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E-TOXM-17
Mus musculus
30 Downloadable Samples
Affymetrix Murine Genome U74A Version 2 Array (mgu74av2)
Description
The multi-lab International Life Sciences Institute (ILSI) project on the application of genomics to risk assessment offered the unique opportunity to investigate the influence of variability within and between laboratories on identifying biologically relevant gene expression changes. We assessed the gene expression profiles of mouse lymphoma L5178Y cells treated with hydroxyurea (HU) in three independent studies from two different laboratories, Sanofi-Aventis and Procter and Gamble. Cells were dosed for 4 hr and harvested immediately at the end of the treatment or after a 20-hr recovery period. Cytotoxicity and genotoxicity were evaluated by standard assays. Statistical analysis of these data revealed that, while gene expression responses to HU treatment were markedly different at 4 hr vs. 24 hr, there was otherwise a consistent pattern of dose-response across the three studies. Therefore, the studies were merged and each time point was evaluated separately. At 4 hr, we identified 173 (P lt 0.0001) dose-responsive genes with a common trend in all three studies. These were mainly associated with the cell cycle, DNA repair and DNA metabolism, and in particular, the intra-S and G2/M phase checkpoints. At 24 hr, we identified 434 dose-responsive genes common across studies. These genes were involved in lymphocyte-specific activities and the activation of apoptosis via the caspase cascade. Our results show that despite inter-laboratory variability, combining the three studies in a single statistical analysis identifies more significantly-modulated genes than in any of the individual studies, due to improved statistical sensitivity. The genes identified in our study provide information that is relevant to HU biology.
Publication Title
Laboratory variability does not preclude identification of biological functions impacted by hydroxyurea.
Sample Metadata Fields
Sex, Disease, Disease stage, Compound, Time
View Samples- Mus musculus
- 28 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
The activation of the transcription factor NF-E2-related factor 2 (Nrf2) maintains cellular homeostasis in response to oxidative stress by the regulation of multiple cytoprotective genes. Without stressors the activity of Nrf2 is inhibited by its interaction with the kelch-like ECH-associated protein 1 (Keap1). Here, we describe RA839, a small molecule that binds non-covalently to the Nrf2-interacting kelch domain of Keap1 with a Kd of approximately 6 M, as demonstrated by X-ray co-crystallization and isothermal titration calorimetry. Whole-genome DNA arrays showed that at 10 M RA839 significantly regulated 105 genes in bone marrow-derived macrophages. Canonical pathway mapping of these genes revealed an activation of pathways linked with Nrf2 signalling. These pathways were also activated after the activation of Nrf2 by the silencing of Keap1 expression. RA839 regulated only two genes in Nrf2 knockout macrophages. Similar to the activation of Nrf2 by either silencing of Keap1 expression or by the reactive compound CDDO-Me, RA839 prevented the induction of both inducible nitric oxide synthase expression and nitric oxide release in response to lipopolysaccharides in macrophages. In mice RA839 acutely induced Nrf2-target gene expression in liver. RA839 is a selective inhibitor of the Keap1/Nrf2 interaction and a useful tool compound to study the biology of Nrf2.
Publication Title
Characterization of RA839, a Noncovalent Small Molecule Binder to Keap1 and Selective Activator of Nrf2 Signaling.
Sample Metadata Fields
Specimen part, Treatment
View Samples- Mus musculus
- 24 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
The transcription factor NF-E2-related factor 2 (Nrf2) induces cytoprotective genes, but has also been linked to the regulation of hepatic energy metabolism. In order to assess the pharmacological potential of hepatic Nrf2 activation in metabolic disease, Nrf2 was activated over 8 weeks in mice on Western diet using two different siRNAs against kelch-like ECH-associated protein 1 (Keap1), the inhibitory protein of Nrf2. Whole genome expression analysis followed by pathway analysis demonstrated that the suppression of Keap1 expression induced genes that are involved in anti-oxidative stress defense and biotransformation, pathways proving the activation of Nrf2 by the siRNAs against Keap1. The expression of neither fatty acid- nor carbohydrate-handling proteins was regulated by the suppression of Keap1. Metabolic profiling of the animals did also not show effects on plasma and hepatic lipids, energy expenditure or glucose tolerance by the activation of Nrf2. The data indicate that hepatic Nrf2 is not a major regulator of intermediary metabolism in mice.
Publication Title
Chronic Activation of Hepatic Nrf2 Has No Major Effect on Fatty Acid and Glucose Metabolism in Adult Mice.
Sample Metadata Fields
Specimen part, Treatment
View Samples- Homo sapiens
- 16 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Histone deacetylases (HDACs) are important regulators of epigenetic gene modification that are involved in the transcriptional control of metabolism. In particular class IIa HDACs have been shown to affect hepatic gluconeogenesis and previous approaches revealed that their inhibition reduces blood glucose in type 2 diabetic mice. In the present study, we aimed to evaluate the potential of class IIa HDAC inhibition as a therapeutic opportunity for the treatment of metabolic diseases. For that, siRNAs selectively targeting HDAC4, 5 and 7 were selected and used to achieve a combinatorial knockdown of these three class IIa HDAC isoforms. Subsequently, the hepatocellular effects as well as the impact on glucose and lipid metabolism were analyzed in vitro and in vivo. The triple knockdown resulted in a statistically significant decrease of gluconeogenic gene expression in a murine hepatic cell line as well as in human primary hepatocytes. Despite a similar HDAC-induced downregulation of hepatic genes involved in gluconeogenesis in mice using a liver-specific lipid nanoparticle siRNA formulation, the in vivo effects on whole body glucose metabolism were only limited and did not outweigh the safety concerns observed by histopathological analysis in spleen and kidney. Mechanistically, Affymetrix gene chip analysis and gene expression studies provide evidence that class IIa HDACs directly target and thus regulate the expression of HNF4α and FOXP1 in the liver, thereby modifying gene regulatory mechanisms mediating glucose and lipid metabolism and transport. In conclusion, the combinatorial knockdown of HDAC4, 5 and 7 by therapeutic siRNAs affected multiple pathways in vitro and in vivo leading to the downregulation of genes involved in gluconeogenesis. However, the effects on the gene expression level were not paralleled by a significant reduction of gluconeogenesis in mice, as shown in pyruvate tolerance tests. However, the liver-specific inhibition of these HDAC isoforms was associated with severe adverse effects in vivo, making this approach not a viable treatment option for chronic metabolic disorders like type 2 diabetes.
Publication Title
No associated publication
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 264 Downloadable Samples
- Affymetrix Human Genome U133A Array (hgu133a)
Description
The aims of this study were to assess the feasibility of prospective pharmacogenomics research in multicenter international clinical trials of bortezomib in multiple myeloma and to develop predictive classifiers of response and survival with bortezomib. Patients with relapsed myeloma enrolled in phase 2 and phase 3 clinical trials of bortezomib and consented to genomic analyses of pretreatment tumor samples. Bone marrow aspirates were subject to a negative-selection procedure to enrich for tumor cells, and these samples were used for gene expression profiling using DNA microarrays. Data quality and correlations with trial outcomes were assessed by multiple groups. Gene expression in this dataset was consistent with data published from a single-center study of newly diagnosed multiple myeloma. Response and survival classifiers were developed and shown to be significantly associated with outcome via testing on independent data. The survival classifier improved on the risk stratification provided by the International Staging System. Predictive models and biologic correlates of response show some specificity for bortezomib rather than dexamethasone. Informative gene expression data and genomic classifiers that predict clinical outcome can be derived from prospective clinical trials of new anticancer agents.
Publication Title
Gene expression profiling and correlation with outcome in clinical trials of the proteasome inhibitor bortezomib.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 172 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Glatiramer Acetate (GA) has provided safe and effective treatment for multiple sclerosis (MS) patients for two decades. It acts as an antigen, yet the precise mechanism of action remains to be fully elucidated, and no validated pharmacokinetic or pharmacodynamic biomarkers exist. In order to better characterize GAs biological impact, genome-wide expression studies were conducted with a human monocyte (THP-1) cell line. Consistent with previous literature, branded GA upregulated antiinflammatory markers (e.g. IL10), and modulated multiple immune-related pathways. Despite some similarities, significant differences were observed between expression profiles induced by branded GA and Probioglat, a differently-manufactured glatiramoid purported to be a generic GA.
Publication Title
Gene expression studies of a human monocyte cell line identify dissimilarities between differently manufactured glatiramoids.
Sample Metadata Fields
Cell line, Treatment, Time
View Samples- Mus musculus
- 103 Downloadable Samples
- Affymetrix Mouse Expression 430A Array (moe430a)
Description
We have developed a total RNA amplification and labeling strategy for use with Affymetrix GeneChips. Our protocol, which we denote BIIB, employs two rounds of linear T7 amplification followed by Klenow labeling to generate a biotinylated cDNA. In benchmarking studies using a titration of mouse universal total RNA, BIIB outperformed commercially available kits in terms of sensitivity, accuracy, and amplified target length, while providing equivalent results for technical reproducibility. BIIB maintained 50 and 44% present calls from 100 and 50 pg of total RNA, respectively. Inter- and intrasample precision studies indicated that BIIB produces an unbiased and complete expression profile within a range of 5 ng to 50 pg of starting total RNA. From a panel of spiked exogenous transcripts, we established the BIIB linear detection limit to be 20 absolute copies. Additionally, we demonstrate that BIIB is sensitive enough to detect the stochastic events inherent in a highly diluted sample. Using RNA isolated from whole tissues, we further validated BIIB accuracy and precision by comparison of 224 expression ratios generated by quantitative real-time PCR. The utility of our method is ultimately illustrated by the detection of biologically expected trends in a T cell/B cell titration of 100 primary cells flow sorted from a healthy mouse spleen.
Publication Title
Accurate and precise transcriptional profiles from 50 pg of total RNA or 100 flow-sorted primary lymphocytes.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 50 Downloadable Samples
- Affymetrix Human Genome U219 Array (hgu219)
Description
Proliferating tumor cells use aerobic glycolysis to support their high metabolic demands. Paradoxically, increased glycolysis is often accompanied by expression of the lower activity PKM isoform, effectively constraining lower glycolysis. Here, we report the discovery of novel PKM activators with a unique allosteric binding mode. Characterization of how these compounds impact cancer cells revealed an unanticipated link between glucose and amino acid metabolism. PKM activation resulted in a metabolic rewiring of cancer cells manifested by a profound dependency on the non-essential amino acid serine for continued cell proliferation. Induction of serine auxotrophy by PKM activation was accompanied by reduced carbon flow into the serine biosynthetic pathway and increased expression of high affinity serine transporters. These data support the hypothesis that PKM expression confers metabolic flexibility to cancer cells that allows adaptation to nutrient stress.
Publication Title
Small molecule activation of PKM2 in cancer cells induces serine auxotrophy.
Sample Metadata Fields
Cell line, Treatment, Time
View Samples- Homo sapiens
- 39 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Transcriptional analysis of the superior temporal cortex (BA22) in schizophrenia: Pathway insight into disease pathology and drug development
Publication Title
Transcription and pathway analysis of the superior temporal cortex and anterior prefrontal cortex in schizophrenia.
Sample Metadata Fields
Sex, Age, Specimen part, Disease, Disease stage
View Samples- Homo sapiens
- 34 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
RSPO is a WNT pathway activator and functions as a potent regulator of stem cell growth in colon. RSPO family members were produced by several human tumors representing multiple tumor types including ovarian, pancreatic, colon, breast and lung cancer. Specific monoclonal antibody antagonists of RSPO family members were developed. In human patient-derived tumor xenograft models, anti-RSPO treatment markedly inhibited tumor growth either as single agents or in combination with chemotherapy. Furthermore, blockade of RSPO signaling reduced the tumorigenicity of cancer cells based on serial transplantation studies.
Publication Title
Therapeutic Targeting of Tumor-Derived R-Spondin Attenuates β-Catenin Signaling and Tumorigenesis in Multiple Cancer Types.
Sample Metadata Fields
Specimen part
View Samples