长链非编码 RNA SNHG1 通过激活 Akt 通路促进索拉非尼耐药，并且在肝癌细胞中受 miR-21 正向调控。
LncRNA SNHG1 contributes to sorafenib resistance by activating the Akt pathway and is positively regulated by miR-21 in hepatocellular carcinoma cells.
机构信息
The Hepatosplenic Surgery Center, The First Affiliated Hospital of Harbin Medical University, Harbin, 150001, China.
Department of General Surgery, The Fourth Affiliated Hospital of Harbin Medical University, Harbin, 150001, China.
出版信息
J Exp Clin Cancer Res. 2019 May 3;38(1):183. doi: 10.1186/s13046-019-1177-0.
BACKGROUND
Acquired resistance to sorafenib greatly limits its therapeutic efficiency in the treatment of hepatocellular carcinoma (HCC). Increasing evidence indicates that long noncoding RNAs (lncRNAs) play important roles in the resistance to anti-cancer drugs. The present study aims to explore the involvement of lncRNA SNHG1 (small nucleolar RNA host gene 1) in sorafenib resistance and how SNHG1 is associated with overexpressed microRNA-21 (miR-21) and the activated Akt pathway, which have been demonstrated to mediate this resistance in HCC cells.
METHODS
Sorafenib-resistant HCC (SR-HCC) cells were generated and their sorafenib-resistant properties were confirmed by cell viability and apoptosis assays. Potential lncRNAs were screened by using multiple bioinformatics analyses and databases. The expression of genes and proteins was detected by qRT-PCR, Western blot and in situ hybridization. Gene silencing was achieved by specific siRNA or lncRNA Smart Silencer. The effects of anti-SNHG1 were evaluated in vitro and in experimental animals by using quantitative measures of cell proliferation, apoptosis and autophagy. The binding sites of miR-21 and SNHG1 were predicted by using the RNAhybrid algorithm and their interaction was verified by luciferase assays.
RESULTS
The Akt pathway was highly activated by overexpressed miR-21 in SR-HCC cells compared with parental HCC cells. Among ten screened candidates, SNHG1 showed the largest folds of alteration between SR-HCC and parental cells and between vehicle- and sorafenib-treated cells. Overexpressed SNHG1 contributes to sorafenib resistance by activating the Akt pathway via regulating SLC3A2. Depletion of SNHG1 enhanced the efficacy of sorafenib to induce apoptosis and autophagy of SR-HCC cells by inhibiting the activation of Akt pathway. Sorafenib induced translocation of miR-21 to the nucleus, where it promoted the expression of SNHG1, resulting in upregulation of SLC3A2, leading to the activation of Akt pathway. In contrast, SNHG1 was shown to have little effect on the expression of miR-21, which downregulated the expression of PTEN, leading to the activation of the Akt pathway independently of SNHG1.
CONCLUSIONS
The present study has demonstrated that lncRNA SNHG1 contributes to sorafenib resistance by activating the Akt pathway and its nuclear expression is promoted by miR-21, whose nuclear translocation is induced by sorafenib. These results indicate that SNHG1 may represent a potentially valuable target for overcoming sorafenib resistance for HCC.
背景
索拉非尼获得性耐药极大地限制了其在肝细胞癌（HCC）治疗中的疗效。越来越多的证据表明，长链非编码 RNA（lncRNA）在抗癌药物耐药中发挥重要作用。本研究旨在探讨 lncRNA SNHG1（核仁小分子 RNA 宿主基因 1）在索拉非尼耐药中的作用，以及 SNHG1 如何与过表达的 microRNA-21（miR-21）和激活的 Akt 通路相关，这些已被证明在 HCC 细胞中介导这种耐药性。
方法
通过细胞活力和凋亡测定法生成索拉非尼耐药性 HCC（SR-HCC）细胞，并证实其索拉非尼耐药性。通过多种生物信息学分析和数据库筛选潜在的 lncRNA。通过 qRT-PCR、Western blot 和原位杂交检测基因和蛋白质的表达。通过特异性 siRNA 或 lncRNA Smart Silencer 实现基因沉默。通过体外和实验动物定量测量细胞增殖、凋亡和自噬来评估抗 SNHG1 的作用。通过 RNAhybrid 算法预测 miR-21 和 SNHG1 的结合位点，并通过荧光素酶测定验证其相互作用。
结果
与亲本 HCC 细胞相比，SR-HCC 细胞中过表达的 miR-21 使 Akt 通路高度激活。在筛选出的十个候选物中，SNHG1 在 SR-HCC 细胞与亲本细胞之间以及在溶剂处理和索拉非尼处理的细胞之间的变化幅度最大。过表达的 SNHG1 通过调节 SLC3A2 激活 Akt 通路，从而有助于索拉非尼耐药。SNHG1 的耗竭通过抑制 Akt 通路的激活增强了索拉非尼诱导 SR-HCC 细胞凋亡和自噬的功效。索拉非尼诱导 miR-21 向核内易位，在核内促进 SNHG1 的表达，导致 SLC3A2 上调，从而激活 Akt 通路。相反，SNHG1 对 miR-21 的表达影响不大，miR-21 下调 PTEN 的表达，从而独立于 SNHG1 激活 Akt 通路。
结论
本研究表明，lncRNA SNHG1 通过激活 Akt 通路促进索拉非尼耐药，其核表达受 miR-21 促进，miR-21 的核易位由索拉非尼诱导。这些结果表明，SNHG1 可能是克服 HCC 索拉非尼耐药的一个有价值的潜在靶点。
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