No abstract

Recently, DNA methylation analysis of FAM19A4 in cervical scrapes has been shown to adequately detect high‐grade cervical intraepithelial neoplasia and cervical cancer (≥CIN3) in high‐risk HPV (hrHPV)‐positive women. Here, we compared the clinical performance of FAM19A4 methylation analysis to cytology and HPV16/18 genotyping, separately and in combination, for ≥CIN3 detection in hrHPV‐positive women participating in a prospective observational multi‐center cohort study. The study population comprised hrHPV‐positive women aged 18–66 years, visiting a gynecological outpatient clinic. From these women, cervical scrapes and colposcopy‐directed biopsies (for histological confirmation) were obtained. Cervical scrapes were analyzed for FAM19A4 gene promoter methylation, cytology and HPV16/18 genotyping. Methylation analysis was performed by quantitative methylation‐specific PCR (qMSP). Sensitivities and specificities for ≥CIN3 were compared between tests. Stratified analyses were performed for variables that potentially influence marker performance. Of all 508 hrHPV‐positive women, the sensitivities for ≥CIN3 of cytology, FAM19A4 methylation analysis, and cytology combined with HPV16/18 genotyping were 85.6, 75.6 and 92.2%, respectively, with corresponding specificities of 49.8, 71.1 and 29.4%, respectively. Both sensitivity and specificity of FAM19A4 methylation analysis were associated with age (p ≤ 0.001 each). In women ≥30 years (n = 287), ≥CIN3 sensitivity of FAM19A4 methylation analysis was 88.3% (95%CI: 80.2–96.5) which was noninferior to that of cytology [85.5% (95%CI: 76.0–94.0)], at a significantly higher specificity [62.1% (95%CI: 55.8–68.4) compared to 47.6% (95%CI: 41.1–54.1)]. In conclusion, among hrHPV‐positive women from an outpatient population aged ≥30 years, methylation analysis of FAM19A4 is an attractive marker for the identification of women with ≥CIN3.

Primary human papillomavirus (HPV)-based screening results in a 2-5% lower specificity for cervical intraepithelial neoplasia Grade 2 or worse (CIN21) compared to Pap cytology. To identify HPV-positive women with CIN21, we retrospectively evaluated the cross-sectional and longitudinal performance of p16/Ki-67 dual-stained cytology in HPV-positive women with normal cytology participating in population-based cervical screening. Conventional Pap cytology specimens of 847 of these women derived from the VUSA-Screen study were dual-stained for p16/Ki-67. Cross-sectional clinical performance in detecting CIN3 or worse (CIN31), and CIN21 was compared to that of baseline HPV genotyping. Moreover, 5-year cumulative incidence risks (CIR) for CIN31 (CIN21) were determined. The sensitivity of p16/Ki-67 dual-stained cytology for CIN31 (CIN21) was 73.3% (68.8%) with a specificity of 70.0% (72.8%). HPV16/18 genotyping showed a sensitivity for CIN31 (CIN21) of 46.7% (43.8%), with a specificity of 78.3% (79.4%). The 5-year CIR for CIN31 in HPV-positive women with normal cytology was 6.9%. Testing these women with p16/Ki-67 dual-stained cytology resulted in a significantly lower CIN31 5-year CIR of 3.3% (p 5 0.017) in case of a negative test result. A negative HPV16/18 genotyping test result also led to a lower 5-year CIN31 CIR of 3.6%. p16/ Ki-67 dual-stained cytology detects more than 70% of underlying CIN31 lesions in HPV-positive women with normal cytology at baseline and is therefore suitable for triaging these women to colposcopy. Furthermore, the CIN31 5-year CIR of 3.3% after a negative dual-stain result is significantly lower compared to the 5-year CIR of 6.9% in women without p16/Ki-67 dualstained cytology triage.Persistent infections with carcinogenic human papillomavirus (HPV) genotypes represent the main causative event in the multistep process of cervical carcinogenesis. Recent efforts to improve cervical screening have focused on introducing HPV testing as a conjunct to Pap cytology testing, or as a primary screening tool. Unlike cytology, HPV testing is objective and has consistently been shown to be more sensitive for the detection of cervical intraepithelial neoplasia Grade 2 (Grade 3) or worse (CIN21/31) compared to cytology-based testing(for CIN31 94% vs. 65%).1,2 Based on this evidence, Key words: p16/Ki-67 dual-stained cytology, cervical cytology, human papillomavirus, genotyping Abbreviations: BMD: borderline mild dyskaryosis; CIN21: cervical intraepithelial neoplasia Grade 2 or worse; CIN31: cervical intraepithelial neoplasia Grade 3 or worse; CIR: cumulative incidence risks; EIA: enzyme immunoassay; HC2: hybrid capture 2; HPV: human papillomavirus; LBC: liquid-based cytology; NPV: negative predictive value; PPV: positive predictive value;

Background:Women with borderline/mildly dyskaryotic (BMD) cytology smears are currently followed up with repeat testing at 6 and 18 months. The objective of this study is to analyse the cross-sectional and longitudinal performance of p16/Ki-67 dual-stained cytology for the detection of cervical intraepithelial neoplasia (CIN) grade 3 or worse (CIN3+) and CIN2+ in women with BMD, and to compare the results with baseline human papillomavirus (HPV) testing.Methods:Conventional Pap cytology specimens of 256 women with BMD were dual stained for p16/Ki-67 retrospectively, and compared with baseline HPV results and long-term follow-up results.Results:p16/Ki-67 dual-stained cytology showed a sensitivity of 100%, a specificity of 64.4% and a negative predictive value (NPV) of 100.% for CIN3+. Human papillomavirus testing demonstrated similar sensitivity (96.3%), and NPV (99.1%), but a significantly lower specificity (57.6% P=0.024) for CIN3+. Sensitivity, specificity and NPV for CIN2+ of dual-stained cytology were 89.7%, 73.1% and 95.1%, respectively, which was similar when compared with HPV testing. Dual-stained cytology showed a significant lower referral rate than HPV testing (43.6% vs 49.1% P=0.043). During long-term follow-up, no CIN3+ lesions developed in HPV-positive, dual-stained negative women.Conclusions:Comparable sensitivity and NPV of dual-stained cytology for CIN3+, combined with a significantly higher specificity, makes p16/Ki-67 dual-stained cytology a viable alternative to HPV testing for triaging BMD.