研究目标

描述高危HPV在研究人群中的型别分布情况，在同一人群中比较以HPV DNA L1区、HPV DNA E6/E7区以及E6/E7 mRNA为靶点的三种HPV核酸检测技术的临床效果。

材料和方法

本研究于2014-2016年在河南新密入组Start-up项目随访妇女，同时从五家三甲医院的体检中心入组进行宫颈癌筛查的妇女，以及门诊或住院病人中病理确诊为宫颈上皮内瘤变或宫颈癌的妇女。采集以上受试者的宫颈脱落细胞标本，并保存在ThinPrep细胞学保存液中。细胞学标本制片、染色后由中国医学科学院肿瘤医院的细胞学医生进行最终细胞学诊断，并采用cobas、Aptima和Onclarity三种HPV检测技术对每位妇女的同一份标本进行检测。病理诊断由中国医学科学院肿瘤医院的病理医生诊断，必要时进行会诊以确定最终病理级别。

研究结果

1.     本研究共纳入2072名研究对象，其平均年龄为47.54岁（标准差为9.67），宫颈病理学检查结果为正常的有1054例，CIN1病变者169例，CIN2病变者110例，CIN3病变者187例，SCC患者506例，ADC患者46例。研究人群中，HPV16和HPV33/58阳性率最高，分别为30.4%和9.5%。HPV16阳性率随宫颈病理级别升高而升高（P<0.05），但在ADC中有所下降；HPV18阳性率在ADC中最高，为30.4%。绝经妇女的HPV阳性率高于未绝经妇女（55.8% VS 51.1%，P=0.046）；饮酒者中HPV阳性率显著高于未饮酒者（68.9% VS 47.6%，P<0.001）；吸烟者中HPV16阳性率显著高于未吸烟者（46.6% VS 9.9%，P=0.007），其余高危型别阳性率在吸烟者和非吸烟者之间未见统计学差异（P>0.05）。

2.     随着宫颈鳞状上皮病变级别的增高，采用三种检测技术所检出的HPV阳性率均呈现出升高趋势；与之相对的是，宫颈腺癌低于CIN2以上的鳞状上皮病变HPV阳性率。细胞学阴性或宫颈组织正常者中，cobas检测HPV阳性率（21.4%）均显著高于Onclarity（18.3%）和Aptima（16.0%）HPV阳性率（P<0.001）。在细胞学阴性或ASC-US以上病变中，Onclarity和cobas检测其他12种hrHPV阳性率之间存在统计学差异，P值分别为0.016和<0.001。

3.     总体而言，Aptima检测CIN2+、CIN3+以及癌症的灵敏度、阳性预测值和阴性预测值均高于Onclarity和cobas，但不存在统计学差异；Aptima检测CIN2+病变的特异度与Onclarity（74.65%：95%CI，72.14%-77.01%）相比无统计学差异，但显著高于cobas（76.61%：95%CI，74.16%-78.90% VS 71.22%：95%CI，68.62%-73.69%）。cobas检测ASC-US人群中CIN2+和CIN3+病变的灵敏度均高于Onclarity，但特异度低于Onclarity（62.87%：95%CI，56.03%-69.24% VS 66.83%：95%CI，60.08%-72.96%），阴性预测值高于Onclarity（92.70%：95%CI，87.08%-95.99% VS 91.84%：95%CI，86.27%-95.27%）；Aptima检测CIN2+和CIN3+病变的灵敏度、特异度、阳性预测值和阴性预测值均高于Onclarity，但均无统计学差异。筛查人群中，Onclarity、cobas和Aptima检测CIN2+病变的灵敏度均为100%（95%CI：83.39%-100%），显著高于液基细胞学检测（65%：95%CI，3.29%-81.88%），Onclarity和Aptima检测CIN2+的阳性预测值分别为12.42%（95%CI：8.19%-1.41%）和12.27（8.09%-18.19%），高于cobas（10.42%，95%CI：6.85%-15.54%）和液基细胞学检测（8.03%，95%CI：4.75%-13.24%）。

4.     对三种检测技术的一致性分析表明，三种检测技术两两之间的总体kappa值均在0.85以上，随着病理级别的升高，kappa值均有降低趋势，但在ADC中，三种检测技术的kappa值为1，一致率为100%。基于三种检测技术的检出限对检测不一致的结果分析表明，cobas和Onclarity检测14种hrHPV结果不一致的由121例降低到42例，Onclarity和Aptima检测结果不一致的从115例降低到74例，但cobas和Aptima检测结果不一致的从150例仅减少到126例，其中96例cobas阳性/Aptima阴性结果中有71例分布在宫颈正常者中；对型别检出不一致的结果比较显示，cobas与Onclarity或Aptima对HPV16或HPV18/45检测结果不一致例数减少较少，而Onclarity和Aptima检测HPV16不一致的由44降低到14例，检测HPV18/45不一致的则由25例降低到13例，Onclarity和cobas检测其他12种高危HPV型别不一致的结果则由141例降低到53例。

研究结论

1.     研究人群中阳性率最高的为HPV16，其次为HPV33/58。吸烟、饮酒和绝经状态对HPV感染率有一定影响。随年龄的变化，hrHPV阳性率呈“双峰”分布。采用不同检测技术时，总体阳性率之间未见明显差异，但Onclarity和cobas对其他12种hrHPV的检测阳性率之间存在一定差异。

2.     三种HPV检测技术的临床效果相差不大，均优于液基细胞学检查。与其他两种以DNA为靶点的检测技术相比，Aptima能够更好地检测与病变相关的感染；Onclarity HPV检测则能够提供更多的型别信息，基于“同等风险，同等管理”的原则，对于优化阳性妇女的管理更有优势。

3.     Onclarity与cobas或Aptima检测的一致性均较高，cobas和Aptima的一致性相对较低。

Objectives

To discribe the distribution of hrHPV in this population, and to compare the clinical performance of three HPV assays that target the HPV DNA L1 region, HPV DNA E6/E7 region and E6/E7 mRNA respectively in one population.

Materials and Methods

During 2014 and 2016, we recruited women who were followed up in the cervical cancer screening program-START-UP in Henan Xinmi County, at the same time, we enrolled women who participated cervical cancer screening in the routine physical examination centers of 5 tertiary hospitals across China, as well as outpatients or inpatients with biopsy-comfired cervical intraepithelial neoplasia (CIN) or cervical cancer. Cervical exfoliated cells specimens were collected from these women, all cytology specimens were stored in PreservCyt solution (Hologic, Inc. Bedford, MA), the cytology slides were processed and stained by trained technicians, and the final interpretation of cytology results was made by expert cytologists in NCC. At the same time, all specimens were tested using cobas, Aptima and Onclarity HPV assays, pathology diagnoses were made by the pathologists in NCC, and panel review was performed in necessary.

Results

1.     2072 women were included into this study, with mean age at 47.54±9.67, among them 1054 were diagnosed with normal histology, 169 with CIN1, 110 with CIN2, 187 with CIN3, 506 with SCC and 46 with ADC. In this population, prevalence of HPV16 and HPV33/58 were higher than other types, estimated as 30.4% and 9.5%, respectively. The positive rate of HPV16 increased with the increasing of cervical lesions, but decreased in ADC; prevalence of HPV 18 was the highest in ADC, estimated as 30.4%. Women who were postmenopausal showed higher HPV positivity compared to those who were not (55.8% VS 51.1，P=0.046); the HPV positivity of women reported alcohol use was statistically higher than those did not (68.9% VS 47.6%，P<0.001); women reported tobacco use implicated higher HPV16 positivity than those did not (46.6% VS 9.9%，P=0.007), but the other hrHPV positivity showed no significant difference (P>0.05).

2.     The positivity of hrHPV tested by three HPV assays all increased with the increasing of cervical squamous lesions, however, the positivity of HPV in ADC was lower than that in cervical squamous intraepithelial grade 2 or worse. In women with negative cytology or normal histology, hrHPV positivity by cobas (21.4%) was higher than Onclarity (18.3%) and Aptima (16.0%) (P<0.001). In women with nagetive cytology or ASC-US+, the positive rates of 12 other hrHPV tested by Onclarity and cobas were significantly different, P values were 0.016 and <0.001, respectively.

3.     Overall, the clinical sensitivity, PPV and NPV of Aptima in detection of CIN2+ or CIN3+ were slightly higher than those of Onclarity and cobas, but there was no statistical difference; the specificity of Aptima in detection of CIN2+ was no different from that of Onclarity (76.61%：95%CI，74.16%-78.90% VS 74.65%：95%CI，72.14%-77.01%), but was significantly higher than cobas (76.61%：95%CI，74.16%-78.90% VS 71.22%：95%CI，68.62%-73.69%), and the specificity of LBC (69.34%：95%CI，66.70-71.86) was statistically lower than Onclarity (74.65%：95%CI，72.14-77.01) and Aptima (76.61%：95%CI，74.16-78.90). In women with cytological ASC-US, the sensitivities of cobas for CIN2+ and CIN3+ were higher than Onclarity, but the specificity was lower than Onclarity (62.87%：95%CI，56.03%-69.24% VS 66.83%：95%CI，60.08%-72.96%), the NPV was higher than Onclarity (92.70%：95%CI，87.08%-95.99% VS 91.84%：95%CI，86.27%-95.27%). The sensitivity, specificity, PPV and NPV of Aptima in detection of CIN2+ or CIN3+ were not statistically higher than those of Onclarity. In the screening population, the sensitivities of Onclarity, cobas and Aptima were 100% (95%CI：83.39%-100%), and were statistically higher than that of LBC (65%：95%CI，3.29%-81.88%), the PPVs of Onclarity and Aptima for detection of CIN2+ were 12.42% (95%CI：8.19%-1.41%) and 12.27% (8.09%-18.19%) respectively, both were non-significantly higher than cobas (10.42%, 95%CI：6.85%-15.54%) and LBC (8.03%, 95%CI：4.75%-13.24%).

4.     The overall consistency analyses of three HPV assays showed that the agreement between any two assays were very high, with kappa values estimated higher than 0.85; and with the increasing of cervical lesions, kappa values decreased, while in women with ADC, the kappa value was 1, with agreement of 100%. Based on the limit of detection of 3 HPV assays, we analyzed the discordant detection results. The number of discordant results reduced from 121 to 42 in cobas/ Onclarity inconsistency, and the number of Onclarity/ Aptima discordance decreased from 115 to 74, however, the inconsistent results by cobas and Aptima decreased from 150 to 126, which showed the least reduction, among whom 71 women tested as cobas+/Aptima- were diagnosed as normal histology. When focused on the agreement of specific types of HPV, we found a less reduction on discordant results of HPV16 or HPV18/45 for cobas and Onclarity/ Aptima, however the discordant results of Onclarity and Aptima for HPV16 decreased from 44 to 14, for HPV18, the discordant results decreased from 25 to 13; as for the other 12hrHPV, the number of different results by Onclarity and cobas reduced from 141 to 53.

Conclusions

1.     The most prevalent type was HPV16, and then was HPV33/58 in our population, HPV positivity were influenced by smoking, drinking and menopause, women who were exposed to these factors showed higher hrHPV positivity than those who were not. With the change of age, the distribution of hrHPV presented as “two-peak”. The overall positive rate of HPV showed no significant difference when using different HPV detection methods, but the detection of the other 12hrHPV indicated significant difference between Onclarity and cobas.

2.     The clinical performance of the 3 HPV assays was not significantly different, but all were better than liquid-based cytology. Compared to the other two assays that target the DNA region of virus, Aptima HPV assay showed better performance in detection of infections related to lesions; Onclarity could provide more typing information, which could optimize the management of positive women based on the principle of “equal management of equal risk”.

3.     Onclarity showed high agreement with cobas or Aptima; the agreement of cobas and Aptima was the lowest in our study.