van Vliet Martin H, Burgmer Pia, de Quartel Linda, Brand Jaap P L, de Best Leonie C M, Viëtor Henk, Löwenberg Bob, Valk Peter J M, van Beers Erik H
Skyline Diagnostics BV, Rotterdam, The Netherlands.
Genet Test Mol Biomarkers. 2013 May;17(5):395-400. doi: 10.1089/gtmb.2012.0437. Epub 2013 Mar 13.
Double (bi-allelic) mutations in the gene encoding the CCAAT/enhancer-binding protein-alpha (CEBPA) transcription factor have a favorable prognostic impact in acute myeloid leukemia (AML). Double mutations in CEBPA can be detected using various techniques, but it is a notoriously difficult gene to sequence due to its high GC-content. Here we developed a two-step gene expression classifier for accurate and standardized detection of CEBPA double mutations. The key feature of the two-step classifier is that it explicitly removes cases with low CEBPA expression, thereby excluding CEBPA hypermethylated cases that have similar gene expression profiles as a CEBPA double mutant, which would result in false-positive predictions. In the second step, we have developed a 55 gene signature to identity the true CEBPA double-mutation cases. This two-step classifier was tested on a cohort of 505 unselected AML cases, including 26 CEBPA double mutants, 12 CEBPA single mutants, and seven CEBPA promoter hypermethylated cases, on which its performance was estimated by a double-loop cross-validation protocol. The two-step classifier achieves a sensitivity of 96.2% (95% confidence interval [CI] 81.1 to 99.3) and specificity of 100.0% (95% CI 99.2 to 100.0). There are no false-positive detections. This two-step CEBPA double-mutation classifier has been incorporated on a microarray platform that can simultaneously detect other relevant molecular biomarkers, which allows for a standardized comprehensive diagnostic assay. In conclusion, gene expression profiling provides a reliable method for CEBPA double-mutation detection in patients with AML for clinical use.
编码CCAAT/增强子结合蛋白α（CEBPA）转录因子的基因发生双（双等位基因）突变，对急性髓系白血病（AML）具有良好的预后影响。可以使用多种技术检测CEBPA中的双突变，但由于其高GC含量，该基因的测序非常困难。在此，我们开发了一种两步基因表达分类器，用于准确、标准化地检测CEBPA双突变。两步分类器的关键特征是它明确排除了CEBPA表达低的病例，从而排除了与CEBPA双突变体具有相似基因表达谱的CEBPA高甲基化病例，否则会导致假阳性预测。在第二步中，我们开发了一个55基因特征来识别真正的CEBPA双突变病例。该两步分类器在一组505例未经选择的AML病例中进行了测试，包括26例CEBPA双突变体、12例CEBPA单突变体和7例CEBPA启动子高甲基化病例，并通过双环交叉验证方案评估了其性能。两步分类器的灵敏度达到96.2%（95%置信区间[CI]81.1至99.3），特异性达到100.0%（95%CI 99.2至100.））。没有假阳性检测。这种两步CEBPA双突变分类器已被整合到一个微阵列平台上，该平台可以同时检测其他相关分子生物标志物，从而实现标准化的综合诊断检测。总之，基因表达谱分析为AML患者临床检测CEBPA双突变提供了一种可靠的方法。
