Breast Cancer Research and Treatment

, Volume 157, Issue 3, pp 405–411 | Cite as

Updated 2013 College of American Pathologists/American Society of Clinical Oncology (CAP/ASCO) guideline recommendations for human epidermal growth factor receptor 2 (HER2) fluorescent in situ hybridization (FISH) testing increase HER2 positive and HER2 equivocal breast cancer cases; retrospective study of HER2 FISH results of 836 invasive breast cancers

  • Kamaljeet SinghEmail author
  • Umadevi Tantravahi
  • Michele M. Lomme
  • Terese Pasquariello
  • Margaret Steinhoff
  • C. James Sung
Preclinical study


For dual probe HER2 FISH assay, the 2013 CAP/ASCO guideline recommendations lowered the HER2/CEP17 ratio cut off for HER2 amplification to ≥2.0 and introduced an average HER2 copy number criterion for HER2 amplification (≥6.0/cell) and HER2 equivocal categories (≥4 and <6/cell). The HER2/CEP17 equivocal category is eliminated. The aim of this study is to assess the impact of 2013 HER2 FISH testing guideline recommendations update on the assignment of HER2 status with dual probe HER2 FISH assay. Dual probe HER2 FISH assay results on breast cancers from 09/2009 to 07/2015 that underwent reflex HER2 FISH testing after equivocal HER2 (2+) immunohistochemistry (IHC) were reviewed. HER2 copy number, CEP17 signals, and HER2/CEP ratios were noted. HER2 status was assigned as HER2 negative (HER2−), HER2 equivocal (HER2e), and HER2 amplified (HER2+) by applying both 2007 and 2013 CAP/ASCO HER2 FISH guideline recommendations and results were compared. New guidelines reclassified HER2 FISH status in a significant proportion of cases (8.3 %, 69/836; p = .021). There were 22 (2.6 %) more HER2+, 17 (2.1 %) more HER2e, and 39 (4.1 %) fewer HER2− tumors. Change of HER2 status correlated significantly with ≥3 CEP17 signals (38 vs. 2 %; p < .001). The 2013 CAP/ASCO guideline recommendations for HER2 FISH testing by dual probe assay increased the HER2 amplified and HER2 equivocal tumors. Increase in HER2 equivocal tumors would potentially increase the frequency of repeat HER2 testing. Tumors with ≥3 CEP17 signals, so-called chromosome 17 polysomy, are more likely to be impacted and classified as HER2 equivocal.


Breast cancer HER2 FISH Equivocal ASCO CAP Polysomy 


Compliance with ethical standards

Conflict of Interest


Financial disclosure



  1. 1.
    Press MF, Pike MC, Chazin VR, Hung G, Udove JA, Markowicz M, Danyluk J, Godolphin W, Sliwkowski M, Akita R et al (1993) Her-2/neu expression in node-negative breast cancer: direct tissue quantitation by computerized image analysis and association of overexpression with increased risk of recurrent disease. Cancer Res 53(20):4960–4970PubMedGoogle Scholar
  2. 2.
    Slamon D, Eiermann W, Robert N, Pienkowski T, Martin M, Press M, Mackey J, Glaspy J, Chan A, Pawlicki M, Pinter T, Valero V, Liu MC, Sauter G, von Minckwitz G, Visco F, Bee V, Buyse M, Bendahmane B, Tabah-Fisch I, Lindsay MA, Riva A, Crown J, Breast Cancer International Research G (2011) Adjuvant trastuzumab in HER2-positive breast cancer. N Engl J Med 365(14):1273–1283. doi: 10.1056/NEJMoa0910383 CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Slamon DJ, Clark GM, Wong SG, Levin WJ, Ullrich A, McGuire WL (1987) Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science 235(4785):177–182CrossRefPubMedGoogle Scholar
  4. 4.
    Slamon DJ, Leyland-Jones B, Shak S, Fuchs H, Paton V, Bajamonde A, Fleming T, Eiermann W, Wolter J, Pegram M, Baselga J, Norton L (2001) Use of chemotherapy plus a monoclonal antibody against HER2 for metastatic breast cancer that overexpresses HER2. N Engl J Med 344(11):783–792. doi: 10.1056/NEJM200103153441101 CrossRefPubMedGoogle Scholar
  5. 5.
    Press MF, Bernstein L, Thomas PA, Meisner LF, Zhou JY, Ma Y, Hung G, Robinson RA, Harris C, El-Naggar A, Slamon DJ, Phillips RN, Ross JS, Wolman SR, Flom KJ (1997) HER-2/neu gene amplification characterized by fluorescence in situ hybridization: poor prognosis in node-negative breast carcinomas. J Clin Oncol 15(8):2894–2904PubMedGoogle Scholar
  6. 6.
    Ellis MJ, Coop A, Singh B, Mauriac L, Llombert-Cussac A, Janicke F, Miller WR, Evans DB, Dugan M, Brady C, Quebe-Fehling E, Borgs M (2001) Letrozole is more effective neoadjuvant endocrine therapy than tamoxifen for ErbB-1- and/or ErbB-2-positive, estrogen receptor-positive primary breast cancer: evidence from a phase III randomized trial. J Clin Oncol 19(18):3808–3816PubMedGoogle Scholar
  7. 7.
    Menard S, Valagussa P, Pilotti S, Gianni L, Biganzoli E, Boracchi P, Tomasic G, Casalini P, Marubini E, Colnaghi MI, Cascinelli N, Bonadonna G (2001) Response to cyclophosphamide, methotrexate, and fluorouracil in lymph node-positive breast cancer according to HER2 overexpression and other tumor biologic variables. J Clin Oncol 19(2):329–335PubMedGoogle Scholar
  8. 8.
    Pritchard KI, Shepherd LE, O’Malley FP, Andrulis IL, Tu D, Bramwell VH, Levine MN, National Cancer Institute of Canada Clinical Trials G (2006) HER2 and responsiveness of breast cancer to adjuvant chemotherapy. N Engl J Med 354(20):2103–2111. doi: 10.1056/NEJMoa054504 CrossRefPubMedGoogle Scholar
  9. 9.
    Hayes DF, Thor AD, Dressler LG, Weaver D, Edgerton S, Cowan D, Broadwater G, Goldstein LJ, Martino S, Ingle JN, Henderson IC, Norton L, Winer EP, Hudis CA, Ellis MJ, Berry DA, Cancer, Leukemia Group BI (2007) HER2 and response to paclitaxel in node-positive breast cancer. N Engl J Med 357(15):1496–1506. doi: 10.1056/NEJMoa071167 CrossRefPubMedGoogle Scholar
  10. 10.
    Owens MA, Horten BC, Da Silva MM (2004) HER2 amplification ratios by fluorescence in situ hybridization and correlation with immunohistochemistry in a cohort of 6556 breast cancer tissues. Clin Breast Cancer 5(1):63–69CrossRefPubMedGoogle Scholar
  11. 11.
    Bast RC Jr, Ravdin P, Hayes DF, Bates S, Fritsche H Jr, Jessup JM, Kemeny N, Locker GY, Mennel RG, Somerfield MR, American Society of Clinical Oncology Tumor Markers Expert P (2001) 2000 update of recommendations for the use of tumor markers in breast and colorectal cancer: clinical practice guidelines of the American Society of Clinical Oncology. J Clin Oncol 19(6):1865–1878PubMedGoogle Scholar
  12. 12.
    Wolff AC, Hammond ME, Schwartz JN, Hagerty KL, Allred DC, Cote RJ, Dowsett M, Fitzgibbons PL, Hanna WM, Langer A, McShane LM, Paik S, Pegram MD, Perez EA, Press MF, Rhodes A, Sturgeon C, Taube SE, Tubbs R, Vance GH, van de Vijver M, Wheeler TM, Hayes DF, American Society of Clinical Oncology/College of American P (2007) American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer. Arch Pathol Lab Med 131(1):18–43. doi: 10.1043/1543-2165(2007)131[18:ASOCCO]2.0.CO;2 PubMedGoogle Scholar
  13. 13.
    Wolff AC, Hammond ME, Hicks DG, Dowsett M, McShane LM, Allison KH, Allred DC, Bartlett JM, Bilous M, Fitzgibbons P, Hanna W, Jenkins RB, Mangu PB, Paik S, Perez EA, Press MF, Spears PA, Vance GH, Viale G, Hayes DF, American Society of Clinical O, College of American P (2014) 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. Arch Pathol Lab Med 138(2):241–256. doi: 10.5858/arpa.2013-0953-SA CrossRefPubMedGoogle Scholar
  14. 14.
    Carlson RW, Moench SJ, Hammond ME, Perez EA, Burstein HJ, Allred DC, Vogel CL, Goldstein LJ, Somlo G, Gradishar WJ, Hudis CA, Jahanzeb M, Stark A, Wolff AC, Press MF, Winer EP, Paik S, Ljung BM, Force NHTiBCT (2006) HER2 testing in breast cancer: NCCN Task Force report and recommendations. J Natl Compr Cancer Netw 4(Suppl 3):S1–S22 quiz S23-24 Google Scholar
  15. 15.
    Romond EH, Perez EA, Bryant J, Suman VJ, Geyer CE Jr, Davidson NE, Tan-Chiu E, Martino S, Paik S, Kaufman PA, Swain SM, Pisansky TM, Fehrenbacher L, Kutteh LA, Vogel VG, Visscher DW, Yothers G, Jenkins RB, Brown AM, Dakhil SR, Mamounas EP, Lingle WL, Klein PM, Ingle JN, Wolmark N (2005) Trastuzumab plus adjuvant chemotherapy for operable HER2-positive breast cancer. N Engl J Med 353(16):1673–1684. doi: 10.1056/NEJMoa052122 CrossRefPubMedGoogle Scholar
  16. 16.
    Paik S, Bryant J, Tan-Chiu E, Romond E, Hiller W, Park K, Brown A, Yothers G, Anderson S, Smith R, Wickerham DL, Wolmark N (2002) Real-world performance of HER2 testing–National Surgical Adjuvant Breast and Bowel Project experience. J Natl Cancer Inst 94(11):852–854CrossRefPubMedGoogle Scholar
  17. 17.
    Stoss OC, Scheel A, Nagelmeier I, Schildhaus HU, Henkel T, Viale G, Jasani B, Untch M, Ruschoff J (2015) Impact of updated HER2 testing guidelines in breast cancer-re-evaluation of HERA trial fluorescence in situ hybridization data. Mod Pathol. doi: 10.1038/modpathol.2015.112 PubMedGoogle Scholar
  18. 18.
    Long TH, Lawce H, Durum C, Moore SR, Olson SB, Gatter K, Troxell ML (2015) The new equivocal: changes to HER2 FISH results when applying the 2013 ASCO/CAP guidelines. Am J Clin Pathol 144(2):253–262. doi: 10.1309/AJCP3Q9WFOQTKUVV CrossRefPubMedGoogle Scholar
  19. 19.
    Bethune GC, van Zanten VD, MacIntosh RF, Rayson D, Younis T, Thompson K, Barnes PJ (2015) Impact of the 2013 American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 (HER2) testing of invasive breast carcinoma: a focus on tumours assessed as ‘equivocal’ for HER2 gene amplification by fluorescence in situ hybridization. Histopathology. doi: 10.1111/his.12723 PubMedGoogle Scholar
  20. 20.
    Espinet B, Puiggros AM, Corominas JM et al (2015) Increase in equivocal in situ hybridization results in ASCO/CAP guidelines for HER2 testing in invasive breast cancer: comparison with the 2007 criteria. Mod Pathol 28:30–75. doi: 10.1038/modpathol.2015.9 Google Scholar
  21. 21.
    Andrade R, Barrera-Herrera L, Plata A, Lopez R, Carvajal M (2015) Impact of the 2013 HER2 guidelines recommendations by FISH, retrospective analysis of 1893 cases at Fundacion Santa Fe De Bogota. Mod Pathol 28:30–75. doi: 10.1038/modpathol.2015.9 Google Scholar
  22. 22.
    Varga Z, Noske A (2015) Impact of modified 2013 ASCO/CAP guidelines on HER2 testing in breast cancer. One year experience. PLoS One 10(10):e0140652. doi: 10.1371/journal.pone.0140652 CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Fulton R, Gown A (2015) Use of 2013 ASCO-CAP breast carcinoma HER2 scoring guidelines results in shifts of scoring categories with slight change in IHC-FISH concordance. Mod Pathol 28(30):75. doi: 10.1038/modpathol.2015.9 Google Scholar
  24. 24.
    Onguru O, Zhang P (2015) How do the differences between 2013 and 2007 ASCO/CAP HER2 guidelines affect HER2 status in breast carcinomas? Mod Pathol 28:30–75. doi: 10.1038/modpathol.2015.9 Google Scholar
  25. 25.
    Muller KE, Marotti JD, Memoli VA, Wells WA, Tafe LJ (2015) Impact of the 2013 ASCO/CAP HER2 guideline updates at an academic medical center that performs primary HER2 FISH testing: increase in equivocal results and utility of reflex immunohistochemistry. Am J Clin Pathol 144(2):247–252. doi: 10.1309/AJCPE5NCHWPSMR5D CrossRefPubMedGoogle Scholar
  26. 26.
    Vanden Bempt I, Van Loo P, Drijkoningen M, Neven P, Smeets A, Christiaens MR, Paridaens R, De Wolf-Peeters C (2008) Polysomy 17 in breast cancer: clinicopathologic significance and impact on HER-2 testing. J Clin Oncol 26(30):4869–4874. doi: 10.1200/JCO.2007.13.4296 CrossRefPubMedGoogle Scholar
  27. 27.
    Viale G (2009) Be precise! The need to consider the mechanisms for CEP17 copy number changes in breast cancer. J Pathol 219(1):1–2. doi: 10.1002/path.2593 CrossRefPubMedGoogle Scholar
  28. 28.
    Marchio C, Lambros MB, Gugliotta P, Di Cantogno LV, Botta C, Pasini B, Tan DS, Mackay A, Fenwick K, Tamber N, Bussolati G, Ashworth A, Reis-Filho JS, Sapino A (2009) Does chromosome 17 centromere copy number predict polysomy in breast cancer? A fluorescence in situ hybridization and microarray-based CGH analysis. J Pathol 219(1):16–24. doi: 10.1002/path.2574 CrossRefPubMedGoogle Scholar
  29. 29.
    Moelans CB, de Weger RA, van Diest PJ (2010) Absence of chromosome 17 polysomy in breast cancer: analysis by CEP17 chromogenic in situ hybridization and multiplex ligation-dependent probe amplification. Breast Cancer Res Treat 120(1):1–7. doi: 10.1007/s10549-009-0539-2 CrossRefPubMedGoogle Scholar
  30. 30.
    Tse CH, Hwang HC, Goldstein LC, Kandalaft PL, Wiley JC, Kussick SJ, Gown AM (2011) Determining true HER2 gene status in breast cancers with polysomy by using alternative chromosome 17 reference genes: implications for anti-HER2 targeted therapy. J Clin Oncol 29(31):4168–4174. doi: 10.1200/JCO.2011.36.0107 CrossRefPubMedGoogle Scholar
  31. 31.
    Hofmann M, Stoss O, Gaiser T, Kneitz H, Heinmoller P, Gutjahr T, Kaufmann M, Henkel T, Ruschoff J (2008) Central HER2 IHC and FISH analysis in a trastuzumab (Herceptin) phase II monotherapy study: assessment of test sensitivity and impact of chromosome 17 polysomy. J Clin Pathol 61(1):89–94. doi: 10.1136/jcp.2006.043562 CrossRefPubMedGoogle Scholar
  32. 32.
    Reinholz M, Jenkins RB, Hillman D et al (2007) The clinical significance of polysomy 17 in the ERBB2+ N9831 intergroup adjuvant trastuzumab trial. Breast Cancer Res Treat 106(supp 1):11Google Scholar
  33. 33.
    Bartlett JM, Munro AF, Dunn JA, McConkey C, Jordan S, Twelves CJ, Cameron DA, Thomas J, Campbell FM, Rea DW, Provenzano E, Caldas C, Pharoah P, Hiller L, Earl H, Poole CJ (2010) Predictive markers of anthracycline benefit: a prospectively planned analysis of the UK National Epirubicin Adjuvant Trial (NEAT/BR9601). Lancet Oncol 11(3):266–274. doi: 10.1016/S1470-2045(10)70006-1 CrossRefPubMedGoogle Scholar
  34. 34.
    Jimenez RE, Wallis T, Tabasczka P, Visscher DW (2000) Determination of Her-2/Neu status in breast carcinoma: comparative analysis of immunohistochemistry and fluorescent in situ hybridization. Mod Pathol 13(1):37–45. doi: 10.1038/modpathol.3880007 CrossRefPubMedGoogle Scholar
  35. 35.
    Persons DL, Tubbs RR, Cooley LD, Dewald GW, Dowling PK, Du E, Mascarello JT, Rao KW, Wilson KS, Wolff DJ, Habegger-Vance G (2006) HER-2 fluorescence in situ hybridization: results from the survey program of the College of American Pathologists. Arch Pathol Lab Med 130(3):325–331. doi: 10.1043/1543-2165(2006)130[325:HFISHR]2.0.CO;2 PubMedGoogle Scholar
  36. 36.
    Vera-Roman JM, Rubio-Martinez LA (2004) Comparative assays for the HER-2/neu oncogene status in breast cancer. Arch Pathol Lab Med 128(6):627–633. doi: 10.1043/1543-2165(2004)128<627:CAFTNO>2.0.CO;2 PubMedGoogle Scholar
  37. 37.
    Kos Z, Craddock K, Berman H et al (2015) Updated HER2 in situ hybridization (ISH) guidelines in breast carcinoma: some equivocal findings. Mod Pathol 28:30–75. doi: 10.1038/modpathol.2015.9 Google Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Kamaljeet Singh
    • 1
    Email author
  • Umadevi Tantravahi
    • 2
  • Michele M. Lomme
    • 1
  • Terese Pasquariello
    • 3
  • Margaret Steinhoff
    • 1
  • C. James Sung
    • 1
  1. 1.Department of Pathology and Laboratory Medicine, Women & Infants HospitalAlpert Medical School of Brown UniversityProvidenceUSA
  2. 2.Pathology and Laboratory Medicine, Cytogenetics and Molecular Laboratory, Women & Infants HospitalAlpert Medical School of Brown UniversityProvidenceUSA
  3. 3.Immunohistochemistry Laboratory, Department of Pathology and Laboratory MedicineWomen & Infants HospitalProvidenceUSA

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