Discovery of Thieno[3,2-d]pyrimidine-6-carboxamides as Potent Inhibitors of SIRT1, SIRT2, and SIRT3

Abstract

The sirtuins SIRT1, SIRT2, and SIRT3 are NAD(+) dependent deacetylases that are considered potential targets for metabolic, inflammatory, oncologic, and neurodegenerative disorders. Encoded library technology (ELT) was used to affinity screen a 1.2 million heterocycle enriched library of DNA encoded small molecules, which identified pan-inhibitors of SIRT1/2/3 with nanomolar potency (e.g., 11c: IC50 = 3.6, 2.7, and 4.0 nM for SIRT1, SIRT2, and SIRT3, respectively). Subsequent SAR studies to improve physiochemical properties identified the potent drug like analogues 28 and 31. Crystallographic studies of 11c, 28, and 31 bound in the SIRT3 active site revealed that the common carboxamide binds in the nicotinamide C-pocket and the aliphatic portions of the inhibitors extend through the substrate channel, explaining the observable SAR. These pan SIRT1/2/3 inhibitors, representing a novel chemotype, are significantly more potent than currently available inhibitors, which makes them valuable tools for sirtuin research.

Carboxamide SIRT1 inhibitors block DBC1 binding via an acetylation-independent mechanism

“…S1A). Both EX-527 and C28 decreased the SIRT1-DBC1 luciferase signal in a dose-dependent manner, with an efficacy that parallels their IC 50 values for SIRT1 inhibition in vitro 36,37 ( Fig. 5C and D).…”

“…37 Titrations of inhibitor spanning several orders in magnitude were performed on cells expressing either SIRT1/DBC1 luciferase-component fusions, or Bach1/Mafk-luciferase component fusions. Bach1 and Mafk have previously been reported to interact in numerous cells types 49 and served as a negative control in these experiments.…”

“…50,51 In this study, we also identify three carboxamide-based molecules with the ability to reduce formation of the SIRT1-DBC1 complex in cells, namely EX-527, SIRT1 inhibitor IV and C28. These molecules have previously been shown to act directly on SIRT1, 36,37 raising the possibility that the effects of these compounds on the SIRT1-DBC1 complex could be due to a direct interaction with SIRT1. The correlation between the dose at which EX-527 and C28 block formation of the SIRT1-DBC1 complex and their respective IC 50 values for SIRT1 in vitro 36,37 supports this possibility.…”

“…33 Conversely, the DBC1-SIRT1 interaction is enhanced during DNA damage and oxidative stress by ATM-mediated phosphorylation of Thr454, which plays an important role in cell fate determination following genotoxic stress. 34 carboxamide-based scaffolds such as EX-527 36,37 interfere with the ability of DBC1 to bind SIRT1 in cells. We demonstrate that the dose at which EX-527 blocks SIRT1-DBC1 binding is comparable to its IC 50 for SIRT1 in vitro, suggesting that the compound acts directly on SIRT1, but that this effect is independent of DBC1 acetylation status.…”

2-Methoxyestradiol Affects Mitochondrial Biogenesis Pathway and Succinate Dehydrogenase Complex Flavoprotein Subunit A in Osteosarcoma Cancer Cells

“…Figure 6 shows the 3D and 2D ligand receptor interactions demonstrated by molecular docking. Interestingly, the crucial interaction regions reported in the literature for known inhibitors (small molecules or peptides) were observed for 2-ME at both the canonical inhibitor and allosteric sites (42)(43)(44)(45)(46) Figure 6B-E.…”