miR-9对鼻咽癌肿瘤干细胞生物学特性的调控作用与机制及靶向干预研究

Recurrence and distant metastasis are the main reasons for the treatment failure of nasopharyngeal carcinoma (NPC); cancer stem cells (CSCs) have a strong resistance to the radiotherapy and chemotherapy, and are the source of tumor recurrence and metastasis. Therefore, finding the therapeutic targets for effectively depleting NPC CSCs are very important to improve the therapeutic effects of NPC. In our previous study, miR-9 overexpression inhibited the proliferation of NPC cells, downregulated the expression of the stem cell-related genes (Oct4, Noang and ABCG2) and inhibited the formation of tumor spheres, and the diameter of tumor spheres in LV-miR-9 group is much smaller than that in vector control group, suggesting that miR-9 suppressed at least self-renewal of NPC CSCs. Firstly, this project plans to comprehensively dissect the effects of miR-9 on self-renewal, apoptosis, differentiation or senescence of NPC CSCs, and further make clear whether the mentioned-above functions of miR-9 are mediated by its target gene Hes1. Secondly, this project plans to explore whether miR-9 can cause c-Myc-Chk1/Chk2 pathway inactivation by down-regulating the expression of its potential target gene MYCBP, followed by inducing radiosensitization of NPC CSCs. Finally, this project plans to explore the in vivo inhibitory effects of miR-9 alone or in combination with radiotherapy on NPC CSCs; the clinical specimens verifies the relationship between miR-9, MYCBP & Hes1 and NPC CSCs, and clinical pathological significance. The expected results will provide new and more effective therapeutic targets for the targeted therapy of NPC CSCs.

复发和远处转移是鼻咽癌治疗失败的主要原因；肿瘤干细胞(CSCs)对放化疗有很强抗性，是肿瘤复发和转移根源。因此寻找有效清除鼻咽癌CSCs的治疗靶点，对提高鼻咽癌治疗效果有重要意义。我们前期研究发现，miR-9过表达抑制鼻咽癌细胞增殖、下调干性基因表达和抑制肿瘤球形成，且过表达组肿瘤球直径小于对照组的，提示miR-9至少抑制了CSCs自我更新。本项目拟进一步全面解析miR-9对鼻咽癌CSCs自我更新、凋亡、分化或衰老等的影响,并明确miR-9上述功能可否由靶基因Hes1介导。其次阐明miR-9可否通过下调潜在靶基因MYCBP,造成c-Myc-CHK1/CHK2通路失活,进而诱导CSCs放射增敏。最后探讨miR-9单药或联合放疗对CSCs体内清除效果；临床标本验证miR-9、Hes1和MYCBP等与鼻咽癌CSCs的关系及临床病理意义。预期成果将为鼻咽癌CSCs靶向治疗提供新的更有效的治疗靶标。

复发和远处转移是鼻咽癌（NPC）治疗失败的主要原因；肿瘤干细胞(CSCs)是肿瘤复发和转移的根源。因此寻找有效清除NPC CSCs的治疗靶点，对提高NPC治疗效果有重要意义。本研究发现，miR-9负向调控NPC细胞体外增殖，同时过表达miR-9显著抑制NPC细胞裸鼠皮下成瘤；miR-9靶向下调其靶基因CCNG1抑制NPC细胞增殖；miR-9过表达显著降低NPC细胞干性；与非癌鼻咽组织相比，Hes1在NPC组织中表达上调，Hes1蛋白表达与临床病理分期关系密切，NPC患者中Hes1高表达组的总体生存率明显低于低表达组；Hes1是miR-9的靶基因；Hes1正向调控NPC细胞体外增殖和干性；miR-9对NPC细胞增殖和干性的调控作用可由Hes1介导；确证PTEN是Hes1的靶基因，Hes1靶向抑制PTEN表达，进而激活PTEN/AKT信号通路，从而增加NPC细胞干性；Hes1促进NPC细胞发生EMT，Hes1正向调控NPC细胞体外迁移与侵袭，Hes1过表达促进CNE2细胞裸鼠体内转移，Hes1通过靶向抑制PTEN激活PTEN/AKT/GSK3/Snail信号通路从而调节EMT相关基因改变，同时Hes1靶向抑制PTEN促进NPC细胞体外迁移与侵袭，我们也发现在高表达Hes1的NPC组织中，E-cadherin低表达，Fibronectin和Snail高表达；移植瘤动物模型结果显示，miR-9高效模拟物 agomir具有很强的体内抑瘤作用，预示其在NPC临床治疗中具有潜在应用价值。总之，本研究揭示了miR-9 对NPC增殖和干性的调控作用与机制，即miR-9靶向下调其靶基因Hes1表达，然后引起Hes1的靶基因PTEN表达上调，进而导致pAKT磷酸化水平下调，从而抑制了NPC细胞的增殖和致瘤性，并降低了干性，同时本研究为NPC CSCs靶向治疗提供了新的治疗靶标——miR-9。课题执行期间，在PLoS Genetics、Oncoimmunology、Laboratory Investigation和Oncotarget等上发表9篇SCI论文，最高影响因子为7.719，另有2篇SCI论文待发表。本课题研究团队在课题执行期间获得2项国家自然科学基金项目资助，均是关于鼻咽癌发病机理研究的。在人才培养方面，共培养博士2人和硕士4人，招收博士后1人，有2人获得副研究员专业技术职务。