Background: The patients with advanced lung adenocarcinoma should select targeted drugs based on the type of tumor epidermal growth factor receptor (EGFR) gene mutation. However, it is difficult to collect tumor tissue of advanced lung adenocarcinoma, and some experts agree that peripheral blood can be used as a substitute for tumor tissue as a test specimen. This paper aimed to investigate the clinical value of ddPCR and super-amplification refractory mutation system (ARMS) in detecting EGFR gene mutation in peripheral blood of patients with advanced lung adenocarcinoma.
Methods: A total of 119 patients diagnosed in Beijing Chest Hospital Affiliated to Capital Medical University from February 2016 to February 2019 were collected, and the sensitivity and specificity of plasma ctDNA EGFR gene mutation detected by ddPCR and super-arms were compared. Some patients with positive EGFR gene mutations received oral treatment with first-line EGFR tyrosine kinase inhibitors (EGFR-TKI). The patients were divided into subgroups according to the test results. In group 1, both ddPCR and super-arms showed positive EGFR gene mutation results, with 21 cases. In group 2, ddPCR and super-arms detection of EGFR gene mutation were all negative, with 16 cases. In group 3, the ddPCR test was positive and the super-arms test was negative, with 5 cases. In group 4, the ddPCR test result was negative while the super-arms test result was positive. Since the number of patients in group 4 was 0, no statistics were included. Objective response rate (ORR) and disease control rate (DCR) were used to evaluate the short-term outcome, and progression-free survival (PFS) was compared with survival analysis to evaluate the long-term outcome.
Results: EGFR mutations were detected in 58 (48.7%) of 119 patients with advanced lung adenocarcinoma. The coincidence rate between ddPCR and EGFR gene mutation in tumor tissues was 82.4% (Kappa=0.647, P<0.001), the sensitivity was 74.1%, and the specificity was 90.2%. However, the coincidence degree of super-arms test and tissue test was 71.4%, the sensitivity was only 58.6%, and the specificity was 83.6%. The ORR and DCR values in group 3 were lower than those in group 1 and 2, but there was no significant difference in ORR between groups (P>0.05). Survival analysis showed that the PFS of the three groups was compared. The difference was not statistically significant (χ²=2.221, P=0.329).
Conclusions: ddPCR, as a high sensitivity and specificity liquid gene detection method, can be used as a reliable method to detect the mutation of plasma ctDNA EGFR gene in patients with advanced lung adenocarcinoma. The results of plasma genetic testing can also be used as the basis for predicting the efficacy of EGFR-TKIs in patients.
【中文题目：微滴数字PCR和Super-ARMS检测晚期肺腺癌患者血浆循环肿瘤DNA表皮生长因子受体基因突变的临床价值研究】 【中文摘要：背景与目的 晚期肺腺癌患者在选择靶向药物时需以肿瘤表皮生长因子受体（epidermal growth factor receptor, EGFR）基因突变类型作为依据，然而晚期肺腺癌肿瘤组织取材较难，有专家共识指出外周血可替代肿瘤组织作为检测标本。本文旨在探讨微滴数字聚合酶链反应法（droplet digital polymerase chain reaction, ddPCR）和超级扩增阻滞突变系统（super-amplification refractory mutation system, super-ARMS）这两种方法检测晚期肺腺癌患者外周血中血浆循环肿瘤脱氧核糖核酸（circulating tumor DNA, ctDNA）中EGFR基因突变的临床价值。方法 收集2016年2月-2019年2月首都医科大学附属北京胸科医院确诊的初治晚期肺腺癌共119例纳入研究，比较ddPCR和Super-ARMS技术检测血浆ctDNA EGFR基因突变的敏感度、特异度。部分EGFR基因经典突变阳性患者接受一线EGFR酪氨酸激酶抑制剂（EGFR tyrosine kinase inhibitors, EGFR-TKI）口服治疗，将患者按检测结果分亚组，组1为ddPCR及Super-ARMS检测EGFR基因突变结果均为阳性，21例。组2为ddPCR及Super-ARMS检测EGFR基因突变结果均为阴性，16例。组3为ddPCR检测结果为阳性而Super-ARMS检测结果为阴性，5例。组4为ddPCR检测结果为阴性而Super-ARMS检测结果阳性，因组4患者个数为0，不纳入统计。以客观缓解率（objective response rate, ORR）及疾病控制率（disease control rate, DCR）评估近期疗效，并用生存分析比较组间无进展生存时间（progression-free survival, PFS）以评估远期疗效。结果 119例晚期肺腺癌组织样本中，共检测到58例（48.7%）EGFR基因突变。ddPCR技术检测与肿瘤组织EGFR基因突变结果的符合率为82.4%（Kappa=0.647, P<0.001），灵敏度为74.1%，特异度为90.2%。而Super-ARMS检测与组织检测的符合度为71.4%，灵敏度仅为58.6%，特异度为83.6%。组3的ORR与DCR值低于组1、2，但组间ORR相比均无统计学意义（P>0.05）。生存分析显示，三组患者PFS比较，差异无统计学意义（χ²=2.221, P=0.329）。结论 ddPCR作为一种高敏感度及特异度的液体基因检测方法，可以作为检测晚期肺腺癌患者血浆ctDNA EGFR基因突变的一种较为可靠的方法。血浆基因检测结果同样可作为EGFR-TKIs药物对患者的疗效预测的依据。】 【中文关键词：肺肿瘤；微滴数字聚合酶链反应；表皮生长因子受体；基因突变；循环肿瘤脱氧核糖核酸；突变扩增系统；Super-突变扩增系统】.
Keywords: Circulating tumor DNA; Droplet digital polymerase chain reaction; Epidermal growth factor receptor; Genetic mutations; Lung tumor; Mutation amplification system; Super-mutation amplification system.