title: Simultaneous detection of HER2/neu gene amplification and protein overexpression in paraffin-embedded breast cancer.
abstract: The determination of HER2/neu status in breast carcinomas has become essential for the selection of breast cancer patients for Herceptin therapy. Herceptin treatment is used in patients with metastatic breast carcinoma with HER2/neu protein overexpression detected by immunohistochemistry (IHC) or gene amplification analysed by fluorescence in situ hybridization (FISH). A multiparametric fluorescent approach based on the simultaneous detection of HER2/neu gene amplification and protein expression was established to increase the accuracy, and to improve the reproducibility, of HER2/neu diagnostics. Based on four paraffin-embedded breast cancer cell lines, a combined fluorescent immunostaining (FIHC) and FISH method was developed by using the PathVysion HER2 DNA Probe Kit (VYSIS) and the polyclonal antibody from the HercepTest (DAKO). Diagnostic applicability was documented on 215 formalin-fixed primary breast carcinomas. Criteria for immunofluorescence quantification were chosen by analogy with the FDA-approved HercepTest scoring, ranging from 0 to 3+. There was 97.7% concordance between conventional IHC and fluorescence IHC. The FISH data resulting from the multiparametric approach did not differ from conventional FISH. Breast carcinomas with HER2/neu protein overexpression and simultaneous gene amplification were detected with 100% sensitivity. In addition, five of the 215 cases (2.3%) had HER2/neu gene amplification without protein overexpression. The main advantage of this novel approach is that polysomy, aneuploidy, gene amplification, and protein content can be analysed simultaneously in the same cell.
authors.affiliation: Institute of Pathology, University of Regensburg, 93042 Regensburg, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Lottner
authors.forename: Stephan
authors.initials: S
authors.lastname: Schwarz
authors.forename: Simone
authors.initials: S
authors.lastname: Diermeier
authors.forename: Arndt
authors.initials: A
authors.lastname: Hartmann
authors.forename: Ruth
authors.initials: R
authors.lastname: Knuechel
authors.forename: Ferdinand
authors.initials: F
authors.lastname: Hofstaedter
authors.forename: Gero
authors.initials: G
authors.lastname: Brockhoff
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000911:Antibodies, Monoclonal / Q000627:therapeutic use; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents / Q000627:therapeutic use; D014408:Biomarkers, Tumor / Q000378:metabolism*; D001943:Breast Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique / Q000379:methods; D018734:Genes, erbB-2; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008207:Lymphatic Metastasis; D008875:Middle Aged; D016612:Paraffin Embedding; D018579:Patient Selection; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism*; D015203:Reproducibility of Results; D000068878:Trastuzumab; D014407:Tumor Cells, Cultured
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D014408:Biomarkers, Tumor; D018719:Erb-b2 Receptor Tyrosine Kinases; D000068878:Trastuzumab