Comparison of the SuperARMS and ARMS for detecting EGFR mutations in liquid-based cytology specimens from NSCLC patients
Comparison of the SuperARMS and ARMS for detecting EGFR mutations in liquid-based cytology specimens from NSCLC patients
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DOI：
10.1186/s13000-019-0910-5
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发表时间：
2020-01-31
影响因子：
2.6
通讯作者：
Wu, Chunyan
中科院分区：
文献类型：
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作者：
Wu, Wei;Cao, Ziyang;Wu, Chunyan
背景非手术细胞学标本不仅可用于准确的组织学亚型分型，而且可用于分子分型。通过优化引物设计，改进了扩增难治突变系统聚合酶链反应（ARMS PCR），即SuperARMS PCR，为血浆游离DNA检测提供了更高的灵敏度和特异性。目前尚不清楚SuperARMS PCR是否能检测细胞学样本中的表皮生长因子受体（EGFR）突变。本研究的目的是比较ARMS PCR和SuperARMS PCR检测晚期非小细胞肺癌（NSCLC）患者细胞学样本中EGFR突变的情况。方法2016年3月至2018年3月，收集234例NSCLC原发灶或转移灶细胞学标本，其中细针穿刺（FNAs）144例，支气管内超声（EBUS）FNAs 36例，经支气管针吸（TBNAs）36例，胸腔积液（PLEs）18例。使用ADx-ARMS EGFR试剂盒（Amoy Diagnostics CO.，有限公司，Xiamen，China）和ADx-SuperARMS EGFR试剂盒（Amoy Diagnostics CO.，有限公司，中国厦门）。进一步使用数字液滴PCR（ddPCR）和下一代测序（NGS）来验证EGFR突变体不一致的样品。结果234例晚期或复发性NSCLC患者均由两名细胞病理学家诊断和评估，ARMS和SuperARMS均能成功检测其EGFR突变状态。重要的是，SuperARMS和ARMS方法显示高度一致的结果，为94.0%（220/234）（95%CI：85.0，95.0%）。SuperARMS在细胞学样本EGFR检测中的阳性率高于ARMS（46.2% vs. 40.2%）。SuperARMS和ARMS检测的EGFR特异性突变位点不完全一致的有16例（6.8%）。当通过SuperARMS检测时，共有14例患者显示EGFR突变，但通过ARMS检测时，存在EGFR野生型。SuperARMS比ARMS检测到2例患者多一个EGFR突变位点。ddPCR和NGS用于进一步确认这些不一致样本中的EGFR突变。8个样本的突变结果与SuperARMS相同，6个样本因剩余DNA不足而未验证。共有78例EGFR突变患者接受酪氨酸激酶抑制剂（TKI）治疗。EGFR TKI治疗的总体客观缓解率（ORR）为88.5%（69/78）。结论SuperARMS检测EGFR具有较高的敏感性和特异性，有望成为临床上的常规检测方法，成为液基细胞学标本中EGFR突变检测的一种广泛、简便、灵敏的方法。
Background Non-surgical cytological specimens are adequate not only for accurate histological subtyping but also for molecular profiling. A modified amplification refractory mutation system polymerase chain reaction (ARMS PCR), known as SuperARMS PCR, was improved by optimizing the primers designation, which provides a higher sensitivity and specificity approach for free plasma DNA detection. It is unclear whether SuperARMS PCR detects epidermal growth factor receptor (EGFR) mutations in cytology samples. The aim of this study was to compare the EGFR mutations detected by ARMS PCR and SuperARMS PCR in cytology samples derived from advanced non-small cell lung cancer (NSCLC) patients. Methods From March 2016 to March 2018, a total of 234 cytological samples were obtained from primary or metastatic lesions of NSCLC, including 144 fine-needle aspirations (FNAs), 36 endobroncheal ultrasonography (EBUS) FNAs, 36 transbronchial needle aspirations (TBNAs) and 18 pleural effusion (PLEs). EGFR mutations were simultaneously detected using an ADx-ARMS EGFR kit (Amoy Diagnostics CO., ltd., Xiamen, China) and an ADx-SuperARMS EGFR kit (Amoy Diagnostics CO., ltd., Xiamen, China). Digital droplet PCR (ddPCR) and next-generation sequencing (NGS) were further used to verify the EGFR mutant inconsistent samples. Results All of the 234 patients with advanced or recurrent NSCLC were diagnosed and assessed by two cytopathologists, and their EGFR mutation statuses were successfully detected by ARMS and SuperARMS. Importantly, the SuperARMS and ARMS methods showed a highly concordant result of 94.0% (220/234) (95%CI: 85.0, 95.0%). The positive rate of the SuperARMS was higher than the ARMS in the cytology samples for EGFR detection (46.2% vs. 40.2%). The specific EGFR mutation sites in 16 samples (6.8%) were not completely consistent between the SuperARMS and ARMS. A total of 14 patients showed EGFR mutations when detected by SuperARMS, but by ARMS there were EGFR wild-type. Two patients were detected as having one more EGFR mutation site by SuperARMS than by ARMS. ddPCR and NGS were used to further confirm the EGFR mutations in these inconsistent samples. Eight samples had the same mutation results as the SuperARMS, and 6 samples were not verified because the remaining DNA was insufficient. A total of 78 EGFR mutation patients received Tyrosine Kinase Inhibitor (TKI) treatment. The overall objective response rate (ORR) was 88.5% (69/78) for EGFR TKI treatment. Conclusion SuperARMS showed a high sensitivity and specificity for EGFR detection and thus, is expected to become a routine test in the clinic to be used as a widely available, easy-to-operate and sensitive method for EGFR mutation detection in liquid-based cytology samples.