Comment[ArrayExpressAccession]	E-GEOD-46824
MAGE-TAB Version	1.1					
Public Release Date	2013-05-10
Investigation Title	Carcinoma-associated fibroblasts’ transcriptomic program predicts clinical outcome in stage II/III colorectal cancer					
Comment[Submitted Name]	Carcinoma-associated fibroblasts’ transcriptomic program predicts clinical outcome in stage II/III colorectal cancer
Experiment Description	We obtained fibroblast cultures from fresh surgical specimen ressected from patients with primary colorectal carcinoma: normal colonic fibroblasts (NCF=9) from the normal colonic mucosa at least 5-10cm from the surgical margin, carcinoma-associated fibroblasts from the primary tumor (CAF-PT=14) and carcinoma-associated fibroblasts (CAF-LM=11) from fresh surgical specimens of liver metastases. We identified 277 probes, in common between the three types of fibroblasts, whose expression level is sequentially deregulated according to cancer progression (NCF→CAF-PT→CAF-LM; fold change Log2 normalized expression>1.5 in each step). Prediction Analysis of Microarrays was applied to obtain a 25-gene signature that better characterizes each fibroblast class. The signature is able to classify patients carrying primary tumors according to prognosis. This fact was exploited to obtain a 19-gene signature (from the 277 deregulated probes) predicting recurrence with high accuracy in stage II/III colorectal cancer patients. Signature validation has been carried out in two independent datasets and in a meta-cohort of 336 stage II/III patients. Since the 25-gene signature was obtained regardless of gene expression data of tumor specimens or patient’s clinical data, the prognostic power of this signature provides strong evidence of the link between the tumor stroma and cancer progression. Furthermore, the 19-gene signature was able to identify low-risk patients with very high accuracy, especially relevant for those high-risk stage-II patients. We hybridised fibroblast RNA in Affymetrix GeneChip 1.0 st arrays					
Term Source Name	ArrayExpress	EFO
Term Source File	http://www.ebi.ac.uk/arrayexpress/	http://www.ebi.ac.uk/efo/efo.owl				
Person Last Name	mollevi	Mollevi	Berdiel			
Person First Name	david	D	M			
Person Mid Initials	g	G				
Person Email	dgmollevi@iconcologia.net					
Person Affiliation	IDIBELL					
Person Address	IDIBELL, Av Gran Via 197-203, L'Hospitalet de Llobregat, Spain					
Person Roles	submitter					
Protocol Name	P-GSE46824-1	P-GSE46824-4	P-GSE46824-5	P-GSE46824-2	P-GSE46824-3	P-GSE46824-6
Protocol Description	We used RMA background correction, quantile normalization and RMA summarization ID_REF =  VALUE = RMA signal	Single-stranded cDNA was generated from the amplified cRNA with the WT cDNA Synthesis Kit (Affymetrix) and then fragmented and labeled with the WT Terminal Labeling Kit (Affymetrix).	Samples were hybridized with GeneChip Human Gene1.0 ST Arrays (Affymetrix) and scanned at the IRB Microarray Core Facility.	We obtained fibroblast cultures from fresh surgical specimen ressected from patients with primary colorectal carcinoma: normal colonic fibroblasts (NCF=9) from the normal colonic mucosa at least 5-10cm from the surgical margin, carcinoma-associated fibroblasts from the primary tumor (CAF-PT=14) and carcinoma-associated fibroblasts (CAF-LM=11) from fresh surgical specimens of liver metastases. All samples were collected under the supervision of the Ethics Committee of the Hospital Universitari de Bellvitge. To establish primary cultures, firstly we homogenated surgical specimens with Collagenase IV and Dispase in a MACS separation unit (Milteny Biotech, Auburn, CA, USA), depleted homogenates from epithelial cells using anti-EPCAM Dynabeads (Invitrogen) and positively selected fibroblasts using anti-fibroblasts microbeads (Milteny Biotech, Auburn, CA, USA). Cells were then cultured in DMEMF12+10%FBS (Gibco) with added penicillin/streptomycin.	Total RNA was isolated from fresh cells using Qiagen Rneasy mini kit	Scanning was done on an Affymetrix GeneChip Scanner 7G
Protocol Type	normalization data transformation protocol	labelling protocol	hybridization protocol	growth protocol	nucleic acid extraction protocol	array scanning protocol
Experimental Factor Name	CELL TYPE	STAGE	SEX	AGE	ORGANISM PART	
Experimental Factor Type	cell type	Stage	sex	age	organism part	
Comment[SecondaryAccession]	GSE46824					
Comment[GEOReleaseDate]	2013-05-10					
Comment[ArrayExpressSubmissionDate]	2013-05-10					
Comment[GEOLastUpdateDate]	2013-05-10					
Comment[AEExperimentType]	transcription profiling by array					
SDRF File	E-GEOD-46824.sdrf.txt