Gene expression profile of cancer cell lines of breast, lung, pancreatic, gasctric, ovarian, hepatocellular, prostate carcinomas and melanomas.
Gene expression profiling of 30 cancer cell lines predicts resistance towards 11 anticancer drugs at clinically achieved concentrations.
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View SamplesThe routine workflow for invasive cancer diagnostics is based on biopsy processing by formalin fixation and subsequent paraffin embedding. Formalin-fixed paraffin-embedded (FFPE) tissue samples are easy to handle, stable and particularly suitable for morphologic evaluation, immunohistochemistry and in situ hybridization. However, it has become a paradigm that these samples cannot be used for genome-wide expression analysis with microarrays. To oppose this view, we present a pilot microarray study using FFPE core needle biopsies from breast cancers as RNA source. We found that microarray probes interrogating sequences near the poly-A-tail of the transcribed genes were well suitable to measure RNA levels in FFPE core needle biopsies. For the ER and the HER2 gene, we observed strong correlations between RNA levels measured in these probe sets and protein expression determined by immunohistochemistry (p = 0.000003 and p = 0.0022). Further, we have identified a signature of 364 genes that correlated with ER protein status and a signature of 528 genes that correlated with HER2 protein status. Many of these genes (ER: 60%) could be confirmed by analysis of an independent publicly available data set. Finally, a hierarchical clustering of the biopsies with respect to three recently reported gene expression grade signatures resulted in widely stable low and high expression grade clusters that correlated with the pathological tumor grade. These findings support the notion that clinically relevant information can be gained from microarray based gene expression profiling of FFPE cancer biopsies. This opens new opportunities for the integration of gene expression analysis into the workflow of invasive cancer diagnostics as well as translational research in the setting of clinical studies.
Genome-wide gene expression profiling of formalin-fixed paraffin-embedded breast cancer core biopsies using microarrays.
Disease stage
View SamplesIn osteosarcoma patients, the development of metastases, often to the lungs, is the most frequent cause of death. To improve this situation, a deeper understanding of the molecular mechanisms governing osteosarcoma development and dissemination and the identification of novel drug targets for an improved treatment are needed. Towards this aim, we characterized osteosarcoma tissue samples compared to primary osteoblast cells using Affymetrix HG U133A microarrays.
De novo expression of EphA2 in osteosarcoma modulates activation of the mitogenic signalling pathway.
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View SamplesWe sequenced mRNA from FACS purified hair follicle bulge stem cells from 21 d old control and ILK-deficient mice, 3 biological replicates each Overall design: Examination of mRNA levels in control and ILK-deficient hair follicle bulge stem cells
Integrin-linked kinase regulates the niche of quiescent epidermal stem cells.
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View SamplesThe glycopeptide antibiotic vancomycin (VCM) represents one of the last lines of defense against methicillin-resistant Staphylococcus aureus infections. However, vancomycin is nephrotoxic, but the mechanism of toxicity is still unclear.
Gene expression analysis reveals new possible mechanisms of vancomycin-induced nephrotoxicity and identifies gene markers candidates.
Specimen part
View SamplesProtein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation and translation. We have developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing (Baltz and Munschauer et al. 2012). Our current work focuses on streamlining and extending protein occupancy profiling on poly(A)-RNA. Our objectives are to identify previously unknown protein-bound transcripts and, more importantly, to assess global and local differences in protein occupancy across different biological conditions. To this end, we have implemented poppi, the first pipeline for differential analysis of protein occupancy profiles. We have applied our analysis pipeline to pinpoint changes in occupancy profiles of MCF7 cells against already published HEK293 cells [GSE38157]. Overall design: We generated protein occupancy cDNA libraries for two biological replicates. Briefly, we crosslinked 4SU-labeled MCF7 cells and purified protein-mRNA complexes using oligo(dT)-beads. The precipitate was treated with RNAse I to reduce the protein-crosslinked RNA fragments to a length of about 30-60 nt. To remove non-crosslinked RNA, protein-RNA complexes were precipitated with ammonium sulfate and blotted onto nitrocellulose. The RNA was recovered by Proteinase K treatment, ligated to cloning adapters, and reverse transcribed. The resulting cDNA libraries were PCR-amplified and next-generation sequenced.
Differential protein occupancy profiling of the mRNA transcriptome.
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View SamplesRNA helicases are important regulators of gene expression that act by remodeling RNA secondary structures and as RNA-protein interactions. Here, we demonstrate that MOV10 has an ATP-dependent 5'' to 3'' in vitro RNA unwinding activity and determine the RNA-binding sites of MOV10 and its helicase mutants using PAR-CLIP. We find that MOV10 predominantly binds to 3'' UTRs upstream of regions predicted to form local secondary structures and provide evidence that MOV10 helicase mutants are impaired in their ability to translocate 5'' to 3'' on their mRNA targets. MOV10 interacts with UPF1, the key component of the nonsense-mediated mRNA decay pathway. PAR-CLIP of UPF1 reveals that MOV10 and UPF1 bind to RNA in close proximity. Knockdown of MOV10 resulted in increased mRNA half-lives of MOV10-bound as well as UPF1-regulated transcripts, suggesting that MOV10 functions in UPF1-mediated mRNA degradation as an RNA clearance factor to resolve structures and displace proteins from 3'' UTRs. Overall design: Flp-In T-REx HEK293 cells expressing FLAG/HA-tagged MOV10 WT, MOV10 K530A, MOV10 D645N and UPF1 were used to determine the protein-RNA interaction sites of RNA helicases MOV10 and UPF1 as well as MOV10 inactive variants using PAR-CLIP in combination with next generation sequencing. mRNA half-life changes of MOV10-targeted mRNA were determined by measuring mRNA half-lives by mRNA sequencing of mock and MOV10-depleted HEK293 cells.
MOV10 Is a 5' to 3' RNA helicase contributing to UPF1 mRNA target degradation by translocation along 3' UTRs.
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View SamplesOvarian carcinoma has the highest mortality rate among gynecological malignancies. In this project, we investigated the hypothesis that molecular markers are able to predict outcome of ovarian cancer independently of classical clinical predictors, and that these molecular markers can be validated using independent data sets. We applied a semi-supervised method for prediction of patient survival. Microarrays from a cohort of 80 ovarian carcinomas (TOC cohort) were used for the development of a predictive model, which was then evaluated in an entirely independent cohort of 118 carcinomas (Duke cohort). A 300 gene ovarian prognostic index (OPI) was generated and validated in a leave-one-out approach in the TOC cohort (Kaplan-Meier analysis, p=0.0087). In a second validation step the prognostic power of the OPI was confirmed in an independent data set (Duke cohort, p=0.0063). In multivariate analysis, the OPI was independent of the postoperative residual tumour, the main clinico-pathological prognostic parameter with an adjusted hazard ratio of 6.4 (TOC cohort, CI 1.8 23.5, p=0.0049) and 1.9 (Duke cohort, CI 1.2 3.0, p=0.0068). We constructed a combined score of molecular data (OPI) and clinical parameters (residual tumour), which was able to define patient groups with highly significant differences in survival. The integrated analysis of gene expression data as well as residual tumour can be used for optimised assessment of prognosis. As traditional treatment options are limited, this analysis may be able to optimise clinical management and to identify those patients that would be candidates for new therapeutic strategies.
A prognostic gene expression index in ovarian cancer - validation across different independent data sets.
Specimen part, Disease stage
View Samplesin vitro microarray study of transcriptional changes of jejunal cells
Deoxynivalenol Affects Cell Metabolism and Increases Protein Biosynthesis in Intestinal Porcine Epithelial Cells (IPEC-J2): DON Increases Protein Biosynthesis.
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View Samplesin vitro microarray study of transcriptional changes of jejunal cells
Deoxynivalenol Affects Cell Metabolism and Increases Protein Biosynthesis in Intestinal Porcine Epithelial Cells (IPEC-J2): DON Increases Protein Biosynthesis.
No sample metadata fields
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