improving diagnostic capability for hpv disease ... · medical education to provide continuing...

Am J Clin Pathol 2013;140:881-889 881881 DOI: 10.1309/AJCPIBIS19QIYHJY 881

© American Society for Clinical Pathology

AJCP / Original ArticleCM

E/SAM

Improving Diagnostic Capability for HPV Disease Internationally Within the NIH-NIAID Division of AIDS Clinical Trial NetworksCatherine C. Godfrey, MD, FRACP,1 Pamela M. Michelow, MD,2 Mandana Godard, MBA, MS, MT (ASCP),3 Vikrant V. Sahasrabuddhe, MBBS, MPH, DrPH,4 Janice Darden, MS,5 Cynthia S. Firnhaber, MD, MS, DTM&H,6,7 Neal T. Wetherall, PhD, MSc, SM (ASCP),8 James Bremer, PhD,9 Robert W. Coombs, MD, PhD, FRCPC,10 and Timothy Wilkin, MD,11 for the A5282 Study Team

From the 1National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD; 2Cytology Unit, Faculty of Health Sciences, University of the Witwatersrand and National Health Laboratory Service, Johannesburg, South Africa; 3Johns Hopkins University, Patient Safety Monitoring in International Laboratories (SMILE), Baltimore, MD; 4Vanderbilt Institute for Global Health and Department of Medicine, Vanderbilt University School of Medicine, Nashville, TN; 5Social & Scientific Systems, Silver Spring, MD; 6Clinical HIV Research Unit, Faculty of Health Sciences, Department of Internal Medicine, University of Witwatersrand, Johannesburg, South Africa; 7Right to Care, Johannesburg, South Africa; 8HJF-DAIDS, a Division of The Henry M. Jackson Foundation for the Advancement of Military Medicine, Bethesda, MD; 9Rush University Medical Center, Chicago, IL; 10Departments of Laboratory Medicine and Medicine, University of Washington, Seattle, WA; and 11Division of Infectious Diseases, Weill Cornell Medical College, New York, NY.

Key Words: External quality assurance; EQA; HPV; Cervical pathology; Cervical cytology; Histopathology; HPV DNA testing

DOI: 10.1309/AJCPIBIS19QIYHJY

ABSTRACT

Objectives: To evaluate an external quality assurance (EQA) program for the laboratory diagnosis of human papillomavirus (HPV) disease that was established to improve international research capability within the Division of AIDS at the National Institute of Allergy and Infectious Disease–supported Adult AIDS Clinical Trials Group network.

Methods: A three-component EQA scheme was devised comprising assessments of diagnostic accuracy of cytotechnologists and pathologists using available EQA panels, review of quality and accuracy of clinical slides from local sites by an outside expert, and HPV DNA detection using a commercially available HPV test kit.

Results: Seven laboratories and 17 pathologists in Africa, India, and South America participated. EQA scores were suboptimal for EQA proficiency testing panels in three of seven laboratories. There was good agreement between the local laboratory and the central reader 70% of the time (90% confidence interval, 42%-98%). Performance on the College of American Pathologists’ HPV DNA testing panel was successful in all laboratories tested.

Conclusions: The prequalifying EQA round identified correctable issues that will improve the laboratory diagnosis of HPV-related cervical disease at the participating international study sites and will provide a mechanism for ongoing education and continuous quality improvement.

Cervical cancer is the third most common cancer in women globally, and most of the burden occurs in resource-limited settings. These areas have a disproportionate mortal-ity: the age-specific mortality rates for women in resource-limited countries are at least triple those of women in resource-rich environments. Human immunodeficiency virus (HIV)–infected women bear a greater burden of disease than uninfected women, and the high mortality rates from cervical cancer in the younger age groups in resource-limited countries may reflect the burden of HIV disease.1-5 In much of Africa, cervical cancer is the leading cancer cause of mortality for women.6

Cervical diagnostic testing allows for early diagnosis and treatment of precursor lesions. Cytology screening programs have had a substantial impact on mortality in those countries where access to regular screening is available and where

Upon completion of this activity you will be able to:•describetheroleofcytology-basedcervicalscreeninginresource-limitedsettings.

• outlineaninitialapproachthatcanimprovelaboratorydiagnosisofcervicaldiseaseinresource-limitedcountries.

• listthebenefitsofanexternalqualityassuranceprogramfordiagnosingcervicaldiseaseglobally.

TheASCPisaccreditedbytheAccreditationCouncilforContinuingMedicalEducationtoprovidecontinuingmedicaleducationforphysicians.TheASCPdesignatesthisjournal-basedCMEactivityforamaximumof1 AMA PRA Category 1 Credit ™perarticle.Physiciansshouldclaimonlythecreditcommensuratewiththeextentoftheirparticipationintheactiv-ity.ThisactivityqualifiesasanAmericanBoardofPathologyMaintenanceofCertificationPartIISelf-AssessmentModule.

Theauthorsofthisarticleandtheplanningcommitteemembersandstaffhavenorelevantfinancialrelationshipswithcommercialintereststodisclose.

Questionsappearonp916.Examislocatedatwww.ascp.org/ajcpcme.

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882 Am J Clin Pathol 2013;140:881-889882 DOI: 10.1309/AJCPIBIS19QIYHJY


Godfrey et al / HPV EQA at International ACTG Sites

there is an organized approach to cervical cancer prevention. Suspicious cytology usually requires histologic confirmation, but the approach is imperfect—a single test using standard cytology detects high-grade squamous intraepithelial lesions between 55% and 85% of the time.7-9 When abnormali-ties occur, colposcopy-directed biopsy is the gold standard against which cervical screening modalities are measured, but the sensitivity of colposcopy-directed biopsy for cervical intraepithelial neoplasia stage 3 (CIN3) varies from 50% to 65%. It is presumed that the sensitivity for the detection of CIN2 is lower.10

Countries with limited resources have turned to alterna-tive screening strategies because of the substantial clinical and pathologic infrastructure required for cytology-based screen-ing for cervical cancer prevention. Visual inspection methods using acetic acid (VIA) to find suspicious lesions have been adopted by a number of countries; these methods have a range of reported sensitivity for high-grade lesions of 64% to 90% and a specificity of 75% to 83% compared with biopsy in research settings where practitioners are heavily supervised. Results are operator dependent, and interrater reliability is quite variable.11,12 VIA and human papillomavirus (HPV) testing have been shown to reduce the incidence of cervical cancer and its mortality.13-15

HPV testing is being incorporated in screening in the United States16,17 to lengthen the cytologic screening interval, but the technology is attractive in resource-limited settings because it may allow for better triage of women with the highest risk of clinical disease. The test has been proposed as a primary screen for cervical disease, and its sensitivity and specificity for detecting CIN2 have been compared with the histologic gold standard, with good sensitivity (91.9%) and poor specificity (51.4%).3,11 HPV testing with cryotherapy and visual inspection with cryotherapy have been evaluated in a large clinical trial of both HIV-infected and uninfected women in South Africa, and both methods are effective in preventing CIN2+.18 Importantly, when cytology and histo-pathology results are not optimal, the results of HPV DNA testing cannot be validated accurately.

AIDS Clinical Trial Group (ACTG) A5282 is a multi-center randomized phase II trial that compares screening with HPV testing with a cytology-based strategy (NCT01315353). The study will screen 700 women and randomize 280 HIV-infected women to compare cumulative CIN2 or greater rates using the two different strategies. It is being conducted at ACTG sites in Africa (Botswana, Malawi, South Africa, Zam-bia, and Zimbabwe), Haiti, India, and Peru, where screen-and-treat strategies are being considered by the national programs.

The Division of AIDS (DAIDS) at the National Institute of Allergy and Infectious Disease conducted an audit of histopathology capabilities in 2009-2010 at potential sites to assess the abilities of the laboratories to support research in

this area. Significant concerns were raised about the ability of potential participating sites to perform the required evalu-ations in cytology, histopathology, and HPV virology. Spec-imen preparation was an issue; specimen record-keeping, including requisition and report tracking, were also prob-lematic at various laboratories. It was unclear whether local pathologists with large clinical workloads, which include a wide range of anatomic specimens, would be able to perform the cervical cytology and histology to the standard required for this protocol.

External quality assurance (EQA) is an accepted method for ensuring the validity of test results and comparability across different sites, and there are robust EQA programs in cervical cytology.19 Histology-based EQA programs are less developed and occur in the context of continuing professional development or recertification of pathologists. Histologic diagnosis often relies on clinical judgment, and interrater reli-ability testing is performed for few clinical entities. Available schemes for both cytology and histology are costly, time-consuming, and not implemented in most countries where A5282 is being conducted. Where voluntary schemes are in place, there is often little or no oversight of adherence.20-28

In contrast, there is an expectation that virology perfor-mance will be monitored. The use of automated viral DNA and RNA testing means that the development of specific standards for performance is now less complex. Assay-related internal quality assurance is commonly undertaken. Many tests include an internal standard in each test-well, and the manufacturer of the assay provides assay controls.29

A comprehensive EQA program was developed to improve research capacity and clinical care and to support the laboratories during the conduct of A5282. This article reports the prestudy assessments and activities.

Materials and Methods

All international ACTG sites were invited to participate in the A5282 protocol. Each site was asked to identify a pathology laboratory and a pathologist; DAIDS visited the laboratories and considered the capacity proposed by the site using an inspection tool developed for the purpose. A three-part EQA scheme to support HPV-related trials in the ACTG was developed to assess cytopathology (Papanicolaou [Pap] smear), histology (biopsy), and HPV virology (Abbott m2000 Real Time High-Risk HPV DNA assay; Abbott Molecular, Wiesbaden, Germany). Both specimen preparation and diag-nostic accuracy were assessed. Scoring schemes were devel-oped to standardize and quantify results ❚Table 1❚, ❚Table 2❚, and ❚Table 3❚. Data were collected and tabulated by Patient Safety Monitoring in International Laboratories (SMILE), a DAIDS contractor charged with laboratory monitoring.

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AJCP / Original Article

Cytology Proficiency Testing (Scheme I)Participation by cytologists and supervising pathologists

in an internationally recognized EQA program was required (eg, College of American Pathologists [CAP] or Royal Col-lege of Pathologists Australia [RCPA]) and performance was tabulated. Pathologists who scored greater than 90% were allowed to participate in the protocol and followed the schedule of quality assurance activities recommended by the testing program. Those pathologists who scored between 80% and 90% could participate but were required to take part in remediation activities. Pathologists who scored less than 80% could not participate in the protocol until remediation was complete. Remediation was tailored to the site but always included the specific web-based educational activities pro-vided by either CAP or RCPA.

Cytology and Histology Specimen Adequacy and Concurrence (Scheme II)

An independent expert (P.M.M.) with significant exper-tise in the evaluation of cervical pathology in HIV-infected women undertook review of locally collected and prepared specimens. Local guidelines for the collection and prepara-tion of specimens were used rather than introducing poten-tially unsustainable new practices. Laboratories submitted conventional (ie, not liquid-based) cytology slides from five women, preferably HIV infected; at least two slides were to have abnormal cellular results. The central reviewer assessed specimen adequacy and smear preparation and offered a diagnosis based on the Bethesda classification. Discrepancies were adjudicated by a third reader. The agreement between local and central interpretation of cytol-ogy specimens was scored in two ways. First, the cytologic results were dichotomized as normal or abnormal (atypical cells of uncertain significance or worse), and the number of concordant specimens was counted. An acceptable result was defined as having four or more concordant results on five separate specimens. In addition, a scoring system, the “modified cytology score,” for discrepant results was devel-oped in which clinically important discordant results were weighted more heavily. A modified cytology score of 60% or greater was acceptable for protocol participation, and less than 60% required remediation prior to interpreting study specimens. Scores between 60% and 75% required remedia-tion, but participation was allowed. The scoring schemes are presented in Table 3.

The histology assessment was similar to the cytology assessment described above. One histology slide from each of five separate colposcopies, in which one or more cervical biopsy specimens were obtained, was submitted along with the corresponding site-interpreted cervical cytology result. At least two of the five cases were to have had CIN2 or greater on histology. The central expert assessed slides for adequacy

of specimen collection and slide preparation and submitted a diagnosis based on commonly accepted criteria. The local interpretation was categorized as concordant vs discordant ❚Table 4❚ and ❚Table 5❚ as above and was given a weighted score reflecting clinically important differences, referred to as the “modified histology score.” A score of 80% was consid-ered acceptable and allowed protocol participation. Patholo-gists who scored less than 80% were not allowed to participate until they obtained a passing score.

All laboratories were provided the results of the central review, and specific feedback was given. All laboratories are required to participate in EQA activities and provide study slides for interpretation every 6 months. For those laboratories that did not achieve acceptable scores, remediation plans were developed that were tailored to the particular site and could

❚Table 1❚Program Description

Scheme Description

I Cytology proficiency testing (procured commercially)II Assessment of clinical material from each siteIII Virology (human papillomavirus DNA testing)

❚Table 3❚Cytology Scoringa

Local

ASC-H/ Invasive Central NILM ASC-US LSIL HSIL Cancer

Unsatisfactory 0 0 0 0 0NILM 20 10 0 0 0ASCUS 10 20 20 15 10LSIL 0 20 20 10 10ASC-H/HSIL –10 10 10 20 10Invasive cancer –10 0 0 10 20

ASC-H, atypical squamous cells—cannot exclude high-grade squamous intraepithelial lesions; ASC-US, atypical squamous cells of undetermined significance; HSIL, high-grade squamous intraepithelial lesions; LSIL, low-grade squamous intraepithelial lesions; NILM, negative for intraepithelial lesions or malignancy.

a Cytology scoring reflects clinically important differences.

❚Table 2❚Cytology Grading

Local

Central Normal ASC-US LSIL HSIL

Normal Concordant Discordant Discordant DiscordantASC-US Discordant Concordant Concordant ConcordantLSIL Discordant Concordant Concordant ConcordantHSIL Discordant Concordant Concordant Concordant

ASC-US, atypical squamous cells of undetermined significance; HSIL, high-grade squamous intraepithelial lesions; LSIL, low-grade squamous intraepithelial lesions.

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include being asked to submit more samples for the prequali-fying round and ongoing remediation with more frequent on-study EQA. When discrepancies occurred, corrections with specific teaching points were provided to the sites. An example is provided in ❚Image 1❚.

HPV DNA Testing (Scheme III)The DNA test used for HPV testing was the Abbott

m2000 Real Time High-Risk HPV DNA assay, which tests 14 high-risk HPV types (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and 68). The test is an automated real-time polymerase chain reaction assay with internal controls. The DAIDS-sponsored Virology Quality Assurance (VQA) program was asked to develop an external quality control validation panel that was prepared from dilutions of the Abbott High-Risk HPV DNA kit.

The DAIDS-sponsored VQA program developed a vali-dation panel. For the panel, Abbott provided multiple vials of HPV-positive and HPV-negative kit controls and HPV Cervicollect medium as a diluent. C33A cells (HPV-negative cervical epithelial cells) were added to the diluent and to the HPV-positive and HPV-negative controls to achieve a final concentration of approximately 100,000 cells/mL. The diluent (with cells) was then used to dilute the HPV-positive kit con-trol 1:5, 1:10, or 1:20. Over four runs, each laboratory tested the negative control approximately eight times, the positive

control (undiluted) eight times, the positive control (1:5) 10 times, the positive control (1:10) 11 times, and the positive control (1:20) 11 times.

Positive and negative controls were provided to each site laboratory. The validation panel was pretested at two dif-ferent reference laboratories: the Abbott m2000 Real Time High-Risk HPV DNA assay was tested at Abbott Molecular Delkenheim Laboratory in Wiesbaden, Germany, and the Roche Linear Array HPV Test (Roche Molecular Systems, Indianapolis, IN) and a liquid bead microassay based on Luminex technology (Luminex, Austin, TX) for HPV geno-typing30 at the University of Washington, Seattle.

Results

Proficiency Testing (Scheme I)Seven laboratories and 16 pathologists in Africa, India,

and South America participated in the program. Sixteen pathologists from five laboratories used the CAP program purchased by SMILE, and two sites used the RCPA pro-gram already in use at their site. Sixteen pathologists at five sites used the CAP program, and two sites subscribed to the RCPA program and submitted consensus results in which multiple pathologists participated. Eight pathologists from five sites had scores of greater than 90%. Five patholo-gists participating in CAP required remediation but were allowed to participate (scores 80%-90%), and three required substantial remediation, one of whom was ultimately suc-cessful. Of the two laboratories using the RCPA scheme, one was fully successful and the second was allowed to participate with remediation.

Central Review (Scheme II)Cervical smears and biopsy specimens were prepared

according to local protocols and were not standardized. Details on the individual laboratories from the prequalifying rounds of EQA are presented in ❚Table 6❚.

❚Table 4❚Histology Gradinga

Local

Central Normal CIN1 (Low-Grade CIN) CIN2 (High-Grade CIN) CIN3 (High Grade)

Normal Concordant Concordant Discordant DiscordantLow-grade CIN (CIN1) Concordant Concordant Discordant DiscordantHigh-grade CIN (CIN2) Discordant Discordant Concordant ConcordantHigh-grade CIN (CIN3) Discordant Discordant Concordant Concordant

CIN, cervical intraepithelial neoplasia.a Note that a reading called “normal” and “CIN1” was considered concordant, reflecting the understanding of the pathophysiology. Modified grading schemes: The schemes were

developed to reflect gradations in interpretation and to attach greater importance to clinically important diagnoses. There were separate schemes for cytology and histology, and pathologists and cytotechnologists have different scoring grids.

❚Table 5❚Histology Scoring

Local

CIN1 (Low- CIN2/3 (High- Invasive Central NILM Grade CIN) Grade CIN) Cancer

NILM 20 10 –10 –10CIN1 10 20 10 –10CIN2/3 –10 10 20 0Invasive cancer –10 –10 0 20

CIN, cervical intraepithelial neoplasia; NILM, negative for intraepithelial lesions or malignancy.

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CytologyConventional cytology was used since liquid-based

smears were not available at the sites. Eleven (31%) of 35 slides examined had problems with smear processing such as suboptimal fixation and air-drying artifact. In six (17%) instances, poor cellularity was noted, and there were insuf-ficient cells for diagnosis in one case. The diagnoses at two sites had good concordance with the central reviewer, two

had borderline concordance, and three required remediation. When categorized as normal vs abnormal, overall there was agreement between the central reader and the local laboratory 71% of the time (90% confidence interval, 42%-98%). Slides with discordant results between the central reviewer and the site had a third reader who concurred with the central reviewer in every instance. The mean modified cytology score for all laboratories was 60%, reflecting suboptimal performance by

A B

C D

❚Image 1❚ A, Local result: atypical squamous cells of undetermined significance/inflammation. Central result: high-grade squamous intraepithelial lesion (HSIL). Although this smear is degenerate and poorly stained, parabasal and metaplastic cells with a very high nuclear/cytoplasmic ratio are noted. A syncytia of squamous cells, with ill-defined cell borders, shows HSIL (arrowhead). The nuclear borders are irregular and the chromatin is hyperchromatic. These cytologic features are consistent with HSIL. Koilocyte (low-grade squamous intraepithelial lesion)–intermediate squamous cells with a well-defined perinuclear halo and enlarged nucleus are noted (arrow). B and C, Metaplastic cells, some of which have a higher nuclear/cytoplasmic ratio with irregular, hyperchromatic nuclei (arrow), most in keeping with atypical squamous cells cannot exclude HSIL, although the local result was negative. D, A syncytium of squamous cells with ill-defined cell borders and enlarged, hyperchromatic nuclei with irregular nuclear borders and occasional longitudinal grooves (arrow) consistent with HSIL, although the local result was negative.

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three laboratories. One laboratory was considered to have borderline performance.

HistologyNine (26%) of 35 histologic specimens had significant

issues with sample collection, including cautery artifact and/or absence of the transformation zone. Nine (26%) of 35 specimens were suboptimally processed. For 28 (80%) of 35 slides, there was concordance between the local and central reviewer. The aggregate mean modified histology score was 81%. Three laboratories had complete concordance with the central reviewer, two had acceptable scores, and two required further remediation.

Virology (Scheme III)Nine laboratories participated over a two-year period, and

all were successful. Specimens were shipped from laboratory 2 to another site for HPV DNA testing ❚Table 7❚.

RemediationImprovement plans were specifically tailored to the site

and were separate for EQA schemes I and II. For scheme I, among those sites participating in the CAP Pap proficiency testing program, only one laboratory did not require reme-diation; however, during the follow-up round of EQA, this laboratory achieved only provisional approval to participate in the protocol, thus emphasizing the requirement for con-tinuous quality improvement as part of protocol execution. The CAP program provides web-based educational activi-ties, and all pathologists with scores less than 90% were required to participate in this program. Additional intensive training activities were provided to selected sites by the central reviewer. Of the sites participating in the RCPA program, one site required remediation. Pathologists who scored less than 80% were not allowed to participate, and those with scores between 80% and 90% could participate as long as remediation occurred.

❚Table 6❚Laboratory Scores: Scheme II

Cytology Histology

Laboratory Specimen Modified Modified No. Adequacy/Quality % Agreement Score, % Specimen Adequacy/Quality % Agreement Score, %

1 Acceptable 100 90 Specimen collection: 4/5 specimens 40 70 cautery effect/lack of epithelium; specimen processing: 4/5 suboptimal staining 2 5/5 Poor staining 60 30 1/5 Insufficient tissue 80 70 and cellularity 3 Acceptable 80 65 2/5 Insufficient tissue; repeat: 40; Repeat: 80 50; Repeat: 90 acceptable 4 Acceptable/ 60; Repeat: 80 25; Repeat: 65 Acceptable 100 100 minor issues 5 Acceptable/ 0; Repeat 1: 40 15; Repeat 1: 20; Acceptable 100 80 minor issues (failing); repeat repeat 2: 80 2: 80 (passing) 6 Acceptable 80 90 Acceptable 100 907 Acceptable 100 100 Acceptable 80 80

❚Table 7❚Scheme III Virology Resultsa

2011 2012

Laboratory No. A B C A B C

1 No data No data No data No data 100 2 (Sent specimens to a different library) No data No data No data No data No data No data3 No data No data No data No data 100 1004 No data No data 100 100 100 5 No data No data No data No data 100 6 No data 100 100 100 100 7 No data 100 100 100 100

Values are presented as percentages.

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specimen reached the laboratory. For example, a laboratory with poorly preserved biopsy specimens was noted to be using formalin that had expired. A faulty colposcope and subopti-mal biopsy equipment resulted in poorly directed biopsies at another site. Specimen preparation was not standardized, and there were common, easily modified errors in speci-men preparation, such as length of fixation and standardized formalin that improved slide quality. Other resources for col-lection and processing of specimens were identified and pro-vided to improve the existing processes. This included expert advice on biopsy collection as well as the purchase of new colposcopes for a number of sites. In some instances, refer-ral patterns were established for both clinical and pathologic consultation. Although these modifications occurred within the context of the study, these improvements will likely have a significant impact on clinical care at the study sites.

Accurate diagnosis is essential for the conduct of clinical trials. There is an expectation that laboratory results will be comparable and reproducible within and between laboratories. A common approach in cancer and cancer prevention trials is to verify each diagnosis with a central review. Our analysis of the prequalifying EQA program has emphasized that in our environment, central review would need to occur in near real time with discordant results being adjudicated and transmitted to the patient to make clinical management decisions. This would present significant logistic and methodologic difficul-ties for the protocol team. Most important, however, the team and sponsors felt strongly about the need to build or improve

For scheme II, no issues were identified in two of the seven laboratories. Four of those requiring remediation need-ed technical support for collection and staining procedures. In all five laboratories requiring remediation for scheme II, there were significant diagnostic discrepancies between the site and the central reader; these pathologists were provided specific and individual feedback ❚Table 8❚. During the conduct of the study, ongoing EQA will be performed, with these sites hav-ing intensive oversight. The remediation plans for cytology and histology are presented in Table 8.

Discussion

To our knowledge, this is the first report of a compre-hensive assessment of protocol prequalifying EQA for the investigation of HPV-related disease at selected ACTG inter-national clinical trial sites. Our approach of certifying research site laboratories, rather than using a central diagnostic labora-tory for making the diagnosis of HPV-related cervical disease, allows for in-study performance monitoring and is a powerful educational tool for strengthening HPV cytologic and histo-pathologic diagnostics. The EQA is an ongoing and regular process that will continue for the duration of the study, and it is expected that gains made for the A5282 clinical trial will be translated into improved clinical diagnostics at the trial sites.

This prestudy certification program identified important factors that compromised specimen evaluation before the

❚Table 8❚Remediation Requirements

Laboratory No. Remediation Cytology Remediation Histology

1 Review cytology collection procedures Loop excision procedures and staining protocol reviewed Reagents identified and used Submit five additional slides2 Collection and staining techniques reviewed Remediation plan required Submit five additional slides Plan not fully articulated, will include quarterly follow-up Protocol team to send illustrative pictures and additional feedback 3 Submit five additional slides Review procedure for sectioning tissue blocks Quarterly follow-up Submit five additional slides Protocol team to give additional feedback4 Submit five additional slides None required Protocol team to send illustrative pictures and additional feedback Quarterly follow-up 5 Submit five additional slides None required, although poor staining noted and histology staining protocols shared Protocol team to send illustrative pictures Digital photographs shared Quarterly follow-up Site visit being considered6 No issues No issues7 No issues None required None required Protocol team recommends that “insufficient biopsy material” be communicated to the site with a plan for rebiopsy One sample with insufficient specimen, communicated to site

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MPH, PhD, S. P. Tripathy, MD, MBBS, and Z. M. Chirenje, MD The laboratory directors are C. Ganoza, MD, M. Khandekar, MBBS, MD, P. Katundu, MHS, C. Wallis, PhD, M. W. Munjoma, PhD, W. Stevens, MBBCh, and J. Trusler, MD. Acknowledgments: We thank S. Michelle Dubb, who was the adjudicating third reader for the histologic and cytologic evaluations. We thank the College of American Pathologists and the Royal College of Pathologists of Australasia for their help in reviewing pertinent quality assurance documents and procedures. We also thank the participating sites and laboratories for their commitment to quality assurance processes. We especially thank Joe Fitzgibbon at the Division of AIDS and Katie Lammersen at the Network Coordinating Center for their continuous and substantial support of quality assurance processes.

References 1. Abraham AG, Strickler HD, Jing Y, et al, for the North

American AIDS Cohort Collaboration on Research and Design (NA-ACCORD) of IeDEA. Invasive cervical cancer risk among HIV-infected women: a North American multi-cohort collaboration prospective study. J Acquir Immune Defic Syndr. 2013;62:405-413.

2. Parham GP, Sahasrabuddhe VV, Mwanahamuntu MH, et al. Prevalence and predictors of squamous intraepithelial lesions of the cervix in HIV-infected women in Lusaka, Zambia. Gynecol Oncol. 2006;103:1017-1022.

3. Sahasrabuddhe VV, Bhosale RA, Kavatkar AN, et al. Comparison of visual inspection with acetic acid and cervical cytology to detect high grade cervical neoplasia among HIV-infected women in India. Int J Cancer. 2012;130:234-240.

4. Hawes SE, Critchlow CW, Sow PS, et al. Incident high-grade squamous intraepithelial lesions in Senegalese women with and without human immunodeficiency virus type1 (HIV-1) and HIV-2. J Natl Cancer Inst. 2006;98:100-109.

5. Minkoff H, Zhong Y, Burk RD, et al. Influence of adherent and effective antiretroviral therapy use on human papillomavirus infection and squamous intraepithelial lesions in human immunodeficiency virus–positive women. J Infect Dis. 2010;201:681-690.

6. WHO/ICO Information Centre on HPV and Cervical Cancer (HPV Information Centre). Human Papillomavirus and Related Cancers in World: Summary Report 2009. http://www.hpvcentre.net/statistics/reports/XWX.pdf. Accessed December 17, 2012.

7. Miller AB, Lindsay J, Hill GB. Mortality from cancer of the uterus in Canada and its relationship to screening for cancer of the cervix. Int J Cancer. 1976;17:602-612.

8. Johannesson G, Geirsson G, Day N. The effect of mass screening in Iceland, 1965-74, on the incidence and mortality of cervical carcinoma. Int J Cancer. 1978;21:418-425.

9. National Cancer Institute. Cervical cancer screening (PDQ). Accessed January 10, 2013, from http://www.cancer.gov/cancertopics/pdq/screening/cervical/HealthProfessional/page2

10. Pretorius RG, Belinson JL. Colposcopy [review]. Minerva Ginecol. 2012;64:173-180.

11. Firnhaber C, Mayisela N, Mao L, et al. Validation of cervical cancer screening methods in HIV positive women from Johannesburg South Africa. PLoS One. 2013;8:e53494.

12. Gage JC, Rodriguez AC, Schiffman M, et al. An evaluation by midwives and gynecologists of treatability of cervical lesions by cryotherapy among human papillomavirus–positive women. Int J Gynecol Cancer. 2009;19:728-733.

capacity at each site for both research and clinical care. For this reason, laboratories rather than clinical diagnoses were certified. It is expected that a significant number of specimens will have central review during the EQA process; this is being monitored by both the protocol team and the data safety monitoring board. It is likely that we will be able to verify the wisdom of the approach of monitoring laboratories rather than diagnoses both during the trial and in aggregate at the end of the trial. The EQA process is a powerful educational tool and complements other ongoing quality improvement efforts in the ACTG Network Laboratory program. We expect that substantial gains in the laboratory diagnosis of HPV-related cervical disease will continue to be made with other simple interventions.

Finally, cervical disease is the most common HPV-related cancer in HIV-infected individuals, but there is a growing lit-erature suggesting that HPV-related cancers at other anatomic sites may become important. For example, a strong associa-tion between HIV and anal cancer has been documented in the United States, with case rates in HIV-infected men 45 to 170 times that of age- and risk-matched populations.31 As the life expectancy for HIV-infected individuals improves because of more universal access to effective antiretroviral therapy, it is likely that these cancers will become more important.32 It is therefore prudent to ensure that the clinical and laboratory infrastructure is in place for accurate diagnosis and treatment of these conditions.

Address reprint requests to Dr Godfrey: National Institute of Allergy and Infectious Diseases, National Institutes of Health, 6700 B Rockledge Dr, Bethesda MD 20892; [email protected]. The project described was supported by award numbers UM1 AI068636 (C.S.F., R.W.C., and T.W.), UM1 AI069463-07 (C.S.F.), AI38858 (C.S.F., R.W.C., and T.W.), AI27757 (R.W.C.) from the National Institute of Allergy and Infectious Diseases and supported by National Institute of Mental Health and National Institute of Dental and Craniofacial Research. This project has been funded in whole or in part with federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under contract numbers HHSN272200800014C (N.T.W.), HHSN272201300004C (M.G.), and HHSN272201200023C (J.B.). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of Allergy and Infectious Diseases or the National Institutes of Health. A5282 team members other than the coauthors are as follows: R. Bhosale, MD, B. Grinsztejn, MD, PhD, and M. Mwanahamuntu, MBBS, MMed investigators; R. T. Allen, MA clinical trials specialist; S. Evans, PhD, and R. Matining, MS, statisticians; A. Walawander, data manager; C. Blanchard-Horan, PhD, MA, CCRP, international program specialist; C. Megill, PA-C, and J. Nicotera, RN, BSN, field representatives; L. Blair, BS, MT, ASCP, laboratory technologist; and T. Behm, BS, J. Nowak, and D. Weibel Laboratory data coordinators. The site principal investigators are V. Mave, MD, MPH, N. Kumwenda,

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16. US Preventive Services Task Force. Screening for cervical cancer. http://www.uspreventiveservicestaskforce.org/uspstf/uspscerv.htm. Accessed February 10, 2013.

17. American Congress of Obstetricians and Gynecologists. New cervical cancer screening recommendations from the U.S. Preventive Services Task Force and the American Cancer Society/American Society for Colposcopy and Cervical Pathology/American Society for Clinical Pathology. http://www.acog.org/About%20ACOG/Announcements/New%20Cervical%20Cancer%20Screening%20Recommendations.aspx. Accessed February 10, 2013.

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