Abstract
Trastuzumab is the cornerstone for treatment of women with HER2-overexpressing breast cancer, both in the adjuvant and in the metastatic settings. The accurate assessment of HER2 is, therefore, critical to identifying patients who may benefit from trastuzumab-based therapy. This project aimed to determine the optimal scoring method for fluorescence in situ hybridization (FISH) assay. FISH assay was done on 893 samples of breast cancer. Three scoring methods were evaluated: Her2/CEP17> or =2, Her2>4, or Her2>6. Protein and gene expression were evaluated by immunohistochemistry (n = 584) and mRNA/assay/nucleic acid sequence-based amplification (NASBA; n = 90). Samples were divided into five groups based on FISH results: disomic amplified and nonamplified, polysomic amplified, nonamplified, and discordant (10.8% of cases, mostly positive with Her2>4 scoring, but negative with the others). Her2/CEP17> or =2 and Her2>6 scoring methods showed the best association (a) with regard to FISH scoring (kappa = 0.906, P < 10(-6)) and (b) between FISH and immunohistochemistry (3+ as positive; kappa > 0.650, P < 10(-6)) or NASBA (kappa > 0.536, P < 10(-6)). Polysomy had an effect on Her2 copy number (P < 10(-6)), but had no effect on protein and mRNA content. Therefore, within the discordant subgroup, for which additive Her-2 gene copies are due to high polysomy, protein and mRNA levels were similar to those of the nonamplified samples. For this subgroup, the best concordance between FISH/immunohistochemistry/NASBA was observed with the Her2/CEP17 ratio and Her-2>6 scoring (68% and 58% perfect matches, respectively). No perfect matches were observed using the Her2>4 scoring method. Correction for chromosome-17 is the method of choice for clinical practice; Her-2>6, but not Her-2>4, could be used as an alternative.
Highlights
The Her-2/neu oncogene, located on chromosome-17, encodes a transmembrane-tyrosine kinase receptor protein belonging to the epidermal growth factor receptor family
To determine the optimal fluorescence in situ hybridization (FISH) scoring system to select patients that might benefit from a trastuzumab-based treatment, three criteria for FISH positivity were considered, and the ensuing status was compared with immunohistochemistry and mRNA/nucleic acid sequence – based amplification (NASBA) data, two techniques evaluating HER-2 expression status at a different level and whose results seemed to be significantly associated
Similar to previous studies [26, 27], we found a good concordance between Her-2/CEP17z2 ratio and Her-2>4 FISH scoring methods, for disomic and low polysomic cases, as well as for cases with increased Her-2 gene copies consecutive to gene amplification
Summary
The Her-2/neu oncogene, located on chromosome-17, encodes a transmembrane-tyrosine kinase receptor protein belonging to the epidermal growth factor receptor family. Overexpression and amplification have been linked to poor prognosis and response to therapy with the anti – HER-2-humanized monoclonal antibody, trastuzumab (Herceptin), in patients with advanced metastatic breast cancer, when used either as single agent [1, 2] or in combination with chemotherapy [3]. HercepTest (Dako), a more standardized immunohistochemical method, was developed to overcome or decrease this variability, with sometimes disappointing results [9]. These concerns regarding the accuracy of immunohistochemistry using standard formalin-fixed paraffin-embedded tissue sections [10] have stimulated the use of fluorescence in situ hybridization (FISH) assay.
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