RNA (E6 and E7) Assays versus DNA (E6 and E7) Assays for Risk Evaluation for Women Infected with Human Papillomavirus

Abstract

In the majority of cases, high-risk human papillomavirus (HR HPV) infections regress spontaneously, with only a small percentage progressing to high-grade lesions. Current screening methods are based on DNA detection. An alternative would be to monitor expression of the E6 and E7 viral oncogenes continuously expressed by malignant phenotypes. In the work reported in this paper, we compared the two methods for a group of women with high-risk HPV infections. Cervical specimens from 400 women, previously found to be HPV DNA positive, were analyzed for HPV DNA by a liquid hybridization assay and typed by multiplex PCR (for types 16, 18, 31, and 33). Identification of HR HPV E6 and E7 RNA transcripts was performed using real-time reverse transcription-PCR and nucleic acid sequence-based amplification assays. Results were compared with concurrent cytological data. HR HPVs were found in 61.2% of patients. The most common genotype was HPV type 16 (HPV-16) (47.1%), followed by HPV-18, HPV-31, and HPV-33. Nine percent of cases involved other genotypes. Among 223 HPV DNA-positive samples, only 118 were positive in the RNA test. Among HPV DNA-positive patients with normal cytology, we detected E6 and E7 RNA transcripts in two cases (18.2%). The rate of detection increased gradually with the grade of the observed lesions, rising from 20% for patients with atypical squamous cells of undetermined significance to 48.1% for women with low-grade squamous intraepithelial lesions and 86.3% for those with high-grade squamous intraepithelial lesions. These results suggest that testing for HPV E6 and E7 transcripts could be a useful tool for screening and patient management, providing more accurate predictions of risk than those obtained by DNA testing.Cancer of the cervix is the second most frequent gynecological malignancy in the world. It is well established that the main cause is infection with human papillomavirus (HPV) (30,43,46,48). However, only certain specific types of virus lead to cancer. We can thus distinguish between high-risk and low-risk HPVs (HR HPV and LR HPV, respectively) (1,6,12,31,43). Epidemiological studies of HPV show that more than 70% of cervical cancers worldwide are caused by HPV type 16 (HPV-16) and HPV-18. The remaining cases of malignancy are associated with other types of HR HPV (mainly . Today, these viruses are among the most important known risk factors for human cancer (6,20,31,38).Most HR HPV infections regress spontaneously 6 to 12 months after their appearance, probably due to successful attack by the immune system (33). Only a small percentage of infections persist. Without surgical treatment, these infections can progress to high-grade lesions and to squamous cell carcinoma or adenocarcinoma of the cervix (37,47,48).The introduction of the Pap smear test by Papanicolaou made it possible to identify precursor lesions and to significantly reduce mortality. However, the test cannot reliably predict whether a mild dysplasia will regress or progress (5, 36). Recently, it was suggested that cervica...

Clinical Performance of Human Papillomavirus E6 and E7 mRNA Testing for High-Grade Lesions of the Cervix

“…These findings are consistent with the HPV DNA prevalence rates reported by other Italian groups (1,6,14,51) and match the rates of prevalence from combined DNA-and RNA-based studies (2,7,10,13,24). As reported earlier, the study detected a number of mixed infections ( Table 4).…”

“…This suggests that the virus was present only at very low copy numbers and/or that only a specific region of viral DNA was integrated into the host genome. The loss of the L1 gene and its impact on viral replication after integration could lead to relatively low viral loads (7,15,24,32). Given that malignant phenotypes require continuous expression of the E6 and E7 oncogenes (13,27,39,57) and that they produce transcripts throughout the epithelium and the surface layers (12,15), it is not surprising that RNA assays detect more clinically significant infections than DNA testing, especially when samples come from the surface layers of the cervical epithelium, as in the present study.…”

“…The most common were mixed infections with HPV-16 and HPV-45 (6.1% of cases). Although previous studies have suggested that disease progression may depend on the overexpression of E6 and E7 RNA by a single dominant type (7,13), the precise role of these infections in carcinogenesis remains unclear.…”

HPV mRNA test in women with minor cervical lesions: Experience of the University Hospital of North Norway

“…The sensitivity for moderate dysplasia or worse is lower, but the specificity is higher (Cattani et al, 2009a;Keegan et al, 2009;Lie et al, 2005;Szarewski et al, 2008). It is well-known that many cervical lesions with moderate or severe dysplasia will regress spontaneously.…”

Nucleic Acid Sequence-Based Amplification Assay for Human Papillomavirus mRNA Detection and Typing: Evidence for DNA Amplification

“…Quite a few studies have assessed the clinical performance of commercial NASBA-based HPV RNA detection assays in comparison to HPV DNA tests and showed a higher specificity for the mRNA test (6,7,10,18,20,22,24,35,36). However, some of these studies (6, 10) do not constitute a direct comparison of HPV DNA and RNA for the detection of clinical end points but have been correlative; i.e., the samples were already selected due to their HPV DNA positivity so that the corresponding transcript could be sought.…”