Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Isolation and expression profiling of circulating tumor cells in breast cancer patients part 3

ABSTRACT: We developed a method to isolate pure circulating tumor cells (CTC). RNA from such CTCs isolated from the peripheral blood of metastatic breats cnacer patients and gene expression was performed using cDNAmicroarray. we used cDNA array to compare gene expression of CTCs with normal epithelial and breast tumor samples CTCs vs. breast tumors

ORGANISM(S): Homo sapiens

SUBMITTER: Vasu Punj

PROVIDER: E-GEOD-45964 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Expression profiling of circulating tumor cells in metastatic breast cancer.

Lang Julie E JE Scott Janet H JH Wolf Denise M DM Novak Petr P Punj Vasu V Magbanua Mark Jesus M MJ Zhu Weizhu W Mineyev Neal N Haqq Christopher M CM Crothers Julia R JR Esserman Laura J LJ Tripathy Debasish D van 't Veer Laura L Park John W JW

Breast cancer research and treatment 20141129 1

Circulating tumor cells (CTCs) are prognostic in all stages of breast cancer. However, since they are extremely rare, little is known about the molecular nature of these cells. We report a novel strategy for the isolation and expression profiling of pure populations of CTCs derived from peripheral blood. We developed a method to isolate CTCs based on immunomagnetic capture followed by fluorescence-activated cell sorting (IE/FACS). After assay validation using the BT474 cell line spiked into bloo ...[more]

PMID: 25432738

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Project description:Introduction: Targeting of the tumor vasculature has evolved into an integral part of existing standard anti-cancer therapies. However, recent pre-clinical and clinical studies have shown that the efficacy of these treatments is temporary at most and frequently followed by the renewed tumor growth, indicating that these targets may not be robust. Methods: We transcriptionally profiled laser capture microdissection isolated microvessels from tumor and matching adjacent normal tissue samples obtained from breast cancer patients diagnosed with infiltrating ductal carcinoma. Gene expression profiles were generated and analyzed using Genespring and the web-based WebGestalt bioinformatics Toolkit. Several differentially expressed genes were validated by immunohistochemistry. To further mine our data set we used a novel integrative systems biology network approach to identify a gene signature that could robustly stratify breast cancer patient populations. Results: Vascular enrichment of the LCM samples was confirmed by Q-PCR of CD31. Hierarchical two-dimensional clustering of the transcriptome data identified 219 significantly upregulated transcripts in at least 4/8 patient samples (cut-off >1.3-fold, p<0.05). Several genes AGRN, FLNA and ILR4 were selected and their protein overexpression was confirmed in the tumor tissue. Additional mining of the data set using the clinical information associated with these tissues by applying an integrative bioinformatics network method generated a 15-gene M-bM-^@M-^XVascular-Derived Prognostic PredictorM-bM-^@M-^Y that is able to stratify patient populations in to high- and low- risk groups. Conclusions: The overall heterogeneity observed in the vascular gene expression patterns in the infiltrating ductal carcinoma samples poses a significant problem for these individual genes to be used as a robust vascular breast cancer specific marker or therapeutic target. Using an integrative bioinformatics network approach allowed us to identify a 15-gene M-bM-^@M-^XVascular-Derived Prognostic PredictorM-bM-^@M-^Y that can robustly identify patients with an increased risk of local recurrence. Stratifying patient populations using this gene signature may lead to better patient-tailored treatment regimes. Two-color microarrays comparing laser-captured microvessels from the tumors and matching adjacent normal tissues of 8 breast cancer patients

Project description:CCAAT/enhancer binding protein beta (C/EBPb) is a member of a family of highly conserved transcription factors that regulates numerous genes involved in proliferation and differentiation in a variety of tissues. C/EBPb is deregulated in human breast cancer and germline deletion of this gene results in multiple defects in mammary gland development. We hypothesized that C/EBPb regulates mammary stem cell self-renewal, maintenance and/or differentiation through the regulation of multiple target genes that coordinate mammary gland development. Utilizing both a germline knockout mouse model and a conditional knockout strategy, we demonstrated that mammosphere formation was significantly decreased in C/EBPb-deficient mammary epithelial cells (MECs). The ability of C/EBPb-deleted MECs to regenerate the mammary gland in vivo was severely impaired when transplanted at limiting dilution. Furthermore, serial transplantation of C/EBPb-null mammary tissue resulted in decreased outgrowth potential when compared to wildtype, and an early senescence phenotype. Flow cytometric analysis revealed that C/EBPb-null MECs contain a lower frequency of repopulating stem cells accompanied by an increase in committed, differentiated luminal cells as compared to wildtype. Microarray analysis of stem/progenitor cell populations was performed and revealed an alteration in cell fate specification in C/EBPb-null glands, exemplified by the aberrant expression of basal markers in the luminal cell compartment. Collectively, our studies demonstrate that C/EBPb is a critical regulator of mammary stem cell differentiation, and an important determinant of luminal cell fate specification. Experiment Overall Design: To identify potential signaling pathways regulated by C/EBPb in stem/progenitor cells, microarray analysis was performed on two stem/progenitor cell subpopulations. For this analysis, subpopulations defined by LIN-CD24+CD29hi and LIN-CD24hiCD29lo were FACS sorted from wildtype and germline C/EBPb-/- glands, and RNA was isolated from each group.

Dataset Information

Isolation and expression profiling of circulating tumor cells in breast cancer patients part 3

Publications

Expression profiling of circulating tumor cells in metastatic breast cancer.

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