Clinical impact of the analytical specificity of the hybrid capture 2 test: data from the New Technologies for Cervical Cancer (NTCC) study.

The Hybrid Capture 2 (HC2) test targets 13 human papillomavirus (HPV) types. Here, cross-reactivity with non-HC2-targeted HPV types is described. We aimed to define the proportion of HC2-positive women who had negative results with HC2-targeted HPV types and estimate its determinants and impact on women's health management. The New Technologies for Cervical Cancer (NTCC) trial was followed in two predetermined phases. Women in the experimental arm were tested for the presence of HPV DNA by HC2 following a sample collection in PreservCyt (first phase) or Digene specimen transport medium (STM) (second phase). HPV genotyping was performed on DNA samples from HC2-positive women by PCR with GP5(+)/GP6(+) primers and reverse line blot (RLB) hybridization. Untyped samples were submitted to direct sequencing or restriction fragment length polymorphism. Multivariate logistic regression analysis estimated the adjusted odds ratios (ORs) between the presence of HC2-targeted types and age, viral load, and type of transport medium. Out of 2,920 HC2-positive samples, 2,310 (79.1%) were positive on RLB for HC2-targeted types, 396 were positive (13.6%) for only non-HC2-targeted types (mostly represented by HPV-53, HPV-66, and HPV-70), and in 214 (7.33%) samples, no HPV types were detected. The probability of detecting HC2-targeted types increased with increasing viral load expressed as the relative light unit/positive-control specimen ratio (RLU/PC) (OR for unitary increase of log RLU/PC, 1.35; 95% confidence interval [CI], 1.30 to 1.42) and with STM versus PreservCyt (OR, 1.56; 95% CI, 1.25 to 1.84). If only the samples containing HC2-targeted types tested positive, the positive predictive value (PPV) would have increased from 7.0% (95% CI, 6.1% to 8.0%) to 8.4% (95% CI, 7.3 to 9.6), although 4.9% (95% CI, 2.4% to 8.8%) of cervical intraepithelial neoplasia grade 2(+) (CIN2(+)) cases would have been missed. In conclusion, STM use and an increased cutoff would reduce the HC2 analytical false-positive rate and increase the positive predictive value for high-grade CIN. The gain in clinical sensitivity by detecting non-HC2-targeted HPV types is limited.

Oral human papillomavirus and human herpesvirus-8 infections among human immunodeficiency virus type 1-infected men and women in Italy.

BACKGROUND: Oral human papillomavirus (HPV) and human herpesvirus-8 (HHV8) infections are sexually transmitted and respectively associated with the development of oropharyngeal carcinoma and Kaposi sarcoma. The aim of the study was to evaluate HPV prevalence and its possible correlation with HHV8 oral shedding, in relation to sex, human immunodeficiency virus (HIV) behavioral risk factor, and immune function. METHODS: The study population comprised 100 HIV-infected individuals divided into 3 groups: (1) 38 men who have sex with men (MSM), (2) 24 heterosexual men, and (3) 38 women. DNA was obtained from cells of unstimulated whole saliva. Human papillomavirus sequences were searched for by polymerase chain reaction (PCR) with MY09/MY11 primers or by nested PCR with GP5+/GP6+ primers as the second step. Typing was accomplished by restriction fragment length polymorphism analysis or by direct sequencing or by reverse line blot. Human herpesvirus-8 sequences were detected and quantified by nested PCR and real-time PCR, respectively. RESULTS: Oral HPV infection was present in 37 (prevalence, 37%) of 100 (13 with high-risk and 24 with low-risk types) patients; the most frequent types were HPV16, HPV6, HPV10, HPV61, HPV66, and HPV83. Human herpesvirus-8 DNA was detected in 46 (46%) of 100 subjects. Both infections had the highest prevalence among MSM and the lowest among women; women had a lower prevalence of high-risk HPV types than did both male groups (P = 0.05). An inverse correlation was observed with concomitant oral HHV8 infection (P = 0.007). CONCLUSIONS: High prevalence of oral HPV and HHV8 infections was observed; MSM had the highest figures, despite better control of HIV infection.

Triage of women with atypical squamous cells of undetermined significance (ASC-US): results of an Italian multicentric study.

OBJECTIVES: To compare the performance of immediate colposcopy, repeat Pap test and HPV test as triage options for women diagnosed as having atypical squamous cells of undetermined significance (ASC-US) while attending organised screening for cervical carcinoma in five centres of the Veneto region. METHODS: Women consecutively diagnosed as having ASC-US were included in a prospective study, and underwent colposcopy and collection of cervico-vaginal cells for conventional Pap test and HPV test (Hybrid Capture 2, High-risk probe set, Digene). Repetition of all three tests was scheduled for 12 months later. DNA was subsequently extracted from residual cells of positive samples, and analysed by polymerase chain reaction with several primers for typing of HPV sequences. Sensitivity, specificity and positive predictive value (PPV) of the different triage options for histology-confirmed cervical intraepithelial neoplasia, grade 2 or worse (CIN2+) were calculated among all women and by age (under and above 35 years). RESULTS: Seven hundred forty-nine women 25-64 years old (median age 42 years) were enrolled in the study. Pap smears at enrolment were read as ASC-US or more severe in 211 (29.4%) cases, colposcopy disclosed an atypical transformation zone in 254 (34.2%) women, and HPV test was positive in 181 (24.2%). High-grade cervical lesions developed in 29/749 (3.9%) women. HPV typing was possible in 163 (90%) of the samples, and carcinogenic types were present in 123. CONCLUSIONS: HPV test showed the best performance; overall, it had the highest sensitivity (92.3%), specificity (78.6%) and PPV (14.9%).

HPV-16 E6 L83V variant in squamous cell carcinomas of the upper aerodigestive tract.

PURPOSE: The aim of this prospective case series study was to determine the prevalence of HPV-DNA, analyze the E6 mRNA expression, identify intra-type variation in the E6 oncogene in upper aerodigestive tract (UADT) squamous cell carcinoma (SCC), and correlate the presence of HPV-DNA with several clinical parameters and outcome. METHODS: Frozen samples of UADT-SCC were analyzed for the presence and characterization of HPV-DNA and RNA sequences by means of polymerase chain reaction (PCR), reverse transcriptase-PCR, and direct sequencing of amplified products. RESULTS: HPV-DNA sequences were detected in 10% of the tumors, all of which were typed as HPV-16. Positivity for HPV-16 E6/E7 mRNA was observed in five of the eight HPV-positive tumors (62.5%). The HPV-16 E6 L83V variant was present in five cases. Multivariate analysis identified a history of absence of smoking (P = 0.009) as a predictor of HPV-positive tumor. No significant differences in overall and disease free survival curves were observed between patients with HPV-positive tumors and patients with tumors without detectable HPV-DNA. CONCLUSION: Our findings support the etiological participation of HPV-16 in a subset of UADT-SCCs from patients lacking traditional risk factors. The potential prognostic significance of HPV-16 E6 L83V variant in HPV-16 positive UADT-SCCs should be more extensively investigated.