©American Society of Clinical Oncology All rights reserved American Society of Clinical Oncology (ASCO)/ College of American Pathologists (CAP) Guideline Recommendations for Immunohistochemical Testing of Estrogen/Progesterone Receptors in Breast Cancer

©American Society of Clinical Oncology All rights reserved INTRODUCTION ASCO and the College of American Pathologists (CAP) previously collaborated on a guideline on HER2 testing, published in 2007 Subject: Estrogen (ER) and Progesterone (PgR) testing Rationale: Evidence of wide variability in test performance and inaccurate results ASCO and CAP decided to produce the first ever evidence-based ER-PgR testing guideline, based on a systematic review

©American Society of Clinical Oncology All rights reserved Background Hormone receptor-positive is the most common breast cancer phenotype worldwide Access to accurate and reliable ER/PgR testing and to established and relatively affordable endocrine therapies could have a profound impact on breast cancer outcomes in high and low/middle income countries across the globe

©American Society of Clinical Oncology All rights reserved Guideline Methodology: Systematic Review ASCO and Cancer Care Ontario jointly conducted a systematic review of the medical literature available from 1990-May 2008 –Ovid (Medline) –EMBASE –Cochrane Database of Systematic Reviews Primary outcome: correlation of hormone receptor status and outcome of endocrine treatment ASCO/CAP Expert Panel made recommendations based on this review

©American Society of Clinical Oncology All rights reserved Clinical Questions 1. What is the optimal testing algorithm for testing ER and PgR status? 1.1 What are the clinically validated methods that can be used in this assessment? 2.What strategies can ensure optimal performance, interpretation, and reporting of established assays? 2.1 What are the preanalytic, analytic and postanalytic variables that must be controlled to ensure that assay results reflect tumor ER and PgR status?

©American Society of Clinical Oncology All rights reserved Clinical Questions, cont’d 2.2. What is the optimal internal quality management regimen to ensure ongoing accuracy of ER and PgR testing? 2.3. What is the regulatory framework that permits application of external controls such as proficiency testing and on-site inspection? 2.4.How can internal and external control efforts be implemented and their effects measured?

©American Society of Clinical Oncology All rights reserved Special Questions 1.Should ER/PgR be done in DCIS or recurrent tumor? 2.Does PgR influence the choice of endocrine therapy?

©American Society of Clinical Oncology All rights reserved Recommendations Optimal algorithm for ER/PgR testing Positive - if finding of ≥ 1% of tumor cell nuclei are immunoreactive Negative - if finding of < 1% of tumors’ cell nuclei are immunoreactive in the presence of evidence that the sample can express ER or PgR (positive intrinsic controls are seen) Uninterpretable - finding that no tumor nuclei are immunoreactive and that internal epithelial elements present in the sample or separately submitted from the same sample lack any nuclear staining Clinical Question 1. What is the optimal testing algorithm for testing ER and PgR status?

©American Society of Clinical Oncology All rights reserved Recommendations Optimal testing conditions Large, preferably multiple core biopsies of tumor are preferred for testing if they are representative of the tumor (grade and type) at resection Interpretation follows guideline recommendation Accession slip and report must include guideline- detailed elements Clinical Question 2. What strategies can ensure optimal performance, interpretation, and reporting of established assays? Clinical Question 2.1 What are the preanalytic, analytic, and postanalytic variables that must be controlled to ensure that assay results reflect tumor ER and PgR status?

©American Society of Clinical Oncology All rights reserved Required reporting elements 1.Percent/proportion of tumor cells staining positively. Percentage either by: 1.Estimation 2.Quantitation (counting cells or image analysis) 3.If cytology specimen, count ≥100 cells 2.Intensity of staining – weak, moderate, or strong – representing an estimate of average of intensity of positive cells relative to positive controls on same batch 3.Interpretation of the assay (+, -, or uninterpretable)

©American Society of Clinical Oncology All rights reserved Two optional report elements are recommended 1.If negative ER and PgR interpretations when the histopathology of the tumor is almost always associated with ER+ and PgR+ results, including: tubular, lobular, and mucinous histological types or Nottingham grade 1 tumors Then an optional cautionary statement should indicate that while the patient’s tumor tested as ER-negative, tumors with the same histological type or Nottingham grade almost always test +

©American Society of Clinical Oncology All rights reserved Optional report elements, cont’d 2. -Pathologist may also provide a composite score e.g. the H score, Allred score, or Quick score -Using the percent and intensity measurements provided -Since each of these is somewhat differently calculated and may lead to confusion across institutions -Scoring is not required

©American Society of Clinical Oncology All rights reserved Recommendations Optimal tissue handling requirements Time from tissue acquisition to start of fixation process should be as short as possible Samples for ER and PgR testing are fixed in 10% neutral buffered formalin (NBF) for 6-72 hours Samples should be sliced at 5 mm intervals after appropriate gross inspection and margins designation and placed in sufficient volume of NBF formalin of a sufficient volume to allow adequate tissue penetration Clinical Question 2.1 What are the preanalytic, analytic, and postanalytic variables that must be controlled to ensure that assay results reflect tumor ER and PgR status?

©American Society of Clinical Oncology All rights reserved Recommendations - Optimal tissue handling requirements, cont’d If tumor comes from remote location, it should be bisected on removal and sent to the laboratory immersed in a sufficient volume of NBF Cold ischemia time, fixative type, and time sample placed in NBF must be recorded

©American Society of Clinical Oncology All rights reserved Recommendations- Optimal tissue handling requirements, cont’d Storage of unstained slides for more than 6 weeks prior to analysis is not recommended Time tissue is removed from patient, time tissue is placed in fixative (cold ischemia time), duration of fixation, and fixative type must be recorded and noted on accession slip or in report

©American Society of Clinical Oncology All rights reserved Additional information in re: Clinical Question 2.1 Standardization of Analytical Variables Antibody Selection Antibodies should have well-established specificity and sensitivity and have been clinically validated (good correlation with patient outcomes) Alternatively, results of lab-selected antibodies should be ≥90% concordant with clinically validated antibodies for ER and PgR-positive category and ≥95% concordant with clinically validated antibodies for ER or PgR negative category Include: ER: 1D5, 6F11, SP1, ER D5; PgR: 1294, 1A6, 312

©American Society of Clinical Oncology All rights reserved Recommendations Optimal internal validation procedure Validation of any test must be done before test is offered Validation must be done using a clinically validated ER or PgR test method Revalidation should be done whenever there is a significant change to the test system, such as a change in the primary antibody clone or introduction of new antigen retrieval or detection systems. Clinical Question 1.1 What are the clinically validated methods that can be used in this assessment?

©American Society of Clinical Oncology All rights reserved Recommendations Optimal internal QA procedures Initial test validation Ongoing quality control and equipment maintenance Initial and ongoing laboratory personnel training and competency assessment Use of standardized operating procedures including routine use of external control materials with each batch of testing and routine evaluation of internal normal epithelial elements or the inclusion of normal breast sections on each tested slide, wherever possible. Regular, ongoing assay reassessment should be done at least semiannually. Revalidation is needed whenever there is a significant change to the test system. Ongoing competency assessment and education of pathologists Clinical Question 2.3. What is the optimal internal quality management regimen to ensure ongoing accuracy of ER and PgR testing?

©American Society of Clinical Oncology All rights reserved Recommendations Optimal external proficiency assessment Mandatory participation in external proficiency testing program with at least two testing events (mailings)/year Satisfactory performance requires at least 90% correct responses on graded challenges for either test Unsatisfactory performance will require laboratory to respond according to accreditation agency program requirements Clinical Question 2.4. What is the regulatory framework that permits application of external controls such as proficiency testing and on-site inspection? Clinical Question 2.5. How can internal and external control efforts be implemented and their effects measured?

©American Society of Clinical Oncology All rights reserved Recommendations Optimal laboratory accreditation On-site inspection every other year with annual requirement for self-inspection Reviews laboratory validation, procedures, QA results and processes, results and reports Unsuccessful performance results in suspension of laboratory testing for ER or PgR

©American Society of Clinical Oncology All rights reserved Special Questions 1.Should ER/PgR be done in DCIS or recurrent tumor? ER and PgR status should be determined on all newly diagnosed invasive breast cancers (primary and/or metastatic site) Lack of validation studies on testing for people with DCIS. Panel saw value, but could not make formal recommendation. Women with breast recurrences accessible to biopsy should also always be tested –To check prior negative results not false negative –To check specimen for emergence of negative clones

©American Society of Clinical Oncology All rights reserved Special Questions 2. Does PgR correlate with or influence the choice of endocrine therapy? The precise role of PgR in patient management has not been strongly established Do not withhold endocrine treatment from women w/ ER-rich, PgR poor tumor Women w/ ER-/PgR+ tumors may respond to endocrine therapy

©American Society of Clinical Oncology All rights reserved How can these efforts be implemented and the effects measured? Educational opportunities from ASCO and CAP CAP certification program for pathologists Coordination of recommendations with NCCN, Commission of Cancer of the American College of Surgeons, the American Joint Committee on Cancer, and patient advocacy groups CAP will: –Review and publish results of proficiency testing and laboratory accreditation –Inclusion of quality monitoring activities on ER/PgR testing in ongoing quality assessment programs

©American Society of Clinical Oncology All rights reserved Guideline Methodology: Panel Members Panel MemberInstitution M. Elizabeth H. Hammond, MD, Co-Chair* Intermountain Health Care, University of Utah School of Medicine, UT Daniel F. Hayes, MD, Co-Chair* University of Michigan Comprehensive Cancer Center, University of Michigan Health and Health System, MI Mitch Dowsett, PhD*Royal Marsden Hospital, UK D. Craig Allred, MD* Washington University School of Medicine, St. Louis, MO Jared N. Schwartz, MD, PhD, FACP, Co-Chair*Presbyterian Hospital, NC Antonio C. Wolff, MD, FACP, Co-Chair* The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins University, MD Sunil Badve, MDECOG, Indiana University, IN Robert L. Becker, MD, Ex-Officio US Food and Drug Administration, Center for Devices and Radiological Health, Office of In Vitro Diagnostic Device Evaluation and Safety Patrick L. Fitzgibbons, MD, FACPSt. Jude Medical Center, CA Glenn Francis, MBBS, FRCPA, MBAPrincess Alexandra Hospital, Australia *Steering Committee Member

©American Society of Clinical Oncology All rights reserved Guideline Methodology: Panel Members, cont’d Panel MemberInstitution Neal S. Goldstein, MDAdvanced Diagnostics Laboratory, MI Malcolm Hayes, MDUniversity of British Columbia, Canada David G. Hicks, MD, FCAPUniversity of Rochester, NY Susan Lester, MDBrigham and Women’s Hospital, MA Richard Love, MDOhio State University, OH Lisa McShane, PhDNCI, Biometric Research Branch, DCTD Keith Miller, MDUK NEQAS C. Kent Osborne, MDBaylor College of Medicine, TX Soonmyung Paik, MDNational Surgical Adjuvant Breast and Bowel Project, PA Jane Perlmutter, PhD, Patient RepresentativeGemini Group, MI Anthony Rhodes, PhDUniversity of the West of England, Bristol, UK NEQAS

©American Society of Clinical Oncology All rights reserved Guideline Methodology: Panel Members, cont’d Panel MembersInstitution Hironobu Sasano, MDTohoku University School of Medicine, Japan Fred C. G. J. Sweep, PhDRadboud University, Nijmegen, Netherlands Sheila Taube, PhDST Consulting, MD Emina Emilia Torlakovic, MD, PhDRoyal University Hospital, Saskatoon, Canada Paul Valenstein, MD, FCAPSt. Joseph Mercy Hospital, Ann Arbor, MI Giuseppe Viale, MD, FRCPathEuropean Institute of Oncology, Milan, Italy Daniel Visscher, MDUniversity of Michigan, Ann Arbor, MI Thomas Wheeler, MD, FCAPBaylor College of Medicine, TX R. Bruce Williams, MD, FCAPThe Delta Pathology Group, Shreveport, LA James L. Wittliff, MD, PhDUniversity of Louisville, KY Judy Yost, MA, MT (ASCP), Ex OfficioCMS, Division of Laboratory Services (CLIA)

©American Society of Clinical Oncology All rights reserved Guideline Methodology: Guests invited to open portion of meeting Invited GuestsAffiliation Richard Bender, MDAgendia Inc Kenneth J. Bloom, MDClarient Allen M. Gown, MDPhenoPath Laboratories, Seattle, WA David L. Rimm, MD, PhDYale University Patrick Roche, PhDVentana Medical Systems Steven Shak, MDGenomic Health Roseanne WelcherDAKO Hadi Yaziji, MDAncillary Pathways, Miami, FL

©American Society of Clinical Oncology All rights reserved Additional ASCO Resources The full text of the guideline, an abridged version of the guideline, an Executive Summary, this slide set, and additional clinical tools and resources can be found at: A patient guide, “What to Know” about this guideline, is available at:

©American Society of Clinical Oncology All rights reserved ASCO Guidelines