A novel aryl pyrazole series of arginine methyltransferase inhibitors has been identified. Synthesis of analogues within this series yielded the first potent, selective, small molecule PRMT6 inhibitor tool compound, EPZ020411. PRMT6 overexpression has been reported in several cancer types suggesting that inhibition of PRMT6 activity may have therapeutic utility. Identification of EPZ020411 provides the field with the first small molecule tool compound for target validation studies. EPZ020411 shows good bioavailability following subcutaneous dosing in rats making it a suitable tool for in vivo studies. KEYWORDS: PRMT6, protein methyltransferase, oncology, tool compound PRMT6 is a member of the protein arginine methyltransferase (RMT) family, which comprises 45 enzymes, nine of which are known to catalyze protein arginine N-methylation reactions. These post-translational modifications are important regulators of RNA processing, transcriptional regulation, signal transduction, and other cellular processes. 1,2 PRMT6 is a nuclear-localized RMT capable of creating omega-N(G)-monomethylarginine and asymmetric omega-N(G),N(G)-dimethylarginine derivatives on histone and other protein substrates containing a GAR motif; 3 it is the only RMT known to methylate the H3R2 mark. 4,5 This mark can act in opposition to the activating H3K4me3 mark, effectively acting as a transcriptional repressor. 6 PRMT6 has been reported to play a role in a variety of cellular processes including maintenance of stem cell pluripotency, 7 regulation of cell cycle, 8 DNA repair, 9 regulation of nuclear receptor-mediated transcription, 10 and viral transactivation. 11 PRMT6 overexpression has been reported in several cancer types including melanoma 12 and bladder, lung, 13 and prostate 14 carcinoma, suggesting that inhibition of PRMT6 may have therapeutic utility and supporting development of small molecule inhibitors for use as tool compounds for in vitro and in vivo target validation studies.An aryl pyrazole bearing a diamine side-chain, 1, was found to have potent PRMT1, PRMT6, and PRMT8 inhibitory activity through screening of the Epizyme internal library.A 2.4 Å resolution crystal structure of a ternary complex of 1, SAH, and PRMT6 was obtained and is shown in Figure 1a,b (4Y2H). The diamine side-chain occupies the putative site of the substrate arginine side-chain. The terminal nitrogen atom is 3.4 Å away from the sulfur atom of SAH. The terminal NH 2 group makes multiple direct hydrogen bonds to the Glu155 side-chain and backbone carbonyl and water-mediated hydrogen bond interactions with the backbone carbonyl of Trp156