Inhibition of Human Sirtuins by in Situ Generation of an Acetylated Lysine-ADP-Ribose Conjugate
Inhibition of Human Sirtuins by in Situ Generation of an Acetylated Lysine-ADP-Ribose Conjugate
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DOI：
10.1021/ja807083y
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发表时间：
2009-05-27
影响因子：
15
通讯作者：
Miyata, Naoki
中科院分区：
文献类型：
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作者：
Asaba, Tomomi;Suzuki, Takayoshi;Miyata, Naoki
根据提出的sirtuin催化乙酰化赖氨酸底物去乙酰化的机理，设计了一种新型的小分子sirtuin抑制剂。在设计和合成的化合物中，我们发现，在乙酰化赖氨酸底物类似物1的乙酰胺的a位上含有乙氧羰基的2k，在重组SIRT1的体外实验中显示出强大的抑制活性，对SIRT2和SIRT3具有高选择性。通过动力学分析、质谱分析和计算等机制研究表明，化合物2k的烯醇形式亲核攻击sirt活性位点的NAD(+)，生成稳定的化合物2k- adp -核糖偶联物5，从而抑制酶活性。化合物2k也引起人结肠癌HCT116细胞中p53乙酰化的剂量依赖性增加，表明细胞中SIRT1受到抑制。这些结果对通过基于机制的药物设计开发选择性sirtuin抑制剂具有启示意义。
A new type of small-molecular sirtuin inhibitor was designed on the basis of the proposed catalytic mechanism for deacetylation of acetylated lysine substrates by sirtuins. Among the compounds thus designed and synthesized, we found that 2k, which contains an ethoxycarbonyl group at the a position to the acetamide of acetylated lysine substrate analogue 1, showed potent inhibitory activity in an in vitro assay using recombinant SIRT1, with high selectivity over SIRT2 and SIRT3. Mechanistic study by means of kinetic analysis, mass spectroscopy, and computation indicated that the enol form of compound 2k nucleophilically attacks NAD(+) in the active site of SIRTs to afford the stable compound 2k-ADP-ribose conjugate 5, leading to inhibition of the enzyme activity. Compound 2k also caused a dose-dependent increase of p53 acetylation in human colon cancer HCT116 cells, indicating inhibition of SIRT1 in the cells. These results have implications for the development of selective sirtuin inhibitors by means of mechanism-based drug design.