Chen Mingqiu, Liu Pingping, Chen Yuangui, Chen Zhiwei, Shen Minmin, Liu Xiaohong, Li Xiqing, Li Anchuan, Lin Yu, Yang Rongqiang, Ni Wei, Zhou Xin, Zhang Lurong, Tian Ye, Li Jiancheng, Chen Junqiang
Department of Radiation Oncology, Fujian Medical University Union Hospital and Fujian Provincial Platform for Medical Laboratory Research of First Affiliated Hospital, Fujian, China.
Department of Radiation Oncology, The Second Affiliated Hospital of Soochow University, Suzhou, China.
Front Genet. 2018 Dec 5;9:611. doi: 10.3389/fgene.2018.00611. eCollection 2018.
The aim of the present study was to identify the potential long non-coding (lnc.)-RNA and its associated molecular mechanisms involved in the regulation of the radiosensitivity of esophageal squamous cell cancer (ESCC) in order to assess whether it could be a biomarker for the prediction of the response to radiotherapy and prognosis in patients with ESCC. Microarrays and bioinformatics analysis were utilized to screen the potential lncRNAs associated with radiosensitivity in radiosensitive ( = 3) and radioresistant ( = 3) ESCC tumor tissues. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed in 35 ESCC tumor tissues (20 radiosensitive and 15 radioresistant tissues, respectively) to validate the lncRNA that contributed the most to the radiosensitivity of ESCC (named the candidate lncRNA). MTT, flow cytometry, and western blot assays were conducted to assess the effect of the candidate lncRNA on radiosensitivity in ECA109/ECA109R ESCC cells. A mouse xenograft model was established to confirm the function of the candidate lncRNA in the radiosensitivity of ESCC . The putative downstream target genes regulated by the candidate lncRNA were predicted using Starbase 2.0 software and the TargetScan database. The interactions between the candidate lncRNA and the putative downstream target genes were examined by Luciferase reporter assay, and were confirmed by PCR. A total of 113 aberrantly expressed lncRNAs were identified by microarray analysis, of which family with sequence similarity 201-member A (FAM201A) was identified as the lncRNA that contributed the most to the radiosensitivity of ESCC. FAM201A was upregulated in radioresistant ESCC tumor tissues and had a poorer short-term response to radiotherapy resulting in inferior overall survival. FAM201A knockdown enhanced the radiosensitivity of ECA109/ECA109R cells by upregulating ataxia telangiectasia mutated (ATM) and mammalian target of rapamycin (mTOR) expression via the negative regulation of miR-101 expression. The mouse xenograft model demonstrated that FAM201A knockdown improved the radiosensitivity of ESCC. The lncRNA FAM201A, which mediated the radiosensitivity of ESCC by regulating ATM and mTOR expression via miR-101 in the present study, may be a potential biomarker for predicting radiosensitivity and patient prognosis, and may be a therapeutic target for enhancing cancer radiosensitivity in ESCC.
本研究的目的是鉴定参与调节食管鳞状细胞癌（ESCC）放射敏感性的潜在长链非编码（lnc.）RNA及其相关分子机制，以评估其是否可作为预测ESCC患者放疗反应和预后的生物标志物。利用微阵列和生物信息学分析筛选放射敏感（n = 3）和放射抗性（n = 3）ESCC肿瘤组织中与放射敏感性相关的潜在lncRNAs。在35例ESCC肿瘤组织（分别为20例放射敏感组织和15例放射抗性组织）中进行逆转录定量聚合酶链反应（RT-qPCR），以验证对ESCC放射敏感性贡献最大的lncRNA（命名为候选lncRNA）。进行MTT、流式细胞术和蛋白质免疫印迹分析，以评估候选lncRNA对ECA109/ECA109R ESCC细胞放射敏感性的影响。建立小鼠异种移植模型，以证实候选lncRNA在ESCC放射敏感性中的功能。使用Starbase 2.0软件和TargetScan数据库预测由候选lncRNA调控的假定下游靶基因。通过荧光素酶报告基因检测检查候选lncRNA与假定下游靶基因之间的相互作用，并通过PCR进行确认。通过微阵列分析共鉴定出113个异常表达的lncRNAs，其中序列相似性家族201成员A（FAM201A）被鉴定为对ESCC放射敏感性贡献最大的lncRNA。FAM-201A在放射抗性ESCC肿瘤组织中上调，对放疗的短期反应较差，导致总体生存率较低。敲低FAM201A通过负调控miR-101的表达上调共济失调毛细血管扩张突变（ATM）和雷帕霉素哺乳动物靶标（mTOR）的表达，从而增强ECA109/ECA109R细胞的放射敏感性。小鼠异种移植模型表明，敲低FAM201A可提高ESCC的放射敏感性。在本研究中，lncRNA FAM201A通过miR-101调节ATM和mTOR的表达介导ESCC的放射敏感性，可能是预测放射敏感性和患者预后的潜在生物标志物，也可能是增强ESCC癌症放射敏感性的治疗靶点。
