A Platform for Rapid Detection of Multiple Oncogenic Mutations With Relevance to Targeted Therapy in Non-Small-Cell Lung Cancer
A Platform for Rapid Detection of Multiple Oncogenic Mutations With Relevance to Targeted Therapy in Non-Small-Cell Lung Cancer
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DOI：
10.1016/j.jmoldx.2010.11.010
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发表时间：
2011-01-01
影响因子：
4.1
通讯作者：
Pao, William
中科院分区：
文献类型：
--
作者：
Su, Zengliu;Dias-Santagata, Dora;Pao, William
体细胞获得性肿瘤突变的识别在癌症的临床治疗中变得越来越重要，因为靶向药物的敏感性与个体肿瘤的基因组成有关。因此，肿瘤的突变谱可以帮助优先考虑抗癌治疗。我们在此报告了两种多重检测的开发和验证，该检测旨在检测 FFPE 组织 DNA 中与现有和新兴肺癌靶向治疗相关的 9 个基因中的 40 多个反复突变。该平台涉及两种方法：基于多重 PCR、引物延伸和毛细管电泳的筛选 (SNaPshot)，旨在评估 8 个基因（AKT1、BRAF、EGFR、KRAS、MEK1、NRAS、PIK3C4 和 PTEN）的 38 个体细胞突变；以及基于 PCR 的大小测定，用于评估 EGFR 外显子 19 缺失、EGFR 外显子 20 插入和 HER2 外显子20 个插入。 SNaPshot 和大小测定都可以快速进行，并且使用最少量的遗传物质。与直接测序相比，突变DNA需要占总DNA的25%或更多才能轻松检测到突变，SNaPshot和尺寸分析可以分别检测突变DNA占总DNA的1.56%至12.5%和1.56%至6.25%的样本中的突变。这些稳健、可靠且相对便宜的检测方法应有助于加速基因型驱动方法在肺癌治疗中的采用。 (J Mol Diagn 2011, 13:74-84; DOI: 10.1016/j.jmoldx.2010.11.010)
The identification of somatically acquired tumor mutations is increasingly important in the clinical management of cancer because the sensitivity of targeted drugs is related to the genetic makeup of individual tumors. Thus, mutational profiles of tumors can help prioritize anticancer therapy. We report herein the development and validation of two multiplexed assays designed to detect in DNA from FFPE tissue more than 40 recurrent mutations in nine genes relevant to existing and emerging targeted therapies in lung cancer. The platform involves two methods: a screen (SNaPshot) based on multiplex PCR, primer extension, and capillary electrophoresis that was designed to assess for 38 somatic mutations in eight genes (AKT1, BRAF, EGFR, KRAS, MEK1, NRAS, PIK3C4, and PTEN) and a PCR-based sizing assay that assesses for EGFR exon 19 deletions, EGFR exon 20 insertions, and HER2 exon 20 insertions. Both the SNaPshot and sizing assays can be performed rapidly, with minimal amounts of genetic material. Compared with direct sequencing, in which mutant DNA needs to compose 25% or more of the total DNA to easily detect a mutation, the SNaPshot and sizing assays can detect mutations in samples in which mutant DNA composes 1.56% to 12.5% and 1.56% to 6.25% of the total DNA, respectively. These robust, reliable, and relatively inexpensive assays should help accelerate adoption of a genotype-driven approach in the treatment of lung cancer. (J Mol Diagn 2011, 13:74-84; DOI: 10.1016/j.jmoldx.2010.11.010)