多潜能核心调控因子Nanog在胰腺癌干细胞特性调控中的作用及清胰化积方干预研究

Pluripotency-regulating factor Nanog plays a crucial role in embryonic development. Studies in rescent years have indicated that gene NANOG is damatically affected clinical prognosis of cancers. One-tenth of late-stage pancreatic cancer patients can survive more than 3 years after Qing Yi Hua Ji (QYHJ) formula based on integrated therapy.In addition,the results of animal experiment have revealed that the growth of cancer stem cells as well as Nanog protein expression was suppressed by QYHJ treatment. Therefore,whether QYHJ down-regulates Nanog expression, results in low capability in growth and metastasis and finally contributes to the good outcome of "Survive with tumor"? In this project, it is planned to construct SW1990/Nanog RNAi cells to explore the role of gene Nanog in self-newral, invasion, metastase and chemoresistance of pancreatic cancer stem cells. Then to verify the results of in vitro experiment, subcutaneous xenograft model and liver metastasis model will be established to investigate the different regulative effect of QYHJ on potency of self-newral, differentiation, invasion and metastase of pancreatic cancer stem cells with different expression status of Nanog. The project will explorate the mechanism of "survive with tumor" by Qingyihuaji formula treatment and also provide with new idea and the treatment target for pancreatic cancer by traditional Chinese herbs.

多潜能核心调控因子Nanog在胚胎发育中起关键作用，近年研究提示Nanog表达与肿瘤预后密切相关。已有研究提示，清胰化积方为主治疗的晚期胰腺癌约10%患者获得3年生存；前期实验提示经清胰化积方干预后胰腺癌干细胞生长受抑，同时伴有Nanog蛋白表达下调，提示清胰化积方可能通过下调Nanog表达导致胰腺癌干细胞自我更新、侵袭转移能力下降，从而使胰腺癌患者能够长期"带瘤生存"。本项目拟构建SW1990/Nanog RNAi细胞，明确Nanog基因在胰腺癌干细胞自我更新、分化、侵袭、转移及耐药中的作用；建立裸鼠皮下及肝转移模型，观察清胰化积方对Nanog不同表达程度的胰腺癌干细胞自我更新、分化、侵袭转移能力的影响，探索清胰化积方治疗后带瘤生存的机理，为中医药治疗胰腺癌提供新思路和治疗靶点。

肿瘤干细胞假说认为，肿瘤的发生、发展起源于存在于体内的一小部分肿瘤干细胞（Cancer stem cell， CSC）。非编码RNA是CSC细胞干性维持和诱导肿瘤细胞分化所必须的，CSC相关核转录因子表达高低与肿瘤患者预后有着密切的联系。先前研究提示，肿瘤干细胞的自我更新、多向分化受到长链非编码RNA (Long non-coding RNA， lncRNA)的调控，lncRNAs具有多种生物学功能，在胚胎发生和肿瘤的发生发展中扮演重要的角色，因此lncRNAs不仅调控肿瘤细胞的凋亡等命运，而且在CSC的自我更新和多向分化中发挥着重要的作用。已有研究证实，lncRNA-ROR与多种恶性肿瘤进展有关，但lncRNA-ROR与胰腺癌的关系尚未见报道，值得深入研究。中医药在肿瘤治疗历史悠久且发挥独特作用，具有改善患者生存质量、延长临床病人生存期的作用。但是，客观来看，在临床中存在着肿瘤患者对中医药治疗的疗效反应存在着不同程度的差异的现象。中药复方以多靶标的治疗方式与目前临床的靶向治疗药物有着众多相似之处，lncRNA在恶性肿瘤中发挥着关键作用因而成为目前研究的热点。淸胰化积方(Qingyihuaji formula，QYHJ)作为课题组给予临床疗效总结多年实践的中药复方，内含有数以百计的单体成分，故我们通过指纹图谱技术检测并分析了QYHJ方中存在的有效单体成分，经过前期筛选和实验后发现，芹菜素对胰腺癌干细胞抑制作用最强，故本课题将API作为研究对象，探索该单体成分干预后lncRNA表达差异以及对胰腺癌干细胞的作用，探索药物干预在非编码RNA和CSC层面的内在作用机制。