Vol. 19 • Issue 1
• Page 12
The Molecular Edge
Human papillomaviruses (HPV) infect the epidermis and mucous membranes of humans worldwide. More than 120 distinct genotypes have been described, each differing from one another by >10% of sequence in their DNA genome.1Most do not cause disease. Some (e.g., HPV1 and 2) are associated with common warts of the skin. About 40 types are sexually transmitted and infect anogenital epithelia.
HPV is considered the most common sexually transmitted infection in the U.S. and ~75% of sexually active individuals will be infected some time during their lifetime.2Some types (e.g., HPV6 and 11) are associated with external genital warts or minor epithelial abnormalities; however, 13 to 15 types (the exact number and types vary depending on the source but all include HPV16 and 18) are considered high-risk (hr) or oncogenic (cancer-causing) in that they are commonly found in precancers and cancers of the cervix, vagina, vulva, anus and penis. HPV16 and 18 are almost exclusively associated with cancers of the larynx, oropharynx, mouth and eye, although not all of these cancers are HPV-related.
Which HPVs are Carcinogenic?
HPV carcinogenicity is most convincingly illustrated for cervical cancer; virtually all cases are associated with hr-HPV. Epidemiologic studies have categorically shown the carcinogenic potential of HPV16 and to a lesser extent, HPV18.3Collectively, these two types are associated with ~70% of cervical cancers and ~90% of HPV-positive head and neck cancers.
Determining the carcinogenic potential of other HPV genotypes has been difficult. There is a paucity of large, well-controlled studies that include testing for a broad range of genotypes. Multiple HPV genotypes are often present in individual cancers, making it difficult to know which one initiated cancer transformation, and some genotypes are rare. Moreover, laboratory HPV genotyping tests are far from standardized across studies, each showing differential sensitivities and specificities. At the 2009 meeting of the International Agency for Research on Cancer (IARC), 13 types, including HPV16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 39 and 59, were defined as carcinogens in the context of cervical cancer.3However, IARC also acknowledged that even so-called "low-risk types" may cause cervical cancer in rare circumstances.
HPV Testing Strategies
A large clinical trial started in the 1990s called the ASCUS-LSIL Triage Study (ALTS) set the benchmark for establishing the clinical utility of hr-HPV testing of cervical specimens. In 2001 the American Society of Colposcopy and Cervical Pathology (ASCCP) recommended incorporation of hr-HPV testing into standard cervical cancer screening. Guidelines have evolved into complex algorithms involving combinations of cytology, hr-HPV testing and colposcopy (visual assessment of the cervix) for different patient populations.4
One of the earliest recognized benefits of HPV testing was the increased clinical sensitivity that it provided over the standard repeat Pap of women with equivocal (ASC-US cytology) Paps. Women with an ASC-US Pap and a positive hr-HPV test have an increased risk for progression to high-grade lesions and should undergo colposcopy for assessment of visible disease. More recent guidelines recommend hr-HPV testing concurrent with the Pap test when screening women 30 years and older. HPV prevalence is relatively low in these patients (~10%) and discovering a persistent HPV infection puts them at significant risk.5,6So, performing cytology and HPV testing concurrently rather than reflexively allows one to risk-stratify these patients into those who require further assessment (if either test is positive) and those who don't need to be re-screened for at least three years (if both tests are negative). Women less than 30 years of age are more sexually active and have higher HPV infection rates; however, most infections are transient and cleared by natural immunological responses. Therefore, annual Pap screening with reflex HPV testing of ASC-US Paps is the best way to detect cervical disease in these younger women.
Current HPV Tests
Two FDA-approved hr-HPV tests are used in the U.S.: Qiagen Digene HPV Hybrid Capture 2 DNA Test (hc2), which detects 13 hrHPV genotypes, and the Hologic Cervista HPV HR test that detects the same genotypes as hc2, plus HPV66. Both are used reflexively for testing of liquid-based cervical cytology specimens from women with equivocal cytologies and for primary screening of women 30 years and older with any cytology. The Cervista HPV16/18 test is the only FDA-approved, genotype-specific test and is designed to be run reflexively on specimens positive by Cervista HPV HR. Other commercial and lab-developed genotyping assays vary greatly in the spectrum of HPV types detected.
Clinical Utility of HPV Genotyping
Since the highest risk for development of cervical cancer is attributed to HPV16 and 185,6 (all other genotypes account for only ~30% of cervical cancers), women who are persistently infected with these genotypes should be closely monitored clinically. This is why HPV16 and 18 are the sole genotypes represented in the FDA-approved genotyping test. Genotyping for HPV16/18 in women 30 years and older who are cytology-negative and hr-HPV-positive may permit referral to colposcopy sooner or more aggressive monitoring. Given the imprecision of colposcopy to detect precancerous lesions, it may be justified to treat women persistently infected with HPV16 or 18, even in the absence of a positive biopsy.7Less aggressive management strategies may be appropriate for women with other hr-HPV infections.
Another potential use of HPV genotyping is the longitudinal monitoring of younger (25- to 30-year-old) women for genotype-specific infections during primary screening.7Persistence of hr-HPV infection is strongly linked to the progression to cervical high-grade lesions and cancer. HPV genotyping, used as a general screening test, could identify a subset of women who are at high risk because they are persistently infected with a specific hr-HPV. By contrast, the risk is significantly lower for women who become infected with one hr-HPV genotype, transiently clear the infection, then become re-infected with different hr-HPV genotypes on a repeated basis. Monitoring for persistence of specific hr-HPVs could improve the sensitivity of HPV testing in predicting cervical disease in this population, but this needs to be practically demonstrated and the cost-effectiveness evaluated.
HPV genotyping may provide additional information in monitoring treatment success for high-grade cervical disease. The absence of detectable HPV or presence of a different HPV type(s) pre- versus post-treatment may be used in assessing treatment efficacy. Ultimately these tests could also be used to evaluate patients who have received prophylactic HPV16/18 vaccination; HPV infections in these individuals will most likely be different from the unvaccinated population and positive cytology screens may be more ambiguous.
Recent evidence indicates further utility for detection of HPV16/18 in oral and oropharyngeal cancers.8Overall, HPV is infrequent in oral (15-25%) and oropharyngeal (35-50%) cancers relative to cervical cancer (99%). HPV-related oropharyngeal tumors respond well to chemo-radiation and patients show a 60-80% reduction in risk of death relative to HPV-negative tumors. Because paraffin-embedded biopsies are frequently presented for analysis not amenable to hr-HPV screening assays and viral copy number is low in these tumors, PCR-based HPV 16/18 genotyping assays are the common method of choice.
Dr. Dunn is professor of Pathology and director of the Molecular Pathology Laboratory, University of Oklahoma Health Sciences Center.
For a list of references, go to www.advanceweb.com/labmanager
Potential Clinical Applications of HPV Genotyping:
Risk-stratifying women 30 years and older with cytology-negative results
Monitoring for persistence of specific hr-HPV genotypes in women 25 to 30 years old
Monitoring effectiveness of treatment of high-grade cervical disease
Evaluating HPV-vaccinated patients with equivocal or abnormal cytology
Risk-stratifying patients with oropharyngeal squamous cell carcinoma