Detecting HPV means better understanding of the risk of cervical cancer was the major focus of Dr. Dinesh Gupta. He spoke on “Opportunistic Screening for Cervical Precancer Lesions” and informed why the combination screening is vital for prevention and detection of cervical cancer. According to Dr. Gupta, combined screening with liquid based cytology and hybrid capture2 HPV DNA test would identify who’s at risk for high-grade disease and cancer and reduce missed disease caused by false-negative Pap Smear. HPV DNA test is the only FDA approved test to detect 13 high risk HPVs associated with virtually all cervical cancer, he added.

1. Screening & Early Detection of Cervical Precancers: A Clinical Role for HPV DNA Testing Dinesh Gupta, PhD Director, NCRL, India Executive Member, IACR Life Member, AOGIN “ We are what we repeatedly do. EXCELLENCE, then, is a HABIT.”

4. HPV-mediated progression to cervical cancer

5. NATURAL HISTORY OF SQUAMOUS INTRAEPITHELIAL LESIONS Oscar, 1992 > 12% 5% 1% PROGRESS Ca Cx 56% 35% 32% PERSIST - 22% 11% PROGRESS CIN 3 32% CIN 3 (SEVEVER DYSPIASIA) 43% CIN 2 (MODERATE DYSPIASIA) 57% CIN 1 (MILD DYSPIASIA) REGRESS Clinical LESION

8. Squamous cell maturation

9. Changes in squamous epithelium Normal Cytology HSIL

13. Gynaecological Cytopathology advanced… Liquid Based Cytology (LBC) ThinPrep 2000 Processor 1 st to US FDA Approval LiquiSmear Processor

16. Based on a cohort analysis of 5,671 women above age 30 within a study that included 7932 women ages 15 to 76 (median 34) RE: C Clavel, 2003 LBC: Sensitivity improved … Monolayer cells on 19mm dia circle

17. Key Elements/ Features: A- Superficial Sq Cells B- Intermediate Cells C- Parabasal Cells D- Metaplastic (Basal) cells

18. Colposcopy Technique

21. The Transformation Zone

23. The HPV TEST Because of the strong association (>99.7%) between HPV and cervical cancer, this test helps DETERMINE THE NEXT STEP for a woman with lower genital abnormality among symptomatic, or identifies latent disease among healthy. This test checks for HPV genetic material, or DNA, within the cervical cells. A swab of cervical cells is taken and the specimen is sent to a laboratory and examined for HPV . If the HPV test is positive for a high-risk strains, the abnormalities are more likely precancerous. If the test is negative, the cervical changes are more likely benign or regress or maintain.

26. Specimen Collection Using the DNA Collection Device 1 2 3 4

27. digene Hybrid Capture 2 ® Denature Physiologic specimen Hybridize with full-length RNA probe Capture and detect hybrids Sensitive Luminescence

34. Viral load cut-off by HC2 is clearly indicative of higher disease stage (likely to be not missed at Colposcopy) J Pathol 2003; 201 : 1–6. 5 August 2003 Very low levels of viral DNA may not be clinically meaningful! Clinical sensitivity of hc2 hpv Viral load threshold for CIN development Viral load threshold for no clinical infection

35. Adapted from Snijders et al. Journal of Pathology 2003; 201:1-6 Analytical versus Clinical Sensitivity The Analytical Sensitivity of PCR methods can be <10 copies of HPV DNA The Analytical Sensitivity of HC2 is 5,000 copies of HPV DNA

36. “ HC2 was more sensitive…than PCR for detecting 2-year cumulative CIN3 or cancer”* * Schiffman et al, Am J Clin Path 2005; 124: 722-732 Lorincz, HPV Testing by Hybrid Capture. Monsonego J(ed): Emerging Issues of HPV Infections: From Science to Practice. Basel, Karger, 2006: 54-62 Data from 16 published studies showed a median Clinical Sensitivity of 82% for PCR Analytical versus Clinical Sensitivity

37. HPV Genome: locations of general primer sets AMPLIMER SIZE Claim: Utilises sequence variation in the 22-bp interprimer region permitting identification of HPV genotypes. (also claim it does not permit formal classification of HPV genotypes. Ref: Bernhardt Kleter et al, J Clin Microbiol Aug. 1999, 2508-17 ) 65 bp E6 E7 L1 L2 E4 E5 E2 E1 1 7900 4000 5000 6000 7000 1000 2000 3000 MY 9/11 GP 5+/6+ SPF 1/2 150 bp 450 bp PRIMER SETS

38. Clinical Efficacy of Screening Options for Cervical Precursors Generally accepted detection range of various diagnostic tests normal cervix HPV infection HPV persistence high grade neoplasia invasion low grade neoplasia Pap Smear (cytology) HC2 HPV DNA Testing VIA/ VILI Histopathology Expert Colposcopy

39. Emerging paradigm: Ca Cx Screening

40. “ There is sufficient evidence that testing for human papillomavirus infection as the primary screening modality can reduce cervical cancer incidence and mortality rates.” – IARC/WHO, 2005 IARC, WHO. IARC Handbooks of Cancer Prevention: Cervical Cancer Screening. Volume 10. IARC Press; 2005.

45. Does Pretreatment Human Papillomavirus (HPV) Titers Predict Radiation Response and Survival Outcomes in Cancer Cervix? – A pilot study. Gynaecological Oncology: 2006 Niloy R. Datta, MD, DNB  , Piyush Kumar, MD  , Shalini Singh, MD, DNB  , Dinesh Gupta, PhD # , Anurita Srivastava, MD * , Tapan N. Dhole, MD, Departments of Radiotherapy  and Microbiology+, Regional Cancer Centre, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Rae Barelli Road, Lucknow, India and Gentech Laboratories, New Delhi HPV in Ca Cx Treatment

53. Lynnette Denny AORTIC May 2006 “ secondary prevention (cytology, colposcopy, biopsy, histology), has not been successful in any developing countries in the world due to the complexity of the infrastructure required to make this approach work and the substantial human, financial and logistical resources required. Consequently cervical cancer prevention efforts in developing countries either do not exist or they are extremely limited.

55. “ Dhanyawad ” Thank U! Delhi Moms are going HPV-way as much as make overs... India Times for a big change Testing for HPV DNA once in a life time reduces the risk of CERVICAL CANCER almost 100%

56. What is the incidence of High-Grade CIN in women with abnormal smears? 90% Refer to HPV or Colpo and directed Biopsy CIN 3 (Severe dysplasia) 70% hc2 HPV +ve Refer to Colpo CIN 1/ CIN 2 (Moderate dysplasia) 30% (ASC-H) hc2 HPV / Repeat smear 6 mo/ refer to Colpo when either is positive ASC-US (Mild Dysplasia) Incidence of HG CIN (CIN2/3) Action?

58. Table 2. Staging System for Cervical Cancer