Expansion of the Structure–Activity Relationship Profile of Triaminopyrimidines as Inhibitors of Caspase-1

Caspase-1 is a sought-after therapeutic target for inflammatory conditions due to its role in activation and release of pro-inflammatory cytokines, but there has been little success getting drugs into the clinic. We have previously shown triaminopyrimidines such as CK-1-41 are potent, reversible small molecule inhibitors of caspase-1, likely binding in an allosteric site within the enzyme. A series of analogs of CK-1-41 were synthesized and tested against caspase-1 to develop a more robust structure–activity relationship profile. In general, alkyl and aryl groups were well tolerated via an ethylene or methylene linkage to the piperazine nitrogen, with IC₅₀ values ranging from 13 to 200 nM. The most potent compounds were methylene linked o-tolyl (AE-2-21) and ethylene linked 4-trifluoromethylphenyl (AE-2-48) with IC₅₀ values of 18 and 13 nM, respectively. Derivatives with electrophilic covalent warheads linked via an amide bond to the piperazine nitrogen were synthesized and characterized. CA-1-11 and EM-1-10 were semi-reversible, non-competitive inhibitors of caspase-1 with slightly reduced potencies of 134 and 144 nM, respectively. All derivatives docked well into the allosteric site, supporting our hypothesis that this family of caspase-1 inhibitors function via an allosteric non-competitive mechanism of inhibition.