FAQs

Questions and Answers on HER2 Testing in Clinical Practice

  • Q: Does HER2 status have prognostic or predictive value in breast cancer?
  • A: HER2 is recognized as an important predictive and prognostic factor in breast cancer.6,8 HER2 gene amplification is a permanent genetic change that results in the continuous overexpression of the HER2 receptor (HER2 protein).5,10 Several studies have shown that HER2 overexpression (either extra copies of the gene itself, or an excess amount of the gene’s protein product) is associated with decreased overall survival.11,13 HER2 status is predictive of a response to Herceptin.1 In clinical trials, in patients with HER2 positive metastatic breast cancer, Herceptin achieved significant survival benefits when used first-line in combination with chemotherapy until disease progression.1,3 A retrospective analysis of 660 of 691 (96%) patients enrolled in the clinical studies (whose tumors scored 2+ or 3+ by IHC) suggested that the beneficial treatment effects were greatest in patients whose tumors tested FISH+.1 Median survival increased to 26.2 months for FISH+ patients receiving Herceptin plus chemotherapy compared with 20.0 months for FISH+ patients receiving chemotherapy alone.2 The results for patients testing IHC+ in the pivotal trial were 25.1 months for Herceptin plus chemotherapy versus 20.3 months for chemotherapy alone.3 This is consistent with previous studies that have shown that the clinical benefit of Herceptin was greater for patients whose tumors scored IHC3+ for HER2.1
    Patients with HER2-driven disease often have a poor response to standard chemotherapy and hormonal therapy.12,14 Two recent studies of metastatic breast cancer have suggested that HER2-overexpressing tumors are less responsive to treatment with hormonal therapies such as tamoxifen.15,16 In addition, some data suggest that HER2-positive cancers do not respond well to CMF-based therapies.7,14 The management of this aggressive disease presents a unique treatment challenge.

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  • Q: Which HER2 tests are FDA approved to select patients for treatment with Herceptin?
  • A: Several FDA-approved commercial assays are available to determine HER2 status and aid in the selection of patients for Herceptin. Approved immunohistochemistry (IHC) assays, which measure the level of expression of the HER2 protein, include HercepTest® and Pathway™.1 Approved FISH (fluorescence in situ hybridization) assays,which detect the underlying gene alteration in the patient's tumor cells, include PathVysion® and HER2 FISH pharmDx™.1 Users should refer to the package inserts of specific assay kits for information on the validation and performance of each assay.

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  • Q: How does IHC testing work?
  • A: In HER2-positive tumors, there is an excess amount of the HER2 protein on the cell surface. This is referred to as HER2 overexpression. IHC assays measure the amount of HER2 protein expressed on the surface of tumor cells. This technique uses antibodies to HER2 and a chemical detection method to stain HER2 protein in the breast tissue samples. IHC tests are scored on a scale of 0 to 3+ based on the reviewer's interpretation of staining intensity and completeness of cell membrane staining.9 The test is usually performed on formalin-fixed, paraffin-embedded tissues.

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  • Q: How do FISH tests work?
  • A: In HER2-positive tumors, there are 2 or more copies of the HER2/neugene per chromosome 17, i.e., there is gene amplification of HER2/neu.4 FISH assays measure the number of HER2/neu gene copies.1 The test includes an internal control, a chromosome 17 control probe that allows one to determine if the increased copy number is due to amplification of the HER2/neu gene or to extra copies of a chromosome on which the gene resides. In a normal cell, there are 2 copies of a chromosome and one copy of each gene per chromosome. FISH test results are determined by taking the ratio of the number of HER2 signals to the number of chromosome 17 signals among 20 interphase nuclei in tumor cells. Normal specimens show a ratio of <2.0, while specimens with amplification of HER2/neu have a ratio of greater than or equal to 2.0 and are HER2-positive.4

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  • Q: Which variables can affect the performance of IHC and FISH?
  • A: IHC detects HER2 overexpression at the protein level, and may be affected by conditions of the testing procedures. These include: time to fixation, duration of fixation, processing, denaturation, heating, antigen retrieval, the staining procedure used, and the interpretation of staining.17,18 Although there are antigen retrieva ltechniques in use, these may result in false-positive IHC results. FISH measures HER2 DNA. Some fixatives, chemicals or heat, may interfere with the FISH assay. However, an internal control is used to distinguish between a FISH-negative and a non-informative result.4

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  • Q: How are test results interpreted by a board-certified pathologist?
  • A: Interpretation of IHC relies on a qualitative scoring system on a scale of 0 to 3+. A patient may score a 0 (negative), 1+ (negative), 2+ (borderline), or 3+ (positive) on an IHC test based on the reviewer's interpretation of staining intensity and completeness of membrane staining.9 With FISH testing, the results are quantitative instead of qualitative; tumors are interpreted as HER2 "negative" or "positive" by enumerating the HER2/neu gene copy number.4 Limitations in assay precision (particularly for the IHC method) and in the direct linkage between assay result and overexpression of the Herceptin target (for the FISH method) make it inadvisable to rely on a single method to rule out potential Herceptin benefit.1

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  • Q: If a patient is IHC 2+ or 3+ and FISH-, are they HER2+?
  • A: Overexpression of the HER2 protein rarely occurs in the absence of gene amplification.19 FISH analysis reveals that some patients with apparent protein overexpression (IHC 2+ or 3+) do not have gene amplification (FISH-), suggesting that these patients may be "false positives." Approximately 2%-4% of patients who demonstrated HER2 protein overexpression by molecular techniques did not have gene amplification.19,20 In current laboratory testing, variability in pre-analytical tissue processing, reagent variability, antigen retrieval, and scoring may result in IHC false-positives.17,18

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  • Q: How can I help to ensure an accurate HER2 test result?
  • A: HER2 testing should be performed by laboratories with demonstrated proficiency in the specific technology being utilized. Improper assay performance may result from the use of suboptimally fixed tissue, failure to utilize specified reagents, deviation from specific assay instructions, and failure to include appropriate controls for assay validation and can lead to unreliable results.1,21 Assessment for HER2 expression or gene amplification should be performed by laboratories with demonstrated proficiency in the specific technology being utilized.1 Recent findings suggest the need to improve quality control measures in laboratories that use IHC assays, including periodic testing for concordance with FISH. These studies also suggest that large-volume reference laboratories performing HER2 tests are more reliable than small-volume laboratories.17,21

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  • Adjuvant indications
  • Herceptin is indicated for adjuvant treatment of HER2-overexpressing node-positive or node-negative (ER/PR-negative or with one high-risk feature)† breast cancer:
  • As part of a treatment regimen containing doxorubicin, cyclophosphamide, and either paclitaxel or docetaxel
  • With docetaxel and carboplatin
  • As a single agent following multi-modality anthracycline-based therapy

† High-risk is defined as ER/PR positive with one of the following features: tumor size >2 cm, age <35 years, or tumor grade 2 or 3.

  • Metastatic indications
  • Herceptin is indicated:
  • In combination with paclitaxel for first-line treatment of HER2-overexpressing metastatic breast cancer
  • As a single agent for treatment of HER2-overexpressing breast cancer in patients who have received one or more chemotherapy regimens for metastatic disease

Boxed WARNINGS and Additional Important Safety Information

  • Cardiotoxicity
  • Herceptin administration can result in sub-clinical and clinical cardiac failure manifesting as congestive heart failure (CHF) and decreased left ventricular ejection fraction (LVEF).
    • The incidence and severity of left ventricular cardiac dysfunction was highest in patients who received Herceptin concurrently with anthracycline-containing chemotherapy regimens.
    • Discontinue Herceptin treatment in patients receiving adjuvant therapy for breast cancer and strongly consider discontinuation of Herceptin in patients with metastatic breast cancer who develop a clinically significant decrease in left ventricular function.
  • Herceptin can cause left ventricular cardiac dysfunction, arrhythmias, hypertension, disabling cardiac failure, cardiomyopathy, and cardiac death.
    • In one adjuvant clinical trial, cardiac ischemia or infarction occurred in the Herceptin-containing regimens.
    • Herceptin can also cause asymptomatic decline in LVEF.
  • Cardiac Monitoring
  • Candidates for treatment with Herceptin should undergo a thorough baseline cardiac assessment, including history, physical examination, and an assessment of LVEF by echocardiogram or MUGA scan.
    • Patients should undergo frequent monitoring for decreased left ventricular function during and after Herceptin treatment.
    • More frequent monitoring should be employed if Herceptin is withheld in patients who develop significant left ventricular cardiac dysfunction.
  • Infusion reactions and Pulmonary Toxicity
  • Serious infusion reactions and pulmonary toxicity have occurred; fatal infusion reactions have been reported.
    • In most cases, symptoms occurred during or within 24 hours of administration of Herceptin. Herceptin infusion should be interrupted for patients experiencing dyspnea or clinically significant hypotension.
    • Patients should be monitored until signs and symptoms completely resolve.
    • Discontinue Herceptin for infusion reactions manifesting as anaphylaxis, angioedema, interstitial pneumonitis, or acute respiratory distress
  • Infusion reactions consist of a symptom complex characterized by fever and chills, and on occasion include nausea, vomiting, pain (in some cases at tumor sites), headache, dizziness, dyspnea, hypotension, rash, and asthenia.
    • In postmarketing reports, serious and fatal infusion reactions have been reported. Discontinue Herceptin in all patients with severe or life-threatening infusion reactions.
  • Herceptin use can result in serious and fatal pulmonary toxicity, which includes dyspnea, interstitial pneumonitis, pulmonary infiltrates, pleural effusions, non-cardiogenic pulmonary edema, pulmonary insufficiency and hypoxia, acute respiratory distress syndrome, and pulmonary fibrosis.
    • Such events can occur as sequelae of infusion reactions.
    • Patients with symptomatic intrinsic lung disease or with extensive tumor involvement of the lungs, resulting in dyspnea at rest, appear to have more severe toxicity.
  • Neutropenia
  • Exacerbation of chemotherapy-induced neutropenia has also occurred
    • In controlled clinical trials, severe neutropenia and febrile neutropenia occurred more frequently in metastatic breast cancer patients receiving Herceptin with myelosuppressive chemotherapy compared to chemotherapy alone.
    • The incidence of septic death was not significantly increased.
  • Pregnancy Category D
  • Herceptin can cause fetal harm when administered to a pregnant woman.
  • Post-marketing reports suggest that Herceptin use during pregnancy increases the risk of oligohydramnios during the second and third trimester.
  • Most Common Adverse Events
  • The most common adverse reactions associated with Herceptin use were fever, nausea, vomiting, infusion reactions, diarrhea, infections, increased cough, headache, fatigue, dyspnea, rash, neutropenia, anemia, and myalgia.

Please see the Herceptin full Prescribing Information including Boxed WARNINGS and additional important safety information.

  • References:
  • 1. Herceptin Prescribing Information. Genentech, Inc. March 2009.
  • 2. Mass Robert D, Press MF, Anderson S, et al. Evaluation of Clinical Outcomes According to HER2 Detection by Fluorescence In Situ Hybridization in Women with Metastatic Breast Cancer Treated with Trastuzumab. Clinical Breast Cancer, 2005; Vol. 6, No. 3: 240-246.
  • 3. Slamon DJ, Leyland-Jones B, Shak S, et al. Use of chemotherapy plus a monoclonal antibody against HER2 for metastatic breast cancer that overexpresses HER2. N Engl J Med. 2001;344:783-792.
  • 4. PathVysion® HER-2 DNA Probe Kit Package Insert, Vysis, Inc.; April 2007.
  • 5. Simon R, Nocito A, Hübscher T, et al. Patterns of HER-2/neu amplification and overexpression in primary and metastatic breast cancer. J Natl Cancer Inst. 2001;93:1141-1146.
  • 6. Slamon DJ, Godolphin W, Jones LA, et al. Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer. Science. 1989;244:707-712.
  • 7. Gusterson BA, Gelber RD, Goldhirsch A, et al, for the International (Ludwig) Breast Cancer Study Group. Prognostic importance of c-erbB-2 expression in breast cancer. J Clin Oncol. 1992;10:1049-1056.
  • 8. Sjögren S, Inganäs M, Lindgren A, et al. Prognostic and predictive value of c-erbB-2 overexpression in primary breast cancer, alone and in combination with other prognostic markers. J Clin Oncol. 1998;16:462-469.
  • 9. DAKO HercepTest® Information Web site. Summary of Procedure. Available at: http://www.dako.com/prod_productrelatedinformation?url=support_ herceptest_elearning_interpreting_guidelin.htm. Accessed September 2, 2008.
  • 10. Sliwkowski MX, Lofgren JA, Lewis GD, et al. Nonclinical studies addressing the mechanism of action of trastuzumab (Herceptin). Semin Oncol. 1999;26(suppl 12):60-70.
  • 11. Slamon DJ, Clark GM, Wong SG, et al. Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science. 1987;235:177-182.
  • 12. Lohrisch C, Piccart M. HER2/neu as a predictive factor in breast cancer. Clin Breast Cancer. 2001;2:129-135.
  • 13. Paik S, Hazan R, Fischer ER, et al. Pathologic findings from the National Surgical Adjuvant Breast and Bowe l Project: prognostic significance of erbB-2 protein overexpression in primary breast cancer. J Clin Oncol. 1990;8:103-112.
  • 14. Ross JS, Fletcher JA. HER-2/neu (c-erbB-2) gene and protein in breast cancer. Am J Clin Pathol. 1999;112(suppl 1):S53-S67.
  • 15. Leitzel K, Teramoto Y, Konrad K, et al. Elevated serum c-erbB-2 antigen levels and decreased response to hormone therapy of breast cancer. J Clin Oncol. 1995;13:1129-1135.
  • 16. Houston SJ, Plunkett TA, Barnes DM, et al. Overexpression of c-erbB-2 is an independent marker of resistance of c-erbB-2 expression in breast cancer. International (Ludwig) Breast Cancer Study Group. J Clin Oncol. 1992;10:1049-1056.
  • 17. Paik S, Bryant J, Tan-Chiu E, et al. Real-world performance of HER2 testing -National Surgical Adjuvant Breast and Bowel Project experience. J Natl Cancer Inst. 2002;94:852-854.
  • 18. O'Leary TJ. Standardization of immunohistochemistry. Appl Immunohistochem Mol Morphol. 2001;9:3-8.
  • 19. Pauletti G, Godolphin W, Press MF, Slamon DJ. Detection and quantitation of HER-2/neu gene amplification in human breast cancer archival material using fluorescence in situ hybridization. Oncogene. 1996;13:63-72.
  • 20. Kallioniemi O-P, Kallioniemi A, Kurisu W, et al. ERBB2 amplification in breast cancer analyzed by fluorescence in situ hybridization. Proc Natl Acad Sci U S A. 1992;89:5321-5325.
  • 21. Roche PC, Suman VJ, Jenkins RB, et al. Concordance between local and central laboratory HER2 testing in the Breast Intergroup Trial N9831. J Natl Cancer Inst. 2002;94:855-857.


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