Comparing triage algorithms using HPV DNA genotyping,HPV E7 mRNA detection and cytology in high-risk HPV DNA-positive women

BackgroundHigh-risk human papillomavirus (hrHPV) DNA positive women require triage testing to identify those with high-grade cervical intraepithelial neoplasia or cancer (≥CIN2).ObjectiveComparing three triage algorithms (1) E7 mRNA testing following HPV16/18/31/33/45/52/58 genotyping (E7 mRNA test), (2) HPV16/18 DNA genotyping and (3) cytology, for ≥CIN2 detection in hrHPV DNA-positive women.Study designhrHPV DNA-positive women aged 18–63 years visiting gynecology outpatient clinics were included in a prospective observational cohort study. From these women a cervical scrape and colposcopy-directed biopsies were obtained. Cervical scrapes were evaluated by cytology, HPV DNA genotyping by bead-based multiplex genotyping of GP5+6+-PCR-products, and presence of HPV16/18/31/33/45/52/58 E7 mRNA using nucleic acid sequence-based amplification (NASBA) in DNA positive women for respective HPV types. Sensitivities and specificities for ≥CIN2 were compared between E7 mRNA test and HPV16/18 DNA genotyping in the total group (n = 348), and E7 mRNA test and cytology in a subgroup of women referred for non-cervix-related gynecological complaints (n = 133).ResultsSensitivity for ≥CIN2 of the E7 mRNA test was slightly higher than that of HPV16/18 DNA genotyping (66.9% versus 60.9%; ratio 1.10, 95% CI: 1.0002–1.21), at similar specificity (54.8% versus 52.3%; ratio 1.05, 95% CI: 0.93–1.18). Neither sensitivity nor specificity of the E7 mRNA test differed significantly from that of cytology (sensitivity: 68.8% versus 75.0%; ratio 0.92, 95% CI: 0.72–1.17; specificity: 59.4% versus 65.3%; ratio 0.91, 95% CI: 0.75–1.10).ConclusionFor detection of ≥CIN2 in hrHPV DNA-positive women, an algorithm including E7 mRNA testing following HPV16/18/31/33/45/52/58 DNA genotyping performs similar to HPV16/18 DNA genotyping or cytology.     

Predominance of High-Risk Human Papillomavirus Genotype 16 and 39 in Women with Premalignant and Malignant Cervical Pathology from Raipur,Chhattisgarh: Clinical Evaluation of Tagging Oligonucleotide Cleavage and Extension Mediated Genotyping Assay

Background: Identification of 14 high-risk human papillomavirus (HR-HPV) is immensely important in elucidating molecular epidemiology, patient monitoring and evidence-based treatment. There is paucity of such data from Chhattisgarh state of Central India. The present study has evaluated tagging oligonucleotide cleavage and extension-mediated Anyplex HR-HPV genotyping assay in identification of 14 HR-HPV genotypes attributable to premalignant and malignant cervical lesion in comparison to GP5+/6+ assay, cytology and colposcopy. Materials and Methods: A total of 185 clinically suspected cases of premalignant and malignant cervical lesion were investigated by HR-HPV genotyping, GP5+/6+, cytology and colposcopy. Results: Genotyping assay showed clinical sensitivity and specificity of 86.5% (confidence interval [CI]: 80.7–91.0) and 100% (CI: 86.3–100) respectively and found noninferior to GP5+/6+ assay (P > 0.05). HR-HPV prevalence was 76.3%, 88.4%, 94.8%, 100% and 100% among cervical intraepithelial neoplasia (CIN) Grade I–III, squamous cell carcinoma and adenocarcinoma cases, respectively. The four most common genotypes detected in CIN I–III were HPV 16 (63.9%), HPV 39 (15.0%), HPV 18 (6.0%) and HPV 33 (5.3%). In cervical cancer (CC) cases, HPV 16 (44.4%), HPV 39 (11.1%), dual infection of HPV 16, 18 (11.1%) and triple infection of HPV 16, 18, 33 (11.1%) were the four most identified genotypic aetiologies. A novel coinfection of HR-HPV 35, 39 were found in two and one cases of CIN I and II. Finding of HPV 39 as the second most prevalent genotype was unusual and underscores the importance of genotyping screening. Conclusion: Anyplex HR-HPV assay is arguably the useful assay for better patient management and can be useful for HR-HPV screening by its unique individual genotype identification of all HR-HPV. Finding of HPV 16, 39, 18, 33 and coinfection of 16,18 and 16, 18, 33 in CIN and CC would help vaccine manufacturer to design specific future HPV polyvalent vaccine preparation to curb down the CC-associated mortality.     

Feasibility study of a human papillomavirus E6 oncoprotein test for diagnosis of cervical precancer and cancer

In a feasibility study using a prototype, lateral-flow test for human papillomavirus type 16, 18, and/or 45 (HPV16/18/45) E6 oncoproteins, 51 of 75 (68%; 95% confidence interval [95% CI] of 56 to 78%) of HPV16/18/45 DNA-positive specimens from women with a diagnosis of CIN3+ (cervical intraepithelial neoplasia grade 3+ or cervical cancer) tested positive for HPV16/18/45 E6 oncoprotein. None of 16 (95% CI of 0 to 37%) HPV16/18/45 DNA-positive cervical specimens from women with a negative or CIN1 diagnosis tested positive for HPV16/18/45 E6 oncoprotein.     

Aptima HPV E6/E7 mRNA test is as sensitive as Hybrid Capture 2 Assay but more specific at detecting cervical precancer and cancer

Detection of human papillomavirus (HPV) E6/E7 oncogene expression may be more predictive of cervical cancer risk than testing for HPV DNA. The Aptima HPV test (Gen-Probe) detects E6/E7 mRNA of 14 oncogenic types. Its clinical performance was compared with that of the Hybrid Capture 2 DNA test (HC2; Qiagen) in women referred for colposcopy and those routinely screened. Aptima was also compared with the PreTect HPV-Proofer E6/E7 mRNA assay (Proofer; Norchip) in the referral population. Cervical specimens collected in PreservCyt (Hologic Inc.) were processed for HPV detection and genotyping with the Linear Array (LA) method (Roche Molecular Diagnostics, Laval, Quebec, Canada). Histology-confirmed high-grade cervical intraepithelial neoplasia (CIN 2) or worse (CIN 2+) served as the disease endpoint. On the basis of 1,418 referral cases (CIN 2+, n = 401), the sensitivity of Aptima was 96.3% (95% confidence interval [CI], 94.4, 98.2), whereas it was 94.3% (95% CI, 92.0, 96.6) for HC2. The specificities were 43.2% (95% CI, 40.2, 46.2) and 38.7% (95% CI, 35.7, 41.7), respectively (P < 0.05). In 1,373 women undergoing routine screening (CIN 2+, n = 7), both Aptima and HC2 showed 100% sensitivity, and the specificities were 88.3% (95% CI, 86.6, 90.0) and 85.3% (95% CI, 83.5, 87.3), respectively (P < 0.05); for women ≥ 30 years of age (n = 845), the specificities were 93.9% (95% CI, 92.3, 95.5) and 92.1% (95% CI, 90.3, 93.9), respectively (P < 0.05). On the basis of 818 referral cases (CIN 2+, n = 235), the sensitivity of Aptima was 94.9% (95% CI, 92.1, 97.7) and that of Proofer was 79.1% (95% CI, 73.9, 84.3), and the specificities were 45.8% (95% CI, 41.8, 49.8) and 75.1% (95% CI, 71.6, 78.6), respectively (P < 0.05). Both Aptima and Proofer showed a higher degree of agreement with LA genotyping than HC2. In conclusion, the Aptima test is as sensitive as HC2 but more specific for detecting CIN 2+ and can serve as a reliable test for both primary cervical cancer screening and the triage of borderline cytological abnormalities.     

Anyplex II HPV28 detection and Anyplex II HPV HR detection assays are highly concordant with other commercial assays for detection of high-risk HPV genotypes in women with high grade cervical abnormalities

Purpose: to evaluate the performance of Anyplex II HPV28 and HPV HR Detection assays against the EuroArray HPV, Cobas 4800 HPV (Cobas), HPV Amplicor (Amp), Linear Array HPV (LA) and Hybrid Capture 2 (HC2) in detection of high-risk HPV (HR-HPV) from liquid-based cervical cytology samples. Methods: cervical specimens from 404 women undergoing management of high-grade cytological abnormality were evaluated by Anyplex II HPV28 and HPV HR Detection assays for detection of HR-HPV genotypes and prediction of histologically-confirmed cervical intraepithelial neoplasia grade 2 or higher (≥CIN2). The results were compared to EuroArray, HC2, Cobas, Amp, and LA. Results: specimens were evaluated from 404 women with an average age of 30 years, including 336 with a histological diagnosis of?≥?CIN2 and 68 with?≤?CIN1. Concordance of HR-HPV detection between Anyplex II HPV28 and other genotyping assays was 94.79 % (κ?=?0.84; EuroArray) and 97.27 % (κ?=?0.91; LA); and between Anyplex II HPV HR and other HR-HPV detection assays was 86.35 % (κ?=?0.62; HC2), 96.03 % (κ?=?0.87; Cobas) and 96.77 % (κ?=?0.89; Amp). Using HR-HPV detection for prediction of?≥?CIN2 by Anyplex II HPV28 and HPV HR, sensitivity (90.18, 95 % CI 86.48–93.14; 90.77, 95 % CI 87.16–93.65) and specificity (both 67.16, 95 % CI 54.60–78.15) were not significantly different to the other HPV assays tested, with one exception. Both Anyplex assays had significantly higher sensitivity than HC2 (p?<?0.0001), with a specificity of 96 % (p?>?0.05) of HC2 in this high-risk population. Conclusions: both Anyplex II HPV detection assays were concordant with other commercial assays for HR-HPV detection, with comparable sensitivity and specificity for?≥?CIN2 detection.     

Clinical validation of Anyplex™ II HPV HR Detection according to the guidelines for HPV test requirements for cervical cancer screening

BackgroundAnyplex™ II HPV HR Detection (Seegene, Seoul, Korea) is a multiplex real-time PCR using tagging oligonucleotide cleavage and extension (TOCE) technology for simultaneous detection and genotyping of 14 high-risk (HR) HPV types, including HPV16 and HPV18.ObjectivesTo evaluate whether the clinical performance and reproducibility of Anyplex™ II HPV HR Detection meet the international consensus guidelines for HPV test requirements for cervical cancer screening [1].Study designThe clinical performance of Anyplex™ II HPV HR Detection for detecting cervical intraepithelial neoplasia grade 2 or worse (CIN2+) was determined relative to that of the reference assay, i.e., HR HPV GP5+/6+-PCR-EIA, by analysis of a total of 879 cervical liquid based cytology (LBC) specimens from a screening population, of which 60 were from women with CIN2+. The intra-laboratory reproducibility and inter-laboratory agreement were determined on 509 LBC samples, of which 172 were positive by the reference assay.ResultsAnyplex™ II HPV HR Detection showed a clinical sensitivity for CIN2+ of 98.3% (59/60; 95% CI: 89.1–99.8) and a clinical specificity for CIN2+ of 93.6% (764/816; 95% CI: 89.8–96.1). The clinical sensitivity and specificity were non-inferior to those of HR HPV GP5+/6+-PCR-EIA (non-inferiority score test: P = 0.005 and P = 0.023, respectively). Both intra-laboratory reproducibility (96.8%; 95% CI: 95.3–98.1; kappa value of 0.93) and inter-laboratory agreement (96.0%; 95% CI: 94.3–97.4; kappa value of 0.91) were high.ConclusionsAnyplex™ II HPV HR Detection performs clinically non-inferior to HR HPV GP5+/6+-PCR-EIA. Anyplex™ II HPV HR Detection complies with international consensus validation metrics for HPV DNA tests for cervical cancer screening [1].     

Clinical Validation of the HPV-Risk Assay,a Novel Real-Time PCR Assay for Detection of High-Risk Human Papillomavirus DNA by Targeting the E7 Region

The HPV-Risk assay is a novel real-time PCR assay targeting the E7 region of 15 high-risk human papillomavirus (HPV) types (i.e., HPV16, -18, -31, -33, -35, -39, -45, -51, -52, -56, -58, -59, -66, -67, and −68), and provides additional genotype information for HPV16 and HPV18. This study evaluated the clinical performance and reproducibility of the HPV-Risk assay with cervical scraping specimens and its utility with self-collected (cervico)vaginal specimens. The clinical performance of the HPV-Risk assay for cervical intraepithelial neoplasia of grade 2 or worse (CIN2+) with cervical scraping specimens was evaluated by a noninferiority analysis, relative to high-risk HPV GP5+/6+ PCR, following international guidelines for HPV test requirements for cervical cancer screening. The HPV-Risk assay showed clinical sensitivity for CIN2+ of 97.1% (95% confidence interval [CI], 89.1 to 99.3%; 67/69 samples) and a clinical specificity for CIN2+ of 94.3% (95% CI, 92.5 to 95.7%; 777/824 samples). The clinical sensitivity and specificity were noninferior to those of GP5+/6+ PCR (noninferiority score test, P = 0.006 and 0.0003, respectively). Intralaboratory reproducibility over time (99.5% [95% CI, 98.6 to 99.8%]; 544/547 samples, kappa = 0.99) and interlaboratory agreement (99.2% [95% CI, 98.6 to 99.8%]; 527/531 samples, kappa = 0.98) for the HPV-Risk assay with cervical scraping specimens were high. The agreement of the HPV-Risk assay results for self-collected (cervico)vaginal specimens and clinician-obtained cervical scraping specimens was also high, i.e., 95.9% (95% CI, 85.1 to 99.0%; 47/49 samples, kappa = 0.90) for self-collected lavage samples and 91.6% (95% CI, 84.6 to 95.6%; 98/107 samples, kappa = 0.82) for self-collected brush samples. In conclusion, the HPV-Risk assay meets the cross-sectional clinical and reproducibility criteria of the international guidelines for HPV test requirements and can be considered clinically validated for cervical screening purposes. The compatibility of the HPV-Risk assay with self-collected specimens supports its utility for HPV self-sampling.     

Clinical significance of signal pattern of high-risk human papillomavirus using a novel fluorescence in situ hybridization assay in cervical cytology

The present study aimed to evaluate a novel fluorescence in situ hybridization (FISH) assay for detecting the high-risk human papillomavirus (HR-HPV) DNA and signal pattern in cervical cytology specimens and for identifying cervical intraepithelial neoplasia (CIN) lesions. One hundred and ninety-six liquid-based cytology specimens with CIN were recruited. The signal pattern (punctate, mixed punctate and diffuse, and diffuse) detected by FISH was compared with E6 mRNA and correlated with histological classification. FISH and E6-type specific polymerase chain reaction (PCR) had fair to good agreement for detecting HPV DNA across all grades of CIN (kappa coefficient, 0.37–0.73). Among 44 samples of negative FISH and positive E6 type-specific PCR in HPV 16, 18, 31, 33, 52 and 58, 82% (36/44) of E6 mRNA were not detected, in contrast to 41% (48/118) of positive FISH and positive E6 type-specific PCR (p <0.0001). Among HR-HPV DNA positive cases tested by the FISH assay, the specificity of predicting CIN3 using the punctuate pattern is higher than that using E6 mRNA (96.3% vs. 44.8%). The punctate pattern was 0% in patients with <CIN1 lesions, 8.7% for CIN1 lesions, 6.1% for CIN2 lesions, and 34.0% for CIN3 lesions (p 0.001). The odds ratios were 8.7-fold higher (2.7–27.8, p <0.0001) for the punctate pattern versus the mixed punctate and diffuse pattern, and the diffuse pattern, for predicting CIN3 lesions. The novel FISH assay is comparable to PCR for detecting HPV DNA in cervical cytology with CIN lesions. The punctate signal pattern detected by the FISH assay can be more biologically and clinically relevant for clinically detecting CIN3 lesions.     

Aberrant expression of VEGF-C is related to grade of cervical intraepithelial neoplasia (CIN) and high risk HPV, but does not predict virus clearance after treatment of CIN or prognosis of cervical cancer

AIMS: Increased angiogenesis leads to invasion in cervical cancer. Vascular endothelial growth factors (VEGFs) are involved in angiogenesis, but molecular links to the most important aetiological agent, human papillomavirus (HPV), need clarifying. MATERIAL/METHODS: Archival samples-150 squamous cell carcinomas (SCCs) and 152 cervical intraepithelial neoplasia (CIN) lesions-were examined immunohistochemically for anti-VEGF-C antibody and for HPV by polymerase chain reaction (PCR). Follow up data were available for all SCC cases, and 67 CIN lesions were monitored with serial PCR to assess HPV clearance/persistence after treatment. RESULTS: High risk (HR) HPV types were closely associated with CIN (odds ratio, 19.12; 95% confidence interval, 2.31 to 157.81) and SCC (27.25; 3.28 to 226.09). There was a linear increase of VEGF-C expression-weak in CIN1 and intense in CIN3 and SCC (20.49; 8.69 to 48.26). VEGF-C upregulation was a sensitive (93.5%; 95% CI, 90.1% to 96.9%) marker of HR-HPV type (4.70; 2.17 to 10.21), but lost its significance in multivariate regression-p16(INK4a) and survivin were equally strong independent predictors of HR-HPV. Aberrant expression of VEGF-C did not predict clearance/persistence of HR-HPV after treatment of CIN. In cervical cancer, VEGF-C had no prognostic value in univariate or multivariate survival analysis. After adjustment for HR-HPV, FIGO stage, age, and tumour grade, only FIGO stage and age remained independent prognostic predictors. CONCLUSIONS: VEGF-C is an early marker of cervical carcinogenesis, with linearly increasing expression starting from low grade CIN. VEGF-C expression is closely related to HR-HPV in cervical lesions, probably because of its p53 independent upregulation by the E6 oncoprotein of HR-HPV.     

Evaluation of the clinical performance of OncoPredict HPV® SCR assay within the VALGENT-2 framework

Human papillomavirus (HPV) assays used in cervical cancer screening should be clinically validated according to international criteria. OncoPredict HPV® Screening (SCR) is a partial genotyping multiplex real-time PCR assay targeting E6/E7 genes of 13 high-risk (hr) HPVs. OncoPredict HPV® SCR (index assay) identifies HPV-16 and HPV-18 separately, 11 other hrHPV in aggregate and includes quality controls for sample adequacy, DNA extraction efficiency and PCR inhibition. 1300 VALGENT-2 study samples (from women aged 20–60 attending the Scottish cervical cancer screening program) were tested with the index assay and the GP5+/6+ PCR enzyme immunoassay (standard comparator assay). Non-inferior accuracy detecting cervical intraepithelial neoplasia of grade 2 or worse (CIN2+) of the index versus comparator was verified. Intra- and interlaboratory reproducibility of the index was evaluated by overall concordance and Cohen's kappa, using a sub-population (n = 526). Relative sensitivity and specificity for CIN2+ of the index versus comparator were 1.01 (95% confidence interval [CI]: 0.99–1.03) and 1.02 (95% CI: 1.0–1.04), respectively. Noninferiority p values were all ≤0.05, except for CIN3+ in patients ≥30 years. Excellent intra- and interlaboratory reproducibility was shown with concordance >98% and kappas >0.95. OncoPredict HPV® SCR fulfills the three international validation criteria for hrHPV DNA tests in cervical cancer screening.     

Human papillomavirus genotyping as a predictor of high-grade cervical dysplasia in women with mildly cytologic abnormalities: a two-year follow-up report

High-risk human papillomavirus (HR-HPV) DNA testing has emerged as another testing modality for women with mildly cytologic abnormalities. We conducted a two-year follow-up study of 108 women with mildly abnormal cervical cytology for detection of CIN 2/3. A cervical swab sample was obtained for HPV genotyping by a HPV blot and histologic follow-up results were correlated with HR-HPV types. Of the 108 cases, 93 (86.1%) were positive for HR-HPV DNA. HPV-16 was detected in 45.1% of patients. CIN grade 2 or 3 was confirmed in 25 (23.1%) of the 108 women during the two-year follow-up period. The two-year cumulative incidence rates of CIN 2/3 were 38.6% (17/44) among HPV-16- positive women, but only 5.6% among HR-HPV-positive women without HPV-16 or HPV-18. The sensitivity of a positive HPV-16 test for CIN 2/3 was 68.0%, the specificity was 67.5%. Our results demonstrated that the type-specific HPV-16 test increased sensitivity of detecting high-grade cervical dysplasia for women who have mildly cytologic abnormalities. The implication of the present findings is that HPV genotyping may identify women with the greatest risk of high-grade CIN.     

Immunohistochemical expression of p16(INK4a) is predictive of HR-HPV infection in cervical low-grade lesions.

The p16(INK4a) is a cyclin-dependent kinase inhibitor that decelerates the cell cycle by inactivating the cyclin-dependent kinases involved in the phosphorylation of the retinoblastoma protein (RB). Expression of E6 and E7 oncogenes of high-risk (HR) human papillomavirus (HPV), affecting the RB-p16 pathway, leads to p16 upregulation. Although it is widely reported that p16 is overexpressed in a high percentage of preneoplastic lesions and in almost all carcinomas of the uterine cervix, protein upregulation and its correlation with HPV infection in low-grade lesions is still being debated. In this study, we investigated in parallel, p16 expression and HPV infection in 100 cervical biopsies (17 normal tissues, 54 CIN1, 10 CIN2, 11 CIN3, eight invasive squamous cancers). Results obtained demonstrated that none of the 17 normal cervical tissues, evaluated by immunohistochemistry, presented p16 positivity whereas, starting from CIN1 (31%) to CIN2 (90%), CIN3 (100%) and carcinomas (100%), a constant and significant increase of protein overexpression (P<0.0001) was observed. In addition, p16 overexpression consistently showed elevated sensitivity (84%) and specificity (98%) in detecting HR-HPV infection with a high positive predictive value (97%) and negative predictive value (86%). Of interest, 93% of the p16-positive CIN1 were also HR-HPV infected. Our findings confirmed that p16 overexpression is associated to high-grade precancerous lesions and cervical carcinomas, and further demonstrated that immunohistochemical evaluation of p16 may be a useful biomarker in identifying HR-HPV-infected low-grade lesions.     

The Aptima HPV Assay Fulfills the Cross-Sectional Clinical and Reproducibility Criteria of International Guidelines for Human Papillomavirus Test Requirements for Cervical Screening

The Aptima HPV assay (Hologic Gen-Probe, San Diego, CA) is an FDA-approved assay for detecting human papillomavirus (HPV) E6/E7 mRNA from 14 high-risk HPV types. This study evaluated the clinical performance of the Aptima HPV assay for cervical intraepithelial neoplasia of grade 2 or worse (CIN2+), relative to the high-risk HPV GP5+/GP6+ PCR, in a cross-sectional clinical equivalence analysis using the noninferiority score test with cervical samples from population-based screening, i.e., 69 cervical scraping samples from women with CIN2+ and 843 from women without evidence of CIN2+. In addition, intralaboratory reproducibility over time and interlaboratory agreement of the Aptima HPV assay results were assessed with another set of 548 cervical samples. The Aptima HPV assay showed a clinical sensitivity for CIN2+ of 94.2% (95% confidence interval [CI], 85.5 to 97.8%) and a clinical specificity for CIN2+ of 94.5% (95% CI, 92.8 to 95.9%); by comparison, these figures were 97.1% (95% CI, 89.1 to 99.3%) (67/69 samples) and 93.6% (95% CI, 91.7 to 95.0%) (785/839 samples), respectively, for GP5+/GP6+ PCR. The clinical sensitivity and specificity of the Aptima HPV assay were noninferior to those of GP5+/GP6+ PCR (P = 0.039 and 0.00016, respectively). In addition, high reproducibility of the Aptima HPV assay, as reflected by the intralaboratory reproducibility over time of 96.0% (95% CI, 94.4 to 97.3%) (526/548 samples; kappa = 0.89) and interlaboratory agreement of 96.7% (95% CI, 95.4 to 98.1%) (531/548 samples; kappa = 0.91), was found. Altogether, these data show that the Aptima HPV assay meets the cross-sectional clinical and reproducibility criteria of the international guidelines for HPV test requirements for cervical screening. Longitudinal data are needed to ensure that the long-term negative predictive value of this mRNA assay is similar to those of validated HPV DNA tests.     

Antibodies against human papillomavirus type 16 and 18 E6 and E7 proteins in cervicovaginal washings and serum of patients with cervical neoplasia.

Serum antibodies against the E6 and E7 proteins of human papillomavirus (HPV) 16 and 18 are associated with cervical cancer. The aim of this study was to investigate the presence of local antibodies against HPV in cervicovaginal washings (CWs). In this study antibodies against the native HPV16 and HPV18 E6/E7 proteins were detectable in CWs (48%) and sera (29%) from patients with cervical cancer (n = 21) utilizing a sandwich protein enzyme-linked immunosorbent assay (ELISA). In paired CWs and sera from patients with cervical intraepithelial neoplasia (n = 38) and from healthy women (n = 22) no antibodies against these proteins were found. In 10 of 11 patients, the antibody response corresponded with the HPV type in the cervical smear and/or tumor tissue, which indicates the HPV type specificity of the assay. In 7 of 11 patients with antibody reactivity against HPV16 or HPV18 E6 and/or E7 proteins a higher level of antibody reactivity in the CWs than in the paired serum samples was found at similar inputs of total IgG. This suggests that the antibodies in the CWs against the investigated HPV proteins in these patients were locally produced.     

Comparison of Abbott RealTime High Risk HPV and Hybrid Capture 2 for the detection of high-risk HPV DNA in a referral population setting

Background

The Abbott RealTime High Risk HPV assay (ART) is an automated multiplex real-time PCR test for detection of DNA from 14 high risk (HR) HPV types in cervical specimens and simultaneous distinction of HPV16 and HPV18 from other HR-HPV.

Objectives

To evaluate the performance of the ART assay in specimens referred for HPV testing to our laboratory (referral population) by comparison with historical data from HC2 and INNO-LiPA as well as histological status, if available.

Study design

412 cervical specimens were collected from women between 18 and 70 years of age: 301 previously tested by HC2 without clinical data and 111 previously tested by HC2 and INNO-LiPA with histological diagnosis of CIN3+.

Results

Our study demonstrated good overall agreement between ART, HC2 and INNO-LiPA. In the group of the CIN3+ specimens HR-HPV was detected by ART in 93.07% (95% CI: 88.12-98.02), while HR-HPV detection rates with HC2 and INNO-LiPA were 91.09% (95% CI: 85.53-96.65) and 95.05% (95% CI: 90.82-99.28), respectively. The typing capability of ART for HPV16, HPV18 and a pool of twelve other HR-HPV types was investigated by comparison with INNO-LiPA demonstrating high overall assay concordance (89.81%; k 0.87).

Conclusions

The Abbott RealTime assay showed similar clinical performance for detection of CIN3+ compared with HC2. The high level of automation and ability to identify HPV16, HPV18 and other HR-HPV make this assay a very attractive option for HR-HPV testing, potentially improving patient management by risk stratification of cytological abnormal populations.     

Comparison of Onclarity Human Papillomavirus (HPV) Assay with Hybrid Capture II HPV DNA Assay for Detection of Cervical Intraepithelial Neoplasia Grade 2 and 3 Lesions

Analytical and clinical performance validation is essential before introduction of a new human papillomavirus (HPV) assay into clinical practice. This study compares the new BD Onclarity HPV assay, which detects E6/E7 DNA from 14 high-risk HPV types, to the Hybrid Capture II (HC2) HPV DNA test, to concurrent cytology and histology results, in order to evaluate its performance in detecting high-grade cervical lesions. A population of 567 women, including 325 with ≥ASCUS (where ASCUS stands for atypical cells of undetermined significance) and any HC2 result and 242 with both negative cytology and negative HC2 results, were prospectively enrolled for the study. The overall agreement between Onclarity and HC2 was 94.6% (95% confidence intervals [CI], 92.3% to 96.2%). In this population with a high prevalence of disease, the relative sensitivities (versus adjudicated cervical intraepithelial neoplasia grades 2 and 3 [CIN2+] histology endpoints) of the Onclarity and HC2 tests were 95.2% (95% CI, 90.7% to 97.5%) and 96.9% (95% CI, 92.9% to 98.7%), respectively, and the relative specificities were 50.3% (95% CI, 43.2% to 57.4%) for BD and 40.8% (95% CI, 33.9%, 48.1%) for HC2. These results indicate that the BD Onclarity HPV assay has sensitivity comparable to that of the HC2 assay, with a trend to an increased specificity. Moreover, as Onclarity gives the chance to discriminate between the different genotypes, we calculated the genotype prevalence and the absolute risk of CIN2+: HPV 16 was the most prevalent genotype (19.8%) with an absolute risk of CIN2+ of 77.1%.