Thiobarbiturates as sirtuin inhibitors: virtual screening, free-energy calculations, and biological testing.

NAD+-dependent histone deacetylases (sirtuins) are enzymes that cleave acetyl groups from lysine residues in histones and other proteins. Potent selective sirtuin inhibitors are interesting tools for the investigation of the biological functions of these enzymes and may be future drugs for the treatment of cancer or neurodegenerative diseases. Herein we present the results from a protein-based virtual screen of a commercial database with subsequent biological testing of the most promising compounds. The combination of docking and in vitro experimental testing resulted in the identification of novel sirtuin inhibitors with thiobarbiturate structure. To rationalize the experimental results, free-energy calculations were carried out by molecular mechanics Poisson-Boltzmann/surface area (MM-PBSA) calculations. A significant correlation between calculated binding free energies and measured Sirt2 inhibitory activities was observed. The analyses suggested a molecular basis for the interaction of the identified thiobarbiturate derivatives with human Sirt2. Based on the docking and MM-PBSA calculations we synthesized and tested five further thiobarbiturates. The MM-PBSA method correctly predicted the activity of the novel thiobarbiturates. The identified compounds will be used to further explore the therapeutic potential of sirtuin inhibitors.

Inhibitors of NAD+ dependent histone deacetylases (sirtuins).

Histone deacetylases (HDACs) are enzymes that deacetylate acetyl lysines in histones and various non-histone proteins. Three classes of histone deacetylases have been described in humans: class I, II and IV were shown to be zinc dependent amidohydrolases and eleven subtypes are known today (HDAC1-11). Class III enzymes depend in their catalysis on NAD+ with the subsequent formation of nicotinamide and O acetyl-ADP ribose. Based on the homology to the yeast histone deacetylase Sir2p the NAD+-dependent deacetylases have been termed sirtuins and seven members (SIRT1-7) have been described in humans. Whereas class I and II HDACs have been identified as valid anticancer targets and clinical studies of their inhibitors as new anticancer agents are under way much less is known about the consequences of class III histone deacetylase inhibition. Sirtuins have been linked to ageing and overexpression of sirtuins leads to a prolonged lifespan in yeast. Lately, sirtuin activity has been tied to the pathogenesis of HIV, cancer and neurodegenerative disease. In the last two years several reports of new sirtuin inhibitors have emerged. Additionally, sirtuin activators have been identified and have been implicated as potential drugs for the amelioration of metabolic diseases. Thus, the field of sirtuin biology can be investigated with these new tools which will allow in turn to assess the therapeutic potential of those compounds. We will present an overview over sirtuins and their available inhibitors and activators.

Structure-activity studies on splitomicin derivatives as sirtuin inhibitors and computational prediction of binding mode.

NAD (+)-dependent histone deacetylases (sirtuins) are enzymes that cleave acetyl groups from lysines in histones and other proteins. Potent selective sirtuin inhibitors are interesting tools for the investigation of the biological functions of those enzymes and may be future drugs for the treatment of cancer. Splitomicin was among the first two inhibitors that were discovered for yeast sirtuins but showed rather weak inhibition on human enzymes. We present detailed structure-activity relationships on splitomicin derivatives and their inhibition of recombinant Sirt2. To rationalize our experimental results, ligand docking followed by molecular mechanics Poisson-Boltzmann/surface area (MM-PBSA) calculations were carried out. These analyses suggested a molecular basis for the interaction of the beta-phenylsplitomicins with human Sirt2. Protein-based virtual screening resulted in the identification of a novel Sirt2 inhibitor chemotype. Selected inhibitors showed antiproliferative properties and tubulin hyperacetylation in MCF7 breast cancer cells and are promising candidates for further optimization as potential anticancer drugs.