A Novel Peptide Isolated from a Phage Display Peptide Library with Trastuzumab Can Mimic Antigen Epitope of HER-2

Abstract

Trastuzumab, a humanized antibody to HER-2, has been shown to be effective in the treatment of breast cancer in which HER-2 overexpression and metastasis occurs. In our search for an effective mimic epitope of HER-2 binding with trastuzumab and to develop HER-2 peptide vaccine, we screened a phage display 12-mer peptide library with trastuzumab as the target. A mimetic peptide (mimotope) H98 (LLGPYELWELSH) that could specifically recognize trastuzumab was isolated. The DNA encoding peptide H98 was cloned and expressed as the fusion protein GST-H98 in Escherichia coli BL21. The purified GST-H98 could specifically bind to trastuzumab and block the binding of trastuzumab to HER-2 protein. Moreover, H98 could significantly block the function of trastuzumab inhibiting the growth of cancer cells. Mice that were immunized with GST-H98 made specific antibody to H98 as well as to HER-2. In addition, T-cell proliferation occurred in mice immunized with GST-H98. Although no sequence homology was found between H98 and HER-2, through the use of structure analysis we were able to determine that peptide H98 contributed to a conformational epitope of HER-2. Furthermore, we determined that the last two amino acids at the C terminus, and the third together with the fourth amino acid at the N terminus of peptide H98 are critical to the binding of H98 to trastuzumab. As a result, we conclude that peptide H98 has potential for being developed as a HER-2 vaccine for biotherapy of cancer with HER-2 overexpression.

Novel Engineered Trastuzumab Conformational Epitopes Demonstrate In Vitro and In Vivo Antitumor Properties against HER-2/neu

“…In addition, several HER-2 B cell mimotopes have been identified through phage display (11,52,53). Riemer et al (52) used a constrained 10-mer random peptide phage display library to identify peptide mimotopes to trastuzumab; Abs raised against one of these peptides recognized HER-2/neu and caused internalization of the receptor from the cell surface in a similar manner as trastuzumab (52).…”

“…Although this peptide sequence bears no sequence homology to HER-2, it was matched to the third loop of HER-2 at the HER-2/trastuzumab interface using computational methods (54). Jiang et al (53) identified another mimotope that matched to an epitope between loops 1 and 2 of HER-2 at the HER-2/trastuzumab interface. These studies indicate that all three loops are important for trastuzumab-binding HER-2.…”

Isolation of cross-reacting antigen candidates by mRNA-display using a mixed cDNA library

“…These five amino acids are considered to be the minimal sequence essential for binding to the antibody. Epitope mappings using random peptide display libraries have been reported for several antibodies, and a number of binding peptides for each antibody have been isolated (Stephen et al 1995;Jiang et al 2005;Ja et al 2005;Bimder et al 2006). However, the identification of cross-reacting antigen candidates as annotated polypeptides has not been reported.…”

“…Many protein-display systems with random peptide libraries have been used for epitope mapping (Stephen et al 1995, Jiang et al 2005Ja et al 2005;Bimder et al 2006). However, by using random peptide libraries, it has been difficult to identify cross-reacting antigens as annotated proteins.…”

Filamentous phage as an immunogenic carrier to elicit focused antibody responses against a synthetic peptide

“…The goal of these studies is to use a peptide immunogen, instead of whole antigen (Agn), to produce Abs that cross-react with a single restricted epitope on a protein. For example, peptides are being used to produce Abs against specific epitopes on tumor Agns that decrease cell proliferation [10][11][12][13]. Peptides are also used as alternatives to CHO Agns because they can elicit a T-cell dependent immune response along with CHO-cross-reactive Abs [1][2][3][4][5][6].…”