长链非编码RNA-HOTAIR与miR-214-3p的表达调控以及相互作用介导澳洲茄边碱对肺腺癌生长抑制作用的机制研究

Lung cancer is one of the most common malignant tumors in the world. The non-surgical treatment still faces chemotherapy drug resistance, toxicities,low sensitivity and other adverse issues. The large numbers of studies show that solamargine has anti-cancer effect for a wide variety of tumor including lung cancer. Our previous work showed that lncRNA-HOTAIR and miR-214-3p might mediate the effect of solamargine-inhibited lung cancer cell growth. Moreover，SP1 was a direct target of miR-214-3p，overexpression of miR-214-3p upregulated the IGFBP1 protein expression by suppressing the SP1 protein expression，finally，inhibited the cell growth and metastasis of lung cancer. To further clarify the specific molecular mechanism and regulatory network，this project will conduct research from the following two aspects: 1) The use of modern molecular biology techniques, explore the potential molecular mechanisms which solamargine inhibited the cell growth and metastasis in non small cell lung carcinoma (NSCLC)，and illuminate the key role of lncRNA-HOTAIR/miR-214-3p/SP1/IGFBP1 Signaling pathways in solamargine-inhibited cell growth. To explore the effect of lncRNA-HOTAIR on miR-214-3p，and the interaction between lncRNA-HOTAIR and SP1 or IGFBP1 protein. 2) In animal model, to observe the inhibition effect of solamargine on lung cancer tissues with up/down-regulate the expression of the lncRNA-HOTAIR，miR-214-3p and IGFBP1.

肺癌是世界上最常见的恶性肿瘤之一。目前非手术治疗存在化疗药物抵抗，耐药以及敏感性低等问题。研究表明，澳洲茄边碱对包括肺癌在内的多种肿瘤具有抗癌活性。我们前期结果表明，长链非编码RNA-HOTAIR和miR-214-3p可能介导澳洲茄边碱对肺腺癌的生长抑制效应，且miR-214-3p可直接靶向调控核转录因子SP1的蛋白表达，进而刺激IGFBP1的表达和活性，最终抑制肺腺癌的生长和转移。为深入明确其具体的分子机制及调控网络，本项目拟进行：1）体外实验研究HOTAIR/miR-214-3p/SP1/IGFBP1信号通路在澳洲茄边碱抑制肺腺癌生长中的关键作用；探索HOTAIR，miR-214-3p与SP1和IGFBP1等蛋白之间的相互作用及调控机制；2）建立裸鼠肿瘤模型，体内观察澳洲茄边碱对肺腺癌的生长抑制效应，明确HOTAIR，miR-214-3p和IGFBP1等异常表达对肺腺癌生长表型的影响。

肺癌是世界上最常见的恶性肿瘤之一，非手术治疗存在各种各样的问题。急需进一步探索有关导致或促进肺癌发生和发展的分子机制，以便确定新的临床治疗靶点。本研究揭示了澳洲茄边碱可显著抑制非小细胞肺癌的生长、增殖和侵袭，阻滞细胞周期并促进细胞凋亡。深入研究后，我们发现IGFBP1和PDPK1可能是澳洲茄边碱发挥抑癌作用的关键靶分子。以此为出发点，本项目详细阐述了澳洲茄边碱抑制肺癌生长的分子作用机制。一方面，澳洲茄边碱通过抑制长链非编码RNA HOTAIR，同时上调miR-214-3p的表达，HOTAIR和miR-214-3p相互作用并直接调控PDPK1（磷酸肌醇依赖性蛋白激酶1)基因的启动子活性和蛋白水平，最终抑制肺癌的生长。另一方面，Stat3磷酸化蛋白激活后介导澳洲茄边碱抑制转录调控因子Sp1和FOXO3a的蛋白表达。Sp1和FOXO3a分别与IGFBP1（胰岛素样生长因子结合蛋白1）启动子区域结合后，共同调控IGFBP1基因启动子活性和蛋白表达水平。澳洲茄边碱显著上调IGFBP1的蛋白表达，siRNA沉默IGFBP1后有效逆转澳洲茄边碱对肺癌细胞的生长抑制作用，提示IGFBP1可能是该过程的关键靶分子。总之，本项目成功揭示了中药单体澳洲茄边碱抑制肺腺癌细胞生长的新的分子信号通路，即基于Stat3/Sp1-FOXO3a/IGFBP1和lncRNA-HOTAIR/miR-214-3p/PDPK1信号通路。并深入研究靶蛋白之间的‘交叉对话’和整体调控网络。通过体外和体内实验双重验证澳洲茄边碱的抗癌活性和多靶效应，为中药单体的抗肿瘤研究提供了切实可行的研究方案。