An Enterokinase-Activatable Bioluminescent Probe for In Vivo Tumor Imaging

Abstract The detection of enterokinase (ENTK) activity is essential for the investigation of ENTK-associated diseases, including cancer. However, existing methods are often limited by complex procedures or autofluorescence interference. Here, we report a bioluminescent probe, Ac-Asp-Asp-Asp-Asp-Lys-Luc (DDDDK-Luc), for real-time and sensitive imaging of ENTK activity both in vitro and in vivo. Characterized by a high signal-to-noise ratio and low detection limit, the probe exhibited a 59.7-fold bioluminescence enhancement upon ENTK cleavage in vitro with a detection limit of 5.58 U/L. Furthermore, DDDDK-Luc enabled real-time monitoring of endogenous ENTK activity in living HeLa cells and in a subcutaneous tumor xenograft mouse model. We anticipate that this probe holds great potential for applications in biopharmaceutical process control and ENTK-related disease diagnosis.