Franziska Meier-Stiegen

Heinrich-Heine-Universität Düsseldorf, Düsseldorf, North Rhine-Westphalia, Germany

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Publications (13)85.05 Total impact

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    ABSTRACT: BACKGROUND: Circulating tumor cells (CTCs) are promising surrogate markers for systemic disease, and their molecular characterization might be relevant to guide more individualized cancer therapies. To enable fast and efficient purification of individual CTCs, we developed a work flow from CellSearch (TM) cartridges enabling high-resolution genomic profiling on the single-cell level. METHODS: Single CTCs were sorted from 40 CellSearch samples from patients with metastatic breast cancer using a MoFlo XDP cell sorter. Genomes of sorted single cells were amplified using an adapter-linker PCR. Amplification products were analyzed by array-based comparative genomic hybridization, a gene-specific quantitative PCR (qPCR) assay for cyclin D1 (CCND1) locus amplification, and genomic sequencing to screen for mutations in exons 1, 9, and 20 of the phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA) gene and exons 5, 7, and 8 of the tumor protein p53 (TP53) gene. RESULTS: One common flow-sorting protocol was appropriate for 90% of the analyzed CellSearch cartridges, and the detected CTC numbers correlated positively with those originally detected with the CellSearch system (R-2 = 0.9257). Whole genome amplification was successful in 72.9% of the sorted single CTCs. Over 95% of the cells displayed chromosomal aberrations typical for metastatic breast cancers, and amplifications at the CCND1 locus were validated by qPCR. Aberrant CTCs from 2 patients harbored mutations in exon 20 of the PIK3CA gene. CONCLUSIONS: This work flow enabled effective CTC isolation and provided insights into genomic alterations of CTCs in metastatic breast cancer. This approach might facilitate further molecular characterization of rare CTCs to increase understanding of their biology and as a basis for their molecular screening in the clinical setting. (C) 2014 American Association for Clinical Chemistry
    Clinical Chemistry 10/2014; 60(10):1290-7. DOI:10.1373/clinchem.2014.222331 · 7.77 Impact Factor
  • Geburtshilfe und Frauenheilkunde 09/2014; 74(S 01). DOI:10.1055/s-0034-1388344 · 0.96 Impact Factor
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    ABSTRACT: An imbalance between cell proliferation and programmed cell death can result in tumor growth. Although most systemic cytotoxic agents induce apoptosis in tumor cells, a high apoptotic rate in primary breast cancer correlates with poor prognosis. The aim of this study was to investigate the incidence and the prognostic significance of apoptotic disseminated tumor cells (DTC) in the bone marrow (BM) of breast cancer patients who either underwent primary surgery or primary systemic chemotherapy (PST).
    BMC Cancer 06/2014; 14(1):394. DOI:10.1186/1471-2407-14-394 · 3.32 Impact Factor
    This article is viewable in ResearchGate's enriched format
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  • Cancer Research 03/2014; 73(24 Supplement):PD6-6-PD6-6. DOI:10.1158/0008-5472.SABCS13-PD6-6 · 9.28 Impact Factor
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    ABSTRACT: Background We aimed to assess the clinical validity of circulating tumour cell (CTC) quantifi cation for prognostication of patients with metastatic breast cancer by undertaking a pooled analysis of individual patient data. Methods We contacted 51 European centres and asked them to provide reported and unreported anonymised data for individual patients with metastatic breast cancer who participated in studies between January, 2003, and July, 2012. Eligible studies had participants starting a new line of therapy, data for progression-free survival or overall survival, or both, and CTC quantifi cation by the CellSearch method at baseline (before start of new treatment). We used Cox regression models, stratifi ed by study, to establish the association between CTC count and progression-free survival and overall survival. We used the landmark method to assess the prognostic value of CTC and serum marker changes during treatment. We assessed the added value of CTCs or serum markers to prognostic clinicopathological models in a resampling procedure using likelihood ratio (LR) χ² statistics. Findings 17 centres provided data for 1944 eligible patients from 20 studies. 911 patients (46·9%) had a CTC count of 5 per 7·5 mL or higher at baseline, which was associated with decreased progression-free survival (hazard ratio [HR] 1·92, 95% CI 1·73–2·14, p<0·0001) and overall survival (HR 2·78, 95% CI 2·42–3·19, p<0·0001) compared with patients with a CTC count of less than 5 per 7·5 mL at baseline. Increased CTC counts 3–5 weeks after start of treatment, adjusted for CTC count at baseline, were associated with shortened progression-free survival (HR 1·85, 95% CI 1·48–2·32, p<0·0001) and overall survival (HR 2·26, 95% CI 1·68–3·03) as were increased CTC counts after 6–8 weeks (progression-free survival HR 2·20, 95% CI 1·66–2·90, p<0·0001; overall survival HR 2·91, 95% CI 2·01–4·23, p<0·0001). Survival prediction was signifi cantly improved by addition of baseline CTC count to the clinicopathological models (progression-free survival LR 38·4, 95% CI 21·9–60·3, p<0·0001; overall survival LR 64·9, 95% CI 41·3–93·4, p<0·0001). This model was further improved by addition of CTC change at 3–5 weeks (progression-free survival LR 8·2, 95% CI 0·78–20·4, p=0·004; overall survival LR 11·5, 95% CI 2·6–25·1, p=0·0007) and at 6–8 weeks (progression-free survival LR 15·3, 95% CI 5·2–28·3; overall survival LR 14·6, 95% CI 4·0–30·6; both p<0·0001). Carcinoembryonic antigen and cancer antigen 15-3 concentrations at baseline and during therapy did not add signifi cant information to the best baseline model. Interpretation These data confi rm the independent prognostic eff ect of CTC count on progression-free survival and overall survival. CTC count also improves the prognostication of metastatic breast cancer when added to full clinicopathological predictive models, whereas serum tumour markers do not.
    The Lancet Oncology 03/2014; · 24.73 Impact Factor
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    ABSTRACT: We aimed to assess the clinical validity of circulating tumour cell (CTC) quantification for prognostication of patients with metastatic breast cancer by undertaking a pooled analysis of individual patient data. We contacted 51 European centres and asked them to provide reported and unreported anonymised data for individual patients with metastatic breast cancer who participated in studies between January, 2003, and July, 2012. Eligible studies had participants starting a new line of therapy, data for progression-free survival or overall survival, or both, and CTC quantification by the CellSearch method at baseline (before start of new treatment). We used Cox regression models, stratified by study, to establish the association between CTC count and progression-free survival and overall survival. We used the landmark method to assess the prognostic value of CTC and serum marker changes during treatment. We assessed the added value of CTCs or serum markers to prognostic clinicopathological models in a resampling procedure using likelihood ratio (LR) χ(2) statistics. 17 centres provided data for 1944 eligible patients from 20 studies. 911 patients (46·9%) had a CTC count of 5 per 7·5 mL or higher at baseline, which was associated with decreased progression-free survival (hazard ratio [HR] 1·92, 95% CI 1·73-2·14, p<0·0001) and overall survival (HR 2·78, 95% CI 2·42-3·19, p<0·0001) compared with patients with a CTC count of less than 5 per 7·5 mL at baseline. Increased CTC counts 3-5 weeks after start of treatment, adjusted for CTC count at baseline, were associated with shortened progression-free survival (HR 1·85, 95% CI 1·48-2·32, p<0·0001) and overall survival (HR 2·26, 95% CI 1·68-3·03) as were increased CTC counts after 6-8 weeks (progression-free survival HR 2·20, 95% CI 1·66-2·90, p<0·0001; overall survival HR 2·91, 95% CI 2·01-4·23, p<0·0001). Survival prediction was significantly improved by addition of baseline CTC count to the clinicopathological models (progression-free survival LR 38·4, 95% CI 21·9-60·3, p<0·0001; overall survival LR 64·9, 95% CI 41·3-93·4, p<0·0001). This model was further improved by addition of CTC change at 3-5 weeks (progression-free survival LR 8·2, 95% CI 0·78-20·4, p=0·004; overall survival LR 11·5, 95% CI 2·6-25·1, p=0·0007) and at 6-8 weeks (progression-free survival LR 15·3, 95% CI 5·2-28·3; overall survival LR 14·6, 95% CI 4·0-30·6; both p<0·0001). Carcinoembryonic antigen and cancer antigen 15-3 concentrations at baseline and during therapy did not add significant information to the best baseline model. These data confirm the independent prognostic effect of CTC count on progression-free survival and overall survival. CTC count also improves the prognostication of metastatic breast cancer when added to full clinicopathological predictive models, whereas serum tumour markers do not. Janssen Diagnostics, the Nuovo-Soldati foundation for cancer research.
    The Lancet Oncology 03/2014; DOI:10.1016/S1470-2045(14)70069-5 · 25.12 Impact Factor
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    ABSTRACT: Evaluation and characterization of circulating tumor cells (CTCs) have become a major focus of translational cancer research. Presence of CTCs predicts worse clinical outcome in early and metastatic breast cancer. Whether all cells from the primary tumor have potential to disseminate and form subsequent metastasis remains unclear. As part of the metastatic cascade, tumor cells lose their cell-to-cell adhesion and undergo epithelial-mesenchymal transition (EMT) in order to enter blood circulation. During EMT epithelial antigens are downregulated; thus, such tumor cells might elude classical epithelial marker-based detection. Several researchers postulated that some CTCs express stem cell-like phenotype; this might lead to chemoresistance and enhanced metastatic potential of such cells. In the present review, we discuss current data on EMT and stem cell markers in CTCs of breast cancer and their clinical significance.
    BioMed Research International 01/2014; 2014:415721. DOI:10.1155/2014/415721 · 2.71 Impact Factor
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    ABSTRACT: The frequently altered phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway is involved in the regulation of cellular processes required for breast carcinogenesis. The aim of the project was to develop a method to identify hotspot mutations in the PIK3CA gene in circulating tumor cells (CTCs) of metastatic breast cancer (metBC) patients. From 44 enrolled CTC-positive metBC patients a total number of 57 peripheral blood samples were analysed by CellSearch(®). Genomic DNA of enriched CTCs was isolated, amplified and analyzed for PIK3CA mutations in exons 9 and 20 which lead to E542K, E545K or H1047R amino acid changes and result in increased PI3K activity. The mutations were detected by using SNaPshot-methodology comprising PCR amplification and single nucleotide primer extension. SNaPshot analysis was established using genomic DNA from different breast cancer cell lines and then successfully transferred to investigate blood samples and single cells. Overall, twelve hotspot mutations in either exon 9/E545K (6/12, 50%) or exon 20/H1047R (6/12, 50%) could be determined within 9 out of 57 (15.8%) blood samples from 7 out of 44 (15.9%) patients; CTC counts ranged from 1 to 9748. PIK3CA variants E542K, E545G and E545A were not detected. Analysing the PIK3CA genotype of CTCs has clinical relevance with respect to drug resistance, e.g. against HER2-targeted therapy. The herein described approach including SNaPshot technology provides a simple method to characterize hotspot mutations within CTCs enriched from peripheral blood and can be easily adopted for analysing further therapeutically relevant SNPs.
    Molecular oncology 07/2013; DOI:10.1016/j.molonc.2013.07.007 · 6.70 Impact Factor
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    ABSTRACT: Overexpression of the HER2-receptor in early breast cancer (EBC) patients is associated with aggressive tumor behavior. However, women suffering from HER2-positive EBC benefit from trastuzumab treatment. As the HER2 status of the primary tumor may differ from that of disseminated tumor cells (DTC) in bone marrow (BM), the aim of this study was (1) to compare the HER2 status of the primary tumor (prim-HER2-status) with that of DTC (DTC-HER2-status) and (2) to analyze the influence of the DTC-HER2-status on patient survival. For this purpose, BM aspirates from 569 EBC patients were analyzed for the presence of DTC. The DTC-HER2-status was identified by a double-staining procedure against cytokeratin and the HER2-receptor. DTC were detected in 151 (27 %) patients. The concordance between the HER2 status of DTC and the primary tumor was 51 %. In patients with detectable DTC, mean disease-free survival was 77.44 (95 % CI 74.72-80.17) months for DTC-HER2-negative and 55.15 (95 % CI 48.57-61.79) months for DTC-HER2-positive patients (p = 0.044). The multivariate analysis showed that the DTC-HER2-status was an independent predictor of disease-free survival. In conclusion, the presence of HER2-positive DTC in EBC patients is associated with an increased risk of relapse. Due to the low concordance between the HER2 status of the primary tumor and DTC, only a minority (13 %) of the DTC-HER2-positive patients was treated with trastuzumab. These patients might, however, benefit from HER2-directed therapy.
    Breast Cancer Research and Treatment 03/2013; 138(2). DOI:10.1007/s10549-013-2470-9 · 4.47 Impact Factor
  • T. Fehm, M. Banys, F. Meier-Stiegen, A. Hartkopf, V. Müller
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    ABSTRACT: Isolierte Tumorzellen können sowohl im peripheren Blut als auch im Knochenmark (KM) von Patientinnen mit Mammakarzinom detektiert werden. Während der Nachweis von disseminierten Tumorzellen (DTZ) im Knochenmark von primären Mammakarzinompatientinnen einen unabhängigen Prognosefaktor (Level-I-Evidenz) darstellt, ist die prognostische Bedeutung von zirkulierenden Tumorzellen (ZTZ) im peripheren Blut in diesem Kollektiv noch nicht eindeutig geklärt. Im Beitrag werden die Relevanz von ZTZ beim primären wie beim metastasierten Mammakarzinom und die aktuelle Studienlage zu diesem Thema erläutert. Abstract Isolated tumor cells can be detected in peripheral blood and bone marrow of breast cancer patients. The prognostic significance of disseminated tumor cells (DTC) in bone marrow (BM) has been demonstrated in a large pooled analysis (level I evidence). Since BM biopsies are not well tolerated by many patients, evaluation of circulating tumor cells (CTC) in blood may represent an important alternative. Recent studies have indicated a prognostic significance of CTC in both the primary and metastatic setting. The evaluation of CTC may become one of the crucial markers for prediction of survival and therapy monitoring, and its characterization might enable specific targeting of minimal residual as well as metastatic disease. This report provides an overview of ongoing studies regarding CTC as well as the clinical relevance of CTC in primary and metastatic breast cancer.
    Der Gynäkologe 07/2012; 45(7):563-567. DOI:10.1007/s00129-012-3015-8
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    ABSTRACT: Tumor dormancy describes a prolonged quiescent state in which tumor cells are present, but disease progression is not yet clinically apparent. Breast cancer is especially known for long asymptomatic periods, up to 25 years, with no evidence of the disease, followed by a relapse. Factors that determine the cell's decision to enter a dormant state and that control its duration remain unclear. In recent years, considerable progress has been made in understanding how tumor cells circulating in the blood interact and extravasate into secondary sites and which factors might determine whether these cells survive, remain dormant, or become macrometastases. The mechanisms of tumor cell dormancy are still not clear. Two different hypotheses are currently discussed: tumor cells persist either by completely withdrawing from the cell cycle or by continuing to proliferate at a slow rate that is counterbalanced by cell death. Because dormant disseminated tumor cells may be the founders of metastasis, one hypothesis is that dormant tumor cells, or at least a fraction of them, share stem cell-like characteristics that may be responsible for their long half-lives and their suggested resistance to standard chemotherapy. Therefore, knowledge of the biology of tumor cell dormancy may be the basis from which to develop innovative targeted therapies to control or eliminate this tumor cell fraction. In this review, we discuss biological mechanisms and clinical implications of tumor dormancy in breast cancer patients.
    Breast Cancer: Targets and Therapy 01/2012; 4:183-191. DOI:10.2147/BCTT.S26431

Publication Stats

37 Citations
85.05 Total Impact Points

Institutions

  • 2013–2014
    • Heinrich-Heine-Universität Düsseldorf
      Düsseldorf, North Rhine-Westphalia, Germany
    • University of Tuebingen
      • Department of Gynecology and Obstetrics
      Tübingen, Baden-Württemberg, Germany