Our objective was to assess the sensitivity and specificity of human papillomavirus (HPV) testing for cervical cancer screening in randomized trials. We conducted a systematic literature search of the following databases: MEDLINE, CINAHL, EMBASE, and Cochrane. Eligible studies were randomized trials comparing HPV-based to cytology-based screening strategies, with disease status determined by colposcopy/biopsy for participants with positive results. Disease rates (cervical intraepithelial neoplasia [CIN]2 or greater and CIN3 or greater), sensitivity, and positive predictive value were abstracted or calculated from the articles. Six studies met inclusion criteria. Relative sensitivities for detecting CIN3 or greater of HPV testing-based strategies vs cytology ranged from 0.8 to 2.1. The main limitation of our study was that testing methodologies and screening/management protocols were highly variable across studies. Screening strategies in which a single initial HPV-positive test led to colposcopy were more sensitive than cytology but resulted in higher colposcopy rates. These results have implications for cotesting with HPV and cytology as recommended in the United States.
Cytology, the primary modality for cervical cancer screening in the United States, has resulted in significant declines in cervical cancer morbidity and mortality. Nonetheless, cervical cytology has limitations, including a false-negative rate for cancer of at least 20%, leading to a search for more sensitive screening strategies. Human papillomavirus (HPV) is the causal factor of cervical cancer, and 18 HPV types including HPV16 and HPV18 have been associated with invasive cancer. HPV DNA testing has therefore been proposed as an alternative or adjunct for cervical cancer screening, with advantages that the test is more objective and sensitive than cytology.
Most women acquire HPV soon after sexual debut and spontaneously clear the virus within 1-2 years after infection; only approximately 10% of women remain HPV positive 5 years after acquisition. Although incident HPV infection is common, the risk for cervical cancer is associated with persistent infection. Therefore, the prognostic value of a single positive HPV test in young women is limited.
Recent guidelines in the United States recommend that in women older than 30 years, cervical cancer screening can be performed with either cytology every 3 years or cytology plus HPV cotesting every 5 years. In cases in which the HPV test is positive and cytology normal, repeating both tests in 12 months is recommended, unless HPV16 or HPV18 is present. In addition to cotesting as used in the United States, other strategies under evaluation globally include HPV testing alone without cytology and HPV testing first followed by cytology triage for positive results.
Metaanalyses comprised largely of observational studies have shown that HPV testing-based strategies are more sensitive but less specific than cytology-based screening strategies. Although observational studies are useful in determining test accuracy, performance estimates from observational studies can be biased when evaluating 2 diagnostic tests simultaneously in a manner that differs from how the tests are used separately. Potential bias is minimized when sensitivity and specificity of different strategies (single or combined tests) are analyzed in randomized trials. More reliable comparisons of sensitivity and specificity can be made in the context of randomized clinical trials conducted over 1 or more screening episodes.
Our objective was to assess the sensitivity and specificity of HPV testing in randomized trials by summarizing data from randomized trials of various cervical cancer screening strategies that incorporate HPV testing.
Materials and methods
We conducted a systematic literature search in 2010 of the following electronic databases: MEDLINE, CINAHL, EMBASE, and the Cochrane Library. We used the following subsets of search terms combined by the word “and:” (papillomavirus/papillomaviridae/papilloma virus/hpv) and (cytodiagnosis/cytolo*/pap smear/papanicolaou/colposcopy/cervical smear/uterine smear/cervicovagina smear/cervix uteri smear/endocervix smear) and (cervical cancer/cervical intraepithelial neoplasia/CIN/cervical dysplasia/cervical neoplasm/uterine cervical neoplasm/uterine cervix tumor/cervix cancer/uterine cervix carcinoma-in situ/uterine cervix dysplasia/cervix dysplasia/cervix neoplasm/uterine cervix cancer/mass screening/cancer screening). The following limit was placed on all searches to retrieve primarily randomized studies: “random*.” No language restrictions were included. The references cited in the articles selected for study inclusion were hand searched for additional citations. For each primary study, citations of relevant articles were searched for in MEDLINE, and a cited reference search was conducted in the Institute for Scientific Information Web of Knowledge electronic database. As a secondary analysis of published data, the study was exempt from institutional review board approval.
Study selection
We applied the following inclusion criteria: (1) the study was a randomized trial comparing HPV-based strategies to cytology-based strategies for primary cervical cancer screening and (2) disease status was determined by colposcopy/biopsy for study participants in whom treatment was warranted. Our study was limited to screening strategies and therefore did not assess the use of reflex HPV testing for a cytology result of atypical squamous cells of undetermined significance (ASCUS). None of the studies in this review used genotyping assays recently approved by the US Food and Drug Administration.
Studies were selected with a 2-step method. First, resulting titles and abstracts from literature searches were analyzed, and citations that were likely to meet the aforementioned criteria were chosen. The full manuscripts of these citations were then evaluated to determine whether full inclusion criteria were met.
Each study was abstracted onto pretested data abstraction forms by at least 2 reviewers with a third reviewer for adjudication of discrepancies. In cases of multiple publications from a single study, the publication analyzing the most recent dataset was used. If this was unclear, the study authors were contacted for clarification; repeat data abstraction was performed if indicated by the authors’ response. In addition, although another study (the HPV FOCAL trial: a randomized trial of human papillomavirus testing for cervical cancer screening) met our inclusion criteria, colposcopy results were not published; therefore, it was excluded from the systematic review. The final results from one trial were published after the performance of the systematic literature review; results from this publication were abstracted and used in the analysis primarily rather than the interim results.
Study classification
We devised a simplified classification system of different HPV testing strategies to determine whether differences in outcomes were related to testing strategy. These strategies, illustrated in Figure 1 , included the following: (1) HPV testing alone (no cytology) with referral colposcopy for positive HPV test results, (2) HPV testing with cytology triage for positive HPV test results, (3) combination of cytology and HPV testing (cotesting) with an active response to positive HPV testing results (women with a positive initial HPV test were referred to colposcopy), and (4) cotesting with a passive response to positive HPV testing results (women with a positive HPV test but normal cytology underwent more frequent surveillance but were not referred to colposcopy based on the initial positive HPV test result).
Study quality
Quality was assessed using the following markers for internal validity: method of randomization, blinding, intention-to-treat analysis, and loss to follow-up. Blinding was assessed with a point given for each entity blinded: patient, clinician, pathologist, cytologist, and statistician.
Study quality was determined by using the Quality Assessment of Diagnostic Accuracy (QUADAS) questionnaire, an evidence-based tool that assesses the quality of diagnostic accuracy studies. Although a 14-point QUADAS tool is used primarily to assess the quality of observational studies rather than randomized trials of diagnostic tests, we used a modified 11-point assessment to provide additional information on study quality. The modified QUADAS tool covered the following topics: representativeness of the study population to a screening population, description of selection criteria, validity of reference standard, the delay between index and reference tests, the reproducibility of these tests, blinding, clinical information available to clinicians, and reporting of inadequate results and loss to follow-up. Based on the criteria used in previous studies, we chose a score of 50% or higher (at least 6 of 11 points) to denote a high-quality study.
Values abstracted and computed
Disease outcome measures for the purposes of this study were cervical intraepithelial neoplasia (CIN)2 or worse (CIN2 or greater) and CIN3 or worse (CIN3 or greater). For each study, the rates of HPV test positivity/abnormal cytology rates or percent referred to colposcopy were either abstracted or calculated. When the positivity/abnormality rate was not recorded, the colposcopy referral rate was used instead because these values should theoretically be the same. The only exception was for passive-response studies in which a positive HPV test did not result in a referral to colposcopy. The only passive-response study that did not give a colposcopy referral rate was phase 1 of the New Technologies for Cervical Cancer Screening (NTCCS) trial in the 25-34 year age group. Hereafter, the term, test positivity rate, will signify HPV positivity rate, abnormal cytology rate, or colposcopy referral rate.
Using these values, the rates of CIN2 or greater and CIN3 or greater per positive test result were calculated by multiplying the test positivity rate by the total number of participants in each group and using that result as the denominator and the total number of CIN cases as the numerator. To calculate the rate of disease per person screened, the total number of participants was used as the denominator. Most of these data were not available for the second round of studies with multiple screening rounds; therefore, these measures were reported only for the first round of screening.
To assess the performance of the different strategies, the values for relative sensitivity, specificity, and relative positive predictive value (PPV) for HPV testing vs cytology-based strategies were directly abstracted from the publication if available. If not stated in the text, relative sensitivity was computed by taking the ratio of disease rate in the HPV based-testing group divided by the disease rate in the cytology-based testing group.
Comparisons of test specificity were reported for the A Randomised Trial of HPV Testing in Primary Cervical Screening (ARTISTIC) trial and the Finnish trial. The relative PPV was used as a marker for specificity and was calculated with the number of cases found divided by the number of participants who underwent colposcopy used as the denominator. The relative sensitivities for the Population Based Screening Study Amsterdam (POBASCAM) trial and relative sensitivities and relative PPVs for the India trial were calculated in the manner described in the previous text. The relative sensitivities and PPVs were calculated for the ARTISTIC trial using published sensitivities and PPVs.
One potential harm associated with cervical cancer screening is the excess number of colposcopies and biopsies needed to detect 1 case of disease (number needed to colposcopy [NNC]). The NNC for CIN2 or greater and CIN3 or greater was calculated for round 1 as the reciprocal of the number of CIN cases per positive test result. The NNC was not calculated for the NTCCS phase 1 study in 25-34 year olds because the colposcopy rate was not given. The SwedeScreen study was also not included because overall sensitivities and PPVs were not published. The colposcopy rate used for the POBASCAM trial was reported in the earlier publication.
Statistical analysis
Statistical methods used to evaluate sensitivities varied between individual studies and included a χ 2 test, a Fisher exact test, and Poisson regression analysis. For the studies in which we calculated relative sensitivities based on the published data, statistical comparison was not performed.
The NNC values were compared statistically by calculating the z-statistic on the SE of the difference between the NNC for the HPV arm and the cytology arm. A P value was then determined for this z-statistic, and the statistical significance was defined as P < .05.
Results
Study characteristics and quality
Six studies met inclusion criteria and were chosen after undergoing the evaluation process described in the previous text ( Figure 2 ). Covariate characteristics such as study location, study size, and exclusion criteria are summarized in Table 1 . The sample sizes ranged from 12,410 to 131,746 women; combined the studies included a total of 422,084 women. Four studies had data available from 2 screening rounds. A round is defined by a screening episode; in the second round, participants from the first round were rescreened. The time between screening intervals ranged from 3 to 5 years. Outcome measures ranged from CIN1 or greater to cancer death. Cancer is rare in countries with established screening programs and the numbers of cases were small in most studies.
| Author, year, country | Comparison | Exclusion criteria | HPV test used a | Cytology method | Outcome measured | Threshold for colposcopy | Size, N C /N H b |
|---|---|---|---|---|---|---|---|
| Ronco et al, 2010, Italy (NTCCS trial) Total n = 94,370 | Phase 1, 25-34 y old: cytology alone vs HPV plus cytology (passive response) | Pregnant, history of hysterectomy, treatment for CIN in past 5 y, age 25 y or younger or 60 y or older | hc2 | Conventional and LBC | CIN2 or greater, CIN3 or greater, cancer c | Cyto: ASCUS or greater (7/9 centers) or LSIL or greater (2/9 centers) HPV: cytology positive as above or HPV test remains positive 12 months after initial testing | 12,410 5808/6602 |
| Phase 1, 35-60 y old: cytology alone vs HPV plus cytology (active response) | Cyto: ASCUS or greater (7/9 centers) or LSIL (2/9 centers) HPV: positive HPV test or positive cytology, as above | 33,364 16,658/16,706 | |||||
| Phase 2, all ages: cytology alone vs HPV alone | Cyto: ASCUS or greater (7/9 centers) or LSIL or greater (2/9 centers) HPV: positive HPV test | 49,196 24,535/24,661 | |||||
| Leinonen et al, 2009, Finland (Finnish trial) | Cytology alone vs HPV plus cytology triage | Age 25 y or younger or 65 y or older | hc2 | LBC | CIN1 or greater, CIN2 or greater, CIN3 or greater | Cyto: LSIL or greater HPV: positive HPV test followed by cytology LSIL or greater | 71,337 35,500/35,837 |
| Kitchener et al, 2009, United Kingdom (ARTISTIC trial) | Cytology alone vs HPV plus cytology (passive response) | Age 20 y or younger or 60 y or older | hc2 | Conventional | CIN2 or greater, CIN3 or greater | Cyto: HSIL or greater, 2 consecutive LSIL results, or 3 consecutive ASCUS results HPV: HPV test remains positive 12 months after initial testing or HSIL or greater | 24,510 6124/18,386 |
| Sankaranarayanan et al, 2009, India (India trial) | Cytology alone vs HPV alone | Pregnant; history of cervical cancer, hysterectomy, or uterine prolapse; no history of marriage; age 30 y or younger or 59 y or older | hc2 | Conventional | Cancer, cancer death | Cyto: ASCUS or greater HPV: positive HPV test | 131,746 32,058/34,126 |
| Rikjaart et al, 2012, Netherlands (POBASCAM trial) | Cytology alone vs HPV plus cytology (passive response) | History of hysterectomy, CIN2 or greater or worse, or abnormal cytology in previous 2 y, age 29 y or younger or 56 y or older | GP5/GP6 | Conventional | CIN2 or greater, CIN3 or greater | Cyto: HSIL or greater initially or repeated ASCUS results at 6 months HPV: HPV test remains positive at 6 or 18 months after initial testing or positive cytology, as above | 40105 19999/20106 |
| Naucler et al, 2007, Sweden (SwedeScreen trial) | Cytology alone vs HPV plus cytology (passive response) | Age 32 y or younger or 38 y or older | GP5/GP6 | Conventional | CIN2 or greater, CIN3 or greater | Cyto: ASCUS or greater (Stockholm) or HSIL or greater (all other cities) HPV: HPV test remains positive 12 months after initial testing or positive cytology, as above | 12527 6270/6257 |
a The hc2 (hybrid capture 2) was considered positive if 1 relative luciferase unit or greater;
b N C /N H is the number of study subjects in the cytology group/number of study subjects in the HPV group;
c In most recent publication, CIN2+ signifies CIN2, CIN3, and AIS combined. CIN3+ signifies CIN3 and AIS combined.
The NTCCS study evaluated 3 different HPV testing–based methodologies compared with cytology in separate populations imbedded in 1 large study. Phase 1 evaluated a combination of HPV and cytology with differing responses to the HPV test based on age: younger women received passive follow-up, whereas older women received active follow-up ( Figure 1 ). Phase 2 enrolled additional participants to evaluate HPV testing alone compared with cytology in all ages. The different trials were analyzed individually. Although the NTCCS trials categorized results based on age, most studies did not do this for all of their outcomes.
The studies differed in important methodological aspects such as HPV testing methods and thresholds for colposcopy ( Table 1 ). The POBASCAM and SwedeScreen studies used general primer pair GP5-GP6 for HPV DNA polymerase chain reaction testing, whereas the other studies used the Hybrid Capture 2 (hc2) high-risk HPV test (Qiagen, Gaithersburg, MD).
Some studies used liquid based-cytology (LBC), whereas others used conventional cytology. Phase 1 of the NTCCS study used conventional cytology in the control arm and LBC combined with HPV testing in the experimental arm. Also, NTCCS used HPV testing and/or cytology in round 1 but only cytology in round 2 for all participants, meaning that the randomized screening strategy was not continued into round 2. Similarly, POBASCAM performed both HPV testing and cytology in all participants in round 2. Most importantly, the cytological and temporal thresholds for colposcopy referral varied between studies.
Two studies used ASCUS, 1 used low-grade squamous intraepithelial lesion (LSIL), and 3 used high-grade squamous intraepithelial lesion (HSIL) as the threshold for colposcopy. One study with passive response retested at 6 and 18 months, and 3 studies with passive response retested at 12 months ( Table 1 ).
The presence of study quality measures is summarized in Table 2 . Studies ranged in duration from 6.5 to 8 years. All studies had 2 rounds over this time period with the exception of the India study. The second round of the Finnish study is still in progress. Most studies were of good quality, scoring 9-10 of 11 with the QUADAS tool. Although still of high quality, the Finnish study had the lowest QUADAS score (6 of 11) because participant selection criteria were not clearly defined, the delay between positive index test and colposcopy was not described, colposcopy methods were not described in detail, and unsatisfactory test results and loss to follow-up were not reported.
| Author, year | Rounds, n | Time frame | Randomization | Blinding a | Intention to screen | Loss to follow-up (HPV %/Cyto %) | QUADAS score (x/11) |
|---|---|---|---|---|---|---|---|
| Ronco et al, 2010 (NTCCS trial) | 2 | February 2002 through November 2008 | Computerized blocks and series of random numbers | 1/5 | Yes | Varies by phase | 9 |
| Leinonen et al, 2009 (Finnish trial) | 1 b | January 2003 to present | Based on national ID number | None | Yes | NR | 6 |
| Kitchener et al, 2009 (ARTISTIC trial) | 2 | July 2001 through July 2008 | Series of random numbers | 2/5 | Yes | 34.2/34.9 | 9 |
| Sankaranarayanan et al, 2009 (India trial) | 1 | January 2000 through December 2007 | Method not stated | None | Yes | 20.3/20.3 | 9 |
| Rikjaart et al, 2012 (POBASCAM trial) | 2 | January 1999 through February 2007 | Series of random numbers | 2/5 | Yes | 14.1/14.9 c | 9 |
| Naucler et al, 2007 (SwedeScreen trial) | 2 | May 1997 through August 2005 | Series of random numbers | 3/5 | Yes | 12.9/12.2 | 10 |
a A point was assigned for each entity blinded: patient, clinician, pathologist, cytologist, statistician;
b Data from 1 round is available, whereas the second round is still in progress;
c Percentages calculated from raw numbers regarding eligibility and attendance published in paper.
Given that the HPV testing strategy, the HPV testing method, the cytology threshold for colposcopic referral, the type of cytology (LBC vs conventional), and age were heterogeneous in the included studies, a formal meta-analysis could not be conducted.
Accuracy of cytology vs HPV testing
To compare the numbers of interventions required to find disease, the test positivity rates and the rates of disease per woman screened and per woman who tested positive in round 1 are presented in Table 3 . The test positivity for the HPV-based strategies ranged from 1.2% to 13.1% and the test positivity for the cytology-based strategies ranged from 1.2% to 7.0%. The wide range in positivity rates for HPV-based strategies were most likely related to population differences in HPV prevalence and different responses to positive HPV tests.
| Author | Comparison | Age range, y | Positivity rate, (%) a | Rate per woman screened (CIN2 or greater), % | Rate per positive result (CIN2 or greater), % | Rate per woman screened (CIN3 or greater), % | Rate per positive result (CIN3 or greater), % | |||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| HPV | Cyto | HPV | Cyto | HPV | Cyto | HPV | Cyto | HPV | Cyto | |||
| Ronco et al (2010) (NTCCS) | HPV plus cytology (passive response) vs cytology alone | 25–34 | 11.7 | 4.6 | 1.2 | 0.6 | 10.1 | 17.7 | 0.4 | 0.4 | 3.0 | 11.0 |
| Ronco et al (2010) (NTCCS) | HPV plus cytology (active response) vs cytology alone | 35–60 | 10.6 | 3.6 | 0.6 | 0.3 | 6.0 | 9.2 | 0.3 | 0.2 | 2.8 | 4.5 |
| Ronco et al (2010) (NTCCS) | HPV alone vs cytology alone | 25–34 | 13.1 | 4.0 | 1.7 | 0.4 | 12.7 | 9.2 | 0.6 | 0.2 | 4.8 | 4.1 |
| Ronco et al (2010) (NTCCS) | HPV alone vs cytology alone | 35–60 | 5.8 | 3.1 | 0.5 | 0.3 | 9.5 | 8.4 | 0.3 | 0.1 | 4.7 | 3.6 |
| Leinonen et al (2009) (Finnish trial) | HPV plus cytology triage vs cytology alone | 25–65 | 1.2 | 1.2 | 0.4 | 0.3 | 37.0 | 25.4 | 0.1 | 0.1 | 10.7 | 8.0 |
| Kitchener et al (2009) (ARTISTIC trial) | HPV plus cytology (passive response) vs cytology alone | 20–64 | 6.8 | 5.2 | 2.5 | 2.2 | 36.2 | 41.8 | 1.3 | 1.3 | 18.6 | 25.1 |
| Sankaranarayanan et al (2009) (India trial) b | HPV alone vs cytology alone | 30–59 | 10.3 | 7.0 | 0.9 | 1.1 | 9.1 | 15.4 | 0.4 | 0.5 | 3.6 | 6.8 |
| Rikjaart et al (2012) (POBASCAM trial) c | HPV plus cytology (passive response) vs cytology alone | 29–56 | 2.3 | 1.3 | 1.1 | 0.7 | 47 | 49 | 0.8 | 0.5 | 33 | 32 |
| Naucler et al (2007) (SwedeScreen trial) | HPV plus cytology (passive response) vs cytology alone | 32–38 | NR | NR | 1.8 | 1.2 | NA | NA | 1.2 | 0.9 | NA | NA |
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