Identification of a Peptide from In vivo Bacteriophage Display with Homology to EGFL6: A Candidate Tumor Vasculature Ligand in Breast Cancer

BACKGROUND: A crucial step in tumorigenesis is the recruitment of novel vasculature to the site of neoplasia. Currently, a number of high throughput techniques are employed to identify genes, mRNA and proteins that are aberrantly expressed in tumor vasculature. One drawback of such techniques is the lack of functional in vivo data that they provide. Bacteriophage (phage) display has been demonstrated in vivo to select peptides that home to tumors and tumor vasculature. The peptides can be compared to sequences of putative cancer-related proteins, in order to identify novel proteins essential for tumorigenesis. OBJECTIVES: It was hypothesized that an in vivo selection for phage which targeted human breast cancer xenografts could identify peptides with homology to cancer-related proteins for in vivo imaging of breast cancer. METHODS: Following four rounds of in vivo selection in human MDA-MB-435 breast cancer xenografted mice, peptide 3-G03 was discovered with significant homology to a putative secreted protein termed EGFL6. Egfl6 mRNA is upregulated in several transcriptomic analyses of human cancer biopsies, and the protein may play a role in tumor vascularization. RESULTS: Egfl6 mRNA expression was demonstrated in MDA-MB-435 cells and EGFL6 protein was secreted from these cells. Based on homology of 3-G03 to EGFL6, an EGFL6 peptide was synthesized and shown to target MDA-MB-435 cells. EGFL6 peptide was radiolabeled with (111)In and analyzed for biodistribution and tumor imaging capabilities. Single photon emission computed tomography imaging revealed uptake of the peptide in a manner consistent with other tumor vasculature targeting agents.