Department of Respiratory Medicine, Liaocheng Infectious Disease Hospital, No.45, Jianshe East Road, Liaocheng, 252000, Shandong, China.
Biotechnol Lett. 2020 Jul;42(7):1123-1135. doi: 10.1007/s10529-020-02846-9. Epub 2020 Mar 13.
Lung cancer was one of the most deadly cancers around the world. Circular RNA AKT3 (CircAKT3) was highly expressed in lung cancer and could inhibit cell proliferation, but there were few studies on the mechanism of specific regulation of drug resistance. Therefore, we aimed to provide new ideas and perspectives for the role of circAKT3 in the mechanism of tumor resistance.
The levels of circAKT3, miR-516b-5p and STAT3 in lung cancer tissues and cells were examined using quantitative real-time polymerase chain reaction (qRT-PCR) or western blot assays. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay was used to examine the sensitivity of cells treated under different conditions to cisplatin (DDP). A glucose assay kit and lactate assay kit were used to assess glycolysis and lactate production of cells treated with different plasmids and 2-deoxy-glucose (2-DG). Western blot analysis was used to detect the expression level of the hypoxia-inducible factor (HIF-1α) in A549 and H1299 cells. Starbase 3.0 predicted a targeted relationship between circAKT3 and miR-516b-5p, STAT3 and miR-516b-5p, and the relationship was proved by a dual-luciferase reporter assay. Knockdown of circAKT3 was used to study the effects of circAKT3 on tumor development in vivo.
The levels of circAKT3 and STAT3 were upregulated, miR-516b-5p was downregulation in lung cancer tissues and cells. Functionally, circAKT3 knockdown improved cell sensitivity to DDP, and repressed glycolysis in lung cancer cells. Meanwhile, inhibition of HIF-1α-dependent glycolysis attenuated the circAKT3-induced increase of chemo-resistance in A549 cells. Mechanistically, miR-516b-5p was found to possess some binding sites with circAKT3. Noticeably, the inhibitory action of circAKT3 knockdown on DDP resistance and glycolysis was overturned through inhibitor of miR-516b-5p in lung cancer cells. Furthermore, besides, circAKT3 knockdown suppressed lung tumor cell growth by the miR-516b-5p/STAT3 axis in vivo.
CircAKT3 inhibit cisplatin sensitivity of lung cancer cells at least partly through regulating miR-516b-5p/STAT3 axis-mediated glycolysis balance, providing a possible long noncoding RNA -targeted therapy for lung cancer.
肺癌是全球最致命的癌症之一。环状 RNA AKT3（CircAKT3）在肺癌中高表达，可抑制细胞增殖，但关于其耐药特异性调控机制的研究较少。因此，我们旨在为 circAKT3 在肿瘤耐药机制中的作用提供新的思路和视角。
采用实时定量聚合酶链反应（qRT-PCR）或 Western blot 检测肺癌组织和细胞中 circAKT3、miR-516b-5p 和 STAT3 的水平。3-(4,5-二甲基-2-噻唑基)-2,5-二苯基-2-H-四唑溴盐（MTT）检测不同条件下顺铂（DDP）处理细胞的敏感性。葡萄糖测定试剂盒和乳酸测定试剂盒检测不同质粒和 2-脱氧葡萄糖（2-DG）处理后细胞的糖酵解和乳酸生成。Western blot 分析检测 A549 和 H1299 细胞中缺氧诱导因子（HIF-1α）的表达水平。Starbase 3.0 预测 circAKT3 与 miR-516b-5p、STAT3 与 miR-516b-5p 之间存在靶向关系，并通过双荧光素酶报告基因实验验证。敲低 circAKT3 研究 circAKT3 对体内肿瘤发生的影响。
肺癌组织和细胞中 circAKT3 和 STAT3 水平上调，miR-516b-5p 下调。功能上，circAKT3 敲低可提高 DDP 处理细胞的敏感性，抑制肺癌细胞的糖酵解。同时，抑制 HIF-1α 依赖性糖酵解可减弱 A549 细胞中 circAKT3 诱导的化疗耐药增加。机制上，发现 miR-516b-5p 与 circAKT3 具有一些结合位点。值得注意的是，在肺癌细胞中，circAKT3 敲低对 DDP 耐药和糖酵解的抑制作用通过 miR-516b-5p 抑制剂得到逆转。此外，circAKT3 敲低通过 miR-516b-5p/STAT3 轴抑制体内肺肿瘤细胞生长。
CircAKT3 通过调节 miR-516b-5p/STAT3 轴介导的糖酵解平衡，至少部分抑制肺癌细胞对顺铂的敏感性，为肺癌提供了一种潜在的长链非编码 RNA 靶向治疗方法。
