Rational design and development of a novel and highly specific near-infrared fluorogenic substrate for sensing and imaging of human pancreatic lipase in living systems

Abstract

• A novel activatable near-infrared (NIR) fluorogenic substrate ( DDAO-ol ) for human pancreatic lipase (hPL) was devised. • DDAO-ol is found with the best combination of stability, specificity, sensitivity and reactivity towards hPL. • DDAO-ol offers a practical tool for high-throughput screening and characterizing hPL inhibitors. • DDAO-ol offers a powerful visualization tool for in situ sensing and imaging of PL in living cells, tissues and organs. Human pancreatic lipase (hPL), a crucial digestive enzyme responsible for the digestion of dietary lipids in humans, has been validated as a key target for treating obesity and as a potential biomarker for diagnosis of pancreatic diseases. This study aims to devise a novel and highly specific near-infrared (NIR) fluorogenic substrate for sensing and imaging of human pancreatic lipase (hPL) in biological systems. To achieve this aim, the oleate esters of various NIR fluorophores are rationally designed on the basis of the enzymatic properties and the preferred substrates of hPL. Following virtual screening and experimental validation, DDAO-ol is found with the best combination of stability, specificity, sensitivity and reactivity towards hPL. Under physiological conditions, DDAO-ol can be rapidly hydrolyzed by hPL to release the fluorophore DDAO , which emits strong NIR fluorescence signal around 660 nm. DDAO-ol offers a novel and practical tool to construct the assay for high-throughput screening and characterizing hPL inhibitors, which strongly facilitates the discovery of novel anti-obesity agents targeting hPL. Meanwhile, DDAO-ol is also a powerful visualization tool for sensing and imaging of mammalian pancreatic lipase (PL) in living systems (including living cells, tissues and organs) with low cytotoxicity and high imaging resolution. Collectively, this study reports a novel and highly specific NIR fluorogenic substrate for hPL, which offers a practical and reliable tool for deciphering the biological functions of hPL and the relevance of this key digestive enzyme to diseases.