DNA methylation biomarkers for blood-based colorectal cancer screening
DNA methylation biomarkers for blood-based colorectal cancer screening
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DOI：
10.1373/clinchem.2007.095992
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发表时间：
2008-02-01
影响因子：
9.3
通讯作者：
Sledziewski, Andrew
中科院分区：
文献类型：
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作者：
Lofton-Day, Catherine;Model, Fabian;Sledziewski, Andrew
背景技术背景：除非在标记物发现和开发的每个阶段采取措施以最大限度地提高性能特征，否则难以开发敏感、特异的血液检测。我们描述了一种筛选策略，用于确定高性能的标记检测，检测结直肠癌（CRC）特异性甲基化DNA在plasma.METHODS：我们首先使用限制性内切酶为基础的发现方法，以确定标记候选人具有明显不同的甲基化模式在CRC组织和非病理组织。然后，我们使用结合微阵列和/或组织样品的实时PCR分析的选择过程来进一步测试标记候选物，以在CRC组织中实现最大甲基化，并在来自健康个体和患有其他疾病的患者的组织中实现最小扩增。实时检测3个选定的标志物进行了验证与血浆样本133 CRC患者和179名健康对照个体在相同的年龄范围。通过微阵列和实时PCR检测，该组减少到6例。用血浆样品测试三种标志物TMEFF 2、NGFR和SEPT 9。在65% [95%置信区间，56%-73%]的CRC患者血浆样本中检测到TMEFF 2甲基化，而在69%（62%-76%）的对照中未检测到。NGFR的相应结果分别为51%（42%-60%）和84%（77%-89%）; SEPT 9的相应值分别为69%（60%-77%）和86%（80%-91%）。结论：在选择和验证过程的所有步骤中应用的严格标准能够成功识别和排名基于血液的标记物候选物。(C)2007年美国临床化学协会。
BACKGROUND: Sensitive, specific blood-based tests are difficult to develop unless steps are taken to maximize performance characteristics at every stage of marker discovery and development. We describe a sieving strategy for identifying high-performing marker assays that detect colorectal cancer (CRC)-specific methylated DNA in plasma.METHODS: We first used restriction enzyme-based discovery methods to identify marker candidates with obviously different methylation patterns in CRC tissue and nonpathologic tissue. We then used a selection process incorporating microarrays and/or real-time PCR analysis of tissue samples to further test marker candidates for maximum methylation in CRC tissue and minimum amplification in tissues from both healthy individuals and patients with other diseases. Real-time assays of 3 selected markers were validated with plasma samples from 133 CRC patients and 179 healthy control individuals in the same age range.RESULTS: Restriction enzyme-based testing identified 56 candidate markers. This group was reduced to 6 with microarray and real-time PCR testing. Three markers, TMEFF2, NGFR, and SEPT9, were tested with plasma samples. TMEFF2 methylation was detected in 65% [95% confidence interval, 56%-73%] of plasma samples from CRC patients and not detected in 69% (62%-76%) of the controls. The corresponding results for NGFR were 5 1% (42%-60%) and 84% (77%-89%); for SEPT9, the values were 69% (60%-77%) and 86% (80%-91%).CONCLUSIONS: The stringent criteria applied at all steps of the selection and validation process enabled successful identification and ranking of blood-based marker candidates. (C) 2007 American Association for Clinical Chemistry.