Lysine demethylase KDM5 (JARID1) family enzymes are 2-oxoglutarate (2-OG)-dependent hydroxylases that play critical roles in removing the methyl group(s) from histone H3K4Me3. Recently, we described the identification of potent and selective KDM5 inhibitor (compound 1) which was shown to eliminate drug-tolerant persister (DTP) cells in vitro. Excited with the potential to overcome drug resistance in the clinic, we sought to optimize the initial lead, with a particular focus on improving its cellular potency (PC9 H3K4Me3 EC50 = 5.2 uM) and mouse PK profile (unbound plasma Cmax / cell EC50 = 0.4x at 100 mg/kg PO), in order to identify an in vivo “tool” compound that could be studied in animal models. KDM5A co-crystal structure with compound 1 showed the phenyl group binds in a nice hydrophobic pocket and points toward solvent. Our initial SAR focused on different substitutions on the phenyl ring, to better “fill” this pocket and also in hope of picking up interactions with surrounding residues such as Tyr409 and Arg73. This turned out to be an unproductive endeavor, as no improvement in enzyme or cell potency was observed with such modifications. Instead, it was discovered that the phenyl group in 1 can be replaced with a heteroaryl, such as N-methyl pyrazole (compound 2) which led to small improvement in cell potency (EC50 = 2.0 uM). However, 2 showed high clearance in mouse (CLp 60 mL/min/kg). Metasite analysis of compound 2 suggested N-demethylation was the major route of metabolism. To block this N-demethylation pathway, tert-butyl analog compound 3 was prepared, which showed better cell potency (EC50 = 0.34 uM). More importantly, compound 3 displayed low clearance (CLp 5 mL/min/kg) and excellent oral bioavailability in mouse. When dosed in mouse at 100 mg/kg, the molecule achieved unbound plasma Cmax concentrations that were15-fold greater than the corresponding cell EC50 values. Furthermore, compound 3 was found to be highly selective for KDM5, as well as against broad kinase and receptor panels (Invitrogen and Cerep). As a result, compound 3 was selected for target validation in vivo.

Structure-activity-relationship (SAR) optimization of a pyrazolo[1,5-a]pyrimidin-7(4H)-one scaffold that led to potent, selective and cellularly-active KDM5 inhibitors with excellent pharmacokinetic profile suitable for in vivo biological studies

251st National Meeting (2016)

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.