Ahmad Syed, St Hilaire Valentine R, Dandepally Srinivasa R, Johnson Gary L, Williams Alfred L, Scott John E
Department of Pharmaceutical Sciences, Biomanufacturing Research Institute and Technology Enterprise (BRITE), North Carolina Central University, Durham, NC, USA.
Department of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, NC, USA; Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, NC, USA.
Biochem Biophys Res Commun. 2018 Jan 29;496(1):205-211. doi: 10.1016/j.bbrc.2018.01.027. Epub 2018 Jan 5.
The kinase MEKK2 (MAP3K2) activates the MEK5/ERK5 cell signaling pathway and may play an important role in tumor growth and metastasis. Thus, MEKK2 may represent a novel kinase target for cancer. In order to identify inhibitors of MEKK2, we screened a library of compounds using a high throughput MEKK2 intrinsic ATPase enzyme assay. We identified two hits with validated structures and confirmed activity in the primary assay (IC values = 322 nM and 7.7 μM) and two orthogonal MEKK2 biochemical assays. Compound 1, the more potent hit, was the subject of further investigation. Limited structure-activity relationship (SAR) studies were performed on this iminocoumarin hit which resulted in ≥20-fold more potent analogs (e.g. 8 and 16 nM IC). Two analogs had improved selectivity in a 50-member kinase profiling panel compared to the hit. These studies suggested that substitutions around the phenoxy ring of this scaffold can impart improved potency and selectivity for MEKK2. Analog Compound 1s (16 nM IC) was further verified by external testing to inhibit MEKK2 and MEKK3 with similar potencies. Compound 1s displayed activity in cell-based assays in which it inhibited ERK5 pathway activation in cells and inhibited cell migration in a scratch assay. Thus, we have identified a scaffold that has promising potential to be developed into a highly selective and potent inhibitor of MEKK2. Information from these SAR studies provides specific guidance for the future design of MEKK2 inhibitor probes.
激酶MEKK2（MAP3K2）激活MEK5/ERK5细胞信号通路，可能在肿瘤生长和转移中发挥重要作用。因此，MEKK2可能是一种新型的癌症激酶靶点。为了鉴定MEKK2的抑制剂，我们使用高通量MEKK2内在ATP酶测定法筛选了一个化合物库。我们在初次测定（IC值=322 nM和7.7 μM）以及两种正交的MEKK2生化测定中鉴定出两种结构经过验证且活性得到确认的命中化合物。化合物1是活性更强的命中化合物，是进一步研究的对象。对这种亚氨基香豆素命中化合物进行了有限的构效关系（SAR）研究，结果得到了活性提高≥20倍的类似物（例如IC值为8和16 nM）。与命中化合物相比，两种类似物在一个包含50种激酶的分析面板中具有更高的选择性。这些研究表明，该支架苯氧环周围的取代可以提高对MEKK2的活性和选择性。类似物化合物1s（IC值为16 nM）经外部测试进一步验证，对MEKK2和MEKK3具有相似的抑制活性。化合物1s在基于细胞的测定中表现出活性，它在细胞中抑制ERK5途径的激活，并在划痕试验中抑制细胞迁移。因此，我们已经鉴定出一种有潜力开发成MEKK2高选择性和强效抑制剂的支架。这些SAR研究的信息为未来MEKK2抑制剂探针的设计提供了具体指导。
