Small Molecule Inhibitors of the Human Histone Lysine Methyltransferase NSD2 / WHSC1 / MMSET Identified from a Quantitative High-Throughput Screen with Nucleosome Substrate

Abstract

The activity of the histone lysine methyltransferase NSD2 is thought to play a driving role in oncogenesis. Both overexpression of NSD2 and point mutations that increase its catalytic activity are associated with a variety of human cancers. While NSD2 is an attractive therapeutic target, no potent, selective and cell-active inhibitors have been reported to date, possibly due to the challenging nature of developing high-throughput assays for NSD2. To establish a platform for the discovery and development of selective NSD2 inhibitors, multiple assays were optimized and implemented. Quantitative high-throughput screening was performed with full-length wildtype NSD2 and a nucleosome substrate against a diverse collection of known bioactives comprising 16,251 compounds. Actives from the primary screen were further interrogated with orthogonal and counter assays, as well as activity assays with the clinically relevant NSD2 mutants E1099K and T1150A. Five confirmed inhibitors were selected for follow-up, which included a radiolabeled validation assay, surface plasmon resonance studies, methyltransferase profiling, and histone methylation in cells. The identification of NSD2 inhibitors that bind the catalytic SET domain and demonstrate activity in cells validates the workflow, providing a template for identifying selective NSD2 inhibitors.

Target validation and structure-based virtual screening to Discover potential lead molecules against the oncogenic NSD1 histone methyltransferase

“…Recently, NSD1 was reported to promote the migration, invasion or tumorigenesis of gastric cancer cells via WNT10B/βcatenin axis, and therefore validated NSD1 as a promising target for gastric cancer (Li and Han 2021). Therefore, the development of epigenetic inhibitors against NSDs in general and NSD1 in particular is urgently pursued, as it represents a novel therapeutic hope for cancers with poor prognosis (Coussens et al 2017; Morishita et al 2017) in the eld of cancer epigenetics. Recently, covalent inhibition by BT5 lead molecule was developed against NSD1 SET domain demonstrating activity against leukemia cells (Huang et al 2020).…”

Discovery of Small-Molecule Antagonists of the PWWP Domain of NSD2

“…While NSD2 is an attractive therapeutic target, efforts to target the catalytic SET domain with small molecule inhibitors have so far met with little success 6–9 and only recently was the first selective inhibitor of an NSD family protein reported, a compound that binds covalently to the catalytic site of NSD1 10 . Aside from the catalytic domain, NSD2 has multiple protein-protein interaction (PPI) domains that may be clinically relevant, including PHD (plant homeodomain) and PWWP (proline-tryptophan-tryptophan-proline) domains.…”