Investigations on small molecule inhibitors targeting the histone H3K4 tri-methyllysine binding PHD-finger of JmjC histone demethylases

Abstract

Plant homeodomain (PHD) containing proteins are important epigenetic regulators and are of interest as potential drug targets. Inspired by the amiodarone derivatives reported to inhibit the PHD finger 3 of KDM5A (KDM5A(PHD3)), a set of compounds were synthesised. Amiodarone and its derivatives were observed to weakly disrupt the interactions of a histone H3K4me3 peptide with KDM5A(PHD3). Selected amiodarone derivatives inhibited catalysis of KDM5A, but in a PHD-finger independent manner. Amiodarone derivatives also bind to H3K4me3-binding PHD-fingers from the KDM7 subfamily. Further work is required to develop potent and selective PHD finger inhibitors.

Keywords: Epigenetics; Histone demethylases; JmjC-KDMs; PHD-finger inhibitor; Plant Homeodomain.

Graphical abstract

Fig. 1

PHD-finger domains associated with the JmjC-KDMs. A) Phylogenetic tree of the PHD-finger domains in human JmjC-KDM family proteins. Branch lengths are indicated as a cladogram, and recognized histone marks in green. B) Domain architectures of selected JmjC-KDMs with PHD-fingers.

Fig. 2

Design of potential PHD-finger binders from the structure of amiodarone.

Scheme 1

Synthesis of derivatives of 1a.

Scheme 2

Modification of the quaternary ammonium group of potential PHD inhibitors.

Fig. 3

Screening of AMI analogues for binding and catalytic inhibition of KDM5A. (A) AlphaScreen assay for H3K4me3 and His-KDM5A(PHD3) interactions. (B) Normalised dose–response inhibition curves for displacement of H3K4me3-Bn from KDM5A(PHD3) by representative AMI derivatives. Average ± StdDev (N ≥ 3 independent replicates). (C, D) Dose-response inhibition curves of H3K4me3 demethylation activity by AMI derivatives for KDM5A using a MALDI-TOF MS-based assay. AMI derivatives were tested against two active KDM5A constructs, KDM5A_c1 (M1-L801), and KDM5A_c2 (ΔARID/PHD1, L88-G353). KDOAM25a is a small molecule JmjC-domain inhibitor of KDM5.

Fig. 4

AlphaScreen binding assay for H3K4me3 and His-KDM7(PHD). Normalised dose–response inhibition curves for the displacement of H3K4me3-Bn from KDM7A/B/C by representative AMI derivatives.