Cell affinity screening combined with nanoLC-MS/MS based peptidomics for identifying cancer cell binding peptides from Bufo Bufo gargarizans

Animal venoms contain many peptides with high specificity and selectivity against their protein targets, a characteristic which makes venoms an invaluable source of potential drugs. High-sensitivity mass spectrometry (MS)- based peptidomic platform has evolved as a predominant method for natural peptide drug discovery due to its strength for direct and rapid identification of peptides and peptide-associated post-translational modifications (PTMs). In this study, we used cell-affinity assays combined with nanoLC-MS/MS based peptidomics to identify cancer cell binding peptides (CBPs) from Bufo Bufo gargarizans. We identified 76 potential cell binding peptides and 237 non-affinity peptides in venom extracts from Asiatic toads, and some were verified with MS-parallel reaction monitoring (PRM) mode. These peptides were further analyzed and internalized within human cells and some demonstrated anti-tumor properties in vitro. These specific peptides might be used as templates for peptide-based drug design or optimization.