Clinical and Analytical Evaluation of the Anyplex II HPV HR Detection Assay within the VALGENT-3 Framework

Abstract

In 2012, VALidation of human papillomavirus (HPV) GENotyping Tests (VALGENT) was initiated to provide a formalized and uniform study framework for comparison and validation of HPV assays with genotyping capability. In VALGENT-3, the clinical and analytical performance of Anyplex II HPV HR detection (Anyplex) was compared to that of the Hybrid Capture 2 HPV DNA test (hc2) and the cobas 4800 HPV test (cobas). The panel comprises 1,300 stored samples that were obtained from women 25 to 64 years old who participated in the Slovenian cancer screening program, enriched with 300 samples from women with abnormal cervical cytology. The sensitivity and specificity of Anyplex were noninferior to those of hc2, with a relative sensitivity of 1.01 (95% confidence interval [CI], 0.97 to 1.04) for cervical intraepithelial neoplasia grade 2 or worse (CIN2+) and 1.01 (95% CI, 0.97 to 1.06) for CIN3+ and relative specificity of 1.02 (95% CI, 1.00 to 1.03) for a CIN grade of ≤1. The clinical sensitivity of Anyplex for CIN2+ and CIN3+ was comparable to that of hc2 ( values for McNemar test [ ] of 0.655 and 0.564, respectively), but its specificity was significantly higher ( = 0.008). The sensitivity and specificity of Anyplex were also noninferior to those of cobas, with relative sensitivity of 1.01 (95% CI, 0.98 to 1.04) for CIN2+ and 1.01 (95% CI, 0.99 to 1.04) for CIN3+ and relative specificity of 1.00 (95% CI, 0.99 to 1.01) ( value of>0.05 in all cases). Regardless of the clinical outcome (CIN2+ or CIN3+), age restriction (women ≥30 years old), or comparator test used, Anyplex consistently showed excellent clinical performance and can be considered validated for primary cervical cancer screening.

Clinical and Analytical Performance of the BD Onclarity HPV Assay with SurePath Screening Samples from the Danish Cervical Screening Program Using the VALGENT Framework

“…DNA was extracted from the VALGENT4 panel samples as described previously (8) and was shipped to the DDL Diagnostics Laboratory (Rijswijk, The Netherlands), where all GP5ϩ/6ϩ PCR testing was performed. The clinically validated hrHPV GP-EIA for pooled detection of 14 oncogenic types (genotypes 16,18,31,33,35,39,45,51,52,56,58,59, 66, and 68) was used as a comparator for the clinical performance of the Onclarity assay. For genotype concordance analysis, a Luminex-based readout was used (GP-LMNX) for individual genotyping of the 14 HPV types (14).…”

“…HPV assay development over the past decade has led to a host of PCR-based assays reporting high-risk HPV (hrHPV) findings with various degrees of data resolution. In this regard, HPV assays can be stratified into four categories: (i) consensus assays that report only positive or negative outcomes resulting from measurement of the presence of the 13 or 14 most common oncogenic HPV genotypes (genotypes 16,18,31,33,35,39,45,51,52,56,58,59, 68, and sometimes 66); (ii) consensus assays with limited genotype reporting, often for HPV16 and HPV18; (iii) HPV assays with extended genotyping, typically identifying HPV16 and -18 combined with more, but not all, of the oncogenic genotypes; and (iv) full genotyping assays with individual reporting of 14 oncogenic HPV genotypes. The Hybrid Capture 2 assay (HC2; Qiagen, Hilden, Germany) and GP5ϩ/6ϩ PCR-enzyme immunoassay (GP-EIA; DDL Diagnostic Laboratory, Rijswijk, The Netherlands) have been considered standard comparator assays, since both have demonstrated longitudinal evidence of protection against cervical precancer and cancer through randomized trials (5,6).…”

“…To date, four VALGENT panels have been collected from the Belgian (VALGENT1) (9-11), Scottish (VALGENT2) (12)(13)(14)(15), Slovenian (VALGENT3) (16)(17)(18)(19)(20), and Danish (VAL-GENT4) (8) cervical cancer screening programs. VALGENT4 specifically comprises a panel of samples collected in SurePath medium; previously, the majority of clinically validated HPV assays for use in screening had been undertaken on samples collected with the ThinPrep system.…”

Accuracy of genotyping for HPV16 and 18 to triage women with low-grade squamous intraepithelial lesions: a pooled analysis of VALGENT studies

“…Up to now, three VALGENT panels have been completed with a fourth ongoing. VALGENT-1 were provided by the AML laboratory using SurePath collected samples (Antwerp, Belgium) [9][10][11][12][13], VALGENT-2 by the Scottish HPV Reference Laboratory using ThinPrep collected samples (Edinburgh, Scotland) [14][15][16], VALGENT-3 were performed using ThinPrep collected samples from the Laboratory for Molecular Microbiology of University of Ljubljana (Ljubljana, Slovenia) [17][18][19][20][21], and VALGENT-4 using fresh SurePath collected samples by the Molecular Pathology Laboratory of Copenhagen University Hospital (Copenhagen, Denmark) [22]. In each VALGENT panel, the study population comprised a continuous series of 1,000 or 1,300 cervical specimen (archived or fresh) from women participating in the local cervical cancer screening programme supplemented with 300 abnormal pathological samples (100 ASC-US, 100 LSIL and 100 high-grade squamous intraepithelial lesions [HSIL]) [5].…”

“…In each VALGENT panel, the study population comprised a continuous series of 1,000 or 1,300 cervical specimen (archived or fresh) from women participating in the local cervical cancer screening programme supplemented with 300 abnormal pathological samples (100 ASC-US, 100 LSIL and 100 high-grade squamous intraepithelial lesions [HSIL]) [5]. Detailed information about each panel collection, processing and manipulation can be found in previously published VALGENT reports [9][10][11][12][13][14][15][16][17][18][19][20][21][22].…”