Bcl-2 phosphorylation confers resistance on chronic lymphocytic leukaemia cells to the BH3 mimetics ABT-737, ABT-263 and ABT-199 by impeding direct binding
Bcl-2 phosphorylation confers resistance on chronic lymphocytic leukaemia cells to the BH3 mimetics ABT-737, ABT-263 and ABT-199 by impeding direct binding
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Bcl-2 磷酸化通过阻止直接结合赋予慢性淋巴细胞白血病细胞对 BH3 模拟物 ABT-737、ABT-263 和 ABT-199 的抗性
DOI：
10.1111/bph.13370
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发表时间：
2016-02-01
影响因子：
7.3
通讯作者：
Zhang, Zhichao
中科院分区：
文献类型：
--
作者：
Song, Ting;Chai, Gaobo;Zhang, Zhichao
背景和目的尽管ABT-263和ABT-199在慢性淋巴细胞白血病（CLL）中正在进行的临床试验表明BH 3模拟物具有相当大的希望，但了解CLL对BH 3模拟物耐药的机制仍然是一个挑战。实验方法ABT-737、ABT-263和ABT-199的LD 50值在来自40名患者的许多原代CLL细胞中进行了测定。Bcl-2家族蛋白，包括磷酸化Bcl-2（pBcl-2）和它们之间的相互作用的水平通过免疫印迹和免疫共沉淀法测定。通过等温滴定量热法和ELISA进行体外结合测定。BH 3在分离的线粒体分析analysed.Key ResultsThe（Mcl-1 + pBcl-2）Bcl-2表达的比例提供了最显着的预测标记的细胞毒性潜力的ABT-737，ABT-263和ABT-199在面板的CLL样品。在机制上，pBcl-2抑制ABT化合物对Bax和Bim从Bcl-2的置换的作用，从而抑制线粒体凋亡。ABT化合物对Bcl-2的谷氨酸、磷酸模拟物、突变体（T69 E、S70 E和S87 E; EEE-Bcl-2）表现出低100-300倍的结合亲和力。BH 3肽与全长WT-Bcl-2和全长EEE-Bcl-2的结合亲和力顺序不同。结论与意义我们的研究表明BH 3结合沟的结构改变是由Bcl-2的磷酸化引起的。我们的数据还提供了一个框架，以克服CLL细胞的耐药性ABT化合物结合pBcl-2激酶抑制剂与ABT化合物。
Background and PurposeAlthough the ongoing clinical trials of ABT-263 and ABT-199 in chronic lymphocytic leukaemia (CLL) have indicated that BH3 mimetics hold considerable promise, understanding the mechanism of CLL resistance to BH3 mimetics remains a challenge.Experimental ApproachThe LD50 values of ABT-737, ABT-263 and ABT-199 in a number of primary CLL cells from 40 patients, were determined. The levels of Bcl-2 family proteins, including phosphorylated Bcl-2 (pBcl-2) and their interactions were measured by immunoblotting and co-immunoprecipitation. In vitro binding assays were performed by isothermal titration calorimetry and ELISA. BH3 profiling in isolated mitochondria was analysed.Key ResultsThe ratio of (Mcl-1 + pBcl-2) to Bcl-2 expression provided the most significant predictive marker for the cytotoxic potential of ABT-737, ABT-263 and ABT-199 in the panel of CLL samples. Mechanistically, pBcl-2 inhibited the effects of the ABT compounds on the displacement of Bax and Bim from Bcl-2, thereby suppressing mitochondrial apoptosis. The ABT compounds exhibited 100-300-fold lower binding affinity to the glutamic acid, phosphomimetic, mutant of Bcl-2 (T69E, S70E and S87E; EEE-Bcl-2). BH3 peptides exhibited different rank orders of binding affinities to full-length WT-Bcl-2 and full-length EEE-Bcl-2.Conclusions and ImplicationsOur study suggested that a structural alteration in the BH3-binding groove was induced by phosphorylation of Bcl-2. Our data also provided a framework to overcome resistance of CLL cells to the ABT compounds by combining pBcl-2 kinase inhibitors with the ABT compounds.