Wang Haifeng, Guo Min, Ding Dongxiao, Yang Feng, Chen Zhongjie
Department of Hematology and Oncology, Beilun Branch of the First Affiliated Hospital of Medical College of Zhejiang University, Ningbo, Zhejiang, People's Republic of China.
Department of Respiratory Medicine, Ningbo Zhenhai Longsai Hospital, Ningbo, Zhejiang, People's Republic of China.
Onco Targets Ther. 2020 Apr 30;13:3641-3652. doi: 10.2147/OTT.S237576. eCollection 2020.
Lung cancer is one of the most prevailing human cancers worldwide. Emerging evidence implies that long non-coding RNA nicotinamide nucleotide transhydrogenase-antisense RNA1 (NNT-AS1) is implicated in the tumorigenesis of lung cancer. Herein, we aimed to expose the impact of NNT-AS1 on the drug resistance of lung cancer.
Levels of NNT-AS1, microRNA (miR)-1236-3p and autophagy-related gene 7 (ATG7) were evaluated using quantitative real-time polymerase chain reaction (qRT-PCR) assay. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was implemented to detect cell proliferation and the half maximal inhibitory concentration (IC) of cisplatin (DDP) in vitro. Moreover, flow cytometry was performed to assess cell apoptosis. Cell migration and invasion were examined utilizing transwell assay in lung cancer cells. Besides, levels of ATG7 and cell behavior-related proteins were determined via Western blot. Dual-luciferase reporter assay was administrated to identify the interaction between miR-1236-3p and NNT-AS1 or ATG7. The biological role of NNT-AS1 in DDP resistance of lung cancer was examined by xenograft tumor model in vivo.
NNT-AS1 and ATG7 were upregulated, whereas miR-1236-3p was curbed in lung cancer tissues and in with or without DDP-resistant cell lines. NNT-AS1 detection significantly constrained cell growth, metastasis, and the IC of DDP in A549/DDP and H522/DDP cells. Interestingly, the influence of miR-1236-3p mimic on DDP resistance was overturned via NNT-AS1 upregulation in vitro. Reintroduction of miR-1236-3p inhibitor relieved the effect of ATG7 silencing on DDP sensitivity in A549/DDP and H522/DDP cells. Importantly, NNT-AS1 was a sponge of miR-1236-3p to separate ATG7. Besides, NNT-AS1 silencing enhanced DDP sensitivity of lung cancer in vivo.
NNT-AS1/miR-1236-3p/ATG7 axis regulated DDP resistance in lung cancer cells and might supply a probable target and prognostic marker in lung cancer treatment.
肺癌是全球最常见的人类癌症之一。新出现的证据表明，长链非编码核糖核酸烟酰胺核苷酸转氢酶反义核糖核酸1（NNT-AS1）与肺癌的肿瘤发生有关。在此，我们旨在揭示NNT-AS1对肺癌耐药性的影响。
采用定量实时聚合酶链反应（qRT-PCR）检测NNT-AS1、微小核糖核酸（miR）-1236-3p和自噬相关基因7（ATG7）的水平。采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐（MTT）法检测体外细胞增殖和顺铂（DDP）的半数最大抑制浓度（IC）。此外，通过流式细胞术评估细胞凋亡。利用Transwell实验检测肺癌细胞的迁移和侵袭能力。此外，通过蛋白质免疫印迹法检测ATG7水平和细胞行为相关蛋白。采用双荧光素酶报告基因实验确定miR-1236-3p与NNT-AS1或ATG7之间的相互作用。通过体内异种移植瘤模型研究NNT-AS1在肺癌顺铂耐药中的生物学作用。
在肺癌组织以及有或无顺铂耐药的细胞系中，NNT-AS1和ATG7上调，而miR-1236-3p受到抑制。检测NNT-AS1显著抑制了A549/DDP和H522/DDP细胞的生长、转移以及顺铂IC。有趣的是，在体外通过上调NNT-AS1可逆转miR-1236-3p模拟物对顺铂耐药性的影响。重新导入miR-1236-3p抑制剂可缓解A549/DDP和H522/DDP细胞中ATG7沉默对顺铂敏感性的影响。重要的是，NNT-AS1是miR-1236-3p的海绵，可隔离ATG7。此外，沉默NNT-AS1可增强体内肺癌对顺铂的敏感性。
NNT-AS1/miR-1236-3p/ATG7轴调节肺癌细胞的顺铂耐药性，可能为肺癌治疗提供一个潜在靶点和预后标志物。
