Abstract
BackgroundHPV oncogenes mRNA detection gains momentum as an adjuvant for HPV-related cervical abnormalities diagnosis, but is based on costly detection assays not allowing viral type targeting.ObjectiveTo assess detection rate of HPV oncogenes E6/E7 mRNA from cervical specimens using a manual, open technology, fully customizable protocol and determine whether HPV-related epidemiological features influence mRNA retrieval. We reviewed literature and compared our retrieval rate with automated technologies.MethodsWe used 60 samples positive for HPV DNA types 16, 18, 31 and/or 45. We extracted mRNA with a TRizol-based protocol, and tested mRNA purity and concentration using light absorbance. We reverse-transcribed mRNA into cDNA for E6/7 detection.ResultsHPV oncogenes E6/E7 mRNA was retrieved from 36 (60%) out of 60 specimens. No HPV load-related clinical or epidemiological feature was significantly associated with mRNA retrieval. Presence of HPV-DNA 16/18 was associated with mRNA retrieval (OR = 9.08; 95% CI 1.26 to 65.32 for HPV 16; and 18.2; IC95% 1.86 to 391.44 for HPV 18).ConclusionsThe open-technology protocol yielded an mRNA detection rate similar to that of automated technologies. Advantages are lower costs and target HPV type customization.
Highlights
In the last two decades, the detection of Human papillomavirus (HPV) DNA in cervical samples has proven to be a good diagnostic and risk predictor tool for cervical intraepithelial neoplasia (CIN) and cervical cancer (Castle et al 2011)
This may be due to the fact that the oncogenic potential of the high-risk HPV types relies on the actions of oncoproteins coded by E6/E7, which bind to and modulate different gene products, in particular tumor suppressors p53 and pRb (Halfon et al 2010; Kraus et al 2006)
We retrieved oncogenes E6 and E7 mRNA from 36 (60%) of the 60 specimens previously found to be positive for the relevant HPV genotypes
Summary
In the last two decades, the detection of Human papillomavirus (HPV) DNA in cervical samples has proven to be a good diagnostic and risk predictor tool for cervical intraepithelial neoplasia (CIN) and cervical cancer (Castle et al 2011). Some studies suggest that HPV oncogenes E6 and E7 mRNA levels in the uterine cervix may be more specific early indicators of predisposition to carcinogenesis than DNA levels (Benevolo et al 2011; Ratnam et al 2011; Winer et al 2009) This may be due to the fact that the oncogenic potential of the high-risk HPV types (hr-HPV) relies on the actions of oncoproteins coded by E6/E7, which bind to and modulate different gene products, in particular tumor suppressors p53 and pRb (Halfon et al 2010; Kraus et al 2006). HPV oncogenes mRNA detection gains momentum as an adjuvant for HPV-related cervical abnormalities diagnosis, but is based on costly detection assays not allowing viral type targeting
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