Discovery of a Dual PRMT5–PRMT7 Inhibitor

Abstract

The protein arginine methyltransferases PRMT7 and PRMT5, respectively, monomethylate and symmetrically dimethylate arginine side-chains of proteins involved in diverse cellular mechanisms, including chromatin-mediated control of gene transcription, splicing, and the RAS to ERK transduction cascade. It is believed that PRMT5 and PRMT7 act in conjunction to methylate their substrates, and genetic deletions support the notion that these enzymes derepress cell proliferation and migration in cancer. Using available structures of PRMT5, we designed DS-437, a PRMT5 inhibitor with an IC 50 value of 6 μM against both PRMT5 and PRMT7 that is inactive against 29 other human protein-, DNA-, and RNA-methyltransferases and inhibits symmetrical dimethylation of PRMT5 substrates in cells. This compound behaves as a cofactor competitor and represents a valid scaffold to interrogate the potential of the PRMT5−PRMT7 axis as a target for therapy.KEYWORDS: PRMT5, PRMT7, inhibitor, methyltransferases P rotein arginine methyltransferases are a class of enzymes that transfer a methyl group from the cofactor Sadenosylmethionine (SAM) onto the arginine omega nitrogens of substrate proteins, including histones, and can be divided into three subclasses based on their product specificity: type I, II, and III PRMTs asymmetrically dimethylate, symmetrically dimethylate, and monomethylate their substrates, respectively. 1Of the nine confirmed human PRMTs, PRMT5 is currently the only known type II PRMT, and PRMT7 the only monomethylase.2,3 PRMT9 is uncharacterized, and other PRMTs are type I enzymes. 1 PRMT5 has been reported to methylate a number of substrates, both nuclear and cytoplasmic: methylation of arginine 3 of histone 4 (H4R3) by PRMT5 generates a binding site for DNMT3A and is associated with gene silencing; 4 PRMT5-mediated methylation of RAF proteins regulates the RAS to ERK signal transduction cascade and promotes cell proliferation; 5 methylation of Sm ribonucleoproteins (Sm) by PRMT5 mediates recruitment of SMN, and deletion of PRMT5 results in aberrant splicing/degradation of MDM4, associated with derepression of p53-mediated apoptosis. 6,7 These results suggest that PRMT5 may represent an attractive cancer target. Further supporting this notion, PRMT5-targeting siRNAs lead to cell-cycle arrest, apoptosis, and loss of cell migratory activity in glioblastoma cell lines, and increase survival in mice xenograft models of glioblastoma. 8Similar to PRMT5, PRMT7 plays a role in the methylation of H3R2 as well as Sm proteins.2,9 PRMT7 only monomethylates arginine side-chains 3 and interacts with PRMT5, suggesting that the two enzymes may function in conjunction to symmetrically dimethylate protein substrates.1 Genetic silencing of PRMT7 reduces symmetrical dimethylation of H4R3 (H4R3me2s), derepresses E-cadherin expression, and attenuates cell migration and invasion in breast cancer cells. 10 Potent, selective, and cell-active chemical probes targeting PRMT5 and PRMT7 would be useful tools to investigate molecular pathways, ...

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