ASCO CAP乳腺癌HER2检测指南2023

Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer: ASCO – College of American Pathologists Guideline Update

;

Antonio C. Wolff, MD 1 ; Mark R. Somer fi eld, PhD 2 ; Mitchell Dowsett, PhD 3 ; M. Elizabeth H. Hammond, MD 4 ; Daniel F. Hayes, MD 5 Lisa M. McShane, PhD 6 ; Thomas J. Saphner, $\mathbb{M}\mathbb{D}^{7}\mathbb{G}$ ; Patricia A. Spears, BS 8 ; and Kimberly H. Allison, MD 9

DOI https://doi.org/10.1200/JCO.22.02864

ABSTRACT

PURPOSE To update ASCO – College of American Pathologists (CAP) recommendations for human epidermal growth factor receptor 2 (HER2) testing in breast cancer. The Panel is aware that a new generation of antibody-drug conjugates (ADCs) targeting the HER2 protein is active against breast cancers that lack protein overexpression or gene ampli fi cation.

METHODS

An Update Panel conducted a systematic literature review to identify signals for updating recommendations.

RESULTS

The search identi fi ed 173 abstracts. Of fi ve potential publications reviewed, none constituted a signal for revising existing recommendations.

RECOMMENDATIONS

DISCUSSION

The 2018 ASCO-CAP recommendations for HER2 testing are af fi rmed.

HER2 testing guidelines have focused on identifying HER2 protein overexpression or gene ampli fi cation in breast cancer to identify patients for therapies that disrupt HER2 signaling. This update acknowledges a new indication for trastuzumab deruxtecan when HER2 is not overexpressed or ampli fi ed but is immunohistochemistry (IHC) $^{1+}$ or $^{2+}$ without ampli fi cation by in situ hybridization. Clinical trial data on tumors that tested IHC 0 are limited (excluded from DESTINY-Breast04), and evidence is lacking that these cancers behave differently or do not respond similarly to newer HER2 ADCs. Although current data do not support a new IHC 0 versus $^{1+}$ prognostic or predictive threshold for response to trastuzumab deruxtecan, this threshold is now relevant because of the trial entry criteria that supported its new regulatory approval. Therefore, while it is premature to create new result categories of HER2 expression (eg, HER2-Low, HER2-Ultra-Low), best practices to distinguish IHC 0 from $^{1+}$ are now clinically relevant. This Update af fi rms prior HER2 reporting recommendations and offers a new HER2 testing reporting comment to highlight the current relevance of IHC 0 versus $^{1+}$ results and best practice recommendations to distinguish these often subtle differences.

Additional information is available at www.asco.org/breast-cancer-guidelines .

ACCOMPANYING CONTENT

$\mathcal{O}$ Appendix Data Supplement

Accepted March 29, 2023 Published June 7, 2023

J Clin Oncol 41:3867-3872 © 2023 by American Society of Clinical Oncology and College of American Pathologists

INTRODUCTION

ASCO and the College of American Pathologists (CAP) fi rst published a practice guideline on human epidermal growth factor receptor 2 (HER2) testing in breast cancer in 2007. 1 The guideline was updated in $2013^{2}$ and again in 2018 3 based on targeted literature searching, Panel expertise, and new signals. 4 These guidelines were developed to standardize and ensure accurate detection of HER2 gene ampli fi ed or protein overexpressed breast cancers for prediction of bene fi t from HER2-targeted therapies. The clinical utility of

HER2 testing to identify patients for therapy with the HER2 antibody trastuzumab has since expanded to other antibodies (pertuzumab added to trastuzumab, margetuximab), small molecule tyrosine kinase inhibitors (lapatinib, neratinib, or tucatinib), and antibody-drug conjugates (ADCs) (trastuzumab emtansine or trastuzumab deruxtecan).

The impetus for revisiting the ASCO-CAP guideline was the 2022 publication of the DESTINY-Breast04 trial. Modi et al 5 showed in an open-label phase III study a signi fi cant improvement in survival in patients with breast cancers without

THE BOTTOM LINE

Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer: ASCO – College of American Pathologists Guideline Update

Guideline Question

What is the optimal testing algorithm for the assessment of human epidermal growth factor receptor 2 (HER2) status and what strategies can help ensure optimal performance, interpretation, and reporting of established assays?

Target Population

Patients with breast cancer.

Target Audience

Medical oncologists, surgical oncologists, radiologists, pathologists, oncology nurses, patients, caregivers, advocates, and oncology advanced practice providers.

Methods

A systematic review of the literature was performed, and relevant evidence was evaluated for inclusion in this updated clinical practice guideline (Data Supplement, online only).

Recommendations

The 2018 ASCO – College of American Pathologists (CAP) recommendations for HER2 testing are af fi rmed.

HER2 Immunohistochemistry (IHC) Testing Best Practices Recommendations

• HER2 testing should still be optimized for the predictive purpose of identi fi cation of breast cancers with protein overexpression and/or gene ampli fi cation who could bene fi t from therapies aimed at disrupting HER2 signaling pathways. • While it is premature to change reporting terminology for lower levels of HER2 IHC expression (eg, HER2-Low), pathology labs should include a footnote in their HER2 testing reports (IHC and in situ hybridization [ISH]) with the following recommended comment: s “ Patients with breast cancers that are HER2 IHC $^{3+}$ or IHC 2 1 /ISH ampli fi ed may be eligible for several therapies that disrupt HER2 signaling pathways. Invasive breast cancers that test ’ HER2-negative ’ (IHC 0, 1 1 or 2 1 /ISH not-ampli fi ed) are more speci fi cally considered ’ HER2-negative for protein overexpression/gene ampli fi cation ’ since non-overexpressed levels of the HER2 protein may be present in these cases. Patients with breast cancers that are HER2 IHC $^{1+}$ or IHC 2 1 / ISH not-ampli fi ed may be eligible for a treatment that targets non-ampli fi ed/non-overexpressed levels of HER2 expression for cytotoxic drug delivery (IHC 0 results do not result in eligibility currently). ” • HER2 IHC 1 1 or 0 results are still both interpreted as HER2-negative (HER2 is not overexpressed) using the previously recommended scoring criteria ( Fig 1 ). Importantly, the semiquantitative IHC score must always be reported as well to ensure patients who meet eligibility criteria for trastuzumab deruxtecan can be identi fi ed. $\bigcirc$ Example: HER2-negative for protein overexpression ( $^{\prime}\!{}^{+}$ staining present). • Since eligibility for trastuzumab deruxtecan (IHC 1 1 or IHC 2 1 /ISH not-ampli fi ed) may hinge around the IHC 0/IHC $^{1+}$ threshold (although the clinical validity of this threshold remains untested), pathologists can make best practice efforts to distinguish IHC 1 1 results from 0 by the following practices: 1. Examining HER2 IHC stained slides using standardized ASCO-CAP guidelines scoring criteria (see Fig 1 for interpretation). 2. Examining HER2 IHC at high power $(40\times)$ when discriminating 0 from $^{1+}$ staining 3. Considering second pathologist review when results are close to the 0 versus $^{1+}$ interpretive threshold $(>\!10\%$ of cells with incomplete membrane staining that is faint/barely perceptible). 4. Usingcontrolswitharangeofproteinexpression(including1 1 )tohelpensuretheassayhasanappropriatelimitofdetection. 5. Careful attention to preanalytic conditions of breast cancer tissue samples from both primary and metastatic sites. • Medical oncologists can also consider HER2 IHC results on prior or concurrent primary samples (or other metastatic sites) because there may be heterogeneity in HER2 expression levels between samples and because metastatic cancer tissue samples may suffer from preanalytic conditions that are not as well monitored as in primary breast tissue samples.

HER2 overexpression or ampli fi cation, but with immunohistochemistry (IHC) $^{1+}$ or IHC $^{2+}$ with in situ hybridization (ISH) not-ampli fi ed results, treated with the ADC famtrastuzumab-deruxtecan-nxki compared with physician ’ s

choice of chemotherapy after progression on other therapies for metastatic disease. Participants in the control arm did not have access to trastuzumab deruxtecan after progression, and patients with IHC 0 results were excluded from the trial.

These data extended the US Food and Drug Administration – approved label indication of this drug and resulted in premarket approval of the monoclonal IHC antibody testing system used in DESTINY-Breast04 (Ventana PATHWAY antiHER2/neu 4B5 rabbit monoclonal antibody on the BenchMark ULTRA instrument) for a new use as a semiquantitative assay, to identify patients with breast cancer without HER2 overexpression or ampli fi cation who could be eligible for treatment with trastuzumab deruxtecan. 6 The implications of these data for the ASCO-CAP breast cancer HER2 testing guideline recommendations were reviewed.

EVIDENCE REVIEW

Although there is clearly a new role for IHC assays to most accurately identify tumors that test HER2 IHC $^{1+}$ or $_{2+}/\mathrm{ISH}$ not-ampli fi ed, this clinical need is based on DESTINY-Breast04 clinical trial entry criteria rather than the demonstration of a new predictive or prognostic threshold for HER2 IHC test results below overexpression (IHC 3 1 ). 5 As patients whose cancers tested IHC 0 in a central laboratory were ineligible for the trial, and trial data did not identify a differential bene fi t between patients with IHC 1 1 and 2 1 /ISH not-ampli fi ed treated with trastuzumab deruxtecan, no new predictive biomarker threshold for response (yes or no) has been identi fi ed among tumors historically classi fi ed as HER2-negative for overexpression or ampli fi cation. Instead, a new threshold has been artifactually created between a result of IHC 0 and IHC at least $^{1+}$ to determine access to the drug based on trial eligibility.

The terminology HER2-Low was used in the trial as shorthand for IHC $^{1+}$ or $_{2+}/\mathrm{ISH}$ not-ampli fi ed cases. However, other than renaming test results to fi t trial eligibility for this new treatment indication, there is no evidence that HER2-Low is a new or reproducibly de fi ned subtype of breast cancer with distinct prognostic or predictive implications. 7 - 13 HER2 IHC 0 versus Low status also appears to be unstable across patient samples, with close to $\angle0\%$ of cases switching between IHC 0 and IHC $^{1+}$ or $_{2+}/\mathrm{ISH}$ not-ampli fi ed (HER2-Low) results when paired primary and metastatic are compared. 8

There are data to suggest that IHC 0 cases may also have low levels of HER2 protein expression by more sensitive testing methods. 11 Higher frequencies of HER2 protein detection in fresh tissue samples also suggest that preanalytic factors during tissue processing likely affect protein detection rates at low levels. 14 Data from one single-arm phase II study (the DAISY trial [ClinicalTrials.gov identi fi er: NCT04132960 ] reported in abstract form) suggest that IHC 0 and HER2-Low cancers have similar response to trastuzumab deruxtecan. Although these results require con fi rmation, they suggest that an IHC 0 result may not truly mean a cancer has no targetable HER2 protein present as HER2 assays are semiquantitative and were optimized to detect overexpression. Additional clinical trial data are needed to determine if IHC 0 samples also include targetable levels of HER2 protein needed

for clinical response, to test if new, more sensitive assays can accurately quantify it, and to investigate if there is differential clinical bene fi t based on protein expression levels. Currently, there is a risk that available IHC assays are suboptimal for detection of these low levels of protein expression and could result in false-negative and false-positive test determinations around the IHC 0/IHC 1 1 threshold that would incorrectly in fl uence treatment recommendations and potentially impact data from ongoing clinical trials that still rely on them.

The Panel (Appendix Table A1 , online only) also notes that adoption of HER2-Low terminology in IHC reporting is problematic because it would require changing the reporting schema for IHC $^{2+}$ results (currently reported as equivocal for protein overexpression with re fl ex ISH testing required to determine gene ampli fi cation status), such that the fi nal IHC result category could not be reported (as HER2-Low v HER2-positive) until re fl ex ISH results are back. Since the DESTINY-Breast04 trial used current standard IHC scoring de fi nitions for 0, 1 1 , 2 1 , and $^{3+}$ , there is also no evidence to support changing these at this time.

Based on the lack of new data to support a change to current HER2 scoring and reporting recommendations, the prior HER2 guideline recommendations for classic anti-HER2 therapies are af fi rmed, and no new reporting terminology is adopted. However, a new HER2 testing reporting footnote and additional best practices to identify candidates that may be eligible for trastuzumab deruxtecan are offered (The Bottom Line box).

RECOMMENDATIONS

The recommendations in previous (2013 and 2018) ASCO-CAP HER2 testing guideline updates are af fi rmed for classic antiHER2 therapies that conventionally target HER2 signaling (Appendix Table A2 , online only). Although no changes are made to prior recommendations, there should be awareness that, for metastatic patients without HER2 overexpression or gene ampli fi cation, an $\mathrm{{Im}C1+}$ or $^{2+}$ result may make patients eligible for treatment targeting nonampli fi ed/nonoverexpressed levels of HER2 expression (and IHC 0 results would not), for which trastuzumab-deruxtecan is the only currently available agent. A new HER2 testing reporting footnote and best practices for identi fi cation and reporting of IHC 0 versus IHC $^{1+}$ results are offered in the bulleted Bottom Line box.

Guideline Disclaimer

The Clinical Practice Guidelines and other guidance published herein are provided by ASCO and the CAP to assist providers in clinical decision making. The information herein should not be relied upon as being complete or accurate, nor should it be considered as inclusive of all proper treatments or methods of care or as a statement of the standard of care. With the rapid development of scienti fi c knowledge, new evidence may emerge between the time information is developed and when it

 FIG 1. 2018 Algorithm for evaluation of HER2 protein expression by IHC assay of the invasive component of a breast cancer specimen. NOTE. The fi nal reported results assume that there is no apparent histopathologic discordance observed by the pathologist. Unusual staining patterns of HER2 by IHC can be encountered that are not covered by these de fi nitions. In practice, these patterns are rare and if encountered should be considered $^{11C2+}$ equivocal. As one example, some speci fi c subtypes of breast cancers can show IHC staining that is moderate to intense but incomplete (basolateral or lateral) and can be found to be HER2 ampli fi ed. Another example is circumferential membrane IHC staining that is intense but within ${\tt\Delta}{\tt V}^{\tt I}{\tt V}^{\%}$ of tumor cells (heterogeneous but very limited in extent). Such cases can be considered $^{2+}$ equivocal but additional samples may reveal different percentages of HER2-positive staining. \*Readily appreciated using a low-power objective and observed within a homogeneous and contiguous invasive cell population. This algorithm is reprinted from recommendations in Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer: ASCO/College of American Pathologists Clinical Practice Guideline Focused Update . 3 This is a tool based on an ASCO and College of American Pathologists guideline and is not intended to substitute for the independent professional judgment of the treating physician. Practice guidelines do not account for individual variation among patients. This tool does not purport to suggest any particular course of medical treatment. Use of the guideline and this tool are voluntary. www.asco.org/breast-cancer-guidelines . $\copyright$ 2023 American Society of Clinical Oncology and College of American Pathologists. All rights reserved. For licensing opportunities, contact licensing@asco.org. See reporting “ Recommended comment ” and “ Example ” in The Bottom Line box. HER2, human epidermal growth factor receptor 2; IHC, immunohistochemistry; ISH, in situ hybridization.

is published or read. The information is not continually updated and may not re fl ect the most recent evidence. The information addresses only the topics speci fi cally identi fi ed therein and is not applicable to other interventions, diseases, or stages of diseases. This information does not mandate any particular course of medical care. Further, the information is not intended to substitute for the independent professional judgment of the treating provider, as the information does not account for individual variation among patients. Recommendations specify the level of con fi dence that the recommendation re fl ects the net effect of a given course of action. The use of words like “ must, ” “ must not, ” “ should, ” and “ should not ” indicates that a course of action is recommended or not recommended for either most or many patients, but there is latitude for the treating physician to select other courses of action in individual cases. In all cases, the selected course of action should be considered by the treating provider in the context of treating

the individual patient. Use of the information is voluntary. ASCO and the CAP do not endorse third party drugs, devices, services, or therapies used to diagnose, treat, monitor, manage, or alleviate health conditions. Any use of a brand or trade name is for identi fi cation purposes only. ASCO and the CAP provide this information on an “ as is ” basis and make no warranty, express or implied, regarding the information. ASCO and the CAP speci fi cally disclaim any warranties of merchantability or fi tness for a particular use or purpose. ASCO and the CAP assume no responsibility for any injury or damage to persons or property arising out of or related to any use of this information, or for any errors or omissions.

Guideline and Con fl icts of Interest

The Expert Panel was assembled in accordance with ASCO ’ s Con fl ict of Interest Policy Implementation for Clinical Practice

Guidelines ( “ Policy, ” found at www.asco.org/guidelinemethodology ). All members of the Expert Panel completed ASCO ’ s disclosure form, which requires disclosure of fi nancial and other interests, including relationships with commercial entities that are reasonably likely to experience direct regulatory or commercial impact as a result of promulgation of the guideline. Categories for disclosure include employment; leadership; stock or other ownership; honoraria, consulting or advisory role; speaker ’ s bureau; research funding; patents, royalties, other intellectual property; expert testimony; travel, accommodations, expenses; and other relationships. In accordance with the Policy, the majority of the members of the Expert Panel did not disclose any relationships constituting a con fl ict under the Policy.

ADDITIONAL RESOURCES

More information, including a supplement with additional evidence tables, slide sets, and clinical tools and resources, is available at www.asco.org/breast-cancer-guidelines . The Methodology Manual (available at www.asco.org/guidelinemethodology ) provides additional information about the methods used to develop this guideline. Patient information is available at www.cancer.net .

ASCO believes that cancer clinical trials are vital to inform medical decisions and improve cancer care, and that all patients should have the opportunity to participate.

AFFILIATIONS

1 Johns Hopkins Sidney Kimmel Comprehensive Cancer Center, Baltimore, MD 2 American Society of Clinical Oncology, Alexandria, VA 3 The Royal Marsden NHS Foundation Trust, London, United Kingdom 4 Intermountain Healthcare and University of Utah School of Medicine, Salt Lake City, UT 5 University of Michigan, Ann Arbor, MI 6 National Cancer Institute, Bethesda, MD 7 Vince Lombardi Cancer Clinic, Two Rivers, WI 8 University of North Carolina, Chapel Hill, NC 9 Stanford University School of Medicine, Stanford, CA

RELATED ASCO GUIDELINES

• Integration of Palliative Care into Standard Oncology Care 15 ( http://ascopubs.org/doi/10.1200/JCO.2016.70.1474 ) • Patient-Clinician Communication 16 ( http://ascopubs.org/ doi/10.1200/JCO.2017.75.2311 ) • Chemotherapy and Targeted Therapy for HER2-Negative Metastatic Breast Cancer 17 ( https://ascopubs.org/doi/ 10.1200/JCO.22.01533 )

CORRESPONDING AUTHOR

American Society of Clinical Oncology, 2318 Mill Rd, Suite 800, Alexandria, VA 22314; e-mail: guidelines@asco.org.

COPYRIGHT

$@2023$ by American Society of Clinical Oncology and College of American Pathologists. This guideline update was developed through collaboration among the College of American Pathologists and the American Society of Clinical Oncology and has been jointly published by invitation and consent in the Archives of Pathology & Laboratory Medicine and the Journal of Clinical Oncology . It has been edited in accordance with style standards established at the Journal of Clinical Oncology . All rights reserved.

EDITOR ’ S NOTE

This American Society of Clinical Oncology and College of American Pathologists Clinical Practice Guideline provides recommendations, with comprehensive review and analyses of the relevant literature for each recommendation. Additional information, including a supplement with additional evidence tables, slide sets, clinical tools and resources, and links to patient information at www.cancer.net , is available at www.asco.org/survivorship-guidelines .

EQUAL CONTRIBUTION

A.C.W. and K.H.A. were Expert Panel co-chairs.

AUTHORS ’ DISCLOSURES OF POTENTIAL CONFLICTS OF INTEREST

Disclosures provided by the authors are available with this article at DOI https://doi.org/10.1200/JCO.22.02864 .

AUTHOR CONTRIBUTIONS

Conception and design: Antonio C. Wolff, Mitchell Dowsett, M. Elizabeth H. Hammond, Daniel F. Hayes, Kimberly H. Allison Administrative support: Mark R. Somer fi eld Collection and assembly of data: Antonio C. Wolff, Mark R. Somer fi eld, Kimberly H. Allison Data analysis and interpretation: Antonio C. Wolff, Mark R. Somer fi eld, Mitchell Dowsett, M. Elizabeth H. Hammond, Lisa M. McShane, Thomas J. Saphner, Patricia A. Spears, Kimberly H. Allison Manuscript writing: All authors Final approval of manuscript: All authors Accountable for all aspects of the work: All authors

ACKNOWLEDGMENT

The Expert Panel wishes to thank Alexi A. Wright, MD, MPH, Praveen Vikas, MBBS, and the Evidence Based Medicine Committee for their thoughtful reviews of and insightful comments on this guideline.

REFERENCES

1. Wolff AC, Hammond MEH, Schwartz JN, et al: American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer. J Clin Oncol 25:118-145, 2007 2. Wolff AC, Hammond MEH, Hicks DG, et al: Recommendations for human epidermal growth factor receptor 2 testing in breast cancer: American Society of Clinical Oncology/College of American Pathologists clinical practice guideline update. J Clin Oncol 31:3997-4013, 2013 3. Wolff AC, Hammond MEH, Allison KH, et al: Human epidermal growth factor receptor 2 testing in breast cancer: American Society of Clinical Oncology/College of American Pathologists clinical practice guideline focused update. J Clin Oncol 36:2105-2122, 2018 4. Shojania KG, Sampson M, Ansari MT, et al: How quickly do systematic reviews go out of date? A survival analysis. Ann Intern Med 147:224-233, 2007 5. Modi S, Jacot W, Yamashita T, et al: Trastuzumab deruxtecan in previously treated HER2-low advanced breast cancer. N Engl J Med 387:9-20, 2022 6. US Food and Drug Administration: US Food and Drug Medical Device Database of Premarket Approvals. https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpma/pma.cfm?id 5 p990081s047 7. Fernandez AI, Liu M, Bellizzi A, et al: Examination of low ERBB2 protein expression in breast cancer tissue. JAMA Oncol 8:607-614, 2022 8. Miglietta F, Griguolo G, Bottosso M, et al: Evolution of HER2-low expression from primary to recurrent breast cancer. NPJ Breast Cancer 7:137, 2021 9. Tarantino P, Jin Q, Tayob N, et al: Prognostic and biologic signi fi cance of ERBB2-low expression in early-stage breast cancer. JAMA Oncol 8:1177-1183, 2022 10. Hurvitz SA: DESTINY-changing results for advanced breast cancer. N Engl J Med 387:75-76, 2022 11. Mouta fi M, Robbins CJ, Yaghoobi V, et al: Quantitative measurement of HER2 expression to subclassify ERBB2 unampli fi ed breast cancer. Lab Invest 102:1101-1108, 2022 12. Hein A, Hartkopf AD, Emons J, et al: Prognostic effect of low-level HER2 expression in patients with clinically negative HER2 status. Eur J Cancer 155:1-12, 2021 13. Schettini F, Chic N, Braso-Maristany F, et al: Clinical, pathological, and PAM50 gene expression features of HER2-low breast cancer. NPJ Breast Cancer 7:2021, 2021 14. Slamon DJ, Godolphin W, Jones LA, et al: Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer. Science 244:707-712, 1989 15. Ferrell BR, Temel JS, Temin S, et al: Integration of palliative care into standard oncology care: American Society of Clinical Oncology clinical practice guideline update. J Clin Oncol 35:96-112, 2017 16. Gilligan T, Coyle N, Frankel RM, et al: Patient-clinician communication: American Society of Clinical Oncology consensus guideline. J Clin Oncol 35:3618-3632, 2017 17. Moy B, Rumble RB, Carey LA, et al: Chemotherapy and targeted therapy for human epidermal growth factor receptor 2-negative metastatic breast cancer that is either endocrine-pretreated or hormone receptor-negative: ASCO guideline rapid recommendation update. J Clin Oncol 40:3088-3090, 2022

AUTHORS ’ DISCLOSURES OF POTENTIAL CONFLICTS OF INTEREST

Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer: ASCO – College of American Pathologists Guideline Update

The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated unless otherwise noted. Relationships are self-held unless noted. $|=$ Immediate Family Member, Inst $=$ My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO ’ s con fl ict of interest policy, please refer to www.asco.org/rwc or ascopubs.org/jco/authors/author-center .

Open Payments is a public database containing information reported by companies about payments made to US-licensed physicians ( Open Payments ).

Antonio C. Wolff

Research Funding: Genentech (Inst), Merck Sharp & Dohme (Inst), Array BioPharma (Inst)

Patents, Royalties, Other Intellectual Property: A.C.W. has been named as inventor on one or more issued patents or pending patent applications relating to methylation in breast cancer, and has assigned his rights to JHU, and participates in a royalty sharing agreement with JHU

Mitchell Dowsett

Consulting or Advisory Role: AstraZeneca/MedImmune, Besins Healthcare, Roche, Rovi, Lilly Research Funding: Lilly (Inst) Patents, Royalties, Other Intellectual Property: AIR-CIS — a molecular pro fi le for predicting sensitivity of breast cancer patients to CDK4/6 inhibitors among patients resistant to an aromatase inhibitor (Inst), Share of royalties from the invention of abiraterone

M. Elizabeth H. Hammond Consulting or Advisory Role: Daiichi Sankyo/Astra Zeneca

Daniel F. Hayes Stock and Other Ownership Interests: InBiomotion Honoraria: Tempus Consulting or Advisory Role: Cepheid, Freenome, Epic Sciences, Cellworks, BioVica, Oncocyte, Turnstone Bio, Predictus Biosciences, Guardant Health, L-Nutra, Macrogenics, Tempus, Xilis, Exact Sciences Research Funding: AstraZeneca (Inst), P fi zer (Inst), Menarini Silicon Biosystems (Inst), Cepheid/Danaher (Inst) Patents, Royalties, Other Intellectual Property: Royalties from licensed technology, Diagnosis and Treatment of Breast Cancer. Patent No.: US

8,790,878 B2. Date of Patent: July 29, 2014. Applicant Proprietor: University of Michigan. D.F.H. is designated as inventor/coinventor, Circulating Tumor Cell Capturing Techniques and Devices. Patent No.: US 8,951,484 B2. Date of Patent: February 10, 2015. Applicant Proprietor: University of Michigan. D.F.H. is designated as inventor/ coinventor, Title: A method for predicting progression free and overall survival at each follow-up timepoint during therapy of metastatic breast cancer patients using circulating tumor cells. Patent no.: 05725638.0- 1223-US2005008602

Other Relationship: Menarini, UpToDate Uncompensated Relationships: UpToDate

Patricia A. Spears Consulting or Advisory Role: P fi zer

Kimberly H. Allison Consulting or Advisory Role: Mammotome Expert Testimony: Kaiser Permanente No other potential con fl icts of interest were reported.

APPENDIX

TABLE A1.

Human Epidermal Growth Factor Receptor 2 Testing in Breast Cancer: ASCO-CAP Update Expert Panel Membership

|Name Affiliation Role or Area of Expertise|||| |---|---|---|---| | Kimberly H. Allison, MD (co-chair) Stanford University School of Medicine, Pathology Stanford, CA |||| |Antonio C. Wolff, MD (co-chair)| Johns Hopkins Sidney Kimmel Comprehensive Cancer Center, Baltimore, MD ||Medical Oncology| || Johns Hopkins Sidney Kimmel Comprehensive Cancer Center, Baltimore, MD ||Medical Oncology| |Mitchell Dowsett, PhD | The Royal Marsden NHS Foundation Trust, London, United Kingdom ||Molecular Pathology| |M. Elizabeth H. Hammond, MD| Intermountain Healthcare and University of Utah School of Medicine, Salt Lake City, UT ||Pathology| || Intermountain Healthcare and University of Utah School of Medicine, Salt Lake City, UT ||Pathology| | Daniel F. Hayes, MD |University of Michigan, Ann Arbor, MI ||Medical Oncology| |Lisa M. McShane, PhD|National Cancer Institute, Bethesda, MD||Biostatistics| |Thomas J. Saphner, MD|Vince Lombardi Cancer Clinic, Two Rivers, WI||Community Oncology| | Patricia A. Spears, BS|University of North Carolina, Chapel Hill, NC||Patient Advocacy| | Mark R. Somerfield, PhD ASCO, Alexandria, VA ASCO Practice Guideline Staff (Health Research Methods) ||||

Abbreviation: CAP, College of American Pathologists.

TABLE A2. Af fi rmed 2018 ASCO-CAP Recommendations

|Topic|Recommendations| |---|---| |Specimens to be tested| All newly diagnosed patients with breast cancer must have a HER2 test performed. Patients who then develop metastatic disease must have a HER2 test performed in a metastatic site, if tissue sample is available. | ||Optimal algorithm for HER2 testing 1. IHC 2+ (equivocal) is invasive breast cancer with weak to moderate complete membrane staining observed in >10%| ||of tumor cells.| ||| || 2. On the basis of some criteria (including a tumor grade 3), if the initial HER2 test result in a core needle biopsy specimen of a primary breast cancer is negative, a new HER2 test may be ordered on the excision specimen. | || specimen of a primary breast cancer is negative, a new HER2 test may be ordered on the excision specimen. 3. If a case has a HER2/CEP17 ratio is ≥2.0 but the average HER2 signals/cell is |

TABLE A2. Af fi rmed 2018 ASCO-CAP Recommendations (continued)

|Topic|Recommendations| |---|---| |IHC rejection criteria| Test is rejected and repeated or tested by FISH if Controls are not as expected | |IHC rejection criteria| Controls are not as expected Artifacts involve most of sample | ||Sample has strong membrane staining of normal breast ducts (internal controls)| |IHC interpretation criteria| Should interpret IHC test using a threshold of more than 10% of tumor cells that must show homogeneous, dark circumferential (chicken wire) pattern to call result 3+, HER2 positive. | | Reporting requirements for all assay types | Report must include guideline-detailed elements except for changes to reporting requirement and algorithms defined in this table. | | Optimal tissue handling requirements | Time from tissue acquisition to fixation should be as short as possible; samples for HER2 testing are fixed in 10% neutral buffered formalin for 6-72 hours; cytology specimens must be fixed in formalin. | || Samples should be sliced at 5- to 10-mm intervals after appropriate gross inspection and margin designation and placed in sufficient volume of neutral buffered formalin. Any exceptions to this process must be included in report. | | Optimal tissue sectioning requirements | Sections should ideally not be used for HER2 testing if cut >6 weeks earlier; this may vary with primary fixation or storage conditions Validation of test must be performed before test is offered | | Optimal internal validation procedure || |Optimal initial test validation| Laboratories performing these tests should be following all accreditation requirements, one of which is initial testing validation. The laboratory should ensure that initial validation conforms to the published 2010 ASCO-CAP Recommendations for IHC Testing of ER and PgR guideline validation requirements with 20 negative and 20 positive for FDA-approved assays and 40 negative and 40 positive for LDTs. This requirement does not apply to assays that were previously validated in conformance with the 2007 ASCO-CAP HER2 testing guideline, and who are routinely participating in external proficiency testing for HER2 tests, such as the program offered by the CAP. | | Optimal initial test validation| Laboratories are responsible for ensuring the reliability and accuracy of their testing results, by compliance with accreditation and proficiency testing requirements for HER2 testing assays. Specific concordance requirements are not required. | | Optimal monitoring of test concordance between methods |See text following under "Optimal Laboratory Accreditation" below.| |Optimal internal QA procedures| Should review and document external and internal controls with each test and each batch of tests. Ongoing quality control and equipment maintenance | || Use of standardized operating procedures including routine use of control materials Revalidation of procedure if changed Should perform ongoing competency assessment and document the actions taken as a part of the laboratory record. Participation in and successful completion of external proficiency testing program with at least two testing events | | Optimal external proficiency assessment | Participation in and successful completion of external proficiency testing program with at least two testing events (mailings) a 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 | |Optimal laboratory accreditation| On-site inspection every other year with annual requirement for selfinspection Reviews laboratory validation, procedures, QA results and processes, results, and reports Unsatisfactory performance results in suspension of laboratory testing for HER2 for that method |

Abbreviations: CAP, College of American Pathologists; CEP17, chromosome enumeration probe 17; ER, estrogen receptor; FDA, US Food and Drug Administration; FISH, fl uorescent in situ hybridization; HER2, human epidermal growth factor receptor 2; IHC, immunohistochemistry; ISH, in situ hybridization; LDT, laboratory-developed test; PgR, progesterone receptor; QA, quality assurance.

a Evidence is limited on the ef fi cacy of HER2-targeted therapy in the small subset of cases with a HER2 /CEP17 ratio ${\ge}2.0$ and an average HER2 copy number of ${<\!4.0}$ per cell. In the fi rst generation of adjuvant trastuzumab trials, patients in this subgroup who were randomly assigned to the trastuzumab arm did not seem to derive an improvement in disease-free or overall survival, but there were too few such cases to draw de fi nitive conclusions. IHC expression for HER2 should be used to complement ISH and de fi ne HER2 status. If the IHC result is not $^{3+}$ positive, it is recommended that the specimen be considered HER2 negative because of the low HER2 copy number by ISH and the lack of protein overexpression.

b There are insuf fi cient data on the ef fi cacy of HER2-targeted therapy in cases with a HER2 ratio of $<\!2.0$ in the absence of protein overexpression because such patients were not eligible for the fi rst generation of adjuvant trastuzumab clinical trials. When concurrent IHC results are negative (0 or $^{1+}$ ), it is recommended that the specimen be considered HER2 negative.

c It is uncertain whether patients with an average of ${\tt>}4.0$ and <6.0 HER2 signals per cell and a HER2 /CEP17 ratio of $<\!2.0$ bene fi t from HER2-targeted therapy in the absence of protein overexpression $\left(\mathsf{I H C}3+\right)$ ). If the specimen test result is close to the ISH ratio threshold for positive, there is a higher likelihood that repeat testing will result in different results by chance alone. Therefore, when IHC results are not $^{3+}$ positive, it is recommended that the sample be considered HER2 negative without additional testing on the same specimen.