliated Hosp 1, Dept Rheumatol & Clin Immunol, Xiamen 361003, Fujian, Peoples R China. Peking Univ, Peoples Hosp, Clin Immunol Ctr, Dept Rheumatol & Immunol, Beijing 100044, Peoples R China. Capital Med Univ, Sch Basic Med Sci, Dept Immunol, 10 Xitoutiao, Beijing 100069, Peoples R China. |
| Keywords | Humanized antibody Tumor necrosis factor-alpha (TNF-alpha) Receptor activator of NF-kappa B ligand (RANKL) Dual targets COLLAGEN-INDUCED ARTHRITIS RHEUMATOID-ARTHRITIS MONOCLONAL-ANTIBODY DISEASE |
| Issue Date | 2014 |
| Publisher | 生物化学与生物物理学研究通讯 |
| Citation | BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS.2014,450,(1),717-722. |
| Abstract | To neutralize the pathological activities of tumor necrosis factor-a (TNF-alpha) and receptor activator of NF-kappa B ligand (RANKL), we engineered and characterized a humanized 8G12 (h8G12) antibody that targeted TNF-alpha and RANKL. Standard molecular biological and complementarity determining region (CDR)-grafting techniques were used to engineer the h8G12 antibody, and enzyme-linked immunosorbent assays (ELISAs) and Western blotting were employed to determine its binding activation and specificity. TNF-alpha-mediated cytotoxicity and RANKL-induced osteoclastogenesis assays were used to evaluate the neutralizing effects of the antibody. The cDNA sequences were established by grafting the murine monoclonal antibody (mAb) 8G12 CDRs into the heavy and light chain (HC and LC) variable regions (VH and VL) of the human mAbs 3DGG_B and 1I9R_L, respectively. The recombinant plasmids were transfected into Chinese hamster ovary (CHO) cells to produce the h8G12 antibody, which could simultaneously recognize TNF-alpha and RANKL. In addition, the h8G12 antibody reduced the TNF-alpha-mediated apoptosis of L929 cells by 25.84%. Furthermore, the h8G12 antibody significantly inhibited leukocyte infiltration in a murine allergic contact inflammation model. Concurrent with the inhibition of apoptosis, the h8G12 antibody significantly reduced the number of osteoclast-like cells in a dose-dependent manner. These results demonstrated that the h8G12 antibody neutralized the activities of TNF-alpha and RANKL and that it might be a potential candidate for the treatment of inflammatory bone diseases, such as rheumatoid arthritis (RA). (C) 2014 Elsevier Inc. All rights reserved. |
| URI | http://hdl.handle.net/20.500.11897/188935 |
| ISSN | 0006-291X |
| DOI | 10.1016/j.bbrc.2014.06.046 |
| Indexed | SCI(E) PubMed |
| Appears in Collections: | 人民医院 |
