Abstract:
Cervical cancer is the fourth most common cancer in women, which is associated in >95% with a high-risk human papillomavirus (HPV) infection. Methylation of specific genes has been closely associated with the progress of cervical high-grade dysplastic lesions to invasive carcinomas. Therefore, DNA methylation has been proposed as a triage for women infected with high-risk HPV. Methylation analyses of cervical cancer tissue have shown that cell adhesion molecule 1 (CADM1) and myelin and lymphocyte protein (MAL) methylation are present in over 90% of all cervical high-grade neoplasias and invasive cervical cancers. Here, we established a liquid biopsy-based assay to detect MAL and CADM1 methylation in cell free (cf)DNA of cervical cancer. Methylation of the target gene was validated on bisulfite converted smear-DNA from cervical dysplasia patients and afterward applied to cfDNA using quantitative real-time PCR. In 52 smears, a combined analysis of CADM1 and/or MAL (CADM1/MAL) showed methylation in 86.5% of the cases. In cfDNA samples of 24 cervical cancer patients, CADM1/MAL methylation was detected in 83.3% of the cases. CADM1/MAL methylation was detected already in 81.8% of stage I-II patients showing the high sensitivity of this liquid biopsy assay. In combination with a specificity of 95.5% towards healthy donors (HD) and an area under the curve (AUC) of 0.872 in the receiver operating characteristic (ROC) analysis, CADM1/MAL cfDNA methylation detection might represent a novel and promising liquid biopsy marker in cervical cancer.
摘要:
宫颈癌是女性第四常见的癌症,95%与高危型人乳头瘤病毒(HPV)感染有关。特定基因的甲基化与宫颈高级别发育不良病变向浸润性癌的进展密切相关。因此,DNA甲基化已被提议作为感染高危HPV的女性的分类。宫颈癌组织的甲基化分析表明,细胞粘附分子1(CADM1)和髓磷脂和淋巴细胞蛋白(MAL)甲基化存在于所有宫颈高级别肿瘤和浸润性宫颈癌的90%以上。这里,我们建立了一种基于液体活检的检测方法来检测宫颈癌无细胞(cf)DNA中的MAL和CADM1甲基化.在来自宫颈发育不良患者的亚硫酸氢盐转化的涂片DNA上验证了靶基因的甲基化,然后使用定量实时PCR将其应用于cfDNA。在52次涂片中,CADM1和/或MAL(CADM1/MAL)的联合分析显示86.5%的病例发生甲基化.在24名宫颈癌患者的cfDNA样本中,83.3%的病例检测到CADM1/MAL甲基化。在81.8%的I-II期患者中已经检测到CADM1/MAL甲基化,表明该液体活检测定具有很高的敏感性。结合受试者工作特征(ROC)分析中对健康供体(HD)的95.5%的特异性和0.872的曲线下面积(AUC),CADM1/MALcfDNA甲基化检测可能是宫颈癌中一种新颖且有前途的液体活检标志物。