血浆Exosomes中长链非编码RNA作为肺腺癌早期诊断和进展评价标志物的研究

Exosomes are cell-derived small vesicles, which play important roles in the occurrence and development of various cancer. Tumor-derived Exosomes may provide significant information on tumor diagnosis and progression with regard to a large number of tumor associated molecules in them. Results from our preliminary experiment have found many long non-coding RNAs (LncRNAs) with significantly different expression between the lung adenocarcinoma cell line A549-derived Exosomes and the lung normal epithelial cell line 16HBE-derived Exosomes. Moreover, the LncRNAs’ expression in Exosomes was quite similar with their expression in the host cells. All these indicated that Exosomes may contain lung adenocarcinoma-associated LncRNAs. Tumor-derived Exosomes have easier access to the blood. Its membrane structure can effectively prevent the embedded LncRNAs to be degraded. Therefore, these lung adenocarcinoma-associated LncRNAs in plasma Exosomes may provide practicable bioinformation on early diagnosis and progression evaluation for lung adenocarcinoma. Here, with a relatively large population study, this project intends to construct a system based on Exosomes LncRNAs in plasma for early diagnosis and progression evaluation of lung adenocarcinoma; and to dynamically observe the change of Exosomes LncRNAs in plasma during the lung cancerization of animal model to validate the results from population study. This project will help to afford biomarkers for early diagnosis and progression evaluation of lung adenocarcinoma.

外泌体（Exosomes）是一种由细胞分泌的小囊泡，其与肿瘤发生发展密切相关。肿瘤细胞分泌的Exosomes内含大量的肿瘤相关分子，可为肿瘤诊断和进展评价提供重要信息。申请者预实验比较肺腺癌细胞A549和肺正常细胞16HBE分泌的Exosomes，发现多个长链非编码RNA（LncRNAs）表达差异显著，且LncRNAs表达与宿主细胞具有高度一致性，提示Exosomes中存在肺腺癌相关的LncRNAs。Exosomes易于进入血液，其膜结构可防止LncRNAs被降解，血浆Exosomes中肺腺癌特异的LncRNAs可对肺癌的早期诊断和进展评价提供可用的生物信息。本项目拟通过大样本的人群研究，建立基于血浆Exosomes LncRNAs的肺腺癌诊断和进展评价体系，并动态观察动物肺癌变模型中血浆Exosomes LncRNAs的变化，验证人群研究结果，为肺腺癌的早期诊断和进展评价提供生物标志物。

肿瘤细胞分泌的外泌体（Exosomes）内含大量的肿瘤分子，如长链非编码RNA（lncRNA），可为肿瘤诊断和进展评价提供重要信息。本课题通过收集一定样本的肺腺癌病例和对照，结合二代测序技术（RNA-seq）和荧光定量PCR（RT-PCR）技术，对血浆Exosomes内长链非编码RNA分子（包括lncRNA的特殊类型环状RNA即circRNA）在肺腺癌诊断和进展评价中的意义进行了分析。首先通过RNA-seq技术检测了6例肺腺癌病例和5例肺炎对照的lncRNA表达谱，筛选差异表达的lncRNA分子；经与组织RNA-seq数据比对，挑选出14个lncRNAs，进而在50对肺癌组织与癌旁组织中进行表达验证，确定LINC01125、 lnc-TBC1D8-6:1、hsa_circ_0047921、hsa_circ_0007761、hsa_circ_0056285 5个lncRNAs为血浆Exosomes中肺腺癌关联的lncRNAs标志。最后通过RT-PCR方法在500余对肺腺癌与非肺癌对照（包括健康对照、肺结核、哮喘、慢阻肺患者等）中检测这些标志的表达，并分析其诊断和进展评价意义。结果显示LINC01125、hsa_circ_0047921、hsa_circ_0007761、hsa_circ_0056285在肺腺癌和非肺癌来源血浆的Exosomes中的表达有显著差异，受试者工作曲线分析（ROC）显示各标志单独对我国人群肺腺癌具有显著的诊断价值，其组合能显著增加诊断的准确性，提高灵敏度和特异度。然而，判别分析显示上述lncRNAs组合对肺腺癌进展的判别准确性较低。综合上述结果，本研究为血浆Exosomes内lncRNAs应用于肺腺癌诊断奠定了坚实的工作基础。项目共资助发表SCI论文12篇，总影响因子49分。培养青年老师2名、硕士研究生4人。项目投入经费24万元，支出17.9486万元，各项支出基本与预算相符。剩余经费6.0514万元，剩余经费计划用于本项目研究后续支出。