基于CD44/Ras通路探讨益气解毒颗粒联合盐霉素抑制鼻咽癌干细胞的分子机制

One of the biggest problems in cancer treatment is tumor recurrence and metastasis, which may be related with cancer stem cells (CSC).Targeting CSC to find new methods of cancer treatment has important clinical significance. According to the latest CSC research results and preliminary research results of our group, the project intends to carry out at three aspects: (1) Ras signaling pathway characteristics in nasopharyngeal carcinoma(NPC) CSC and the regulation of CD44 on the signaling pathway will be studied deeply by building CD44, HER2 overexpression nasopharyngeal carcinoma cell model, CD44, HER2 gene knockout nasopharyngeal CSC model and its animal model; (2) Effects of Yiqi Jiedu granule combined with salinomycin on NPC CSC proliferation, apoptosis, invasion and metastasis in vivo and in vitro will be observed; (3) Molecular mechanisms of Yiqi Jiedu granule combined with salinomycin will be studied further from the CD44 / Ras signaling pathway by detecting the protein expression levels and activity of CD44, HER2, Ras signaling pathway and their downstream effector proteins. The research can provide theoretical basis for the new targeting drug development of the treatment of nasopharyngeal carcinoma，is also the further clarification of the role of Yiqi Jiedu granule and can lay the foundation for its wide applications. It provides scientific basis for synergies effects of the combination of Chinese and Western medicines, and offers a new way for anticancer.

肿瘤治疗面临的最大瓶颈之一是肿瘤复发和转移，其真正原因可能是肿瘤干细胞（CSC），以CSC为靶点寻找新的肿瘤治疗方法，具有重要临床意义。根据最新CSC研究成果和本课题组前期研究基础，本项目拟进行以下研究：（1）通过构建CD44、HER2高表达鼻咽癌细胞模型，CD44、HER2基因敲除鼻咽癌CSC模型以及其动物模型，深入研究鼻咽癌CSC Ras信号通路特征及CD44对该信号通路的调控作用；（2）观察益气解毒颗粒联合盐霉素对体内外鼻咽癌CSC增殖、凋亡、侵袭与转移的影响；（3）通过检测CD44、HER2、Ras信号通路蛋白及其下游效应蛋白表达水平和活性，从CD44/Ras信号通路深入研究益气解毒颗粒与盐霉素联合作用分子机制。该研究可为鼻咽癌治疗靶点药物开发提供理论依据；为益气解毒颗粒广泛应用奠定基础；为中西医联合用药协同增效提供科学依据；从肿瘤干细胞研究中西医抗癌作用，为肿瘤防治开辟新途径。

肿瘤治疗面临最大瓶颈之一是肿瘤复发和转移，其原因可能是肿瘤干细胞(CSCs)，以 CSCs 为靶点寻找肿瘤治疗新方法具有重要意义。益气解毒颗粒联合盐霉素具有协同效应，其作用机制是什么。项目采用了细胞培养、HE染色、Western-blot、免疫荧光、RNA干扰和流式细胞术等技术分八部分进行了研究，主要结果如下：（1）与普通鼻咽癌细胞比较，鼻咽癌干细胞具有生长增殖速率低、克隆形成率高和克隆小、迁移能力强、粘附率能力强的生物学行为特征；（2）与普通鼻咽癌细胞比较，鼻咽癌干细胞CD44/Ras信号通路异常高表达，从而p-MEK1/2、p-ERK1/2、N-cadherin表达水平显著升高，PTEN、E-cadherin表达水平显著降低；（3）益气解毒方联合盐霉素可协同抑制鼻咽癌干细胞增殖、迁移，诱导凋亡；（4）益气解毒方联合盐霉素可通过调控CD44/Ras信号通路抑制鼻咽癌干细胞增殖、迁移和诱导其凋亡；（5）益气解毒方联合盐霉素能协同抑制鼻咽癌细胞迁移、侵袭和诱导凋亡，能抑制细胞周期相关蛋白的表达；（6）益气解毒方抑制人鼻咽癌CNE-2Z细胞增殖并诱导细胞凋亡和自噬，其诱导细胞自噬与激活PI3K/AKT/mTOR通路有关；益气解毒方诱导的细胞自噬可以促进细胞凋亡；（7）益气解毒方水提物通过MAPK/ERK信号通路抑制鼻咽癌细胞增殖和诱导凋亡；（8）益气解毒方能有效调节小鼠 Treg/Th17 的平衡，恢复Treg 的功能，增强小鼠化疗期间T 细胞的免疫功能，产生抑癌效应，联合顺铂可减轻化疗毒性。提示益气解毒方和盐霉素的联合应用可作为治疗鼻咽癌的方案，其作用机制与调控CD44/Ras信号通路密切相关；提示益气解毒方与顺铂联合应用治疗鼻咽癌也是一种可选择方案。本项目已发表期刊论文23篇，待发表期刊论文2篇，2篇论文撰写中，会议论文4篇；参加国际会议1人次，国内会议23人次,省内会议15人次；会议报告5次；4篇论文获得奖励；培养毕业硕士研究生3名，博士研究生1名，博士后1名。