Circulating breast tumor cells exhibit dynamic changes in epithelial and mesenchymal composition - PubMed (original) (raw)

. 2013 Feb 1;339(6119):580-4.

doi: 10.1126/science.1228522.

Aditya Bardia, Ben S Wittner, Shannon L Stott, Malgorzata E Smas, David T Ting, Steven J Isakoff, Jordan C Ciciliano, Marissa N Wells, Ajay M Shah, Kyle F Concannon, Maria C Donaldson, Lecia V Sequist, Elena Brachtel, Dennis Sgroi, Jose Baselga, Sridhar Ramaswamy, Mehmet Toner, Daniel A Haber, Shyamala Maheswaran

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Circulating breast tumor cells exhibit dynamic changes in epithelial and mesenchymal composition

Min Yu et al. Science. 2013.

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Abstract

Epithelial-mesenchymal transition (EMT) of adherent epithelial cells to a migratory mesenchymal state has been implicated in tumor metastasis in preclinical models. To investigate its role in human cancer, we characterized EMT in circulating tumor cells (CTCs) from breast cancer patients. Rare primary tumor cells simultaneously expressed mesenchymal and epithelial markers, but mesenchymal cells were highly enriched in CTCs. Serial CTC monitoring in 11 patients suggested an association of mesenchymal CTCs with disease progression. In an index patient, reversible shifts between these cell fates accompanied each cycle of response to therapy and disease progression. Mesenchymal CTCs occurred as both single cells and multicellular clusters, expressing known EMT regulators, including transforming growth factor (TGF)-β pathway components and the FOXC1 transcription factor. These data support a role for EMT in the blood-borne dissemination of human breast cancer.

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Figures

Fig. 1

Fig. 1

RNA-ISH analysis of EMT markers in human breast tumors. Representative RNA-ISH analysis of pooled epithelial (E) (red dots, arrowheads) and mesenchymal (M) (dark blue dots, arrows) markers in (A) primary tumor and (B) tumor-infiltrated lymph node of a patient with ductal ER+/PR+ type breast cancer. (C) RNA-ISH analysis of HER2 (red dots, arrowheads) and M (dark blue dots, arrows) expression in a HER2+ primary breast tumor. (D) Quantitation of E and M dual-positive tumor cells (percentage of total tumor cells) in a TMA consisting of premalignant DCIS (N = 7 cases) and ER+/PR+ (N = 20 cases), HER2+ (N = 9 cases), and TN (N = 16 cases) breast cancers. A representative image from a TN case is shown on the right. E, red dots; M, dark blue dots; nuclei are stained with hematoxylin, light blue. Scale bars: (A) to (D), 20 μm; inserts, 10 μm.

Fig. 2

Fig. 2

RNA-ISH analysis of EMT markers in CTCs from patients with metastatic breast cancer. (A) Representative images of five types of CTCs isolated from patients with metastatic breast cancer, based on RNA-ISH staining of E (green dots) and M (red dots) markers. Scale bar, 5 μm (B) Quantitation of EMT features in CTCs based on E and M RNA-ISH staining of histological subtypes of breast cancer [lobular, ductal, and U (unknown)], along with molecular classification (ER/PR, HER2, TN). CTC numbers per 3 ml of blood based on RNA (E+M) or protein (CK) staining are listed below. (C) Fractionation of CTCs according to E/M ratios in five patients who were clinically responding to treatment (top) and five patients who had progressive disease on treatment (bottom). The subtype of breast cancer, each patient’s treatment regimen, and the number of days on treatment are shown. The drugs used to inhibit the signaling pathways shown on the figure are as follows: MET + VEGF (vascular endothelial growth factor), cabozantinib; AI (aromatase inhibitor), letrozole; PI3K (phosphatidylinositol 3-kinase), BKM120, INK1117, and BYL719; PI3K/mTOR (mammalian target of rapamycin), SAR245409; MEK (MAP kinase kinase), MSC193639B; EGFR/HER3 (human epidermal growth factor receptor 3), MEHD7945A. The chemotherapeutic drugs used were cisplatin, taxol, and adriamycin. Tumor genotypes are given in table S2.

Fig. 3

Fig. 3

Longitudinal monitoring of EMT features in CTCs from an index patient. Plot of CTC counts per 3 ml of blood based on RNA (E and M markers) detection methods in a patient with KRAS- and PIK3CA-mutant ER/PR+ lobular breast cancer, who was serially sampled during treatment with inhibitors targeting the PI3K (GDC0941) and MEK (GDC0973) pathways, followed by adriamycin chemotherapy. Color-coded quantitation of EMT features based on RNA-ISH staining is shown above each time point. Treatment history and clinical responses are noted on the chart. P, disease progression; R, treatment response). M+ clusters were detected at time points 1, 8, and 12. Images of CTCs staining for E (green) and M (red) markers and protein staining for CK (red), CD45 (green), or platelet marker CD61 (green) from different time points are shown below the chart. The number of single CTCs (S-CTC) detected on the entire CTC-chip upon processing 3 ml of blood and the number of CTCs within the CTC clusters (C-CTC) are indicated. Nuclei are stained with 4′,6-diamidino-2-phenylindole (DAPI) (blue). Scale bar, 10 μm. Criteria for disease progression (P) or treatment response (R) are described in the supplementary materials.

Fig. 4

Fig. 4

RNA-sequence analysis of transcripts enriched in CTCs. Heat map representing transcripts enriched in CTCs captured from the index patient, who was sampled at multiple time points during treatment. A CTC signature of 45 genes was derived by comparing 5 time points from the patient (rows 1 to 5) with identically processed blood specimens from 10 healthy donors (HDs) (rows 6 to 15). An EMT-specific signature of 170 genes was derived from comparing M+ cluster-enriched CTCs (row 4) with E+ CTCs. Red and blue colors indicate relative high and low gene expression, respectively. Categories of gene signatures in the GSEA database are shown for both the 45 gene CTC signature and the 170 gene EMT-cluster CTC signature, with genes contributing to the enrichment highlighted in green. The number of enriched signatures within each category is given in parentheses.

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References

    1. Nguyen DX, Bos PD, Massagué J. Nat Rev Cancer. 2009;9:274. - PubMed
    1. Thiery JP. Nat Rev Cancer. 2002;2:442. - PubMed
    1. Kalluri R, Weinberg RA. J Clin Invest. 2009;119:1420. - PMC - PubMed
    1. Brabletz T. Nat Rev Cancer. 2012;12:425. - PubMed
    1. Ledford H. Nature. 2011;472:273. - PubMed

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