KRAS mutation analysis on low percentage of colon cancer cells: the importance of quality assurance
KRAS mutation analysis on low percentage of colon cancer cells: the importance of quality assurance
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DOI：
10.1007/s00428-012-1356-2
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发表时间：
2013-01-01
期刊：
影响因子：
3.5
通讯作者：
van Krieken, J. H. J. M.
中科院分区：
文献类型：
--
作者：
Dijkstra, J. R.;Heideman, D. A. M.;van Krieken, J. H. J. M.
KRAS突变检测对于有资格接受表皮生长因子受体靶向药物治疗的转移性结直肠癌患者是强制性的，因为具有突变的肿瘤对药物不敏感。目前正在使用几种突变检测方法，并已证明需要外部质量保证。在外部质量保证计划中，一个经常很少解决但很重要的问题是测试样品中肿瘤细胞的百分比很低，其中大多数测试的分析灵敏度变得至关重要。使用基于具有已知KRAS基因突变状态的细胞系混合物的人工样本，我们评估了一系列常用方法（桑格测序、高分辨率解链、焦磷酸测序和扩增难治性突变系统-聚合酶链反应）对具有0、2.5、5、10和15%突变细胞的样本的可靠性。整个欧洲的九个实验室参加了这次活动，共提交了十个数据集。每种方法的检测限不同，范围为> 15-5%肿瘤细胞。所有方法均显示出随着肿瘤细胞百分比的降低而成比例地降低的正确突变调用率。我们的研究结果表明，实验室和临床医生需要意识到正确突变检出率的下降与肿瘤细胞百分比的下降成比例，外部质量保证计划需要解决测试样本中肿瘤细胞百分比低的问题。
KRAS mutation testing is mandatory for patients with metastatic colorectal cancer who are eligible for treatment with an epidermal growth factor receptor targeting agent, since tumors with a mutation are not sensitive to the drug. Several methods for mutation testing are in use and the need for external quality assurance has been demonstrated. An often little addressed but important issue in external quality assurance schemes is a low percentage of tumor cells in the test samples, where the analytical sensitivity of most tests becomes critical. Using artificial samples based on a mixture of cell lines with known mutation status of the KRAS gene, we assessed the reliability of a series of commonly used methods (Sanger sequencing, high resolution melting, pyrosequencing, and amplification refractory mutation system-polymerase chain reaction) on samples with 0, 2.5, 5, 10, and 15% mutated cells. Nine laboratories throughout Europe participated and submitted a total of ten data sets. The limit of detection of each method differed, ranging from > 15-5% tumor cells. All methods showed a decreasing correct mutation call rate proportionally with decreasing percentage of tumor cells. Our findings indicate that laboratories and clinicians need to be aware of the decrease in correct mutation call rate proportionally with decreasing percentage of tumor cells and that external quality assurance schemes need to address the issue of low tumor cell percentage in the test samples.