一个功能未知的环状RNA circ-0000211调控肺腺癌侵袭转移的机制研究

By the application of high-throughput microarray technology, we identified an unknown circular RNA, circ-0000211(CIR) which is highly expressed in human lung adenocarcinoma. Our pre-experiments suggested that CIR was significantly overexpressed in lung adenocarcinoma, and was closely related with lymph node metastasis. By silencing siRNA, we found CIR can inhibit invasion and metastasis of lung adenocarcinoma cell lines. Further bioinformatics analysis revealed that CIR mainly promotes the invasion and metastasis of lung adenocarcinoma by competing with miR-622. In turn, miR-622 upregulates the downstream gene, HIF1-α. Therefore, we propose the scientific hypothesis that CIR promotes the invasion and metastasis of lung adenocarcinoma by endogenous miR-622 competition which upregulating HIF1-α expression. The study was designed to evaluate the biological function of CIR which promoting the invasion and metastasis of lung adenocarcinoma with vivo and in vitro experiments. In addition, along with rescue experiment , we aimed to clarify the molecular mechanism that CIR competitively adsorbs miR-622，which leading to upregulating HIF1-α expression. Finally, with clinical samples, we comprehensively evaluated the relationship between CIR/miR-622/HIF1-α regulatory networks and lung adenocarcinoma invasion and metastasis. CIR related functions and mechanism in lung adenocarcinoma have not been reported. Thus, this study may provide a new target for the prevention and treatment of lung adenocarcinoma.

circ-0000211（简称CIR）是我们应用高通量芯片技术筛选获得的一条功能未知、在人肺腺癌组织中差异高表达的环状RNA。前期预实验提示：CIR高表达与肺腺癌淋巴结转移显著正相关；体外沉默CIR可抑制肺腺癌细胞株侵袭转移能力。生物信息学预测及预实验提示CIR可作为内源竞争性RNA（ceRNA）吸附miR-622、上调促转移基因HIF1-α表达。故我们提出“CIR通过竞争性吸附miR-622上调HIF1-α表达、从而促进肺腺癌侵袭转移”的科学假说。本研究拟通过体内外实验，明确CIR促进肺腺癌侵袭转移的生物学功能；设计拯救实验，阐明CIR可竞争性吸附miR-622上调HIF1-α表达、从而促进肺腺癌侵袭转移的分子机制；最后结合临床样本全面评价CIR/miR-622/HIF1-α调控网络与肺腺癌侵袭转移的相关性。有关CIR促进肺腺癌侵袭转移的功能及机制未见报道，本研究可为肺癌防治提供新靶标。

背景：环状RNA与多种肿瘤的侵袭以及进展密切相关。在我们前期的芯片研究结果提示环状RNA hsa-circ-0000211在肺腺癌组织中异常表达。但是其在肺腺癌中起何种作用以及作用机制尚未明确。研究方法：我们利用RT-qPCR技术检测Hsa-circ-0000211在肺腺癌组织以及细胞系中的表达水平。我们通过生物信息学分析以及荧光素酶报告基因的方法证实Hsa-circ-0000211与 miR-622直接相结合。通过RNA沉默以及过表达的方法证实hsa-circ-0000211调节miR-622以及HIF1A.表达水平。我们利用transwell assay技术发现hsa-circ-0000211具有促进侵袭以及转移的功能。Western blot 被用来检测HIF1A的蛋白表达水平。结果：相较于正常肺组织以及肺上皮细胞Hsa-circ-0000211在肺腺癌以及细胞株中高表达。沉默Hsa-circ-0000211表达能够抑制肺腺癌的侵袭以及转移。进一步的研究提示has-circ- 0000211主要通过miR-622海绵作用来促进肺腺癌的转移及侵袭。随后，我们也发现has-circ- 0000211主要通过作用于miR-622来上调HIF1-α表达。结论：Has-circ-0000211主要通过hsa-circ-0000211/miR-622/HIF1-α调节网络来促进肺腺癌的侵袭转移。我们的研究为肺腺癌的治疗提供了新的靶点。