The use of cellular thermal shift assay (CETSA) to study Crizotinib resistance in ALK-expressing human cancers.
The use of cellular thermal shift assay (CETSA) to study Crizotinib resistance in ALK-expressing human cancers.
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DOI：
10.1038/srep33710
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发表时间：
2016-09-19
影响因子：
4.6
通讯作者：
Lai R
中科院分区：
文献类型：
--
作者：
Alshareef A;Zhang HF;Huang YH;Wu C;Zhang JD;Wang P;El-Sehemy A;Fares M;Lai R
已在各种类型的人类癌症中鉴定出各种形式的致癌ALK蛋白。虽然已发现克唑替尼（一种ALK抑制剂）对ALK+肿瘤的一个亚组具有治疗作用，但已充分认识到对该药物的临床耐药性，并且尚未完全了解该现象的机制。使用细胞热位移试验（CETSA），我们测量了一组ALK+细胞系中的克唑替尼-ALK结合，并将结果与ALK结构及其与特异性结合蛋白的相互作用相关联。克唑替尼IC 50与克唑替尼-ALK结合显著相关。克唑替尼耐药细胞中的次优克唑替尼-ALK结合不是由于细胞特异性环境，因为将NPM-ALK转染到这些细胞中显示了大量的克唑替尼-NPM-ALK结合。有趣的是，我们发现耐药细胞表达更高蛋白水平的β-连环蛋白，siRNA敲低恢复了克唑替尼-ALK结合（与IC 50显著降低相关）。晶体结构的计算分析表明，β-连环蛋白对克唑替尼-ALK结合产生空间位阻。总之，通过CETSA可测量的克唑替尼-ALK结合可用于预测克唑替尼敏感性，而克唑替尼-ALK结合又取决于ALK及其一些结合伴侣的结构。
Various forms of oncogenic ALK proteins have been identified in various types of human cancers. While Crizotinib, an ALK inhibitor, has been found to be therapeutically useful against a subset of ALK+ tumours, clinical resistance to this drug has been well recognized and the mechanism of this phenomenon is incompletely understood. Using the cellular thermal shift assay (CETSA), we measured the Crizotinib—ALK binding in a panel of ALK+ cell lines, and correlated the findings with the ALK structure and its interactions with specific binding proteins. The Crizotinib IC50 significantly correlated with Crizotinib—ALK binding. The suboptimal Crizotinib—ALK binding in Crizotinib-resistant cells is not due to the cell-specific environment, since transfection of NPM-ALK into these cells revealed substantial Crizotinib—NPM-ALK binding. Interestingly, we found that the resistant cells expressed higher protein level of β-catenin and siRNA knockdown restored Crizotinib—ALK binding (correlated with a significant lowering of IC50). Computational analysis of the crystal structures suggests that β-catenin exerts steric hindrance to the Crizotinib—ALK binding. In conclusion, the Crizotinib—ALK binding measurable by CETSA is useful in predicting Crizotinib sensitivity, and Crizotinib—ALK binding is in turn dictated by the structure of ALK and some of its binding partners.
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影响因子：
4.8
作者：
Hegazy, Samar A.;Alshareef, Abdulraheem;Lai, Raymond
通讯作者：
Lai, Raymond
影响因子：
5.2
作者：
Ceccon, Monica;Mologni, Luca;Gambacorti-Passerini, Carlo
通讯作者：
Gambacorti-Passerini, Carlo
DOI：
10.3324/haematol.2010.027086
发表时间：
2011-02-01
期刊：
HAEMATOLOGICA-THE HEMATOLOGY JOURNAL
影响因子：
--
作者：
Anand, Mona;Lai, Raymond;Gelebart, Pascal
通讯作者：
Gelebart, Pascal
影响因子：
7.3
作者：
Cui, J. Jean;Tran-Dube, Michelle;Edwards, Martin P.
通讯作者：
Edwards, Martin P.
影响因子：
5.7
作者：
Christensen, James G.;Zou, Helen Y.;Los, Gerrit
通讯作者：
Los, Gerrit