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

Abstract

Increased activity of the lysine methyltrans-ferase NSD2 driven by translocation and activating mutations is associated with multiple myeloma and acute lymphoblastic leukemia, but no NSD2-targeting chemical probe has been reported to date. Here, we present the first antagonists that block the protein-protein interaction between the N-terminal PWWP domain of NSD2 and H3K36me2. Using virtual screening and experimental validation, we identified the small-molecule antagonist 3f, which binds to the NSD2-PWWP1 domain with a Kd of 3.4 μM and abrogates histone H3K36me2 binding in cells. This study establishes an alternative approach to targeting NSD2 and provides a small-molecule antagonist that can be further optimized into a chemical probe to better understand the cellular function of this protein.

ZMYND8-regulated IRF8 transcription axis is an acute myeloid leukemia dependency

“…Given that PHD and PWWP domains can read methylated amino acids and BDs can recognize acetylated lysine residues, it may be possible that ZMYND8's reader cassette recognizes combinatorial PTMs on the BRD4 ET domain, though those exact PTMs remain unclear and require further investigation. Recent advancements in chemical biology have demonstrated that the discrete reader pocket of the BD and PWWP domains are amenable to chemical perturbation (Clegg et al, 2019;Cochran et al, 2019;Filippakopoulos et al, 2010;de Freitas et al, 2020;Wimalasena et al, 2020). These findings highlight that therapeutic targeting of the reader modules in ZMYND8 can potentially be achieved by selective chemical probes.…”

Pharmacological targeting of a PWWP domain demonstrates cooperative control of NSD2 localization

“…We recently described the use of virtual screening, target class screening, and ligand-based scaffold hopping approaches to identify ligands of the NSD2 PWWP domains as starting points for further development 19 . This initial effort led to compound 3f which binds NSD2-PWWP1 with a K d of 3.4 ± 0.4 µM as determined by surface plasmon resonance (SPR).…”

“…Specifically, we reported a potent chemical probe targeting the N-terminal PWWP domain of NSD3 that repressed MYC mRNA levels and reduced the proliferation of leukemia cell lines 18 . Additionally, we recently described the development of the first antagonist of NSD2-PWWP1 which binds with modest potency and abrogates H3K36me2 binding 19 . Therefore, we hypothesized that targeting the PWWP domain(s) of NSD2 with highly potent and selective chemical probes may be a strategy to modulate NSD2 engagement with chromatin, subcellular localization, and/or catalytic function.…”

“…Our data demonstrate that UNC6934 is a potent and selective drug-like molecule suitable as a high-quality chemical probe to interrogate the function of NSD2-PWWP1. 19 led to MRT866 and finally the chemical probe UNC6934. UNC7145 is a structurally similar negative control compound.…”

“… (a) Domain architecture of the three splicing isoforms, NSD2-long, NSD2-short, and REIIBP (b) Chemical series progression: successive rounds of optimization starting from compound 3f 19 led to MRT866 and finally the chemical probe UNC6934. UNC7145 is a structurally similar negative control compound.…”