多功能纳米复合物靶向肺癌干细胞调控Wnt/β-catenin信号通路对NSCLC的抑制作用

Lung cancer stem cell (CSC) is closely related with the relapse and metastasis of non-small cell lung cancer (NSCLC). Lung CSCs can be identified and isolated using cell surface marker CD133. In addition, lung CSCs are regulated by Wnt/β-catenin signal pathway. Our preliminary work has demonstrated the suppression of tumorigenesis of NSCLC and inhibition of the self-renewal and colony formation of lung CSCs by miR-34a. Furthermore, we established a nanoparticle complex system to deliver medicines or genes to lung cancer cells in vitro and in vivo with high efficiency. Based on these work, we hypothesize that a nanoparticle complex carrying anti-CD133 and miR-34a mimic can regulate lung cancer stem cells and inhibit the relapse and metastasis of NSCLC by targeting Wnt1 and TCF7, two key components in Wnt/β-catenin signal pathway. In this proposal, by analyses on lung cancer cell lines, lung cancer stem cells, lung cancer clinical samples and lung cancer mice model we will determine the regulatory function and molecular mechanisms of miR-34a-Wnt/β-catenin pathway in regulating lung CSC and NSCLC. In addition, we will test the delivery efficiency and therapeutic effects of the nanoparticle complex to treat lung cancer stem cells by application to animal models. The results from this project will provide strong experimental evidence for targeting therapy of NSCLC and targeting delivery of medicines to lung CSC.

肺癌干细胞（CSC）与非小细胞肺癌（NSCLC）复发转移密切相关，CD133+CSC受Wnt/β-catenin信号通路调控。前期研究证实miR-34a属Wnt/β-catenin负调控分子，并成功构建纳米脂质体向肺癌细胞高效载药递送系统。据此推测纳米-anti-CD133-miR-34a复合物靶向肺CSC，通过调控Wnt/β-catenin信号通路的关键基因Wnt1、TCF7抑制肺CSC的增殖。课题拟构建纳米复合物靶向肺CSC，观察CSC固有Wnt/β-catenin信号通路中的关键分子Wnt1/TCF7的表达变化及肺CSC的更新能力；借助肺癌动物模型，验证纳米复合物载药递送系统对肺CSC的靶向性、杀瘤效率及抑癌效果，并检测肿瘤组织miR-34a-Wnt/β-catenin等分子的表达，阐明纳米复合物调控肺CSC的分子机制，为靶向治疗肺CSC提供方法学借鉴实验依据。

大量研究证实癌干细胞（Cancer Stem Cells, CSCs）是导致癌症复发转移的元凶。我们从CD133+的非小细胞肺癌A549和H1299细胞中筛选出数个表达异常且参与Wnt/β-catenin信号通路的miRNA，如miR-34c、miR-221/222、miR-508等。研究发现miR-221/222临床肺癌患者中二三期病人的肿瘤组织也明显高于一期患者，相对于正常肺上皮细胞BEAS-2B，miR-221/222表达水平在A549和H1299癌细胞中表达明显升高。在A549和H1299细胞中过表达或敲低miR-221和miR-222，CD133阳性细胞明显降低，而过表达miR-221和miR-222则增加CD133阳性细胞，过表达或敲低miR-221和miR-222导致干性相关基因OCT4，Nanog和H-tert表达上调或下降，进一步提示miR-221/222的促细胞中的干性调节功能。我们构建了miR-221/222前体过表达的慢病毒载体，建立了A549稳转细胞系，体内裸鼠成瘤实验也证实了miR-221/222的促癌功能。我们研究发现miR-221/222能够直接靶向抑制ERCK的转录，进而解除其对Notch1基因的抑制作用，激活Notch1信号通路，促进肺癌的发生发展。miR-221/222调控肺癌细胞的增殖以及对肺癌干细胞的干性调控能力，为探索miRNAs作为非小细胞肺癌生物标志物及靶向治疗肺癌干细胞提供新的线索和思路。