Identification of 1,2,3,4,5,6-hexabromocyclohexane as a small molecule inhibitor of Jak2 tyrosine kinase autophophorylation
Identification of 1,2,3,4,5,6-hexabromocyclohexane as a small molecule inhibitor of Jak2 tyrosine kinase autophophorylation
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DOI：
10.1021/jm049470k
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发表时间：
2005-04-07
影响因子：
7.3
通讯作者：
Sayeski, PP
中科院分区：
文献类型：
--
作者：
Sandberg, EM;Ma, XY;Sayeski, PP
市售的Jak2抑制剂α -氰基-3,4-二羟基- n -苄基肉桂酸（AG490）已被广泛用于研究Jak2激酶的功能。虽然-氰基-3,4-二羟基- n -苄基肉桂酸是一种有效的Jak2抑制剂，但它也可以抑制许多其他激酶信号通路。为了避免这个问题，我们试图鉴定Jak2酪氨酸激酶活性的新型小分子抑制剂。为此，我们构建了Jak2激酶结构域的同源性模型，并在结构表面鉴定了溶剂可达的口袋。使用DOCK程序，我们在硅中测试了6451种已知化学结构的化合物与位于激活环附近的口袋相互作用的能力。我们从美国国家癌症研究所获得了得分最高的7种化合物，并在体外测试了它们抑制Jak2自磷酸化的能力。通过Western blot分析，我们发现其中一种化合物1,2,3,4,5,6-六溴环己烷能够有效且直接地抑制Jak2的自磷酸化。该化合物的表征表明，它以时间和浓度依赖的方式抑制Jak2酪氨酸的自磷酸化。它大大降低了生长激素介导的Jak2自磷酸化，但不阻断表皮生长因子受体的自磷酸化。此外，高达100 μ M的剂量对细胞没有毒性，通过它们排除碘化丙啶的能力来测量。因此，我们相信该化合物可以作为新一代Jak2抑制剂的先导化合物，并且可能有助于阐明Jak2激酶功能的机制。
The commercially available Jak2 inhibitor, alpha-cyano-3,4-dihydroxy-N-benzylcinnamide (AG490), has been used extensively to study Jak2 kinase function. While alpha-cyano-3,4-dihydroxy-N-benzylcinnamide is a potent Jak2 inhibitor, it can inhibit a number of other kinase signaling pathways as well. To circumvent this problem, we sought to identify novel small molecule inhibitors of Jak2 tyrosine kinase activity. For this, we constructed a homology model of the Jak2 kinase domain and identified solvent accessible pockets on the surface of the structure. Using the DOCK program, we tested 6451 compounds of known chemical structure in silico for their ability to interact with a pocket positioned adjacent to the activation loop. We attained the top seven scoring compounds from the National Cancer Institute and tested their ability to inhibit Jak2 autophosphorylation in vitro. Using Western blot analysis, we found that one of the compounds, 1,2,3,4,5,6-hexabromocyclohexane, was able to potently, and directly, inhibit Jak2 autophosphorylation. Characterization of this compound revealed that it inhibits Jak2 tyrosine autophosphorylation in both a time- and concentration-dependent manner. It greatly reduced growth hormone-mediated Jak2 autophosphorylation but did not block autophosphorylation of the epidermal growth factor receptor. Furthermore, doses as high as 100 mu M were not toxic to cells as measured by their ability to exclude propidium iodide. As such, we believe that this compound could serve as a lead compound for a new generation of Jak2 inhibitors and, perhaps, be useful in elucidating the mechanisms of Jak2 kinase function.