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Evaluation of HER-2/neu Gene Amplification and Overexpression: Comparison of Frequently Used Assay Methods in a Molecularly Characterized Cohort of Breast Cancer Specimens

From the Breast Cancer Research Program of the Lee Breast Center, Department of Pathology and Department of Preventive Medicine, Norris Comprehensive Cancer Center, University of Southern California; Division of Hematology and Oncology, University of California Los Angeles School of Medicine, Los Angeles, CA; and Vysis, Inc, Downers Grove, IL.

Address reprint requests to: Michael F. Press, MD, PhD, Norris Topping Tower Room 5409, 1441 Eastlake Ave, U.S.C./Norris Comprehensive Cancer Center, Los Angeles, CA 90033; email: press@ hsc.usc.edu.

PURPOSE: To compare and evaluate HER-2/neu clinical assay methods.

MATERIALS AND METHODS: One hundred seventeen breast cancer specimens with known HER-2/neu amplification and overexpression status were assayed with four different immunohistochemical assays and two different fluorescence in situ hybridization (FISH) assays.

RESULTS: The accuracy of the FISH assays for HER-2/neu gene amplification was high, 97.4% for the Vysis PathVision assay (Vysis, Inc, Downers Grove, IL) and 95.7% for the the Ventana INFORM assay (Ventana, Medical Systems, Inc, Tucson, AZ). The immunohistochemical assay with the highest accuracy for HER-2/neu overexpression was obtained with R60 polyclonal antibody (96.6%), followed by immunohistochemical assays performed with 10H8 monoclonal antibody (95.7%), the Ventana CB11 monoclonal antibody (89.7%), and the DAKO HercepTest (88.9%; Dako, Corp, Carpinteria, CA). Only the sensitivities, and therefore, overall accuracy, of the DAKO Herceptest and Ventana CB11 immunohistochemical assays were significantly different from the more sensitive FISH assay.

CONCLUSION: Based on these findings, the FISH assays were highly accurate, with immunohistochemical assays performed with R60 and 10H8 nearly as accurate. The DAKO HercepTest and the Ventana CB11 immunohistochemical assay were statistically significantly different from the Vysis FISH assay in evaluating these previously molecularly characterized breast cancer specimens.

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