Hansel Alfred, Steinbach Daniel, Greinke Christiane, Schmitz Martina, Eiselt Juliane, Scheungraber Cornelia, Gajda Mieczyslaw, Hoyer Heike, Runnebaum Ingo B, Dürst Matthias
Department of Gynaecology, Jena University Hospital, Jena, Germany.
Institute of Pathology, Jena University Hospital, Jena, Germany.
PLoS One. 2014 Mar 19;9(3):e91905. doi: 10.1371/journal.pone.0091905. eCollection 2014.
High-risk human papillomavirus (hrHPV)-DNA testing is frequently performed parallel to cytology for the detection of high-grade dysplasia and cervical cancer particularly in women above 30 years of age. Although highly sensitive, hrHPV testing cannot distinguish between HPV-positive women with or without clinically relevant lesions. However, in principle discrimination is possible on the basis of DNA methylation markers. In order to identify novel DNA regions which allow an effective triage of hrHPV-positive cases, hypermethylated DNA enriched from cervical cancers was compared with that from cervical scrapes of HPV16-positive cases with no evidence for disease by CpG island microarray hybridization. The most promising marker regions were validated by quantitative methylation-specific PCR (qMSP) using DNA from archived cervical tissues and cervical scrapes. The performance of these markers was then determined in an independent set of 217 hrHPV-positive cervical scrapes from outpatients with histopathological verification. A methylation signature comprising the 5' regions of the genes DLX1, ITGA4, RXFP3, SOX17 and ZNF671 specific for CIN3 and cervical cancer (termed CIN3+) was identified and validated. A high detection rate of CIN3+ was obtained if at least 2 of the 5 markers were methylated. In the subsequent cross-sectional study all cervical carcinomas (n = 19) and 56% (13/23) of CIN3 were identified by this algorithm. Only 10% (11/105) of hrHPV-positive women without histological evidence of cervical disease were scored positive by the methylation assay. Of note is that the detection rate of CIN3 differed between age groups. Eight of nine CIN3 were detected among women ≥30 years of age but only five of fourteen among <30 year old group (p = 0.03). The specificity for CIN3+ in the older age group was 76.6% (95% CI 65.6-85.5%). Clinical validation studies are required to determine the usefulness of these novel markers for triage after primary hrHPV testing in a cervical cancer screening setting.
高危型人乳头瘤病毒（hrHPV）DNA检测常与细胞学检查同时进行，用于检测高级别发育异常和宫颈癌，尤其是30岁以上女性。尽管hrHPV检测高度敏感，但无法区分有无临床相关病变的HPV阳性女性。然而，原则上可根据DNA甲基化标志物进行鉴别。为了确定能够有效分流hrHPV阳性病例的新DNA区域，通过CpG岛微阵列杂交，将从宫颈癌中富集的高甲基化DNA与HPV16阳性且无疾病证据的宫颈刮片中的DNA进行比较。使用存档宫颈组织和宫颈刮片的DNA，通过定量甲基化特异性PCR（qMSP）对最有前景的标志物区域进行验证。然后在一组217例经组织病理学验证的门诊患者的hrHPV阳性宫颈刮片中确定这些标志物的性能。确定并验证了一个甲基化特征，该特征包括DLX1、ITGA4、RXFP3、SOX17和ZNF671基因的5'区域，这些区域对CIN3和宫颈癌（称为CIN3+）具有特异性。如果5个标志物中至少有2个甲基化，则CIN3+的检出率较高。在随后的横断面研究中，该算法识别出了所有宫颈癌（n = 19）和56%（13/23）的CIN3。在无宫颈疾病组织学证据的hrHPV阳性女性中，只有10%（11/105）通过甲基化检测呈阳性。值得注意的是，CIN3的检出率在不同年龄组之间存在差异。在≥30岁的女性中，9例CIN3中有8例被检测到，但在<30岁的组中，14例中只有5例（p = 0.03）。老年组中CIN3+的特异性为76.6%（95%CI 65.6 - 85.5%）。需要进行临床验证研究，以确定这些新标志物在宫颈癌筛查中初次hrHPV检测后分流的有用性。
