Assessment of the Roche Linear Array HPV Genotyping Test within the VALGENT framework

doi:10.1016/j.jcv.2017.12.001

Journal of Clinical Virology

Volume 98, January 2018, Pages 37-42

https://doi.org/10.1016/j.jcv.2017.12.001 Get rights and content

Highlights

•
Cervical cancer screening with HPV tests should use only clinically validated assays.
•
This VALGENT study is the first to demonstrate that Linear Array HPV assay fulfils the international validation criteria for screening.
•
Linear Array HPV assay could become the standard comparator for HPV assays with genotyping capacity in the future.

Abstract

Background

Cervical cancer screening programs are switching from cytology-based screening to high-risk (hr) HPV testing. Only clinically validated tests should be used in clinical practice.

Objectives

To assess the clinical performance of the Roche Linear Array HPV genotyping test (Linear Array) within the VALGENT-3 framework.

Study design

The VALGENT framework is designed for comprehensive comparison and clinical validation of HPV tests that have limited to extended genotyping capacity. The Linear Array enables type-specific detection of 37 HPV types. For the purpose of this study, Linear Array results were designated as positive only if one of the 13 hrHPV types also included in the Hybrid Capture 2 (HC2) was detected. The VALGENT-3 framework comprised 1600 samples obtained from Slovenian women (1300 sequential cases from routine cervical cancer screening enriched with 300 cytological abnormal samples). Sensitivity for cervical intraepithelial neoplasia of grade 2 or worse (CIN2 + ) (n = 127) and specificity for <CIN2 (n = 1216) were assessed for Linear Array and for HC2 and non-inferiority of Linear Array relative to HC2 was checked. In addition, the prevalence of separate hrHPV types in the screening population, as well as the concordance for presence of HPV16, HPV18 and other hrHPV types between Linear Array and the Abbott RealTime High Risk HPV test (RealTime) were assessed.

Results

The clinical sensitivity and specificity for CIN2+ of the Linear Array in the total study population was 97.6% (95% CI, 93.3–99.5%) and 91.7% (95% CI, 90.0–93.2%), respectively. The relative sensitivity and specificity of Linear Array vs HC2 was 1.02 [95% CI, 0.98–1.05, (p < 0.001)] and 1.02 [95% CI, 1.01–1.03, (p < 0.001)], respectively The overall prevalence of hrHPV using the Linear Array in the screening population was 10.5% (95% CI, 8.9–12.3%) with HPV16 and HPV18 detected in 2.3% and 0.9% of the samples, respectively. Excellent agreement for presence or absence of HPV16, HPV18 and other hrHPV between Linear Array and RealTime was observed.

Conclusions

Linear Array showed similar sensitivity with higher specificity to detect CIN2+ compared to HC2. Detection of 13 hrHPV types with Linear Array fulfils the clinical accuracy requirements for primary cervical cancer screening.

Section snippets

Background and objectives

In 2012, the International Agency for Research on Cancer (IARC) concluded that at least 12 high-risk human papillomavirus (hrHPV) types (HPV16, HPV18, HPV31, HPV33, HPV35, HPV3, HPV45, HPV51, HPV52, HPV56, HPV58 and HPV59), were carcinogenic to humans for the development of cervical cancer (IARC-2009 hrHPV types)[1]. Furthermore, it has been demonstrated through several randomized controlled trials that hrHPV DNA testing is more effective than cervical cytology in primary screening of women

Sample collection

The collation of specimens used for the present iteration of VALGENT-3 project was performed in Slovenia, as previously described [7], [24]. Briefly, throughout December 2009 and August 2010, a total of 1300 consecutive cervical samples were obtained from women aged 25–64 years who participated in the national cervical cancer screening programme (screening population). Additionally, according to the VALGENT protocol [16], 300 cytological abnormal samples were collected between January 2014 and

Study population characteristics

The demographics and cytopathological results of the study population have already been described [24]. Briefly, the average age of women in the total study population (screening and enrichment population) was 39 years (range, 20–77), with 18.4% of the population <30 years. In the screening population (n = 1300), the cytological stratification was as follows²⁴: NILM (95.2%), ASC-US (2.4%), ASC-H (0.2%), atypical glandular cells (AGC, <0.1%), LSIL (1.0%), and HSIL (1.1%). When the total study

Discussion

The Linear Array is a frequently used HPV genotyping tests, which enables consensus and type-specific detection of 37 HPV types. Although the Linear Array was not intended for the use in primary HPV-based cervical cancer screening, it is often used in epidemiological studies and as a test to verify type coverage and cross-reactivity of hrHPV DNA assays with untargeted types [30], [31], [32]. For the purpose of this study, Linear Array was considered positive if one or more of the 13 hrHPV types

Conflict of interest

MA’s and LX's institution has received support from VALGENT (1–4) projects, as described previously in details in methodological VALGENT protocol paper (Arbyn2016JClinVirol16). MP’s and AO's institution received research grants from Abbott Molecular. Manufacturers of the evaluated HPV tests were not involved in the study design, data collection, data analysis and interpretation, or writing the manuscript.

Acknowledgement

All authors are supported by the seventh framework programme of DG Research of the European Commission, through the COHEAHR Network (grant No. 603019).

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Citation Excerpt :
This study provided data for two candidate screening assays that individually genotype multiple HRHPV types, Anyplex HPV28 and EuroArray, compared with the now defunct LA. Validation of these assays using a process based on the VALGENT (VALidation of HPV GENotyping Tests) framework for cervical cancer screening [8] should be considered. In this cohort of HIV-positive and HIV-negative Australian GBM, all four assays used to detect HPV on anal swabs performed similarly well for anal HPV16, with high levels of agreement for HPV16 detection between assays and lesion-level HPV16 detection.
Anal cancer is preceded by high-risk human papillomavirus (HRHPV) infection, predominantly HPV16. No HPV assay is licenced for use in anal screening. We aimed to determine the sensitivity and specificity of four anal canal swab HPV assays to predict high-grade squamous epithelial lesions (HSIL).
In a cohort of Australian HIV-positive and negative gay and bisexual men, we compared the sensitivity and specificity of detection of 13 anal HRHPV genotypes by Linear Array (LA), Cobas 4800, EuroArray, and Anyplex II HPV28 (+ and ++ cut offs), compared their ability to predict prevalent anal HSIL, and compared anal canal HRHPV detection with HRHPV isolated from HSIL using laser capture microdissection (LCM).
A total of 475 participants had baseline results available for all four assays (166, 35.0% HIV positive), and 169 participants had a diagnosis of cytological and/or histological HSIL. The HPV16 and any HRHPV detection were highest with Anyplex II HPV28 (+) (156, 32.8% 95% CI 28.6–37.2 and 359, 75.6%, 95% CI 71.5–79.4, respectively). For detection of concurrent HSIL and HPV16, the assay sensitivity was similar, ranging from 49.1%, 95% CI 41.4–56.9 (Anyplex II HPV28 ++) to 55.0%, 95% CI 47.2–62.7 (Anyplex II HPV28 +). For concurrent HSIL and any HRHPV detection, EuroArray was more specific than Anyplex II HPV28 (+) (45.9% 95% CI 40.2–51.7 vs 36.7%, 95% CI 31.3–42.4, p = 0.021) and had comparable specificity with Anyplex II HPV28 (++) (45.9% vs 47.2%, 95% CI 41.5–53.0, p = 0.75). All assays had high sensitivities for predicting HPV16 detected on LCM (92.5–97.5%). Anyplex II HPV28 and EuroArray were significantly more sensitive than LA for lesions caused by non-HPV16 HRHPV types on LCM.
Anyplex II HPV28 and EuroArray detected more non-16 HRHPV genotypes than LA. Increasing the Anyplex II HPV28 cutoff improved specificity without compromising sensitivity for detection of concurrent HSIL.
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The clinical performance evaluation of the novel MassARRAY human papillomavirus (MA-HPV) assay was performed using Danish SurePath cervical cancer screening samples under the fourth Validation of HPV Genotyping Tests (VALGENT) framework. The MA-HPV assay is a mass array–based assay that individually detects 14 oncogenic HPV genotypes and five nononcogenic types. The MA-HPV assay was validated using the VALGENT4 panel, which constitutes 997 consecutive samples from a screening population in addition to 297 disease-enriched samples with abnormal cytology findings. The clinical accuracy of the MA-HPV assay for sensitivity and specificity was assessed relative to that of the general primer 5+/6+ PCR enzyme immunoassay (GP-EIA), by a noninferiority test. The type-specific concordance of the MA-HPV assay was assessed as well. The relative sensitivity of the MA-HPV assay for cervical intraepithelial neoplasia ≥2 or ≥3 was 1.02 (95% CI, 0.98–1.05) and 1.01 (95% CI, 0.99–1.04), respectively. The sensitivity of the MA-HPV was noninferior to that of the GP-EIA (P = 0.0001), whereas the specificity of the MA-HPV was inferior (0.89; 95% CI, 0.85–0.91; P > 0.99). The MA-HPV assay is a clinical sensitive assay with a lower clinical specificity compared with the GP-EIA. The assay in its current form seems more suited to play a role where specificity is of lesser importance but where high sensitivity is paramount.
2020 list of human papillomavirus assays suitable for primary cervical cancer screening
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Only clinically validated HPV assays can be accepted in cervical cancer screening.
To update the list of high-risk HPV assays that fulfil the 2009 international validation criteria (Meijer-2009).
PubMed/Medline, Embase, Scopus, references from selected studies; published in January 2014 to August 2020.
HPV test validation studies and primary screening studies, involving testing with an index HPV test and a comparator HPV test with reporting of disease outcome (occurrence of histologically confirmed cervical precancer; CIN2+).
Women participating in cervical cancer screening.
Testing with an index and a comparator HPV test of clinician-collected cervical specimens and assessment of disease outcome (<CIN2, CIN2+). Comparator HPV assays were HC2, GP5+/6+ PCR-EIA, recommended in validation guidelines, or tests with consistent previous validations.
Assessment of relative clinical accuracy (including non-inferiority statistics index vs comparator assay) and test reproducibility in individual studies; random effects meta-analyses of the relative clinical sensitivity and specificity of index vs comparator tests.
Seven hrHPV DNA tests consistently fulfilled all validation criteria in multiple studies using predefined test positivity cut-offs (Abbott RealTime High Risk HPV, Anyplex II HPV HR Detection, BD Onclarity HPV Assay, Cobas 4800 HPV Test, HPV-Risk Assay, PapilloCheck HPV-Screening Test and Xpert HPV). Another assay (Alinity m HR HPV Assay) was fully validated in one validation study. The newer Cobas 6800 HPV Test, was validated in two studies against Cobas 4800. Other tests partially fulfilled the international validation criteria (Cervista HPV HR Test, EUROArray HPV, Hybribio's 14 High-Risk HPV, LMNX Genotyping Kit GP HPV, MALDI-TOF, RIATOL qPCR and a number of other in-house developed assays) since the non-inferior accuracy was reached after a posteriori cut-off optimization, inconsistent accuracy findings in different studies, and/or insufficient reproducibility assessment. The APTIMA HPV Assay targeting E6/E7 mRNA of hrHPV was fully validated in one formal validation study and showed slightly lower pooled sensitivity but higher specificity than the standard comparator tests in seven screening studies. However, the current international validation criteria relate to DNA assays. The additional requirement for longitudinal performance data required for non-DNA based HPV assays was not assessed in this review.
Eleven hrHPV DNA assays fulfil all requirements for use in cervical cancer screening using clinician-collected specimens.
Evaluation of HarmoniaHPV test for detection of clinically significant Human Papillomavirus infection using the VALGENT framework
2021, Journal of Virological Methods
The VALidation of HPV GENotyping Tests (VALGENT) is a framework for comparison and validation of HPV tests with genotyping capabilities. In this study, the clinical performance of a single tube HPV test -HarmoniaHPV- was assessed in SurePath™ samples and compared to a clinically validated reference test, the GP5+/6+ Enzyme ImmunoAssay (GP5+/6 + EIA).
HarmoniaHPV test is a real-time, PCR based, limited genotyping HPV test which detects 14 high-risk HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68 with HPV16, and HPV 18 reported individually. Clinical performance was assessed using 998 unselected, cervical screening samples enriched with 297 cytologically abnormal specimens (100 atypical squamous cells of unspecified significance, 100 low-grade squamous intraepithelial lesions, 97 high-grade squamous intraepithelial lesions). Cases were defined as women diagnosed with histologically confirmed cervical intraepithelial neoplasia 2 or more (≥CIN2, N = 122).
Using the manufacturer recommended (un-adjusted) cut-offs, HarmoniaHPV had non-inferior sensitivity for detection of ≥ CIN2 but showed inferior specificity. A cut-off optimisation exercise was therefore carried out and optimised cut-offs for each individual channel rendered a sensitivity and specificity of HarmoniaHPV that was non-inferior to GP5+/6 + EIA. Analytically, the test showed excellent intra- and inter-laboratory reproducibility, which improved further with the use of the optimised cut-offs.
HarmoniaHPV when operated with optimised cut-offs fulfils the international clinical criteria for use in cervical cancer screening on SurePath samples. The optimised cut-offs warrant additional testing and independent validation.
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2021, Journal of Virological Methods
Citation Excerpt :
By design, a VALGENT panel comprises a cohort combining a consecutively collected screening population (called the Screening population) and a cohort of women with abnormal cytology (called the Enriched population). To date, four VALGENT panels have been collected; VALGENT1/Belgium (Schmitt et al., 2013a, b; Schmitt et al., 2013c), VALGENT2/Scotland (Akbari et al., 2018; Cuschieri et al., 2016, 2015; Geraets et al., 2014; Heard et al., 2016), VALGENT3/Slovenia (Benoy et al., 2019; Ostrbenk et al., 2018; Polman et al., 2017; Viti et al., 2018; Xu et al., 2018a, b; Xu et al., 2020), and VALGENT4/Denmark (Bonde et al., 2018; Heideman et al., 2019; Bonde et al., 2019c; Ejegod et al., 2020b). Numerous HPV assays with different levels of genotyping have been assessed in the VALGENT studies, using the same comparator assays across all VALGENT installments.
The CLART HPV4S (CLART4S) is a novel full genotyping assay, based on PCR/microarray technology. We assessed the clinical accuracy of the CLART4S assays under the fourth installment of the VALGENT framework. The VALGENT cohort comprised 998 consecutive cervical samples from women participating in the Danish screening programme enriched with 297 samples with abnormal cytology (100 ASCUS, 100 LSIL, 97 HSIL). The CLART4S assay detects 16 HPV genotypes individually: 14 oncogenic and two non-oncogenic HPV types. The GP5+/6+ PCR Enzyme-Immuno-Assay (GP-EIA) and GP5+/6+ PCR with Luminex genotyping (GP-LMNX) were used as comparator tests for clinical accuracy and HPV genotype concordance, respectively. The sensitivity for ≥ CIN2 for the CLART4S assay was 96.7 % (GP-EIA: 92.6 %) with a relative sensitivity of 1.04 (1.00−1.09). The sensitivity for ≥ CIN3 was 98.8 % (GP-EIA: 94.0 %), with relative sensitivity of 1.05 (1.00−1.10). The specificity for <CIN2 was 88.6 % (GP-EIA: 89.2 %) with a relative specificity of 0.99 (0.98−1.01). The CLART 4S was found to be non-inferior to that of GP-EIA for both sensitivity (p < 0.0001) and specificity (p = 0.0452). The overall oncogenic HPV concordance between CLART4S and GP-LMNX was high, however when looking at individual genotype agreement the concordance was more diverse, with the highest agreement found in the Screening population.
Clinical validation of the Cobas 4800 HPV assay using cervical samples in SurePath medium under the VALGENT4 framework
2020, Journal of Clinical Virology
The VALidation of HPV Genotyping Tests (VALGENT) framework is an international cooperation designed for comparison and clinical validation of HPV assays with genotyping capabilities.
Here we addressed the accuracy of the Roche cobas 4800 HPV test using SurePath samples from the Danish cervical cancer screening program under the VALGENT framework.
The VALGENT4 panel comprises 998 consecutive SurePath cervical samples from routine screening and 297 SurePath samples enriched for disease (100 ASC-US, 100 LSIL, 97 HSIL). The cobas HPV test is a real-time PCR assay which detects HPV16 and 18 individually and 12 other high-risk (hr) HPV genotypes in one bulk.
The clinical performance of the cobas test was assessed relative to that of the comparator assay GP5+/6 + PCR Enzyme ImmunoAssay (GP-EIA) by a non-inferiority test. The relative sensitivity for ≥ CIN2 was 1.00 (95% CI: 0.97-1.04) and relative specificity for the control group was 1.02 (95% CI: 1.01-1.04). The cobas test was found non-inferior to that of GP-EIA for both sensitivity and specificity (p-0.0006 and p < 0.0001, respectively). The type specific performance of the cobas test was evaluated using the GP5+/6 + PCR with Luminex genotyping (GP-LMNX) as comparator. The cobas test showed excellent to good concordance (Kappa: 0.70 to 0.90) with GP-LMNX for all three genotype groups in the overall VALGENT population but good to moderate concordance in the Screening population (kappa from 0.56 to 0.80).
The cobas HPV test demonstrated non-inferiority to the comparator assay on cervical SurePath screening samples using the VALGENT4 panel.

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Assessment of the Roche Linear Array HPV Genotyping Test within the VALGENT framework

Highlights

Abstract

Background

Objectives

Study design

Results

Conclusions

Section snippets

Background and objectives

Sample collection

Study population characteristics

Discussion

Conflict of interest

Acknowledgement

Lancet Oncol.

Vaccine

Lancet Oncol.

Lancet Oncol.

J. Clin. Virol.

Clin. Microbiol. Infect.

J. Clin. Virol.

J. Clin. Virol.

Lancet Oncol.

Gynecol. Oncol.

Papillomavir Res.

Working group on the evaluation of carcinogenic risks to humans. biological agents: a review of human carcinogens

Guidelines for human papillomavirus DNA test requirements for primary cervical cancer screening in women 30 years and older

Int. J. Cancer

HPV screening for cervical cancer in Rural India

N. Engl. J. Med.

Comparison of clinical and analytical performance of the abbott RealTime high risk HPV test to the performance of hybrid capture 2 in population-Based cervical cancer screening

J. Clin. Microbiol.

Cross-sectional comparison of an automated hybrid capture 2 assay and the consensus GP5+/6+ PCR method in a population-based cervical screening program

J. Clin. Microbiol.

Efficacy of HPV DNA testing with cytology triage and/or repeat HPV DNA testing in primary cervical cancer screening

J. Natl. Cancer Inst.

Commercially available molecular tests for human papillomaviruses (HPV): 2015 update

J. Clin. Virol.

Commercially available assays for multiplex detection of alpha human papillomaviruses

Expert Rev. Anti Infect. Ther.