Abstract
BackgroundPrimary testing for high-risk HPV (hrHPV) is increasingly implemented in cervical cancer screening programs. Many hrHPV-positive women, however, harbor clinically irrelevant infections, demanding additional disease markers to prevent over-referral and over-treatment. Most promising biomarkers reflect molecular events relevant to the disease process that can be measured objectively in small amounts of clinical material, such as miRNAs. We previously identified eight miRNAs with altered expression in cervical precancer and cancer due to either methylation-mediated silencing or chromosomal alterations. In this study, we evaluated the clinical value of these eight miRNAs on cervical scrapes to triage hrHPV-positive women in cervical screening.ResultsExpression levels of the eight candidate miRNAs in cervical tissue samples (n = 58) and hrHPV-positive cervical scrapes from a screening population (n = 187) and cancer patients (n = 38) were verified by quantitative RT-PCR. In tissue samples, all miRNAs were significantly differentially expressed (p < 0.05) between normal, high-grade precancerous lesions (CIN3), and/or cancer. Expression patterns detected in cervical tissue samples were reflected in cervical scrapes, with five miRNAs showing significantly differential expression between controls and women with CIN3 and cancer. Using logistic regression analysis, a miRNA classifier was built for optimal detection of CIN3 in hrHPV-positive cervical scrapes from the screening population and its performance was evaluated using leave-one-out cross-validation. This miRNA classifier consisted of miR-15b-5p and miR-375 and detected a major subset of CIN3 as well as all carcinomas at a specificity of 70%. The CIN3 detection rate was further improved by combining the two miRNAs with HPV16/18 genotyping. Interestingly, both miRNAs affected the viability of cervical cancer cells in vitro.ConclusionsThis study shows that miRNA expression analysis in cervical scrapes is feasible and enables the early detection of cervical cancer, thus underlining the potential of miRNA expression analysis for triage of hrHPV-positive women in cervical cancer screening.
Highlights
Primary testing for high-risk HPV is increasingly implemented in cervical cancer screening programs
Microarray-based differential miRNA expression in cervical tissue specimens can be verified by qRT-PCR To confirm our previously obtained microarray results, we used qRT-PCR to determine the expression levels of the eight genetically or epigenetically deregulated miRNAs in normal cervical squamous epithelium (n = 8), high-grade Cervical intraepithelial neoplasia (CIN) lesions (CIN2–3, n = 18), squamous cell carcinomas (SCC) (n = 22), and AC (n = 11) tissue specimens, of which 44 had been analyzed by microarray [17]
In conclusion, present data show that analysis of differentially expressed miRNAs may provide an alternative molecular triage strategy in high-risk HPV (hrHPV)-based cervical screening
Summary
Primary testing for high-risk HPV (hrHPV) is increasingly implemented in cervical cancer screening programs. As hrHPV testing detects women with clinically irrelevant transient infections, additional triage markers are required to identify women with high-grade CIN lesions who are in need of treatment given their risk of developing cancer. Should be objective (non-morphological), available in a high-throughput format and feasible in low-resource countries. For this purpose, HPV16/18 genotyping has previously been investigated as triage test in cervical cancer screening [7,8,9]. Additional molecular markers reflecting the underlying carcinogenic process are highly appealing alternatives These include DNA copy number aberrations and DNA methylation changes in the host cell genome that result in altered coding and non-coding gene expression [3]
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