High-contrast tumor imaging via a de novo designed aminopeptidase N targeted fluorogenic probe

Cancer, a leading cause of death worldwide, still needs surgery as essential curation in over 80% population. However, complete resection of cancerous tissues remains a great challenge due to the difficulty in tumor margin identification. Fluorescent probe based imaged-guided surgery (IGS) for in-situ local disease focus imaging is thought to be an ideal method for intraoperation tumor identification due to its high spatiotemporal resolution and real-time manner. By targeting tumor microenvironments (TME) or tumor-associated biomolecules, fluorescent probes could "light-up" tumor tissues and navigate surgical operation. Among which, biosensors targeting aminopeptidase N (APN) have attracted intense attention due to its high relevance with progression of tumor. However, most of the probes adopting its natural substrate (L-Alaine) as the recognition unit may be vulnerable toward other widely distributed aminopeptidase such as leukotriene A4 hydrolase (LTA4H) and leucine aminopeptidase (LAP). In order to acquire high specific APN targeted probe for accurate detection of APN activity in biological samples and fulfill high contrast tumor imaging, herein a non-natural amino acid (P-Chloro-L-Phenylalanine, PCPA) based fluorescent probe was firstly reported for APN targeted bioimaging. The acquired named as TMN-PCPA showed high affinity (Km = 3.27 ± 0.23 μM) and dramatically improved selectivity toward APN than previously reported APN probe. TMN-PCPA was successfully applicated into discrimination tumor bio-samples from normal bio-samples in both cellular and ex vivo level with significantly increased tumor to normal (T/N) ratio, thus showed great potential in intraoperation application such imaged-guided surgery and providing structural basis to high specific APN targeted biosensor construction.