Understanding HPV testing’s
  place in primary screening

January 2007
PAP/NGC Programs Review

Patricia G. Wasserman, MD
Mary R. Schwartz, MD
Teresa M. Darragh, MD

Cervical cancer is the second most common cancer in women worldwide and is the most common female cancer in developing countries. In 2000, more than 450,000 women around the world developed cervical cancer, and more than 200,000 died from the disease.³⁴ These figures are less gloomy for American women: 10,370 cases of cervical carcinoma were diagnosed in 2005 with about 3,710 related deaths.⁸ Countries with well established cervical cancer screening programs have been extremely successful in lowering the incidence of and mortality from cervical cancer, thanks to the enormous success of the Pap test. However, this test is imperfect and prone to a fairly high false-negative rate. Yet finally, for the first time in the year 2006, the American Cancer Society estimated that fewer than 10,000 women would be diagnosed with cervical cancer in the United States.

Why are there cervical cancer screening failures in the U.S.?

• While more than 50 percent of cervical cancer patients were never screened, and 10 percent to 20 percent of patients were not screened in at least the previous five years, there still are 30 percent false-negative screening events (that is, the patient has disease but it is not detected).^1–3
• Of those false-negatives, 50 percent are due to sampling errors, and the remainder are due to screening or interpretive errors.⁵
• Any single Pap test event is only 50 percent sensitive for disease prevalence.⁴
• Recurring independent Pap test events make a screening program successful. Nevertheless, even with perfect compliance, this system is flawed because of less than ideal sensitivity, interpretive variance, and morphological subjectivity.^6,7

How do we reduce false-negatives and increase accuracy?

There are at least four major lines of attack¹: enriching the sample, increasing the detection of abnormal cells, refining the accuracy of the morphologic interpretation, and eliminating morphology from the entire equation.

The first two approaches are addressed by the introduction of thin-layer cytology, which has increased the sensitivity and specificity of the Pap test. New automation systems are designed to help locate abnormal cells, lessening fatigue and lowering false-negative rates due to screening errors.⁹ Quality improvement programs and continuing medical education courses improve the morphologic acumen of cytotechnologists and cytopathologists. All of these strategies have had a minimal impact on the rate of false-negative cytology. So what about removing morphology entirely from our “hands”? Removal of the reliable, but imperfect, morphologic analysis should be addressed by a test that does not require the subjectivity of the cytological process but would improve the performance of cervical cancer screening. HPV testing is the best contender for that role at this point in time.

What do we know so far about human papillomavirus?

• HPV has been shown to be the causative agent of about 95 percent of all cervical carcinomas worldwide.^{10, 11, 24}
• The vast majority of HPV infections with both high-risk and low-risk types are benign in nature and will regress spontaneously with no serious sequelae.²⁹
• Cervical cancer is an unusual complication of a persistent cervical infection with a high-risk HPV type.
• Women younger than age 30 have a high rate of infection with high-risk HPV (15–46 percent), and most infections will be transient.²⁹
• The clearance time reported for transient HPV infection is about six to 18 months.
• A significant drop in HPV prevalence is noted in patients after the third decade of life (10 percent or less).^{9, 12}
• High-risk HPV types are HPV 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, and 68. Since these types are known to be oncogenic, they are the only clinically relevant types that should be tested for. The only FDA-approved test is Hybrid Capture 2 (HC2, Digene, Gaithersburg, Md.).^{17, 20, 21}
• Persistence of high-risk HPV infection is essential for the development, continuation, and progression to CIN 3.

Is HPV testing a useful tool to identify patients at risk for developing cervical neoplasia?^13–17

• Several studies have demonstrated that HPV testing is more sensitive and more reproducible and has a better predictive and prognostic value than cytology alone.
• The addition of HPV testing to cytology screening has achieved sensitivities of over 95 percent with specificity analogous to or only slightly less than cytology alone (90 percent).
• These studies have led the way to revised practice guidelines with the addition of HPV testing as an adjunct to the Pap test in the primary screening of women age 30 and older.
• This strategy may reduce the false-negative rate to nearly zero, allowing for an increase in the screening interval to up to three years for women who are negative on both Pap and high-risk HPV tests.

What are the current uses of HPV testing?¹⁸

• HPV testing is recommended as a reflex test in women of any age for the triage of those with equivocal cytological results (ASC-US).
• HPV testing is recommended as an adjunct to cervical cytology in the screening of women more than 30 years of age.
• HPV testing may be used in the post-colposcopy followup of women with HPV-positive ASC-US, ASC-H, or LSIL, without histological confirmation of CIN 2/3.
• HPV testing is recommended for the initial followup of treated CIN 2/3, replacing colposcopy or cytology or both.
• HPV testing is not recommended as a screening tool in conjunction with the Pap in women younger than 30 years of age.
• Negative Pap and HPV tests in women older than 30 years allow for a longer interval between cervical screenings (three-year interval).

What is the management of women who are screened with a combination of cytology and HPV testing?¹⁹

• Normal Pap test and negative HPV-DNA (double negative): Negative results on combined screening have a high negative predictive value (99.1–100 percent) for cervical cancer. A repeat Pap and HPV is recommended within three years. The risk of unidentified CIN 2/3 is approximately one in 1,000.
• Normal Pap test and positive HPV-DNA: These cases constitute three percent to five percent of the population screened, and have a 20-fold increased chance of developing CIN 2/3 than women with negative HPV-DNA tests. It is recommended to re-test with a combination of Pap and HPV in six to 12 months. Approximately 40 to 60 percent of these patients will clear their HPV infection over this short followup. The risk for CIN 2/3 is 2.8 to 4.2 percent.
• ASC-US Pap and negative HPV-DNA: The recommendation is for retesting with both Pap and HPV combination in 12 months. The risk for CIN 2/3 is six percent to 12 percent, recognizing that even HPV testing is imperfect.
• ASC-US Pap and positive HPV-DNA: The risk for CIN 2/3 is 15 to 20 percent within a two-year period, justifying the recommendation of colposcopic evaluation.
• ASC-H and LSIL cases have a high frequency of high-risk HPV-DNA detection, thus diminishing the value of HPV testing as a colposcopic triage test.

What are the major concerns of adding HPV testing as a tool for cervical cancer screening in clinical practice?

Laboratory-related issues^{23, 25, 26}

• HPV testing should be clinically validated to ensure that results are objective and highly reproducible.
• It is potentially clinically deleterious to attain the highest analytical sensitivity for the detection of HR-HPV molecules. HPV presence does not equate to clinical evidence of disease. Therefore, an issue vital to the validation question is the need for an equilibrium between analytical sensitivity and clinical sensitivity. Maximal sensitivity has the potential of detecting clinically insignificant infection and thus increasing the false-positive rate. Hence, the current FDA-approved HPV Hybrid Capture 2 has been calibrated to balance sensitivity and specificity toward the detection of a significant copy number of HPV DNA molecules, which is more likely to correlate with a clinically significant disease state.

Clinical issues²²

• The need for counseling and educating patients about the significance of HPV as a sexually transmitted infection, and the significance of its presence in the setting of a negative cytology.
• The change in practice due to the lengthening of screening intervals to every three years.
• The potential for drastic increases in cervical cancer screening costs.
• The risk of expanding superfluous interventions and treatment, thus diminishing the quality of life.

Public health issue^{27, 28}

• The high prevalence of HPV infection will have the effect of large numbers of women in need of advice and support. This requires a well-organized, accessible, and massive public health education for women and health care providers.
• Cost-effectiveness analyses models indicate promising outcome by providing increased protection against cervical cancer while reducing the average woman lifetime costs associated with screening, opening the horizon for a significant positive impact worldwide.
• By improving the clinical performance of screening and enhancing the efficiency and cost-effectiveness of the test, it will be possible to direct assets toward efforts to lessen disparities and increase screening coverage.

What is the future of cervical cancer screening?^{30–33, 35–38}

• HPV screening as the primary screening modality could be implemented in countries with high-quality cytology labs, with cytology reserved for triage of HPV-positive cases. This approach has the following benefits: (1) HPV assays are automated, objective, and highly sensitive, allowing for improved quality control and reducing false-negative rates; (2) cytology may be used for HPV-positive women as a reflex test; (3) the longer screening intervals proposed may prove to be safer due to fewer false-positives; and (4) primary screening with HPV-DNA testing needs to be evaluated in large trials to understand and predict its full impact on cancer incidence and mortality.
• Screen-and-treat method: HPV as a standalone test (self-sampled or physician-collected) may be used in developing countries where cytology-based screening programs are not available. A positive result would be followed by immediate treatment, maximizing the use of sparse health care resources and patient compliance. Field testing of this process will begin this year in China and India.

There are two issues related to this screening method: (1) Large-scale trials must concentrate on the optimal time to initiate screening, intervals between visits, and management of positive HPV results; and (2) if the premise of this program is that all HPV-positive women should be treated, the risk of overtreatment with its serious and potentially permanent adverse effects must be examined closely.

• Increase HPV specificity: The need to increase the lower specificity of HPV testing vis-à-vis cytology screening may be tackled by several methods: (1) HPV typing, especially for HPV types 16 and 18, may increase the correlation of a positive test with true disease; (2) viral loads may separate clinically relevant infections from cases unlikely to be linked with CIN 2/3; (3) surrogate markers, such as proliferative markers (p16 and mRNA) or cell cycle markers (cdc6 and mcm5 proteins), may help distinguish those infections that have the potential to progress toward cancer; and (4) host-cell biomarkers may be detected by comparative genomic hybridization, or CGH, microarray analyses, possibly aiding in a superior risk stratification of HPV-positive women.

What is the current state of cervical cancer screening?

• Women now have improved cervical cancer screening.
• There are clearer and more meaningful cytological interpretations because of standardization by the 2001 Bethesda nomenclature.
• There are coherent and easier evidence-based management algorithms.
• There are powerful molecular diagnostics, such as HPV testing, that improve outcomes and potentially reduce costs.
• A newly approved HPV vaccine may prevent 98 percent of persistent infections with high-risk HPV types 16 and 18, potentially reducing cervical cancer by up to 51 percent even in the absence of cytological screening.

What is the path of success?

• Patients must come in for the test (most important reason for “missed” diagnoses).
• Clinicians must take adequate, representative, well-prepared samples.
• Cytologists must correctly identify dysplastic lesions.
• Results must reach the clinicians.
• Clinicians must act appropriately on results based on published evidence-based guidelines.
• Patients must return for therapy or appropriate clinical followup.

Overall, our approach to cervical cancer screening has been a major success. In the last decade or so, advances in the understanding of the role of HPV in the pathogenesis of cervical cancer have started to revolutionize this approach. Primary screening with molecular diagnostic tests for HPV and other potentially specific markers are on the horizon, and they promise to provide even better prevention of cervical cancer to women worldwide.

References

1. Stoler MH. Cervical cancer screening in the HPV era: what is the standard of care? Program and abstracts of Pathology Today: American Society for Clinical Pathology 2005 Annual Meeting; Oct. 8–11; Seattle. Session SP 20.
2. Kinney W, Sung HY, Kearney KA, Miller M, Sawaya G, Hiatt RA. Missed opportunities for cervical cancer screening of HMO members developing invasive cervical cancer. Gynecol Oncol. 1998;71:428–430.
3. Kinney WK, Manos MM, Hurley LB, Ransley JE. Where is the high grade cervical neoplasia? The importance of minimally abnormal Papanicolau diagnoses. Obstet Gynecol. 1998;91:973–976.
4. Nanda K, McCrory DC, Myers ER. Accuracy of the Papanicolau test in screening for and follow-up of cervical cytological abnormalities: a systematic review. Ann Intern Med. 2000;132:810–819.
5. Koss LG. The Papanicolau test for cervical cancer detection. A triumph and a tragedy. JAMA 1989;261:737–743.
6. Miller MG, Sung HY, Sawaya GF, Kearney KA, Kinney W, Hiatt RA. Screening interval and risk of invasive squamous cell cancer. Obstet Gynecol. 2003;101:29–37.
7. Sawaya GF, McConnell KJ, Kulasingam SL, et al. Risk of cervical cancer associated with extending the interval between cervical cancer screenings. N Engl J Med. 2003;349:1501–1509.
8. American Cancer Society. Estimated new cancer cases and deaths by sex for all sites, U.S., 2005 in Cancer Facts and Figures. www.cancer.org.
9. Stoler MH. Advances in cervical screening technology. Mod Pathol. 2000;13:275–284.
10. Jacobs MV, Walboomers JM, Snijders PJ, et al. Distribution of 37 mucosotropic HPV types in women with cytologically normal cervical smears: the age-related patterns for high-risk and low-risk types. Int J Cancer. 2000;87:221–227.
11. Bosch FX, Manos MM, Munoz N, et al. Prevalence of HPV in cervical cancer: a worldwide perspective. International Biological Study on Cervical Cancer (IBSCC) Study Group. J Natl Cancer Inst. 1995;87:796–802.
12. Sellors JW, Karwalajtys TL, Kaczorowski JA, et al. Survey of HPV in older Ontario women (SHOOW) group. Prevalence of infection with carcinogenic human papillomavirus among older women. CMAJ. 2002;167:871–873.
13. Lorincz AT, Richart RM. HPV-DNA testing as an adjunct to cytology in cervical screening programs. Arch Pathol Lab Med. 2003;127:959–968.
14. Kulasingam SL, Hughes JP, Kiviat NB, et al. Evaluation of HPV testing in primary screening for cervical abnormalities: comparison of sensitivity, specificity, and frequency of referral. JAMA. 2002;288:1749–1757.
15. Smith RA, Cokkinides V, Eyre HJ. American Cancer Society guidelines for the early detection of cancer. CA Cancer J Clin. 2003;53:27–43.
16. ACOG Practice Bulletin: clinical management guidelines for OB-GYNs. No. 45, August 2003. Cervical cytology screening. Obstet Gynecol. 2003;102:417–427.
17. ACOG Practice Bulletin: clinical management guidelines for OB-GYNs. No. 61, April 2005. Human papillomavirus. Obstet Gynecol. 2005;105:905–918.
18. Wright TC, Schiffman M, Solomon D, et al. Interim guidance for the use of HPV-DNA testing as an adjunct to cervical cytology for screening. Obstet Gynecol. 2004;103:304–309.
19. Cox JT. HPV testing in primary cervical screening and abnormal Papanicolau management. Obstet Gynecol Surv. 2006; 61 (6), s15–s25.
20. Wright TC, Cox JT, Massad LS, et al. 2001 consensus guidelines for the management of women with cervical cytological abnormalities. JAMA. 2002; 287:2120–2129.
21. Wright TC, Cox JT, Massad LS, et al. ASCCP 2001 consensus guidelines for the management of women with cervical intraepithelial neoplasia. Am J Obstet Gynecol. 2003;189:295–304.
22. Anhang R, Goodman A, Goldie SJ. HPV communication: review of existing research and recommendations for patient education. CA Cancer J Clin. 2004;54:248–259.
23. Titus K. Making a valid point about HPV tests. CAP TODAY. 2005; 19 (9): 1.
24. Munoz N, et al. Epidemiologic classification of HPV types associated with cervical cancer. N Engl J Med. 2003;348:518–527.
25. Schiffman M, et al. A comparison of a prototype PCR assay and Hybrid Capture 2 for detection of carcinogenic HPV-DNA in women with equivocal or mildly abnormal Papanicolaou smears. Am J Clin Pathol. 2005;124:722–732.
26. Stoler MH. HPV testing in cervical cytology practice: it’s all about choice. Acta Cytol. 2005;49:117–119.
27. Goldie SJ, Kim JJ, Wright TC. Cost-effectiveness of HPV-DNA testing for cervical cancer screening in women aged 30 years or more. Obstet Gynecol. 2004; 103:619–631.
28. Goldie SJ, Kim JJ, Myers E. Cost-effectiveness of cervical cancer screening. Vaccine. 24S3, 2006, S3/164–S3/170.
29. Woodman CB, Collins S, Winter H, et al. Natural history of cervical HPV infection in young women: a longitudinal cohort study. Lancet. 2001;357:1831–1836.
30. Sasieni P, Cuzick J. Can HPV testing become the sole primary cervical screening test? J Med Screen. 2002;9:49–51.
31. Cuzick J, Clavel C, Petry KU, et al. Overview of the European and North American studies on HPV testing in primary cervical cancer screening. Int J Cancer. 2006;119:1095–1101.
32. Cuzick J, Mayrand MH, Ronco G, Snijders P, Wardle J. New dimensions in cervical cancer screening. Vaccine. 24S3, 2006; S3/90–S3/97.
33. Carcopino X, et al. HPV load may help determine cervical cancer risk. J Med Virol. 2006;78:1131–1140.
34. Ferlay J, Bray F, Pisani P, Parkin DM. GLOBOCAN 2000: Cancer incidence. Mortality and prevalence worldwide. Version 1.0 IARC Cancer Base No. 5. Lyon; IARC Press, 2001.
35. Davies P, Arbyn M, Dillner J, Kitchener H, Meijer C, Ronco G. A report on the current status of European Research on the use of HPV testing for primary cervical cancer screening. Int J Cancer. 2006;118:791–796.
36. Mayrand MH, Duarte-Franco E, Coutlee F, et al. Randomized controlled trial of HPV testing versus Pap cytology in the primary screening for cervical cancer precursors: design, methods and preliminary accrual results of the Canadian cervical cancer screening trial (CCCaST). Int J Cancer. 2006;119:615–623.
37. Kitchener HC, Almonte M, Wheeler P, et al. HPV testing in routine cervical screening: cross sectional data from the ARTISTIC trial. Br J Cancer. 2006;95:56–61.
38. Uribe P, Kaplan K, Zahn C. Evidence for the role of HPV-DNA testing in primary screening for cervical dysplasia. Clin Obst Gynecol. 2005;48:120–126.