Identification of Peptides Binding Specifically to Colon Cancer Cells through Phage Display Peptide Library Screening

Objective: Colorectal cancer ranks among the most frequently occurring cancers. Addressing the diagnosis and treatment challenges requires the advancement of alternative approaches that can more precisely differentiate between cancerous and healthy cells. Materials and Methods: We identified colon cancer cell surface specific 12-mer peptides using subtractive phage display technique. Four rounds of biopanning were conducted using the M13 bacteriophage PhD-12 peptide library with the human colon cancer cell line HCT-116. Subtractive selection involved the use of the human rectal cancer cell line SW837 and the healthy CCD112-CoN cell line. Binding levels of selected phage clones were tested by ELISA-HRP binding assays. Experimentally identified peptide sequences were synthesized by solid-phase chemistry. Peptides were labelled at the N-terminus with a fluorescent dye for visualization of cells by confocal microscopy. Results: Peptide COL419 (Serine-Valine-Alanine-Histidine-Leucine-Serine-Proline-Histidine-Serine-Threonine-Threonine-Alanine; SVAHLSPHSTTA) can differentiate colon and rectal cancers from healthy cells. In addition, the COL-AA peptide (Histidine-Tyrosine-Proline-Threonine-Asparagine-Leucine-Histidine-Tyrosine-Proline-Threonine-Asparagine-Leucine; HYPTNLHYPTNL), which was designed from the amino acid composition analysis of the experimentally selected peptide sequences bound to colon cancer cells and neither rectal cancer cells nor healthy colorectal cells. Conclusion: The peptides determined specifically target colon cancer cells. These findings enhance our understanding of the differences between these cancer types. The selected peptides have the potential for clinical application in targeted diagnosis and treatment of colon cancer.