Xu Mei, Zheng Mengmeng, Liu Guizhen, Zhang Meng, Kang Jingwu
State Key Laboratory of Bio-organic and Natural Products Chemistry, Center for Excellence in Molecular Synthesis, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Lingling Road 345, Shanghai 200032, China.
State Key Laboratory of Bio-organic and Natural Products Chemistry, Center for Excellence in Molecular Synthesis, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Lingling Road 345, Shanghai 200032, China; University of Chinese Academy of Sciences, Yuquan Road 19, Beijing 100049, China.
J Chromatogr A. 2018 Feb 16;1537:128-134. doi: 10.1016/j.chroma.2018.01.019. Epub 2018 Jan 8.
In the present study, a capillary electrophoresis (CE) method was developed for screening of inhibitors against the break point cluster region Abelson tyrosine kinase (BCR-ABL). The screening method was established by using 5-carboxyfluorescein labeled peptide substrate of BCR-ABL (F-ABLS), a known BCR-ABL tyrosine kinase inhibitor dasatinib, as well as a small chemical library consisting of 37 natural products. Thus, the inhibition of BCL-ABL kinase by small inhibitors was assayed by a CE system equipped with the laser induced fluorescence detector. The yield of phosphorylated product could be precisely measured through the separation by CE. The method is competent for enzymatic inhibition assay as well as the measurement of the inhibition kinetics. For screening BCR-ABL tyrosine kinase inhibitors, the hits were readily identified once the peak area of the phosphorylated products was reduced in comparison with the negative control. By taking the advantage of the screening method, luteolin and epicatechin gallate were discovered as the new BCR-ABL inhibitors.
在本研究中，开发了一种毛细管电泳（CE）方法用于筛选针对断裂点簇集区阿贝尔森酪氨酸激酶（BCR-ABL）的抑制剂。该筛选方法是通过使用5-羧基荧光素标记的BCR-ABL肽底物（F-ABLS）、一种已知的BCR-ABL酪氨酸激酶抑制剂达沙替尼以及一个由37种天然产物组成的小型化学文库建立的。因此，通过配备激光诱导荧光检测器的CE系统测定小分子抑制剂对BCL-ABL激酶的抑制作用。通过CE分离可以精确测量磷酸化产物的产量。该方法适用于酶抑制测定以及抑制动力学的测量。对于筛选BCR-ABL酪氨酸激酶抑制剂，一旦与阴性对照相比磷酸化产物的峰面积减小，就很容易鉴定出活性化合物。利用该筛选方法，发现木犀草素和表儿茶素没食子酸酯为新型BCR-ABL抑制剂。
