Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei, Taiwan; Department of Obstetrics and Gynecology, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
Int J Cancer. 2014 Jul 1;135(1):117-27. doi: 10.1002/ijc.28658. Epub 2013 Dec 17.
Using DNA methylation biomarkers in cancer detection is a potential direction in clinical testing. Some methylated genes have been proposed for cervical cancer detection; however, more reliable methylation markers are needed. To identify new hypermethylated genes in the discovery phase, we compared the methylome between a pool of DNA from normal cervical epithelium (n = 19) and a pool of DNA from cervical cancer tissues (n = 38) using a methylation bead array. We integrated the differentially methylated genes with public gene expression databases, which resulted in 91 candidate genes. Based on gene expression after demethylation treatment in cell lines, we confirmed 61 genes for further validation. In the validation phase, quantitative MSP and bisulfite pyrosequencing were used to examine their methylation level in an independent set of clinical samples. Fourteen genes, including ADRA1D, AJAP1, COL6A2, EDN3, EPO, HS3ST2, MAGI2, POU4F3, PTGDR, SOX8, SOX17, ST6GAL2, SYT9, and ZNF614, were significantly hypermethylated in CIN3+ lesions. The sensitivity, specificity, and accuracy of POU4F3 for detecting CIN3+ lesions were 0.88, 0.82, and 0.85, respectively. A bioinformatics function analysis revealed that AJAP1, EDN3, EPO, MAGI2, and SOX17 were potentially implicated in β-catenin signaling, suggesting the epigenetic dysregulation of this signaling pathway during cervical cancer development. The concurrent methylation of multiple genes in cancers and in subsets of precancerous lesions suggests the presence of a driver of methylation phenotype in cervical carcinogenesis. Further validation of these new genes as biomarkers for cervical cancer screening in a larger population-based study is warranted.
使用 DNA 甲基化生物标志物进行癌症检测是临床检测的一个潜在方向。一些甲基化基因已被提出用于宫颈癌检测，但需要更可靠的甲基化标记物。为了在发现阶段识别新的高甲基化基因，我们使用甲基化珠阵列比较了一组来自正常宫颈上皮（n=19）的 DNA 和一组来自宫颈癌组织（n=38）的 DNA 的甲基组。我们将差异甲基化基因与公共基因表达数据库进行整合，结果得到 91 个候选基因。基于细胞系中去甲基化处理后的基因表达，我们确认了 61 个基因进行进一步验证。在验证阶段，我们使用定量 MSP 和焦磷酸测序检测了这些基因在独立的临床样本中的甲基化水平。在验证阶段，我们使用定量 MSP 和焦磷酸测序检测了这些基因在独立的临床样本中的甲基化水平。包括 ADRA1D、AJAP1、COL6A2、EDN3、EPO、HS3ST2、MAGI2、POU4F3、PTGDR、SOX8、SOX17、ST6GAL2、SYT9 和 ZNF614 在内的 14 个基因在 CIN3+病变中显著高甲基化。POU4F3 检测 CIN3+病变的敏感性、特异性和准确性分别为 0.88、0.82 和 0.85。生物信息学功能分析显示，AJAP1、EDN3、EPO、MAGI2 和 SOX17 可能与β-连环蛋白信号有关，提示在宫颈癌发展过程中，该信号通路存在表观遗传失调。癌症和癌前病变亚组中多个基因的同时甲基化表明，在宫颈癌发生过程中存在甲基化表型的驱动因素。在更大的基于人群的研究中进一步验证这些新基因作为宫颈癌筛查的生物标志物是必要的。
