Discovery of Novel Inhibitors and Fluorescent Probe Targeting NAMPT.

Discovery of Novel Inhibitors and Fluorescent Probe Targeting NAMPT.

Nicotinamide phosphoribosyltransferase (NAMPT) is a promising antitumor target. Novel NAMPT inhibitors with diverse chemotypes are highly desirable for development of antitumor agents. Using high throughput screening system targeting NAMPT on a chemical library of 30000 small-molecules, we found a non-fluorescent compound F671-0003 and a fluorescent compound M049-0244 with excellent in vitro activity (IC50: 85 nM and 170 nM respectively) and anti-proliferative activity against HepG2 cells. These two compounds significantly depleted cellular NAD levels. Exogenous NMN rescued their anti-proliferative activity against HepG2 cells. Structure-activity relationship study proposed a binding mode for NAMPT inhibitor F671-0003 and highlighted the importance of hydrogen bonding, hydrophobic and π-π interactions in inhibitor binding. Imaging study provided the evidence that fluorescent compound M049-0244 (3 μM) significantly stained living HepG2 cells. Cellular fluorescence was further verified to be NAMPT dependent by using RNA interference and NAMPT over expression transgenic mice. Our findings provide novel antitumor lead compounds and a “first-in-class” fluorescent probe for imaging NAMPT.

烟酰胺磷酸核糖转移酶（NAMPT）是一个有前景的抗肿瘤靶点。具有多种化学类型的新型NAMPT抑制剂对于抗肿瘤药物的研发是非常理想的。利用针对化学文库中30000个小分子的NAMPT高通量筛选系统，我们发现了一种非荧光化合物F671 - 0003和一种荧光化合物M049 - 0244，它们具有优异的体外活性（半数抑制浓度分别为85 nM和170 nM）以及对HepG2细胞的抗增殖活性。这两种化合物显著降低了细胞内的NAD水平。外源性NMN挽救了它们对HepG2细胞的抗增殖活性。构效关系研究提出了NAMPT抑制剂F671 - 0003的一种结合模式，并强调了氢键、疏水作用和π - π相互作用在抑制剂结合中的重要性。成像研究提供的证据表明，荧光化合物M049 - 0244（3 μM）可显著对活的HepG2细胞染色。通过使用RNA干扰和NAMPT过表达转基因小鼠，进一步证实细胞荧光依赖于NAMPT。我们的研究结果提供了新型抗肿瘤先导化合物以及一种用于NAMPT成像的“首创”荧光探针。