Promising Fusion Protein Design to Target the U87 MG Glioma Cell Line

Abstract

Gliomas, with a poor clinical course, account for 30% to 40% of all intracranial tumors. Immunotherapywith monoclonal antibodies has emerged as a promising area of investigation and recently it has been shown thatantibodies utilize complementarity-determining regions (CDRs) of their variable domains to bind to antigenswith high affinity and specificity. Here, we designed an antibody mimetic fused with diphtheria toxin to targetthe U87 MG glioma cell line. VHCDR1 and VLCDR3, together with 5 amino acid residues on both side of theCDRs, through a cognate framework region (VHFR2) yielded a mimetic of BT32/A6 (United States Patentnumber: 5639863). We fused the mimetic with the first 388 amino acid residues of diphtheria toxin and E. colistrain BL21 (ED3) was used to express the soluble immunotoxin DT-MG. The immunotoxin DT-MG alone didnot kill Raji up to the maximal concentration tested (10-6M) in vitro. By contrast, concentrations ≥10-9M, of thefused DT-MG killed more than 95% of U-87 MG cells. It is suggested that the mimetic maintained the synergicinteractions and high-affinity associated with the parent antibody. This construct holds promise for targetingspecific cancer epitopes and may be useful when incorporated into diagnostic and therapeutic regimens.