长链非编码RNA SNHG6在肝癌发生发展中的作用及其机制研究

We have demonstrated that the expression of long noncoding RNAs (lncRNAs) has been dysregulated in hepatocellular carcinoma (HCC) and may serve as novel prognostic markers and therapeutic targets (Gastroenterology，2015). Recently, our preliminary data showed that lncRNA SNHG6 was highly expressed in HCC tissues compared with controls and was associated with portal vein tumor thrombus, tumor stage, metastasis, and shorter overall survival of HCC patients. Ectopic expression of SNHG6 in HCC cells promoted cell proliferation and inhibited apoptosis. Furthermore, SNHG6 could become a sponge for miR-26a/b. Thus, based on our data, we proposed that SNHG6 may be functioned as competing endogenous RNA (ceRNA) to promote HCC progression. To confirm the hypothesis, firstly, the effect of SNHG6 will be identified by clinical multicenter sample in HCC. Secondly, we will screen the target mRNA and explore the relationship among SNHG6, miR-26a/b, and target mRNA. Lastly, we will investigate the functions of ceRNA network involving SNHG6 by in vitro and in vivo tumor assays and SNHG6 transgene mice. In conclusion, SNHG6 may act as a biomarker for poor prognosis in HCC and confer a malignant phenotype to tumor cells. The ceRNA network involving SNHG6 may contribute to a better understanding of liver carcinogenesis and tumor progression.

我们已发现长链非编码RNA（lncRNA）在肝癌发生、发展中发挥重要调控功能（Gastroenterology，2015）。近来，我们预实验结果表明：lncRNA SNHG6在肝癌中高表达且与患者预后不良密切相关；SNHG6可促进肝癌细胞增殖、抑制凋亡；SNHG6可作为miR-26a/b的“海绵”发挥生物学功能。基于前期研究结果，我们提出SNHG6作为竞争性内源RNA（ceRNA）发挥癌基因作用的科学假说。为验证假说，本项目将利用临床多中心、大样本分析SNHG6与肝癌发生发展的关系；其次，筛选SNHG6调控的mRNA，并阐明SNHG6－miR-26a/b－靶mRNA之间互为调控的网络关系，并通过肿瘤体内外实验及转基因动物验证SNHG6的ceRNA网络；最后，探讨SNHG6二级结构与生物学功能的关系。据此，阐明SNHG6在肝癌中确切的生物学功能及调控机制。

已有研究表明长链非编码RNA（lncRNA）在肝细胞癌（HCC）中表达失调。然而，lncRNA在肝癌中的功能和作用机制仍然未知。在此研究中，我们确定了SNHG6 与肝癌发生、发展及预后的关系。发现SNHG6的5个转录本在HCC组织中差异表达，并利用慢病毒载体过表达或siRNA沉默肝癌细胞株中SNHG6 表达，通过肿瘤体外实验方法和体内实验方法观察SNHG6对肝癌细胞增殖、粘附、侵袭及转移的影响，确定SNHG6在肝癌中的生物学功能，发现只有SNHG6-003具有促癌功能。SNHG6-003在HCC细胞中的过表达促进细胞增殖并诱导其抗药性；而敲低SNHG6-003可促进肝癌细胞凋亡。此外，通过双荧光素酶基因报告实验，我们确定了miR-26a/b与靶基因TAK1 3′-UTR 的直接结合关系，并明确了过表达或沉默SNHG6-003后对miR-26a/b与TAK1mRNA结合能力的影响。最终证明SNHG6-003作为竞争性内源RNA（ceRNA）起作用，可以作为“海绵”吸附miR-26a/b，从而调节转化生长因子-β-活化激酶1（TAK1）的表达。重要的是，表达分析显示SNHG6-003和TAK1均在人类癌症中上调，表现出共表达模式。在HCC患者中，SNHG6-003的高表达与肿瘤进展密切相关并且SNHG6-003高表达的肝癌患者生存时间短。因此，靶向涉及SNHG6-003的ceRNA网络可以用作针对HCC的治疗策略。