Highly sensitive method for genomewide detection of allelic composition in nonpaired, primary tumor specimens by use of affymetrix single-nucleotide-polymorphism genotyping microarrays
Highly sensitive method for genomewide detection of allelic composition in nonpaired, primary tumor specimens by use of affymetrix single-nucleotide-polymorphism genotyping microarrays
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DOI：
10.1086/518809
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发表时间：
2007-07-01
影响因子：
9.8
通讯作者：
Ogawa, Seishi
中科院分区：
文献类型：
--
作者：
Yamamoto, Go;Nannya, Yasuhito;Ogawa, Seishi
杂合性缺失（洛），伴随或不伴随拷贝数缺失，是癌症基因组的主要特征，与癌症的发展密切相关。然而，洛缺失的检测经常受到肿瘤标本中正常细胞成分的存在和组成型DNA可用性的限制的阻碍。在这里，我们描述了一种简单但高度灵敏的方法，用于全基因组检测等位基因的组成，基于Affyphase单核苷酸多态性基因分型微阵列平台，而不依赖于组成型DNA的可用性。通过使用匿名对照检测由等位基因不平衡引起的等位基因特异性信号的细微失真，即使在存在高达70%-80%的正常细胞污染的情况下，也能够通过精确测定等位基因特异性拷贝数来灵敏地检测洛。通过检测大量急性白血病样本中的拷贝数中性洛或单亲二体性（UPD），证明了称为“AsCNAR”（使用匿名参考的等位基因特异性拷贝数分析）的新算法的性能。我们接下来将该技术应用于检测骨髓增生性疾病（MPD）中涉及9 p臂的UPD，其与纯合JAK 2突变密切相关。它揭示了9 p UPD的意外高频率，否则将无法检测到，并且还揭示了在同一MPD标本中存在具有不同9 p UPD的多个亚群。总之，AsCNAR应该大大提高我们解剖癌症基因组复杂性的能力，并有助于我们理解人类癌症的遗传基础。
Loss of heterozygosity (LOH), either with or without accompanying copy-number loss, is a cardinal feature of cancer genomes that is tightly linked to cancer development. However, detection of LOH is frequently hampered by the presence of normal cell components within tumor specimens and the limitation in availability of constitutive DNA. Here, we describe a simple but highly sensitive method for genomewide detection of allelic composition, based on the Affymetrix single-nucleotide-polymorphism genotyping microarray platform, without dependence on the availability of constitutive DNA. By sensing subtle distortions in allele-specific signals caused by allelic imbalance with the use of anonymous controls, sensitive detection of LOH is enabled with accurate determination of allele-specific copy numbers, even in the presence of up to 70%-80% normal cell contamination. The performance of the new algorithm, called "AsCNAR" (allele-specific copy-number analysis using anonymous references), was demonstrated by detecting the copy-number neutral LOH, or uniparental disomy ( UPD), in a large number of acute leukemia samples. We next applied this technique to detection of UPD involving the 9p arm in myeloproliferative disorders ( MPDs), which is tightly associated with a homozygous JAK2 mutation. It revealed an unexpectedly high frequency of 9p UPD that otherwise would have been undetected and also disclosed the existence of multiple subpopulations having distinct 9p UPD within the same MPD specimen. In conclusion, AsCNAR should substantially improve our ability to dissect the complexity of cancer genomes and should contribute to our understanding of the genetic basis of human cancers.