Selection of peptides and synthesis of pentameric peptabody molecules reacting specifically with ERBB-2 receptor

The HER-2/ErbB-2 oncoprotein is overexpressed in human breast and ovarian a denocarcinomas and is clearly associated with the malignant phenotype, Alth ough no specific ligand for this receptor has been positively identified, E rbB-2 was shown to play a central role in a network of interactions with th e related ErbB-1, ErbB-3 and ErbB-4 receptors, We have selected new peptide s binding to ErbB-2 extracellular domain protein (ECD) by screening 2 newly developed constrained and unconstrained random hexapeptide phage libraries . Out of 37 phage clones, which bound specifically to ErbB-2 ECD, we found 6 constrained and 10 linear different hexapeptide sequences. Among the latt er, 5 consensus motifs, all with a common methionine and a positively charg ed residue at positions 1 and 3, respectively, were identified. Furthermore , 3 representative hexapeptides were fused to a coiled-coil pentameric reco mbinant protein to form the so-called peptabodies recently developed in our laboratory. The 3 peptabodies bound specifically to the ErbB-2 ECD, as det ermined by enzyme-linked immunosorbent assay and BIA-core analysis and to t umor cells overexpressing ErbB-2, as shown by flow cytometry, Interestingly , one of the free selected linear peptides and all 3 peptabodies inhibited the proliferation of tumor cells overexpressing ErbB-2, In conclusion, a no vel type of ErbB-2-specific ligand is described that might complement prese ntly available monoclonal antibodies, (C) 2001 Wiley-Liss, Inc.