An accurate and rapid detection of lymph node metastasis in non-small cell lung cancer patients based on one-step nucleic acid amplification assay
An accurate and rapid detection of lymph node metastasis in non-small cell lung cancer patients based on one-step nucleic acid amplification assay
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DOI：
10.1016/j.lungcan.2012.08.018
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发表时间：
2012-12-01
期刊：
影响因子：
5.3
通讯作者：
Okumura, Meinoshin
中科院分区：
文献类型：
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作者：
Inoue, Masayoshi;Hiyama, Kayo;Okumura, Meinoshin
叶下切除术目前被认为是早期小尺寸非小细胞肺癌（NSCLC）的一种选择，术中快速准确的淋巴结评估是完全切除的必要条件。为了解决这一问题，我们研究了一步核酸扩增（one-step nucleic acid amplification， OSNA）技术——一种自动快速分子诊断方法及其最佳mRNA标记物在肺癌淋巴结转移检测中的临床应用。我们从基因数据库中提取了16个在肺癌中高表达的目标候选mRNA标记物，然后用手术切除的有或没有转移的淋巴结，用定量RT-PCR方法量化了它们的表达水平。在定量rt - pcr筛选中，细胞角蛋白19 （CK19）、细胞角蛋白7 （CK7）、层结蛋白（SFN）和前梯度同源物2 (AGR2) mRNA在转移阴性和阳性淋巴结中的表达差异有统计学意义。最终选择CK19和CK7作为潜在的靶标记物，并利用49例肺癌患者的165个淋巴结的OSNA检测结果进行量化。CK19和CK7的OSNA检测在40分钟内完成，与苏木精-伊红染色和免疫组织化学的病理诊断相比，其阳性预测值、阴性预测值和准确性分别为95.0%、99.3%和98.8%,85.0%、97.9%和96.4%，使用截断值为250拷贝/ μ l。1例假阴性是由于癌细胞大量坏死造成的，1例假阳性是由于胸膜播散患者取样中癌细胞分配偏倚造成的。以CK19作为标记时表现最佳，而添加CK7 mRNA作为标记并未增加敏感性或特异性。综上所述，基于CK19的OSNA检测可有效地用于肺癌淋巴结转移的分子诊断。这是第一个提示OSNA检测在术中快速诊断肺癌淋巴结状态的潜在临床应用的报告。2012爱思唯尔爱尔兰有限公司版权所有。
A sublobar resection is currently recognized as an option for early small-sized non-small cell lung cancer (NSCLC), and intraoperative rapid and accurate lymph node assessment is required for a complete resection. To solve this issue, we investigated the clinical utility of one-step nucleic acid amplification (OSNA) assay, an automated rapid molecular diagnostic method and its optimal mRNA marker for detection of lymph node metastasis in lung cancer. We extracted 16 target candidate mRNA markers with high expression in lung cancer from a genetic database, and then quantified their expression levels by quantitative RT-PCR using surgically dissected lymph nodes with or without metastasis. Cytokeratin 19 (CK19), cytokeratin 7 (CK7), stratifin (SFN), and anterior gradient homolog 2 (AGR2) showed significant differences for mRNA expression between metastasis-negative and -positive lymph nodes in quantitative-RT-PCR screening. CK19 and CK7 were finally selected as potential target markers and were quantified using OSNA assay findings of 165 dissected lymph nodes obtained from 49 lung cancer patients. The OSNA assay with CK19 and CK7 were completed within 40 min and their positive predictive value, negative predictive value, and accuracy comparing to pathological diagnosis with hematoxylin-eosin staining and immuno-histochemistry were shown to be 95.0%, 99.3%, and 98.8%, and 85.0%, 97.9%, and 96.4%, respectively, using a cut-off value of 250 copies/mu L. Among the 165 lymph nodes tested, 1 false negative result was due to massive necrosis of cancer cells and 1 false positive was caused by the allocation bias of cancer cells in the sampling in patient with pleural dissemination. The best performance was observed when CK19 was used as a marker, while the addition of CK7 mRNA as a marker did not increase sensitivity or specificity. In conclusion, an OSNA assay using CK19 could be effective for molecular diagnosis of lymph node metastasis in lung cancer. This is the first report suggesting the potential clinical utility of OSNA assay for intraoperative rapid diagnosis of nodal status in lung cancer. (c) 2012 Elsevier Ireland Ltd. All rights reserved.