Sensitivity For CIN2 Research Articles

Efficiency of CIN2+ Detection by Thyrotropin-Releasing Hormone (TRH) Site-Specific Methylation.

Cervical cancer screening typically involves a Pap smear combined with high-risk human papillomavirus (hr-HPV) detection. Women with hr-HPV positivity but normal cytology, as well as those with precancerous abnormal cytology, such as low-grade squamous intraepithelial lesions (LSIL) and high-grade SIL (HSIL), are referred for colposcopy and histology examination to identify abnormal lesions, such as cervical intraepithelial neoplasia (CIN) and cervical cancer. However, in order to enhance the accuracy of detection, bioinformatics analysis of a microarray database was performed, which identified cg01009664, a methylation marker of the thyrotropin-releasing hormone (TRH). Consequently, a real-time PCR assay was developed to distinguish CIN2+ (CIN2, CIN3, and cervical cancer) from CIN2- (CIN1 and normal cervical epithelia). The real-time PCR assay utilized specific primers targeting methylated cg01009664 sites, whereas an unmethylated reaction was used to check the DNA quality. A cut-off value for the methylated reaction of Ct < 33 was established, resulting in improved precision in identifying CIN2+. In the first cohort group, the assay demonstrated a sensitivity of 93.7% and a specificity of 98.6%. In the cytology samples identified as atypical squamous cells of undetermined significance (ASC-US) and LSIL, the sensitivity and specificity for detecting CIN2+ were 95.0% and 98.9%, respectively. However, when self-collected samples from women with confirmed histology were tested, the sensitivity for CIN2+ detection dropped to 49.15%, while maintaining a specificity of 100%. Notably, the use of clinician-collected samples increased the sensitivity of TRH methylation testing. TRH methylation analysis can effectively identify women who require referral for colposcopy examinations, aiding in the detection of CIN2+.

Performance of p16/Ki67 Immunostaining for Triage of Elderly Women with Atypical Squamous Cells of Undetermined Significance.

The p16/Ki67 technique has been poorly studied in postmenopausal women with ASC-US cytology. The objective of this study was to compare the accuracy of p16/Ki67 staining, HPV testing and HPV 16 genotyping for the identification of CIN2 + lesions in postmenopausal women with ASC-US cytology. A total of 324 postmenopausal women with positive ASC-US were included. The women underwent HPV test, colposcopy, and biopsy. The slides were discolored and then stained with the CINtec Plus Kit for p16/Ki67. The HPV test results were classified as HPV16 +, hrHPV+ (other hrHPV genotypes), or HPV negative. The p16/Ki67 sensitivity for CIN2+ was 94.5%, the specificity 86.6%, PPV of 59% and NPV of 95.9%. The HPV test showed a sensitivity of 96.4% for CIN2+, a specificity of 62.8%, a PPV of 35% and a NPV of 98.8%. In postmenopausal women, the prevalence of genotype 16 decreases in favor of the other high-risk genotypes. Given the low sensitivity of cytology and the low percentage of HPV16-positive cancers among elderly women, triage via cytology and genotyping is not the best strategy; double staining cytology shows high profiles of sensibility and specificity for CIN2+ in ASCUS postmenopausal women.

Performance of standardised colposcopy to detect cervical precancer and cancer for triage of women testing positive for human papillomavirus: results from the ESTAMPA multicentric screening study

Colposcopy, currently included in WHO recommendations as an option to triage human papillomavirus (HPV)-positive women, remains as the reference standard to guide both biopsy for confirmation of cervical precancer and cancer and treatment approaches. We aim to evaluate the performance of colposcopy to detect cervical precancer and cancer for triage in HPV-positive women. This cross-sectional, multicentric screening study was conducted at 12 centres (including primary and secondary care centres, hospitals, laboratories, and universities) in Latin America (Argentina, Bolivia, Colombia, Costa Rica, Honduras, Mexico, Paraguay, Peru, and Uruguay). Eligible women were aged 30-64 years, sexually active, did not have a history of cervical cancer or treatment for cervical precancer or a hysterectomy, and were not planning to move outside of the study area. Women were screened with HPV DNA testing and cytology. HPV-positive women were referred to colposcopy using a standardised protocol, including biopsy collection of observed lesions, endocervical sampling for transformation zone (TZ) type 3, and treatment as needed. Women with initial normal colposcopy or no high-grade cervical lesions on histology (less than cervical intraepithelial neoplasia [CIN] grade 2) were recalled after 18 months for another HPV test to complete disease ascertainment; HPV-positive women were referred for a second colposcopy with biopsy and treatment as needed. Diagnostic accuracy of colposcopy was assessed by considering a positive test result when the colposcopic impression at the initial colposcopy was positive minor, positive major, or suspected cancer, and was considered negative otherwise. The main study outcome was histologically confirmed CIN3+ (defined as grade 3 or worse) detected at the initial visit or 18-month visit. Between Dec 12, 2012, and Dec 3, 2021, 42 502 women were recruited, and 5985 (14·1%) tested positive for HPV. 4499 participants with complete disease ascertainment and follow-up were included in the analysis, with a median age of 40·6 years (IQR 34·7-49·9). CIN3+ was detected in 669 (14·9%) of 4499 women at the initial visit or 18-month visit (3530 [78·5%] negative or CIN1, 300 [6·7%] CIN2, 616 [13·7%] CIN3, and 53 [1·2%] cancers). Sensitivity was 91·2% (95% CI 88·9-93·2) for CIN3+, whereas specificity was 50·1% (48·5-51·8) for less than CIN2 and 47·1% (45·5-48·7) for less than CIN3. Sensitivity for CIN3+ significantly decreased in older women (93·5% [95% CI 91·3-95·3] in those aged 30-49 years vs 77·6% [68·6-85·0] in those aged 50-65 years; p<0·0001), whereas specificity for less than CIN2 significantly increased (45·7% [43·8-47·6] vs 61·8% [58·7-64·8]; p<0·0001). Sensitivity for CIN3+ was also significantly lower in women with negative cytology than in those with abnormal cytology (p<0·0001). Colposcopy is accurate for CIN3+ detection in HPV-positive women. These results reflect ESTAMPA efforts in an 18-month follow-up strategy to maximise disease detection with an internationally validated clinical management protocol and regular training, including quality improvement practices. We showed that colposcopy can be optimised with proper standardisation to be used as triage in HPV-positive women. WHO; Pan American Health Organization; Union for International Cancer Control; National Cancer Institute (NCI); NCI Center for Global Health; National Agency for the Promotion of Research, Technological Development, and Innovation; NCI of Argentina and Colombia; Caja Costarricense de Seguro Social; National Council for Science and Technology of Paraguay; International Agency for Research on Cancer; and all local collaborative institutions.

Performance of visual inspection of the cervix with acetic acid (VIA) for triage of HPV screen-positive women: results from the ESTAMPA study.

VIA is recommended for triage of HPV-positive women attending cervical screening. In the multicentric ESTAMPA study, VIA performance for detection of cervical intraepithelial neoplasia grade 3 or worse (CIN3+) among HPV-positive women was evaluated. Women aged 30-64 years were screened with HPV testing and cytology and referred to colposcopy if either test was positive. At colposcopy visit, study-trained midwives/nurses/GPs performed VIA ahead of colposcopy. VIA was considered positive if acetowhite lesions were observed in or close to the transformation zone. Ablative treatment eligibility was assessed for VIA positives. Performance indicators were estimated. Three thousand one hundred and forty-two HPV-positive women were included. Sensitivity for CIN3+ was 85.9% (95% CI 81.2-89.5) among women <50 years and, although not significant, slightly lower in women 50+ (78.0%, 95% CI 65.9-86.6). Overall specificity was 58.6% (95% CI 56.7-60.5) and was significantly higher among women 50+ (70.3%, 95% CI 66.8-73.5) compared to women <50 (54.3%, 95% CI 52.1-56.5). VIA positivity was lower among women 50+ (35.2%, 95% CI 31.9-38.6) compared to women <50 (53.2, 95% CI 51.1-55.2). Overall eligibility for ablative treatment was 74.5% and did not differ by age. VIA sensitivity, specificity, and positivity, and ablative treatment eligibility varied highly by provider (ranges: 25%-95.4%, 44.9%-94.4%, 8.2%-65.3%, 0%-98.7%, respectively). VIA sensitivity for cervical precancer detection among HPV-positive women performed by trained providers was high with an important reduction in referral rates. However, scaling-up HPV screening triaged by VIA will be challenging due to the high variability of VIA performance and providers' need for training and supervision.

Validation of BD Onclarity HPV Assay on Vaginal Self-Samples versus Cervical Samples Using the VALHUDES Protocol.

In this study, we evaluated accuracy of HPV testing on self-samples versus clinician-taken samples through the VALHUDES protocol. VALHUDES was designed as a diagnostic test accuracy study, where women referred to colposcopy collected self-samples followed by clinician-taken cervical samples. Four hundred eighty-five women recruited in five colposcopy clinics (median age = 40 years; IQR, 31-49) with valid results for all specimens were included in the main analysis: 230 vaginal self-samples were collected with Evalyn Brush and 255 with Qvintip. Cervical samples were taken by the gynecologist with the Cervex-Brush. HPV testing was performed with BD Onclarity HPV assay (Onclarity). Colposcopy and histology were used as the reference standard for accuracy estimation. The sensitivity for CIN2+ on vaginal self-samples overall was not different from cervical samples (ratio = 0.96; 95% CI, 0.90-1.03), whereas specificity was significantly higher (ratio = 1.09; 95% CI, 1.02-1.16). However, the relative accuracy (self- vs. clinician sampling) differed by vaginal collection device: relative sensitivity and specificity ratios of 1.00 (95% CI, 0.94-1.06) and 1.15 (95% CI, 1.05-1.25), respectively for Evalyn-Brush; 0.91 (95% CI, 0.79-1.04) and 1.03 (95% CI, 0.95-1.13), respectively for Qvintip. Clinical accuracy of BD Onclarity HPV assay on vaginal self-samples was not different from cervical samples. VALHUDES study showed that HPV testing with Onclarity HPV on vaginal self-samples is similarly sensitive compared with cervical specimens. However, differences in accuracy by self-sampling devices, although not significant, were noted. Onclarity HPV testing on vaginal self-samples following validated collection and handling procedures may be used in primary cervical cancer screening.

Clinical performance of DNA and RNA based HPV tests for test of cure (TOC) post treatment for cervical intraepithelial neoplasia (CIN) - a retrospective study.

HPV testing as a "test of cure" (TOC) of women treated for cervical high-grade lesions can support and inform appropriate clinical management pathways. However, there is a lack of studies that report the discrete performance of different HPV assays in this context, including HPV mRNA based assays. To address this, we performed an analysis of the clinical performance of two hrHPV assays in the (TOC) setting; the recently launched DNA based Alinity m HR HPV (Abbott Molecular) and RNA based Aptima HPV assay (Hologic). Using a retrospective case-control design, two panels of archived cervical liquid based cytology samples, originally taken as per routine TOC protocols in Scotland were assessed. Each panel contained 63 cases, where cervical intraepithelial neoplasia 2 or worse (CIN2+) was detected and 160 controls (women with no CIN2+ and two subsequent cytology negative results (minimum) 3 years apart or women who had histologically confirmed ≤CIN1). All samples were previously tested using the RealTime High Risk HPV assay (Abbott Molecular) as per national TOC protocol. Panel A and Panel B were tested using Alinity and Aptima assay respectively. Both assays showed similar performance to the original RealTime assay. Aptima had sensitivity for CIN2+ of 96.8% (95% CI: 89.0- 99.6) compared to RealTime (93.7% (95% CI: 84.5 - 98.2)). Alinity had sensitivity for CIN2+ of 92.1% (95% CI: 82.4- 97.4) compared to RealTime (98.4% (95% CI: 91.5- 99.95)). Both mRNA based and DNA based HPV tests show robust performance for the monitoring of residual disease post-treatment.

Evaluation of dry specimen transport and processing time using an isothermal amplification high-risk human papillomavirus assay

BackgroundDry specimen transport has shown equivalence to traditional liquid transport using a novel high-risk Human papillomavirus assay. Considering that dry transport might cross obstacles during cervical cancer screening in low and middle resource settings, this study was designed evaluate different processing time of dry specimen transport using the same isothermal amplification hrHPV assay. MethodsThere were 564 women between the ages of 30–55 recruited from colposcopy clinic. For each patient, two endocervical samples were collected and placed into empty collection tubes by physician. Samples were stored at room temperature until analyzed for hrHPV using the AmpFire assay at two time points: 2 days and 2 weeks. 511 of the 564 participants with positive hrHPV were provided colposcopy exam and quadrant biopsy. ResultsA total of 1128 endocervical samples from 564 patients were detected by the Ampfire assay. Good agreement was found between two time periods (Kappa ​± ​Standard error ​= ​0.67 ​± ​0.04). Sensitivity (2days/2weeks) for CIN2+ was 95.28% (95% CI: 92.14%–98.42%) vs 90.57% (CI (86.65%–94.49%) and specificity (2days/2weeks) was 22.47% (CI 19.33%–25.61%) vs 28.15% (CI 24.23%–32.07%) respectively. The difference for Ampfire HPV detection in sensitivity for CIN2+ for the two time periods was not significant (P ​= ​0.227), while the difference in specificity for CIN2+ was significant (P ​= ​0.001). The difference in Ct values 29.23 (CI 28.15–30.31) and 29.27 (CI 28.19–30.35) between two time points was not significant (P ​= ​0.164). ConclusionProcessing dry brush specimens can be delayed up to 2 weeks. Using the AmpFire assay platform which supports cervical cancer prevention programs in low-to-middle-income countries (LMICs).

Comparison of Different HPV-based Strategies and Cytology in Routine Cervical Cancer Screening Programme in China: A Population-based Study.

The study aimed to compare the performance of human papillomavirus (HPV)-based strategies to cytology for detecting cervical intraepithelial neoplasia grade 3 or worse (CIN3+) in routine program in China. The study included 50,434 women ages 30-64 years from January 2015 to December 2019, to assess four strategies: cytology with HPV triage (strategy 1), primary HPV testing with reflex cytology (strategy 2), primary HPV testing with HPV-16/18 genotyping and reflex cytology for non-16-18 high-risk HPV genotypes (strategy 3), and co-testing (strategy 4). The main outcomes were sensitivity for CIN3+ and colposcopy referral rate. Overall, the rates of HPV positivity and cytologic abnormality were 7.0% [95% confidential interval (CI), 6.8-7.2] and 3.5% (95% CI, 3.3-3.6], respectively. The sensitivity for CIN3+ were 88.5% for strategy 4, 83.2% for strategy 3, 64.6% for strategy 1, and 60.1% for strategy 2. The relative sensitivity of strategy 4 and strategy 3 compared with strategy 1 for detecting CIN3+ were 1.38 (95% CI, 1.24-1.54) and 1.29 (95% CI, 1.14-1.46), respectively. The colposcopy referral rates of strategies 4 and 3 were significantly higher than that of strategy 1 (2.4% and 2.2% vs. 1.4%, P < 0.001). In conclusion, co-testing and primary HPV testing with HPV-16/18 genotyping and reflex cytology improved the sensitivity for CIN3+ compared with cytology but increased the colposcopy referral rate. Long-term negative predicted value for HPV-negative women should be studied to determine the screening interval. Our study provides further evidence to the introduction of HPV-based strategies in China. PREVENTION RELEVANCE: Both co-testing and primary HPV testing with HPV-16/18 genotyping and reflex cytology triage provided higher sensitivity for detecting CIN3+; however, the number of colposcopy referrals also increased compared with cytology in a routine program. It has great public health implications for the introduction of HPV-based screening strategies in China.

The role of immunocytochemical biomarkers in diagnostics of precancerous pathology of cervix

The last decades showed the worldwide tendency to finding consensus between diagnostics improvement and constant increase of cost of medical services in conditions of restricted financing. The aim of the article was to analyze the diagnostic value of p16 and Ki-67 biomarkers in diagnostics of precancerous diseases of cervix. Data of 80 patients with cervical dysplasia of varying degree who received excisional treatment were analyzed. It was shown that cytological study has a high sensitivity (79.17%) for the diagnosis of CIN 2-3, but low specificity (53.57%). The p16 immunocytochemical biomarker has a high sensitivity for the diagnosis of CIN 2 (0.92; 95% CI: 0.76-0.98) with good specificity (0.78; 95% CI: 0.67-0.82), for the diagnosis of CIN 3 both sensitivity (0.93; 95% CI: 0.82-0.98) and specificity (0.93; 95% CI: 0.82-0.98) is high. The immuno­cytochemical biomarker Ki-67 has a high sensitivity for CIN 2 (0.92; 95% CI: 0.65-0.99), but insufficient specificity (0.62; 95% CI: 0.54-0.64), for the diagnosis of CIN 3 the sensitivity is very high (0.96; 95% CI: 0.80-0.99) as well as specificity (0.78; 95% CI: 0.69-0.81). The combined use of p16 and Ki-67 biomarkers can significantly increase the diagnostic accuracy of the diagnosis of high-grade precancerous pathology of cervix and justify timely surgical intervention. Such an approach for the differential diagnosis of severe dysplasia, on the one hand, may contribute to a decrease in the risk of developing cervical cancer, and on the other hand, it will allow to avoid unnecessary operations and preserve reproductive function of women, reduce the economic costs of treatment.

Clinical relevance of partial HPV16/18 genotyping in stratifying HPV-positive women attending routine cervical cancer screening: a population-based cohort study.

ObjectiveTo evaluate partial HPV16/18 genotyping as a possible biomarker to select women attending HPV‐based cervical cancer screening at higher risk to be referred to colposcopy.DesignPopulation‐based cohort study.SettingOrganised cervical cancer screening programmes (Italy).PopulationWomen with high‐risk HPV infection (period: 2015–2019).MethodsWe analysed the association between partial HPV16/18 genotyping, cytology triage and histologically confirmed diagnosis of high‐grade cervical intraepithelial neoplasia (CIN3+) lesions.Main outcome measuresDetection rate (DR) and positive predictive value (PPV) for histologically confirmed CIN3+ (any episode in the 2 years after baseline); sensitivity for CIN3+ and number of colposcopies needed for lesion detection.ResultsThe study included 145 437 women screened with HPV testing by the clinically validated COBAS 4800 HPV assay (Roche). Overall, 9601 (6.6%) women were HPV+ at baseline; HPV16 and HPV18 were present in 1865 and 594 samples, respectively. The cumulative (baseline plus 1‐year repeat) cytology positivity was 42.8% and high‐grade cytology was significantly higher (P < 0.0001) among women with HPV16 infection at baseline (15.2%). The cumulative CIN3+ DR for women with HPV16, HPV18 and other HPV‐type infections was 9.8%, 3.4% and 1.8%, respectively.ConclusionsPartial HPV16 genotyping may play a role in triage, whereas HPV18 seems to behave much more similarly to the other HPV types and does not provide additional stratification. HPV16 genotyping combined with high‐grade cytology can be envisaged as a triage biomarker in cervical screening to maximise CIN3+ detection while minimising colposcopy at baseline or 1‐year repeat.Tweetable abstractHPV16 genotyping combined with high‐grade cytology can be used as triage biomarker for CIN3+ in HPV‐positive women.

Extended HPV Genotyping to Compare HPV Type Distribution in Self- and Provider-Collected Samples for Cervical Cancer Screening.

Primary high-risk human papillomavirus (hr-HPV) testing of self-collected cervico-vaginal swabs could increase cervical cancer screening coverage, although triage strategies are needed to reduce unnecessary colposcopies. We evaluated the use of extended hr-HPV genotyping of self-collected samples for cervical cancer screening. We recruited women ages 25-65 years at two colposcopy clinics in North Carolina between November 2016 and January 2019, and obtained self-collected cervico-vaginal samples, provider-collected cervical samples, and cervical biopsies from all enrolled women. Self- and provider-collected samples were tested for 14 hr-HPV genotypes using the Onclarity Assay (Becton Dickinson). We calculated hr-HPV genotype-specific prevalence and assessed agreement between results in self- and provider-collected samples. We ranked the hr-HPV genotypes according to their positive predictive value (PPV) for the detection of cervical intraepithelial neoplasia (CIN) grade 2 or higher (CIN2+). A total of 314 women participated (median age, 36 years); 85 women (27%) had CIN2+. More women tested positive for any hr-HPV on self-collected (76%) than on provider-collected samples (70%; P = 0.009) with type-specific agreement ranging from substantial to almost perfect. HPV-16 was the most common genotype in self-collected (27%) and provider-collected samples (20%), and HPV-16 prevalence was higher in self- than provider-collected samples (P < 0.001). In self- and provider-collected samples, HPV-16 had the highest PPV for CIN2+ detection. Overall sensitivity for CIN2+ detection was similar for both sample types, but the higher HPV-16 prevalence in self-collected samples could result in increased colposcopy referral rates. Additional molecular markers might be helpful to improve the triage of women who are hr-HPV positive on self-collected samples.

An evaluation of solid versus liquid transport media for high-risk HPV detection and cervical cancer screening on self-collected specimens

BackgroundThe solid transport media is a small size card that allows fast, easy DNA extraction from a variety of biological samples. In 2016 we developed a solid media transport card; for that pilot study to control the self-collection we used a pseudo-self-collection technique. The current study expands this prior work using true self-collections and only the POI card, and aims to evaluate the solid media transport card to detect HR-HPV in self-samples compared to liquid transport media.MethodsTen thousand eight hundred eighty-five women between the ages of 30–59 with no screening for 3 years were enrolled. The self-collected sample was first applied to a new solid media transport card (Labeled as SC) then the brush placed in 6 ml ThinPrep liquid (Labeled as SL). Then a physician collected a direct endocervical specimen into ThinPrep liquid (Labeled as DL). Samples were tested with Cobas 4800 and the SeqHPV NGS assay for HR-HPV. Patients positive on any test were recalled for colposcopy and biopsy.ResultsTen thousand three hundred thirty-nine participants had complete data. The mean age was 43.9 years. CIN 2+ rates were 1.4% (142/10339). The agreement in HPV detection between the two different self-sample collection media was also good (Cobas HPV kappa = 0.86; SeqHPV kappa = 0.98). Tested with Cobas, the sensitivity of Cobas-SL and Cobas-SC for CIN 2+ was95.07 and 94.37%; and for CIN3+ was 96.30, 96.30% respectively. The specificity of Cobas-SC, and Cobas-SL for CIN2+ was 88.74 and 87.35%; for CIN3 was 88.04and 86.65% respectively. Tested with SeqHPV, the sensitivity for CIN2+ of Seq-SC and Seq-SL was 95.77 and 96.48%; for CIN3+, both the SC and SL specimens had a sensitivity of 100%. The specificity for CIN2+ of Seq-SC and Seq-SL was 89.54 and 89.53%; for CIN3+ was 88.84,88.82% respectively. For both HR-HPV assays, the sensitivities were similar for the two self-sample media (SC vs SL, p = 1.00).ConclusionsThe solid transport card for collecting vaginal self-samples as accurate as liquid transport media assayed by two different PCR based HR-HPV tests. The solid transport media is a suitable medium for collecting and storing vaginal self-samples.

Evaluation of an isothermal amplification HPV detection assay for primary cervical cancer screening

ObjectiveThe aim of this research was to evaluate independently the performance of a new isothermal amplification assay for cervical cancer screening compared to two previously validated PCR-based assays and histologic endpoints.MethodsThis is a sub-study from the Chinese multi-center screening trial (CHIMUST). The self-collected and clinician-collected specimens stored in PreservCyt at − 4 °C from 6042 women with complete data were tested with the AmpFire assay. These specimens had been previously tested with Cobas and SeqHPV assays. In the primary study all patients with an abnormal test were referred to colposcopy where all had directed and/or random biopsies plus ECC. No additional patients were called back based on the AmpFire results.Results6042/6619 women had complete data (mean age 44.1). There were 57 cases of CIN 2, 35 cases of CIN 3 and 2 cancers. The sensitivity for CIN2+ and CIN3+ were similar among the three assays (both direct and self-collected). For the specificities in all categories (CIN2+/CIN3+ and self and direct collection), isothermal amplification assay was either equal to or more specific than Cobas but consistently less specific than SeqHPV.ConclusionThe AmpFire HPV assay showed similar sensitivity to Cobas and SeqHPV for CIN2+ and CIN3+ on both self and clinician-collections (P>0.05), with good specificity. The speed, low cost, and simplicity of this assay will make it particularly suited for low and middle resource settings. Its accuracy with self-collection makes it applicable for mass screening programs.

Methylation markers FAM19A4 and miR124-2 as triage strategy for primary human papillomavirus screen positive women: A large European multicenter study.

In human papillomavirus (HPV) cervical cancer screening, cytology is used as triage to counter the low specificity of HPV testing. VALID‐SCREEN is a EU‐multicenter, retrospective study conducted to evaluate the clinical performance of the FAM19A4/miR124‐2 methylation‐based molecular triage test as a substitute or addition to cytology as reflex testing of HPV screen positive women. FAM19A4/miR124‐2 methylation test (QIAsure Methylation Test) was evaluated in 2384 HPV‐positive cervical screening samples, from women 29‐76 years of age, derived from four EU countries. Specimens were collected in ThinPrep or SurePath media, HPV‐status, concurrent cytology, and histology diagnosis were provided by the parent institutes. The control population consisted of women with no evidence of disease within 2 years of follow‐up. A total of 899 histologies were retrieved; 527 showed no disease, 124 CIN2 (5.2%), 228 CIN3 (9.6%) and 20 cervical cancers (0.8%); 19 of 20 screen‐detected cervical cancers were found methylation‐positive (sensitivity 95%). Overall specificity of FAM19A4/miR124‐2 methylation test was 78.3% (n = 2013; 95%CI: 76‐80). The negative predictive value of hrHPV positive, methylation‐negative outcomes were 99.9% for cervical cancer (N = 1694; 95%CI: 99.6‐99.99), 96.9% for ≥CIN3 (95%CI: 96‐98), and 93.0% for ≥CIN2 (95%CI: 92‐94). Overall sensitivity for CIN3 using FAM19A4/miR124‐2 methylation test was 77% (n = 228; 95%CI: 71‐82). CIN3 sensitivity was uniform between centers independent of sample collection medias, DNA extraction methods and HPV screening tests. Being objectively reported compared to the subjectivity of cytology, equally performing across settings and screening methods, the FAM19A4/miR124‐2 methylation constitute an alternative/supplement to cytology as triage method to be investigated in real‐life pilot implementation.

Interobserver variability and accuracy of p16/Ki-67 dual immunocytochemical staining on conventional cervical smears

Backgroundp16/Ki-67 dual immunocytochemical staining (DS) has been proven as a sensitive and specific test for triage of HPV positive women with good reproducibility and accuracy. However, implementation of the test into an organized screening program (OSP) is not easy. The aims of this study were to compare the performance and agreement of DS results among three Slovenian cytopathological laboratories involved in the national OSP, and to define cases where staining results can be difficult to interpret.MethodsCervical smears were obtained for DS from 129 women referred to colposcopy. Smears were evaluated blindly in three laboratories by a cytotechnologist and a cytopathologist after initial training. Results were positive, suspicious, negative or inadequate. Five characteristics of DS staining were recorded. After primary evaluation, an extensive expert-led additional training was undertaken, including a discussion of difficult cases and a practical exam. Smears were re-evaluated and results compared to primary evaluation.ResultsAfter the additional training, the overall percentage of agreement among the three laboratories increased from 77.5 to 89.9% and kappa increased from 0.70 to 0.86. Sensitivity for CIN2+ increased in two laboratories, to 90.5 and 85.7%, without the loss of specificity (75.8%). In one laboratory, the sensitivity slightly decreased from 90.5 to 88.9%, but the specificity increased from 63.6 to 68.2%. Difficult cases had significantly less DS cells, weak intensity of p16 staining, suboptimal cell morphology and background staining compared to positive cases.ConclusionAdditional expert-led training and discussion of difficult cases are necessary for accurate interpretation of DS in laboratories involved in OSP. The most difficult cases were those with single stained cells and weak p16 staining. Training protocol for safe implementation of p16/Ki-67 DS in OSP is proposed.

Analysis of cervical cancer screening results in women aged 50 and older in Beijing and Guizhou

Objective: To analyze the screening results of cervical cancer in elderly women in Beijing and Guizhou. To compare the screening effectiveness of cervical cancer in elderly women in different areas. Methods: A retrospective analysis was performed on the screening data of elderly women (≥50 years) who had examination at cervical lesion clinic in Peking University People's Hospital and Affiliated Hospital of Guizhou Medical University from January 2013 to January 2014, with histopathology as the gold standard, to compare screening effectiveness of cervical cytology or HPV detection for elderly women in cervical cancer screening in the two regions. Results: Among the patients with cervical intraepithelial neoplasia grade 2 and above (CIN2 + ) detected by colposcopy in Peking University People's Hospital and Affiliated Hospital of Guizhou Medical University, 44.1% (82/186) and 57.0% (98/172) were women aged 50 and older. In Beijing group 39.0% (32/82) of CIN2 + patients had clinical symptoms, which in Guizhou group was 31.6% (31/98). In Beijing group and Guizhou group, 50.0% (19/38) and 73.2% (30/41) patients with cervical cancer had never been screened, respectively. In Beijing group and Guizhou group, 5.4% (4/74) and 19.2% (10/52) of the patients with CIN2 + had cervical cytology negative for intraepithelial lesion or malignancy (NILM), respectively (P=0.015). The cervical cytology showed atypical squamous cells of undetermined significance (ASC-US) in 10.2% (13/127) and 24.6% (14/57), respectively, with significant difference (P=0.01). The cervical cytology showed low-grade squamous intraepithelial lesions (LSIL) in 14.9% (11/74) and 37.5% (6/16) with no significant difference (P=0.08). Atypical squamous cells cannot exclude high-grade squamous intraepithelial lesions (ASC-H) were 41.7% (5/12) and 71.4% (20/28) with no statistically significant difference (P=0.15). High-grade squamous intraepithelial lesions (HSIL) were 78.9% (15/19) and 86.4% (19/22) with no significant difference (P=0.83). In cases with HPV detection information, the sensitivity of CIN2+ diagnosis by high-risk HPV detection in Beijing group and Guizhou group was 95.2% (40/42) and 73.6% (39/53) that was statistically significant (P<0.05). Conclusions: In both Beijing and Guizhou, the incidence of CIN2+ in elderly women is at a relatively high level. There were differences in the detection of CIN2+ by cervical cytology or HPV detection in Beijing and Guizhou.

DNA methylation markers as a triage test for identification of cervical lesions in a high risk human papillomavirus positive screening cohort.

Objective triage strategies are required to prevent unnecessary referrals for colposcopy in population‐based screening programs using primary high‐risk human papillomavirus (hrHPV) testing. We have identified several DNA methylation markers with high sensitivity and specificity for detection of high‐grade cervical intraepithelial neoplasia or worse (CIN2+) in women referred for colposcopy. Our study assessed diagnostic potential of these methylation markers in a hrHPV‐positive screening cohort. All six markers (JAM3, EPB41L3, C13orf18, ANKRD18CP, ZSCAN1 and SOX1) showed similar association across histology in the hrHPV‐positive cohort when compared to the Dutch cohort (each p > 0.15). Sensitivity for CIN2+ was higher using methylation panel C13orf18/EPB41L3/JAM3 compared to the other 2 panels (80% vs. 60% (ANKRD18CP/C13orf18/JAM3) and 63% (SOX1/ZSCAN1), p = 0.01). For CIN3+ all three methylation panels showed comparable sensitivity ranging from 68% (13/19) to 95% (18/19). Specificity of SOX1/ZSCAN1 panel (84%, 167/200) was considerably higher compared to ANKRD18CP/C13orf18/JAM3 (68%, 136/200, p = 2 × 10−5) and C13orf18/EPB41L3/JAM3 (66%, 132/200, p = 2 × 10−7). High negative predictive value (NPV) (91–95% and 96–99%) was observed for CIN2+ and CIN3+, for all three methylation panels, while positive predictive value (PPV) varied from 25 to 40% for CIN2+ and 15–27% for CIN3+. Interestingly, 118/235 samples were negative for all six markers (including 106 controls (89.8%), 6 CIN1 (5.1%), 5 CIN2 (4.2%) and 1 CIN3 (0.8%)). Methylation results from both independent cohorts were comparable as well as high sensitivity for detection of cervical cancer and its high‐grade precursors in hrHPV‐positive population. Our study therefore validates these methylation marker panels as triage test either in hrHPV‐based or abnormal cytology‐based screening programs.

Clinical Evaluation of INNO-LiPA HPV Genotyping EXTRA II Assay Using the VALGENT Framework.

In this diagnostic test validation study, we assessed the clinical accuracy and HPV genotyping performance of the INNO-LiPA HPV Genotyping Extra II (INNO-LiPA) within the VALGENT-3 framework. VALGENT is designed to assess the analytical and clinical performance of HPV tests with genotyping capacity. The VALGENT-3 panel comprised 1300 consecutive cervical cell specimens enriched with 300 samples with abnormal cytology obtained from women attending the Slovenian cervical cancer screening programme. The INNO-LiPA allows type-specific detection of 32 HPV types; however, for the clinical accuracy assessment, we considered it as high-risk (hr)HPV positive when at least one of the following HPV types was present: HPV16, HPV18, HPV31, HPV33, HPV35, HPV39, HPV45, HPV51, HPV52, HPV56, HPV58, HPV59, and HPV68. Clinical accuracy for detection of cervical intraepithelial neoplasia grade 2 or worse (CIN2+) was compared between INNO-LiPA and Hybrid Capture 2 (HC2), which is a standard comparator test for HPV tests used in cervical cancer screening. In addition, hrHPV and type-specific detection HPV types were compared between INNO-LiPA and Linear Array HPV Genotyping Test (Linear Array). The prevalence of hrHPV determined by INNO-LiPA was 17.1% (95% CI, 15.0–19.2%) in the screening population. HrHPV testing with INNO-LiPA had a sensitivity for CIN2+ of 96.9% (95% CI, 92.1–99.1%) which was non-inferior to HC2 (relative sensitivity of 1.01; 95% CI, 0.97–1.04; pn.inf = 0.0002) and a specificity for ≤CIN1 of 85.3% (95% CI, 83.2–87.3%) which was inferior to HC2 (relative specificity of 0.95; 95% CI, 0.93–0.97; pn.inf = 0.9998). Genotyping agreement between INNO-LiPA and Linear Array was excellent for hrHPV, HPV16, HPV18, HPV35, HPV45, HPV58 and HPV59, but good or fair for other HPV types. To conclude, INNO-LiPA demonstrated non-inferior clinical sensitivity but lower specificity compared to HC2 in addition to excellent concordance compared to Linear Array for hrHPV and some genotypes.

Clinical Utility of CINtec PLUS Triage in Equivocal Cervical Cytology and Human Papillomavirus Primary Screening.

While cervical cytology is accepted triage for human papillomavirus (HPV)-positive women, the efficiency of cervical screening could be improved by exploiting disease markers with higher specificity. CINtec PLUS triage alone and combined with HPV 16/18 genotyping was performed on ThinPrep samples from HPV-positive women. Clinical performance and the potential to reduce or expedite colposcopy referrals were evaluated. The 2-year sensitivity and specificity for cervical intraepithelial neoplasia grade 3 or worse (CIN3+) were 90% and 42%, respectively. Specificity was improved over HPV testing in equivocal cytology and could cut the referral rate by about 40%. When combined with HPV 16/18 genotyping, CINtec PLUS triage of the 12 other high-risk HPV genotypes generally demonstrated better sensitivity for CIN3+ than separate triage of non-type-specific HPV-positive women. This strategy could reduce colposcopy referrals by 31%. These findings highlight the potential of CINtec PLUS to improve management pathways in HPV-positive women. CINtec PLUS cytology represents a sensitive and efficient triage in HPV-positive women. The clinical performance of the dual-stain was most notable in women younger than 25 years and could potentially improve management pathways.

Evaluating Colposcopy with Dynamic Spectral Imaging During Routine Practice at Five Colposcopy Clinics in Wales: Clinical Performance

Background: The diagnostic accuracy of colposcopy is poor for detecting precancerous cervical lesions. Objectives: We assessed the performance of colposcopy for identifying cervical intraepithelial neoplasia grade 2 or worse (CIN2+), before and after including a dynamic spectral imaging (DSI) map that quantifies and maps acetowhitening to assist subsequent biopsy of suspicious lesions. Methods: Four hundred and twenty-five women were examined at a multi-center setting in Wales, of which 393 women were included in the final analysis. Results: For all referrals, the sensitivity of conventional colposcopy for histologically confirmed CIN2+ was 51.5%, the specificity was 92.0%, the positive predictive value was 56.7%, and the negative predictive value (NPV) was 90.4%. With the incorporation of the DSI map in predicting CIN2+, these became 84.8, 61.5, 30.8, and 95.3% respectively. The increase of sensitivity was statistically significant (p < 0.001). For the 236 women having colposcopy after low-grade (LG) cytology, with the incorporation of DSI, the sensitivity for CIN2+ increased from 27.3 to 86.4% (p < 0.001) and the NPV from 92.6 to 97.8%. Conclusions: Colposcopy with DSI results in improved sensitivity to detect CIN2+ and maintains a high NPV for all referrals and especially for those with LG referral cytology.