粪便DNA中SFRP2启动子甲基化检测的原型反向杂交分析与甲基化特异性荧光定量PCR法的比较

Comparison of a prototype reverse hybridization assay and MethyLight for detection of SFRP2 promotor methylation in fecal DNA.

作者信息

Kriegshäuser Gernot, Enko Dietmar, Zitt Matthias, Oberwalder Michael, Oberkanins Christian, Öfner Dietmar, Zeillinger Robert, Maximilian Müller Hannes

机构信息

Institute of Clinical Chemistry and Laboratory Medicine, General Hospital Steyr, Steyr - Austria.

Clinical Institute of Medical and Laboratory Diagnostics, Medical University of Graz, Graz - Austria.

出版信息

Int J Biol Markers. 2017 Oct 31;32(4):e467-e470. doi: 10.5301/ijbm.5000289.

DOI:10.5301/ijbm.5000289

PMID:28777432

Abstract

BACKGROUND

This study aimed to evaluate the diagnostic performance of a novel nonquantitative methylation-specific reverse hybridization (MSRH) assay to detect secreted frizzled-related protein 2 (SFRP2) promotor methylation in fecal DNA.

METHODS

SFRP2 promoter methylation was investigated in stool DNA isolated from 18 colorectal cancer (CRC) patients and 22 healthy controls using the MSRH assay based on methylation-specific DNA amplification followed by reverse hybridization of biotinylated amplicons to sequence-specific methylation detection probes, with MethyLight serving as a reference method.

RESULTS

SFRP2 promotor methylation as determined by MSRH vs. MethyLight showed a sensitivity and specificity of 61.1% and 86.3% vs. 77.7% and 77.3%, respectively. Moderate agreement (ĸ = 0.54, 95% confidence interval [95% CI], 0.29-0.80, p<0.001) was observed between the 2 methods. However, the differences in SFRP2 promotor methylation observed between CRC patients and healthy individuals by both assays were statistically significant (p<0.001).

CONCLUSIONS

Our findings, although limited by the small sample size, do not support the use of the MSRH assay for CRC screening in stool.

摘要

背景

本研究旨在评估一种新型非定量甲基化特异性反向杂交（MSRH）检测法在粪便DNA中检测分泌型卷曲相关蛋白2（SFRP2）启动子甲基化的诊断性能。

方法

采用基于甲基化特异性DNA扩增的MSRH检测法，随后将生物素化扩增子与序列特异性甲基化检测探针进行反向杂交，以甲基化荧光定量法（MethyLight）作为参考方法，对18例结直肠癌（CRC）患者和22例健康对照者的粪便DNA中的SFRP2启动子甲基化进行研究。

结果

MSRH法与MethyLight法检测SFRP2启动子甲基化的灵敏度和特异性分别为61.1%和86.3%，以及77.7%和77.3%。两种方法之间观察到中度一致性（ĸ = 0.54，95%置信区间[95%CI]，0.29 - 0.80，p<0.001）。然而，两种检测方法在CRC患者和健康个体之间观察到的SFRP2启动子甲基化差异具有统计学意义（p<0.001）。