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| Status |
Public on Sep 26, 2011 |
| Title |
Prolonged Drug Selection of Breast Cancer Cells and Enrichment of Cancer Stem Cell Characteristics. |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by array
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| Summary |
Background: Cancer stem cells are presumed to have virtually unlimited proliferative and self-renewal abilities and to be highly resistant to chemotherapy, a feature that is associated with overexpression of ATP-binding cassette transporters. We investigated whether prolonged continuous selection of cells for drug resistance enriches cultures for cancer stem-like cells.
Methods: Cancer stem cells were defined as CD44+/CD24– cells that could self-renew (ie, generate cells with the tumorigenic CD44+/CD24– phenotype), differentiate, invade, and form tumors in vivo. We used doxorubicin-selected MCF-7/ADR cells, weakly tumorigenic parental MCF-7 cells, and MCF-7/MDR, an MCF-7 subline with forced expression of ABCB1 protein. Cells were examined for cell surface markers and side-population fractions by microarray and flow cytometry, with in vitro invasion assays, and for ability to form mammospheres. Xenograft tumors were generated in mice to examine tumorigenicity (n = 52). The mRNA expression of multidrug resistance genes was examined in putative cancer stem cells and pathway analysis of statistically significantly differentially expressed genes was performed. All statistical tests were two-sided.
Results: Pathway analysis showed that MCF-7/ADR cells express mRNAs from ABCB1 and other genes also found in breast cancer stem cells (eg, CD44, TGFB1, and SNAI1). MCF-7/ADR cells were highly invasive, formed mammospheres, and were tumorigenic in mice. In contrast to parental MCF-7 cells, more than 30% of MCF-7/ADR cells had a CD44+/CD24– phenotype, could self-renew, and differentiate (ie, produce CD44+/CD24– and CD44+/CD24+ cells), and overexpressed various multidrug resistance-linked genes (including ABCB1, CCNE1, and MMP9). MCF-7/ADR cells were statistically significantly more invasive in Matrigel than parental MCF-7 cells (MCF-7 cells = 0.82 cell per field and MCF-7/ADR = 7.51 cells per field, difference = 6.69 cells per field, 95% confidence interval = 4.82 to 8.55 cells per field, P<.001). No enrichment in the CD44+/CD24– or CD133+ population was detected in MCF-7/MDR.
Conclusion: The cell population with cancer stem cell characteristics increased after prolonged continuous selection for doxorubicin resistance.
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| Overall design |
PARALLEL study design with 4 samples Parental MCF-7 cell line versus Doxorubicin Resistant MCF-7 cell sublines Biological replicates: 2 parental controls, 2 drug resistant, independently grown and harvested.
agent:Selection agent is multi-step doxorubicin selection: MCF7226ng, MCF7262ng
biological replicate: MCF71, MCF72
biological replicate: MCF226ng, MCF7262ng
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| Contributor(s) |
Calcagno AM, Salcido CD, Guillet J, Wu C, Fostel JM, Mumau MD, Gottesman MM, Varticovski L, Ambudkar SV |
| Citation(s) |
20935265 |
| Submission date |
Sep 30, 2010 |
| Last update date |
Dec 06, 2018 |
| Contact name |
Suresh V. Ambudkar |
| E-mail(s) |
ambudkar@helix.nih.gov
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| Organization name |
National Cancer Institute, NIH
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| Lab |
Laboratory of Cell Biology
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| Street address |
37 Convent Dr., Room 2120
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| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
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| Platforms (1) |
| GPL571 |
[HG-U133A_2] Affymetrix Human Genome U133A 2.0 Array |
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| Samples (4)
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| GSM602658 |
MCF-7_parental_cell_rep_1 |
| GSM602659 |
MCF-7_parental_cell_rep_2 |
| GSM602660 |
MCF-7ADR_multi-step_doxorubicin_selected_subline_rep_1 |
| GSM602661 |
MCF-7ADR_multi-step_doxorubicin_selected_subline_rep_2 |
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| Relations |
| BioProject |
PRJNA132705 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE24460_RAW.tar |
7.5 Mb |
(http)(custom) |
TAR (of CEL) |
| Processed data included within Sample table |
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