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RNA profiling of EpCAM-positive cells in bone marrow of early breast cancer (EBC) patients

ABSTRACT: We developed a novel approach to isolate tumor cells with high purity from bone marrow which was subjected to immunomagnetic enrichment using EpCAM beads followed by fluorescence activated cell sorting (IE/FACS) to isolate EpCAM-positive cells away from leukocytes (CD45+). For RNA profiling, QPCR analysis was performed on sixty four (64) cancer-related genes using Taqman® low density arrays. For non-tumor controls, RNA profiling was performed on matched leukocytes (CD45+) isolated from the same enriched bone marrow samples from 17 of the 30 patients.

ORGANISM(S): Homo sapiens

PROVIDER: GSE112757 | GEO | 2019/04/04

REPOSITORIES: GEO

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DNA profiling of EpCAM-positive cells in bone marrow of early breast cancer (EBC) patients

Project description:Parallel DNA and RNA profiling of EpCAM-positive cells in bone marrow and primary tumor tissue with positive disseminated tumor cell (DTC) count via immunomagnetic Enrichment/Flow Cytometry (IE/FC) of metastatic breast cancer (MBC) patients confirm their malignant nature We developed a novel approach to isolate tumor cells with high purity from bone marrow which was subjected to immunomagnetic enrichment using EpCAM beads followed by fluorescence activated cell sorting (IE/FACS) to isolate EpCAM-positive cells away from leukocytes (CD45+). For DNA profiling, sorted cells were subjected to BAC array comparative genomic hybridization analysis following whole genome amplification. For RNA profiling, QPCR analysis was performed on sixty four (64) cancer-related genes using Taqman® low density arrays. For non-tumor controls, RNA profiling was performed on matched leukocytes (CD45+) isolated from the same enriched bone marrow samples.

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Expanded Genomic Profiling of Circulating Tumor Cells in Metastatic Breast Cancer Patients to Assess Biomarker Status and Biology Over Time (CALGB 40502 and CALGB 40503, Alliance).

Project description:We developed a novel approach to isolate tumor cells with high purity from blood which was subjected to immunomagnetic enrichment using EpCAM beads followed by fluorescence activated cell sorting (IE/FACS) to isolate EpCAM-positive cells away from leukocytes (CD45+). Duplicate samples of 20 cells were isolated from the same enriched blood from MBC patients and then subjected to DNA and RNA profiling in parallel. For DNA profiling, sorted cells were subjected to BAC array comparative genomic hybridization analysis following whole genome amplification. For RNA profiling, QPCR analysis was performed on sixty four (64) cancer-related genes using Taqman® low density arrays.

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MicroRNA expression signatures of CD14-CD45-EpCAM+ cells in human bone marrow for colorectal cancer liver metastasis

Project description:To further development of our gene expression approach to biodosimetry, we have employed microRNA microarray expression profiling to identify genes with the potential to distinguish liver metastasis related microRNA. Colorectal cancer patients were administered anesthesia and 20 mL BM was taken from the right and left anterior iliac crests before surgery. Mononucleated cells were collected using a standard Ficoll-Hypaque gradient technique. To enrich for EpCAM+ cells, CD14+ cells were removed from the whole bone marrow using auto MACSTM pro (Milteny Biotec, Bergisch Gladbach, Germany) with anti-CD14 immunomagnetic beads (clone; TÜK4, Milteny Biotec). Next, CD45+ cells were removed by treatment with anti-CD45 immunomagnetic beads (clone; 5B1; Milteny Biotec). The residual CD14?CD45? cells were then incubated with FcR blocking reagent (Milteny Biotec), followed by incubation with anti-EpCAM immunomagnetic beads (clone; HEA-125, Milteny Biotec), and the CD14?CD45?EpCAM+ cells were taken up. Total RNA of these cells we analyzed the microRNA levels of CD14?CD45?EpCAM+ cells obtained from non-metastasis patients (n = 12) and liver metastasis patients (n = 7). Ten-microRNA consensus signature was identified that distinguished between CD14?CD45?EpCAM+ cells from liver metastasis patients and CD14?CD45?EpCAM+ cells from non-liver metastasis patients. MicroRNA expression of CD14-CD45-EpCAM+ cells in human bone marrow was measured. RNA of these cells we analyzed the microRNA levels of CD14?CD45?EpCAM+ cells obtained from non-metastasis patients (n = 12) and liver metastasis patients (n = 7).

2013-11-11 | E-GEOD-51716 | biostudies-arrayexpress

MicroRNA expression signatures of CD14-CD45-EpCAM+ cells in human bone marrow for colorectal cancer liver metastasis

Project description:To further development of our gene expression approach to biodosimetry, we have employed microRNA microarray expression profiling to identify genes with the potential to distinguish liver metastasis related microRNA. Colorectal cancer patients were administered anesthesia and 20 mL BM was taken from the right and left anterior iliac crests before surgery. Mononucleated cells were collected using a standard Ficoll-Hypaque gradient technique. To enrich for EpCAM+ cells, CD14+ cells were removed from the whole bone marrow using auto MACSTM pro (Milteny Biotec, Bergisch Gladbach, Germany) with anti-CD14 immunomagnetic beads (clone; TÜK4, Milteny Biotec). Next, CD45+ cells were removed by treatment with anti-CD45 immunomagnetic beads (clone; 5B1; Milteny Biotec). The residual CD14−CD45− cells were then incubated with FcR blocking reagent (Milteny Biotec), followed by incubation with anti-EpCAM immunomagnetic beads (clone; HEA-125, Milteny Biotec), and the CD14−CD45−EpCAM+ cells were taken up. Total RNA of these cells we analyzed the microRNA levels of CD14−CD45−EpCAM+ cells obtained from non-metastasis patients (n = 12) and liver metastasis patients (n = 7). Ten-microRNA consensus signature was identified that distinguished between CD14−CD45−EpCAM+ cells from liver metastasis patients and CD14−CD45−EpCAM+ cells from non-liver metastasis patients.

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2012-12-31 | E-GEOD-27511 | biostudies-arrayexpress

microRNA expression profiles in bone marrow fractions from gastric cancer patients

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2013-08-23 | E-GEOD-40325 | biostudies-arrayexpress

microRNA expression profiles in bone marrow fractions from gastric cancer patients

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Single cell analysis of Villin-Cre;Apc min;Lef1 fl/fl (VApcMinL) and ApcMin Epcam- cells and CD45-Epcam- stromal cells from lamina propria at the age of 11 wks.

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