Comment[ArrayExpressAccession] E-GEOD-46294 MAGE-TAB Version 1.1 Public Release Date 2015-05-01 Investigation Title Integrative miRnome and transcriptome analysis in human malignant prolactin pituitary tumors identify miR-183 as driving gene for tumor progression to malignancy. Comment[Submitted Name] Integrative miRnome and transcriptome analysis in human malignant prolactin pituitary tumors identify miR-183 as driving gene for tumor progression to malignancy. Experiment Description miRNAs expression is known to be deregulated in many types of cancer leading to suspect them implication in crucial steps occurring during tumor progression and maybe as potential driving genes. In this study we tried to decipher the place of miRNA in the hierarchical events cascade occurring in prolactinoma-tumours progression toward malignancy by using an integrative genomic approach performed on the same samples associating global miRNA expression, transcriptomic, DNA structure and methylation analyses. By this approach, we observed the down-regulation of 3 miRNAs (miR-183, miR-98, and miR-744) in A vs. NA tumors. MiR-98 and miR-744 were demonstrated to regulate respectively E2F2 and TGFB1 which are up-regulated in the A vs. NA prolactinomas. Moreover we demonstrated that KIAA0101 is a new target of miR-183 and that miR-183 is regulated by the transcription factor ETS2. KIAA0101 is up-regulated in the A vs. NA prolactinomas. KIAA0101, E2F2 and TGFB1 are interconnected in a pathway controlling cell proliferation and invasion that is over-activated in A PRL tumors. Thus the down-regulation of these three miRNAs is involved in the tumoral progression of human prolactinoma towards aggressiveness. The integrative genomic approach used here allowed us to propose a model of PRL tumoral progression towards aggressiveness. Moreover it revealed the potential of integrative genomic strategy applied in the same human tumors to identify the molecular mechanisms responsible for tumor progression even from a small cohort of patients. miRNA signatures was assessed on 4 aggressive, 8 non-aggressive human prolactin secreting pituitary tumors and 1 normal pituitary for data normalization. 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 roche Roche Nazaret Lachuer Person First Name magali M N J Person Email magali.roche@lyon.unicancer.fr Person Affiliation university lyon 1 Person Address university lyon 1, 8 avenue Rockfeller, Lyon, France Person Roles submitter Protocol Name P-GSE46294-1 P-GSE46294-3 P-GSE46294-4 P-GSE46294-2 P-GSE46294-5 Protocol Description The scanned images were analyzed with Feature Extraction Software 10.7 (Agilent) using default parameters. Raw data files (from Feature Extraction) were then normalized using labelling spikes and each sample from one array was normalized by the normal pituitary of same arrray in Genespring Software 7.0 (Agilent Technologies). It is expected that the values are 1 for GSM1128224 and GSM112822. We have normalized each sample in the batch by the normal pituitary (eg. batch 1 is normalized by sample 0_1 and batch 2 is normalized by sample 0_2). Thus sample 0_1 is normalized by itself and so for sample 0_2. ID_REF = VALUE = Normalized signal intensity 100 ng of total RNA added with labelling Spike-In (from Agilent's microRNA Spike- In kit p/n 5190- 1934) was dephosphorylated, denaturated and then labeled with Cyanine 3-pCp following manufacturer's instructions (Agilent Technologies, miRNA Microarray System with miRNA Complete Labeling and Hyb Kit protocol, version 2.2 october 2009). Then labeled samples were purified using Micro Bio-Spin 6 column from Bio-Rad (optional in protocol). Before hybridization, samples were dried and resuspent in a total volume of 45 µl containing 1X GE Blocking Agent, 1X Hi-RPM Hybridization Buffer, 1 µl of Hybridaton Spike-In (from Agilent's microRNA Spike- In kit p/n 5190- 1934) and incubated 5 min at 100°C. Total mixture was hybridized on Human miRNA Microarray v3, 8x15k for 20 hours at 55°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately. Total RNA from pituitary tumors and normal pituitary was extracted using miRNeasy Mini kit with DNAse treatment (Qiagen, Venlo, Netherlands) according to the manufacturer’s protocol. Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner Type C using AgilentHD_miRNA profile for settings (default). Protocol Type normalization data transformation protocol labelling protocol hybridization protocol nucleic acid extraction protocol array scanning protocol Experimental Factor Name tumor stage sex Experimental Factor Type tumor stage sex Comment[SecondaryAccession] GSE46294 Comment[GEOReleaseDate] 2015-05-01 Comment[ArrayExpressSubmissionDate] 2013-04-23 Comment[GEOLastUpdateDate] 2015-05-01 Comment[AEExperimentType] transcription profiling by array SDRF File E-GEOD-46294.sdrf.txt