Virtual High-Throughput Screening To Identify Novel Activin Antagonists.
Virtual High-Throughput Screening To Identify Novel Activin Antagonists.
复制标题
虚拟高通量筛选以鉴定新型激活素拮抗剂。
DOI：
10.1021/acs.jmedchem.5b00753
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发表时间：
2015
影响因子：
7.3
通讯作者：
Woodruff,TeresaK
中科院分区：
文献类型：
--
作者：
Zhu,Jie;Mishra,RamaK;Schiltz,GaryE;Makanji,Yogeshwar;Scheidt,KarlA;Mazar,AndrewP;Woodruff,TeresaK
激活素属于 TGFβ 超家族，与多种疾病有关，包括癌症相关的恶病质、早产和骨质疏松症。靶向激活素及其相关信号通路有望成为这些疾病的治疗方法。在两个激活素 βA 亚基之间的界面中发现了一个小分子配体结合凹槽，并用于 ZINC 数据库的虚拟高通量计算机筛选以识别命中。在两种成熟的激活素测定中测试了 39 种没有显着毒性的化合物：FSHβ 转录和 HepG2 细胞凋亡。该筛选工作流程产生了两种先导化合物：NUCC-474 和 NUCC-555。这些潜在的激活素拮抗剂随后被证明可以在离体卵巢培养物中抑制激活素 A 介导的细胞增殖。体内测试表明，我们最有效的化合物 (NUCC-555) 导致卵巢切除小鼠的 FSH 水平呈剂量依赖性下降。 Blitz 竞争结合测定证实了 NUCC-555 与激活素 A:ActRII 的靶结合，从而以剂量依赖性方式破坏激活素 A:ActRII 复合物与 ALK4-ECD-Fc 的结合。与其他 TGFβ 超家族成员肌肉生长抑制素 (GDF8) 相比，NUCC-555 还特异性结合激活素 A。这些数据证明了一种新的基于计算机的策略来识别小分子激活素拮抗剂。我们的方法是第一个鉴定出与 ALK4 结合的激活素的一流小分子拮抗剂，这开辟了抑制 TGFβ 受体超家族成员活性的全新方法。此外，先导化合物可以作为先导化合物优化的起点，以实现对激活素介导的疾病有效的化合物的目标。
Activin belongs to the TGFβ superfamily, which is associated with several disease conditions, including cancer-related cachexia, preterm labor with delivery, and osteoporosis. Targeting activin and its related signaling pathways holds promise as a therapeutic approach to these diseases. A small-molecule ligand-binding groove was identified in the interface between the two activin βA subunits and was used for a virtual high-throughput in silico screening of the ZINC database to identify hits. Thirty-nine compounds without significant toxicity were tested in two well-established activin assays: FSHβ transcription and HepG2 cell apoptosis. This screening workflow resulted in two lead compounds: NUCC-474 and NUCC-555. These potential activin antagonists were then shown to inhibit activin A-mediated cell proliferation in ex vivo ovary cultures. In vivo testing showed that our most potent compound (NUCC-555) caused a dose-dependent decrease in FSH levels in ovariectomized mice. The Blitz competition binding assay confirmed target binding of NUCC-555 to the activin A:ActRII that disrupts the activin A:ActRII complex’s binding with ALK4-ECD-Fc in a dose-dependent manner. The NUCC-555 also specifically binds to activin A compared with other TGFβ superfamily member myostatin (GDF8). These data demonstrate a new in silico-based strategy for identifying small-molecule activin antagonists. Our approach is the first to identify a first-in-class small-molecule antagonist of activin binding to ALK4, which opens a completely new approach to inhibiting the activity of TGFβ receptor superfamily members. in addition, the lead compound can serve as a starting point for lead optimization toward the goal of a compound that may be effective in activin-mediated diseases.