Abstract
García-Caballero T, Grabau D, Green A R, Gregory J, Schad A, Kohlwes E, Ellis I O, Watts S & Mollerup J (2010) Histopathology56, 472–480 Determination of HER2 amplification in primary breast cancer using dual-colour chromogenic in situ hybridization is comparable to fluorescence in situ hybridization: a European multicentre study involving 168 specimensAims:Fluorescence in situ hybridization (FISH) can be used to reveal several genomic imbalances relevant to proper cancer diagnosis and to the correct treatment regime. However, FISH requires expensive and advanced fluorescence microscopes in addition to expertise in fluorescence microscopy. To determine whether a newly developed dual-colour chromogenic in situ hybridization (CISH) method is a suitable alternative to FISH, we analysed the human epidermal growth factor receptor 2 gene (HER2) amplification level of 168 breast cancer specimens using dual-colour CISH and FISH and compared the results.Methods and results:We found 100% agreement between HER2 status determined by FISH and dual-colour CISH. Furthermore, we observed that the time used to score slides was significantly reduced by 28% in dual-colour CISH compared with the FISH protocol. Concordance between HER2 protein status and dual-colour CISH or FISH was equally good with an overall agreement of 96.8%. Correlation between the HER2/centromere 17 gene ratios obtained with dual-colour CISH and FISH was highly significant with an overall correlation coefficient (ρ) of 0.96.Conclusions:We conclude that dual-colour CISH and bright field microscopy are excellent alternatives to FISH when analysing the HER2 status of primary breast cancer.
Highlights
Fluorescence in situ hybridization (FISH) analysis is based on specific recognition of denatured target DNA sequences by fluorescent labelled sequence pairing probes
Gene copy numbers correspond to the mean copy number per cell of the 20 nuclei counted (n = 167). *Significantly different from the FISH human growth factor receptor 2 (HER2):centromere 17 (CEN-17) ratio (P < 0.001, paired two-tailed t-test). †Significantly different from the FISH HER2 copy number (P < 0.001, paired two-tailed t-test)
Identification of breast cancer patients that may benefit from treatment with trastuzumab or lapatinib (TykerbÒ; GlaxoSmithKline, Brentford, UK),[11,15] and the response to endocrine treatment of metastatic breast cancer appears to be dependent on HER2 status.[16,17]
Summary
Fluorescence in situ hybridization (FISH) analysis is based on specific recognition of denatured target DNA sequences by fluorescent labelled sequence pairing probes. The large potential for diagnostic and prognostic use of FISH is not implemented in all routine pathological laboratories This could be due to disadvantages previously put forward,[1,2,3,4] which include certain fixatives interfering with fluorescence detection and limited community experience with tissue-based FISH. Comparisons of FISH with chromogenic in situ hybridization (CISH) have revealed several advantages with the use of CISH.[2,3] The CISH technique is a simple extension of the FISH protocol that allows bright field microscopic evaluation of slides with the additional benefit that morphological features can be observed, and that archiving of slides is possible. Conversion of the FISH to a CISH signal is achieved using fluorochrome- or hapten-specific antibodies as an extension to the FISH probes These antibodies are conjugated to an enzymatic marker such as horseradish peroxidase (HRP) or alkaline phosphatase (AP). We compared the results obtained with FISH and a newly available dual-colour CISH assay for the determination of HER2 status in 168 cases of primary breast cancer
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