Gene Transcript Assay by Real-Time Rt-PCR in Epithelial Breast Cancer Cells Selected by Laser Microdissection

Becette, Véronique; Vignaud, Sophie; Régnier, Catherine H.; Labroquère, M.; Fourme, Emmanuelle; Menet, Emmanuelle; Bièche, Ivan; Spyratos, Frédérique

doi:10.1177/172460080401900203

Cited by 8 publications

(1 citation statement)

References 13 publications

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“…Such a methodological approach has been shown to allow accurate quantification of mRNA expression for ERα, PgR and HER2 when compared to protein expression (15)(16)(17). We therefore developed a Qrt-PCR multi-gene assessment standard protocol for the quantification of ERα, PgR and HER2 as well as HER1/EGFR, HER3 and HER4, which are key biomarkers for treatment decision in BC (18,19).…”

Section: Introductionmentioning

confidence: 99%

Simultaneous Quantitative Detection of Relevant Biomarkers in Breast Cancer by Quantitative Real-Time PCR

Labuhn¹,

Vuaroqueaux²,

Fina

et al. 2006

Int J Biol Markers

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Section: Introductionmentioning

confidence: 99%

Simultaneous Quantitative Detection of Relevant Biomarkers in Breast Cancer by Quantitative Real-Time PCR

Labuhn¹,

Vuaroqueaux²,

Fina

et al. 2006

Int J Biol Markers

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Sample parameters affecting the clinical relevance of RNA biomarkers in translational breast cancer research

et al. 2012

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In the frame of translational breast cancer research, eligibility criteria for formalin-fixed paraffin-embedded tissue (FFPE) material processing for gene expression studies include tumor cell content (TCC) and sample site (primary vs metastatic tumors). Herein we asked whether the observed differences in gene expression between paired samples with respect to TCC and sample site also have different clinical significance. We assessed ESR1, ERBB2, MAPT, MMP7, and RACGAP1 mRNA expression with real time PCR in paired samples before (NMD) and after macrodissection (MD) from 98 primary tumors (PMD, PNMD) and 72 metastatic lymph nodes (LNMD, LNNMD), as well as from 93 matched P (mP) and LN (mLN). TCC range was 2.5–75 % in the NMD series and 28–98 % in the MD and in the mP/mLN series. The prognostic effect of these markers, individually or in clusters, remained stable between paired PMD/NMD. In comparison, cluster classification failed in the LNNMD group with lower TCC. In the mP/mLN cohort, RACGAP1 mRNA expression was of prognostic significance when tested in mLN samples (p < 0.001). Similarly, luminal B, HER2, and triple negative tumors were of dismal prognosis when classified in the LN component of the same series (mLN, overall survival: p = 0.013, p = 0.034, and p = 0.007, respectively). In conclusion, the clinical relevance of the RNA markers examined may be affected by TCC in metastatic LN samples but not in primary tumors, while it differs between primary tumors and matched metastases. These data will facilitate the design of translational studies involving FFPE sample series.Electronic supplementary materialThe online version of this article (doi:10.1007/s00428-012-1357-1) contains supplementary material, which is available to authorized users.

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Comparative evaluation of non-informative HER-2 immunoreactions (2+) in breast carcinomas with FISH, CISH and QRT-PCR

et al. 2007

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The routine assessment of HER-2 expression can be affected by many immunohistological preanalytical and analytical variables. The evaluation of non-informative HER-2 tests, because of 2(+) scores, has been addressed in studies using in situ hybridization (fluorescent in situ hybridization (FISH) or chromogenic in situ hybridization (CISH)). There are very few studies that additionally checked 2(+) cases by quantitative reverse transcription-PCR (QRT-PCR). We analyzed totally 195 breast carcinoma cases, 70 of them showing 2(+) immunoreaction, with FISH/CISH and QRT-PCR. Confirmed amplification in 2(+) cases fell within the reported range (12.8% vs. 8-44%) and some of them showed lower mRNA levels indicating a genuine decrease of HER-2 protein as a mechanism for the non-informative score. In other cases, increased mRNA levels could be ascribed to HER-2 polysomy, verifying previous observations of immunohistologically detectable HER-2 polysomy. A remarkable subset of the 2(+) cases showed "normal" mRNA levels without amplification or polysomy and technical parameters as well as heterogeneity could be incriminated. The overall concordance of QRT-PCR and FISH was 93.8%, highest than most previously reported. Yet, the lack of clear cut-off mRNA values and the challenge of sample microdissection hinder QRT-PCR from claiming the status of a gold standard test for HER-2 evaluation.

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Gene Transcript Assay by Real-Time Rt-PCR in Epithelial Breast Cancer Cells Selected by Laser Microdissection

Cited by 8 publications

References 13 publications

Simultaneous Quantitative Detection of Relevant Biomarkers in Breast Cancer by Quantitative Real-Time PCR

Simultaneous Quantitative Detection of Relevant Biomarkers in Breast Cancer by Quantitative Real-Time PCR

Sample parameters affecting the clinical relevance of RNA biomarkers in translational breast cancer research

Comparative evaluation of non-informative HER-2 immunoreactions (2+) in breast carcinomas with FISH, CISH and QRT-PCR

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