1887

Abstract

Purpose. Human papillomavirus (HPV) E6/E7 mRNA is a more specific marker for cervical lesion screening than HPV DNA. Here, we aimed to develop a new one-step multiplex reverse transcript real-time PCR (MRT-PCR) to detect E6/E7 mRNA from 14 high-risk HPV (hrHPV) genotypes.

Methodology. The analytical sensitivity and specificity of the MRT-PCR system were validated. Its clinical performance was evaluated by comparing the results with bDNA signal amplification assay and histopathological results.

Results. The detection limit of MRT-PCR was 20 to 200 copies per reaction of different HPV genotypes, and no cross-reactivity was observed with any other low-risk HPV or other pathogens commonly found in the female genital tract. Using the bDNA signal amplification assay for comparison, a test on 166 clinical samples showed that the overall agreement between the two methods was 95.18 % and the one-step MRT-PCR was more sensitive. Further, compared with the histopathological results for the 166 clinical samples, the sensitivity and specificity of the MRT-PCR method were 88.9 and 71.6 %, respectively, and the positive rate for hrHPV E6/E7 mRNA increased with the severity of the cervical lesions.

Conclusion. The one-step multiplex RT-PCR for E6/E7 mRNA detection is a simple, fast, universally applicable, sensitive and highly specific method for hrHPV E6/E7 mRNA detection. It is reliable for cervical lesion screening and of potential value in future clinical applications.

Loading

Article metrics loading...

/content/journal/jmm/10.1099/jmm.0.000824
2018-08-20
2024-03-29
Loading full text...

Full text loading...

/deliver/fulltext/jmm/67/10/1509.html?itemId=/content/journal/jmm/10.1099/jmm.0.000824&mimeType=html&fmt=ahah

References

  1. Li B, He Y, Han X, Zhang S, Xu Y et al. Aberrant promoter methylation of SH3GL2 gene in vulvar squamous cell carcinoma correlates with clinicopathological characteristics and HPV infection status. Int J Clin Exp Pathol 2015; 8:15442–15447[PubMed]
    [Google Scholar]
  2. Bartholomew DA, Luff RD, Quigley NB, Curtis M, Olson MC. Analytical performance of Cervista HPV 16/18 genotyping test for cervical cytology samples. J Clin Virol 2011; 51:38–43 [View Article][PubMed]
    [Google Scholar]
  3. Gustinucci D, Giorgi Rossi P, Cesarini E, Broccolini M, Bulletti S et al. Use of cytology, E6/E7 mRNA, and p16INK4a-Ki-67 to define the management of human papillomavirus (HPV)-positive women in cervical cancer screening. Am J Clin Pathol 2016; 145:35–45 [View Article][PubMed]
    [Google Scholar]
  4. Muñoz N, Castellsagué X, de González AB, Gissmann L. Chapter 1: HPV in the etiology of human cancer. Vaccine 2006; 24:S1–S10 [View Article][PubMed]
    [Google Scholar]
  5. Cara MB, John OL. Histology of cervical intraepithelial neoplasia and the role of biomarkers. Best Pract Res Clin Obstet Gynaecology 2011; 25:605–615
    [Google Scholar]
  6. Hiller T. Comparative analysis of 19 genital human papillomavirus types with regard to p53 degradation, immortalization, phylogeny, and epidemiologic risk classification. Cancer Epidemoil Biomarkers 2006; 15:1262–1267 [View Article]
    [Google Scholar]
  7. McLaughlin-Drubin ME, Münger K. The human papillomavirus E7 oncoprotein. Virology 2009; 384:335–344 [View Article][PubMed]
    [Google Scholar]
  8. Dong WL, Caldeira S, Sehr P, Pawlita M, Tommasino M. Determination of the binding affinity of different human papillomavirus E7 proteins for the tumour suppressor pRb by a plate-binding assay. J Virol Methods 2001; 98:91–98 [View Article][PubMed]
    [Google Scholar]
  9. Castle PE, Dockter J, Giachetti C, Garcia FA, McCormick MK et al. A cross-sectional study of a prototype carcinogenic human papillomavirus E6/E7 messenger RNA assay for detection of cervical precancer and cancer. Clin Cancer Res 2007; 13:2599–2605 [View Article][PubMed]
    [Google Scholar]
  10. Cuschieri K, Wentzensen N. Human papillomavirus mRNA and p16 detection as biomarkers for the improved diagnosis of cervical neoplasia. Cancer Epidemiol Biomarkers Prev 2008; 17:2536–2545 [View Article][PubMed]
    [Google Scholar]
  11. Evans MF, Peng Z, Clark KM, Adamson CS, Ma XJ et al. HPV E6/E7 RNA in situ hybridization signal patterns as biomarkers of three-tier cervical intraepithelial neoplasia grade. PLoS One 2014; 9:e91142 [View Article][PubMed]
    [Google Scholar]
  12. Ho CM, Lee BH, Chang SF, Chien TY, Huang SH et al. Type-specific human papillomavirus oncogene messenger RNA levels correlate with the severity of cervical neoplasia. Int J Cancer 2010; 127:622–632 [View Article][PubMed]
    [Google Scholar]
  13. Fontecha N, Basaras M, Hernáez S, Andía D, Cisterna R. Assessment of human papillomavirus E6/E7 oncogene expression as cervical disease biomarker. BMC Cancer 2016; 16:852 [View Article][PubMed]
    [Google Scholar]
  14. Jie H, Ting Y, Jin-Yin Z, Li-Ping G, Shang-Xian G et al. The establishment of E6/E7 quality products from high-risk human papillomavirus. Chin J Pharm Anal 2015; 35:595–599
    [Google Scholar]
  15. Shen Y, Gong J, He Y, Cheng G, Okunieff P et al. Quantivirus® HPV E6/E7 RNA 3.0 assay (bDNA) is as sensitive, but less specific than hybrid capture 2 test. J Virol Methods 2013; 187:288–293 [View Article][PubMed]
    [Google Scholar]
  16. Walboomers JM, Jacobs MV, Manos MM, Bosch FX, Kummer JA et al. Human papillomavirus is a necessary cause of invasive cervical cancer worldwide. J Pathol 1999; 189:12–19 [View Article][PubMed]
    [Google Scholar]
  17. Sherman ME, Lorincz AT, Scott DR, Wacholder S, Castle PE et al. Baseline cytology, human papillomavirus testing, and risk for cervical neoplasia: a 10-year cohort analysis. J Natl Cancer Inst 2003; 95:46–52 [View Article][PubMed]
    [Google Scholar]
  18. Boulet GA, Horvath CA, Berghmans S, Bogers J. Human papillomavirus in cervical cancer screening: important role as biomarker. Cancer Epidemiol Biomarkers Prev 2008; 17:810–817 [View Article][PubMed]
    [Google Scholar]
  19. Bansal N, Wright JD, Cohen CJ, Herzog TJ. Natural history of established low grade cervical intraepithelial (CIN 1) lesions. Anticancer Res 2008; 28:1763–1766[PubMed]
    [Google Scholar]
  20. Yim EK, Park JS. The role of HPV E6 and E7 oncoproteins in HPV-associated cervical carcinogenesis. Cancer Res Treat 2005; 37:319–324 [View Article][PubMed]
    [Google Scholar]
  21. Molden T, Kraus I, Skomedal H, Nordstrøm T, Karlsen F. PreTect HPV-Proofer: real-time detection and typing of E6/E7 mRNA from carcinogenic human papillomaviruses. J Virol Methods 2007; 142:204–212 [View Article][PubMed]
    [Google Scholar]
  22. Wang J, Tian Q, Zhang S, Lyu L, Dong J et al. Clinical significance of HPV L1 capsid protein detection in cervical exfoliated cells in high-risk HPV positive women. Zhonghua Fu Chan Ke Za Zhi 2015; 50:253–257[PubMed]
    [Google Scholar]
  23. Lindh M, Görander S, Andersson E, Horal P, Mattsby-Balzer I et al. Real-time Taqman PCR targeting 14 human papilloma virus types. J Clin Virol 2007; 40:321–324 [View Article][PubMed]
    [Google Scholar]
  24. Okodo M, Okayama K, Teruya K, Sasagawa T. Uniplex E6/E7 PCR method detecting E6 or E7 genes in 39 human papillomavirus types. J Med Virol 2018; 90:981–988 [View Article][PubMed]
    [Google Scholar]
  25. Cattani P, Siddu A, D'Onghia S, Marchetti S, Santangelo R et al. RNA (E6 and E7) assays versus DNA (E6 and E7) assays for risk evaluation for women infected with human papillomavirus. J Clin Microbiol 2009; 47:2136–2141 [View Article][PubMed]
    [Google Scholar]
  26. Wang HY, Park S, Lee D, Kim S, Kim G et al. Prevalence of type-specific oncogenic human papillomavirus infection assessed by HPV E6/E7 mRNA among women with high-grade cervical lesions. Int J Infect Dis 2015; 37:135–142 [View Article][PubMed]
    [Google Scholar]
  27. Wang HY, Lee D, Park S, Kim G, Kim S et al. Diagnostic performance of HPV E6/E7 mRNA and HPV DNA assays for the detection and screening of oncogenic human papillomavirus infection among woman with cervical lesions in China. Asian Pac J Cancer Prev 2015; 16:7633–7640 [View Article][PubMed]
    [Google Scholar]
  28. Shi WJ, Liu H, Wu D, Tang ZH, Shen YC et al. E6/E7 proteins are potential markers for the screening and diagnosis of cervical pre-cancerous lesions and cervical cancer in a Chinese population. Oncol Lett 2017; 14:6251–6258 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/jmm.0.000824
Loading
/content/journal/jmm/10.1099/jmm.0.000824
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error