Rev4.hpv dna testing for s2011

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Rev4.hpv dna testing for s2011

  1. 1. HPV DNA Testing “Molecular Smear” Abdulmohsen Alkushi, MD, Msc, FRCPC Consultant Pathologist, National Guard Hospital, KAMC-Riyadh The Role of Pathologists in 21st Century
  2. 2. HPV DNA Testing n  Introduction n  Techniques n  Potential value
  3. 3. Cervical Cancer n  Cervical cancer prevention programs have relied on cytological testing using the Papanicolaou (Pap) smear test.
  4. 4. Cervical Cancer n  Research worldwide has clearly shown that virtually all cervical cancer is caused by human papillomavirus (HPV) infection.
  5. 5. HPV & Cervical Cancer n  The direct detection of HPV in cervical specimens may offer an alternative or complement to population-based cytological screening. n  Recent studies have demonstrated that HPV test results are more sensitive (although they are less specific) than Pap smears in detecting high-grade dysplasia in older women. • Wright TC Jr, et al. Journal of the American Medical Association 283:81–86 (2000). • Schiffman M, et al. Journal of the American Medical Association 283:87–93 (2000).
  6. 6. HPV Detection n  HPV cannot be cultured reliably in a laboratory setting. n  Molecular techniques: n  Non-amplification technologies n  Amplification technologies
  7. 7. HPV DNA Detection n  Amplification techniques can be further divided into n  (1) target amplification n  (2) signal amplification n  (3) probe amplification (for example, ligase chain reaction). n  To date, target and signal amplification techniques, in addition to non-amplified techniques, have been applied to the detection of HPV.
  8. 8. HPV DNA Testing n  Because there are many HPV types with differing oncogenic potential, diagnostic tests must not only detect HPV DNA, they also must determine the type(s) present in each specimen.
  9. 9. Signal-amplified techniques n  Hybrid Capture Technology: n  developed by the Digene Corporation, detects nucleic acid targets directly, using signal amplification n  Digene has developed two products for the detection of HPV: the first-generation Hybrid Capture Tube (HCT) test and the more recent Hybrid Capture II (hc2) assay n  Both assays detect high-risk HPV types.
  10. 10. Hybrid Capture Technology n  HCT test detects high-risk types (16, 18, 31, 33, 35, 45, 51, 52, and 56). n  Four additional viral types were added to the high-risk category in the hc2 test: 39, 58, 59, and 68. n  The level of detection of hc2 is rated at 5,000 viral copies per sample (one picogram of HPV DNA per sample) in contrast to HCT, which detects 10 picograms).
  11. 11. Hybrid Capture Technology n  To perform the HC assay cervical or vaginal clinical specimens collected through self-sampling or obtained by a health care provider during a pelvic examination are combined with an extraction buffer. n  Once samples are taken, they can be stored at room temperature for two weeks, at 4°C for one additional week, and at -20°C for up to three months. n  hc2 test involves a five-part process takes an estimated 6 to 7 hours,
  12. 12. Hybrid Capture Technology
  13. 13. Hybrid Capture Technology
  14. 14. US FDA Approval n  Hybrid Capture Tube (HCT) was granted US FDA approval in May 1995. n  In March 1999, the US FDA approved Digene s second-generation HPV detection kit (hc2). n  In March 2003, the FDA approved hc2 test to be used simultaneously with the Pap test to screen for cervical cancer in women age 30 and older.
  15. 15. US FDA Approval n  The Digene® HPV Test holds two US FDA approved indications: n  Primary adjunctive screening with a Pap test for women age 30 and older to detect the presence or absence of high–risk HPV for use as a guide for patient management in cervical cancer screening programs. n  Reflex HPV testing: Triage for women with (ASC–US) Pap results to determine appropriate follow–up.
  16. 16. HPV & Cervical Cancer Based on a cohort analysis of 5,671 women age >30 years within a study that included 7,932 women ages 15–76 (median age of 34). Clavel C, Masure M, Bory J–P, et al. Brit J Cancer, 2001; 89 (12): 1616–1623.
  17. 17. Sensitivity of HPV + Pap test 98% Sensitivity of HPV test 94% Sensitivity of Pap test 75%
  18. 18. HPV & Cervical Cancer n  The prevalence of HPV in women generally decreases over time, while the incidence of cervical cancer increases. n  The point at which HPV prevalence meets cervical cancer incidence is around the age of 30 n  This supports the rationale for primary adjunctive screening (HPV + Pap) in women 30 and older.
  19. 19. Target-amplified techniques n  DNA target amplification is a laboratory-based procedure that duplicates DNA fragments from a target sequence of a gene. n  PCR is the most commonly used by this method n  GenoID® Reveal HPV® Real-Time HPV Detection Kit n  PapilloCheck® HPV-Screening
  20. 20. Non-amplified techniques n  Southern blot n  Dot blot hybridizations
  21. 21. Non-amplified techniques n In situ hybridization
  22. 22. Non-amplified techniques n  Several factors limit the incorporation of non-amplified techniques into large- scale screening programs: n  Relatively low sensitivity n  Time-consuming n  Require trained technicians n  The demand for an array of laboratory reagents and equipment required
  23. 23. DNA test may be more effective than Pap smear for detecting cervical cancer, studies suggest. The CBS Evening News (10/17, Couric) reported, "Doctors are telling us tonight [that] a new test could replace the Pap smear for detecting cervical cancer."
  24. 24. Potential for HPV DNA Testing
  25. 25. Potential Protocols for HPV DNA Testing n  As a means of triage for women whose Pap test indicate ASC-US n  As a means of surveillance of women treated for HSIL or microinvasive cancer. n  As a primary screen for HSIL in older women (women age 35 or older)
  26. 26. Result Obtained on Cytology & HPV DNA Testing Wright, et al. HPV Testing as Adjunct to Cytology. Obstet Gynecol Feb 2004,103:2 This algorithm represents interim guidance developed by members of ASCCP, NIH/ NCI, and ACS
  27. 27. Future of Gyne Cytology Modern Uterine Cytopathology Moving to Molecular Smear
  28. 28. Leading the News Specific HPV test may prove useful in countries with few resources, investigators say. Bloomberg News (4/2, Chase) reports that, "for the first time," a test manufactured by Dutch drugmaker Qiagen called careHPV may become an essential and influential component of "a real strategy for providing" human papilloma virus (HPV) "screening to women globally."
  29. 29. Thank you for your attention!

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