Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of cancer mammospheres and bulk tumors

ABSTRACT: Tumorigenic breast cancer cells characterized by CD44 expression and low or undetectable CD24 levels (CD44+/CD24-/low) may be resistant to chemotherapy and therefore responsible for cancer relapse. Paired breast cancer core biopsies before and after neoadjuvant chemotherapy or lapatinib were obtained and as single cell suspensions stained using antibodies against CD24, CD44, and lineage markers, and then analyzed by flow cytometry. Mammosphere (MS) formation in culture was compared before and after treatment. Global gene expression differences between cancer cells bearing CD44+/CD24-/low cells and all other sorted cells, and between cancer MS and the primary bulk invasive cancers were analyzed. We report that CD44+/CD24-/low tumorigenic breast cancer cells were intrinsically chemoresistant - chemotherapy led to increased CD44+/CD24-/low cells, increased self-renewal capacity on MS assays, and enhanced tumorigeneicity in immunocompromised SCID/Beige mice. Conversely, in patients with HER2 overexpressing tumors, the EGFR/HER2 tyrosine kinase inhibitor, lapatinib decreased CD44+/CD24-/low cells, with the majority of these patients after conventional therapy achieving pathologic complete response, a validated surrogate marker for long-term survival. Gene transcription pathways that underlie chemoresistant, MS-forming CD44+/CD24-/low cells involve genes belonging to stem cell self-renewal, Wnt signaling, and early development pathways. Experiment Overall Design: Cells from human breast tumors were grown as mammospheres (MS). Experiment Overall Design: Isolated single cell suspensions from primary breast cancers were plated onto non-adherent (polyhema-coated) plastic, counted with a hematocytometer, and 20,000 cells were then seeded into a 6-well ultra-low attachment plate supplemented with 2mL MEGM, with the addition of 2 mL of freshly unfrozen MEGM every 3-4 days. Gene expression profiles were taken of both MS and primary bulk tumors and compared with each other.

ORGANISM(S): Homo sapiens

SUBMITTER: Chad Creighton

PROVIDER: E-GEOD-7515 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Residual breast cancers after conventional therapy display mesenchymal as well as tumor-initiating features.

Creighton Chad J CJ Li Xiaoxian X Landis Melissa M Dixon J Michael JM Neumeister Veronique M VM Sjolund Ashley A Rimm David L DL Wong Helen H Rodriguez Angel A Herschkowitz Jason I JI Fan Cheng C Zhang Xiaomei X He Xiaping X Pavlick Anne A Gutierrez M Carolina MC Renshaw Lorna L Larionov Alexey A AA Faratian Dana D Hilsenbeck Susan G SG Perou Charles M CM Lewis Michael T MT Rosen Jeffrey M JM Chang Jenny C JC

Proceedings of the National Academy of Sciences of the United States of America 20090803 33

Some breast cancers have been shown to contain a small fraction of cells characterized by CD44(+)/CD24(-/low) cell-surface antigen profile that have high tumor-initiating potential. In addition, breast cancer cells propagated in vitro as mammospheres (MSs) have also been shown to be enriched for cells capable of self-renewal. In this study, we have defined a gene expression signature common to both CD44(+)/CD24(-/low) and MS-forming cells. To examine its clinical significance, we determined whet ...[more]

PMID: 19666588

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Project description:Tumorigenic breast cancer cells characterized by CD44 expression and low or undetectable CD24 levels (CD44+/CD24-/low) may be resistant to chemotherapy and therefore responsible for cancer relapse. Paired breast cancer core biopsies before and after neoadjuvant chemotherapy or lapatinib were obtained and as single cell suspensions stained using antibodies against CD24, CD44, and lineage markers, and then analyzed by flow cytometry. Mammosphere (MS) formation in culture was compared before and after treatment. Global gene expression differences between cancer cells bearing CD44+/CD24-/low cells and all other sorted cells, and between cancer MS and the primary bulk invasive cancers were analyzed. We report that CD44+/CD24-/low tumorigenic breast cancer cells were intrinsically chemoresistant â chemotherapy led to increased CD44+/CD24-/low cells, increased self-renewal capacity on MS assays, and enhanced tumorigeneicity in immunocompromised SCID/Beige mice. Conversely, in patients with HER2 overexpressing tumors, the EGFR/HER2 tyrosine kinase inhibitor, lapatinib decreased CD44+/CD24-/low cells, with the majority of these patients after conventional therapy achieving pathologic complete response, a validated surrogate marker for long-term survival. Gene transcription pathways that underlie chemoresistant, MS-forming CD44+/CD24-/low cells involve genes belonging to stem cell self-renewal, Wnt signaling, and early development pathways. Experiment Overall Design: Human breast tumor samples were sorted using flow cytometry to select for cells that were CD44+ and CD24-. Gene expression profiles of these cells were compared with profiles of the other sorted cells (CD24+ and CD44-/CD24-). Experiment Overall Design: Core biopsies of primary breast tumors were taken and placed immediately in cold RPMI-1640 supplemented with 10% heat-inactivated newborn calf serum (HINCS, Invitrogen, Carlsbad, CA). Within an hour, the samples were minced and then digested in 10-15 mL of MEGM with 250-300 units/mL collagenase at 370C. The samples were filtered, washed, and then subjected to hypotonic shock to lyse red blood cells. About 106 single cells were re-suspended, incubated for 15 min at 40C with anti-CD44 (APC), anti-CD24 (FITC), and anti-lineage cocktail antibodies (PE-conjugated anti-CD2, CD3, CD10, CD16, CD18, CD31 and CD 140B) (Pharmingen, San Diego, CA) using the manufacturerâs suggested concentrations. The cells were then washed twice, re-suspended with the viability dye propidium iodide, and analyzed using Dako MoFlo flow cytometry. Side- and forward- scatter were used to eliminate debris and cell doublets, and the Lin- cells were further analyzed by CD44 and CD24 markers.

Project description:Breast cancers contain a minority population of cancer cells characterized by CD44 expression but low or undetectable levels of CD24 (CD44+CD24-/low) that have higher tumorigenic capacity than other subtypes of cancer cells. METHODS: We compared the gene-expression profile of CD44+CD24-/low tumorigenic breast-cancer cells with that of normal breast epithelium. Differentially expressed genes were used to generate a 186-gene invasiveness gene signature (IGS), which was evaluated for its association with overall survival and metastasis-free survival in patients with breast cancer or other types of cancer. RESULTS: There was a significant association between the IGS and both overall and metastasis-free survival (P<0.001, for both) in patients with breast cancer, which was independent of established clinical and pathological variables. When combined with the prognostic criteria of the National Institutes of Health, the IGS was used to stratify patients with high-risk early breast cancer into prognostic categories (good or poor); among patients with a good prognosis, the 10-year rate of metastasis-free survival was 81%, and among those with a poor prognosis, it was 57%. The IGS was also associated with the prognosis in medulloblastoma (P=0.004), lung cancer (P=0.03), and prostate cancer (P=0.01). The prognostic power of the IGS was increased when combined with the wound-response (WR) signature. CONCLUSIONS: The IGS is strongly associated with metastasis-free survival and overall survival for four different types of tumors. This genetic signature of tumorigenic breast-cancer cells was even more strongly associated with clinical outcomes when combined with the WR signature in breast cancer. Expression profling was performed on 6 tumorigenic, 3 non tumorigenic samples of breast tumors and 3 normal breast samples on two different platforms GPL96 and GPL97. A gene signature was derived by comparing the gene expressions of 6 tumorigenic samples with 3 normal breast samples.

Dataset Information

Transcription profiling of cancer mammospheres and bulk tumors

Publications

Residual breast cancers after conventional therapy display mesenchymal as well as tumor-initiating features.

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