Beijing Obstetrics and Gynecology Hospital, Capital Medical University, Beijing 100026, PR China.
Oncol Rep. 2013 Jan;29(1):125-32. doi: 10.3892/or.2012.2077. Epub 2012 Oct 9.
The aim of the present study was to identify novel methylation markers for cervical cancer screening and to test the clinical application of the most promising biomarker in cervical scrapings. Methylated-CpG island recovery assay-based microarray analysis was carried out on a discovery set consisting of cervical cancer tissue and normal cervical tissue to identify significantly hypermethylated genes. Five hundred and four CpG islands, corresponding to 378 genes, were differentially methylated between cervical cancer tissue and normal cervical tissue. Among them, 30 genes were significantly hypermethylated. Of the 30 genes, SOX9, PKLR and DLX4 were selected for further validation by direct bisulfite sequencing. The SOX9 gene revealed complete methylation in the cervical cancer tissue and complete non-methylation in the normal control tissue. A TaqMan-based real-time PCR assay was performed to detect the methylation levels of the SOX9 gene in 156 cervical scrapings, including 48 normal cervical scrapings, 30 scrapings with cervical intraepithelial neoplasia 1 (CIN1), 30 scrapings with CIN2-3 and 48 scrapings with squamous cell carcinoma (SCC). The methylation levels (methylation score) of the SOX9 gene increased significantly with the severity of cervical squamous lesions. The area under the receiver operating characteristic (ROC) curve (AUC) revealed that the methylation score of the SOX9 gene could be used to segregate SCC/CIN2-3 from CIN1/normal (AUC, 0.961; p=0.000). At the optimal cut-off value, a sensitivity of 92.3% and a specificity of 89.7% were obtained. In conclusion, SOX9 methylation is frequently involved in cervical carcinogenesis, and may provide a valuable molecular biomarker for early detection of cervical cancer.
本研究旨在寻找新的宫颈癌筛查甲基化标志物，并在宫颈刮片中检测最有前途的生物标志物的临床应用。采用基于甲基化-CpG 岛回收分析的微阵列分析方法，对宫颈癌组织和正常宫颈组织的发现集进行分析，以确定显著过甲基化的基因。在宫颈癌组织和正常宫颈组织之间，有 504 个 CpG 岛（对应 378 个基因）存在差异甲基化。其中，有 30 个基因显著过甲基化。在这 30 个基因中，SOX9、PKLR 和 DLX4 被选为进一步通过直接亚硫酸氢盐测序验证的基因。SOX9 基因在宫颈癌组织中完全甲基化，在正常对照组织中完全非甲基化。采用 TaqMan 实时 PCR 检测了 156 例宫颈刮片中 SOX9 基因的甲基化水平，其中包括 48 例正常宫颈刮片、30 例宫颈上皮内瘤变 1 级（CIN1）、30 例 CIN2-3 和 48 例鳞状细胞癌（SCC）。SOX9 基因的甲基化水平（甲基化评分）随着宫颈鳞状病变的严重程度显著增加。接收者操作特征（ROC）曲线下面积（AUC）显示，SOX9 基因的甲基化评分可用于将 SCC/CIN2-3 与 CIN1/正常区分开（AUC，0.961；p=0.000）。在最佳截断值时，获得了 92.3%的敏感性和 89.7%的特异性。总之，SOX9 甲基化频繁参与宫颈癌的发生，可能为宫颈癌的早期检测提供有价值的分子生物标志物。
