Quantitative real-time RT-PCR and chromogenic in situ hybridization: precise methods to detect HER-2 status in breast carcinomaReport as inadecuate

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BMC Cancer

, 9:90

First Online: 23 March 2009Received: 11 June 2008Accepted: 23 March 2009DOI: 10.1186-1471-2407-9-90

Cite this article as: Rosa, F.E., Silveira, S.M., Silveira, C.G. et al. BMC Cancer 2009 9: 90. doi:10.1186-1471-2407-9-90


BackgroundHER-2 gene testing has become an integral part of breast cancer patient diagnosis. The most commonly used assay in the clinical setting for evaluating HER-2 status is immunohistochemistry IHC and fluorescence in situ hybridization FISH. These procedures permit correlation between HER-2 expression and morphological features. However, FISH signals are labile and fade over time, making post-revision of the tumor difficult. CISH chromogenic in situ hybridization is an alternative procedure, with certain advantages, although still limited as a diagnostic tool in breast carcinomas.

MethodsTo elucidate the molecular profile of HER-2 status, mRNA and protein expression in 75 invasive breast carcinomas were analyzed by real time quantitative RT-PCR qRT-PCR and IHC, respectively. Amplifications were evaluated in 43 of these cases by CISH and in 11 by FISH.

ResultsThe concordance rate between IHC and qRT-PCR results was 78.9%, and 94.6% for qRT-PCR and CISH. Intratumoral heterogeneity of HER-2 status was identified in three cases by CISH. The results of the three procedures were compared and showed a concordance rate of 83.8%; higher discordances were observed in 0 or 1+ immunostaining cases, which showed high-level amplification 15.4% and HER-2 transcript overexpression 20%. Moreover, 2+ immunostaining cases presented nonamplified status 50% by CISH and HER-2 downexpression 38.5% by qRT-PCR. In general, concordance occurred between qRT-PCR and CISH results. A high concordance was observed between CISH-qRT-PCR and FISH. Comparisons with clinicopathological data revealed a significant association between HER-2 downexpression and the involvement of less than four lymph nodes P = 0.0350.

ConclusionBased on these findings, qRT-PCR was more precise and reproducible than IHC. Furthermore, CISH was revealed as an alternative and useful procedure for investigating amplifications involving the HER-2 gene.

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2407-9-90 contains supplementary material, which is available to authorized users.

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Author: Fabíola E Rosa - Sara M Silveira - Cássia GT Silveira - Nádia A Bérgamo - Francisco A Moraes Neto - Maria AC Domi

Source: https://link.springer.com/

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