Probing the methotrexate–protein interactions by proteomics and thermostability assay for drug resistance study,Analytical Methods

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Probing the methotrexate–protein interactions by proteomics and thermostability assay for drug resistance study
Analytical Methods ( IF 2.7 ) Pub Date : 2020-12-7 , DOI: 10.1039/d0ay02099k
Wenbo Zhang _{1,

2,

3,

4,

5} , Xiaoying Li _{1,

2,

3,

4,

5} , Xiaolei Zhang _{1,

2,

3,

4,

5} , Yan Dong _{1,

2,

3,

4,

5} , Lianghai Hu _{1,

2,

3,

4,

5}

Affiliation

Screening of drug targets is critical to understand the mechanism of action of the drug. The aim of this study is to screen the drug-resistant target proteins of the anticancer drug methotrexate (MTX) by using chemical proteomics and to further study the interaction between MTX and its target protein in vitro and in vivo according to the principle of the increasing thermal stability of the target protein after binding with the drug molecule. The results showed that 21 drug resistance related proteins of MTX including phosphoglycerate kinase 1 (PGK1) were detected by quantitative proteomics. The expression of PGK1 increased with the prolongation of incubation time of MTX, indicating PGK1 protein is affected by MTX time dependently in cells. Further the results of the study on the interaction between MTX and PGK1 in vitro and in vivo using cellular thermal shift assay (CETSA) showed that the level of PGK1 in MTX-treated groups was higher than that in the control group under the stimulation of higher temperature conditions, indicating that PGK1 has direct interactions with MTX. The present study provided the data and theoretical support for the study of the resistant target proteins of MTX and a novel point for the extension application of MTX.

中文翻译：

通过蛋白质组学和热稳定性测定法检测甲氨蝶呤与蛋白质的相互作用，以进行耐药性研究

筛选药物靶标对于了解药物的作用机制至关重要。本研究的目的是通过使用化学蛋白组学和进一步研究MTX和其靶蛋白之间的相互作用筛选抗癌药物氨甲蝶呤（MTX）的耐药性靶蛋白在体外和体内根据与靶蛋白结合后增加靶蛋白热稳定性的原理。结果表明，定量蛋白质组学检测到21种MTX耐药蛋白，包括磷酸甘油酸激酶1（PGK1）。PGK1的表达随MTX温育时间的延长而增加，表明PGK1蛋白在细胞中受MTX时间的影响。进一步地，关于MTX和PGK1之间的相互作用的研究的结果，在体外和体内使用细胞热位移分析（CETSA）表明，在较高温度条件下，MTX治疗组中PGK1的水平高于对照组，表明PGK1与MTX有直接相互作用。本研究为MTX的抗性靶蛋白的研究提供了数据和理论依据，为MTX的延伸应用提供了新的思路。

更新日期：2021-01-07

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