Human Papillomavirus (HPV) DNA Triage of Women with Atypical Squamous Cells of Undetermined Significance with cobas 4800 HPV and Hybrid Capture 2 Tests for Detection of High-Grade Lesions of the Uterine Cervix

Abstract

T he primary screening of cancer of the uterine cervix is based in most countries on Pap cytology testing. Women with an abnormal cytology are referred for colposcopy-directed biopsy to assess the presence and grade of cervical disease. In North America, for each new case of cervical cancer, there are between 50 and 100 cases of abnormal smears consistent with low-grade (LSIL) or high-grade squamous intraepithelial lesions (HSIL) (11). Additionally, there are at least twice as many cases of "atypical squamous cells of undetermined significance" (ASC-US). Up to 15% of women with ASC-US have underlying high-grade cervical intraepithelial neoplasia 2 (CIN2) or CIN3 (2). Three clinical options have been proposed by the American Society for Colposcopy and Cervical Pathology to refer women with ASC-US to colposcopy, namely, repeat cytology, immediate referral to colposcopy, and reflex testing with generic assays for detection of high-risk (HR) human papillomavirus (HPV) genotypes (7, 21).When liquid-based cytology is used or when cocollected samples for cytology and HR HPV DNA testing can be obtained, HR HPV testing is the preferred triage approach (21). Since the introduction of the Hybrid Capture 2 assay (HC2; Qiagen, Inc., Mississauga, Ontario, Canada), several new HPV generic assays have been developed for the detection of HR HPV for ASC-US triage (17). The cobas 4800 HPV test (Roche Diagnostics, Laval, Québec, Canada) is a novel assay that detects HPV66 in addition to the 13 HR HPV genotypes detected by HC2. Nevertheless, HPV66 is detected by HC2 due to cross-hybridization (7). One report found that cross-reactivity with low-risk genotypes was more frequently encountered with HC2 than with the cobas 4800 HPV test (14). Few studies have evaluated the screening performance of the cobas 4800 HPV test (4,6,12,14,15,19,22).In this work, we compared the clinical performance of cobas 4800 HPV to that of HC2 on clinical specimens collected from women referred for colposcopy because of an ASC-US smear. HPV genotypic analysis was also performed with the Linear Array HPV genotyping (LA) assay (Roche Diagnostics, Laval, Canada) on all samples to assess cross-reactivity of the HPV generic assays. MATERIALS AND METHODSStudy design and population. Participants were recruited consecutively from December 2005 to December 2007 if they were referred for colposcopy because of at least one ASC-US cytology, were Ն24 years old, and had not received treatment for CIN in the last 2 years (10). Cervical cells were first collected with a Cytobrush for a conventional cytology. A second cervical specimen was collected with a Cytobrush prior to colposcopic examination. The Cytobrush was washed into PreservCyt collection medium (Hologic, Inc., Marlborough, MA) and kept at room temperature for 1 week and at Ϫ20°C thereafter. Samples in PreservCyt were processed for HPV DNA testing within 1 week for HC2 testing and LA and 3 to 5 years later depending on patients' inclusion dates for cobas 4800 HPV (Roche Diagnostics) testing. Colposcopy-guided b...

Comparison of the Abbott RealTi m e High-Risk Human Papillomavirus (HPV), Roche Cobas HPV, and Hybrid Capture 2 Assays to Direct Sequencing and Genotyping of HPV DNA

“…In recently published studies, Cobas HPV also showed comparable performances to HC2, with kappa coefficients from 0.69 to 0.87, but there also were discrepant results between HC2 and Cobas HPV in approximately 6% to 15% of the samples (10,20,34). In another recent study, Cobas HPV showed a clinical sensitivity of 89.7% and a specificity of 66.7% for detecting cases with CIN2 or worse-grade CINs, whereas HC2 demonstrated a better clinical sensitivity of 93.1% and a specificity of 72.2% (18). Differences in concordance rates between the HPV detection assays among the studies might result from differences in the analytical sensitivities for different HPV genotypes and differences in HPV genotypes among the constituent of subjects with HPV infections.…”

“…The two real-time PCR assays are suggested by their respective manufacturers to be able to detect 14 HR HPV genotypes (HPV types 16,18,31,33,35,39,45, 51, 52, 56, 58, 59, 66, and 68). The results of the two assays are reported as either positive or negative for HPV types 16 and 18 and a pooled result for the remaining 12 HR genotypes.…”

“…HC2 is a sandwich capture hybridization assay with chemiluminescence using unlabeled single-stranded RNA probes (23). After denaturation of nucleic acids, the single-stranded HPV DNA from the specimen was hybridized with mixed RNA probes specific for 13 HR HPV genotypes (HPV types 16,18,31,33,35,39,45,51,52,56,58,59, and 68). RNA-DNA hybrids were captured on the surface of antibody-coated microplates.…”

“…More than 150 HPV genotypes have been identified, and approximately 50 of them are known to infect cervical epithelia. Recently, the International Agency for Research on Cancer (IARC) defined 12 high-risk (HR) HPV genotypes (HPV types 16,18,31,33,35, 39, 45, 51, 52, 56, 58, and 59) as group 1 carcinogens (3). In particular, HPV16 is known as the most common HPV type, contributing to approximately half of all invasive cervical cancer cases.…”

Evaluation of the Clinical Performance of the cobas 4800 HPV Test in Patients Referred for Colposcopy

“…Furthermore, the specificity of cobas was comparable to that of HC2 in women referred with ASCUS. Most previous studies have evaluated the cobas for triage of ASCUS smears (3,4,5); few, if any, have included LSIL. Interestingly, in women referred with LSIL, the cobas test was more specific than HC2 for detection of CIN2ϩ (55.2% compared to 35.1%; McNemar P ϭ 0.001).…”