非小细胞肺癌胸水中癌细胞EGFR基因TK区突变研究

目的:检测非小细胞肺癌(NSCLC)患者胸水标本中癌细胞EGFR基因TK区突变。方法:收集21例NSCLC患者胸水标本,经密度梯度离心后,将胸水中细胞包埋,制成蜡块;巢式聚合酶链式反应(PCR)扩增EGFR基因18-21号外显子;应用PCR-LIS-SSCP及直接测序方法检测EGFR基因TK区突变。结果:21例NSCLC患者胸水癌细胞标本,19例为腺癌,1例为腺鳞癌,1例为大细胞癌;9例患者存在EGFR基因TK区突变,均为腺癌,突变率为42.9%(9/21),腺癌患者突变率为47.4%(9/19);9例患者中8例接受TKIs治疗,疗效评价为PR(部分缓解)或SD(稳定)。5例出现TKIs耐药,疾病进展TTP(疾病进展时间)为4～17个月不等。应用TKIs治疗前后对比胸水及癌组织中EGFR基因突变,2例出现获得性突变。结论:NSCLC胸水标本与文献报道的癌组织中EGFR基因TK区突变率基本相同;胸水标本中检测肿瘤细胞基因突变的方法可靠,值得在临床推广、应用。

Study for the mutations of EGFR gene TK domain in cancer cells of pleural fluid from NSCLC patients

Objective：To investigate the mutation status of EGFR exon 18 to 21 in 21 patients of non-small cell lung cancer （NSCLC） in pleural fluid samples. Method： Using density gradient centrifugation method to remove the part of mesothelial cells, inflammatory cells and red cells, collecting centrifugated cells to make the paraffin blocks; extract DNA from the paraffin blocks of pleural fluid. EGFR were tested by mutations in exon 18 to 21. Result： EGFR mutations were detected in 9 of 21 NSCLC patients, mutation rate was 42.9% （19/21）. The mutation rate of ad patients was 47.4% （9/19）. The 8 patients with EGFR mutations were accepted the TKIs therapy. The evaluations for therapeutic effect were partial response （PR） or stable disease （SD）. 5 patients had acquired resistance of TKIs, among them, two were found new acquired EGFR gene mutations. Conclusion： The mutant rate of pleural fluid samples those we report was coincidence with the cancer tissue sample reported. The measuring gene mutation process of pleural fluid samples we report provide a stabilized method for using in clinical and research study. It was suggested that this method should be a stabilized way. It can be widely used in clinical course.