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HER2 gene amplification in patients with breast cancer with equivocal IHC results
  1. Sybren L Meijer1,
  2. Jelle Wesseling2,3,
  3. Vincent T Smit4,
  4. Petra M Nederlof2,
  5. Gerrit K J Hooijer1,
  6. Henrique Ruijter2,
  7. Jan Willem Arends5,
  8. Mike Kliffen6,
  9. Joost M van Gorp7,
  10. Lotus Sterk8,
  11. Marc J van de Vijver1
  1. 1Departments of Pathology of the Academic Medical Center, Amsterdam, The Netherlands
  2. 2The Netherlands Cancer Institute, Amsterdam, The Netherlands
  3. 3University Medical Center, Groningen, The Netherlands
  4. 4Leiden University Medical Center, Leiden, The Netherlands
  5. 5Deventer Hospital, Deventer, The Netherlands
  6. 6Erasmus University Medical Center, Rotterdam, The Netherlands
  7. 7Diakonessenhuis Utrecht, Utrecht, The Netherlands
  8. 8Laboratory of Pathology, East Netherland, Enschede, The Netherlands
  1. Correspondence to Marc J van de Vijver, Department of Pathology, Academic Medical Center Meibergdreef 9, Amsterdam 1105 AZ, Netherlands; m.j.vandevijver{at}


Aims Equivocal human epidermal growth factor receptor 2 protein (HER2) (2+) immunohistochemistry (IHC) is subject to significant interobserver variation and poses a challenge in obtaining a definitive positive or negative test result. This equivocal test result group accounts for approximately 15% of all tumours, and for optimal guidance of HER2 targeted therapy, a further analysis of quantification of gene copy number and amplification status is needed for patients with early or metastatic breast cancer.

Methods 553 breast-cancer specimens with equivocal HER2 IHC(2+) test results were collected and subsequently centrally retested by chromogenic in situ hybridisation (CISH), and HER2 gene copy numbers per tumour cell nucleus were determined.

Results Using CISH, 77 of 553 equivocal HER2 IHC(2+) test result cases (13.9% of total) showed high levels of HER2 gene amplification (≥10.0 gene copies per nucleus), and 41 of 553 (7.4% of total) showed low-level HER2 gene amplification (6.0–9.9 gene copies per nucleus). In 73.6% of cases, no amplification of the HER2 gene was shown, and in only 4.9% of cases was an equivocal test result by CISH observed (4.0–5.9 gene copies per nucleus).

Conclusions Testing by CISH of all equivocal HER2 IHC(2+) test result provides a definitive guidance in HER2 targeted therapy in 95.1% of cases. A significant proportion (21.3%) of patients with equivocal IHC(2+) test results show amplification of the HER2 gene.

  • Breast cancer
  • HER2
  • equivocal test result
  • immunohistochemistry
  • CISH
  • breast cancer
  • breast pathology
  • GI neoplasms
  • gynaecological pathology
  • molecular oncology
  • molecular pathology
  • cancer research
  • image analysis
  • oncology
  • comparative genomic hybridisation
  • cancer genetics
  • diagnostics
  • gene amplification
  • familial cancers
  • colorectal cancer
  • gall bladder
  • p53
  • pancreas
  • cerb 2
  • immunocytochemistry
  • in situ hybridisation

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  • Funding The study was supported by an unrestricted educational grant from F Hoffmann-La Roche AG.

  • Competing interests MvdV is a member of the F Hoffmann-La Roche AG Herceptin Pathology Advisory Board and has received honoraria for attendance at these meetings.

  • Provenance and peer review Not commissioned; externally peer reviewed.