Detection of high-grade cervical neoplasia using extended genotyping: Performance data from the longitudinal phase of the Onclarity trial.

OBJECTIVES: The Onclarity cervical cancer screening trial was designed to establish the clinical validity of the Onclarity HPV assay for extended genotyping (xGT) during detection of high-grade cervical neoplasia grades 2 or 3 (≥CIN2 or ≥CIN3). Here, three-year follow up data is presented to evaluate the overall efficacy of these screening strategies, compared to the baseline data. METHODS: At baseline 29,513 women, ≥25 years, had evaluable cytology and valid high-risk HPV results. Women with atypical squamous cells-undetermined significance or worse cytology or a positive HPV test were referred for colposcopy/biopsy. Participants that did not reach the study end point (treatment for ≥CIN2) continued into the longitudinal phase that included the same protocol as baseline. RESULTS: The three-year cumulative incident risk (CIR) for ≥CIN3 in HPV-negative women was 0.15% [95%CI: 0.06, 0.26] and for HPV- and cytology-negative women was 0.12% [95% CI: 0.03,0.23]. HPV16 carried the highest baseline and three-year ≥CIN3 CIR, followed by HPV31 and HPV18. At least one year of genotype-specific persistence increased ≥CIN3 risk for xGT results compared to genotype non-persistence, HPV clearance, or new infection over the same time period. Risk-based screening with immediate colposcopy for HPV16/18/31 and further xGT triage resulted in better ≥CIN3 sensitivity (79.2% versus 72.3%; relative difference of 6.9 [95%CI: 3.3, 10.4]) and a lower colposcopy/≥CIN3 ratio (9.2 versus 11.2; relative difference of -1.9 [95%CI: -2.6, -1.3]) when compared to primary HPV16/18-based screening. CONCLUSIONS: An HPV-negative result offers the same assurance of no disease over three years of follow up as that offered by a negative co-testing result. xGT facilitates risk-based screening and persistence tracking and can help optimize disease detection during screening without excessive colposcopic procedures.

Analytical and Clinical Sample Performance Characteristics of the Onclarity Assay for the Detection of Human Papillomavirus.

The objective of this study was to determine the result reproducibility and performance of the BD Onclarity human papillomavirus (HPV) assay (Onclarity) on the BD Viper LT platform using both contrived and clinical specimens. Reproducibility was assessed in BD SurePath liquid-based cytology (LBC) medium (SurePath) using contrived panels (HPV genotype 16 [HPV16] positive, HPV18 positive, or HPV45 positive) or clinical specimens (HPV16, -18, -31, -33/58, -45, or -52 positive or HPV negative). In addition, specimens from 3,879 individuals from the Onclarity trial were aliquoted prior to or following cytology processing and tested for HPV. Finally, specimens were collected using either the Cervex-Brush or Cytobrush (or Cytobrush/spatula) for comparison of HPV results. Contrived specimens showed >95% concordance with the expected results, and pooled clinical specimens had standard deviations and coefficients of variation ranging from 0.87 to 1.86 and 2.9% to 5.6%, respectively. For precytology and postcytology aliquot analyses, specimens showed >98.0% overall agreement and mean differences in cycle threshold (CT ) scores for HPV ranging from -0.07 to 0.31. Positivity rates were close between the Cervex-Brush and Cytobrush/spatula for all age groups tested. Onclarity results are reproducible and reliable, regardless of sample collection before or after cytology aliquoting. Onclarity performs well regardless of the method of specimen collection (Cervex-Brush or Cytobrush/spatula) for cervical cancer screening.

HPV infections and cytologic abnormalities in vaccinated women 21-34 years of age: Results from the baseline phase of the Onclarity trial.

OBJECTIVES: Countries with school-based human papillomavirus (HPV) vaccination have seen significant reductions in vaccine-targeted HPV infections, cytologic abnormalities, and high-grade cervical intraepithelial neoplasia (≥CIN2). However, the impact of HPV vaccination in the United States (where vaccination is largely opportunistic) may be less due to lower coverage rates and vaccination in patients at ages beyond the recommended routine vaccination age. METHODS: The Onclarity trial enrolled 33,858 subjects ≥21 years who were screened with cytology and the BD Onclarity HPV Assay. HPV positive women or those with cytologic abnormalities underwent colposcopy and biopsy. The prevalence of HPV, cytologic abnormalities, and ≥CIN2 was compared in a subset of 14,153, vaccinated and unvaccinated women, 21-34 years. Results were compared by vaccination status; Mantel-Haenszel analysis was performed to determine the association between vaccination status and prevalence, adjusting for age. RESULTS: The prevalence of overall HPV, HPV16, 18, 31, and 33/58 were all lower in vaccinated women for each age group; a significant difference (p < 0.001) was observed in vaccinated women for all ages combined. Cytologic low-grade squamous intraepithelial lesion (LSIL) or worse was lower in vaccinated women (p = 0.021), as was ≥CIN2 prevalence associated with HPV 16 or 18 (p = 0.011). CONCLUSIONS: Women with a prior history of HPV vaccination have a lower prevalence of any high-risk HPV, HPV 16, 18, 31, and 33/58; a cytology result of ≥LSIL, and ≥CIN2 associated with HPV 16/18 compared to unvaccinated women. A lower HPV prevalence in older, vaccinated women suggests that "catch-up" vaccination provides benefit.

Stratified risk of high-grade cervical disease using onclarity HPV extended genotyping in women, ≥25 years of age, with NILM cytology.

OBJECTIVES: Increasing evidence suggests that extended human papillomavirus (HPV) genotyping (beyond 16/18) is effective for risk stratification in women with normal cytology. This report provides extended genotyping results, using the BD Onclarity HPV Assay, for individual genotypes HPV16, 18, 31, 45, 51, and 52 ̶ and three pooled genotype results for HPV33/58, 35/39/68, and 56/59/66. METHODS: 27,037 women with normal cytology, ≥25 years, were enrolled into the Onclarity HPV trial during routine screening. Women positive for any HPV genotype were referred to colposcopy/biopsy. Hierarchical-ranked prevalence and risk values, associated with cervical intraepithelial neoplasia, grade 2 or worse (≥CIN2) or ≥CIN3, were calculated based on extended genotyping results. RESULTS: HPV 16 and 31 carried the highest risk for ≥CIN2 (11.6% and 12.1%, respectively) and ≥CIN3 (8.1% and 7.5%, respectively); these genotypes were the most prevalent in both ≥CIN2 (29.6% and 19.3%, respectively) and ≥CIN3 (43.7% and 22.5%, respectively). Of the other 12 genotypes, HPV 18, 33/58, and 52 comprised an intermediate risk band (≥CIN2 risk range: 4.9-6.8%; ≥CIN3 risk range: 3.9-5.0%). Genotypes 45, 51, 35/39/68, and 56/59/66 constituted the lowest risk band for both disease grades (≥CIN2 value risk range: 1.7-3.0%; ≥CIN3 value risk range: 1.2-3.6%). CONCLUSIONS: Extended genotyping stratifies risk for ≥CIN2/3 in the ≥25 year-old, normal cytology population. While baseline HPV 16/31 values exceeded the risk threshold for colposcopy referral, the management of women with normal cytology who were positive for the intermediate- or lower-risk genotypes may evolve based on refined risk estimates as well as clinical factors.

Detection of Cervical Neoplasia by Human Papillomavirus Testing in an Atypical Squamous Cells-Undetermined Significance Population: Results of the Becton Dickinson Onclarity Trial.

Objectives: To determine clinical utility of Onclarity human papillomavirus (HPV) assay for atypical squamous cells-undetermined significance (ASC-US) triage, and the value of HPV genotyping within ASC-US. Methods: Women (n = 33,858; 21 years or older) had HPV testing using Onclarity and Hybrid Capture 2 (HC2). ASC-US individuals (n = 1,960, 5.8%) were referred to colposcopy. Results: Of ASC-US, 39.1% were HPV positive by Onclarity; HPV 16 was the most prevalent genotype (7.4%). Cervical intraepithelial neoplasia grade 2 (CIN 2) and CIN 3+ prevalences were 4.4% and 2.2%, respectively. Onclarity had sensitivity for CIN 2+ (85.7%) and CIN 3+ (91.4%), and specificities for CIN 2+ (64.1%) and CIN 3+ (62.0%), similar to HC2. Risks for CIN 3+ were 16.1%, 2.8%, 2.5%, and 2.7% with HPV 16, 18, 45, and 11 other genotypes, respectively. Conclusions: Onclarity is clinically validated for ASC-US triage. Through risk stratification, genotyping could help identify women at highest risk for CIN 3+.

Risk of Cervical Intraepithelial Neoplasia 2 or Worse by Cytology, Human Papillomavirus 16/18, and Colposcopy Impression: A Systematic Review and Meta-analysis.

OBJECTIVE: To calculate pooled risk estimates for combinations of cytology result, human papillomavirus (HPV) 16/18 genotype and colposcopy impression to provide a basis for risk-stratified colposcopy and biopsy practice. DATA SOURCE: A PubMed search was conducted on June 1, 2016, and a ClinicalTrials.gov search was conducted on June 9, 2018, using key words such as "uterine cervical neoplasms," "cervical cancer," "mass screening," "early detection of cancer," and "colposcopy." METHODS OF STUDY SELECTION: Eligible studies must have included colposcopic impression and either cytology results or HPV 16/18 partial genotype results as well as a histologic biopsy diagnosis from adult women. Manuscripts were reviewed for the following: cytology, HPV status, and colposcopy impression as well as age, number of women, and number of cervical intraepithelial neoplasia (CIN) 2, CIN 3, and cancer cases. Strata were defined by the various combinations of cytology, genotype, and colposcopic impression. TABULATION, INTEGRATION, AND RESULTS: Of 340 abstracts identified, nine were eligible for inclusion. Data were also obtained from three unpublished studies, two of which have since been published. We calculated the risk of CIN 2 or worse and CIN 3 or worse based on cytology, colposcopy, and HPV 16/18 test results. We found similar risk patterns across studies in the lowest risk groups such that risk estimates were similar despite different referral populations and study designs. Women with a normal colposcopy impression (no acetowhitening), less than high-grade squamous intraepithelial lesion cytology, and HPV 16/18-negative were at low risk of prevalent precancer. Women with at least two of the following: high-grade squamous intraepithelial lesion cytology, HPV16- or HPV18-positive, and high-grade colposcopic impression were at highest risk of prevalent precancer. CONCLUSION: Our results support a risk-based approach to colposcopy and biopsy with modifications of practice at the lowest and highest risk levels.

The Onclarity Human Papillomavirus Trial: Design, methods, and baseline results.

OBJECTIVES: The baseline phase of the Onclarity trial was conducted to determine the screening performance of the Onclarity human papillomavirus (HPV) assay for detecting cervical cancer and precancer (≥CIN2) during triage of women ≥21 years with ASC-US cytology, as an adjunct test in women ≥30 years with normal cytology and for primary screening (HPV alone) in women ≥25 years. METHODS: 33,858 women ≥21 years were enrolled during routine clinic visits. All women with abnormal cytology, women ≥25 years that were high-risk HPV positive, and a random subset of women ≥25 years, negative by cytology and for HPV, were referred for colposcopy and cervical biopsy. Verification bias adjustment with 95% confidence intervals was applied. RESULTS: ASC-US prevalence was 5.8%. The overall HPV prevalence was 14.7%; for HPV 16, 18, and the 12 other HPV types it was 2.7%, 0.8%, and 11.2%, respectively. The prevalence of ASC-US and HPV was inversely proportional with age. The verification bias adjusted prevalence of ≥CIN2 and ≥CIN3 was 1.8% and 0.8%, respectively. Overall, five cases of cervical cancer were identified (all were HPV positive). The odds ratios associated with any HPV positive genotype, or with individual genotypes HPV 16, HPV 18, and HPV 31, for ≥CIN3, were statistically significant when compared to negative histology (p < 0.0001 for all). CONCLUSIONS: This report provides demographic information, cytology findings, HPV genotype information, and histopathology for participants in the baseline phase of this trial and offers further evidence to support genotype-specific screening for cervical cancer and precancer. Clinical Trial Registry URL:https://clinicaltrials.gov/ct2/show/NCT01944722.

Clinical Validation of a Test for the Diagnosis of Vaginitis.

OBJECTIVE: Vaginitis may be diagnosed as bacterial vaginosis, vulvovaginal candidiasis, trichomoniasis, or coinfection. A new molecular test assays the vaginal microbiome and organisms that cause three common infections. The objective of the trial was to evaluate the clinical accuracy of the investigational test for vaginal swabs collected by patients (self) or clinicians. The primary and secondary outcomes were to compare the investigational test with reference methods for the three most common causes of vaginitis and compare clinician-collected with self-collected swabs. METHODS: We conducted a cross-sectional study in which women with symptoms of vaginitis were recruited at ten clinical centers and consented to the investigation between May and September 2015. The woman collected a vaginal swab, sheathed, and then handed it to the clinician. These swabs were to evaluate how self-collected swabs compared with clinician-collected swabs. The clinician collected an investigational test swab and reference test swabs. From 1,740 symptomatic patients, clinician-collected and self-collected vaginal swabs were evaluated by the molecular test and six tests. The reference methods for bacterial vaginosis were Nugent's score and Amsel's criteria for intermediate Nugent results. The reference methods for Candida infection were isolation of any potential Candida microorganisms from inoculation of two culture media: chromogenic and Sabouraud agar and sequencing. The reference methods for trichomoniasis were wet mount and culture. RESULTS: For clinician-collected swabs, by reference methods, bacterial vaginosis was diagnosed in 56.5%, vaginal candidiasis in 32.8%, trichomoniasis in 8%, and none of the three infections in 24% with a coinfection rate of 20%. The investigational test sensitivity was 90.5% (95% confidence interval [CI] 88.3-92.2%) and specificity was 85.8% (95% CI 83.0-88.3%) for bacterial vaginosis. The investigational test sensitivity was 90.9% (95% CI 88.1-93.1%) and specificity was 94.1% (95% CI 92.6-95.4%) for the Candida group. Sensitivity for Candida glabrata was 75.9% (95% CI 57.9-87.8%) and specificity was 99.7% (95% CI 99.3-99.9%). Investigational test sensitivity was 93.1% (95% CI 87.4-96.3%) and specificity was 99.3% (95% CI 98.7-99.6%) for trichomoniasis. Results from self-collected swabs were similar to clinician-collected swabs. CONCLUSION: A molecular-based test using vaginal swabs collected by clinicians or patients can accurately diagnose most common bacterial, fungal, and protozoan causes of vaginitis. Women and their clinicians seeking accurate diagnosis and appropriate selection of efficacious treatment for symptoms of vaginitis might benefit from this molecular test.