SETDB1 Triple Tudor Domain Ligand, ( R,R )-59, Promotes Methylation of Akt1 in Cells.

Increased expression and hyperactivation of the methyltransferase SETDB1 are commonly observed in cancer and central nervous system disorders. However, there are currently no reported SETDB1-specific methyltransferase inhibitors in the literature, suggesting this is a challenging target. Here, we disclose that the previously reported small-molecule ligand for SETDB1's Triple Tudor Domain, ( R,R )-59, is unexpectedly able to increase SETDB1 methyltransferase activity both in vitro and in cells. Specifically, ( R,R )-59 promotes in vitro SETDB1-mediated methylation of lysine 64 of the protein kinase Akt1. Treatment with ( R,R )-59 also increased Akt1 threonine 308 phosphorylation and activation, a known consequence of Akt1 methylation, resulting in stimulated cell proliferation in a dose-dependent manner. ( R,R )-59 is the first SETDB1 small-molecule positive activator for the methyltransferase activity of this protein. Mechanism of action studies show that full-length SETDB1 is required for significant in vitro methylation of an Akt1-K64 peptide, and that this activity is stimulated by ( R,R )-59 primarily through an increase in catalytic activity rather than a change in SAM binding.