{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"omics_type":["Unknown","Transcriptomics","Genomics","Proteomics"],"submitter":["Louise Showe"],"study_type":["transcription profiling by array"],"organism":["Homo sapiens"],"species":["Homo sapiens"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-GEOD-37614"],"description":["Breast cancer is a heterogeneous disease for which prognosis and treatment strategies are largely governed by the receptor status (estrogen, progesterone and Her2-neu) of the tumor cells.  Gene expression profiling of whole breast tumors further stratifies breast cancer into several molecular subtypes which also co-segregate with the receptor status of the tumor cells.  We postulated that cancer associated fibroblasts (CAFs) within the tumor stroma may exhibit subtype specific gene expression profiles and thus contribute to the biology of the disease in a subtype specific manner. Several studies have reported gene expression profile differences between CAFs and normal breast fibroblasts but in none of these studies were the results stratified based on tumor subtypes.  To address whether gene expression in breast cancer associated fibroblasts varies between breast cancer subtypes, we compared the gene expression profiles of early passage primary CAFs isolated from twenty human breast cancer samples representing three main subtypes; seven ER+, seven triple negative (TNBC) and six Her2+.  We observed significant expression differences between CAFs derived from Her2+ breast cancer and CAFs from TNBC and ER+ cancers, particularly in pathways associated with cytoskeleton and integrin signaling.  In the case of Her2+ breast cancer, the signaling pathways found to be selectively up regulated in CAFs may contribute to the more invasive properties and unfavorable prognosis of Her2+ breast cancer. These data demonstrate that in addition to the distinct molecular profiles that characterize the neoplastic cells, CAF gene expression is also differentially regulated in distinct subtypes of breast cancer. We isolated CAFs from twenty primary breast cancer samples representing three main subtypes (ER+ (n=7), TNBC (n=7), Her2+ (n=6)) and performed gene expression profile analyses on RNA isolated from these early passage CAFs.  Those samples were done in two batches with 4 samples repeated in both batches. One TNBC sample was found to be an outlier and not used in the analysis."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Labeling - Biotin Labeling of RNA at 250ng with Ambion Illumina® TotalPrep RNA Amplification Kit(Cat #AMIL1791)","Hybridization - Hyb on Illumina Human-HT12 v4 Expression BeadChip Kit using Illumina whole-Genome Gene Exprssion with Intellihyb Seal system which detect signal with streptavidin-Cy3.","Nucleic Acid Extraction - The tissues were stored in ice cold medium DMEM/F12 supplemented with 10% fetal bovine serum (FBS), penicillin and streptomycin. The fresh tumor tissue was kept on ice at 4oC until ready for processing within 6 hours from the excision time. If the tumor tissue weighed less than 0.5 gram, the tissue was mechanically dissociated by mincing with scalpel and scissors to 1-2 mm3 in a 10 cm tissue culture plate. Fibroblast growth medium (DMEM supplemented with 10% FBS penicillin and streptomycin) was then added. After several days, outgrowth of spindle shaped cells was observed. Tissue debris and non-adherent cells were removed and medium changed between day 2 - 4. For tissues weighing more than 0.5 gram, the tissue was minced as described above and then enzymatically dissociated in tissue digestion buffer containing collagenase I (Worthington), hyaluronidase (Sigma), Collagenase IV (Worthington) at 1 mg/ml of each enzyme in DMEM/F12 medium in a volume of 1:5 ratio of tumor to buffer (wt/vol) on a gyrating platform at 37o C for 30 min. The digestion was quenched by addition of fibroblast growth medium and filtered through a 70 m cell strainer. Cells were pelleted at 1500 rpm for 10 min. Tissue debris and non-adherent cells were removed during medium change between day 2 or 4. By 10 14 days, near confluent adherent spindle shaped cells were harvested using 0.25% trypsin in versene, washed and replated in fresh fibroblast growth medium. Medium was changed every 4 7 days. CAFs from early passages (passage 2 - 3) were harvested and the cell pellet was stored in RNA later (Applied Biosystems) at -80oC until RNA was isolated. RNA purification was carried out using TRI Reagent (Molecular Research Center) according to manufacturer s recommendations. RNA quality was determined using the Bioanalyzer (Agilent). Only samples with 28S/16S ratios >0.75 were used for further studies. Equal amounts (400ng) of total RNA was amplified as recommended by Illumina and hybridized to the HumanHT-12 v4 human whole genome bead arrays.","Scaning - Illumina Setting with PMTFactor=2"],"figure_sub":["MIAME Score","Raw Data","Organization","Assays and Data","Processed Data","MAGE-TAB Files","Array Designs"],"pubmed_authors":["Louise Showe","AV Kossenkov","C Satija","L Chang","LC Showe","M Herlyn","J Tchou","E Puré"],"data_protocol":["Data Transformation - Illumina BeadStudio v.3.0 software was used to export expression levels and detect p-values for each probe of each sample. Quality control of each array was performed using median Spearman correlation computed against all other arrays. Arrays whose median correlation differed from the global correlation by more than 8 absolute deviations were marked as outliers and not used for further analysis, resulting in the removal of one TNBC sample, TB147.  The remaining 19 arrays were then quantile-normalized between each other and filtered to remove non-informative probes (probes with a detection p-value>0.05 in all samples). Between-batch normalization was performed using Distance Weighted Discrimination (DWD) approach using 4 samples replicated in the 2 microarray batches.  Average expression between replicates was used for data analysis. ID_REF =  VALUE = normalized data","Data Transformation - Illumina BeadStudio v.3.0 software was used to export expression levels and detect p-values for each probe of each sample. Quality control of each array was performed using median Spearman correlation computed against all other arrays. Arrays whose median correlation differed from the global correlation by more than 8 absolute deviations were marked as outliers and not used for further analysis, resulting in the removal of one TNBC sample, TB147. The remaining 19 arrays were then quantile-normalized between each other and filtered to remove non-informative probes (probes with a detection p-value>0.05 in all samples). Between-batch normalization was performed using Distance Weighted Discrimination (DWD) approach using 4 samples replicated in the 2 microarray batches. Average expression between replicates was used for data analysis. ID_REF =  VALUE = normalized data"],"additional_accession":[]},"is_claimable":false,"name":"Human breast cancer associated fibroblasts exhibit subtype specific gene expression profiles","description":"Breast cancer is a heterogeneous disease for which prognosis and treatment strategies are largely governed by the receptor status (estrogen, progesterone and Her2-neu) of the tumor cells.  Gene expression profiling of whole breast tumors further stratifies breast cancer into several molecular subtypes which also co-segregate with the receptor status of the tumor cells.  We postulated that cancer associated fibroblasts (CAFs) within the tumor stroma may exhibit subtype specific gene expression profiles and thus contribute to the biology of the disease in a subtype specific manner. Several studies have reported gene expression profile differences between CAFs and normal breast fibroblasts but in none of these studies were the results stratified based on tumor subtypes.  To address whether gene expression in breast cancer associated fibroblasts varies between breast cancer subtypes, we compared the gene expression profiles of early passage primary CAFs isolated from twenty human breast cancer samples representing three main subtypes; seven ER+, seven triple negative (TNBC) and six Her2+.  We observed significant expression differences between CAFs derived from Her2+ breast cancer and CAFs from TNBC and ER+ cancers, particularly in pathways associated with cytoskeleton and integrin signaling.  In the case of Her2+ breast cancer, the signaling pathways found to be selectively up regulated in CAFs may contribute to the more invasive properties and unfavorable prognosis of Her2+ breast cancer. These data demonstrate that in addition to the distinct molecular profiles that characterize the neoplastic cells, CAF gene expression is also differentially regulated in distinct subtypes of breast cancer. We isolated CAFs from twenty primary breast cancer samples representing three main subtypes (ER+ (n=7), TNBC (n=7), Her2+ (n=6)) and performed gene expression profile analyses on RNA isolated from these early passage CAFs.  Those samples were done in two batches with 4 samples repeated in both batches. One TNBC sample was found to be an outlier and not used in the analysis.","dates":{"release":"2013-12-20T00:00:00Z","modification":"2023-08-21T13:19:18.61Z","creation":"2021-10-04T12:20:19Z"},"accession":"E-GEOD-37614","cross_references":{"GEO":["GSE37614"],"EFO":["EFO_0002768"]}}