A SPR strategy for high-throughput ligand screenings based on synthetic peptides mimicking a selected subdomain of the target protein: A proof of concept on HER2 receptor

The discovery of pharmaceutical agents is a complex, lengthy and costly process, critically depending on the availability of rapid and efficient screening methods. In particular, when targets are large, multidomain proteins, their complexity may affect unfavorably technical feasibility, costs and unambiguity of binding test interpretation. A possible strategy to overcome these problems relies on molecular design of receptor fragments that are: sensible targets for ligand screenings, conformationally stable also as standalone domains, easily synthesized and immobilized on chip for Biacore experiments. An additional desirable feature for new ligands is the ability of selectively targeting alternative conformational states typical of many proteins. To test the feasibility of such approach on a case with potential applicative interest, we developed a surface plasmon resonance (SPR)-based screening method for drug candidates toward HER2, a Tyr-kinase receptor targeted in anticancer therapies. HER2 was mimicked by HER2-DIVMP, a modified fragment of it immobilized onto the sensor surface specifically modeling HER2 domain IV in its bounded form, designed by structural comparison of HER2 alone and in complex with Herceptin, a monoclonal therapeutic anti-HER2 antibody. This design and its implementation in SPR devices was validated by investigating Herceptin-HER2-DIV-MP affinity, measuring its dissociation constant (K-D = 19.2 nM). An efficient synthetic procedure to prepare the HER2-DIVMP peptide was also developed. The HER2-DIVMP conformational stability suggested by experimental and computational results, makes it also a valuable candidate as a mold to design new molecules selectively targeting domain IV of HER2. (C) 2009 Elsevier Ltd. All rights reserved.