Masked Quinone Methides: Suicide Inhibitors of Class II HDACs

Histone deacetylases (HDACs) are promising targets of novel cancer therapeutic agents. Previous screening efforts have identified several classes of HDAC inhibitors, most notably of zinc chelator variety (i.e. hydroxamic acids, ortho-amino anilides, etc.). In this research we describe a group of novel class II HDAC suicide inhibitors that were synthesized trough microwave-assisted Petasis and Mannich reactions. We show that the mechanism of action of these compounds involves initial chelation of a Lewis acidic zinc(II) ion, that then catalyzes decomposition of such compounds to reactive quinone methide (QM) intermediates that are subsequently covalently trapped by a nearby nucleophilic thiol side chain of a cysteine residue. Stability of these compounds in the presence of various Lewis acids was tested in the production of QM intermediates. Biochemical assays have been performed to determine the compounds inhibitory activity against HDACs. A selected few have shown IC50 values in the nanomolar range for class II HDACs, all of which share similar structural features. Protein mass spectrometry confirmed the covalent binding of these mechanism-based inhibitors to a specific cysteine residue. Mutagenesis was carried out to confirm the identified cysteine residue targeted by the HDAC inhibitors. Cell-based profiling of HDAC inhibitors was performed and analyzed by immunofluorescence to observe the differential effects of these compounds on histone and tubulin acetylation. In conclusion, we have demonstrated that QM precursors serve as a new class of potential small-molecule mechanism-based HDAC inhibitors that could be used to further study the relationship between epigenetic modifications in chromatin structure and cell behavior.