Gao Dan, Wang Yini, Xie Weiyi, Yang Ti, Jiang Yuyang, Guo Yuewei, Guan Jin, Liu Hongxia
State Key Laboratory Breeding Base-Shenzhen Key Laboratory of Chemical Biology, Graduate School at Shenzhen, Tsinghua University, Shenzhen 518055, China; Key Laboratory of Metabolomics at Shenzhen, Shenzhen 518055, China.
Department of Chemistry, Tsinghua University, Beijing 100084, China; Key Laboratory of Metabolomics at Shenzhen, Shenzhen 518055, China.
J Chromatogr B Analyt Technol Biomed Life Sci. 2016 Mar 1;1014:17-23. doi: 10.1016/j.jchromb.2016.01.003. Epub 2016 Jan 8.
A marine natural compound flexibilide isolated from the soft coral Sinularia flexibilis has been found to have antitumor activity. However, its pharmacological mechanism on tumor cells has not been studied. Herein, an ultra-performance liquid chromatography coupled to quadrupole time of-flight mass spectrometry (UPLC/Q-TOF MS) based metabolomics approach was established to investigate the antitumor effect of flexibilide on HCT-116 cells and its action mechanism. Q-TOF MS and MS/MS were used to identify significantly different metabolites. Comparing flexibilide-treated HCT-116 cells group with control group (dimethyl sulfoxide), 19 distinct metabolites involved in sphingolipid metabolism, alanine, aspartate and glutamate metabolism, d-glutamine and d-glutamate metabolism, glycerophospholipid metabolism, pyrimidine metabolism and others were discovered and identified. The significant decrease of phosphatidylcholine (PC) and phosphocholine levels and increase of lysophosphatidylcholine (LysoPC) levels in flexibilide treated cells suggested down-regulation of PC biosynthesis pathway. The decrease of sphingolipids reflected the lesions of cell membrane, and the up-regulation of sphingosine-1-phosphate indicated that TRAF2 and caspase-8 were likely to be activated by flexibilide and further caused cell apoptosis. Furthermore, TCA cycle was deemed to be down-regulated after flexibilide treatment, which might lead to an unsustainable of mitochondrial transmembrane potential MMP). The further measured descreased MMP with the increasing concentration of flexibilide treatment indiciated the dysfunction of mitochondrial which might finally lead to apoptosis. The UPLC/Q-TOF MS based metabolomics approach provides new insights into the mechanistic studies of flexibilide on tumor cells, which benefit its further improvement and application.
从软珊瑚柔细角珊瑚中分离出的海洋天然化合物柔扁枝杉烷已被发现具有抗肿瘤活性。然而，其对肿瘤细胞的药理机制尚未得到研究。在此，建立了一种基于超高效液相色谱-四极杆飞行时间质谱联用（UPLC/Q-TOF MS）的代谢组学方法，以研究柔扁枝杉烷对HCT-116细胞的抗肿瘤作用及其作用机制。采用Q-TOF MS和MS/MS鉴定显著差异的代谢物。将柔扁枝杉烷处理的HCT-116细胞组与对照组（二甲基亚砜）进行比较，发现并鉴定了19种参与鞘脂代谢、丙氨酸、天冬氨酸和谷氨酸代谢、D-谷氨酰胺和D-谷氨酸代谢、甘油磷脂代谢、嘧啶代谢等的不同代谢物。柔扁枝杉烷处理的细胞中磷脂酰胆碱（PC）和磷酸胆碱水平显著降低，溶血磷脂酰胆碱（LysoPC）水平升高，提示PC生物合成途径下调。鞘脂的减少反映了细胞膜的损伤，鞘氨醇-1-磷酸的上调表明TRAF2和半胱天冬酶-8可能被柔扁枝杉烷激活，进而导致细胞凋亡。此外，柔扁枝杉烷处理后三羧酸循环被认为下调，这可能导致线粒体跨膜电位（MMP）无法维持。随着柔扁枝杉烷处理浓度的增加，进一步测定的MMP降低表明线粒体功能障碍，最终可能导致细胞凋亡。基于UPLC/Q-TOF MS的代谢组学方法为柔扁枝杉烷对肿瘤细胞的作用机制研究提供了新的见解，有利于其进一步改进和应用。
