参与乳腺肿瘤转移的miRNA筛选及机制研究

Tumor metastasis is primarily responsible for cancer-associated mortality. Epithelial mesenchymal transition (EMT) is a cell reprogramming process, and plays an important role in tumor invasion and metastasis. EMT is regulated by many factors, including the cytokines, transcription factors and miRNAs. So far, the role of miRNA in EMT is still poorly understood. In this project we attempt to identify metastasis associated genes and miRNAs, we designed a serial of screening models, such as in vivo mouse screening model and in vitro transwell screening model, to screen miRNAs involved in EMT and metastasis in SKBR3 and MCF7 cell pools expressing miRNAs libraries, as well as analysis of gene expression profiling of TGF-β and Snail induced EMT model in MCF10A. In addition, we clarify the mechanisms of how miRNAs regulate metastasis. Our preliminary data showed that miR-578j, miR-182, miR-200c, and other miRNAs involved in regulating tumor invasion and metastasis process. Therefore, we need to further clarify the function of these miRNAs at the cellular and animal level, and also to explore its molecular mechanism in the process of invasion and metastasis through indentifying the targets of miRNAs. Moreover, miRNAs signatures identified in this project can delineate their clinical significance for breast cancer, and may provide new molecular approaches for diagnosis and prognosis of patients.

侵袭转移是恶性肿瘤致死的主要原因，上皮-间质转化(EMT)则在肿瘤侵袭转移过程中发挥关键作用。EMT受多因素调控，包括细胞因子、转录因子和miRNA等。目前对miRNA参与侵袭转移调控研究刚刚起步。本课题以乳腺癌为研究对象，通过在低转移乳腺癌细胞系中建立稳定表达miRNA文库再经过体内小鼠转移模型和体外侵袭模型来筛选促进侵袭转移和EMT过程的miRNA。并结合TGF-β和Snail诱导EMT细胞模型的表达谱进行分析来确立参与乳腺肿瘤侵袭转移调节的miRNA。初期筛选研究显示miR-578j、miR-182、200c等多种miRNA参与乳腺肿瘤侵袭转移过程的调节。因此需要进一步在细胞水平、动物水平上研究这些miRNA在肿瘤侵袭转移过程中的功能；同时寻找其靶基因来探索它们发挥作用的分子机制。最后通过在临床组织标本中表达分析来阐明其在临床应用中的意义，为将来能成为临床诊断分子标志提供理论基础。

侵袭转移是恶性肿瘤致死的主要原因，上皮-间质转化(EMT)则在肿瘤侵袭转移过程中发挥关键作用。EMT 受多因素调控，包括细胞因子、转录因子和miRNA 等。目前对miRNA参与侵袭转移调控研究刚刚起步。本课题通过在低转移乳腺癌细胞系中建立稳定表达miRNA 文库再经过体内小鼠转移模型和体外侵袭模型来筛选促进侵袭转移和EMT 过程的miRNA。并结合TGF-β和Snail 诱导EMT 细胞模型的表达谱进行分析来确立参与乳腺肿瘤侵袭转移调节的miRNA。通过多种筛选模型对参与侵袭转移调控的miRNA筛选，我们鉴定出MicroRNA-548j、MicroRNA-182等可以显著促进肿瘤细胞的转移，MicroRNA-92b显著抑制肿瘤细胞的转移。随后在细胞水平以及小鼠动物模型中进行验证。我们对这些MicroRNA参与肿瘤转移调节的分子机制也进行深入探索。发现MicroRNA-548j可以通过调节其下有靶基因Tensin1（TNS1）的表达来发挥其生理学功能，MicroRNA-182可以调节SNAI1的表达。而MicroRNA-92b则是通过抑制integrin αV (ITGAV)的表达从而参与抑制肿瘤细胞的转移能力。最后，我们也完成了在临床乳腺肿瘤转移样本和非转移样本中我们分别检测这些MicroRNA的表达情况，结果也表明MicroRNA-548j在淋巴结转移的肿瘤样本表达显著上调，MicroRNA-182在淋巴结转移灶中的表达水平也显著高于原发灶。而MicroRNA-92b在在淋巴结转移的肿瘤样本表达则显著下调。此项研究筛选调节恶性肿瘤的侵袭和转移的miRNA并探索其作用机制，显示了其潜在的巨大的应用价值，为将来能成为临床诊断分子标志提供理论基础。