deVos Theo, Tetzner Reimo, Model Fabian, Weiss Gunter, Schuster Matthias, Distler Jürgen, Steiger Kathryn V, Grützmann Robert, Pilarsky Christian, Habermann Jens K, Fleshner Phillip R, Oubre Benton M, Day Robert, Sledziewski Andrew Z, Lofton-Day Catherine
Epigenomics Inc., Seattle, WA 98101, USA.
Clin Chem. 2009 Jul;55(7):1337-46. doi: 10.1373/clinchem.2008.115808. Epub 2009 Apr 30.
The presence of aberrantly methylated SEPT9 DNA in plasma is highly correlated with the occurrence of colorectal cancer. We report the development of a new SEPT9 biomarker assay and its validation in case-control studies. The development of such a minimally invasive blood-based test may help to reduce the current gap in screening coverage.
A new SEPT9 DNA methylation assay was developed for plasma. The assay comprised plasma DNA extraction, bisulfite conversion of DNA, purification of bisulfite-converted DNA, quantification of converted DNA by real-time PCR, and measurement of SEPT9 methylation by real-time PCR. Performance of the SEPT9 assay was established in a study of 97 cases with verified colorectal cancer and 172 healthy controls as verified by colonoscopy. Performance based on predetermined algorithms was validated in an independent blinded study with 90 cases and 155 controls.
The SEPT9 assay workflow yielded 1.9 microg/L (CI 1.3-3.0) circulating plasma DNA following bisulfite conversion, a recovery of 45%-50% of genomic DNA, similar to yields in previous studies. The SEPT9 assay successfully identified 72% of cancers at a specificity of 93% in the training study and 68% of cancers at a specificity of 89% in the testing study.
Circulating methylated SEPT9 DNA, as measured in the new (m)SEPT9 assay, is a valuable biomarker for minimally invasive detection of colorectal cancer. The new assay is amenable to automation and standardized use in the clinical laboratory.
血浆中异常甲基化的SEPT9 DNA的存在与结直肠癌的发生高度相关。我们报告了一种新的SEPT9生物标志物检测方法的开发及其在病例对照研究中的验证。开发这种基于血液的微创检测方法可能有助于缩小目前筛查覆盖范围的差距。
开发了一种用于血浆的新型SEPT9 DNA甲基化检测方法。该检测方法包括血浆DNA提取、DNA的亚硫酸氢盐转化、亚硫酸氢盐转化后DNA的纯化、通过实时PCR对转化后DNA进行定量以及通过实时PCR测量SEPT9甲基化。在一项对97例经结肠镜检查证实患有结直肠癌的病例和172例健康对照的研究中确定了SEPT9检测方法的性能。在一项对90例病例和155例对照的独立盲法研究中，基于预定算法的性能得到了验证。
SEPT9检测流程在亚硫酸氢盐转化后产生了1.9微克/升（置信区间1.3 - 3.0）的循环血浆DNA，基因组DNA回收率为45% - 50%，与先前研究的产量相似。在训练研究中，SEPT9检测方法以93%的特异性成功识别出72%的癌症，在测试研究中以89%的特异性成功识别出68%的癌症。
新的（m）SEPT9检测方法所测量的循环甲基化SEPT9 DNA是用于结直肠癌微创检测的有价值的生物标志物。这种新检测方法适用于临床实验室的自动化和标准化使用。
