Nanofluidic Digital PCR for KRAS Mutation Detection and Quantification in Gastrointestinal Cancer
Nanofluidic Digital PCR for KRAS Mutation Detection and Quantification in Gastrointestinal Cancer
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DOI：
10.1373/clinchem.2012.186577
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发表时间：
2012-09-01
影响因子：
9.3
通讯作者：
Capella, Gabriel
中科院分区：
文献类型：
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作者：
Azuara, Daniel;Ginesta, Mireia M.;Capella, Gabriel
背景：多个突变的KRAS(v-ki-ras2 Kirsten鼠肉瘤病毒癌基因同源)等位基因的同时定量可能提供标准突变检测程序所提供的信息之外的信息。方法：我们评估了两组患者(结直肠癌和胰腺疾病)：第一组包括27例结直肠癌患者、14例腺瘤患者和5名对照组；第二组患者包括42例胰腺癌患者、4例壶腹腺癌患者和6例慢性胰腺炎患者。结果：纳米流体数字聚合酶链式反应可检测到0.05%~0.10%的突变等位基因，具体取决于所分析的突变类型。在结直肠疾病组，常规聚合酶链式反应检测到14例腺瘤中KRAS突变阳性者9例(%)，而数字聚合酶链式反应将这一数字增加到L L(79%)14例。用数字聚合酶链式反应检测到另外两例。5例(腺瘤3例，癌2例)突变等位基因总数发生改变。对于胰腺疾病组，数字聚合酶链式反应将阳性例数从26例增加到34例(81%)，并在25例患者中发现了>；=2个突变等位基因，而常规聚合酶链式反应仅在12例患者中发现多个KRAS突变等位基因。肿瘤活检和胰液检测结果之间有很好的相关性。结论：数字聚合酶链式反应为常规标本中KRAS突变等位基因的比例提供了一种可靠的、定量的测量方法。它还可以对肿瘤进行更好的分类，具有潜在的临床相关性。(C)2012年美国临床化学协会
BACKGROUND: Concomitant quantification of multiple mutant KRAS (v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog) alleles may provide information in addition to that provided by standard mutation-detection procedures. We assessed the feasibility of a nanofluidic digital PCR array platform to detect and quantify KRAS mutations simultaneously in clinically relevant samples.METHODS: We assessed 2 groups of patients (colorectal and pancreatic disease): Group 1 consisted of 27 patients with colorectal carcinomas, 14 patients with adenomas, and 5 control individuals; group 2 consisted of 42 patients with pancreatic carcinoma, 4 with adenocarcinomas of the ampulla, and 6 with chronic pancreatitis). Digital PCR was performed with the Digital Array Chip (Fluidigm).RESULTS: Nanofluidic digital PCR detected mutant alleles at 0.05% to 0.10%, depending on the variant analyzed. For the colorectal disease group, conventional PCR detected 9 (64%) of 14 adenomas that were positive for KRAS mutants, whereas digital PCR increased this number to l l (79%) of 14. Sixteen (59%) of 27 carcinomas showed KRAS mutation with conventional PCR. Two additional cases were detected with digital PCR. In 5 cases (3 adenomas, 2 carcinomas), the total number of mutant alleles changed. For the pancreatic disease group, digital PCR increased the number of positive cases from 26 to 34 (81%) and identified >= 2 mutant alleles in 25 cases, compared with conventional PCR, which identified multiple KRAS mutant alleles in only 12 cases. A good correlation was observed between results obtained with tumor biopsies and those obtained with pancreatic juice.CONCLUSIONS: Digital PCR provides a robust, quantitative measure of the proportion of KRAS mutant alleles in routinely obtained samples. It also allows a better classification of tumors, with potential clinical relevance. (C) 2012 American Association for Clinical Chemistry