DNA- versus RNA-based methods for human papillomavirus detection in cervical neoplasia
Introduction
It has now been firmly established that infection with high-risk human papillomavirus (HPV) types is the primary cause of almost all cervical cancers [1], [2], [3]. Persistent infection with high-risk HPV types is the first step in the carcinogenesis [4]. However, the vast majority of women infected by high-risk HPV will not develop cervical cancer, as most HPV infections are transient, especially in the young, sexually active population. Women with active HPV infection will express E6/E7 oncogenes, which are required for malignant transformation by inhibiting the tumor suppressors p53 and RB [4]. E6/E7 mRNA transcripts are detected by mRNA-based molecular techniques [5], [6]. Standardized HPV DNA-based testing has shown that high-risk HPV DNA detection has higher sensitivity for histological verified high-grade cervical neoplasia compared to classical cytology [7], [8], [9], [10], [11], [12], [13]. However, the specificity as well as the positive predictive values (PPV) in these studies is low, especially when young women are tested. Persistent expression of E6/E7 oncogenes could serve as an indicator of progression to cervical intraepithelial neoplasia (CIN) and invasive cancer [14], [15]. RNA-based testing to detect E6/E7 mRNA transcripts may therefore be of higher prognostic value and improve the specificity and PPV compared to HPV DNA testing in screening. While DNA-based methods have been extensively evaluated in screening programs, the predictive values of mRNA based methods are unknown. Few studies concerning the extent of oncogenic expression in cervical neoplasia have so far been published [14], [15], [16], [17], [18], [19].
Recent meta-analyses have shown that the accuracy of PAP test varied greatly with sensitivity ranging from 30–87% [20], [21]. The high rate of false negative cytology has important public health and legal implications. Consequently, there has been substantial interest in the use of HPV testing in order to improve the efficiency of population-based screening programs for cervical cancer. It has been discussed if HPV testing may be used as primary screening for cervical cancer, as triage for equivocal or low-grade lesions and in the follow-up after treatment for CIN [22]. Secondary HPV testing will be implemented in the Norwegian cervical cancer screening program in 2005. The aims of the present study were to evaluate two commercially available assays for HPV testing to detect high-grade cervical neoplasia. The outcome of DNA-based and mRNA-based testing was compared with cytology and histology, using histology as the “gold-standard”.
Section snippets
Study population and collection of specimens
Women were recruited from 5 hospital based outpatients clinics and 10 gynecologists in private practice in Norway. Enrolment took place from October 2002 through October 2003. Included in this study were 383 women with median age 35 years (range, 19–85). The study population comprised women with positive cytology referred for colposcopy or conization. Both squamous and glandular lesions were included in the study. The Regional Committee for Medical Research Ethics, South Region, approved the
Results
Overview of the total material is shown in Fig. 1. The DNA test was positive in 95% (275/291), and the mRNA test was positive in 77% of the histological confirmed high-grade lesions (225/291). All invasive carcinomas were positive with the mRNA test. The outcome of HPV testing is shown in Table 3. High-risk HPV (DNA and/or mRNA) was present in 90% of the cases (344/383). Agreement between the two HPV tests was found in 72% of the cases (274/383). The DNA test was positive and the mRNA test
Discussion
The aim of many ongoing studies is to establish a place for HPV testing in routine screening. Both DNA- and RNA-based molecular techniques are now commercially available and need to be validated before inclusion in clinical practice. The aims of the present study were to compare commercially available DNA-based and mRNA-based tests for detection of high-grade histology among Norwegian women with positive index cytology.
All HPV types included in the two HPV assays can now be considered
Acknowledgments
We thank Shazia Begum, Bente Risberg, and Helen Vålerhaugen for excellent technical assistance, the gynecology clinics at Bærum, Hamar, Elverum, and St. Olavs Hospitals, and the following gynecologist: Alnæs, Benthien, Berild, Dalaker, Fjeld, Molnar, Norling, Riis-Johannessen, Skår, Trosterud.
Consumables for the Pre Tect HPV-Proofer assay were covered by Norchip AS.
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