Wertheim Gerald B W, Smith Catherine, Luskin Marlise, Rager Alison, Figueroa Maria E, Carroll Martin, Master Stephen R
Department of Pathology, Children's Hospital of Philadelphia; Department of Pathology and Laboratory Medicine.
Department of Pathology, Children's Hospital of Philadelphia;
Clin Chem. 2015 Jan;61(1):249-58. doi: 10.1373/clinchem.2014.229781. Epub 2014 Oct 27.
Epigenetic dysregulation involving alterations in DNA methylation is a hallmark of various types of cancer, including acute myeloid leukemia (AML). Although specific cancer types and clinical aggressiveness of tumors can be determined by DNA methylation status, the assessment of DNA methylation at multiple loci is not routinely performed in the clinical laboratory.
We recently described a novel microsphere-based assay for multiplex evaluation of DNA methylation. In the current study, we validated and used an improved assay [termed expedited microsphere HpaII small fragment Enrichment by Ligation-mediated PCR (xMELP)] that can be performed with appropriate clinical turnaround time.
Using the xMELP assay in conjunction with a new 17-locus random forest classifier that has been trained using 344 AML samples, we were able to segregate an independent cohort of 70 primary AML patients into methylation-determined subgroups with significantly distinct mortality risk (P = 0.009). We also evaluated precision, QC parameters, and preanalytic variables of the xMELP assay and determined the sensitivity of the random forest classifier score to failure at 1 or more loci.
Our results demonstrate that xMELP performance is suitable for implementation in the clinical laboratory and predicts AML outcome in an independent patient cohort.
涉及DNA甲基化改变的表观遗传失调是包括急性髓系白血病（AML）在内的各种癌症的一个标志。尽管特定癌症类型和肿瘤的临床侵袭性可由DNA甲基化状态决定，但临床实验室通常不会对多个位点的DNA甲基化进行评估。
我们最近描述了一种基于微球的新型DNA甲基化多重评估检测方法。在本研究中，我们验证并使用了一种改进的检测方法[称为连接介导PCR快速微球HpaII小片段富集法（xMELP）]，该方法可在适当的临床周转时间内完成。
使用xMELP检测方法结合一个新的17位点随机森林分类器（该分类器已使用344个AML样本进行训练），我们能够将70例原发性AML患者的独立队列分为甲基化确定的亚组，这些亚组的死亡风险显著不同（P = 0.009）。我们还评估了xMELP检测方法的精密度、质量控制参数和分析前变量，并确定了随机森林分类器评分对1个或更多位点失败的敏感性。
我们的结果表明，xMELP的性能适合在临床实验室实施，并可预测独立患者队列中的AML预后。
