Magnetic nanoparticles-based lactate dehydrogenase microreactor as a drug discovery tool for rapid screening inhibitors from natural products

摘要

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.