千金子调控肺癌干细胞干性分子的药效物质基础及机制

Seeds of Euphorbia lathyris L. belonging to Euphorbiaceae, named as Caper Euphorbia Seed, have been applied in Traditional Chinese Medicine for treatment of cancer. The compound types include diterpenoid and coumarin. However, the effective substances and the involved mechanism remain unclear. Lung cancer stem cells are important reason leading to failure of lung cancer chemotherapy. Thus,therapy targeting lung cancer stem cells is the available strategy. Our previous study indicated that lathyrane diterpenoid Euphorbia Factor L8 and L1 could affect stemness-associated molecules, including downregulating expression of β-catenin, Bcl-2 and inhibiting function of ABCG2 and ABCB1. On the other hand, active compounds of Caper Euphorbia Seed might contain other types of chemicals, such as coumarins. Generally, transformation mediated by human intestinal bacteria will affect efficacy of Traditional Chinese Medicine. Thus, this project will set up activity index of influencing stemness-associated molecules of lung cancer stem cells. Subsequently, chemical separation will be under guide of activity tracking. Moreover, biotransformation mediated by human intestinal bacteria will be investigated. MTT assay, flow cytometry, western blot, gene transfection, RNA interference, PCR, ATP enzyme activity measurement and nude mouse xenograft model will be applied to investigate the activities and related mechanisms of the effective substances before and after biotransformation. Based on our research results, the structure activity relationship will be summarized. Taken together, this project will help elucidate the effective substances and acting mechanisms of Caper Euphorbia Seed, which will provide new strategy for chemotherapy of lung cancer.

传统中药千金子为大戟科植物续随子Euphorbia lathyris L.的种子，化学成分包括二萜、香豆素等，用于治疗恶性肿瘤，但药效物质、作用机制尚不清楚。肺癌干细胞是肺癌化疗失败的重要原因，靶向肺癌干细胞的治疗是根治肺癌的可行策略。前期研究发现千金子二萜Euphorbia factor L8可以降低肺癌干细胞干性分子β-catenin、Bcl-2的表达，抑制干性分子ABCG2、ABCB1的功能。千金子的活性成分可能还包括香豆素等成分；同时，人肠内生物转化对药效有重要影响。本项目将以调控肺癌干细胞干性分子为活性指标，活性跟踪指导化学成分的分离；研究人肠内菌对活性成分的生物转化。采用流式细胞术、蛋白免疫印迹、基因转染与RNA干扰、ATP酶活性测定、裸鼠移植瘤模型等方法研究原形物及转化产物对干性分子的调节与机制，总结构效关系。本课题将阐明千金子药效物质基础及作用机制，为肺癌化疗提供新思路。

传统中药千金子为大戟科植物续随子Euphorbia lathyris L.的种子，化学成分包括二萜、香豆素等，用于治疗恶性肿瘤，但药效物质、作用机制尚不清楚。肺癌干细胞是肺癌化疗失败的重要原因，靶向肺癌干细胞的治疗是根治肺癌的可行策略。本项目分离得到系列千金子二萜类化合物，鉴定其结构；部分化合物进行了人肠内菌代谢转化研究、Caco-2细胞对千金子二萜的代谢研究。我们的研究发现千金子二萜Euphorbia factor L8 可以降低肺癌干细胞干性分子β-catenin、Bcl-2 的表达，抑制干性分子ABCG2、ABCB1 的功能。千金子二萜Euphorbia factor L2、3-O-苯乙酰基-5, 15-O-二乙酰基续随子醇可以诱导非小细胞肺癌A549细胞发生线粒体途径的凋亡，包括活性氧升高、线粒体膜电位下降、细胞色素c释放、Caspase-9活化、Caspase-3活化等。在本课题资助下，我们还研究了Apicidin、鸦胆子素D等化合物的抗肿瘤作用。本课题初步阐明了千金子药效物质基础及作用机制，为研究抗肺癌新药提供了基础。