title: Human Cavernosal Endothelial Cell Phenotype

status: Public on Sep 28 2009

city: Seattle

first: Hunter

last: Wessells

state: WA

address: 300 Ninth Avenue

department: Urology

institute: University of Washington

summary:
Purpose: To identify the molecular phenotype of endothelial cells (EC) isolated from the unique vasculature of the corpus cavernosum.
Methods: Human EC derived from corpus cavernosum (HCCEC, n=5), coronary artery (HCAEC, n=4) and umbilical vein (HUVEC, n=3) were grown in culture and mRNA transcripts quantified by Affymetrix GeneChip microarrays. Genes differentially expressed across samples were partitioned around medoids to identify genes with highest expression in HCCEC. Several genes were verified by real-time PCR. The role of claudin 11 (CLDN11) in endothelial cell barrier function was examined by in vitro transendothelial electrical resistance assay.
Results: Unsupervised hierarchical clustering of samples using all expressed genes yielded discrete groupings of EC lines according to their in vivo source of origin. 190 genes/transcripts were highly expressed only in cavernosal HCCEC. Gene Ontology classification indicated cavernosal enrichment in genes related to cell adhesion, extracellular matrix (ECM), pattern specification and organogenesis. KEGG pathway analysis showed high expression of gene relating to ECM-receptor interaction, focal adhesions, and cytokine-cytokine receptor interaction. Real-time PCR confirmed expression differences in cadherins 2 and 11, CLDN11, desmoplakin and versican. CLDN11, a component of tight junctions not previously described in ECs, was highly expressed only in HCCEC and its knockdown by siRNA significantly reduced transendothelial electrical resistance in HCCEC.
Conclusions: HCCECs displayed expression of transcripts encoding matrix and adhesion proteins that regulate structural and functional characteristics of blood vessels. High expression of the tight junction protein CLDN11 is novel in endothelial cells and  contributes to barrier function of cultured HCCEC.
Keywords: penis, tight junctions, microarray, gene ontology, cell type comparison

overall_design:
Endothelial cells were grown on 2% gelatin type B (Sigma)/15% fetal bovine serum (Gibco) coated tissue culture dishes in Medium 199 (Gibco) supplemented with 20% FBS, EGM-2MV (Clonetics), L-glutamine and 10,000u/ml Penicillin, 10,000u/ml Streptomycin, and 25mg/ml Fungizone. All cells were grown in a humidified incubator at 37 oC and 5% CO2 with media changed every 2-3 days. Total RNA isolation for the arrays was performed using Qiagen RNeasy mini columns according to manufacturers instructions (RNeasy Mini Handbook). Briefly, cells grown in T-75 culture flasks were trypsinized with 0.25% trypsin/1 mM EDTA (Gibco) between the third and seventh passages and centrifuged to pellet the cells. Next, cells were disrupted and homogenized in RNeasy lysis buffer using a 1ml syringe with 21-gauge needle. Ethanol was added to allow binding of RNA to the silica-gel-based membrane of the mini column. Several washes as well as on-column DNase treatment were used to ensure that contaminants and DNA were effectively removed from the column. Finally, RNA was eluted in approximately 30ul of RNase free water.
Biotin labeled target cRNA was prepared according to the Affymetrix eukaryotic target labeling protocol starting with 5ug total RNA for each sample. Each target was hybridized to an Affymetrix Human U133A GeneChip. GeneChips were washed and scanned at the University of Washington’s Center for Expression Arrays according to procedures developed by the manufacturer.

first: Chris

last: Sullivan

first: Karen

last: Engel

first: Yoshi

last: Tsuboto

first: Bryan

last: Kim

first: Daniel

last: Thorner

first: Kanchan

last: Chitaley


title: RMA expression data for liver samples from subjects with HCV cirrhosis with and without concomitant HCC

status: Public on Sep 11 2009

city: Columbus

first: Kellie

middle: J.

last: Archer

state: OH

address: 1841 Neil Ave

department: Division of Biostatistics

country: USA

institute: Ohio State University

summary:
In this study, we used the Affymetrix HG-U133A 2.0 GeneChip for deriving a multigenic classifier capable of predicting HCV+cirrhosis with vs without concomitant HCC.
We studied gene expression in cirrhotic tissues with (N=16) and without (N=47) HCC.

Keywords: cross-sectional

overall_design: Liver tissue samples were obtained from patients waiting for liver transplantation. For each sample, RNA was extracted and hybridized to an Affymetrix GeneChip.

middle: J

first: Valeria

middle: R

last: Mas

first: Krystle

last: David

middle: G

last: Maluf

last: Bornstein

first: Robert

middle: A

last: Fisher


title: Affymetrix SNP array data for Sézary Syndrome (SS) samples

status: Public on Sep 30 2009

city: Roma

first: Elisabetta

last: Caprini

state: Italy

address: Via dei Monti di Creta 104

country: Italy

institute: IDI-IRCCS

type:
SNP genotyping by SNP array
Genome variation profiling by SNP array

summary: This study used tumour and paired normal samples from 28 Sézary Syndrome (SS) patients to define recurrent regions of chromosomal aberrations. Our data identified recurrent losses of 17p13.2-p11.2 and 10p12.1-q26.3 occurring in 71 and 68% of cases respectively; common gains were detected for 17p11.2-q25.3 (64%) and chromosome 8/8q (50%). Moreover, we identified novel genomic lesions recurring in more than 30% of tumours: loss of 9q13-q21.33 and gain of 10p15.3-10p12.2. In the Sézary Syndrome cases analysed, we could find several small and few large Uniparental Disomies involving interstitial or telomeric regions of LOH occurring mainly for chromosome 10 and to a lesser extent for chromosome 9 and 17. In the attempt to correlate Copy Number data and clinical parameters we find a relationship between complex pattern of chromosomal aberrations, involving at least three recurrent Copy Number alterations, and shorter survival. Integrating mapping and transcriptional data we were able to identify a total of 113 deregulated transcripts in aberrant chromosomal regions that included cancer related genes such as members of the NF-kB pathway (BAG4, BTRC, NKIRAS2, PSMD3, TRAF2) that might explain its constitutive activation in CTCL. Matching this list of genes with those discriminating patients with different survival times we identify several common candidates that might exert critical roles in Sézary Syndrome, like BUB3 and PIP5K1B.

overall_design: Affymetrix SNP arrays were performed, according to the manufacturer's directions, on DNA extracted from cryopreserved lymphomonocytes purified by Ficoll density gradient centrifugation and positive selection using anti-human CD3-conjugated dynabeads (tumour samples) and from granulocytes (normal matched cells) obtained by collecting the upper phase overlying the Ficoll density gradient sediment. Copy number and LOH analyses of Affymetrix 10K SNP arrays were performed for 28 tumour samples and 28 matched normal samples used as references.

first: Diego

last: Arcelli


title: Modeling Inducible Human Tissue Neoplasia Identifies an ECM Interaction Network Involved in Cancer Progression

status: Public on Jun 02 2009

city: Stanford

first: Paul

last: Khavari

state: CA

address: 259 Campus Dr

department: Dermatology

institute: Stanford University

summary: To elucidate mechanisms of cancer progression, we generated inducible human neoplasia in 3-dimensionally intact epithelial tissue. Gene expression profiling of both epithelia and stroma at specific time points during tumor progression revealed sequential enrichment of genes mediating discrete biologic functions in each tissue compartment. A core cancer progression signature was distilled using the increased signaling specificity of downstream oncogene effectors and subjected to network modeling. Network topology predicted that tumor development depends upon specific ECM-interacting network hubs. Blockade of one such hub, the b1 integrin subunit, disrupted network gene expression and attenuated tumorigenesis in vivo. Thus, integrating network modeling and temporal gene expression analysis of inducible human neoplasia provides an approach to prioritize and characterize genes functioning in cancer progression.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
There are 3 experiments in this study: (1a) a time course of  human epidermal tissue transformed by oncogenic ER:H-Ras and IkBaM comprising days 0, 5, 20, and 35 post Ras activation using 4OHT; (1b) a matched time course of adjacent mouse stromal tissue during tumor progression; (2) Arrays comparing 4OHT-induced, Raf-1:ER/IkBaM transformed epidermal tissue and -4OHT controls; and (3) Arrays comparing Ras:ER/IkBaM grafts co-treated with 4OHT and either IgG control antibody or an anti-b1 integrin blocking antibody, P5D2 for 30 days. All arrays were done in biologic duplicate.

Epidermal tissue co-expressing ER:H-RasG12V (ER:Ras) and IkBaM was regenerated on female scid/scid mouse recipients. Grafts were allowed to heal for at least 3 weeks before Ras activation via daily i.p. injections of 730ug of 4OHT (in 110ul of a corn oil/ethanol mixture).  Duplicate grafts were harvested after  0, 5, 20, and 35 days of 4OHT treatment. Laser capture microdissection was utilized to independently isolate epithelial cells of the basal most layers and adajacent stromal tissue for each time point. RNA was subjected to one round of T7-based linear amplification (Ambion Message Amp II enhanced kit) and hybridized to either HG-U133A 2.0 or MG-430A affymetrix oligonucleotide arrays. Arrays from this experiment are labeled Ras and Stroma, respectively. The second experment was performed on Raf-1:ER/IkBaM expressing grafts with and without 4OHT treament for 30 days. The third experiment was done comparing ER:Ras/IkBaM grafts concomitantly treated with 4OHT and 1.5mg/week of either IgG control antibody or a mouse monoclonal blocking antibody against b1 integrin for 30 days. For experiments 2 and 3, HG-U133A 2.0 GeneChips were used; 4OHT treatment and RNA isolation/amplification was performed as above.

first: Jason

last: Reuter


title: Identification of regions and genes important in Sézary syndrome pathogenesis using genomic and expression microarrays

type:
SNP genotyping by SNP array
Genome variation profiling by SNP array
Expression profiling by array

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Expression analysis upon NAMPT knockdown in MCF-7 cells

status: Public on May 28 2009

city: Dallas

first: W. Lee

last: Kraus

state: TX

address: 5323 Harry Hines Blvd.

institute: UT Southwestern Medical Center

summary: NAMPT is an enzyme in the mammalian NAD+ salvage pathway. Expression microarray analysis was used to study the effect of NAMPT knockdown on gene expression in MCF-7 breast cancer cells.

overall_design: Stable knockdown of NAMPT was achieved using a retroviral shRNA construct. An shRNA directed against Luciferase was used to generate the Luc control cells.  Three independent biological replicates with matching Luc controls were analyzed using Affymetrix U133 A2.0 microarrays.

first: Tong

last: Zhang

first: William

middle: L


title: Affymetrix Gene Expression array data for Sézary Syndrome (SS) samples

overall_design: Affymetrix Gene Expression arrays were performed, according to the manufacturer's directions, on RNA extracted from cryopreserved lymphomonocytes purified by Ficoll density gradient centrifugation and positive selection using anti-human CD3-conjugated dynabeads (tumour samples). Gene expression analyses of Affymetrix HG-U133 A arrays were performed for 32 tumour samples.


title: Expression data from prostate cancer epithelial cells from African American and European American men

status: Public on Jul 28 2009

city: Washington

first: Olga

last: Timofeeva

state: DC

address: 3900 Reservoir Rd NW

department: Oncology

institute: Georgetown University Medical Center

summary:
African-American (AA) men experience increased risk of developing prostate cancers as well as increased mortality following treatment as compared to European-American (EA) men. The aim of our study was to identify biological factors with potential to predispose AA men to prostate tumor progression and metastasis.
High-throughput microarrays were used to investigate differences in global gene expression comparing the two groups.

overall_design: To identify cancer-specific gene expression patterns in AA men, we established primary prostate cancer epithelial cells from 14 AA and 13 EA men. Cells were cultured for RNA extraction and hybridization on Affymetrix microarrays.

first: Anatoly

last: Dritschilo


title: Transcription patterns during wheat development

status: Public on Aug 11 2009

city: Ames

first: PLEXdb Curator

middle: Sudhansu

state: IA

address: 1023  Crop Genomes Informatics Lab (Iowa State University)

department: Virtual Reality Application Center

institute: Iowa State University

summary:
The analysis of gene expression during wheat development:
Gene expression measurements were carried out on a developmental tissue
series for wild-type wheat (cv. Chinese Spring) using the Affymetrix
Wheat GeneChip.  Thirteen tissues at defined developmental stages were
chosen to match the barley (cv. Morex) tissue series of Druka et al.
2006 that used the Affymetrix Barley1 GeneChip.  Three replicates of:
root tissue at two different developmental stages, leaf, crown,
caryopsis, anther, pistil, inflorescence, bracts, mesocotyl, endosperm,
embryo and coleoptiles were hybridised.  Comparisons between this wheat
data and the barley dataset were performed and are available at
http://contigcomp.acpfg.com.au

 [PLEXdb(http://www.plexdb.org) has submitted this series at GEO on behalf of the original contributor,  Tim Sutton.  The equivalent experiment is TA3 at PLEXdb.]

overall_design: tissue type: germinating seed, coleoptile(3-replications);  tissue type: germinating seed, root(3-replications);  tissue type: germinating seed, embryo(3-replications);  tissue type: seedling, root(3-replications);  tissue type: seedling, crown(3-replications);  tissue type: seedling, leaf(3-replications);  tissue type: immature inflorescence(3-replications);  tissue type: floral bracts, before anthesis(3-replications);  tissue type: pistil, before anthesis(3-replications);  tissue type: anthers, before anthesis(3-replications);  tissue type: 3-5 DAP caryopsis(3-replications);  tissue type: 22 DAP embryo(3-replications);  tissue type: 22 DAP endosperm(3-replications)

first: Tim

last: Sutton


title: MCF7 time-series upon RITA treatment.

status: Public on May 12 2009

city: Stockholm

first: Martin

last: Enge

address: CCK, Z4

department: Dep of Oncology-Pathology

country: Sweden

institute: Karolinska Institute

summary:
Targeting “oncogene addiction” is a promising strategy for anti-cancer therapy.  Here, we report a potent inhibition of crucial oncogenes by p53 upon reactivation with small molecule RITA in vitro and in vivo. RITA-activated p53 unleashes transcriptional repression of anti-apoptotic proteins Mcl-1, Bcl-2, MAP4, and survivin, blocks Akt pathway on several levels and downregulates c-Myc, cyclin E and B-catenin. p53 ablates c-Myc expression via several mechanisms at transcriptional and posttranscriptional level. We show that transrepression of oncogenes correlated with higher level of p53 bound to chromatin-bound p53 than transactivation of pro-apoptotic targets.  Inhibition of oncogenes by p53 reduces the cell’s ability to buffer pro-apoptotic signals and elicits robust apoptosis. Our study highlights the role of transcriptional repression for p53-mediated tumor suppression.

Keywords: time course

overall_design: Breast carcinoma cell-line MCF7 was treated with the small-molecule p53 activator RITA for 2h, 8h, 16h and 24h.

first: Angela

last: Gluch


title: Prediction of acute multiple sclerosis relapses by transcription levels of peripheral blood cells

status: Public on Jul 15 2009

city: Ramat Gan

first: Michael

last: Gurevich

address: Tel Hashomer

department: Multiple Sclerosis Center

country: Israel

institute: Sheba Medical Center

summary:
Background: The ability to predict the spatial frequency of relapses in multiple sclerosis (MS) would enable treating physicians to decide when to intervene more aggressively and to plan clinical trials more accurately.  Methods: In the current study our objective was to determine if subsets of genes can predict the time to the next acute relapse in patients with MS. Data-mining and predictive modeling tools were utilized to analyze a gene-expression dataset of 94 non-treated patients; 62 patients with definite MS and 32 patients with clinically isolated syndrome (CIS). The dataset included the expression levels of 10,594 genes and annotated sequences corresponding to 22,215 gene-transcripts that appear in the microarray.  Results: We designed a two stage predictor. The first stage predictor was based on the expression level of 10 genes, and predicted the time to next relapse with a resolution of 500 days (error rate 0.079, p< 0.001). If the predicted relapse was to occur in less than 500 days, a second stage predictor based on an additional different set of 9 genes was used, resulting in a prediction with a resolution of 50 days as to the timing of the next relapse. The error rate of this predictor was 2.3 fold lower than the error rate of random predictions (error rate = 0.35, p<0.001). The predictors were further evaluated and found effective not only in untreated patients but were also valid for MS patients which subsequently received immunomodulatory treatments after the initial testing (the error rate of the first level predictor was < 0.18 with p<0.001 for all the patient groups).  Conclusions: We conclude that gene expression analysis is a valuable tool that can be used in clinical practice to predict future MS disease activity.  Similar approach can be also useful for dealing with other autoimmune diseases that characterized by relapsing-remitting nature

Keywords: Disease prediction

overall_design: Data-mining and predictive modeling tools were utilized to analyze a gene-expression dataset of 94 non-treated patients; 62 patients with definite MS and 32 patients with clinically isolated syndrome (CIS). The dataset included the expression levels of 10,594 genes and annotated sequences corresponding to 22,215 gene-transcripts that appear in the microarray. We designed a two stage predictor. The first stage predictor was based on the expression level of 10 genes, and predicted the time to next relapse with a resolution of 500 days. If the predicted relapse was to occur in less than 500 days, a second stage predictor based on an additional different set of 9 genes was used, resulting in a prediction with a resolution of 50 days as to the timing of the next relapse. The predictors were further evaluated and found effective not only in untreated patients but were also valid for MS patients which subsequently received immunomodulatory treatments after the initial testing.

first: Tamir

last: Tuller

first: Udi

last: Rubinstein

first: Rotem

last: Or-Bach

first: Anat

last: Achiron


title: Limitations and possibilities of small RNA digital gene expression profiling: synthetic miRNA replicates (Illumina)

status: Public on Jul 29 2009

city: Utrecht

first: Elzo

last: de Wit

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

type: Non-coding RNA profiling by high throughput sequencing

summary:
Analysis of RNA samples by massive parallel sequencing holds the promise to assay gene expression in both a quantitative and qualitative fashion and therefore allows for digital gene expression (DGE) profiling. We assessed the effect of different experimental approaches by generating small RNA libraries from a biological sample as well as an equimolar pool of synthetic miRNAs and analyzed the results using capillary dideoxy sequencing and next-generation sequencing platforms (Roche/454, AB/SOLiD and Illumina/Solexa). Whereas different sequencing platforms provided highly similar results, large differences in DGE profiles were observed depending on the library preparation method used. Nevertheless, our results indicate that the preferential nature of the library preparation methods is systematic and highly reproducible and we show that DGE is well suited for the quantification of relative expression differences between samples.

Keywords: Transcriptome analysis

overall_design: Examination library method on preparation of equimolar pool of miRNAs, three replicates, sequenced on Illumina GA II

first: Sam

last: Linsen

first: Edwin

last: Cuppen


title: Transcriptomic analysis of the effect of silicon on wheat plants infected or uninfected with powdery mildew

status: Public on Jun 30 2009

city: Jouy en Josas

first: Florian

last: Chain

address: Domaine de Vilvert

department: Institut Micalis

country: France

institute: INRA

summary:
The supply of soluble silicon (Si) to plants has been associated with many benefits that remain poorly explained and often contested. In this work, the effect of Si was studied on wheat plants under both control and pathogen stress (Blumeria graminis f.sp. tritici (Bgt)) conditions by conducting an exhaustive transcriptomic analysis (55,000 genes) aimed at comparing the differential response of plants under four treatments. The response to the supply of Si on control (uninfected) plants was limited to 47 genes providing little evidence of regulation of a specific metabolic process. Plants reacted to inoculation with Bgt by an up-regulation of many genes linked to stress and metabolic processes and a down-regulation of genes linked to photosynthesis. Supplying Si to inoculated plants largely prevented disease development, a phenotypic response that translated into a nearly perfect reversal of genes regulated by the effect of Bgt alone.  These results suggest that Si plays a limited role on a plant’s metabolism in absence of stress, even in the case of a high-Si accumulating monocot such as wheat. On the other hand, the benefits of Si, in the form of biotic stress alleviation, were remarkably aligned with a counter-response to transcriptomic changes induced by the pathogen Bgt.

Keywords: stress type and molecule effect

overall_design:
Wheat culture in hydroponic system :
Hydroponic systems were used to precisely control Si feeding of plants and to reduce external Si contamination. Seeds of wheat cultivar AC Drummond, chosen for its known high susceptibility to Bgt, were sown in 9-cm pots in a nylon bed consisting of nylon stockings cut into small ribbons. Hydroponic systems were set up to immerse roots for 15 minutes every 30 minutes. The plants were grown in a greenhouse (16 h light at 22°C and 8h dark at 18°C, 80% humidity). Plants were immersed in distilled water only during the first week following sowing.

Si amendment and Bgt inoculation:
Wheat plants were grown in the presence (Si+) or absence (Si-) of Si and inoculated (B+) or not (B-) with Bgt for a total of four treatments: Si-B-, Si+B-, Si-B+, and Si+B+. These treatments were applied in the following manner. One week after sowing, distilled water was replaced by a Hoagland solution amended or not with potassium silicate (Kasil 6, PQ Corp etc) at a concentration of 1.7 mM (Si+ Hoagland). Twice a week, Si was added to the Si+ nutrient solution in order to maintain Si concentration at 1.7 mM. Every other week, both Si- and Si+ Hoagland solutions were renewed and the pH adjusted to 5.8 in all systems at each Si addition or nutrient solution renewal.
Four weeks after sowing (i.e. three weeks after Si amendment started), half the plants were inoculated with Bgt as described previously. Briefly, one day prior to inoculation, reservoir wheat plants heavily infected with Bgt were gently shaken to remove old spores and to stimulate the production of fresh ones. The inoculation was performed by shaking these infected wheat plants over the experimental plants .

RNA preparation:
Total RNA was extracted from leaves of three plants per treatment (three biological replicates) with the RNeasy plant kit (Qiagen, Hilden, Germany). Concentration and quality of RNA were assessed on a Nanodrop spectrophotometer (Nanodrop, Wilmington, De, USA) and 2100 Bioanalyzer (Agilent, Palo Alto, Ca, USA). Biotin-labeled cRNAs were synthesized using MessageAmp II-Biotin Enhanced Kit (Ambion, Austin, Tx, USA) following the manufacturer’s instructions using 1 µg total RNA. Labelled cRNA was then fragmented using 5X Array Fragmentation Buffer supplied with the MessageAmp II-Biotin Enhanced Kit.

Microarray hybridization:
The labeled samples were added to an Affymetrix GeneChip® Wheat Genome Array (Affymetrix, Santa Clara, CA, USA), according to the manufacturer’s instructions. The wheat chip contains probe sets representing 55,052 transcripts for all 42 chromosomes in the wheat genome. The chips were hybridized for 16 h at 45 °C in a rotisserie oven at 60 rpm. Following hybridization, the arrays were washed and stained in an Affymetrix Fluidics Station 450, according to the standard protocol from Affymetrix, and scanned using an Affymetrix Scanner 3000. Twelve chips were prepared and corresponded to three biological replications of each of the four treatments (Si-B-, Si+B-, Si-B+, and Si+B+).

first: Caroline

last: Côté-Beaulieu

first: François

last: Belzile

first: James

last: Menzies

first: Richard

last: Bélanger


title: Expression profile analysis of inflammatory response regulated by hepatocyte nuclear factor 4α

status: Public on May 06 2009

city: Boston

first: Peter

last: Burke

state: MA

address: 850 Harrison Avenue, Dowling 2 South

department: Surgery

institute: Boston University School of Medicine

summary: To obtain a genomic view of hepatocyte nuclear factor-4α (HNF-4α) in the regulation of the inflammatory response, microarray analysis was used to probe the expression profile of an inflammatory response induced by cytokines in a model of knock-down HNF-4α HepG2 cells. The results indicate an extensive role for HNF-4α plays in the regulation of a large number of the liver-specific genes. Majority of genes (71%) affected by cytokine treatment are also affected by HNF-4α knock-down. This significant overlap suggests that HNF-4α may play a role in regulating the cytokine-induced inflammatory response.

overall_design: The different treated HepG2 cells were grouped into 4 groups (four replicates in each group): Group1, Control; Group2, HNF-4α shRNA treated cells; Group3, cytokine treated group; Group4, HNF-4α shRNA and cytokine treatments. RNA extraction and hybridization on Affymetrix microarrays were processed. The expression profiles between different groups were analyzed and compared. In order to explore the function of HNF-4α in the inflammatory response, a set of 170 genes annotated as inflammatory response was obtained from GO (geneotology.org), the enrichments of these inflammatory genes were analyzed in different treated groups.

first: Zhongyan

last: Wang

first: Eric

middle: P

last: Bishop


title: Bone marrow gene expression of pediatric acute lymphoblastic leukemia (ALL)

status: Public on Aug 19 2009

city: Beijing

last: Qiao

address: Datun Road

department: System Biology

country: China

institute: Institute of Genetics and Developmental Biology

summary: Pediatric acute lymphoblastic leukemia (ALL) contains cytogenetically distinct subtypes that respond differently to cytotoxic drugs. Subtype classification can be also achieved through gene expression profiling. However, how to apply such classifiers to a single patient and correctly diagnose the disease subtype in an independent patient group has not been addressed. Furthermore, the underlying regulatory mechanisms responsible for the subtype-specific gene expression patterns are still largely unknown. Here, by combining three published microarray datasets (PMIDs: 12086872, 12730115, 17002788) on 535 Caucasian samples and generating a new dataset on 100 Chinese children ALL samples, we were able to 1) identify a 62-gene classifier with 97.6% accuracy from the Caucasian samples and validated it on the completely independent set of 100 Chinese samples, 2) to uncover potential regulatory networks of ALL subtypes. The classifier we identified was so far the only one that could be applied directly to a single sample and sustained validation in a large independent patient group. Our results also suggest that the etiology of ALL is largely the same among different ethnic groups, and that the transcription factor hubs in the predicted regulatory network might play important roles in regulating gene expression and development of ALL.

overall_design:
A total of 100 Chinese pediatric acute lymphoblastic leukemia bone marrow (BM) samples were analyzed, together with five non-ALL BM samples as negative control (the five samples were combined to generate one control, see details in Li et al. Blood, 2009). The diagnosis of ALL was based on morphology, immunology, cytogenetic and molecular (MICM) classification. Cytogenetic ALL subtypes were identified experimentally by G-banding karyotype and multiplex nested RT-PCR. Among the 100 ALL patients, eleven relapsed within five years. All the samples, including those relapsed afterward, were from patients treated on BCH-2003 protocol and were extracted at their initial diagnosis. The five non-ALL BM samples were taken from the removed bones of patients who had plastic surgery for their bone deformity in Beijing Children’s Hospital. And the informed consent was obtained from parents, guardians, or patients (as appropriate). More details are available in Li et al. Gene expression-based classification and regulatory networks of pediatric acute lymphoblastic leukemia, Blood, 2009.
We have generated these ALL and control samples to validate that a high accuracy subtype classifier that we identified could be applied directly to a single sample and sustained validation in a large independent patient group.

first: Zhigang

last: Li

first: Wei

last: Zhang

first: Minyuan

last: Wu

first: Shanshan

last: Zhu

first: Chao

last: Gao

first: Lin

last: Sun

first: Ruidong

first: Huiling

last: Xue

first: Yamei

last: Hu

first: Shilai

last: Bao

first: Huyong

last: Zheng

first: Jing-Dong

last: Han


title: Limitations and possibilities of small RNA digital gene expression profiling

first: Elzo

last: de Wit

summary:
Analysis of RNA samples by massive parallel sequencing holds the promise to assay gene expression in both a quantitative and qualitative fashion and therefore allows for digital gene expression (DGE) profiling. We assessed the effect of different experimental approaches by generating small RNA libraries from a biological sample as well as an equimolar pool of synthetic miRNAs and analyzed the results using capillary dideoxy sequencing and next-generation sequencing platforms (Roche/454, AB/SOLiD and Illumina/Solexa). Whereas different sequencing platforms provided highly similar results, large differences in DGE profiles were observed depending on the library preparation method used. Nevertheless, our results indicate that the preferential nature of the library preparation methods is systematic and highly reproducible and we show that DGE is well suited for the quantification of relative expression differences between samples.

This SuperSeries is composed of the SubSeries listed below.

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

first: Sam

last: Linsen

first: Edwin

last: Cuppen


title: The kinetics of gene expressions of HeLa cells treated with MT7

status: Public on Jun 08 2009

city: Shanghai

first: Zhixiang

address: Rd. Zuchongzhi

institute: SIMM

summary:
Uncontrolled mitosis forms basis for tumor cell proliferation. Interference with normal mitotic progression often leads tumor cells to mitotic arrest. As a consequence, cell death follows, but not at all time. Reasonably, the antimitotic therapy has been proven effective and indispensable in the clinical treatment of cancer.Recently, we generated a 6H-Pyrido[2',1':2,3]imidazo[4,5-c] isoquinolin -5(6H)-one library, several products of which had been shown to possess antimitotic activities in vitro. In an attempt to understand the antitumor mechanism of these novel chemicals, we have chosen one bioactive product of this library, MT7, as a model compound in this study
We used microarrays to detail the kinetics of gene expression in HeLa cells in a time course of MT7 treatment (from 1.5 h to 12 h). The gene expression profiles were further analyzed by GSEA and Connectivity Map, respectively.

overall_design: HeLa cells growing on the 10-cm dishes were treated with 5 mM MT7 for 0(control), 1.5(Time1), 3(Time2), 6(Time3) and 12(Time4) hours, respectively. Cells were then washed three times with PBS and the total RNA was isolated and processed for Affymetrix U133A 2.0 hybridizations.

first: Jian

last: Ding

first: Ze-Hong

last: Miao


title: A synthetic gene-metabolic circuit preferentially increased fatty acid metabolism in human hepatocytes

status: Public on Jun 05 2009

city: Los Angeles

first: Jason

middle: Thaddeus

last: Dean

address: 420 Westwood Plaza

department: Chemical and Biomolecular Engineering

institute: UCLA

summary:
Obesity is becoming increasingly widespread in developed countries, and is often associated with heart diseases and diabetes.  Elevated levels of plasma free fatty acids are a biochemical hallmark of obesity.  Unlike plants and bacteria, mammals cannot utilize fatty acids to generate glucose because of the lack of glyoxylate shunt enzymes. Instead, fatty acids are used for energy storage, and their utilization is regulated at multiple levels ranging from hormonal to metabolic ensuring that glucose is preferentially oxidized before fatty acids.  Here we designed a synthetic gene-metabolic circuit to alter the intracellular signaling and metabolic pathways that control fatty acid metabolism. By introducing the Escherichia coli glyoxylate shunt enzymes, isocitrate lyase (AceA) and malate synthase (AceB), into the mitochondria of human hepatocytes, we demonstrated that fatty acid utilization is preferentially increased while glucose uptake is significantly reduced.  This bacterial pathway diverts signal metabolites (citrate, acetyl-CoA, and malonyl-CoA) that inhibits fatty acid uptake and provides an additional channel for fatty acid utilization. Remarkably, the hepatocytes readily adapted to the synthetic circuit by a series of global transcriptional and post-translational changes in metabolic and signal transduction pathways that further advanced our design objective. This systems approach illustrates the logic of the synergistic interaction between metabolism and signal transduction. The ready acceptance of this non-native pathway by hepatocytes suggests the plasticity of liver metabolism and opens a possibility for the synthetic approach in regulating human metabolism.
Keywords: Fatty acid response, synthetic circuit, carbohydrate metabolism, stably transfected HepG2 cell line, glyoxylate shunt

overall_design:
Wild type (WT) human hepatocyte HepG2 and HepG2 cells stably transfected with pBudaceAB (designated ACE), a vector containing isocitrate lyase and malate synthase from E. coli, were analyzed after 24hour exposure to palmitate.  Cells were seeded at 70% confluency in 10cm plates with DMEM growth media augmented with 300uM palmitate.  After 24hours, total RNA was harvested and hybrized to Affymetrix HG_U133A 2.0 GeneChips.  Data was processed using GCOS 1.2 software.

2 WT biological replicates and 2 ACE biological replicates were analyzed.

middle: T

first: Linh

last: Tran

first: Katrina

middle: M

last: Dipple

first: James

middle: C

last: Liao


title: Functional classes of bronchial mucosa genes that are differentially expressed in asthma

status: Public on Apr 24 2009

city: Chicoutimi

first: Catherine

last: Laprise

state: Qc

address: 555 Boul. Université

country: Canada

institute: Université du Québec à Chicoutimi

summary:
Asthma pathogenesis and susceptibility involves a complex interplay between genetic and environmental factors.
We report the results of expression microarray studies using tissue obtained from bronchial biopsies of healthy controls and of subjects with allergic asthma, both before and following inhaled corticotherapy.

overall_design: Four healthy subjects (2 men and 2 women) without a history of allergy or asthma and four asthmatics (all women) meeting the criteria of the American Thoracic Society for the diagnosis of asthma were recruited to the study. Microarray expression studies were performed using Affymetrix HG-U95Av2 GeneChips.

first: Robert

last: Sladek

first: André

last: Ponton

first: Marie-Claude

last: Bernier

first: Thomas

last: Hudson

first: Michel

last: Laviolette


title: Genome-wide impact of ART-27 loss on androgen-regulated transcription in prostate cancer cells

city: New York

first: Susan

last: Logan

state: NY

address: 550 First Ave., MSB 249

institute: NYU School of Medicine

summary:
The androgen receptor (AR) directs diverse biological processes through interaction with coregulators such as androgen receptor trapped clone-27 (ART-27). The impact of ART-27 on genome-wide transcription was examined. The studies indicate that loss of ART-27 enhances expression of many androgen-regulated genes, suggesting that ART-27 inhibits gene expression. Surprisingly, classes of genes that are upregulated upon ART-27 depletion include regulators of DNA damage checkpoint and cell cycle progression, suggesting that ART-27 functions to keep expression levels of these genes low.

Keywords: LNCaP, cell type comparison, UXT, ART-27, androgen receptor, R1881, AR, androgen-regulated gene expression, prostate cancer

overall_design: Steroid-deprived LNCaP cells were transfected with control or ART-27 siRNA and stimulated with ethanol vehicle or 10 nM R1881 for 18 hrs. 8 samples, 4 conditions, 2 replicates per condition.

first: Jerome

last: Nwachukwu

first: Paolo

last: Mita

first: Rachel

last: Ruoff

last: Ha

first: Qianben

last: Wang

first: S

last: Huang

first: Samir

middle: S

last: Taneja

first: Myles

last: Brown

first: William

last: Gerald

first: Michael

last: Garabedian

middle: K


title: IL1R2 and glucocorticoid responsiveness in patients with AIED

status: Public on Apr 29 2009

city: New Hyde Park

first: Andrea

last: Vambutas

address: 430 Lakeville Road

department: Otolaryngology

institute: Long Island Jewish Medical Center

summary: Glucocorticoid responsive AIED patients demonstrate alternate splicing of IL1R2 in response to dexamethasone prior to treatment.  Identification of IL1R2 as a potential target gene in AIED was determined by a microarray analysis of patients with end-stage AIED and controls with non-immunologic etiologies of hearing loss undergoing cochlear implantation.

overall_design: Patients with either end-stage AIED or controls with non-immunologic etiologies of hearing loss undergoing cochlear implantation were enrolled.  All patients received pneumovax 2 weeks prior to surgery.  At the time of surgery, blood was taken for PBMC isolation and autologous perilymph was harvested from a bloodless field.  For each patient, 3 microarrays were performed: (1) unstimulated PBMC, (2) pneumovax stimulated PBMC, (3) autologous perilymph stimulated PBMC.  All conditions were compared for the AIED and control subjects.  IL1R2 was identified as induced in controls, but not AIED patients, in response to autologous perilymph.  Findings were confirmed by QPCR.

first: James

last: DeVoti

first: Elliot

last: Goldofsky

last: Gordon

first: Marty

last: Lesser

first: Vincent

last: Bonagura


title: Expression analysis upon SIRT1 knockdown in MCF-7 cells

first: W. Lee

last: Kraus

summary: SIRT1 is a nuclear NAD+-dependent protein deacetylase.  Expression microarray analysis was used to study the effect of SIRT1 knockdown on gene expression in MCF-7 breast cancer cells.

overall_design:
SIRT1 stable knockdown was achieved using two retroviral shRNA constructs. An shRNA directed against Luciferase was used to generate the Luc control cells.  Three independent biological replicates with matching Luc controls were analyzed using Affymetrix U133A 2.0 microarrays.

These 6 samples were part of a 15-sample microarray analysis (GSE12971) examining expression regulation by SIRT1, PARP-1, PARG and macroH2A.  All 15 samples were included for the following data normalization steps: data sets obtained using the GCOS software were grouped based on date of experiment and adjusted for batch effect using the parametric empirical Bayes method (Combat R); all values < 0.01 were adjusted to 0.01; the data were log2 transformed, median centered for each array, and median centered for each gene.

first: Tong

last: Zhang

first: Kristine

last: Frizzell

first: Matthew

last: Gamble


title: Gene expression in erythroid cell differentiation

status: Public on Mar 31 2009

city: Raanana

first: Yuval

last: Tabach

address: Hativat Givaty 30

department: Molecular Cell Biology

institute: Weizmann Institute of Science

summary:
Human mononuclear cells were cultured in 2 phases. In the 1st phase the culture medium contained cyclosporine A the 2nd phase contained SCF and erythropoietin. Cells were collected at 3 stages of differentiation; on day 6, 10, 12 and represented early erythroblasts, medium stage and normoblasts.

Keywords: time course

overall_design: RNA was extracted from each sample of cells at a different stage of differentiation and subjected to microarray studies in duplicates.

first: Shoshana

last: Peller


title: MCF7 EGF, HRG stimulation

status: Public on May 26 2009

city: Yokohama

first: Mariko

last: Okada

address: W518, 1-7-22, Suehiro-cho, Tsurumi-ku

country: Japan

institute: RIKEN RCAI

summary:
Sharing common ErbB/HER receptor signaling pathway, heregulin (HRG) induces differentiation of MCF-7 breast cancer cells while epidermal growth factor (EGF) elicits proliferation. Although the cell fate led by those two ligands was totally different, the gene expression profile in early transcription was unexpectedly qualitatively similar, suggesting that the gene expression in late transcription, not early transcription, may reflect a respect of ligand specificity. In this study, based on the data from time-course microarray of all human genes, we predicted and determined a series of transcription factors which may control HRG-specific timed-late transcription and cellular differentiation of MCF-7 cells. Validation analyses showed that one of activator protein 1 (AP-1) families appeared just after c-Fos expression, another AP-1 family partner, induced expression of another  transcription factor through activation of AP-1 complex. Furthermore, expression of this transcription factors caused suppression of extracellular signal-regulated kinase (ERK) phosphorylation which is sustainedly regulated by HRG-initiated ErbB signaling. Overall, our analysis indicated an importance of formation of timed-transcriptional regulatory network and its function to control upstream signaling pathway through negative feedback for cellular differentiation.

Keywords: growth hormone response, time course

overall_design: MCF7 human breast cancer cells were stimulated by the growth hormone (epidermal growth factor (EGF) or heregulin (HRG)). Control was set as non-treated cells.


title: Genetic analysis of radiation-induced changes in human gene expression

status: Public on Apr 06 2009

city: Philadelphia

middle: Patrick

last: Morley

state: PA

address: 3400 Civic Center Blvd, Bldg 421

department: Penn Cardiovascular Institute

institute: Perelman School of Medicine at the University of Pennsylvania

summary: We used microarrays to measure the expression levels of genes in irradiated immortalized B cells, lymphoblastoid cells, from members of Centre d'Etude du Polymorphisme Humain (CEPH) Utah pedigrees.  Data were collected for cells at baseline and 2 hour and 6 hour after exposure to 10 Gy of ionizing radiation (IR).

overall_design: We used microarrays to measure the expression levels of genes in irradiated immortalized B cells, lymphoblastoid cells, from members of  15 Centre d'Etude du Polymorphisme Humain (CEPH) Utah pedigrees (CEPH 1333, 1341, 1346, 1362, 1408, 1416, 1420, 1421, 1423, 1424, 1444, 1447, 1451, 1454, 1582).  Expression data was obtained for cell lines derived from 2 parents and 8 children per each family. Cells were irradiated at 10 Gy in a 137Cs irradiator.  Cells were harvested prior to radiation and at 2 and 6 hours following exposure to IR.

first: Denis

last: Smirnov

first: Eunice

last: Shin

first: Richard

last: Spielman

first: Cheung

last: Vivian


title: Human fibroblast stimulation with PDGF-BB or b-FGF

status: Public on Mar 16 2009

city: Brussels

first: Jean-Baptiste

last: Demoulin

address: av Hippocrate 75, B1.74.05

department: de Duve Institute

country: Belgium

institute: Université catholique de Louvain

summary:
We analyzed gene expression in human fibroblasts stimulated by platelet-derived growth factor-BB (PDGF-BB) or basic fibroblast growth factor (bFGF) for 1h and 24h. The results of two independent experiments were merged. SAM analysis identified 116 relevant probe sets. Hierarchical clustering of these probe sets showed divergent early gene regulation by PDGF and FGF but overlapping late response. We first analyzed genes commonly regulated by PDGF-BB and b-FGF more than 2 fold after 24h of stimulation and we found that these two growth factors repressed FOXO.
We then focused on the early gene expression response induced by both growth factors. We performed a fold change analysis and found 114 probe sets regulated by PDGF-BB and 42 probe sets regulated by b-FGF, 37 of which were shared between the two gene lists .
Keywords: Time course, cell Treatment comparison

overall_design:
Human foreskin fibroblasts (AG01518) were cultured (1.5E6 cells/10 cm dish) in MEM medium with 10% fetal calf serum and glutamine for 24h, then washed and incubated for 47h in serum-free medium. Cells were stimulated for 1h by PDGF-BB (25ng/ml) or b-FGF (10ng/ml, + heparine 50 microg/ml). Alternatively, cells were incubated for 24h in starvation medium and treated for 24h with growth factors, or left untreated for 48h in serum-free medium (control treatment).
Two biological replicates were performed

first: Ahmed

last: Essaghir


title: Effect of Ovarian Hormone on Gene Expression in Rhesus Dorsal Raphe Nucleus with Human U95A Affymetrix Chip

status: Public on May 21 2009

city: Beaverton

first: Arubala

middle: P.

last: Reddy

state: OR

address: 505 NW 185th Ave

department: Oregon National Primate Research Center

institute: Oregon Health and Sciences University

summary: Small blocks of the midbrain containing the dorsal raphe nucleus was obtained from ovariectomized monkeys treated with placebo, estrogen, progesterone or estrogen plus progesterone for one month. The RNA was extracted and hybridized to the human U95A Affymetrix chip.

overall_design: Each chip represents one monkey. N=3 monkeys per treatment.

first: Cynthia

last: Bethea


title: Targetting CD24 for treatment of colorectal and pancreatic cancer by monoclonal antibodies or siRNA

status: Public on Mar 07 2009

city: Tel Aviv

first: Eyal

last: Sagiv

address: 6th Weizmann st

department: The Integrated Cancer Prevention Center

institute: Tel Aviv Medical Center

summary: CD24 is a potential oncogene reported to be overexpressed in a large variety of human malignancies. We have shown that CD24 is overexpressed in 90% of colorectal tumors at a fairly early stage in the multistep process of carcinogenesis. Anti-CD24 monoclonal antibodies (mAb) induce a significant growth inhibition in colorectal and pancreatic cancer cell lines that express the protein. This study is designed to investigate further the effects of CD24 down-regulation using mAb or small interfering RNA in vitro and in vivo. Western blot analysis showed that anti-CD24 mAb induced CD24 protein down-regulation through lysosomal degradation. mAb augmented growth inhibition in combination with five classic chemotherapies. Xenograft models in vivo showed that tumor growth was significantly reduced in mAb-treated mice. Similarly, stable growth inhibition of cancer cell lines was achieved by down-regulation of CD24 expression using short hairpin RNA (shRNA). The produced clones proliferated more slowly, reached lower saturation densities, and showed impaired motility. Most importantly, down-regulation of CD24 retarded tumorigenicity of human cancer cell lines in nude mice. Microarray analysis revealed a similar pattern of gene expression alterations when cells were subjected to anti-CD24 mAb or shRNA. Genes in the Ras pathway, mitogenactivated protein kinase, or BCL-2 family and others of oncogenic association were frequently down-regulated. As a putative new oncogene that is overexpressed in gastrointestinal malignancies early in the carcinogenesis process, CD24 is a potential target for early intervention in the prevention and treatment of cancer.

overall_design: The study compared gene expression profiles between human CRC cells HT29 before and after expression of 1 and 2 shRNA vectors directed at the human CD24 gene, GFP control gene, HT29 cells and Colo357, human pancreatic cancer cells, before and after the inhibition of the CD24 molecule using 72h treatment with anti-CD24 monoclonal antibodies.

first: E

first: A

last: Starr

first: U

last: Rozovski

first: R

last: Khosravi

first: P

last: Altevogt

first: T

last: Wang

first: N

last: Arber


title: Depletion of Erk1 and Erk2 MAP kinases in primary human keratinocytes

status: Public on May 03 2009

city: San Francisco

first: Phillip

middle: Anthony

last: Dumesic

address: 513 Parnassus Ave, HSW1119

department: Diabetes Center

institute: University of California, San Francisco

summary: We assessed the effect of RNAi-mediated MAP kinase cascade signaling blockade in primary human keratinocytes.  Two sets of siRNA targeting different regions of the Erk1/2 genes were used, enabling identification of off-target siRNA effects.

overall_design: Primary human keratinocytes were electroporated with scrambled control siRNA, a pair of siRNA oligomers against Erk1 and Erk2 (set A), or an independent pair of siRNA oligomers against Erk1 and Erk2 (set B).  Four days after electroporation, RNA was harvested.  Three biological replicates were performed for each of the three siRNA groups.

first: Florence

last: Scholl

first: Deborah

middle: I

last: Barragan

first: Paul

last: Khavari


title: Expression data of HL-60 leukemia cells after treatment with Curcumin and TCDD

status: Public on May 19 2009

city: Düsseldorf

first: Charlotte

last: Esser

state: NRW

address: Aufm Hennekamp 50

department: Immunologie

country: Germany

institute: IUF

summary:
Curcumin has antileukemic potential that is mediated by dfifferent pathways. Because we could not confirm that the Arylhydrocarbonreceptor, a ligand activated transcription factor, is involved in curcumins antileukemic effects, we performed microarray experiments to have an overall screening of the pathways affected by curcumin in HL-60 cells.
We identified an involvement of the vitamin D receptor in the effects of curcumin on HL-60 cells by comparing our results with other experiments.

overall_design: HL-60 cells were treated for 24h with 100 µM curcumin, 10 nM 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), or 0.5 % DMSO as control.

first: Andreas

last: Goergens

first: Markus

last: Frericks


title: Wheat "Chinese Spring" natural antisense transcription survey

status: Public on Jun 11 2009

city: Ames

first: PLEXdb Curator

address: 1023  Crop Genomes Informatics Lab (Iowa State University)

institute: Iowa State University

summary: The Affymetrix GeneChip Wheat Genome Array currently provides the most comprehensive coverage of the wheat genome for a microarray. In addition to using this resource for transcript expression studies and hybridization-based DNA marker discovery, we endeavored to use the GeneChip to discover the expression of natural antisense transcript (NAT) pairs. By using alternative target preparation schemes, both the sense- and antisense-strand derived transcripts were labeled and hybridized to the Wheat GeneChip. To enable maximum discovery, five different tissue types were selected for assay, and the wheat cultivar ‘Chinese Spring’ was used considering that most of the GeneChip probe sequences were based on sequencing of this genome. [PLEXdb(http://www.plexdb.org) has submitted this series at GEO on behalf of the original contributor,  Tristan Coram.  The equivalent experiment is TA21 at PLEXdb.]

overall_design: target strand labeled: Sense(1-replications);  target strand labeled: Antisense(1-replications)

first: Tristan

last: Coram


title: Identification of genes effected by GM-CSF treatment in mature human neutrophils

city: Bern

first: Shida

last: Yousefi

state: Bern

address: Inselspital, INO-F

department: Pharmacology

country: Switzerland

institute: University of Bern

summary:
The objective of this study was to compare the transcriptional repertoire of mature human neutrophils before and after GM-CSF treatment by using oligonucleotide microarrays.
Leukotriene B4 (LTB4) is an important pro-inflammatory lipid mediator generated by neutrophils upon activation. Granulocyte/macrophage colony-stimulating  factor (GM-CSF) stimulation is known to enhance agonist-mediated LTB4 production of neutrophils within minutes, a process called “priming”. Here, we  demonstrate that GM-CSF also limits the production of LTB4 by neutrophils via a transcriptional mechanism at later time points. We identified hematopoietic  specific Ras homologous (RhoH)/translocation three four (TTF), which was induced following GM-CSF stimulation in neutrophils, as a key regulator in this  process. Neutrophils derived from RhoH/TTF-deficient (Rhoh-/-) mice demonstrated increased LTB4 production upon activation compared with  normal mouse neutrophils. Moreover, neutrophils from cystic fibrosis patients expressed enhanced levels of RhoH/TTF and generated less LTB4 upon  activation compared with normal human neutrophils. Taken together, these data suggest that RhoH/TTF represents an inducible feedback inhibitor in  neutrophils that is involved in the limitation of innate immune responses.

overall_design: 3 healthy donors' blood were used for each group. Two groups were created. One group ("untreated"=N ) the RNA were extracted immediately after isolation of blood neutrophil. The second group ("Neu") blood neutrophils were incubated with GM-CSF (50 ng/ml) for 7 hours before RNA extraction.

first: Hans-Uwe

last: Simon


title: Time course of MEK inhibition

status: Public on Feb 11 2009

first: Christine

middle: Anne

last: Pratilas

address: 1275 York Avenue

department: Molecular Pharmacology

institute: Memorial Sloan-Kettering Cancer Center

summary:
This study used microarray expression analysis to identify global changes in transcript alteration in response to MEK inhibition. Genes under ERK control were identified in a representative V600E BRAF cell line as a function of time following exposure to a small molecule inhibitor of MEK.
Keywords: Time course

overall_design: SkMel-28 cells growing in culture were treated with the MEK inhibitor PD0325901 for 2, 8, or 24 hours.  Changes in RNA compared to reference (time =0) were measured using microarray analysis.

first: Barry

last: Taylor


title: Effect of wtp53-reactivating drug RITA on HCT116 cells

status: Public on Feb 28 2009

city: Stockholm

first: Martin

last: Enge

address: CCK, Z4

institute: Karolinska Institute

summary:
Since its discovery as a tumour suppressor some fifteen years ago, the transcription factor p53 has attracted paramount attention for its role as “the guardian of the genome”. TP53 mutations occur so frequently in cancer, regardless of patient age or tumour type, that they appear to be part of the life history of at least 50% of human tumours. In most tumours that retain wild-type p53, its function is inactivated due to deregulated HDM2, a protein which binds to p53 and which can inhibit the transcriptional activity of p53 and induce its degradation.
RITA is a low-molecular-weight compound which addresses the second group of tumours retaining functionally reactive wt p53. It was found in a screening of the National Cancer Institute (NCI) library of low-molecular-weight compounds based on its ability to selectively kill wtp53-containing cells. RITA binds directly to p53 and diplaces its main destructor Mdm2, as well as inducing a shift in the conformation of p53. This is in contrast to the wtp53-reactivating compound Nutlin3a, which targets Mdm2, inhibiting its ability to degrade p53. Using microarray technology we have explored the effect of RITA on the transcriptome of isogenic cell-lines with knocked-out (KO) or intact (WT) TP53. While the effects on KO cells are below detection limit, the effects on WT cells are profound. The known p53 targets induced are predominately apoptotic, in contrast to the genes affected by Nutlin3a, which are exclusively growth-arrest genes.

Keywords: Antitumor agent

overall_design: HCT116 parental and HCT116 p53-null (HCT116 TP53-/-) cells were subjected to treatment with 1uM RITA for 12h, or left untreated. The experiment was done in three independent biological replicates.


title: Paired MEK inhibited and control

summary:
This study used microarray expression analysis to identify global changes in transcript alteration in response to MEK inhibition. Genes under ERK control were identified in a panel of V600E BRAF and RTK-activated tumor cells and xenografts, using short-term inhibition of ERK activity using the MEK inhibitor PD0325901 (Pfizer).
Keywords: paired treatment and control

overall_design: Cell lines growing in culture (n=12) and murine xenografts (n=2) were treated with the MEK inhibitor PD0325901 or vehicle alone as control. Paired analysis of MEK inhibited to control samples was performed for two groups of tumor cells, V600E BRAF and RTK.


title: Expression profiling of V600E BRAF and RTK-activated cells upon MEK inhibition

summary:
This study used microarray expression analysis to identify global changes in transcript alteration in response to MEK inhibition. Genes under ERK control were identified in a panel of V600E BRAF and RTK-activated tumor cells and xenografts, using short-term inhibition of ERK activity using the MEK inhibitor PD0325901 (Pfizer).

This SuperSeries is composed of the SubSeries listed below.

overall_design: Cell lines growing in culture (n=12) and murine xenografts (n=2) were treated with the MEK inhibitor PD0325901 or vehicle alone as control. Paired analysis of MEK inhibited to control samples was performed for two groups of tumor cells, V600E BRAF and RTK.  Time course analysis was performed on one representative cell line in order to first determine the optimal time point to detect changes in all cell lines.


title: Spontaneous regression of chronic lymphocytic leukemia: clinical and biological features of 9 cases

status: Public on Apr 22 2009

city: Rome

first: Sabina

last: Chiaretti

address: Via Benevento, 6

department: Cellular Technologies and Hematology

institute: Division of Hematology

summary: We describe 9 CLL patients who underwent a spontaneous clinical regression. CD38 and ZAP-70 were negative in all cases. Immunoglobulin heavy chain variable region (IgVH) genes, mutated in all 7 evaluable patients, were restricted to the VH3 family in 6, with the usage of VH3-30 gene in 2. The light chain variable region genes were mutated in 6/8 cases, with the usage of Vκ4-1 gene in 3. Microarray analysis of CLL cells revealed a distinctive genomic profile. The number of activated T lymphocytes expressing IFN-γ, TNF-α and IL-4 was similar between CLL in spontaneous regression and healthy individuals.

overall_design: We compared gene expression profiles obtained from 4 cases of CLL who underwent spontaneous regression with that of 3 B-cells from healthy donors and 18 CLL cases with either stable or progressive disease.

first: Ilaria

last: Del Giudice

first: Simona

last: Tavolaro

first: Maria

last: De Propris

first: Roberta

last: Maggio

first: Francesca

last: Mancini

first: Nadia

last: Peragine

last: Santangelo

first: Marilisa

last: Marinelli

middle: R

last: Mauro

first: Anna

last: Guarini

first: R

last: Foà


title: Microarray analysis of Huh7 cells treated with IFNa2, OSM or IFNa2 combined with OSM

status: Public on Jan 21 2009

city: Pamplona

first: Iranzu

last: Gonzalez

state: Navarra

address: Avda. Pio XII nº 55

department: Hepathology and Gene Therapy

country: Spain

institute: Center for applied medical research

summary: OSM increases the antiviral effect of IFNα in Huh7 cells infected with hepatitis A virus (HAV) or HCV replicon and synergizes with IFNα in the induction of antiviral genes

overall_design: 16 samples. Four replicates for each experimental condition. 72 hours of treatment in DMEM supplemented with 2% FBS.

first: Esther

last: Larrea

first: Rafael

last: Aldabe

last: González

first: Victor

last: Segura

first: Pablo

last: Sarobe

first: Jesus

last: Prieto


title: Comparing ERG expression and vector control in 293HEK cells

status: Public on Feb 03 2009

first: Brett

last: Carver

address: 1275 York Ave

summary: The goal of this study was to identify potential genes regulated by ERG

overall_design: 293HEK cells were transiently transfected in duplicate with CMV-Tg2B ERG or control vector independently for a total of 4 samples.

first: Pier

last: Pandolfi


title: Gene expression profiles obtained from laser-microdissected human choroid plexus papilloma cells

status: Public on Feb 01 2009

city: Münster

first: Martin

last: Hasselblatt

address: Domagkstraße 19

department: Institute of Neuropathology

institute: University Hospital Münster

summary: Gene expression profiles generated from human tumor cells laser-microdissected from surgical samples of seven choroid plexus papillomas (Grade I WHO) as eight samples of epithelial cells lasermicrodissected from normal choroid plexus obtained at autopsy.

overall_design:
Choroid plexus tumors are rare pediatric brain tumors derrived from the choroid plexus epithelium.
Gene expression profiles of lasermicrodissected tumor cells from 7 individual choroid plexus tumor samples obtained at surgery were compared to gene expression profiles from non-neoplastic choroid plexus epithelial cells lasermicrodissected from normal non-neoplastic choroid plexus obtained at autopsy (Am J Surg Pathol. 2006 Jan;30(1):66-74.) in order to identfy genes differentially expressed in choroid plexus tumor cells.

first: Heike

last: Stegemann


title: HOXA9 is required for survival in human MLL rearranged acute leukemias

status: Public on Jan 01 2009

city: Boston

first: Joerg

last: Faber

address: 1 Blackfan Circle

institute: Children's Hospital Boston

summary: Leukemias that harbor translocations involving the mixed lineage leukemia gene (MLL) possess unique biological characteristics and often have an unfavorable prognosis. Gene expression analyses demonstrate a distinct profile for MLL-rearranged leukemias with consistent high-level expression of select Homeobox genes including HOXA9. Here, we investigated the effects of HOXA9 suppression in MLL-rearranged and MLL-germline leukemias utilizing RNAi. Gene expression profiling after HOXA9 suppression demonstrated co-downregulation of a program highly expressed in human MLL-AML (this study) and murine MLL-leukemia (Krivtsov et al. 2006) stem cells including HOXA10, MEIS1, PBX3 and MEF2C. Our data indicates an important role for HOXA9 in human MLL-rearranged leukemias, and suggests targeting HOXA9 or downstream programs may be a novel therapeutic option.

overall_design: RNA was purified from t(9;11) MOLM-14 AML cells 44h after transduction in triplicates with 2 of the two most effective HOXA9shRNA constructs (3 x 1F3-HOXA9shRNA; 3 x 2A5-HOXA9shRNA) or GFP-controlshRNA (6x).

first: Andrei

middle: V

last: Krivtsov

first: Matthew

last: Stubbs

first: Renee

last: Wright

first: Tina

middle: N

last: Davis

first: Marry

last: van den Heuvel-Eibrink

first: Christian

last: Zwaan

first: Andrew

last: Kung

first: Scott

last: Armstrong


title: Comparison of stable human Treg and Th clones by transcriptional profiling - experiment I

status: Public on Jan 09 2009

first: Sophie

last: Lucas

address: avenue Hippocrate 74

institute: de Duve Institute

summary:
The aim of this work was to identify functional features that are specific of human Treg cells, through the identification of genes that are differentially expressed: 1/ in activated Treg clones versus activated Thelper clones; 2/ in Th clones activated in the presence versus the absence of TGFb; 3/ in suppressed Th clones, i.e. Th clones activated in the presence of Treg clones, versus controls.

Keywords: TCR activation

overall_design: Two to four million cells of human T cell clones were collected at rest (time point = 0) or after activation with coated anti-CD3 and soluble anti-CD28 antibodies (time points = 6, 24 or 41hrs). RNA was extracted for  hybridization on Affymetrix microarrays.

first: Julie

last: Stockis


title: Gene expression profiling of ATL patients

city: Bethesda

first: John

last: Brady

state: MD

institute: NCI

summary:
Abstract:  Adult T-cell leukemia/lymphoma (ATL) is an aggressive and fatal disease.  We have examined 18 ATL patient samples using Affymetrix HG-U133A2.0 arrays.  Using the BRB array program, we identified genes differentially expressed in leukemia cells compared to normal lymphocytes.  Several unique genes were identified that were overexpressed in leukemia cells including TNFSF11, RGS13, MAFb, CSPG2, C/EBPalpha and TCF4.  200 of the most highly overexpressed ATL genes were analyzed by the PathwayStudio 4.0 program.  ATL leukemia cells were characterized by an increase in genes linked to "central" genes CDC2/cyclin B1, SYK/LYN, PCNA and BIRC5.  Because of its potential therapeutic importance, we focused our studies on the regulation and function of BIRC5, whose expression was increased in 13 of 14 leukemia samples.  TCF4 reporter assays and transfection of DN-TCF4 demonstrated that TCF4 regulates BIRC5 gene expression.  Functionally, transfection of ATL cells wi BIRC5 shRNA decreased BIRC5 exprression and cell viability 80%.  Clinical treatment of ATL patients with Zenapax or bortezomib decreased BIRC5 expression and cell viability.  These experiments represent the first direct experimental evidence that BIRC5 plays an important role in ATL cell viability and provides important insight into ATL genesis and potential targeted therapies.

Keywords: disease state analysis

overall_design: Gene expression profiles of 7 control and 18 ATL patient samples were analyzed using Affymetrix HG-U133A2.0 arrays.

first: Cynthia

middle: A

last: Pise-Masison

first: Michael

last: Radonovich

first: Kathleen

last: Dohoney

last: Morris

first: Deirdre

last: O'Mahony

first: Min-Jung

last: Lee

first: Jane

last: Trepel

first: Thomas

last: Waldmann

middle: E

last: Janik


title: Effect of HDAC inhibitors on expression of androgen induced genes

city: New York

first: Yu

last: Chen

address: 1275 York Ave, Box 20

department: Human Oncology and Pathogenesis Program

institute: Memorial Sloan Kettering Cancer Center

summary:
Elevated levels of androgen receptor (AR) in prostate cancer confer resistance to current antiandrogens and play a causal role in disease progression due to persistent target gene activation.  Through pharmacologic and genetic approaches, we show that half of all direct AR target genes, including TMPRSS2, the primary driver of ETS fusion transcripts in 70 percent of human prostate cancers, require histone deacetylase (HDAC) activity for transcriptional activation by AR.  Surprisingly, the HDAC3-NCoR complex, which typically functions to repress gene expression by nuclear receptors, is required for AR target gene activation.  Prostate cancer cells treated with HDAC inhibitors have reduced AR protein levels, but we show that the mechanism of blockade of AR activity is through failure to assemble a coactivator/RNA polymerase II complex after AR binds to the enhancers of target genes.  Failed complex assembly is associated with a phase shift in the cyclical wave of AR recruitment that typically occurs in response to ligand treatment.  HDAC inhibitors retain the ability to block AR activity in hormone refractory prostate cancer models and therefore merit clinical investigation in this setting.  HDAC-regulated AR target genes defined here can serve as biomarkers to ensure sufficient levels of HDAC inhibition.
Keywords: Androgen receptor, histone deacetylase, prostate cancer, dose response

overall_design: LNCaP prostate cancer cells were grown in charcoal stripped, androgen deplete medium. They were then stimulated with or without 1nM R1881 and with or without two doses of HDAC inhibitors TSA, SAHA, and LBH 589. Cyclohexamide was include to block new protein synthesis. Cells were harvested 16 hours after treatment.

first: Derek

last: Welsbie

first: Jin

last: Xu

first: Laetitia

last: Borsul

first: Howard

last: Scher

first: Neal

last: Rosen

first: Charles

middle: L

last: Sawyers


title: Effect of individual HDAC knockdown on expression of androgen induced genes

summary:
Elevated levels of androgen receptor (AR) in prostate cancer confer resistance to current antiandrogens and play a causal role in disease progression due to persistent target gene activation.  Through pharmacologic and genetic approaches, we show that half of all direct AR target genes, including TMPRSS2, the primary driver of ETS fusion transcripts in 70 percent of human prostate cancers, require histone deacetylase (HDAC) activity for transcriptional activation by AR.  Surprisingly, the HDAC3-NCoR complex, which typically functions to repress gene expression by nuclear receptors, is required for AR target gene activation.  Prostate cancer cells treated with HDAC inhibitors have reduced AR protein levels, but we show that the mechanism of blockade of AR activity is through failure to assemble a coactivator/RNA polymerase II complex after AR binds to the enhancers of target genes.  Failed complex assembly is associated with a phase shift in the cyclical wave of AR recruitment that typically occurs in response to ligand treatment.  HDAC inhibitors retain the ability to block AR activity in hormone refractory prostate cancer models and therefore merit clinical investigation in this setting.  HDAC-regulated AR target genes defined here can serve as biomarkers to ensure sufficient levels of HDAC inhibition.

Keywords: Androgen receptor, histone deacetylase, prostate cancer, dose response

overall_design: Lentiviral shRNA mediated knockdown of Luciferase (ctrl), HDAC1, HDAC2, and HDAC8 LnCAP cells were generated. They were  grown in charcoal stripped, androgen deplete medium. They were then stimulated with or without 1nM R1881. Cyclohexamide was include to block new protein synthesis. Cells were harvested 16 hours after treatment.


title: Gene expression analysis of the wheat response to infection by Fusarium pseudograminearum

city: St Lucia

first: Brendan

last: Kidd

state: QLD

address: 306 Carmody Road

department: Plant Industry

country: Australia

institute: CSIRO

summary: Crown rot of wheat, caused by Fusarium pseudograminearum and other Fusarium species is an important disease globally. To understand the host response to challenge by Fp, we examined gene exression changes in the stem base of the wheat variety Kennedy, following inoculation with macroconidia using the Affymetrix GeneChip Wheat Genome Array. Induced genes included mainly those with defensive functions such as genes encoding anti-microbial proteins as well as oxidative stress-related proteins, signalling molecules, and proteins involved in both primary and secondary metabolism. This study is the first comprehensive analysis of the wheat transcriptome during crown rot infection and provides new insights into the host processes involved in plant defence against this pathogen.

overall_design: There are six samples, three F. pseudograminearum inoculated samples and three mock inoculated samples. Each sample consists of 2cm of stem base from approximately 20 plants.

first: Olivia

middle: J

last: Desmond

last: Manners

first: Peer

last: Schenk

first: Donald

last: Maclean

first: Kemal

last: Kazan


title: Effect of actein on the growth of the MDA-MB-453 breast cancer cell lines as a function of time and concentration.

status: Public on Apr 15 2009

first: Linda

middle: Saxe

last: Einbond

address: 701 West 168Th St.

department: Rehabilitation Medicine

institute: Columbia University

summary:
Previous studies indicate that the triterpene glycoside actein from the herb black cohosh inhibits growth of human breast cancer cells. This study seeks to identify genes altered in human breast cancer cells by treatment with actein, using gene expression analysis. We treated MDA-MB-453 human breast cancer cells with actein at 2 doses, 20 or 40 μg/mL, for 6 or 24 h. We identified 5 genes that were activated after each of the treatments that are known to play a role in cellular responses to diverse stresses, including the DNA damage and unfolded protein responses. In addition, four genes that mediate the integrated stress response (ISR), including activating transcription factor 4, were induced under at least one of the 4 treatment conditions. We used hierarchical clustering to define clusters comprising patterns of gene expression. Two ISR genes, activating transcription factor 3 (ATF3) and DNA damage- inducible transcript 3, and lipid biosynthetic genes were activated after exposure to actein at 40 μg/mL for 6 h, whereas the cell cycle genes cyclin E2 and cell division cycle 25A were repressed. Our results suggest that actein induces 2 phases of the ISR, the survival phase and the apoptotic phase, depending on the dose and duration of treatment. We confirmed the results of gene expression analysis with real-time RT-PCR for 18 selected genes and Western blot analysis for ATF3. Since actein activated transcription factors that enhance apoptosis, and repressed cell cycle genes, it may be useful in the prevention and therapy of breast cancer.
Keywords: expression,time course, dose response

overall_design: This study used Affymetrix U133A 2.0 gene chips. We analyzed 3 replicate cultures after treating MDA-MB-453 cells with actein at 20 or 40 mg/mL for 24 h and 2 replicate cultures after treating MDA-MB-453 cells with actein at 20 or 40 mg/mL for 6 h. All samples were normalized to remove chip-dependent regularities using the GCRMA method of Irizarry et al . Chips and controls at each combination of actein concentration and time were normalized together.  The statistical significance of differential expression was calculated using the empirical Bayesian LIMMA (LInear Model for MicroArrays) method of Smyth et al. A cut-off B>0 was used for the statistical significance of gene expression.

first: Tao

last: Su

first: Hsan-au

last: Wu

first: Richard

last: Friedman

first: Xiaomei

last: Wang

first: Alejandro

last: Ramirez

first: Fredi

last: Kronenberg

first: Bernard

last: Weinstein


title: Molecular profiling of classical Hodgkin’s lymphoma tissues

status: Public on Dec 17 2008

city: Marseille

first: Pascal

last: FINETTI

state: BdR

address: 232 Bd Ste Marguerite

department: Centre de cancérologie de Marseille

institute: Institut Paoli-Calmettes

summary:
Previous reports suggest that outcome of cHL patients may be related to the tumor microenvironment, which in turn may be influenced by EBV infection. Gene profiling was used for further characterize the cHL microenvironment. A training set of 73 cHL tissue samples was profiled using Affymetrix DNA microarrays. Supervised analysis provided a gene signature separating EBV+ from EBV- cHL tissues, including genes characteristic of Th1 and antiviral response. Samples from patients with favourable outcome significantly overexpressed genes involved in the function of B-cells and plasmacytoid dendritic cells (pDCs), like BCL11A. A validation set of 146 cHL samples was analyzed using immunohistochemistry (IHC).

Keywords: Disease state analysis

overall_design: A training set of 73 pre-treatment cHL tissue samples was profiled using Affymetrix DNA microarrays. They were collected at the time of diagnosis from 73 different cHL patients (including 10 children under the age of 15), who underwent lymph node surgical biopsy in several French haematological centres (Marseilles, Lille, Dijon, Nancy, Paris) belonging to the network of the Groupe d’Etude des Lymphomas de l’Adulte (GELA). Each patient (or parents for children) gave written informed consent. Samples were snap-frozen in liquid nitrogen within 30 minutes of removal. All cases were de novo reviewed by 2 hematopathologists (BC and LX) before analysis.

first: Bruno

last: Chetaille

first: Luc

last: Xerri


title: Analysis of ectopic human endometrium and peritoneal tissues in nude mice

status: Public on Dec 22 2008

city: Cambridge

first: D. Stephen

last: Charnock-Jones

address: The Rosie Hospital

department: Obstetrics and Gynaecology

country: United Kingdom

institute: University of Cambridge

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: HIV-1 Activates Macrophages Independent of Toll-like Receptors

status: Public on Dec 15 2008

city: Gainesville

first: Maureen

middle: M.

last: Goodenow

state: FL

address: 1376 Mowry Road

department: Pathology, Immunology, & Laboratory Medicine

institute: University of Florida

summary:
Macrophages provide an interface between innate and adaptive immunity and are important long-lived reservoirs for Human Immunodeficiency Virus Type-1 (HIV-1). Multiple genetic networks involved in regulating signal transduction cascades and immune responses in macrophages are coordinately modulated by HIV-1 infection.   To evaluate complex interrelated processes and to assemble an integrated view of activated signaling networks, a systems biology strategy was applied to genomic and proteomic responses by primary human macrophages over the course of HIV-1 infection. Macrophage responses, including cell cycle, calcium, apoptosis, mitogen-activated protein kinases (MAPK), and cytokines/chemokines, to HIV-1 were temporally regulated, in the absence of cell proliferation. In contrast, Toll-like receptor (TLR) pathways remained unaltered by HIV-1, although TLRs 3, 4, 7, and 8 were expressed and responded to ligand stimulation in macrophages. HIV-1 failed to activate phosphorylation of IRAK-1 or IRF-3, modulate intracellular protein levels of Mx1, an interferon-stimulated gene, or stimulate secretion of TNF, IL-1b, or IL-6. Activation of pathways other than TLR was inadequate to stimulate, via cross-talk mechanisms through molecular hubs, the production of proinflammatory cytokines typical of a TLR response. HIV-1 sensitized macrophage responses to TLR ligands, and the magnitude of viral priming was related to virus replication.  HIV-1 induced a primed, proinflammatory state, M1HIV, which increased the responsiveness of macrophages to TLR ligands. HIV-1 might passively evade pattern recognition, actively inhibit or suppress recognition and signaling, or require dynamic interactions between macrophages and other cells, such as lymphocytes or endothelial cells. HIV-1 evasion of TLR recognition and simultaneous priming of macrophages may represent a strategy for viral survival, contribute to immune pathogenesis, and provide important targets for therapeutic approaches.
Affymetrix arrays were used to identify genomic macrophage response to HIV during viral spread in culture.

Keywords: time course

overall_design: An HIV-1 spreading infection was established in primary human macrophages. RNA was extracted from both viral- and mock-infected macrophages cultures over 7 days and hybridized to Affymetrix HG-U95Av2 GeneChips for analysis.

first: Joseph

last: Brown

middle: M


title: Profiling gene expression in HeLa cells  by hsa-mir-98 overexpression

city: Naples

first: Vincenzo

middle: Alessandro

last: Gennarino

state: Italy

address: Via Pietro Castellino,111

department: TIGEM

institute: Telethon

summary:
Total RNA samples from three biological replicates in which the hsa-mir-98 was overexpressed in HeLa cells  were profiled by gene expression. As negative control, we used total RNA samples from HeLa cells transfected with cel-mir-67

Keywords: gene expression array-based (RNA / in situ oligonucleotide)

overall_design: hsa-miR-98 transfection

last: Gennarino.

first: Sandro

last: Banfi.


title: hsa-mir-26b and hsa-mir-98 overexpression


title: Profiling gene expression in HeLa cells  by hsa-mir-26b overexpression

summary:
Total RNA samples from three biological replicates in which the hsa-mir-26b was overexpressed in HeLa cells  were profiled by gene expression. As negative control, we used total RNA samples from HeLa cells transfected with cel-mir-67

Keywords: gene expression array-based (RNA / in situ oligonucleotide)

overall_design: hsa-miR-26b transfection


title: Identification of transcriptional changes induced by FHIT in lung cancer cells NCI-H460

status: Public on Dec 25 2008

city: Milan

first: Luca

last: Roz

address: Via Venezian 1

department: Experimental Oncology and Molecular Medicine

institute: Fondazione IRCCS Istituto Nazionale dei Tumori

summary: The FHIT gene, at 3p14.2, encompasses the common fragile site FRA3B and is frequently inactivated in primary tumors and cell lines of lung, head and neck, stomach, esophagus, cervix and breast cancer. In particular, loss of the FHIT protein is one of the most frequent alteration in lung tumors and pre-invasive lung lesions, suggesting a role for this gene in the early stages of lung carcinogenesis. Adenoviral-mediated restoration of FHIT expression in FHIT-negative cell lines results in cell cycle alteration, induction of apoptosis (through activation of the cytoplasmic apoptotic pathway) and regression of the tumorigenic phenotype. Taken together these observations support the hypothesis of a role for FHIT in human carcinogenesis, but little is known about its mechanism of action. Identification of transcriptional targets of FHIT is therefore critical to understand the pathways by which FHIT promotes growth arrest and apoptosis.

overall_design:
RNA was extracted from NCI-H460 lung cancer cells (FHIT-negative) transduced with AdFHIT or AdLACZ (control), 48 hours post infection at a multiplicity of infection of 10. Three independent infections were harvested and RNA preparations were pooled, transcribed into cDNA, labeled, and hybridized following the manufacturer's protocol to the oligonucleotide microarray Human Gene Chip HG-U95Av2 (Affymetrix). Two independent hybridizations were performed (rep1 and rep2) and scan images aquired before (sc1) and after (sc2) anti-streptavidin antibody enhancement step.
The whole procedure was repeated twice (starting from thawing of new aliquot of NCI-H460 cells) resulting in two independent experiments (exp1 and exp2).

first: Francesca

last: Andriani

first: Gabriella

last: Sozzi

first: Simone

middle: P

last: Minardi

first: Cristina

last: Ferrario


title: Similar gene expression profiles of sporadic, PGL2-, and SDHD-linked paragangliomas

status: Public on Dec 10 2008

city: Leiden

first: Jan

last: Oosting

address: Albinusdreef 2

department: Pathology

institute: Leiden University Medical Center

summary:
The similarity in gene-expression profiles suggest that PGL2, like SDHD, is involved in the functionality of the SDH complex, and that tumor formation in these three subgroups involves the same pathways as in SDH linked paragangliomas. We were not able to clarify the identity of PGL2 on 11q13. The lack of differential gene-expression of chromosome 11 genes might indicate that chromosome 11 loss, as demonstrated in SDHD-linked paragangliomas, is an important feature in the formation of a paraganglioma regardless of the genetic background.

Keywords: disease state analysis

overall_design: We compared the gene expression profiles of sporadic (n=7), SDHD- (n=6) and PGL2-linked (n=4) head and neck paraganglioma.

first: Erik

middle: F

last: Hensen


title: Expression data from IGF-I-stimulated MCF-7 cells

status: Public on Nov 20 2008

city: Houston

first: Chad

last: Creighton

address: One Baylor Plaza, Mail Stop: BCM305

department: Biostatistics, Ducan Cancer Center

institute: Baylor College of Medicine

summary:
Substantial evidence implicates IGF-I signaling in the development and progression of breast cancer.  To identify transcriptional targets of IGF action in breast cancer cells, we performed gene expression profiling (>22,000 RNA transcripts) of IGF-I-stimulated MCF-7 cells, a well characterized breast cancer cell line that is highly responsive to IGFs. We defined an IGF-I gene signature pattern of hundreds of genes either up-regulated or down-regulated at both 3 and 24 hrs in vitro.  After removing genes considered generic to cell proliferation, the signature was examined in four different public profile datasets of clinical breast tumors (representing close to 1000 patients), as well as in profile datasets of experimental models for various oncogenic signaling pathways. Genes with early and sustained regulation by IGF-I were highly enriched for transcriptional targets of the estrogen, Ras, and PI3K/Akt/mTOR pathways.  The IGF-I signature appeared activated in most estrogen receptor-negative (ER-) clinical breast tumors and in a substantial subset (~25%) of ER+ breast tumors.  Patients with tumors showing activation of the IGF-I signature tended to have a shorter time to disease recurrence (including patients not receiving adjuvant therapy), both when considering all patients and the subset of ER+ patients. We found evidence for cross-talk and common transcriptional endpoints between the IGF-I and estrogen systems.  Our results support the idea that the IGF-I pathway is one mechanism by which breast tumors may acquire hormone independence and a more aggressive phenotype.
Keywords: two group comparison

overall_design: MCF-7L cells were routinely maintained in DMEM + 5% FBS.  Cells were plated in 10cm dishes and then next day incubated in serum-free medium.  After an overnight incubation, cells were stimulated in triplicate with or without IGF-I (8nM or 50ng/ml) for 3 or 24hrs.  Cells were then lysed and total RNA isolated using Qiagen midi-prep kit according to the manufacturer’s instructions.  RNA extraction and hybridization was then carried out.

first: Adrian

last: Lee


title: The hematopoietic stem cell in chronic phase CML is characterized by a transcriptional profile resembling normal myeloid progenitor cells and reflecting loss of quiescence

status: Public on Dec 01 2008

city: Düsseldorf

first: Ingmar

last: Bruns

address: Moorenstr. 5

department: Hematology, Oncology and Clinical Immunology

institute: University of Düsseldorf

summary: We found that composition of cell subsets within the CD34+ cell population is markedly altered in chronic phase (CP) chronic myeloid leukemia (CML). Specifically, proportions and absolute cell counts of common myeloid progenitors (CMP) and megakaryocyte-erythrocyte progenitors (MEP) are significantly greater in comparison to normal bone marrow whereas absolute numbers of hematopoietic stem cells (HSC) are equal. To understand the basis for this, we performed gene expression profiling (Affymetrix HU-133A 2.0) of the distinct CD34+ cell subsets from six patients with CP CML and five healthy donors. Euclidean distance analysis revealed a remarkable transcriptional similarity between the CML patients' HSC and normal progenitors, especially CMP. CP CML HSC were transcriptionally more similar to their progeny than normal HSC to theirs, suggesting a more mature phenotype. Hence, the greatest differences between CP CML patients and normal donors were apparent in HSC including downregulation of genes encoding adhesion molecules, transcription factors, regulators of stem-cell fate and inhibitors of cell proliferation in CP CML. Impaired adhesive and migratory capacities were functionally corroborated by fibronectin detachment analysis and transwell assays, respectively. Based on our findings we propose a loss of quiescence of the CML HSC on detachment from the niche leading to expansion of myeloid progenitors.


title: Targeting PKC: A Novel Role for beta-catenin in ER stress and Apoptotic Signaling

status: Public on Nov 11 2008

first: Marc

last: Raab

address: 44 Binney Street

institute: Dana Farber Cancer Institute

summary:
Targeting protein kinase C (PKC) isoforms by the small molecule inhibitor enzastaurin has shown promising pre-clinical activity in a wide range of tumor cells. In this study, we further delineated its mechanism of action in multiple myeloma (MM) cells and found a novel role of b-catenin in regulating growth and survival of tumor cells. Specifically, inhibition of PKC leads to rapid accumulation of b-catenin by preventing the phosphorylation required for its proteasomal degradation. Microarray analysis and siRNA-mediated gene silencing in MM cells revealed that accumulated b-catenin activates early ER stress signaling via eIF2a, CHOP and p21, leading to immediate growth inhibition. Furthermore, accumulated b-catenin contributes to enzastaurin-induced cell death. Both sequential knock-down of b-catenin, c-Jun, and p73, as well as overexpression of  b-catenin or p73 confirmed that accumulated b-catenin triggers c-Jun-dependent induction of p73,  thereby conferring MM cell apoptosis. In summary, our data reveal a novel role of b-catenin in ER stress-mediated growth inhibition, and a new pro-apoptotic mechanism triggered by b-catenin upon inhibition of PKC isoforms. Moreover, we identify p73 as a potential novel therapeutic target in MM. Based on these and previous data, enzastaurin is currently under clinical investigation in a variety of hematologic malignancies including MM.

Keywords: time course

overall_design: MM.1S myeloma cell lines were treated with enzastaurin for 3, 6 or 12 hours. Controls are represented by untreated cells, at the same time points.

first: Kenneth

last: Anderson

first: Klaus

last: Podar


title: ChIP-on-chip significance analysis reveals large-scale binding and regulation by human transcription factor oncogenes

status: Public on Dec 04 2008

first: Adolfo

last: Ferrando

address: 1130 St Nicholas Ave Irving Cancer Research Center 5-505A

department: Institute for Cancer Genetics

summary: ChIP-on-chip has emerged as a powerful tool to dissect the complex network of regulatory interactions between transcription factors and their targets. However, most ChIP-on-chip analysis methods use conservative approaches aimed to minimize false-positive transcription factor targets. We present a model with improved sensitivity in detecting binding events from ChIP-on-chip data. Its application to human T-cells, followed by extensive biochemical validation, reveals that three transcription factor oncogenes, NOTCH1, MYC, and HES1, bind to several thousands target gene promoters, up to an order of magnitude increase over conventional analysis methods. Gene expression profiling upon NOTCH1 inhibition shows broad-scale functional regulation across the entire range of predicted target genes, establishing a closer link between occupancy and regulation. Finally, the increased sensitivity reveals a combinatorial regulatory program in which MYC co-binds to virtually all NOTCH1-bound promoters. Overall, these results suggest an unappreciated complexity of transcriptional regulatory networks and highlight the fundamental importance of genome-scale analysis to represent transcriptional programs.

overall_design: T-ALL cell lines harboring activating mutations in NOTCH1 were treated with vehicle only (DMSO) or a highly active gamma-secretase inhibitor (COMPE, 100nM) for 72 hs and processed for gene expression profiling analysis. Gene expression signatures associated with inhibition of NOTCH1 signaling with CompE were correlated with findings on NOTCH1 direct target genes identified by ChIP-on-chip analysis.

first: Adam

last: Margolin

first: Teresa

last: Palomero

first: Pavel

last: Sumazin

first: Andrea

last: Califano

first: Gustavo

last: Stolovitzky


title: Expression data from PBMC treated with rabbit anti-thymocyte globulin (rATG) or horse ATG (hATG)

status: Public on Nov 10 2008

first: Xingmin

last: Feng

address: 9000 Rockville Pike

institute: National Institutes of Health

summary:
We performed microarray to compare gene expression patterns of PBMC treated with rATG or hATG. Fold changes were compared using 2-way ANOVA tests for untreated, rATG- and hATG-treated PBMC. In PBMC treated with 10 ug/mL rATG, compared with untreated PBMC, 478 genes showed up-regulation, and 341 genes showed down-regulation at 24 hours using 10% FDR and 2-fold change cutoff. Immediately striking was that 10 ug/mL hATG had affected many fewer genes than did rATG: only 3 genes were up-regulated and 6 genes were down-regulated at 24 hours in hATG-treated PBMC.  When we compared rATG with hATG, rATG induced up-regulation of 268 genes and down-regulation of 95 genes. These genes belong to the categories of immune response (64 genes), cytokine-cytokine receptor interaction (36 genes), regulation of cell proliferation (24 genes), cell cycle (23 genes), cell growth (8 genes), apoptosis (7 genes), and others.
Keywords: different treatment

overall_design: PBMC from healthy controls were cultured with rATG or hATG (each at 10 ug/mL) in 10% FCS/RPMI 1640 at 37C for 24 hours, PBMC without drugs were used as controls.  Total RNA was extracted with the RNeasy kit (Qiagen, Valencia, CA).  RNA purity was assessed by spectrophotometry.  Probes were prepared using standard Affymetrix protocols and hybridized to Affymetrix HG-U133A 2.0 arrays (Affymetrix, Santa Clara, CA)

first: Xiuli

last: Xu

first: Nalini

last: Raghavachari

first: Peter

last: Munson

first: Neal

middle: S

last: Young


title: Oral tongue cancer gene expression profiling: Identification of novel potential prognosticators

status: Public on Nov 14 2008

first: Nicholas

middle: D

last: Socci

address: 1275 York Ave

department: Computational & Systems Biology Program

summary:
Background: The present study is aimed at identifying potential candidate genes as prognostic markers in human oral tongue squamous cell carcinoma (SCC) by large scale gene expression profiling.   Methods: The gene expression profile of patients (n=37) with oral tongue SCC were analyzed using Affymetrix HG_U95Av2 high-density oligonucleotide arrays. Hierarchical clustering analyses failed to show significant segregation of patients. In patients (n=20) with available tumor and matched normal mucosa, 77 genes were found to be differentially expressed (P< 0.05) in the tongue tumor samples compared to their matched normal controls.  Among the 45 over-expressed genes, MMP-1 encoding interstitial collagenase showed the highest level of increase (average: 34.18 folds). The 20 patients were then grouped into stage (early vs. late) and nodal disease (node positive vs. node negative) subgroups and genes differentially expressed in tumor vs. normal and between the subgroups were identified. Three genes, GLUT3, HSAL2, and PACE4, were selected for their potential biological significance in a larger cohort of 49 patients by quantitative real-time RT-PCR.    Results: Using the criterion of two-fold or greater as overexpression, 30.6%, 24.5% and 26.5% of patients showed high levels of GLUT3, HSAL2 and PACE4, respectively.  Univariate analyses demonstrated that GLUT3 over-expression correlated with depth of invasion (P<0.0001), tumor size (P=0.024), pathological stage (P=0.009) and recurrence (P=0.038).  HSAL2 was positively associated with depth of invasion (P=0.015) and advanced T stage (P=0.0467).   In survival studies, only GLUT3 showed a prognostic value with disease-specific (P=0.049), relapse-free (P-0.0042) and overall survival (P=0.003).  PACE4 mRNA expression failed to show correlation with any of the relevant parameters.   Conclusions: The characterization of genes identified to be significant predictors of prognosis by oligonucleotide microarray and further validation by real-time RT-PCR offers a powerful strategy for identification of novel targets for prognostication and treatment of oral tongue carcinoma.

Keywords: gene expression profiling

first: Bhuvanesh

last: Singh

first: Cherry

last: Estillo


title: Effect of wt versus mutant hsa-miR-122 overexpression on spontaneous hESC differentiation

status: Public on Nov 05 2008

city: Jerusalem

first: Galit

last: Tzur

address: Kiryat Hadassah

department: Gene Therapy

institute: Hadassah University Hospital

summary: We aimed to determine whether overexpression of endoderm-specific miRNA may affect hESC differentiation. To this end, we analyzed the effect of lentiviral-based overexpression of liver-specific miR-122 on hESC differentiation, using genomewide gene microarrays. Stable overexpression of endoderm-specific miR-122 in hESC resulted in increased expression of a few endodermal markers in spontaneously-differentiating hESC, but had no clear effect on directing differentiation towards an endodermal fate; rather, it delayed the general differentiation of hESC.

overall_design: Lentiviral vectors encoding either wt or mutant (control) hsa-miR-122 were transduced into hES cells (line HES2). The cells differentiated spontaneously in KO medium without feeder cells for 2 weeks, and the gene expression profile of wt- vs mutant-expressing miR-122 was compared. We used for this study 3 arrays for control (mutant miR) and 2 arrays for the treatment (wt miRNA).

first: Asaf

last: Levy

first: Eti

last: Meiri

first: Omer

last: Barad

first: Yael

last: Spector

first: Zvi

last: Bentwich

first: Lina

last: Mizrahi

first: Mark

last: Katzenellenbogen

first: Etti

last: Ben-Shushan

first: Benjamin

last: Reubinoff

first: Eithan

last: Galun