Potential of pyrazolooxadiazinone derivatives as serine protease inhibitors

As a part of an investigation on molecular hybrids as new serine protease i nhibitors, the pyrazolo [4,3-c][1,2,5]oxadiazin-3(5H)-one ring system was s elected as a model of potential mechanism-based inhibitors. Due to the inhe rent reactivity of this system an optimal balance between susceptibility to nucleophilic attack and stability in solvents was sought prior to developm ent as therapeutic agents. Substitutions on N5 and C7 of the supporting pyr azole ring with either aliphatic or aromatic groups (compounds 2 a-m) and t he replacement of the carbonyl oxygen on the reactive oxadiazinone ring wit h sulfur (compounds 3a,i) were explored. Two members (2i and 2k) of this cl ass of inhibitors displayed time-dependent inhibition of HLE suggesting mec hanism-based inhibition. The observation that HLE generated a product(s) fr om compound 2i which displayed an identical UV-Visible spectrum to that obs erved during non-enzymatic hydrolysis further supports this proposal. FlexX -based docking of these compounds into a model of the human leukocyte elast ase (HLE) active site produced a molecular model of the inhibitor-enzyme in teraction.