Long noncoding RNA ANRIL could be transactivated by c-Myc and promote tumor progression of non-small-cell lung cancer

OncoTargets and Therapy Dovepress open access to scientific and medical research Original Research OncoTargets and Therapy downloaded from https://www.dovepress.com/ by 54.38.22.117 on 12-Jul-2018 For personal use only. Open Access Full Text Article Long noncoding RNA ANRIL could be transactivated by c-Myc and promote tumor progression of non-small-cell lung cancer This article was published in the following Dove Press journal: OncoTargets and Therapy 24 May 2016 Number of times this article has been viewed Yi Lu Xiaohui Zhou Ling Xu Chaohui Rong Ce Shen Wei Bian Department of Respiratory Medicine, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai, People’s Republic of China Introduction Correspondence: Wei Bian; Ce Shen Department of Respiratory Medicine, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, 600 Yi Shan Road, Shanghai 200233, People’s Republic of China Fax +86 21 6470 1361 Email ; Lung cancer remains the leading cause of cancer-related deaths around the world.1 Non-small-cell lung cancer (NSCLC), including adenocarcinoma, squamous cell carcinoma, and large cell carcinoma account for .85% of all lung cancer cases.2 Despite the promising advances made in therapeutic technologies, the prognosis of this disease remains dismal, with the 5-year survival rate of NSCLC (all stages combined) being only ~13%.3 Tumor metastasis and recurrence are common and represent major obstacles to the improvement of patient survival. The long-term survival rate is ,50% even after curative resections in NSCLC.4 Therefore, illumination of the mechanisms underlying metastasis and recurrence is the prerequisite for the development of novel therapeutics in NSCLC. Long noncoding RNAs (lncRNAs) are a class of noncoding RNAs longer than 200 nucleotides and with limited protein-coding potential. Studies revealed that lncRNAs could interact with DNA, RNA, or protein and regulate a variety of genes with diversified mechanisms, thereby having an effect on a large number of cellular pathways, including carcinogenesis.5–10 Aberrantly expressed lncRNAs have been demonstrated to play key roles in the progression of NSCLC.11–13 lncRNA ANRIL (CDKN2B antisense RNA 1) is a 3.8 kb lncRNA transcribed from the INK4B-ARF-INK4A gene cluster in the opposite direction.14 ANRIL has been demonstrated to exert oncogenic activity in a variety of carcinomas, including NSCLC13 and gastric cancer.15 However, the factors that lead to its upregulation and the biological roles of ANRIL in NSCLC remain to be explored. 3077 submit your manuscript | www.dovepress.com OncoTargets and Therapy 2016:9 3077–3084 Dovepress © 2016 Lu et al. This work is published and licensed by Dove Medical Press Limited. The full terms of this license are available at https://www.dovepress.com/terms.php and incorporate the Creative Commons Attribution – Non Commercial (unported, v3.0) License (http://creativecommons.org/licenses/by-nc/3.0/). By accessing the work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. For permission for commercial use of this work, please see paragraphs 4.2 and 5 of our Terms (https://www.dovepress.com/terms.php). http://dx.doi.org/10.2147/OTT.S102658 Powered by TCPDF (www.tcpdf.org) Abstract: In recent years, long noncoding RNAs (lncRNAs) have been demonstrated to play important roles in the development of human cancer. We assessed the role of lncRNA ANRIL in non-small-cell lung cancer (NSCLC). Quantitative real-time polymerase chain reaction was employed to detect the expression of ANRIL in NSCLC tissues and paired nontumor tissues. The high expression level of ANRIL was positively correlated with advanced tumor–node– metastasis stage and greater tumor diameter. Furthermore, chromatin immunoprecipitation assays confirmed the physical interaction between c-Myc and ANRIL. ANRIL silencing significantly inhibited NSCLC cell proliferation. Together, we showed that ANRIL is involved in the oncogenesis of NSCLC, and ANRIL may be a potential therapeutic target for patients with NSCLC. Keywords: c-Myc, ANRIL, cell proliferation, NSCLC Dovepress OncoTargets and Therapy downloaded from https://www.dovepress.com/ by 54.38.22.117 on 12-Jul-2018 For personal use only. Lu et al c-Myc is a well-noted transcriptional factor and has been shown to be abnormally overexpressed in a variety of carcinomas, including NSCLC.16 As a vital transcription regulator, c-Myc plays an essential role in the regulation of many biological processes, including cell cycle, proliferation, apoptosis, protein synthesis, and cell metabolism.17 It can bind to the cis-regulatory element E-box (CACGTG) to transactivate targeted genes.18 In this study, we have shown that c-Myc can directly transactivate ANRIL via interacting with putative c-Myc target response element in the promoter region of ANRIL. Moreover, we examined the effect of ANRIL on the biological behaviors of NSCLC cells. Our results demonstrated that ANRIL could promote the proliferation of NSCLC cells. Materials and methods Patient samples Fifty-six NSCLC tissues and their paired adjacent normal tissues in this study were obtained from patients who underwent radical resections at the Sixth Hospital (Shanghai Jiaotong University, Shanghai, People’s Republic of China). The study was approved by Research Ethics Committee of Shanghai Jiaotong University (Shanghai, People’s Republic of China). Informed consent was obtained from all patients. Cell culture An NSCLC squamous carcinoma cell line (SK-MES-1), three NSCLC adenocarcinoma cell lines (A549, SPC-A1, and NCI-H1975), and a normal human bronchial epithelial cell line (16HBE) were purchased from the Institute of Biochemistry and Cell Biology of the Chinese Academy of Sciences (Shanghai, People’s Republic of China). The cells were cultured in Roswell Park Memorial Institute 1640 or Dulbecco’s Modified Eagle’s Medium (DMEM) (GibcoBRL; Grand Island, NY, USA) medium supplemented with 10% fetal bovine serum (Thermo Fisher Scientific, Waltham, MA, USA), 100 U/mL penicillin, and 100 mg/mL streptomycin (Invitrogen) in incubator at 37°C with 5% CO2. RNA extraction and quantitative real-time polymerase chain reaction Total RNA was extracted from cancer cells utilizing the Trizol reagent (Invitrogen) following the manufacturer’s instructions. First strand complementary DNA (cDNA) was generated with the Reverse Transcription System Kit (Invitrogen). The cDNA template was then amplified using the standard SYBR Green polymerase chain reaction (PCR) kit protocol with real-time (RT)-PCR. The quantitative real-time polymerase chain reaction (qRT-PCR) was performed on ABI 3078 Powered by TCPDF (www.tcpdf.org) submit your manuscript | www.dovepress.com Dovepress 7500 system (Applied Biosystems, Foster City, CA, USA). β-Actin was used as the internal control, and (...truncated)