Identification and characterization of Ch806 mimotopes

The chimeric antibody 806 (Ch806) is a promising antitumor agent that recognizes both the epidermal growth factor receptor variant III (EGFRvIII) and the overexpressed epidermal growth factor receptor (EGFR) in cancer tissues but does not recognize the wild type EGFR in normal tissues. However, passive antibody immunization could not produce effective antitumor titers unless the immunization was administered repeatedly over long periods. To overcome this limitation, we generated epitope mimics that bind to Ch806 and tested whether the peptide mimics could induce the production of similar antibodies when actively immunizing mice with the peptides. We used the PH.D-12 phage display peptide library to identify peptides that bind to the monoclonal antibody (mAb) 12H23, which also recognizes similar epitopes of Ch806. Two mimotopes (WHTEILKSYPHE and LPAFFVTNQTQD) were shown to mimic the mAb 12H23 and Ch806 epitope using immunoassays. The mimotopes were conjugated to immunogenic carrier proteins and used to intraperitoneally immunize BALB/c mice. Interestingly, sera from the mice immunized with the isolated mimotopes not only recognize the recombinant or synthetic 806 eptitope, but can also recognize EGFR that is overexpressed in A431 cells and EGFRvIII expressed in Huh7-EGFRvIII cells, whereas sera from mice immunized with the control peptide-KLH (keyhole limpet hemocyanin) and carrier KLH alone failed to show a similar reactivity. Furthermore, in an antibody-dependent cellular cytotoxicity assay (ADCC), the mimotope-induced antibodies specifically lysed human Huh-7-EGFRvIII cells. Our data indicate that the isolated mimotopes reported here may potentially be used as new alternative agents for treating cancer with EGFRvIII expression or EGFR overexpression.