BACKGROUND: An anti-CD146 monoclonal antibody, AA98, has been identified as an inhibitor of tumor angiogenesis. To overcome the inherent immunogenicity of murine antibody as well as to facilitate immunotoxin construction, a single chain AA98 V(H)/L with three-domain fragments was constructed and expressed in mammalian cells. MATERIALS AND METHODS: The genes of the AA98 heavy chain variable region and the light chain were linked with a modified 12 amino acid sequence that was derived from the heavy chain C(H)1 region, thus constituting the three-domain antibody V(H)/L. Soluble AA98 V(H)/L was produced by mammalian cells and purified by affinity chromatography. The specificity of AA98 V(H)/L for the CD146 molecule was detected by ELISA, immunofluorescence staining and flow cytometry. RESULTS: AA98 V(H)/L alone showed anti-angiogenic properties in a chicken chorioallantoic membrane (CAM) assay as the parent mAb AA98 did. CONCLUSION: This newly generated AA98 V(H)/L antibody displays a therapeutic potential for tumor and other angiogenesis disorders, as well as providing a new strategy for antibody engineering for clinical applications.
BACKGROUND: An anti-CD146 monoclonal antibody, AA98, has been identified as an inhibitor of tumor angiogenesis. To overcome the inherent immunogenicity of murine antibody as well as to facilitate immunotoxin construction, a single chain AA98 V(H)/L with three-domain fragments was constructed and expressed in mammalian cells. MATERIALS AND METHODS: The genes of the AA98 heavy chain variable region and the light chain were linked with a modified 12 amino acid sequence that was derived from the heavy chain C(H)1 region, thus constituting the three-domain antibody V(H)/L. Soluble AA98 V(H)/L was produced by mammalian cells and purified by affinity chromatography. The specificity of AA98 V(H)/L for the CD146 molecule was detected by ELISA, immunofluorescence staining and flow cytometry. RESULTS: AA98 V(H)/L alone showed anti-angiogenic properties in a chicken chorioallantoic membrane (CAM) assay as the parent mAb AA98 did. CONCLUSION: This newly generated AA98 V(H)/L antibody displays a therapeutic potential for tumor and other angiogenesis disorders, as well as providing a new strategy for antibody engineering for clinical applications.
Authors: Allal Ouhtit; Mohammed E Abdraboh; Andrew D Hollenbach; Hatem Zayed; Madhwa H G Raj Journal: Cell Commun Signal Date: 2017-11-09 Impact factor: 5.712