High⁃throughput sequencing of driver gene mutations in non⁃small cell lung cancer
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摘要:
目的：探讨高通量测序技术在非小细胞肺癌驱动基因检测中的应用。方法：应用 Ion Torrent 高通量测序平台，检测150例非小细胞肺癌（non-small-cell lung cancer，NSCLC）患者EGFR、KRAS、BRAF、NRAS、Her-2和PIK3CA基因突变，并利用Sanger 测序法和微滴式数字PCR（droplet digital PCR，ddPCR）法检测150例NSCLC患者EGFR基因突变，对结果进行对比分析。结果：EGFR、KRAS、BRAF、NRAS、Her-2和PIK3CA基因突变检出率分别为51.33%（77/150）、7.33%（11/150）、1.33%（2/150）、1.33%（2/150）、2.00%（3/150）和4.67%（7/150）。57例标本未检出任何基因突变，84例标本检出单个驱动基因发生突变。9例标本检出2个及以上驱动基因发生突变。Sanger测序法检测EGFR基因突变，突变检出率为38.67%（58/150），与高通量测序法比较，差异有统计学意义（χ2=4.862，P=0.027），高通量测序法的灵敏度为98.28%，特异度为78.26%。ddPCR法突变检出率为50.00%（75/150），与高通量测序法比较，差异无统计学意义（χ2=0.053，P=0.818），阳性一致率为82.67%，阴性一致率为80.00%，总一致率为81.33%。结论：高通量测序法更适合应用于NSCLC诊疗，Sanger测序法和ddPCR法可作为有益的补充。
Abstract:
Objective：This study aims to investigate the value of detecting driver gene mutations in non-small cell lung cancer（NSCLC） patients using next-generation sequencing（NGS） technology. Methods：Somatic mutations in 150 NSCLC patients including EGFR，KRAS，BRAF，NRAS，Her-2 and PIK3CA were detected by Ion torrent personal genome machine（PGM）. Sanger sequencing and ddPCR was used to test and verify the results of EGFR gene mutations. Results：According to NGS results，mutations were detected in EGFR（51.33%，77/150），KRAS（7.33%，11/150），BRAF（1.33%，2/150），NRAS（1.33%，2/150），Her-2（2%，3/150）and PIK3CA（4.67%，7/150）. There were 57 samples without any somatic mutations in all genes，84 samples had one or more mutations in single gene，while 9 samples harboured mutations in two or more genes. The overall detection rate of EGFR mutation by NGS and Sanger sequencing was 51.33%（77/150）and 38.67%（58/150）. The difference between the two methods was statistically significant（χ2=4.862，P=0.027）. Compared to Sanger sequencing，the sensitivity and specificity of NGS assays was 98.28% and 78.26%，respectively. The overall detection rate of EGFR mutation by NGS and ddPCR was 51.33%（77/150）and 50%（75/150）. There was no significant difference between the two methods（χ2=0.053，P=0.818）. Compared to ddPCR，the positive concordance rate was 82.67%，the negative consistency rate was 80%，and the total concordance rate was 81.33%. Conclusion：Next-generation sequencing technology is more suitable for the diagnosis and treatment of NSCLC. Sanger sequencing and ddPCR can be useful supplements.
