Development of a Novel Inhibitory Chimeric Anti-HER2 Monoclonal Antibody.

Abstract

We have recently produced an inhibitory mouse anti-human HER2 mAb (2A8) which displayed potent anti-tumor activity in combination with trastuzumab. To describe chimerization and functional characterization of 2A8 mAb. The VH and VL genes of 2A8 mAb were amplified from cDNA of the mouse hybridoma, ligated to constant regions of human immunoglobulin, and expressed in CHO cell line. Reactivity with four members of human HER family, the inhibitory effects and antibody-dependent cell cytotoxicity (ADCC) of purified chimeric mAb (c2A8) were assessed by ELISA, XTT, H3-tymidine incorporation and lactate dehydrogenase assays. Inhibition of ERK and AKT downstream signaling pathways by the chimeric antibody were analyzed by Western blotting. Chimeric 2A8 mAb bound to recombinant human HER2 and did not cross-react with the other members of HER family. Moreover, c2A8 was able to recognize HER2-overexpressing cancer cell line and inhibited growth and proliferation of these cells. The binding affinity of c2A8 was comparable to the mouse parental mAb. ADCC and Western blotting results showed that the mouse 2A8 mAb was successfully chimerized and could significantly inhibit phosphorylation of AKT in combination with trastuzumab. The c2A8 mAb is potentially a valuable tool for targeted immunotherapy of HER2 positive cancers.