The central nervous system responds to pathological processes with neuroinflammation – a cellular immune response that protects the brain from pathogens but, when chronic, exacerbates pathological conditions such as Parkinson’s disease, Alzheimer’s disease, multiple sclerosis, and other neurodegenerative disorders and inflammatory diseases. One of the key triggers of neuroinflammation are inflammasomes, which are sensors of the innate immune response. The NLRP3 inflammasome recognizes pathogen- and damage-associated molecular patterns, inducing its assembly, which in turn leads to activation of caspase-1. Caspase-1 subsequently activates the pro-inflammatory cytokines IL-1β and IL-18 and, via gasdermin D (GSDMD), initiates pyroptosis. Excessive activity of the NLRP3 inflammasome and caspase-1 contributes to the progression of neurodegeneration. Numerous studies have shown that inhibition of caspase-1 is a valid therapeutic strategy for managing neurodegenerative and inflammatory diseases. Covalent inhibitors containing electrophilic warheads capable of binding the catalytic cysteine in the caspase-1’s active site are a particularly promising lead compounds for further exploration.
Based on the previously disclosed caspase-1 inhibitor, indolin-1-carbonyl fluoride, we successfully synthesized 30 compounds by replacing fluoride as the leaving group with other leaving groups, substituting the entire carbamoyl fluoride with alternative electrophilic warheads, and preparing the sulfur analogue of the carbamoyl fluoride. We determined the inhibitory potency of the compounds using in vitro biochemical assay against recombinant human caspase-1. The results showed that most compounds do not inhibit caspase-1; only the sulfur analogue of the parent indolin-1-carbonyl fluoride, compound 3, potently inhibited caspase-1, while derivative 17 bearing 1,3,4-thiadiazol-2-thiolate as the leaving group showed weaker inhibition. Further optimization of these two compounds would be needed to improve inhibitory potency and selectivity relative to other caspases. Two derivatives that likely appeared as false-positive caspase-1 inhibitors due to spectral interference should be evaluated in future studies using an orthogonal assay that is not susceptible to fluorescence interference.
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