Magnetic nanoparticles-based lactate dehydrogenase microreactor as a drug discovery tool for rapid screening inhibitors from natural products
Magnetic nanoparticles-based lactate dehydrogenase microreactor as a drug discovery tool for rapid screening inhibitors from natural products
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基于磁性纳米颗粒的乳酸脱氢酶微反应器作为药物发现工具，用于从天然产物中快速筛选抑制剂
DOI：
10.1016/j.talanta.2019.120554
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发表时间：
2020-03-01
期刊：
影响因子：
6.1
通讯作者：
Song, Fengrui
中科院分区：
文献类型：
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作者：
Cheng, Guorong;Pi, Zifeng;Song, Fengrui
乳酸脱氢酶（LDH）在糖酵解过程中催化丙酮酸转化为乳酸，在癌细胞中过度表达。LDH抑制剂是一种有前景的癌症治疗方法。但到目前为止，快速筛选LDH抑制剂的方法有限。在此，本研究首次报道了使用LDH功能化磁性纳米粒子作为一种药物发现工具，从天然产物中选择性富集LDH潜在抑制剂。首先，通过共价结合将LDH固定在氨基修饰的磁性纳米粒子表面。为了获得最大的酶活性，对包括pH、时间和LDH浓度在内的固定化条件进行了优化。在优化条件下，固定在磁性纳米粒子上的LDH量约为49皮克酶/毫克载体。随后，使用由没食子黄素、绿原酸和毛蕊花糖苷组成的模型混合物进行配体垂钓实验，以验证固定化LDH的特异性和选择性。最后，将固定化LDH方法与超高效液相色谱 - 串联质谱技术（UHPLC - MS/MS）相结合，用于从两种富含蒽醌的天然产物（大黄和虎杖）中筛选潜在的LDH抑制剂。分别从大黄和虎杖提取物中鉴定出9种和6种化合物，其中3种化合物在两者中都有。我们的结果证明，LDH功能化磁性纳米粒子在从复杂基质中发现药物方面具有重要的前景。
Lactate dehydrogenase (LDH), catalyzing the conversion of pyruvate to lactate during glycolysis, is over-expressed in cancer cells. LDH inhibitors are a promising approach for the treatment of cancer. But up till now, there is limited method for rapid screening of LDH inhibitors. Herein, the use of LDH functionalized magnetic nanoparticles as a drug discovery tool for the selective enrichment of LDH potential inhibitors from natural products was firstly reported in this study. Firstly, LDH was immobilized onto the surface of amino-modified magnetic nanoparticles via covalent binding. In order to obtain the maximum enzyme activity, the immobilization conditions including pH, time and LDH concentration were optimized. The amount of LDH immobilized on MNPs was about 49 pg enzyme/mg carrier under the optimized conditions. Subsequently, the ligand fishing assay was performed to validate the specificity and selectivity of immobilized LDH using a model mixture, which consisted of galloflavin, chlorogenic acid and verbascoside. Finally, the immobilized LDH approach combined with ultra-high performance liquid chromatography-tandem mass spectrometry technique (UHPLC-MS/MS) was applied to screen potential LDH inhibitors from two anthraquinone-rich natural products (Rhubarb and Polygonum cuspidatum). Nine and six compounds were identified from Rhubarb and Polygonum cuspidatum extracts respectively, of which three compounds were common to both. Our results have proven that LDH functionalized magnetic nanoparticles have a significant prospect for drug discovery from complex matrices.