Synthesis of Novel Phosphonic-Type Activity-Based Probes for Neutrophil Serine Proteases and Their Application in Spleen Lysates of Different Organisms

Neutrophils are a type of granulocyte important in the “first line of defense” of the innate immune system. Upon activation, they facilitate the destruction of invading microorganisms by the production of superoxide radicals, as well as the release of the enzymatic contents of their lysozymes. These enzymes include specific serine proteases: cathepsin G, neutrophil elastase, proteinase 3, as well as the recently discovered neutrophil serine protease 4 (NSP4). Under normal conditions, the proteolytic activity of neutrophil proteases is tightly regulated by endogenous serpins; however, this mechanism can be subverted during tissue stress, thereby resulting in the uncontrolled activity of serine proteases, which induce chronic inflammation and subsequent pathology. Herein, we describe the development of low‐molecular‐weight activity‐based probes that specifically target the active sites of neutrophil proteases. Searching out proteases: We describe the development of probes to detect active proteases associated with the innate immune response. Probes incorporating a phosphonate function were able to detect cathepsin G at concentrations below that detectable by conventional antibodies, and thus offers a valuable alternative to traditional methods.