PURPOSE: The purpose of this research was to develop a quantitative real-time reverse transcription-PCR (RT-PCR) for CK19-mRNA and evaluate its clinical potential for the molecular detection of occult carcinoma cells in the peripheral blood of breast cancer patients. EXPERIMENTAL DESIGN: The method is based on real-time monitoring during PCR of fluorescently labeled specific hybridization probes for CK19-mRNA. The breast cancer cell line MCF-7 was used for the development and analytical evaluation of the assay. We analyzed blood samples from 89 healthy blood donors, 77 patients with early breast cancer (stage I-II) postoperatively, and 47 patients with previously untreated metastatic disease (stage IV) before and after chemotherapy. All of the samples were also analyzed by nested RT-PCR. RESULTS: The method is highly sensitive and specific, because only 2 of 89 (2.2%) of the healthy control subjects had detectable CK19-mRNA+ cells. In 77 patients with early breast cancer, CK19-mRNA+ cells were detected in 24 (31.2%) before and 5 (6.5%) after adjuvant chemotherapy, and their levels differed significantly (P < 0.001, Wilcoxon test). In 47 patients with verified metastases 19 (40.4%) and 20 (42.6%) were found positive before and after chemotherapy, and no significant difference in CK19-mRNA+ cell levels was observed (P = 0.96, Wilcoxon test). Results obtained by the proposed real-time RT-PCR method correlated well with those obtained for the same samples by nested RT-PCR [concordance in 312 of 337 (92.6%); P = 0.69, McNemar test]. CONCLUSIONS: The developed method is highly sensitive and specific, and can be used for high-throughput continuous monitoring and quantification of circulating epithelial cells in the peripheral blood of breast cancer patients.
PURPOSE: The purpose of this research was to develop a quantitative real-time reverse transcription-PCR (RT-PCR) for CK19-mRNA and evaluate its clinical potential for the molecular detection of occult carcinoma cells in the peripheral blood of breast cancerpatients. EXPERIMENTAL DESIGN: The method is based on real-time monitoring during PCR of fluorescently labeled specific hybridization probes for CK19-mRNA. The breast cancer cell line MCF-7 was used for the development and analytical evaluation of the assay. We analyzed blood samples from 89 healthy blood donors, 77 patients with early breast cancer (stage I-II) postoperatively, and 47 patients with previously untreated metastatic disease (stage IV) before and after chemotherapy. All of the samples were also analyzed by nested RT-PCR. RESULTS: The method is highly sensitive and specific, because only 2 of 89 (2.2%) of the healthy control subjects had detectable CK19-mRNA+ cells. In 77 patients with early breast cancer, CK19-mRNA+ cells were detected in 24 (31.2%) before and 5 (6.5%) after adjuvant chemotherapy, and their levels differed significantly (P < 0.001, Wilcoxon test). In 47 patients with verified metastases 19 (40.4%) and 20 (42.6%) were found positive before and after chemotherapy, and no significant difference in CK19-mRNA+ cell levels was observed (P = 0.96, Wilcoxon test). Results obtained by the proposed real-time RT-PCR method correlated well with those obtained for the same samples by nested RT-PCR [concordance in 312 of 337 (92.6%); P = 0.69, McNemar test]. CONCLUSIONS: The developed method is highly sensitive and specific, and can be used for high-throughput continuous monitoring and quantification of circulating epithelial cells in the peripheral blood of breast cancerpatients.
Authors: Giuliana Giribaldi; Simone Procida; Daniela Ulliers; Franca Mannu; Roberta Volpatto; Giorgia Mandili; Laura Fanchini; Oscar Bertetto; Gianruggero Fronda; Luigi Simula; Elena Rimini; Giovanni Cherchi; Lisa Bonello; Milena Maria Maule; Francesco Turrini Journal: J Mol Diagn Date: 2006-02 Impact factor: 5.568
Authors: A Daskalaki; S Agelaki; M Perraki; S Apostolaki; N Xenidis; E Stathopoulos; E Kontopodis; D Hatzidaki; D Mavroudis; V Georgoulias Journal: Br J Cancer Date: 2009-07-21 Impact factor: 7.640
Authors: Galatea Kallergi; Harris Markomanolaki; Vicky Giannoukaraki; Maria A Papadaki; Areti Strati; Evi S Lianidou; Vassilis Georgoulias; Dimitris Mavroudis; Sofia Agelaki Journal: Breast Cancer Res Date: 2009-11-17 Impact factor: 6.466
Authors: I Van der Auwera; D Peeters; I H Benoy; H J Elst; S J Van Laere; A Prové; H Maes; P Huget; P van Dam; P B Vermeulen; L Y Dirix Journal: Br J Cancer Date: 2009-12-01 Impact factor: 7.640