Selecting Peptides Potentially Useful in Breast Cancer Treatment
Suarez, E.R.1; Bonaldi, C.M.2; Spatari, M.V.B.2; Nohara, A.S.2; Theodoro, T.R.1; Takahama, P.H.1; Del Giglio, A.3; Pinhal, M.A.S.1
1Departamento de Bioquímica e Biologia Molecular, FMABC, SP; 2Departamento de Bioquímica, UNIFESP, SP; 3Departamento de Oncologia e Hematologia, FMABC, SP.
ErbB2 (also known as HER-2 or Neu) is a member of the epidermal growth factor receptor, known as HER family of receptor tyrosine kinase. HER receptors are essential mediators of cell proliferation and differentiation in the developing embryo and adult tissues. In patients with breast cancer, ErbB2 overexpression is an independent predictor of survival. It is associated with poor prognosis, aggressive disease, resistance to chemotherapy and hormone therapy. The critical role of ErbB2 in epithelial oncogenesis as well as its selective overexpression on malignant tissues makes it an ideal target for immunotherapy. Trastuzumab, a humanized antibody to ErbB2, has been shown to be effective in the treatment of breast cancer in which ErbB2 overexpression and metastasis occurs. In our studies we search a mimic peptide to trastuzumab. In order to select specific peptides that binds with ErbB2, a cyclic 7 amino acid phage display random peptide library was constructed using the fUSE5 gene III system. The library was panned against MCF-7 cells (human breast cancer cell line), and also against an external domain of ErbB2 (R&D System®). Specific peptides were selected by dislodgment binding assay using trastuzumab solution. After five rounds of panning assays it was identified several ErbB2 binding phage clones that had been selected, sequenced and analyzed by ClustalW program (European Bioinformatics Institute, Cambridge, UK), using matrix ID. The results showed that selected peptides displayed mainly a repeat motif (BXBX) where B represents a basic amino acid and X a non charged amino acid. This specific motif BXBX suggests that such peptides might bind to ErbB2 molecules present at the surface of MCF-7 cells and also at the recombinant ErbB2 protein. It is important to point out that such specific motif BXBX was not observed at phage clones analyzed by specific binding with trastuzumab or by inespecific binding phages. We are cloning and expressing these selected peptides to check the inhibitory activity of these peptides in cell proliferation and adhesion assays, since we had already observed that the monoclonal antibody trastuzumab had an inhibitory effect on MCF-7 adhesion. Supported by FAPESP, CAPES, CNPq and NEPAS.
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