Title

DESIGN OF A NOVEL PEPTIDE FOR HER2 TARGETING BASED ON PERTUZUMAB-HER2 COMPLEX

Lead Author Affiliation

Thomas J. Long School of Pharmacy and Health Sciences

Second Author Affiliation

Thomas J. Long School of Pharmacy and Health Sciences

Third Author Affiliation

Thomas J. Long School of Pharmacy and Health Sciences

Introduction

Human Epidermal Growth Factor Receptor 2 (HER2) is a cell surface receptor tyrosine kinase and plays a role in the signal pathways leading to cell proliferation and differentiation. Overexpression of HER2 protein is found in about 30% of breast cancers. HER2 has been considered as a potential target for receptor-mediated delivery system of drugs and peptides/antibodies for cancer treatment. Pertuzumab is an antibody that inhibits cell proliferation and differentiation by binding to the extracellular domain of HER2 and has been successfully used in the treatment of breast cancer. Compared to antibodies, peptides that selectively recognize tumor cells have advantages in solubility, stability and immunogenicity. To identify a novel peptide that specifically binds to HER2, we designed a peptide with two chains based on the binding interactions of Pertuzumab-HER2 complex.

Purpose

To design a novel peptide that specifically binds to HER2 based on interactions in Pertuzumab-HER2 complex.

Method

Two key interaction sites between Pertuzumab and HER2 were utilized (residues 30-37 and 50-59 on heavy chains of Pertuzumab) to form a new peptide with two chains. The interactions between the new peptide and HER2 were analyzed by MOE based on London dG scoring function that estimated the free energy of binding between the peptide and domain II and III of HER2. Simulation studies were also performed to map the interactions between the peptide and HER2. The peptide with sequences of Thr-Phe-Thr-Asp-Tyr-Thr-Met-Asp-Trp-Val and Asp-Val-Asn-Pro-Asn- Ser-Gly-Gly-Ser-Ile-Tyr linked by Gly-Lys was designed and synthesized by solid phase synthesis. FITC was conjugated to the peptide through a 6-aminohexanoic acid linker (Ahx). A control peptide with sequence of Gly-Ala-Gly-Ala-Gly-Ala-Gly-Ala-Ahx-FITC was also synthesized. The peptides were lyophilized and purified by using HPLC. The purified peptides were characterized using MALDI-TOF mass spectrometry. Binding studies were performed on HER2 overexpressed MDA-MB-361 and ZR-75-1 cells with HEK293 cell as control. Cells were incubated with newly designed peptide and the control peptide at concentration of 10μM at 37C for 30min. Finally the cells were washed with HBSS and fixed with 4% paraformaldehyde for imaging using a fluorescent microscope.

Results

Residues 30–33 on H1 and 52–58 on H2 of pertuzumab formed an extensive network of hydrogen bonds with main chain and side chain atoms of HER2. Based on these interactions, residues 30-37 and 50-59 on heavy chains of Pertuzumab were selected in designing the novel peptide. In MOE docking analysis, thirty different peptide–HER2 complex conformations were developed with lowest and highest docking energy of -9.2704 and -6.6185 kcal/mol. The peptide conformation with lowest docking energy was selected for binding interaction simulation using MOE. A simulation showed that this peptide bound to the domain II and III of HER2 through interactions with Thr256, Glu258, Asp285, His296 and Lys311 of HER2. In vitro binding studies showed that the peptide specifically bound to the HER2 overexpressed cells but not normal cells, while the control peptide showed no binding to all of three cells.

Significance

This presentation illustrated that a HER2 targeting peptide can be designed by utilizing the key sites of interaction between HER2 and Pertuzumab.

Location

DeRosa University Center, Stockton campus, University of the Pacific

Format

Poster Presentation

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Mar 25th, 10:00 AM Mar 25th, 3:00 PM

DESIGN OF A NOVEL PEPTIDE FOR HER2 TARGETING BASED ON PERTUZUMAB-HER2 COMPLEX

DeRosa University Center, Stockton campus, University of the Pacific

Human Epidermal Growth Factor Receptor 2 (HER2) is a cell surface receptor tyrosine kinase and plays a role in the signal pathways leading to cell proliferation and differentiation. Overexpression of HER2 protein is found in about 30% of breast cancers. HER2 has been considered as a potential target for receptor-mediated delivery system of drugs and peptides/antibodies for cancer treatment. Pertuzumab is an antibody that inhibits cell proliferation and differentiation by binding to the extracellular domain of HER2 and has been successfully used in the treatment of breast cancer. Compared to antibodies, peptides that selectively recognize tumor cells have advantages in solubility, stability and immunogenicity. To identify a novel peptide that specifically binds to HER2, we designed a peptide with two chains based on the binding interactions of Pertuzumab-HER2 complex.