Structural basis for CARM1 inhibition by indole and pyrazole inhibitors

Abstract

CARM1 (co-activator-associated arginine methyltransferase 1) is a PRMT (protein arginine N-methyltransferase) family member that catalyses the transfer of methyl groups from SAM (S-adenosylmethionine) to the side chain of specific arginine residues of substrate proteins. This post-translational modification of proteins regulates a variety of transcriptional events and other cellular processes. Moreover, CARM1 is a potential oncological target due to its multiple roles in transcription activation by nuclear hormone receptors and other transcription factors such as p53. Here, we present crystal structures of the CARM1 catalytic domain in complex with cofactors [SAH (S-adenosyl-L-homocysteine) or SNF (sinefungin)] and indole or pyazole inhibitors. Analysis of the structures reveals that the inhibitors bind in the arginine-binding cavity and the surrounding pocket that exists at the interface between the N- and C-terminal domains. In addition, we show using ITC (isothermal titration calorimetry) that the inhibitors bind to the CARM1 catalytic domain only in the presence of the cofactor SAH. Furthermore, sequence differences for select residues that interact with the inhibitors may be responsible for the CARM1 selectivity against PRMT1 and PRMT3. Together, the structural and biophysical information should aid in the design of both potent and specific inhibitors of CARM1.

Modulation of Epigenetic Targets for Anticancer Therapy: Clinicopathological Relevance, Structural Data and Drug Discovery Perspectives

“…Research conducted on new modulating agents of these PRMTs has been documented [72,73]. In particular, CARM1 has been described as a potential oncological target as its interactions with nuclear transcription factors and p53 may represent a new approach for treating cancer [403]. Its crystal structure ( Table 5) has been resolved with a bound indole-based inhibitor, providing new insightful information for the inhibition of this Arg methyltransferase.…”

S-Adenosylmethionine-dependent Protein Methylation Is Required for Expression of Selenoprotein P and Gluconeogenic Enzymes in HepG2 Human Hepatocytes

“…That this relationship may be causal is suggested by findings that insulin signaling in the mouse was impaired by administration of purified SEPP1 but enhanced by genetic deletion or RNA interference-mediated SEPP1 knockdown (20). Circulating concentrations of SEPP1 and other biomarkers of selenium status are related to single-carbon status, plasma total selenium is positively associated with plasma concentrations of vitamin B 12 and folic acid, and plasma levels of SEPP1 are positively associated with plasma homocysteine concentration (21). Plasma SEPP1 concentrations appeared to be diminished in obese subjects (21), many of whom may be expected to have reduced methylation status (10, 11).…”

“…In human subjects, metabolic syndrome reduces levels of SAM, a methyl-donor substrate for numerous methyltransferases (10), whereas non-alcoholic steatohepatitis is characterized by reduced transmethylation through SAM (11). Cellular availability of SAM and SAH determines capacity for protein methylation (12). Methylation by PRMTs increases activity of transcription factors (forkhead box protein O1A, FOXO1A (13), hepatocyte nuclear factor 4␣ (HNF4␣) (14), and peroxisome proliferator-activated receptor ␥ coactivator 1␣ (PPARGC1␣) (15)), which regulate glucose homeostasis as indicated in Scheme 1.…”

Perspectives on the Discovery of Small-Molecule Modulators for Epigenetic Processes

“…43 Crystal structures of these methyltransferases bound to their respective small-molecule inhibitors have been deposited in the Protein Data Bank (PDB). [44][45][46][47] These structures provided insight on how the compounds bind and inhibit catalytic activity (Figure 2). Furthermore, the structures have guided the synthesis of analogs with improved potency.…”

“…46,47 The G9a/GLP inhibitors have reported selectivity versus related lysine methyltransferases such as SUV39H2, SET7/9, and SET8, 44,46 and the CARM1 inhibitors have selectivity versus the structurally related arginine methyltransferases PRMT1 and PRMT3. 47 The potent and specific DOT1L inhibitor EPZ004777, derived from molecular docking of SAM mimetics at the active site of DOT1L, was shown to specifically block cellular H3K79 methylation and selectively kill cells bearing mixed lineage leukemia (MLL) translocations. 48 Chemical probes of HDMs have also been developed to determine biological activity and validate these enzymes as potential therapeutic targets ( Table 1).…”