Abstract
TRIM24 is an epigenetic transcriptional coregulator that "reads" KMe(3) and KAc histone modifications via its tandem plant homeodomain (PHD) and bromodomain (BRD), respectively. The PHD and BRD are potential therapeutic targets due to the roles of TRIM24 in breast cancer progression. However, there are currently no small-molecule ligands for the PHD, and existing TRIM24 BRD inhibitors lack selectivity over the main off-target, BRPF1. Here, we report the development of the first bivalent tool molecules capable of simultaneously engaging both the TRIM24 PHD and BRD. Key to this strategy was the identification of effective KMe(3) bioisosteres that enhance H3 peptide binding to the TRIM24 PHD. The most promising of these was incorporated into a nine amino acid H3-mimicking peptide, and linked to a TRIM24 BRD ligand. The resulting peptide-drug conjugates (PDCs) bind to TRIM24 with picomolar affinity and a slow dissociation rate (k(off)), which is driven by an in cis bivalent binding mode. Although the PDCs showed limited effects on breast cancer cell proliferation in vitro, this work underscores their potential as tools for studying previously unliganded reader domains and consequently advancing our understanding of multivalent epigenetic regulation in disease.