Exp Mol Med.
2008 Feb;40(1):35-42. 10.3858/emm.2008.40.1.35.
Characterization of a novel anti-human TNF-alpha murine monoclonal antibody with high binding affinity and neutralizing activity
- Affiliations
-
- 1Biotech Laboratory, Yuhan Research Institute, Yuhan Corportation, Yongin 446-701, Korea.
- 2Department of Molecular Science and Technology, Ajou University, Suwon 443-749, Korea.
- 3Department of Life Science and Biotechnology, Kyung Hee University, Yongin 446-701, Korea. khbaek@khu.ac.kr
- KMID: 1103586
- DOI: http://doi.org/10.3858/emm.2008.40.1.35
Abstract
- In order to develop an anti-human TNF-alpha mAb, mice were immunized with recombinant human TNF-alpha. A murine mAb, TSK114, which showed the highest binding activity for human TNF-alpha was selected and characterized. TSK114 specifically bound to human TNF-alpha without cross-reactivity with the homologous murine TNF-alpha and human TNF-beta TSK114 was found to be of IgG1 isotype with kappa light chain. The nucleotide sequences of the variable regions of TSK114 heavy and light chains were determined and analyzed for the usage of gene families for the variable (V), diversity (D), and joining (J) segments. Kinetic analysis of TSK114 binding to human TNF-alpha by surface plasmon resonance technique revealed a binding affinity (KD) of ~5.3 pM, which is about 1,000- and 100-fold higher than those of clinically relevant infliximab (Remicade) and adalimumab (Humira) mAbs, respectively. TSK114 neutralized human TNF-alpha-mediated cytotoxicity in proportion to the concentration, exhibiting about 4-fold greater efficiency than those of infliximab and adalimumab in WEHI 164 cells used as an in vitro model system. These results suggest that TSK114 has the potential to be developed into a therapeutic TNF-alpha-neutralizing antibody with picomolar affinity.
Keyword
MeSH Terms
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Amino Acid Sequence
Animals
Antibodies, Monoclonal/chemistry/genetics/*immunology
Antibody Affinity/*immunology
Antibody Specificity
Base Sequence
Blotting, Western
Cell Line
Cytotoxicity, Immunologic
Enzyme-Linked Immunosorbent Assay
Humans
Immunoglobulin Variable Region/genetics
Kinetics
Mice
Mice, Inbred BALB C
Molecular Sequence Data
Neutralization Tests
Sequence Analysis, Protein
Tumor Necrosis Factor-alpha/*immunology
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