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Supplemental Table 1: Screening history of diagnosed cancer cases Supplemental Table 2: Outcome by age and baseline histology among women with conservatively managed CIN1/2, CIN2, CIN2/3 biopsy results

Purpose: Human papillomavirus (HPV) DNA methylation testing is a promising triage option for women testing HPV positive during cervical cancer screening. However, the extent to which methylation indicates precancer for all 12 carcinogenic HPV types has not been evaluated.Experimental Design: In this nested case–control study, we tested up to 30 cases of precancer [cervical intraepithelial neoplasia grade 3 (CIN3)/adenocarcinoma in situ (AIS)] and 30 normal controls for each carcinogenic type (single infections with 16/18/31/33/35/39/45/51/52/56/58/59). Next-generation bisulfite sequencing was performed on CpG sites within the L1 and L2 genes. We calculated differences in methylation, ORs, and AUC. Using a fixed sensitivity of 80%, we evaluated the specificity and the risk of CIN3/AIS for best performing CpG sites, and compared the performance of an explorative multi-type methylation assay with current triage strategies.Results: Methylation was positively associated with CIN3/AIS across all 12 types. AUCs for the top sites ranged from 0.71 (HPV51 and HPV56) to 0.86 (HPV18). A combined 12-type methylation assay had the highest Youden index (0.46), compared with cytology (0.31) and a 5-type methylation assay, including only previously described types (0.26). The 12-type methylation assay had higher sensitivity (80% vs. 76.6%) and lower test positivity compared with cytology (38.5% vs. 48.7%). The risk of CIN3/AIS was highest for methylation positives and lowest for cytology or HPV16/18 positives.Conclusions: HPV DNA methylation is a general phenomenon marking the transition from HPV infection to precancer for all 12 carcinogenic types. Development of a combined multitype methylation assay may serve as a triage test for HPV-positive women. Clin Cancer Res; 24(9); 2194–202. ©2018 AACR.

Background: Human papillomavirus (HPV) RNA detection is reportedly more specific for the detection of anogenital precancer than HPV DNA but it is unknown whether this is due to detection of RNA or due to HPV genotype restriction.Methods: A total of 363 human immunodeficiency virus (HIV)–positive men who have sex with men had two anal cytology samples taken and were evaluated using high-resolution anoscopy and biopsies of visible lesions. Anal specimens were tested for E6/E7 RNA for five carcinogenic HPV genotypes (HPV16, 18, 31, 33, and 45) and tested for the DNA of 13 carcinogenic HPV genotypes.Results: DNA testing was more likely to be positive than RNA testing (53% vs. 48%; P = 0.02) for the same five HPV genotypes in aggregate. When restricted to five HPV genotypes targeted by the RNA test, the sensitivity to detect anal precancer was the same for DNA and RNA (81%), whereas RNA was more specific than DNA (65% vs. 58%; P = 0.007). In comparison, DNA detection of all 13 carcinogenic HPV genotypes was more sensitive (96% vs. 81%; P = 0.001) but much less specific (65% vs. 33%; P < 0.001) as compared with RNA detection of the five HPV genotypes.Conclusion: After controlling for HPV genotypes, RNA was only slightly more specific than DNA detection for anal precancer.Impact: DNA or RNA testing for a subset of the most carcinogenic HPV genotypes may be useful for distinguishing between those HPV-positive men at higher and lower risk of anal precancer and cancer. Cancer Epidemiol Biomarkers Prev; 22(1); 42–9. ©2012 AACR.

Table S1 shows type-specific associations of methylation with CIN3/AIS. Table S2 shows the correlation of methylation across CpG sites, by HPV type

Study population

The majority of cervical cancers after Pap-test–positive, human papillomavirus test–negative co-tests are symptomatic or clinically apparent.

BACKGROUND: State-of-the-art cervical cancer prevention includes human papillomavirus (HPV) vaccination among adolescents and screening/treatment of cervical precancer (CIN3/AIS and, less strictly, CIN2) among adults. HPV testing provides sensitive detection of precancer but, to reduce overtreatment, secondary “triage” is needed to predict women at highest risk. Those with the highest-risk HPV types or abnormal cytology are commonly referred to colposcopy; however, expert cytology services are critically lacking in many regions. METHODS: To permit completely automatable cervical screening/triage, we designed and validated a novel triage method, a cytologic risk score algorithm based on computer-scanned liquid-based slide features (FocalPoint, BD, Burlington, NC). We compared it with abnormal cytology in predicting precancer among 1839 women testing HPV positive (HC2, Qiagen, Germantown, MD) in 2010 at Kaiser Permanente Northern California (KPNC). Precancer outcomes were ascertained by record linkage. As additional validation, we compared the algorithm prospectively with cytology results among 243 807 women screened at KPNC (2016–2017). All statistical tests were two-sided. RESULTS: Among HPV-positive women, the algorithm matched the triage performance of abnormal cytology. Combined with HPV16/18/45 typing (Onclarity, BD, Sparks, MD), the automatable strategy referred 91.7% of HPV-positive CIN3/AIS cases to immediate colposcopy while deferring 38.4% of all HPV-positive women to one-year retesting (compared with 89.1% and 37.4%, respectively, for typing and cytology triage). In the 2016–2017 validation, the predicted risk scores strongly correlated with cytology (P < .001). CONCLUSIONS: High-quality cervical screening and triage performance is achievable using this completely automated approach. Automated technology could permit extension of high-quality cervical screening/triage coverage to currently underserved regions.

Although guidelines have recommended extended interval cervical screening using concurrent human papillomavirus (HPV) and cytology ("cotesting") for over a decade, little is known about its adoption into routine care. Using longitudinal medical record data (2003-2015) from Kaiser Permanente Northern California (KPNC), which adopted triennial cotesting in 2003, we examined adherence to extended interval screening. We analyzed predictors of screening intervals among 491,588 women undergoing routine screening, categorizing interval length into early (

OBJECTIVE To inform impending postcolposcopy guidelines, this analysis examined the subsequent risk of CIN 3+ among women with a grade lower than CIN 2 (< CIN 2) colposcopy results, taking into account the referring results that brought them to colposcopy and cotest results postcolposcopy. METHODS We analyzed 107,005 women from 25 to 65 years old, recommended for colposcopy at Kaiser Permanente Northern California. We estimated absolute risks of CIN 3+ among women: (1) recommended for colposcopy (precolposcopy), (2) following colposcopy and with histology results < CIN 2 (postcolposcopy), and (3) with cotest results 12 months after a < CIN 2 colposcopy (return cotest). RESULTS After colposcopy showing < CIN 2 (n = 69,790; 87% of the women at colposcopy), the 1-year risk of CIN 3+ was 1.2%, compared with 6.3% at the time of colposcopy recommendation. Negative cotest results 1 year after colposcopy identified a large group (37.1%) of women whose risk of CIN 3+ (i.e.

Background The main goal of cervical screening programs is to detect and treat precancer before cancer develops. Human papillomavirus (HPV) testing is more sensitive than cytology for detecting precancer. However, reports of rare HPV-negative, cytology-positive cancers are motivating continued use of both tests (cotesting) despite increased testing costs. Methods We quantified the detection of cervical precancer and cancer by cotesting compared with HPV testing alone at Kaiser Permanente Northern California (KPNC), where 1 208 710 women age 30 years and older have undergone triennial cervical cotesting since 2003. Screening histories preceding cervical cancers (n = 623) and precancers (n = 5369) were examined to assess the relative contribution of the cytology and HPV test components in identifying cases. The performances of HPV testing and cytology were compared using contingency table methods, general estimating equation models, and nonparametric statistics; all statistical tests were two-sided. Results HPV testing identified more women subsequently diagnosed with cancer (P < .001) and precancer (P < .001) than cytology. HPV testing was statistically significantly more likely to be positive for cancer at any time point (P < .001), except within 12 months (P = .10). HPV-negative/cytology-positive results preceded only small fractions of cases of precancer (3.5%) and cancer (5.9%); these cancers were more likely to be regional or distant stage with squamous histopathology than other cases. Given the rarity of cancers among screened women, the contribution of cytology to screening translated to earlier detection of at most five cases per million women per year. Two-thirds (67.9%) of women found to have cancer during 10 years of follow-up at KPNC were detected by the first cotest performed. Conclusions The added sensitivity of cotesting vs HPV alone for detection of treatable cancer affected extremely few women.

As a foundation for the next guidelines revision, we confirmed with additional precision the risk estimates previously reported for combinations of human papillomavirus and cytology screening., Objectives The next round of the American Society for Colposcopy and Cervical Pathology (ASCCP)-sponsored cervical cancer screening and management guidelines will recommend clinical actions based on risk, rather than test-based algorithms. This article gives preliminary risk estimates for the screening setting, showing combinations of the 2 most important predictors, human papillomavirus (HPV) status and cytology result. Materials and Methods Among 1,262,713 women aged 25 to 77 years co-tested with HC2 (Qiagen) and cytology at Kaiser Permanente Northern California, we estimated 0–5-year cumulative risk of cervical intraepithelial neoplasia (CIN) 2+, CIN 3+, and cancer for combinations of cytology (negative for intraepithelial lesion or malignancy [NILM], atypical squamous cells of undetermined significance [ASC-US], low-grade squamous intraepithelial lesion [LSIL], atypical squamous cells cannot exclude HSIL [ASC-H], high-grade squamous intraepithelial lesion [HSIL], atypical glandular cells [AGC]) and HPV status. Results Ninety percent of screened women had HPV-negative NILM and an extremely low risk of subsequent cancer. Five-year risks of CIN 3+ were lower after HPV negativity (0.12%) than after NILM (0.25%). Among HPV-negative women, 5-year risks for CIN 3+ were 0.10% for NILM, 0.44% for ASC-US, 1.8% for LSIL, 3.0% for ASC-H, 1.2% for AGC, and 29% for HSIL+ cytology (which was very rare). Among HPV-positive women, 5-year risks were 4.0% for NILM, 6.8% for ASC-US, 6.1% for LSIL, 28% for ASC-H, 30% for AGC, and 50% for HSIL+ cytology. Conclusions As a foundation for the next guidelines revision, we confirmed with additional precision the risk estimates previously reported for combinations of HPV and cytology. Future analyses will estimate risks for women being followed in colposcopy clinic and posttreatment and will consider the role of risk modifiers such as age, HPV vaccine status, HPV type, and screening and treatment history.

Background Efficient tools are needed to stage liver disease before treatment of patients infected with hepatitis C virus (HCV). Compared to biopsy, several studies demonstrated favorable performance of noninvasive multianalyte serum fibrosis marker panels [fibrosis-4 (FIB-4) index] and aspartate aminotransferase (AST)-to-platelet ratio index (APRI), but suggested cutoffs vary widely. Our objective was to evaluate FIB-4 index and APRI and their component tests for staging fibrosis in our HCV-infected population and to determine practical cutoffs to help triage an influx of patients requiring treatment. Methods Transient elastography (TE) results from 1731 HCV-infected patients were mapped to an F0–F4 equivalent scale. Each patient's APRI and FIB-4 index were calculated. Areas under the receiver operator curve (AUROCs) and false-positive and false-negative rates were calculated to retrospectively compare the performance of the indices and their component tests. Results The highest AUROCs for distinguishing severe (F3–F4) from mild-to-moderate (F0–F2) fibrosis had overlapping 95% CIs: APRI (0.77; 0.74–0.79), FIB-4 index (0.76; 0.73–0.78), and AST (0.74; 0.72–0.77). Cutoffs had false-negative rates of 2.7%–2.8% and false-positive rates of 6.4%–7.4% for all 3 markers. Conclusions AST was as effective as FIB-4 index and APRI at predicting fibrosis. Published cutoffs for APRI and FIB-4 index would have been inappropriate in our population, with false-negative rates as high as 11%. For our purposes, no serum fibrosis marker was sufficiently sensitive to rule-out significant fibrosis, but cutoffs developed for AST, FIB-4 index, and APRI all had specificities of 79.2%–80.3% for ruling-in severe fibrosis and could be used to triage 1/3 of our population for treatment without waiting for TE or liver biopsy.

For cost-effectiveness and efficiency, many large-scale general-purpose cohort studies are being assembled within large health-care providers who use electronic health records. Two key features of such data are that incident disease is interval-censored between irregular visits and there can be pre-existing (prevalent) disease. Because prevalent disease is not always immediately diagnosed, some disease diagnosed at later visits are actually undiagnosed prevalent disease. We consider prevalent disease as a point mass at time zero for clinical applications where there is no interest in time of prevalent disease onset. We demonstrate that the naive Kaplan-Meier cumulative risk estimator underestimates risks at early time points and overestimates later risks. We propose a general family of mixture models for undiagnosed prevalent disease and interval-censored incident disease that we call prevalence-incidence models. Parameters for parametric prevalence-incidence models, such as the logistic regression and Weibull survival (logistic-Weibull) model, are estimated by direct likelihood maximization or by EM algorithm. Non-parametric methods are proposed to calculate cumulative risks for cases without covariates. We compare naive Kaplan-Meier, logistic-Weibull, and non-parametric estimates of cumulative risk in the cervical cancer screening program at Kaiser Permanente Northern California. Kaplan-Meier provided poor estimates while the logistic-Weibull model was a close fit to the non-parametric. Our findings support our use of logistic-Weibull models to develop the risk estimates that underlie current US risk-based cervical cancer screening guidelines. Published 2017. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

BACKGROUND The objective of cervical screening is to detect and treat precancer to prevent cervical cancer mortality and morbidity while minimizing overtreatment of benign human papillomavirus (HPV) infections and related minor abnormalities. HPV/cytology cotesting at extended 5-year intervals currently is a recommended screening strategy in the United States, but the interval extension is controversial. In the current study, the authors examined the impact of a decade of an alternative, 3-year cotesting, on rates of precancer and cancer at Kaiser Permanente Northern California. The effect on screening efficiency, defined as numbers of cotests/colposcopy visits needed to detect a precancer, also was considered. METHODS Two cohorts were defined. The “open cohort” included all women screened at least once during the study period; > 1 million cotests were performed. In a fixed “long-term screening cohort,” the authors considered the cumulative impact of repeated screening at 3-year intervals by restricting the cohort to women first cotested in 2003 through 2004 (ie, no women entering screening later were added to this group). RESULTS Detection of cervical intraepithelial neoplasia 3/adenocarcinoma in situ (CIN3/AIS) increased in the open cohort (2004-2006: 82.0/100,000 women screened; 2007-2009: 140.6/100,000 women screened; and 2010-2012: 126.0/100,000 women screened); cancer diagnoses were unchanged. In the long-term screening cohort, the detection of CIN3/AIS increased and then decreased to the original level (2004-2006: 80.5/100,000 women screened; 2007-2009: 118.6/100,000 women screened; and 2010-2012: 84.9./100,000 women screened). The number of cancer diagnoses was found to decrease. When viewed in terms of screening efficiency, the number of colposcopies performed to detect a single case of CIN3/AIS increased in the cohort with repeat screening. CONCLUSIONS Repeated cotesting at a 3-year interval eventually lowers population rates of precancer and cancer. However, a greater number of colposcopies are required to detect a single precancer. Cancer 2016. © 2016 American Cancer Society.

HPV testing is more sensitive than cytology for cervical screening. However, to incorporate HPV tests into screening, risk-stratification ("triage") of HPV-positive women is needed to avoid excessive colposcopy and overtreatment. We prospectively evaluated combinations of partial HPV typing (Onclarity, BD) and cytology triage, and explored whether management could be simplified, based on grouping combinations yielding similar 3-year or 18-month CIN3+ risks. We typed ∼9,000 archived specimens, taken at enrollment (2007-2011) into the NCI-Kaiser Permanente Northern California (KPNC) HPV Persistence and Progression (PaP) cohort. Stratified sampling, with reweighting in the statistical analysis, permitted risk estimation of HPV/cytology combinations for the 700,000+-woman KPNC screening population. Based on 3-year CIN3+ risks, Onclarity results could be combined into five groups (HPV16, else HPV18/45, else HPV31/33/58/52, else HPV51/35/39/68/56/66/68, else HPV negative); cytology results fell into three risk groups ("high-grade," ASC-US/LSIL, NILM). For the resultant 15 HPV group-cytology combinations, 3-year CIN3+ risks ranged 1,000-fold from 60.6% to 0.06%. To guide management, we compared the risks to established "benchmark" risk/management thresholds in this same population (e.g., LSIL predicted 3-year CIN3+ risk of 5.8% in the screening population, providing the benchmark for colposcopic referral). By benchmarking to 3-year risk thresholds (supplemented by 18-month estimates), the widely varying risk strata could be condensed into four action bands (very high risk of CIN3+ mandating consideration of cone biopsy if colposcopy did not find precancer; moderate risk justifying colposcopy; low risk managed by intensified follow-up to permit HPV "clearance"; and very low risk permitting routine screening.) Overall, the results support primary HPV testing, with management of HPV-positive women using partial HPV typing and cytology.

Purpose Obesity has been inconsistently linked to increased cervical cancer incidence and mortality; however, the effect of obesity on cervical screening has not been explored. We investigated the hypothesis that increased body mass might decrease detection of cervical precancer and increase risk of cervical cancer even in women undergoing state-of-the-art screening. Methods We conducted a retrospective cohort study of 944,227 women age 30 to 64 years who underwent cytology and human papillomavirus DNA testing (ie, cotesting) at Kaiser Permanente Northern California (January 2003 to December 2015). Body mass index was categorized as normal/underweight (< 25 kg/m2), overweight (25 to < 30 kg/m2), or obese (≥ 30 kg/m2). We estimated 5-year cumulative risks of cervical precancer and cancer by category of body mass index using logistic Weibull survival models. Results We observed lower risk of cervical precancer (n = 4,489) and higher risk of cervical cancer (n = 490) with increasing body mass index. Specifically, obese women had the lowest 5-year risk of precancer (0.51%; 95% CI, 0.48% to 0.54% v 0.73%; 95% CI, 0.70% to 0.76% in normal/underweight women; P trend < .001). In contrast, obese women had the highest 5-year risk of cancer (0.083%; 95% CI, 0.072% to 0.096% v 0.056%; 95% CI, 0.048% to 0.066% in normal/underweight women; P trend < .001). Results were consistent in subgroups defined by age (30 to 49 v 50 to 64 years), human papillomavirus status (positive v negative), and histologic subtype (glandular v squamous). Approximately 20% of cervical cancers could be attributed to overweight or obesity in the women in our study who underwent routine cervical screening. Conclusion In this large, screened population, overweight and obese women had an increased risk of cervical cancer, likely because of underdiagnosis of cervical precancer. Improvements in equipment and/or technique to assure adequate sampling and visualization of women with elevated body mass might reduce cervical cancer incidence.

As cervical cancer screening shifts from cytology to human papillomavirus (HPV) testing, a major question is the clinical value of identifying individual HPV types. We aimed to validate Onclarity (Becton Dickinson Diagnostics, Sparks, MD), a nine-channel HPV test recently approved by the FDA, by assessing (i) the association of Onclarity types/channels with precancer/cancer; (ii) HPV type/channel agreement between the results of Onclarity and cobas (Roche Molecular Systems, Pleasanton, CA), another FDA-approved test; and (iii) Onclarity typing for all types/channels compared to typing results from a research assay (linear array [LA]; Roche). We compared Onclarity to histopathology, cobas, and LA. We tested a stratified random sample ( n = 9,701) of discarded routine clinical specimens that had tested positive by Hybrid Capture 2 (HC2; Qiagen, Germantown, MD). A subset had already been tested by cobas and LA ( n = 1,965). Cervical histopathology was ascertained from electronic health records. Hierarchical Onclarity channels showed a significant linear association with histological severity. Onclarity and cobas had excellent agreement on partial typing of HPV16, HPV18, and the other 12 types as a pool (sample-weighted kappa value of 0.83); cobas was slightly more sensitive for HPV18 and slightly less sensitive for the pooled high-risk types. Typing by Onclarity showed excellent agreement with types and groups of types identified by LA (kappa values from 0.80 for HPV39/68/35 to 0.97 for HPV16). Onclarity typing results corresponded well to histopathology and to an already validated HPV DNA test and could provide additional clinical typing if such discrimination is determined to be clinically desirable.

Current U.S. cervical cancer screening and management guidelines do not consider previous screening history, because data on multiple-round human papillomavirus (HPV) and cytology "co-testing" have been unavailable.To measure cervical cancer risk in routine practice after successive negative screening co-tests at 3-year intervals.Observational cohort study.Integrated health care system (Kaiser Permanente Northern California, Oakland, California).990 013 women who had 1 or more co-tests from 2003 to 2014.3- and 5-year cumulative detection of (risk for) cervical intraepithelial neoplasia grade 3, adenocarcinoma in situ, and cervical cancer (≥CIN3) in women with different numbers of negative co-tests, overall and within subgroups defined by previous co-test results or baseline age.Five-year ≥CIN3 risks decreased after each successive negative co-test screening round (0.098%, 0.052%, and 0.035%). Five-year ≥CIN3 risks for an HPV-negative co-test, regardless of the cytology result, nearly matched the performance (reassurance) of a negative co-test for each successive round of screening (0.114%, 0.061%, and 0.041%). By comparison, ≥CIN3 risks for the cytology-negative co-test, regardless of the HPV result, also decreased with each successive round, but 3-year risks were as high as 5-year risks after an HPV-negative co-test (0.199%, 0.065%, and 0.043%). No interval cervical cancer cases were diagnosed after the second negative co-test. Independently, ≥CIN3 risks decreased with age. Length of previous screening interval did not influence future ≥CIN3 risks.Interval-censored observational data.After 1 or more negative cervical co-tests (or HPV tests), longer screening intervals (every 5 years or more) might be feasible and safe.National Cancer Institute Intramural Research Program.

Summary New cervical cancer screening guidelines in the USA and many European countries recommend that women are tested for human papilloma virus (HPV). To inform decisions about screening intervals, we calculate the increase in precancer or cancer risk per year of continued HPV infection. However, both time to onset of precancer or cancer and time to HPV clearance are interval censored, and onset of precancer or cancer strongly informatively censors HPV clearance. We analyse these bivariate informatively interval-censored data by developing a novel joint model for time to clearance of HPV and time to precancer or cancer by using shared random effects, where the estimated mean duration of each woman's HPV infection is a covariate in the submodel for time to precancer or cancer. The model was fitted to data on 9553 HPV positive and negative women undergoing cervical cancer screening at Kaiser Permanente Northern California: data that were pivotal to the development of US screening guidelines. We compare the implications for screening intervals of this joint model with those from population-average marginal models of precancer or cancer risk. In particular, after 2 years the marginal population-average precancer or cancer risk was 5%, suggesting a 2-year interval to control population-average risk at 5%. In contrast, the joint model reveals that almost all women exceeding 5% individual risk in 2 years also exceeded 5% in 1 year, suggesting that a 1-year interval is better to control individual risk at 5%. The example suggests that sophisticated risk models that can predict individual risk may have implications that are different from those of population-average risk models that are currently used for informing medical guideline development.

Purpose: Human papillomavirus (HPV) DNA methylation testing is a promising triage option for women testing HPV positive during cervical cancer screening. However, the extent to which methylation indicates precancer for all 12 carcinogenic HPV types has not been evaluated. Experimental Design: In this nested case–control study, we tested up to 30 cases of precancer [cervical intraepithelial neoplasia grade 3 (CIN3)/adenocarcinoma in situ (AIS)] and 30 normal controls for each carcinogenic type (single infections with 16/18/31/33/35/39/45/51/52/56/58/59). Next-generation bisulfite sequencing was performed on CpG sites within the L1 and L2 genes. We calculated differences in methylation, ORs, and AUC. Using a fixed sensitivity of 80%, we evaluated the specificity and the risk of CIN3/AIS for best performing CpG sites, and compared the performance of an explorative multi-type methylation assay with current triage strategies. Results: Methylation was positively associated with CIN3/AIS across all 12 types. AUCs for the top sites ranged from 0.71 (HPV51 and HPV56) to 0.86 (HPV18). A combined 12-type methylation assay had the highest Youden index (0.46), compared with cytology (0.31) and a 5-type methylation assay, including only previously described types (0.26). The 12-type methylation assay had higher sensitivity (80% vs. 76.6%) and lower test positivity compared with cytology (38.5% vs. 48.7%). The risk of CIN3/AIS was highest for methylation positives and lowest for cytology or HPV16/18 positives. Conclusions: HPV DNA methylation is a general phenomenon marking the transition from HPV infection to precancer for all 12 carcinogenic types. Development of a combined multitype methylation assay may serve as a triage test for HPV-positive women. Clin Cancer Res; 24(9); 2194–202. ©2018 AACR.

Cervical intraepithelial neoplasia grade 2 (CIN2) frequently regresses, is typically slow-growing, and rarely progresses to cancer. Some women forgo immediate treatment, opting for conservative management (heightened surveillance with cytology and colposcopy), to minimize overtreatment and increased risk of obstetric complications; however, there are limited data examining clinical outcomes in these women. We performed a retrospective cohort analysis of younger women diagnosed with initially untreated CIN1/2, CIN2 and CIN2/3 lesions at Kaiser Permanente Northern California between 2003 and 2015. Clinical outcomes were categorized into five mutually exclusive hierarchical groups: cancer, treated, returned to routine screening, persistent high-grade lesion, or persistent low-grade lesion. Median follow-up for the 2,417 women was 48 months. Six women were diagnosed with cancer (0.2%), all with history of high-grade cytology, and none after a negative cotest. Thirty percent of women were treated, and only 20% returned to routine screening; 50% remained in continued intensive follow-up, of which 86% had either low-grade cytology/histology or high-risk human papillomavirus (HPV) positivity, but not necessarily persistence of a single HPV type. No cancers were detected after a single negative cotest in follow-up. Almost half of initially untreated women did not undergo treatment, but remained by protocol in colposcopy clinic for 2 or more years in the absence of persisting CIN2+. Their incomplete return to total negativity was possibly due to sequential new and unrelated low-grade abnormalities. The prolonged colposcopic surveillance currently required to return to routine screening in the absence of persisting CIN2+ might not be necessary after a negative cotest. Significance: Many younger women under conservative management following an initial CIN2 result remain in a clinical protocol of prolonged intensified surveillance without a subsequent diagnosis of CIN2 or more severe diagnoses. More research is needed to determine whether such prolonged management might be unnecessary following a negative cotest for those women with an initial CIN2 but otherwise only low-grade findings. Cancer Prev Res; 11(3); 165–70. ©2018 AACR.

The goal of cervical screening is to detect and treat precancers before some become cancer. We wanted to understand why, despite state-of-the-art methods, cervical cancers occured in relationship to programmatic performance at Kaiser Permanente Northern California (KPNC), where1,000,000 women aged ≥30years have undergone cervical cancer screening by triennial HPV and cytology cotesting since 2003.We reviewed clinical histories preceding cervical cancer diagnoses to assign "causes" of cancer. We calculated surrogate measures of programmatic effectiveness (precancers/(precancers and cancers)) and diagnostic yield (precancers and cancers per 1000 cotests), overall and by age at cotest (30-39, 40-49, and ≥50years).Cancer was rare and found mainly in a localized (treatable) stage. Of 623 cervical cancers with at least one preceding or concurrent cotest, 360 (57.8%) were judged to be prevalent (diagnosed at a localized stage within one year or regional/distant stage within two years of the first cotest). Non-compliance with recommended screening and management preceded 9.0% of all cancers. False-negative cotests/sampling errors (HPV and cytology negative), false-negative histologic diagnoses, and treatment failures preceded 11.2%, 9.0%, and 4.3%, respectively, of all cancers. There was significant heterogeneity in the causes of cancer by histologic category (p0.001 for all; p=0.002 excluding prevalent cases). Programmatic effectiveness (95.3%) and diagnostic yield were greater for squamous cell versus adenocarcinoma histology (p0.0001) and both decreased with older ages (pA state-of-the-art intensive screening program results in very few cervical cancers, most of which are detected early by screening. Screening may become less efficient at older ages.

Inexpensive and easy-to-perform human papillomavirus (HPV) tests are needed for primary cervical cancer screening in lower-resource regions. In a convenience sample of 516 residual exfoliative cervical specimens from the Kaiser Permanente Northern California and U.S. National Cancer Institute Persistence and Progression Study, we assessed the agreement and clinical performance of a simple, inexpensive real-time PCR assay for the detection of 13 carcinogenic HPV types (the H13 assay; Hybribio, Hong Kong) that is marketed in limited-resource settings compared to previous testing by the Hybrid Capture 2 assay (HC2; Qiagen, Germantown, MD) and the Onclarity assay (BD Diagnostics, Sparks, MD). The test set was chosen to include many HPV-positive specimens. The reference standard was a combination of HC2 and Onclarity results for HPV detection and histologic diagnosis of controls (less than cervical intraepithelial neoplasia grade 2 [P < 0.001) and P < 0.001). In conclusion, H13 corresponds well to the combination of HC2 and Onclarity and has good clinical accuracy compared to histologic diagnosis, with less cross-reactivity with untargeted HPV types than HC2. H13 is a lower-cost HPV DNA test that might be useful for primary screening in limited-resource settings.

It is unclear whether a woman's age influences her risk of cervical intraepithelial neoplasia grade 3 or worse (CIN3+) upon detection of HPV. A large change in risk as women age would influence vaccination and screening policies. Among 972,029 women age 30-64 undergoing screening with Pap and HPV testing (Hybrid Capture 2, Qiagen, Germantown, MD) at Kaiser Permanente Northern California (KPNC), we calculated age-specific 5-year CIN3+ risks among women with HPV infections detected at enrollment, and among women with "newly detected" HPV infections at their second screening visit. Women (57,899, 6.0%) had an enrollment HPV infection. Among the women testing HPV negative at enrollment with a second screening visit, 16,724 (3.3%) had a newly detected HPV infection at their second visit. Both enrollment and newly detected HPV rates declined with age (p < 0.001). Women with enrollment versus newly detected HPV infection had higher 5-year CIN3+ risks: 8.5% versus 3.9%, (p < 0.0001). Risks did not increase with age but declined slightly from 30-34 years to 60-64 years: 9.4% versus 7.4% (p = 0.017) for enrollment HPV and 5.1% versus 3.5% (p = 0.014) for newly detected HPV. Among women age 30-64 in an established screening program, women with newly detected HPV infections were at lower risk than women with enrollment infections, suggesting reduced benefit vaccinating women at older ages. Although the rates of HPV infection declined dramatically with age, the subsequent CIN3+ risks associated with HPV infection declined only slightly. The CIN3+ risks among older women are sufficiently elevated to warrant continued screening through age 65.

BACKGROUND Dual-stain cytology for p16 and Ki-67 has been proposed as a biomarker in cervical cancer screening. The authors evaluated the reproducibility and accuracy of dual-stain cytology among 10 newly trained evaluators. METHODS In total, 480 p16/Ki-67–stained slides from human papillomavirus-positive women were evaluated in masked fashion by 10 evaluators. None of the evaluators had previous experience with p16 or p16/Ki-67 cytology. All participants underwent p16/Ki-67 training and subsequent proficiency testing. Reproducibility of dual-stain cytology was measured using the percentage agreement, individual and aggregate κ values, as well as McNemar statistics. Clinical performance for the detection of cervical intraepithelial neoplasia grade 2 or greater (CIN2+) was evaluated for each individual evaluator and for all evaluators combined compared with the reference evaluation by a cytotechnologist who had extensive experience with dual-stain cytology. RESULTS The percentage agreement of individual evaluators with the reference evaluation ranged from 83% to 91%, and the κ values ranged from 0.65 to 0.81. The combined κ value was 0.71 for all evaluators and 0.73 for cytotechnologists. The average sensitivity and specificity for the detection of CIN2+ among novice evaluators was 82% and 64%, respectively; whereas the reference evaluation had 84% sensitivity and 63% specificity, respectively. Agreement on dual-stain positivity increased with greater numbers of p16/Ki-67–positive cells on the slides. CONCLUSIONS Good to excellent reproducibility of p16/Ki-67 dual-stain cytology was observed with almost identical clinical performance of novice evaluators compared with reference evaluations. The current findings suggest that p16/Ki-67 dual-stain evaluation can be implemented in routine cytology practice with limited training. Cancer (Cancer Cytopathol) 2014;122:914–920. © 2014 American Cancer Society.

BACKGROUND Different US practice guidelines have conflicting recommendations for when women should return after a screening result of human papillomavirus (HPV)-negative with an equivocal Papanicolaou (Pap) result of atypical squamous cells of undetermined significance (ASC-US) (ie, return in either 3 or 5 years). One way to determine management is to compare the risk of precancer/cancer after an HPV-negative/ASC-US result with the risks after other negative screening results. For example, if the risk after an HPV-negative/ASC-US result was similar to the risk after a negative Pap test, a 3-year return would be preferred because guidelines agree that women with negative Pap test results should return in 3 years. Alternatively, if the risk after an HPV-negative/ASC-US result is similar to that after a cotest-negative result (HPV negative/Pap test negative), a 5-year return would be preferred because guidelines agree that women testing cotest negative should return in 5 years. METHODS The authors compared risks of cervical intraepithelial neoplasia of grade 3 or higher (CIN3+) and cervical cancer among women aged 30 years to 64 years at Kaiser Permanente Northern California with the following test results from 2003 through 2012: 17,191 women testing HPV negative/ASC-US; 980,268 women testing Pap test negative (regardless of HPV result); and 892,882 women testing cotest negative. RESULTS The 5-year CIN3+ and cancer risks after an HPV-negative/ASC-US result were closer to the risks after a negative Pap test result (CIN3+: 0.48% vs 0.31% [P =.0019]; and cancer: 0.043% vs 0.031% [P =.4]) than after a negative cotest (CIN3+: 0.48% vs 0.11% [P

OBJECTIVE Kaiser Permanente Northern California (KPNC) introduced 3-year Pap and human papillomavirus DNA cotesting for cervical cancer screening in women 30 years or older in 2003 to 2004. Patient and provider willingness to extend screening intervals and the impact on annual cervical cancer incidence after interval extension are evaluated. MATERIALS AND METHODS Age-adjusted cervical cancer rates and screening intervals were calculated from KPNC Regional Laboratory databases and Northern California Cancer Registry from 2000 to 2009. RESULTS The median screening interval between negative cotests was 36 months compared to the 16 months after a negative Pap test alone before the implementation of cotesting. The age-adjusted invasive cancer rate was 6.5 per 100,000 women in 2000 and 6.3 in 2009; there was no difference in the rates of cervical cancer in women 30 years or older from 2000 to 2009 (p(trend) = .7). CONCLUSIONS Patients and providers were compliant with the extension of screening intervals with cotesting. Cervical cancer rates remained constant during the 10-year study period despite extending screening intervals after a negative cotest.

New screening guidelines recommend that human papillomavirus (HPV)-negative/atypical squamous cells of undetermined significance (ASC-US) results be considered as equivalent to HPV-negative/Pap-negative results, leading to rescreening in 5 years. However, despite ample data, the routine clinical performance of HPV testing of women with ASC-US has not been adequately documented.We estimated 5-year risks of cervical intraepithelial neoplasia (CIN) 3+ and of cancer among 2 groups of women between 2003 and 2010 at Kaiser Permanente Northern California: 27,050 aged 30 to 64 years who underwent HPV and Pap cotesting and had an ASC-US Pap result and 12,209 aged 25 to 29 years who underwent HPV triage of ASC-US.Five-year risks of CIN 3+ and of cancer among women aged 30 to 64 years testing HPV-negative/ASC-US and among 923,152 women testing Pap-negative alone were similar although statistically distinguishable (CIN 3+, 0.43% vs 0.26%, p = .001; cancer, 0.050% vs 0.025%, p = .1). The increased risk of cancer after HPV-negative/ASC-US versus Pap-negative alone was confined to women aged 60 to 64 years (0.26% vs 0.035%, p = .3). Five-year risks of CIN 3+ and cancer among women with HPV-negative/ASC-US results were substantially higher than those among women testing HPV-negative/Pap-negative (CIN 3+, 0.43% vs 0.08%, p.0001; cancer, 0.050% vs 0.011%, p = .003). For women aged 30 to 64 years testing HPV-positive/ASC-US, 5-year risks of CIN 3+ and cancer were slightly higher than those among 9,374 women with low-grade squamous intraepithelial lesion (LSIL) (CIN 3+, 6.8% vs 5.2%, p = .0007; cancer, 0.41% vs 0.16%, p = .04). Similar patterns were seen for women aged 25 to 29 years.Women with HPV-negative/ASC-US had a similar risk as women testing Pap-negative alone but had a higher risk than women testing HPV-negative/Pap-negative. Based upon the principle of "equal management of equal risks," our findings support the equal management of women with HPV-negative/ASC-US and those with Pap-negative alone, except for exiting women from screening because cancer risks at ages 60 to 64 years may be higher for women testing HPV-negative/ASC-US. Our findings also support managing HPV-positive/ASC-US and LSIL similarly.

Current US guidelines for cotesting recommend that the large numbers of women who test Pap-negative, but human papillomavirus (HPV)-positive, return in 1 year, and those who remain HPV-positive or have low-grade squamous intraepithelial lesion (LSIL) or worse Pap results be referred for colposcopy. However, the performance of these guidelines in routine clinical practice has not been evaluated.We estimated cumulative 5-year risks of cervical intraepithelial neoplasia grade 3 or worse (CIN 3+) among 32,374 women aged 30 to 64 years with HPV-positive/Pap-negative cotest results at Kaiser Permanente Northern California during 2003 to 2010.The 5-year CIN 3+ risk after an HPV-positive/Pap-negative cotest result, which was found in 3.6% of women, was 4.5% (95% confidence interval [CI] = 4.2%-4.8%). The 5-year cancer risk was 0.34% (95% CI = 0.26%-0.45%), and half of the cases were adenocarcinoma. Overall, 48% of the women remained HPV-positive on return (median = 418 days after baseline), a percentage that varied little over ages 30 to 64 years. At the return after a baseline HPV-positive/Pap-negative result, almost every repeat cotest result predicted greater subsequent 5-year CIN 3+ risk than the same cotest result had at baseline (HPV-positive/LSIL, 9.2% vs 6.1%, p = .01; HPV-positive/atypical squamous cells of undetermined significance [ASC-US], 7.9% vs 6.8%, p = .2; HPV-positive/Pap-negative, 7.4% vs 4.5%, p.0001; HPV-negative/LSIL,1.7% vs 2.0%, p = .8; HPV-negative/ASC-US, 2.9% vs 0.43%, p = .0005; HPV-negative/Pap-negative, 0.93% vs 0.08%, p.0001).Using the principle of "equal management of equal risks," women testing HPV-positive/Pap-negative had a subsequent CIN 3+ risk consistent with risk thresholds for a 1-year return. However, on returning in approximately 1 year, about one-half of women will be referred for colposcopy because of continued HPV positivity or Pap abnormality. Clinicians should keep in mind that cotest results at the return after a baseline HPV-positive/Pap-negative finding are riskier than the same baseline cotest results in the general population, supporting intensified clinical management at return testing.

OBJECTIVE Current US national guidelines recommend beginning screening at age 21 using Pap tests only, with cotesting starting at age 30. To inform the management of Pap test abnormalities among women aged 21 to 24 years, who have extremely low cancer risks, we compared risks of CIN 3+ among women aged 21 to 24 versus 25 to 29 years or 30 to 64 years. METHODS We estimated 5-year risks of CIN 3+ given different Pap test results, with human papillomavirus (HPV) triage of atypical squamous cells of undetermined significance (ASC-US), among 133,947 women aged 21 to 24 years, compared with 135,382 women aged 25 to 29 years and 965,360 women aged 30 to 64 years, between 2003 and 2010 at Kaiser Permanente Northern California. RESULTS There were 3 cancers diagnosed during follow-up in women aged 21 to 24 years. After high-grade Pap results (0.6% of Pap results), the 5-year CIN 3+ risks among women aged 21 to 24 years were comparable to those aged 25 to 29 and 30 to 64 years (atypical glandular cells, 6.9% vs 14% vs 8.5%, p = .8; atypical squamous cells cannot rule out high-grade squamous intraepithelial lesion, 16% vs 24% vs 18%, p = .8; high-grade squamous intraepithelial lesion, 28% vs 28% vs 47%, p = .4). After low-grade squamous intraepithelial lesion, the 5-year CIN 3+ risk was lower among women aged 21 to 24 years (3.0%) than that among women aged 25 to 29 years (5.0%, p = .01) or aged 30 to 64 years (5.2%, p = .0002). Although the 5-year CIN 3+ risk after HPV-negative/ASC-US was similar across all 3 groups (0.57% vs 0.59% vs 0.43%, p = 1), risk after HPV-positive/ASC-US was lower among women aged 21 to 24 years (4.4%) than that among women aged 25 to 29 years (7.1%, p < .0001) or 30 to 64 years (6.8%, p < .0001). CONCLUSIONS Women aged 21 to 24 years had almost zero cancer risk, and positive Pap test results predicted low CIN 3+ risk except for the 0.6% of women with high-grade Pap results. The generally low risk supports conservative management of women aged 21 to 24 years.

Low-grade squamous intraepithelial lesion (LSIL) Pap results do not typically lead to human papillomavirus (HPV) testing. HPV triage is not cost-effective because most cases are HPV-positive. However, under new national guidelines recommending cotesting for women aged 30 to 64 years, clinicians will increasingly receive the HPV test result with LSIL Pap results. Some authors have suggested that HPV triage might be effective at older ages, when the percentage of HPV positivity among women with LSIL declines.We estimated 5-year risks of CIN 2+ and CIN 3+ among 9,033 women aged 30 to 64 years who had both an HPV test and an LSIL Pap result.HPV positivity among women with LSIL decreased only slightly with age (30 to 34 vs 60 to 64 years, 88% vs 72%, p.0001). The 5-year risks of CIN 2+ and CIN 3+ of women aged 30 to 64 years testing HPV-positive/LSIL were larger than those among women testing HPV-negative/LSIL (CIN 2+, 19% vs 5.1%, p.0001; CIN 3+, 6.1% vs 2.0%, p.0001). The 5-year risk of CIN 3+ in HPV-negative/LSIL women was similar to that for women with atypical squamous cells of undetermined significance (ASC-US) Pap test result without knowledge of HPV test results (2.0% vs 2.6%, p = .4).HPV-negative/LSIL posed lower risk than other Pap results that guidelines currently recommend for referral to immediate colposcopy. By the principle of "equal management of equal risks," women with HPV-negative/LSIL might reasonably be managed similarly to those with ASC-US Pap results without knowledge of HPV testing, that is, retesting at 6 to 12 months, rather than immediate colposcopy. Although the HPV test result for LSIL Pap results provides actionable information to clinicians who screen with cotesting, the high HPV positivity of LSIL at even the oldest ages suggests the lack of cost-effectiveness of HPV triage of LSIL for clinicians who do not use routine cotesting.

Two human papillomavirus (HPV) vaccines are available to prevent cervical cancer. One early measure of HPV vaccine impact would be a reduction in vaccine-related HPV types (HPV 6, 11, 16, or 18, or HPV 16, 18) in cervical samples from young women. We aimed to assess feasibility of specimen collection and baseline HPV prevalence in an integrated healthcare delivery system. Residual cervical specimens collected during routine cervical cancer screening (2006–2008) were retained consecutively from eligible females aged 11–29 years, stratified by age group. Specimens were evaluated for 37 HPV genotypes using the Roche Linear Array assay. Of 10,124 specimens submitted, 10,103 (99 %) were adequate for HPV testing. Prevalence of HPV 6, 11, 16, or 18 genotype was 11.4 % overall and was the highest in the youngest age group (18.1 % in the 11–19-year-olds, 12.5 % in the 20–24-year-olds, and 7.0 % in the 25–29-year-olds). HPV types 6, 11, 16, or 18 prevalence could be measured over time to assess early HPV vaccine impact using residual specimens from an integrated healthcare delivery system, particularly if sampling focused on young women.

HPV testing is more sensitive than cytology for cervical screening. However, to incorporate HPV tests into screening, risk-stratification ("triage") of HPV-positive women is needed to avoid excessive colposcopy and overtreatment. We prospectively evaluated combinations of partial HPV typing (Onclarity, BD) and cytology triage, and explored whether management could be simplified, based on grouping combinations yielding similar 3-year or 18-month CIN3+ risks. We typed ∼9,000 archived specimens, taken at enrollment (2007-2011) into the NCI-Kaiser Permanente Northern California (KPNC) HPV Persistence and Progression (PaP) cohort. Stratified sampling, with reweighting in the statistical analysis, permitted risk estimation of HPV/cytology combinations for the 700,000+-woman KPNC screening population. Based on 3-year CIN3+ risks, Onclarity results could be combined into five groups (HPV16, else HPV18/45, else HPV31/33/58/52, else HPV51/35/39/68/56/66/68, else HPV negative); cytology results fell into three risk groups ("high-grade," ASC-US/LSIL, NILM). For the resultant 15 HPV group-cytology combinations, 3-year CIN3+ risks ranged 1,000-fold from 60.6% to 0.06%. To guide management, we compared the risks to established "benchmark" risk/management thresholds in this same population (e.g., LSIL predicted 3-year CIN3+ risk of 5.8% in the screening population, providing the benchmark for colposcopic referral). By benchmarking to 3-year risk thresholds (supplemented by 18-month estimates), the widely varying risk strata could be condensed into four action bands (very high risk of CIN3+ mandating consideration of cone biopsy if colposcopy did not find precancer; moderate risk justifying colposcopy; low risk managed by intensified follow-up to permit HPV "clearance"; and very low risk permitting routine screening.) Overall, the results support primary HPV testing, with management of HPV-positive women using partial HPV typing and cytology.

Estimates of absolute risks and risk differences are necessary for evaluating the clinical and population impact of biomedical research findings. We have developed a linear-expit regression model (LEXPIT) to incorporate linear and nonlinear risk effects to estimate absolute risk from studies of a binary outcome. The LEXPIT is a generalization of both the binomial linear and logistic regression models. The coefficients of the LEXPIT linear terms estimate adjusted risk differences, while the exponentiated nonlinear terms estimate residual odds ratios. The LEXPIT could be particularly useful for epidemiological studies of risk association, where adjustment for multiple confounding variables is common. We present a constrained maximum likelihood estimation algorithm that ensures the feasibility of risk estimates of the LEXPIT model and describe procedures for defining the feasible region of the parameter space, judging convergence, and evaluating boundary cases. Simulations demonstrate that the methodology is computationally robust and yields feasible, consistent estimators. We applied the LEXPIT model to estimate the absolute five-year risk of cervical precancer or cancer associated with different Pap and human papillomavirus test results in 167,171 women undergoing screening at Kaiser Permanente Northern California. The LEXPIT model found an increased risk due to abnormal Pap test in HPV-negative that was not detected with logistic regression. Our R package blm provides free and easy-to-use software for fitting the LEXPIT model.

Objective To characterize the antecedent screening of women 65years of age and older diagnosed with cervical cancer. Methods Screening histories of women 65years of age and older who were diagnosed with cervical cancer between 2003 and 2008 were examined utilizing the organization's databases and the regional Cancer Registry. Stopping screening was recommended at age 65 for members who had either 3 consecutive negative Paps or a single negative Pap plus HPV test ("cotest"). Results From 2003 through 2008 there were 56 Kaiser Permanente Northern California members 65years of age and older diagnosed with cervical cancer. During the same time period there were 1,323,100 woman-years of membership in women age 65 and older. The risk of invasive cancer among women age 65 and older was 4.2/100,000/year in 2003–2008. 33 of 56 (59%) had one or more Pap smears prior to diagnosis. Of the 33, 14 women (25%) had 3 consecutive negative Pap smears prior to diagnosis. Three of 46,401 women with 1 or more negative cotests at age 65 and older were subsequently diagnosed with invasive cancer during 132,639 women-years of follow-up (2.3/100,000/year). Conclusions Most cervical cancers diagnosed at age 65 and older occur in women who have not met our criteria for stopping screening. A few cancers will continue to occur at age 65 and older despite multiple negative tests, as is true in other age groups. We currently have no evidence that these cancers would be prevented with continued screening at ages 65 and older.

BACKGROUND: The majority of anal cancers are caused by persistent infections with carcinogenic human papillomaviruses (HPV). Similar to cervical carcinogenesis, the progression from HPV infection to anal cancer occurs through precancerous lesions that can be treated to prevent invasion. In analogy to cervical cytology, anal cytology has been proposed as a screening tool for anal cancer precursors in high-risk populations. METHODS: The authors analyzed the interobserver reproducibility of anal cytology in a population of 363 human immunodeficiency virus (HIV)-infected men who have sex with men (MSM). Liquid-based cytology (LBC) specimens were collected in the anal dysplasia clinic before the performance of high-resolution anoscopy on all patients. Papanicolaou-stained LBC slides were evaluated by 2 cytopathologists, each of whom was blinded to the clinical outcome and the other pathologist's results, using the revised Bethesda terminology. RESULTS: Overall agreement between the 2 observers was 66% (kappa, 0.54; linear-weighted kappa, 0.69). Using dichotomizing cytology results (atypical squamous cells of undetermined significance [ASC-US] or worse vs less than ASC-US), the agreement increased to 86% (kappa, 0.69). An increasing likelihood of testing positive for markers associated with HPV-related transformation, p16/Ki-67, and HPV oncogene messenger RNA was observed, with increasing severity of cytology results noted both for individual cytologists and for consensus cytology interpretation (P value for trend [ptrend] < .0001 for all). CONCLUSIONS: Moderate to good agreement was observed between 2 cytopathologists evaluating anal cytology samples collected from HIV-positive MSM. A higher severity of anal cytology was associated with biomarkers of anal precancerous lesions. Anal cytology may be used for anal cancer screening in high-risk populations, and biomarkers of HPV-related transformation can serve as quality control for anal cytology. Cancer (Cancer Cytopathol) 2013. Published 2012 by the American Cancer Society.

The cobas human papillomavirus (HPV) test (cobas) was recently approved by the U.S. Food and Drug Administration (FDA) and identifies HPV16 and HPV18 separately as well as detecting a pool of 11 HR-HPV genotypes (HPV31, -33, -35, -39, -45, -51, -52, -56, -58, -59, -68) and also HPV66. We compared cobas, Linear Array (LA), and Hybrid Capture 2 (HC2) assays for detection of carcinogenic HPV DNA, and cobas and LA for detection of HPV16 and HPV18 DNA, among the first 1,852 women enrolled in the HPV Persistence and Progression Cohort (PaP Cohort) study. Specimens were tested by all 3 assays 1 year after an HC2-positive result. In 1,824 specimens with cobas results, cobas had an 85.9% agreement with HC2 and 91.0% agreement with LA for carcinogenic HPV detection. When results between cobas and HC2 disagreed, cobas tended to call more women HPV positive ( P < 0.01). Categorizing cobas and LA results hierarchically according to cancer risk (HPV16, HPV18, other carcinogenic HPV genotypes, or carcinogen negative), there was a 90% agreement for all categories of HPV ( n = 1,824). We found good agreement between the two U.S. FDA-approved HPV tests, with discrepancies between the two assays due to specific characteristics of the individual assays. Additional studies are needed to compare HC2 and cobas for detecting and predicting CIN3 to understand the clinical implications of the discrepant test results between the two tests.

To characterize the cervical cancers diagnosed following a Pap-negative, high risk human papillomavirus (HPV)-positive (Pap-/HPV+) screen in routine clinical practice.Using data from Kaiser Permanente Northern California, we investigated the cases of cervical cancer diagnosed between January, 2003 and January, 2009 following Pap-/HPV+ screen. Two cervical specimens were routinely collected for cervical cancer screening, one for conventional cytology and the other for high risk HPV testing using Hybrid Capture 2 (Qiagen).Forty-four women (median age at diagnosis=44years) were diagnosed with primary invasive cervical cancer with a recent history of one or more Pap-/HPV+ screens. Twenty-six women had one Pap-/HPV+ screen preceding the diagnosis of cancer, 15 had two, and three had three. There were 16 squamous cancers, one small cell cancer, 24 adenocarcinomas, 2 adenosquamous carcinomas, and one case with separate invasive squamous and adenocarcinoma. FIGO Stage was IA in 11 women, IB in 31 women and IIA in 2 women. Treatment included a pelvic node dissection in 30, 2 (6.7%) of whom had positive nodes.HPV testing contributes to early cervical cancer diagnosis detection in women with negative Pap tests. Most women in this cohort have early stage, node negative, treatable and potentially curable disease. Adenocarcinoma predominated as might be expected because cytology misses these cancers and their precursors. The majority of cancers were diagnosed following a single Pap-/HPV+ screen, suggesting that effective triage to colposcopy of women with a Pap-/HPV+ screen would be preferable to retesting in one year as currently recommended.

Background: Human papillomavirus (HPV) testing has been approved as a primary strategy for cervical cancer screening, either alone or in combination with cytology (co-testing), based on its high sensitivity and long-term reassurance against cervical precancer following a negative test result. However, nearly twice as many women will screen positive for HPV compared with cytology-based screening. Thus, effective management of HPV-positive women requires triage markers to distinguish those at high-risk who should be referred to colposcopy from those with benign infections who can safely return to routine screening. p16/Ki-67 dual stain cytology has previously shown good risk stratification for triage of HPV-positive women; however, studies with follow-up extending beyond 3 years are lacking. We evaluated the long-term risk prediction of p16/Ki-67 for detection of cervical precancer (cervical intraepithelial neoplasia grade 3 or worse, CIN3+) in a large population of HPV-positive women. Methods: 1,588 HPV-positive women screened with HPV/cytology co-testing were enrolled in 2012 at Kaiser Permanente Northern California. p16/Ki-67 cytology was performed on residual Surepath material and slides were evaluated for p16/Ki-67 positivity. Cervical histology endpoints were ascertained from the clinical database with follow-up through 2017. We conducted a Kaplan Meier analysis to estimate risk of CIN3+ by p16/Ki-67 and cytology (atypical squamous cells of undetermined significance or worse, ASC-US+, versus normal cytology). Risks were compared to internal benchmarks for colposcopy referral and for a one year return interval. Results: In women testing p16/Ki-67 positive at baseline, the 2-year risk of CIN3+ was 14.3%, compared with 2.2% in p16/Ki-67-negative women. For ASC-US+, the risk was 12.6% and 2.9% for normal cytology. The 5-year risk of CIN3+ in p16/Ki-67-positive women was 21.6% and 5.0% in p16/Ki-67-negatives. The 5-year risk of ASC-US+ was 17.1% compared to 8.2% for normal cytology. Among p16/Ki-67-negatives, the risk remained below the colposcopy referral threshold for 5 years while in women with normal cytology, the colposcopy referral threshold was crossed after year 3. Conclusion: In the first study evaluating long-term risk stratification of p16/Ki-67 dual staining, p16/Ki-67- negativity provided strong reassurance against CIN3+ for at least five years. In contrast, the risk in women with normal cytology crossed the colposcopy referral threshold after three years. These data support use of p16/Ki-67 for triage of HPV-positive women with the possibility of extending surveillance intervals in p16/Ki-67-negative women. Citation Format: Megan A. Clarke, Barbara Fetterman, Mark Schiffman, Philip E. Castle, Eric Stiemerling, Diane Tokugawa, Nancy Poitras, Walter Kinney, Thomas Lorey, Nicolas Wentzensen. Long term risk prediction of p16/Ki-67 dual stain in triage of HPV-positive women [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2202.

Objective. To characterize the risks of cervical intraepithelial neoplasia 3 (CIN 3) and cancer in women aged 21 to 24 with human papillomavirus (HPV)-positive atypical squamous cells of undetermined significance (ASC-US) and low-grade squamous intraepithelial lesion (LSIL) screening results in routine clinical practice. Materials and Methods. Quality assurance databases containing records of screening test and histologic findings from the Regional Laboratory of the Northern California Kaiser Permanente Medical Care Program were reviewed. Numbers of LSIL and HPV-positive ASC-US results and associated cancers and CIN 3 in women aged 21 to 24 during 2003 to 2007 were tabulated, and the corresponding risks were calculated overall and by year of age. Results. During the 5-year period from 2003 to 2007, 1,620 HPV-positive ASC-US and 2,175 LSIL were diagnosed in women aged 21 to 24, for which corresponding histologic finding is available. No invasive cancers were detected in association with LSIL and HPV-positive ASC-US screening results in this age group during this period. The risk of cancer was therefore 0% (95% CI = 0.00%-0.10%). The risk of CIN 3 associated with an HPV-positive ASC-US was 2.90% (95% CI = 2.14%-3.84%), with LSIL was 2.44% (95% CI = 1.83%-3.18%), and, for the 2 combined, the risk was 2.64% (95% CI = 2.15%-3.20%). Conclusions. The risk of CIN 3 and cancer is low enough that management of women aged 21 to 24 with ASC-US and LSIL smears without immediate colposcopy should be considered, as is currently recommended for women aged 20 and younger.

Cervical infections by approximately 15 cancer-associated (carcinogenic) human papillomavirus (HPV) genotypes cause virtually all cervical cancer and its immediate precursor lesions worldwide. Prophylactic vaccines against human papillomavirus (HPV) types HPV16 and HP18, which cause 70% of cervical cancer worldwide, hold great promise for reducing the burden of cervical cancer worldwide. However, current HPV vaccines prevent future infections and related cervical abnormalities and do not treat pre-existing HPV infections. In the U.S., HPV vaccine introduction should be considered in the context of a very successful cervical cancer screening program that has reduced the rates of cervical cancer by 75% or more. Thus, HPV vaccines will only prevent an incremental number of additional cervical cancers in the U.S. The introduction of HPV vaccines can also prevent other HPV-related sequelae, most importantly cervical intraepithelial neoplasia grade 2 or 3 (CIN2/3), which precede the development of cervical cancer and require clinical follow-up and treatment. Examining data from 7 clinical centers in the U.S., the median age of CIN2/3 is typically between 25 and 30 years of age in 2007; if screen-detected CIN2/3 develops on average 5-10 years after the causal infection is acquired, HPV vaccination will only prevent a significant proportion of CIN2/3 if it is given to women before the age of 26 and more so if given to women 18 and younger. It is increasingly evident that prophylactic HPV vaccines will provide the greatest public health or population benefit only when delivered to adolescent, mostly HPV-naive women.

To estimate the 5-year age group-specific test positives for Pap tests and human papillomavirus (HPV) testing in a large, general screening population of women 30 and older.Using data from Kaiser Permanente Northern California, a large health maintenance organization that introduced cotesting in 2003, we evaluated the cotesting results overall and by 5-year age groups. Women (n=580,289) who opted for and underwent cotesting (n cotests=812,598) between January 2003 and April 2008 were included in the analysis. Pap tests interpreted as atypical squamous cells of undetermined significance (ASC-US) or more severe were considered to be positive. Women were tested for carcinogenic HPV using an assay approved by the U.S. Food and Drug Administration. Binomial exact 95% confidence intervals (CIs) were calculated.Overall, 6.27% (95% CI 6.21-6.32%) of cotests were carcinogenic HPV positive, and only 3.99% (95% CI 3.94-4.03%) cotests had normal cytology and were carcinogenic HPV positive. By comparison, 5.18% (95% CI 5.13-5.23%) of cotests had ASC-US or more severe cytology, and 2.87% (95% CI 2.84-2.91%) of cotests had ASC-US or more severe cytology and were carcinogenic HPV negative.In a general screening population, concerns about excessive HPV test positives among women aged 30 years and older are not borne out.

In US cervical screening, immediate colposcopy is recommended for women with HPV-positive ASC-US (equivocal) cytology. We evaluated whether partial typing by Onclarity™ (BD) might identify HPV-positive women with low enough CIN3+ risk to permit 1-year follow-up instead.The NCI-Kaiser Permanente Northern California Persistence and Progression cohort includes a subset of 13,890 women aged 21+ with HC2 (Qiagen)-positive ASC-US at enrollment; current median follow-up is 3.0years. Using stratified random sampling, we typed 2079 archived enrollment specimens including 329 women subsequently diagnosed with CIN3+, 563 with CIN2, and 1187 withCIN2. Adjusting for sampling, we computed 3-year cumulative CIN3+ risks for each Onclarity typing channel, using Kaplan-Meier methods.The 3-year CIN3+ risk for all HC2-positive women with ASC-US was 5.2%; this establishes the "benchmark" risk for colposcopic referral. Hierarchically, 3-year cumulative risks for each typing channel were 16.0% for HPV16, 7.4% for HPV18, 7.0% for HPV31, 7.1% for grouped HPV33/58, 4.3% for HPV52, 3.9% for HPV45, 2.7% for HPV51, 1.6% for HPV39/68/35, and 1.3% for HPV59/56/66.ASC-US linked to HPV16, HPV18, HPV31, or HPV33/58 warrants immediate colposcopy. Optimal management of women with HPV52 or HPV45 is uncertain. Risk of women with only HPV51, HPV39/68/35, or HPV59/56/66 might be low enough to recommend 1-year retesting permitting viral clearance. This strategy would defer colposcopy for 40% of women with HPV-positive ASC-US, half of whom would be cotest-negative at 1-year return. Approximately 10% of those with CIN3 diagnosable at enrollment would be delayed 1year instead. Cost-effectiveness analyses are needed.

Infection with one or more high-risk HPV (hr-HPV) types is a necessary step in the development of cervical cancer, making its detection a useful tool for cervical cancer screening and prevention (Bosch et al., 2002). Hybrid Capture 2 (HC2, Qiagen, Gaithersburg, MD) was the first FDA-approved and is still the most widely used laboratory test for the presence of high-risk HPV DNA as a predictor of who is or is not at risk for cervical cancer and pre-cancer (defined here as cervical intraepithelial neoplasia grade 2 or worse). While HC2 has remarkable sensitivity for cervical intraepithelial neoplasia grade 2 or worse, the transient nature of most hr-HPV infections reduces its specificity as most infections are benign and will resolve without progressing to cervical intraepithelial neoplasia grade 2 or worse. This can be problematic for cervical cancer screening algorithms in which HPV positivity triggers more intensive follow-up; in such algorithms, good specificity is important for reducing the number of costly colposcopic follow-up visits (Cuzick et al., 2008). Although manufacturer specifications indicate a relative light unit (RLU) ratio compared with positive control of 1.0 as the threshold for determining positivity, reclassifying HC2 specimens that are equivocal and fall just above the positive cut-off is one potential way to improve the assay’s specificity (Rijkaart et al., 2010; Rebolj et al., 2011; Sasieni and Castanon, 2011). Some researchers have looked at increasing the RLU cut-off point because HC2 results at this level predict lower risk than higher values (Lorincz et al., 2002). However, equivocal results are not that common and raising the cut-point to 2.0 or 3.0 may only yield a modest improvement in specificity at some cost to sensitivity; i.e., misclassification of women with cervical intraepithelial neoplasia grade 2 or worse lesions whose RLU ratios are equivocal (Rijkaart et al., 2010). Rather than raising the cut-point, a more conservative approach is to repeat the HC2 test on specimens with equivocal results and determine the final test result based on a collective assessment of the test results (Knoepp et al., 2007), however this method requires additional laboratory time and cost to implement, weighing against any savings it yields from reducing the number of colposcopic examinations. In 2001 Kaiser Permanente Northern California added the HC2 assay to its cervical cancer screening program as a triage test for women with a cytologic diagnosis of atypical squamous cells of undetermined significance and, in 2003, as a concurrent test (“cotest”) with cytology for women 30 and older. In the cotesting scheme, women have been managed as follows: 1) women who are HPV-positive with a cytologic diagnosis of atypical squamous cells of undetermined significance, or have more severe (regardless of the HPV test results) are referred for immediate colposcopy; 2) women who test HPV-negative and have cytology of atypical squamous cells of undetermined significance, or who test HPV-positive with negative cytology are rescreened in one year; and 3) if both tests are negative, women are rescreened in three years. For cotests with borderline positive hr-HPV results, interpretation of the repeat test results is used to determine the final hr-HPV test result and hence the need for rescreening visits versus colposcopy. At the time that HC2 was introduced into the Kaiser Permanente Northern California cervical cancer screening algorithm, based on concern over the meaning of borderline-positive HC2-positive results, it was decided that any specimens with borderline or “equivocal” HC2 results, defined as a RLU ratio greater than or equal to 1.0 but less than 3.0, would be re-tested two additional times. The final hr-HPV result was determined by the majority results of the three tests i.e., only one of the two repeats needed to be positive to have the final positive hr-HPV result. To assess the utility of Kaiser Permanente Northern California’s re-testing scheme for equivocal HC2 specimens, HC2 data, cytology results, and histological outcomes (less than cervical intraepithelial neoplasia grade 2, cervical intraepithelial neoplasia grade 2 or worse) were compiled for patients undergoing routine cervical cancer screening at Kaiser Permanente Northern California.

Objectives: Primary HPV testing has been approved in the United States. Screening intervals can be safely extended for HPV-negative women, but the challenge lies in discriminating transient HPV infections from precancers among HPV-positives. In a large study at Kaiser Permanente Northern California (KPNC), candidate strategies for triage of HPV-positive women are being evaluated, including cytology, p16/Ki-67 dual stain (DS, CINtec PLUS) and HPV16/18 genotyping (cobas). Methods: Over 13,000 HPV-positive women participating in cervical cancer screening at KPNC were enrolled. The dual stain and HPV genotyping assays were implemented and conducted at KPNC. Baseline results for 7,124 women are available and detection of CIN3 is currently evaluated only in cytology-positive women. Results: Among 7,124 HPV-positive women, 4,107 (57.7%) were cytology-positive (ASC-US+), 3,056 (42.9%) were DS-positive, and 1,406 (19.7%) were positive for HPV16 or HPV18. Among all 3,017 HPV-positive, cytology-negative women, 911 (30.2%) were DS- positive, and 508 (16.8%) were positive for HPV16 or HPV18. Of 315 CIN3 detected so far, 280 (88.9%) were DS-positive and 176 (55.9%) were positive for HPV16 or HPV18. Major triage strategies including cytology alone, DS alone, as well as combinations of cytology and genotyping, and DS and genotyping will be presented at the meeting. Discussion: In one of the largest clinical implementation studies of triage strategies, we showed that primary HPV screening followed by DS can reduce colposcopy referral compared to HPV-cytology co-testing while achieving high sensitivity. Additional follow-up is underway to evaluate the programmatic performance of several major candidate strategies. Citation Format: Nicolas A. Wentzensen, Barbara Fetterman, Renee Bremer, Philip Castle, Diane Tokugawa, Nancy Poitras, Elizabeth Hosfield, Thomas Lorey, Mark Schiffman, Walter Kinney. Evaluation of p16/Ki-67 dual stain, cytology, and HPV16/18 genotyping for triage of HPV-positive women in a large screening population [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5296. doi:10.1158/1538-7445.AM2017-5296

Introduction: We developed risk models for cancer screening cohorts assembled from electronic health records. We applied them to a cohort of 1.4 million women undergoing cervical cancer screening with human papillomavirus (HPV) and Pap “cotesting” at Kaiser Permanente Northern California (KPNC) to develop risk estimates to inform screening guidelines. Methods: Cohorts assembled using electronic health records present 3 challenges that make it inappropriate to use Kaplan-Meier or Cox models. First, the time of disease onset for an individual is unobserved, and falls between screens (interval-censoring). Second, there is also prevalent disease (left-censoring). Third, prevalent disease is not always immediately diagnosed (e.g. in those with negative screening results), and thus some incident disease is actually missed prevalent disease. To address the challenges, we propose a “logistic-Weibull” model that is a logistic regression for prevalent disease and a Weibull survival regression for interval-censored incident disease. We also propose a non-parametric method (no covariates) to check the assumptions of the logistic-Weibull model. We calculate risks of cervical intraepithelial neoplasia grade 3 and cancer (CIN3+) for each possible abnormal cotesting result. As an illustrative example of the biases that can occur in real data, we present the Kaplan-Meier, logistic-Weibull, and non-parametric cumulative risk curves for 34,261 women at KPNC who test Pap negative and HPV positive at enrollment. Results: The non-parametric method estimates 1.99% prevalent risk of CIN3+ and 5.68% 7-year cumulative risk of CIN3+ among women who are Pap-negative/HPV-positive at enrollment. In contrast, the Kaplan-Meier method estimates merely 0.18% prevalent risk of CIN3+, primarily because of prevalent disease not diagnosed at baseline. Furthermore, the Kaplan-Meier method overestimates 7-year cumulative risk as 7.37% because it equates the time of onset with the time of diagnosis, and thus overestimates the hazards of disease onset at later times. In contrast, the logistic-Weibull estimates of 1.87% prevalent risk of CIN3+ and 5.84% 7-year cumulative risk of CIN3+ are close to the risk estimates from the non-parametric method. Discussion: The Kaplan-Meier method provided poor risk estimates while logistic-Weibull model-based risks were close to the risk estimates from the non-parametric method. Our findings support use of the logistic-Weibull models over Kaplan-Meier methods for developing the risk estimates that underlie current U.S. cervical cancer screening guidelines. Citation Format: Li C. Cheung, Qing Pan, Noorie Hyun, Mark Schiffman, Barbara Fetterman, Philip E. Castle, Thomas Lorey, Hormuzd A. Katki. Risk models for cancer screening cohorts assembled from electronic health records: Application to calculating risks that underlie current cervical cancer screening guidelines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5300. doi:10.1158/1538-7445.AM2017-5300

Background: The natural history of human papillomavirus (HPV) and the steps leading to cervical cancer are well-known; the steps include infection with one of the 13 carcinogenic HPV genotypes, viral persistence, progression to precancer, and invasion. Cervical screening programs target treatable cervical precancer to prevent cancer mortality and morbidity. HPV infections are very common and only those causing precancer pose a risk of cancer. In addition to HPV genotype, multiple established co-factors can be combined to predict with unparalleled accuracy and precision the broad range of risks for the critical transition from common HPV infection to uncommon cervical precancer. Thus, there are three types of factors predicting risk of precancer: viral (e.g., HPV genotype and viral load), host (e.g., age, race/ethnicity) and behavioral (e.g., oral contraceptive use, smoking, BMI, co-infection with other sexually transmitted agents). We are building a risk prediction model for clinical use that reflects the determinants of HPV natural history. The absolute-risk based model will consider the three possible HPV outcomes: HPV progression, else HPV “clearance” (immune suppression) signifying low risk of subsequent precancer from that infection, else persistence of HPV infection without either progression or clearance (i.e., still unresolved outcome). To estimate these competing risks for all the factors, cofactors and their combinations requires very large cohorts of HPV-infected women. Methods: Our analysis makes use of data from a uniquely large cohort study of HPV-infected women, specifically, the 35,000 HPV-positive women, 30 years or older, from the NCI-Kaiser Permanente Northern California Persistence and Progression cohort study. The median time of follow-up is 3 years (maximum >7 years). Risk predictors already recorded include: woman's age, HPV infection status, HPV genotype, viral load, concurrent cervical cytology result, and the range of behavioral cofactors. We will present at the meeting the steps leading to the final model: 1) univariate, then multivariate, absolute risks of progression, clearance, or persistence for each HPV genotype; 2) the same risks accounting for time to event and loss-to-followup; and 3) the novel statistic mean risk stratification (MRS), which measures how well the model predicts the crucial dichotomous outcome (progression vs. not). MRS identifies which combination of variables, by virtue of frequency of positive results and strength of risk stratification, is most promising in deciding risk-based clinical management (i.e., who needs colposcopic biopsy due to high risk of precancer). We present the univariate absolute risks for HPV genotypes here, but will show the full multivariate proportional hazards and MRS analyses at the conference. Results: Risk of progression (29.4% for HPV16 to 7.2% for HPV68) varied inversely with risk of clearance (60.1% for HPV16 to 81.6% for HPV68), by HPV type. Relatively few (~10%) of infections of any carcinogenic type persisted without progression. The most important univariate cofactors in preliminary analyses are viral load (for HPV16 mainly), woman's age, and concurrent cytology. No behavioral risk factors are especially important. Time to clearance and time to progression did not vary by HPV type, with median time to events of 1.5-2 years. Conclusions: Based on our preliminary results, the fate of most HPV infections is determined within a few years of first detection, based mainly on characteristics of the virus. MRS summarizes the average risk discrimination of the prediction model compared to pre-test probability, permitting estimation of its expected benefit. We hypothesize and will test whether multivariate calculations of absolute risks and the use of mean risk stratification can lead to improved risk-based clinical management of HPV-infected women. Citation Format: Maria Demarco, Noorie Hyun, Hormuzd Katki, Brian Befano, Li Cheung, Tina R. Raine-Bennett, Barbara Fetterman, Thomas Lorey, Nancy Poitras, Julia C. Gage, Phillip E. Castle, Nicolas Wentzensen, Mark Schiffman. Risk model for clinical management of HPV-infected women. [abstract]. In: Proceedings of the AACR Special Conference: Improving Cancer Risk Prediction for Prevention and Early Detection; Nov 16-19, 2016; Orlando, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2017;26(5 Suppl):Abstract nr A28.

BACKGROUND Human papillomavirus (HPV)-based cervical cancer screening requires triage markers to decide who should be referred to colposcopy. p16/Ki-67 dual stain cytology has been proposed as a biomarker for cervical precancers. We evaluated the dual stain in a large population of HPV-positive women. METHODS One thousand five hundred and nine HPV-positive women screened with HPV/cytology cotesting at Kaiser Permanente California were enrolled into a prospective observational study in 2012. Dual stain cytology was performed on residual Surepath material, and slides were evaluated for dual stain-positive cells. Disease endpoints were ascertained from the clinical database at KPNC. We evaluated the clinical performance of the assay among all HPV-positive women and among HPV-positive, cytology-negative women. We used internal benchmarks for clinical management to evaluate the clinical relevance of the dual stain assay. We evaluated sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the dual stain compared with Pap cytology. All statistical tests were two-sided. RESULTS The dual stain had lower positivity (45.9%) compared with cytology at an ASC-US threshold (53.4%). For detection of CIN2+, the dual stain had similar sensitivity (83.4% vs 76.6%, P = .1), and statistically higher specificity (58.9% vs 49.6%, P < .001), PPV (21.0% vs 16.6%, P < .001), and NPV (96.4% vs 94.2%, P = .01) compared with cytology. Similar patterns were observed for CIN3+. Women with a positive test had high enough risk for referral to colposcopy, while the risk for women with negative tests was below a one-year return threshold based on current US management guidelines. CONCLUSION Dual stain cytology showed good risk stratification for all HPV-positive women and for HPV-positive women with normal cytology. Additional follow-up is needed to determine how long dual stain negative women remain at low risk of precancer.

Anal human papillomavirus (HPV) infections are common, and the incidence of anal cancer is high in HIV-infected men who have sex with men (MSM). To evaluate the performance of HPV assays in anal samples, we compared the cobas HPV test (cobas) to the Roche Linear Array HPV genotyping assay (LA) and cytology in HIV-infected MSM. Cytology and cobas and LA HPV testing were conducted for 342 subjects. We calculated agreement between the HPV assays and the clinical performance of HPV testing and HPV genotyping alone and in combination with anal cytology. We observed high agreement between cobas and LA, with cobas more likely than LA to show positive results for HPV16, HPV18, and other carcinogenic types. Specimens testing positive in cobas but not in LA were more likely to be positive for other markers of HPV-related disease compared to those testing negative in both assays, suggesting that at least some of these were true positives for HPV. cobas and LA showed high sensitivities but low specificities for the detection of anal intraepithelial neoplasia grade 2/3 (AIN2/3) in this population (100% sensitivity and 26% specificity for cobas versus 98.4% sensitivity and 28.9% specificity for LA). A combination of anal cytology and HPV genotyping provided the highest accuracy for detecting anal precancer. A higher HPV load was associated with a higher risk of AIN2/3 with HPV16 ( P trend < 0.001), HPV18 ( P trend = 0.07), and other carcinogenic types ( P trend < 0.001). We demonstrate that cobas can be used for HPV detection in anal cytology specimens. Additional tests are necessary to identify men at the highest risk of anal cancer among those infected with high-risk HPV.