烟草致癌物NNK诱发长链非编码RNA基因突变在肺癌发生中的作用

Lung cancer is one of the most common cancers in the world and the leading cause of cancer deaths.Development of lung cancer depends on both genetic and environmental factors.The mutations of environmental response genes are the basis of lung cancer development. Non-coding RNA(ncRNA) is an important kind of new environmental response genes discovered in recent years. Although many classes of functional ncRNAs have been recently described(including microRNAs, snRNAs and circular RNAs), the function of long noncoding RNA (lncRNA) have remained elusive in the field of chemical carcinogenesis.In the current study, we identified specific lncRNA related with the induction of tabacco specific carcinogen NNK[4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone] by using lncRNA microarray technology and bioinformatical analysis. Then,these specific lncRNA genes were sequenced by sequence capture array to discover the mutations within these genes. Some important mutations were selected to verify in lung cancer patients. Then,the candidated mutations were introduced into the cell model by recombinant DNA technique and site-directed mutagenesis.Subsequently, we observed the effect of lncRNA gene mutation on the carcinogenesis of lung cancer induced by NNK in vitro. The epidemiological study was also carried out to explore the relationship between lncRNA gene mutation and lung cancer risk in vivo. We hope that the current study will provide novel insights into the fields of environmental carcinogenesis and cancer pathogenesis.

肺癌是对人类生命威胁最大的恶性肿瘤之一，目前普遍认为环境致癌因素致相关蛋白编码基因突变是肺癌启动的分子基础，然而对环境致癌物诱发长链非编码RNA基因突变及其功能的研究尚未见报道。本研究通过检测烟草致癌物NNK诱导肺上皮细胞恶性转变后lncRNA表达谱的变化，采用多种生物信息学方法，综合分析筛选NNK相关的lncRNA基因作为捕获区域，通过目标序列捕获测序技术广泛寻找lncRNA基因突变位点，根据突变位点的重要性评分进行突变位点的筛选，并在肺癌人群中进行验证；通过构建不同基因型细胞和对不同基因型肺癌病例临床资料的分析，从细胞和人群水平定量分析NNK诱发的lncRNA基因突变对lncRNA表达调控和肺癌发生发展的影响，为环境致癌的分子机制和肺癌的病因学研究提供新的思路和实验依据。

肺癌是对人类生命威胁最大的恶性肿瘤之一，环境致癌物暴露是肺癌的重要病因之一。目前普遍认为环境致癌因素致相关蛋白编码基因突变是肺癌启动的分子基础，而对环境致癌物诱发长链非编码 RNA 基因突变及其功能的研究尚未见报道。本研究主要探讨烟草致癌物NNK诱发长链非编码RNA基因突变，通过调控非编码RNA的表达及下游通路参与肺癌发病的机制。本项目采用NNK体外慢性染毒方法，成功建立了 NNK 诱导的BEAS-2B和16-HBE恶性转化细胞株，通过lncRNA 表达谱芯片技术，筛选确定2106个lncRNA，建立NNK诱导相关特异性lncRNA表达谱，其中lncRNA AFAP1-AS1、miR-22HG和uc.77被证实在肺癌发生发展中发挥重要的调控作用；随后采用目标序列捕获测序技术高通量获得NNK相关lncRNA基因突变谱，在细胞和人群水平定量分析并验证了lncRNA基因突变在肺癌发生发展中的重要作用。结果表明烟草致癌物NNK可诱导长链非编码RNA基因发生突变，其中AFAP1-AS1基因 C/T 突变与肺癌患者的吸烟量、肿瘤分期具有相关性，提示AFAP1-AS1基因突变可在肺癌发生发展中发挥重要作用。目前已按计划完成了研究目标和研究任务，项目经费使用合理，本研究结果可为环境致癌的分子机制和肺癌的病因学研究提供了新的思路和实验依据。