Oncol Res Treat. 2018;41(10):634-642. doi: 10.1159/000491441. Epub 2018 Aug 27.
This study evaluated the distribution of epidermal growth factor receptor (EGFR) T790M mutations in treatment-naïve tumor and normal samples obtained from cancer patients.
We utilized allele-specific PCR (AS-PCR), digital droplet PCR (ddPCR) and next generation sequencing (NGS) to detect EGFR T790M allele in several collections of tumor and normal human tissues.
AS-PCR analysis of treatment-naïve tumor samples revealed somatic T790M mutation in 3/394 (1%) non-small cell lung carcinomas (NSCLC) carrying the tyrosine kinase inhibitor (TKI)-sensitizing EGFR mutation, but in none of 334 NSCLC lacking EGFR exon 19 deletions (ex19del) or L858R substitutions and in none of 235 non-lung tumors. Use of highly sensitive and quantitative assays, such as ddPCR and NGS, produced a high number of T790M-specific signals even in presumably T790M-negative DNA specimens. This background noise was evidently higher in degraded DNA isolated from formalin-fixed paraffin-embedded tissues as compared to high molecular weight DNA. A combination of AS-PCR, ddPCR and NGS revealed mosaic EGFR T790M allele in 2/68 (3%) NSCLC treated with the first-generation TKI. Both these tumors produced evident and durable response to gefitinib.
Detection of mosaic EGFR T790M mutation in treatment-naïve samples may be compromised by yet unresolved technical issues and may have limited clinical value.
本研究评估了未经治疗的肿瘤组织和癌症患者正常组织中表皮生长因子受体（EGFR）T790M 突变的分布。
我们利用等位基因特异性 PCR（AS-PCR）、数字液滴 PCR（ddPCR）和下一代测序（NGS）来检测几种肿瘤和正常人类组织中 EGFR T790M 等位基因。
AS-PCR 分析结果显示，3/394（1%）例携带酪氨酸激酶抑制剂（TKI）敏感 EGFR 突变的非小细胞肺癌（NSCLC）患者的未经治疗的肿瘤样本中存在体细胞 T790M 突变，但在 334 例无 EGFR 外显子 19 缺失（ex19del）或 L858R 取代的 NSCLC 患者和 235 例非肺癌肿瘤患者中均未发现 T790M 突变。使用高灵敏度和定量检测方法，如 ddPCR 和 NGS，即使在推测为 T790M 阴性的 DNA 标本中，也产生了大量的 T790M 特异性信号。与高分子量 DNA 相比，从福尔马林固定石蜡包埋组织中分离的降解 DNA 中这种背景噪声显然更高。AS-PCR、ddPCR 和 NGS 的组合显示，在接受第一代 TKI 治疗的 68 例 NSCLC 中有 2 例（3%）存在 EGFR T790M 嵌合体等位基因。这两个肿瘤对吉非替尼均产生了明显和持久的反应。
在未经治疗的样本中检测到 EGFR T790M 嵌合体突变可能受到尚未解决的技术问题的影响，并且可能具有有限的临床价值。
