肺腺癌恶性胸水中外泌体来源双链DNA的鉴定及其在EGFR突变检测中的意义  

Identification of high quality double strands DNA of exosome from maliglant pleural effusion and its significance in EGFR mutation detection

作　　者：张鹤 赵慧霞[1] 曾志艳 李秋文[1] 肖文华[1] ZHANG He;ZHAO Huixia;ZENG Zhiyan;LI Qiuwen;XIAO Wenhua(Department of Oncology,Forth Medical Center,PLA General Hospital,Beijing 100048,China)

机构地区：[1]解放军总医院第四医学中心肿瘤内科,北京100048

出　　处：《临床肿瘤学杂志》2023年第4期365-370,共6页Chinese Clinical Oncology

基　　金：北京市科委科技计划重大项目资助项目(D141100000214603)。

摘　　要：目的探索肺腺癌恶性胸水中外泌体来源DNA(exoDNA)用于表皮生长因子受体(EGFR)突变检测的可行性。方法采用沉淀法收集胸水外泌体,透射电镜和Western blot对外泌体进行鉴定;同时对exoDNA的稳定性和物理特征进行鉴定。收集28例初诊肺腺癌患者的胸水及对应癌组织,采用扩增阻滞突变系统方法对胸水脱落癌细胞、exoDNA及对应癌组织的EGFR基因突变进行检测,比较外泌体用于EGFR基因突变检测的敏感性、特异性和一致性。结果电镜证实提取的外泌体符合外泌体的大小特征;Western blot印证了外泌体富含的特征蛋白CD63。exoDNA含有大片段DNA且纯度符合要求、适合EGFR突变分析;在肺腺癌初治患者的恶性胸水exoDNA中含有EGFR突变,与配对的组织和胸水脱落癌细胞相比,敏感性和特异性为100%和92.9%,符合率为96.4%(Kappa=0.929,P<0.001)。结论肺腺癌胸水exoDNA质量好,稳定性高,是EGFR基因突变检测的另一全新的、重要的DNA来源。Objective To investigate the feasibility for exosomal DNA(exoDNA)in malignant pleural effusions as a novel DNA recourse for epidermal growth factor receptor(EGFR)mutation analysis in patients with lung adenocarcinoma.Methods Twenty-eight newly diagnosed lung adenocarcinoma patientes with maliglant pleural effusions were investigated.All the 28 patients with lung adenocarcinoma were confirmed by pathology.Exosome was confirmed by electric microscope and Western blot.The exoDNA was extracted from exosome of malignant pleural effusion and its quality and physcial characteristics were indentified.We collected pleural effusions and matched tumor tissues,examined EGFR mutation with cell blocks,exoDNA of pleural effusions and tumor tissues by amplification refractory mutation system,and then investigated the sensitivity,specificity and consistency among them.Results Electron microscopy confirmed that the extracted exosomes matched the size characteristics of exosomes.Western blot confirmed the rich characteristic protein CD63 in exosomes.ExoDNA contained large fragments of DNA with satisfactory purity and was suitable for EGFR mutation analysis.The exoDNA in malignant pleural effusions contained EGFR mutation with a sensitivity of 100%and a specificity of 92.9%.A coincidence rate of 96.4%(Kappa=0.929,P<0.001)was observed in comparison with cell blocks and tumor tissues.Conclusion The quality of exoDNA is high and can be used as a reliable,novel resource of DNA for EGFR detection in lung adenocarcinoma.

关 键 词：肺腺癌 恶性胸水 外泌体 表皮生长因子受体 

分 类 号：R734.2[医药卫生—肿瘤]