Design, synthesis, and in vitro testing of α-methylacyl-CoA racemase inhibitors

The enzyme α-methylacyl-CoA racemase (AMACR) is overexpressed in prostate, colon, and other cancers and has been partially validated as a potential therapeutic target by siRNA knockdown of the AMACR gene. Analogs of the natural substrate branched chain α-methylacyl coenzyme A esters, possessing one or more β-fluorine atoms, have been synthesized using Wittig, conjugate addition, and asymmetric aldol reactions and found to be reversible competitive inhibitors. Each diastereomer of the previously reported inhibitor ibuprofenoyl-CoA was also tested. The compounds had K_i values of 0.9-20 μM and are the most potent inhibitors yet known. The presence of β-fluorine on the α-methyl group or the acyl chain results in a significant lowering of the K_i value compared with nonfluorinated analogs, and this is attributed to a lowering of the pK _a of the α-proton, facilitating enolization and binding. Several of the CoA ester inhibitors were formed by incubating the free carboxylic acid precursors with cell free extracts and CoA. α- Trifluoromethyltetradecanoic acid, the precursor to the most potent inhibitor, was shown to inhibit growth of cancer cell lines PC3, CWR22 Rv1, and Du145 in a dose-dependent manner and could be related to the expression level of AMACR.