Abstract
Human papillomaviruses (HPV) play a key role in promoting human anogenital cancers. Current high-risk HPV screening or diagnosis tests involve cytological or molecular techniques mostly based on qualitative HPV DNA detection. Here, we describe the development of a rapid quantitative polymerase chain reaction (qPCR) detection test of HPV16 and HPV18 oncogenes (E6 and E7) normalized on human gene encoding GAPDH. Optimized qPCR parameters were defined, and analytical specificities were validated. The limit of detection was 101 for all genes tested. Assay performances were evaluated on clinical samples (n = 96). Concordance between the Xpert HPV assay and the triplex assay developed here was 93.44% for HPV16 and 73.58% for HPV18. HPV co-infections were detected in 15 samples. The systems developed in the present study can be used in complement to traditional HPV tests for specifically validating the presence of HPV16 and/or HPV18. It can also be used for the follow-up of patients with confirmed infection and at risk of developing lesions, through the quantification of E6 and E7 oncogene expression (mRNA) normalized on the GAPDH expression levels.
Highlights
Human papillomaviruses (HPVs) are non-enveloped icosahedral particles with a size of 50–60 nm d iameter[1]
The progression to precancerous and cancerous lesion up to cervical intraepithelial neoplasia and eventually to cervical cancer occurs with a persistent HR-HPV infection and the integration of the HPV genome into the host chromosome mediated by host DNA replication and r ecombination[10], the loss or disruption of E2 protein and the subsequent upregulation of E6 and E7 oncogenes e xpressions[11]
Results showed that best relative fluorescence unit (RFU) means were obtained with 200 nM of probe (p ≤ 0.05) and that there were no significant differences between cycle threshold (Ct) values and primer concentrations independently of probe concentration (p > 0.05)
Summary
Human papillomaviruses (HPVs) are non-enveloped icosahedral particles with a size of 50–60 nm d iameter[1]. The progression to precancerous and cancerous lesion up to cervical intraepithelial neoplasia and eventually to cervical cancer occurs with a persistent HR-HPV infection and the integration of the HPV genome into the host chromosome mediated by host DNA replication and r ecombination[10], the loss or disruption of E2 protein and the subsequent upregulation of E6 and E7 oncogenes e xpressions[11]. These oncoproteins are involved in malignant transformation by altering cell cycle regulation and telomere maintenance via the telomerase activation and the degradation of the tumor suppressors p53 by E6 and retinoblastoma protein. LR-HPV oncogenes are less able to interfere with p53 and Rb functions than E6/E7 proteins from HR-types[12,15]
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
