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Published on: September 30, 2019
Selection of tumor-targeting agents on freshly excised human breast tumors using a phage display library
1Department of Surgery, Vermont Comprehensive Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington, VT 05405, USA. girja.shukla@uvm.edu
Abstract:
The selective delivery of therapeutic agents to tumor site without harming rest of the body is a major challenge in clinical oncology today. Phage display approach has been used for searching ligands for cell-surface macromolecules on cancer cells so that they can be employed as drug targeting agents. Although basic protocols for biopanning cells are available, they are not as such suitable for screening highly complex and diverse target as whole tumor. Present study is an attempt to select peptide ligands specific to whole tumors. The cells from freshly collected human breast tumors were biopanned with phage displayed disulfide-constrained random heptapeptide library, following subtraction on normal human breast cells. Comparative analysis of amino acid frequencies in tumor-selected peptides and in random peptides from unselected library showed that selection was not random. The binding assessment of tumor-selected clones, using highly sensitive chemiluminescence ELISA, demonstrated that 47-75% of selected clones, depending on a tumor, bound to tumor cells they were panned on. Furthermore, several clones bound exclusively or preferentially to tumor cells in comparison to normal breast cells. It was interesting to note that insert sequences of tumor-binding clones from different tumors shared significant motifs. It shows the possibility of identifying ligands that may bind to tumor-specific targets common in certain tumors. The results of this investigation on seven human breast tumors demonstrated that, using procedures developed in the present study, whole tumors can be panned successfully with phage displayed library and tumor-binding ligands can be identified rapidly in high throughput manner. This is an important enabling step in identifying lead molecules for developing novel, specific, and effective agents that can be used for the diagnosis and treatment of cancer.
Insights
Researchers developed a new method to find tumor-specific peptide ligands using phage display. This approach successfully identified ligands that bind to whole tumors, advancing cancer drug targeting and diagnosis.
Area of Science:
- Oncology
- Molecular Biology
- Biotechnology
Background:
- Targeted drug delivery to tumors remains a significant challenge in cancer therapy.
- Phage display is a tool for identifying ligands for cell-surface targets, but screening whole tumors is complex.
Purpose of the Study:
- To develop and validate a method for selecting peptide ligands specific to whole human breast tumors.
- To identify novel tumor-binding ligands for potential use in cancer diagnosis and treatment.
Main Methods:
- Biopanning a phage-displayed random heptapeptide library against whole human breast tumor cells.
- Subtracting normal breast cells to enrich for tumor-specific ligands.
- Assessing clone binding using chemiluminescence ELISA.
Main Results:
- Selection process yielded peptides with non-random amino acid frequencies compared to the unselected library.
- 47-75% of selected clones demonstrated binding to their corresponding tumor cells.
- Several identified clones exhibited exclusive or preferential binding to tumor cells over normal cells.
- Identified tumor-binding clones shared common sequence motifs, suggesting potential common tumor targets.
Conclusions:
- The developed procedure enables successful high-throughput screening of whole tumors using phage display libraries.
- This method rapidly identifies tumor-binding ligands, representing a key step towards developing specific cancer therapeutics and diagnostics.
