High-Throughput Screening of DPPIV Inhibitors Antagonizing GLP-1 Degradation Using an Enzymatic Activated Fluorescent Probe.

Dipeptidyl peptidase IV (DPPIV, EC 3.4.14.5) is an exopeptidase widely expressed on various cell surfaces that selectively cleaves N-terminal dipeptides from diverse substrates. In recent years, DPPIV inhibitors have been extensively utilized in the treatment of hepatitis mellitus (DM). In this study, we designed a far-red fluorescent probe, , through molecular docking simulations and by leveraging the functional characteristics of DPPIV. This probe enables rapid, highly selective, and real-time monitoring of DPPIV activity both in vitro and in vivo. Using , we developed a visual high-throughput screening technique for the detection of DPPIV inhibitors. From a library of 4828 compounds, three inhibitors (K784-2660, 6484-0066, and E699-0153) were identified for their strong inhibitory effects on DPPIV. These inhibitors not only suppressed DPPIV activity in the ileum of mice, thereby reducing GLP-1 degradation, but also effectively inhibited DPPIV activity in gut microbiota. The successful application of in visual detection technology highlights its potential for evaluating DPPIV activity and identifying novel DPPIV inhibitors for diabetes mellitus treatment.