title: NOTCH1 directly regulates c-MYC and activates a feed-forward-loop transcriptional network promoting leukemic cell growth

status: Public on Nov 13 2006

first: Adolfo

last: Ferrando

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

department: Institute for Cancer Genetics

country: USA

institute: Columbia University

summary:
The NOTCH1 signaling pathway directly links extracellular signals with transcriptional responses in the cell nucleus and plays a critical role during T-cell development and in the pathogenesis over 50% of human T-cell lymphoblastic leukemia (T-ALL) cases. However, little is known about the transcriptional programs activated by NOTCH1. Using an integrative systems biology approach we show that NOTCH1 controls a feed-forward loop transcriptional network that promotes cell growth. Inhibition of NOTCH1 signaling in T-ALL cells led to a reduction in cell size and elicited a gene expression signature dominated by downregulated biosynthetic pathway genes. By integrating gene expression array and ChIP-on-chip data, we show that NOTCH1 directly activates multiple biosynthetic routes and induces c-MYC gene expression. Reverse engineering of regulatory networks from expression profiles showed that NOTCH1 and c-MYC govern two directly interconnected transcriptional programs containing common target genes that together regulate the growth of primary T-ALL cells. These results identify c-MYC as an essential mediator of NOTCH1 signaling and integrate NOTCH1 activation with oncogenic signaling pathways upstream of c-MYC.
Keywords: Drug treatment

overall_design: Duplicated cultures of T-ALL cell lines were treated with Compound E, a gamma-secretase inhibitor or vehicle only (DMSO) for 24 h and analyzed using oligonucleotide microarrays. Gene expression changes were analyzed in the context of loss of NOTCH1 signaling induced by the gamma secretase inhibitor treatment.

first: Teresa

last: Palomero

first: WeiKeat

last: Lim

first: Duncan

last: Odom

first: MariaLuisa

last: Sulis

first: Pedro

middle: J

last: Real

first: Adam

last: Margolin

first: Kelly

middle: C

last: Barnes

first: Jennifer

last: O'Neil

first: Donna

last: Neuberg

first: Andrew

last: Weng

first: Jon

last: Aster

first: Francois

last: Sigaux

first: Jean

last: Soulier

first: Thomas

last: Look

first: Richard

last: Young

first: Andrea

last: Califano


title: ENCODE: two phosphorylation states of RNAP II ChIP-chip from HeLa cells

status: Public on Oct 11 2006

city: PROVIDENCE

first: Alexander

last: Brodsky

state: RI

address: 593 Eddy St.

department: MCB

institute: Rhode Island Hospital/Brown University

type: Genome binding/occupancy profiling by genome tiling array

summary:
Two phosphorylation states of RNAPII in HeLa cells
Keywords: ChIP-chip

overall_design: Study of RNAPII binding patterns to ENCODE regions.  3 arrays using 8wg16 antibody, hypophosphoryated PolII, 3 arrays using 4H8 antibody, phosphorylated PolII CTD, and 6 input DNA arrays.

first: Clifford

last: Meyer

first: Ian

last: Swinburne

first: Giles

last: Hall

first: Benjamin

last: Keenan

first: Xiaole

last: Liu

first: Edward

last: Fox

first: Pamela

last: Silver


title: Effects of RNA interference-mediated c-MYC depletion on gene expression profiles in human cancer cell lines

status: Public on Sep 28 2006

city: VILLEJUIF

first: David

last: CAPPELLEN

address: 39, rue Camille Desmoulins

department: UMR 8126 CNRS

country: France

institute: Institut Gustave Roussy

summary:
Affymetrix Hu133 GeneCHIP Microarray data for Control and c-MYC knock-down (KD) human cancer cell lines.

Data related to article 'Novel c-MYC target genes mediate differential effects on cell proliferation and migration', from D. CAPPELLEN et al., EMBO Reports

Note: Samples GSM136093 and GSM136094 (Hela cell line, expt 1) were hybridized and normalized separately from the other 18 Samples.
Keywords: siRNA analysis

overall_design: The cells were harvested 3 days after LacZ (Control)  and c-MYC (c-MYC KD) small interfering RNA transfection, and total RNA was used for microarray-based global gene expression profiling.

last: Cappellen


title: Expression data from ARPE-19 cells treated with LDL or ox-LDL

status: Public on Sep 01 2006

city: Baltimore

first: Yanqin

last: Yang

state: MD

address: 600 N. Wolfe Street

institute: The Johns Hopkins University

summary:
LDL or Ox-LDL 200ug/ml, which showed no loss of viability after a 48 hour exposure, induced a physiological and pathological transcriptional response, respectively. LDL induced a downregulation of genes associated with cholesterol biosynthesis while ox-LDL induced transcriptional alterations in genes related to inflammation, matrix expansion, lipid metabolism and processing, and apoptosis. Pentraxin-3 was secreted into the culture medium after RPE cells were stimulated with ox-LDL, and immunohistochemically evident in Bruch’s membrane of human macular samples with age-related macular degeneration. ARPE-19 cells exposed to 200?g/ml ox-LDL had a 38% apoptosis rate compared to less than 1% when exposed to LDL or untreated controls (p<0.0001).
While LDL induced a physiologic response by RPE cells, a pathological phenotypic response was seen after treatment with oxidatively modified LDL. The transcriptional, biochemical, and functional data provide initial support of a role for the hypothesis that modified LDLs are one trigger for initiating events that contribute to the development of age-related macular degeneration.
Keywords: treatment with non-treatment control

overall_design: Human ARPE-19 cells were exposed to LDL or oxidatively modified LDL (ox-LDL) for 48 hours for RNA extraction and hybridization on Affymetrix microarrays. We sought to determine whether retina, pigment epithelial cells develop a pathologic phenotype after exposure to low density lipoproteins (LDL) that are oxidatively modified.We have made two comparsions: LDL treatment versus non-treatment; ox-LDL treatment versus non-treatment.

first: Yuko

last: Yamada

first: Jane

last: Tian

first: Roy

last: Cutler

first: Tinghua

last: Wu

first: James

middle: T

last: Handa


title: Analysis of transcripts regulated by Dicer and Argonaute proteins in human HEK-293 cells

city: Basel

first: Daniela

last: Schmitter

address: Maulbeerstrasse 66

department: Epigenetics

country: Switzerland

institute: Friedrich Miescher Institut

summary:
HEK-293-derived cell lines depleted of Dicer or one of four Argonaute proteins were generated. Microarray analysis of these knock down cell lines and various control cell lines was performed to address how large the proportion of genes is that are regulated by RNA silencing in an individual cell type. Comparison of different arrays revealed commonly (up) regulated genes. The highest overlap was observed in Ago2 and Dicer knockdown cells.
Keywords: different timepoints of induction of stable knock down cell lines compared to various control cell lines

overall_design: three control cell lines (T-Rex, shRNA, empty vector), two induvidual Argonaute 1 and 4 (Ago1kd, Ago4kd) knock down cell lines, one Argonaute 2 and 3 (Ago2kd, Ago3kd) and two induvidual Dicer knock down cell lines were analyzed as biological replicates

first: Jody

last: Filkowski

first: Kaifu

last: Tang

first: Ed

last: Oakley

first: Herbert

last: Angeliker

first: Alain

last: Sewer

first: Mihaela

last: Zavolan

first: Petr

last: Svoboda

first: Witold

last: Filipowicz


title: Decidual stromal cell response to paracrine signals from the trophoblast

status: Public on Sep 18 2006

city: San Francisco

first: Said

last: Talbi

state: CA

address: 500 Parnassus

institute: UCSF

summary:
During the invasive phase of implantation, trophoblasts and maternal decidual stromal cells secrete products that regulate trophoblast differentiation and migration into the maternal endometrium.  Paracrine interactions between the extravillous trophoblast and the maternal decidua are important for successful embryonic implantation, including establishing the placental vasculature, anchoring the placenta to the uterine wall, and promoting immuno-acceptance of the fetal allograph.  Global cross-talk between the trophoblast and the decidua has not been elucidated to date, and the current study used a functional genomics approach to investigate these paracrine interactions.
Our data demonstrate a significant induction of pro-inflammatory cytokines and chemokines, as well as angiogenic/static factors in decidualized endometrial stromal cells in response to trophoblast-secreted products.  The data suggest that the trophoblast acts to alter the local immune environment of the decidua to facilitate the process of implantation and assure an enriched cytokine/chemokine environment, while limiting mitotic activity of the stromal cells during the invasive phase of implantation.
Keywords: Gene expression arrays in human stromal cells

overall_design: Human endometrial stromal cells were decidualized with progesterone and were further treated with conditioned media (CM) from human trophoblasts (TCM) or, as a control, with conditioned media (CCM) from non-decidualized stromal cells for 0, 3 and 12 hr. Total RNA was isolated and processed for analysis on whole genome, high density oligonucleotide arrays, containing 54,600 genes.

first: Alex

last: Hess

first: Amy

last: Hamilton

first: Christy

last: Dosiou

first: Mette

last: Nyegaard

first: Nihar

last: Nayak

first: Olga

last: Genbecev

first: P

last: Mavrogianis

first: K

last: Ferrer

first: Jan

last: Kruessel

first: Asgi

last: Fazleabas

first: Susan

last: Fisher

first: Linda

last: Giudice


title: Genomic Pathways of 17-beta-Estradiol Induced Malignant Cell Transformation in Human Breast Epithelial Cells

status: Public on Aug 10 2006

city: Charlottesville

first: Yong

last: Huang

state: VA

address: 1340 Jefferson Park Ave

department: Medicine

institute: University of Viginia

summary:
The estrogen-dependence of breast cancer has long been recognized, however, the role of 17β-estradiol (E2) in cancer initiation was not known until we demonstrated that it induces complete neoplastic transformation of the human breast epithelial cells MCF-10F. E2-treatment of MCF-10F cells progressively induced high colony efficiency and loss of ductulogenesis in early transformed (trMCF) cells and invasiveness in Matrigel invasion chambers. The cells that
crossed the chamber membrane were collected and identified as bsMCF, and their subclones designated bcMCF, and the cells harvested from carcinoma formation in SCID mice designated caMCF. These phenotypes correlated with gene dysregulation during the progression of the
transformation. The highest number of dysregulated genes was observed in caMCF, being slightly lower in bcMCF, and lowest in trMCF. This order was consistent with the extent of chromosome aberrations (caMCF > bcMCF >>> trMCF). Chromosomal amplifications were found in 1p36.12-pter, 5q21.1-qter and 13q21.31-qter. Losses of the complete chromosome 4 and 8p11.21-23.1 were found only in tumorigenic cells. In tumor-derived cell lines, additional losses were found in 3p12.1-14.1, 9p22.1-pter and 18q11.21-qter. Functional profiling of dysregulated genes revealed progressive changes in the integrin signaling pathway, inhibition of apoptosis, acquisition of tumorigenic cell surface markers and epithelial-mesenchymal transition. In tumorigenic cells, the levels of E-cadherin, EMA, and various keratins were low and CD44E/CD24 were negative, whereas SNAI2, vimentin, S100A4, FN1, HRAS, TGFβ1 and
CD44H were high. The phenotypic and genomic changes triggered by estrogen exposure that lead normal cells to tumorigenesis confirm the role of this steroid hormone in cancer initiation.
Keywords: Cell type comparison

overall_design: Human MCF-10F cells were transformed by 70nM 17-beta-estradiol. The transformed cells were selected by Boyden chamber. The selected cells were injected into SCID mice and also cloned and expanded. The thus grown tumors were harvested for cell culture. Genomic DNA and total RNA were isolated from cell lines in each step for 100K SNP and gene expression microarray assay.

first: Sandra

middle: V

last: Fernadez

first: Shirlean

last: Goodwin

first: Patric

middle: A

last: Russo

first: Irma

middle: H

first: Thomas

middle: R

last: Sutter

first: Jose


title: Using GINI2 to identify novel mutations in candidate tumor suppressor genes in colon cancer cells

status: Public on Aug 11 2006

city: Buffalo

first: Ken

last: Lo

state: NY

address: Elm and Carlton Streets

department: Cancer Genetics

institute: Roswell Park Cancer Institute

summary:
Inhibition of the nonsense mediated decay (NMD) mechanism in cells results in stabilization of transcripts carrying premature translation termination codons.  A strategy referred to as gene indentification by NMD inhibition (GINI) has been proposed to identify genes carrying nonsense mutations (Noensie & Dietz, 2001).  Genes containing frameshift mutations in colon cancer cell line have been identifying mutatnt genes using GINI, we have now further improved the strategy.  In this approach, inhibition of NMD with emetine is complemented with inhibiting NMD by blocking the phosphorylation of the hUpf1 protein with caffeine.  In addition, to enhance the GINI strategy, comparing mRNA level alterations produced by inhibiting transcription alone or inhbiiting transcription together with NMD following caffeine pretreatment were used for the efficient identification of false positives produced as a result of stress response to NMD inhibition.  To demonstrate the improved efficiency of this approach, we analyzed colon cancer cell lines showing microstellite instability.  Bi-allelic inactivating mutations were found in the FXR1, SEC1L1, NCOR1, BAT3, PHD14, ZNF294, C190ORF5 genes as well as genes coding for proteins with yet unknown functions.
Keywords: microarray, gene expression, Affymetrix, drug treatment, emetine, caffeine, actinomycin D, novel tumor suppressor gene candidates

overall_design: Using GINI2 as a method of identifying novel biallelic truncating mutations in MSI+ colon cancer cells, various treatment conditions were applied to cultured colon cancer cell lines LS180 (MSI+) and SW480 (MSI-) and gene expression profiling was measured using Affymetrix GeneChip U133 Plus 2 arrays.

first: Igor

last: Ivanov

first: Lesleyann

last: Hawthorn

first: John

middle: K

last: Cowell

first: Yurij

last: Ionov


status: Public on Sep 10 2006

city: São Paulo

first: Renato

middle: David

last: Puga

state: São Paulo

address: Dr Ovidio Pires de Campos,785

department: Instituto de Psiquiatria

country: Brazil

institute: Universidade de São Paulo

summary:
Interactions between epithelial cells and their associated stroma, mainly the fibroblasts therein have been implicated in breast malignancy. Our aim was to verify the effect of soluble factors resulting from the co-culture of normal (NAF) or associated breast cancer fibroblasts (CAF) with a normal (MCF-10A) or a metastatic (MDA-MB231) breast epithelial cell line on fibroblast gene expression profiles. Fibroblast primary cultures were established and co-culture of these cell types separated by inserts, which allow the passage of soluble factors, was performed. Total RNA was extracted, amplified and subjected to microarray gene expression profiling using microarrays in which 4,608 ORESTES (open reading frame expressed sequence tags) were spotted. Differentially expressed genes, at a False Discovery Ratio (FDR) less then 0.01 and fold variation greater than 2, were selected. CAF molecular signature was characterized by down regulation of 62% (29/47) of genes as compared to NAF, represented mainly by those related to vesicular transport, cytoskeleton plasticity and cell motility. CAF responded to co-culture with MCF-10A cells by up regulation of 63.5% (89/140) of genes, including those involved with cytoskeleton remodelling, lipid synthesis and members of the PI3K pathway. Contrarywise, MDA-MB231 cells affected the gene expression profile of CAF by down regulation of 65% (102/156) of genes, including those associated to tumor suppression and antiangiogenic potential. Up regulated genes, were mainly implicated with cell cycle progression and proliferation. NAF signature was altered by MDA-MB231 presence resulting in down regulation of suppressor genes, integrins and angiogenic factors, but it was modestly affected by MCF-10A.
Keywords: Fibroblasts transcriptome altered by breast cell lines

overall_design: 4 samples of CAF and 3 samples of NAF were analyzed. HB4A cells were used as reference, all samples were performed with dye-swaps.


title: Genetic background of the polygenic neural tube defect

status: Public on Nov 02 2006

city: Budapest

first: Balazs

last: Györffy

address: Bókay 53

country: Hungary

institute: Hungarian Academy of Sciences

summary:
In polygenic disorders we do not know exactly, how many genes are involved in the pathomechanism, but the analysis of fetal gene expression can get us closer to the solution. In our study we were searching for the genetic background of the polygenic neural tube defect, which is the second most common birth defect in the world (1 in 1000 live births). Our data revealed novel candidate genes, like SLAP, LST1 and BENE, which can play an important role in the pathogenesis of neural tube defects. We created a data warehouse from the results, suitable for further analysis. This study also demonstrates that a routinely collected amount of amniotic fluid (as small as 6 mL) is enough to successfully hybridize isolated RNA to expression arrays, making the ability to use the technique from normally collected amniotic fluid samples.
Keywords: Prenatal gene expression signature

overall_design: We report an in vivo study in living human fetuses using oligonucleotide microarray analysis of fetal mRNA isolated from amniocytes, which were collected by amniocentesis. RNA was successfully isolated, amplified, labeled, and hybridized to whole-genome Affymetrix transcript arrays.


title: Gene expression profile of VIN lesions in comparison to controls

status: Public on Nov 01 2006

city: Rotterdam

first: Lindy

last: Santegoets

department: Obstetrics & Gynecology

country: Netherlands

institute: Erasmus University Medical Center Rotterdam

summary:
In order to understand the molecular mechanism behind Vulvar Intraepithelial Neoplasia (VIN), we have analyzed the gene expression profile of VIN lesions in comparison to controls.
Keywords: disease state analysis

overall_design: Nine VIN and 10 control samples were collected. RNA of each sample was isolated and analyzed using Affymetrix Human U133plus2 GeneChips.

first: Leen

last: Blok


title: Leukocytes Gene Expression in Correlation to Plasma Lipid Levels

status: Public on Aug 04 2006

city: Richmond Hill

first: Jun

last: Ma

state: Ontario

address: 2 East Beaver Creek Road, Building 2

country: Canada

institute: GeneNews Limited

summary:
Background---For decades, plasma lipid levels have been known risk factors of atherosclerosis. Recently, inflammation has gained acceptance as a crucial event in the pathogenesis and development of atherosclerosis. A number of studies have provided some insights into the relationships between the two aspects of atherosclerosis: plasma lipids --- the risk factors, and circulating leukocytes --- the effectors of inflammation.  In this study, we investigate the relationships between plasma lipids and leukocytes.
Methods and Results---No significant correlation was found between leukocyte counts and plasma lipid levels in 74 individuals. Profiling and analyzing the leukocyte gene expression of 32 individuals revealed distinctive patterns in response to plasma lipid levels: 1) genes involved in lipid metabolism and in the electron transport chain were positively correlated with triglycerides and low-density lipoprotein cholesterol levels, and negatively correlated with high-density lipoprotein cholesterol levels; 2) genes involved in platelet activation were negatively correlated with high-density lipoprotein cholesterol levels; 3) transcription factors regulating lipidgenesis-related genes were correlated with plasma lipid levels; 4) a number of genes correlated to plasma lipid levels were found located in the regions of known QTLs associated with hyperlipemia.
Conclusions--- We discovered interesting patterns of leukocyte gene expression in response to plasma lipid levels. Most importantly, genes involved in lipid metabolism, the electron transportation chain, and platelet activation were found correlated with plasma lipid levels. We suggest that leukocytes respond to changing plasma lipid levels by regulating a network of genes, including genes involved in lipid and fatty acid metabolism, through the activation of key transcription factors, such as sterol regulatory element binding transcription factors and peroxisome proliferative activated receptors.
Keywords: Atherosclerosis, leukocyte, lipid, gene expression

overall_design:
1. Profile gene expression in human peripheral blood cells.
2. Test blood biochemistry and blood cell differential counts
3. Examine the correlation between blood gene expression and blood lipid levels.
4. Explore possible pathways with significant genes.
5. Validate a number of significant genes with RT-PCR

first: Adam

last: Dempsey

first: Dimitri

last: Stamatiou

first: Wayne

last: Marshal

first: Choong-Chin

last: Liew


title: miR-124 transfection time course

status: Public on Nov 03 2006

city: Chicago

first: Xiaowei

last: Wang

state: IL

address: 909 S Wolcott Ave, Rm 5137

institute: University of Illinois at Chicago

summary:
This time course analysis is to study the targets and functions of miR-124. MiR-124 was overexpressed and the effect of miR-124 overexpression was compared to negative controls to identify downregulated genes and potential miRNA targets.
Keywords: time course

overall_design: one array for each time point of miR-124 and negative control transfections

first: X


title: Gene expression of distinct histological components of Wilms tumor analyzed under the perspective of kidney development

status: Public on Aug 01 2006

summary:
Gene expression analyses through cDNA microarray of unique Wilms tumor (WT) histological components, blastema (BL), epithelia (EP) and stroma (ST), from different patients were performed and compared with non-neoplastic mature and pool of fetal kidney (FK). We used a customized cDNA array containing 4 608 human genes and demonstrated that BL had over representation of genes with similar expression behavior to the earliest stage of normal renal development. Moreover, since WT is a result of loss of developmental control and gain of tumorigenic potential in a successive way, herein we identified genes whose expression level is altered during the kidney development and also in WT and classified as WT-kidney development set. From this set, a smaller group of 36 differentially expressed genes was derived, which was enriched by genes involved in signal transduction, such as APC, BZRP, MET, PLAU, GPR35 and TRADD. An over representation of genes belonging to the WNT signaling pathway were observed. Immunostaining assays of APC and beta-catenin were carried out in 108 specimens showing differential labeling localization in WT. Altogether our data show molecular evidences confirming the recapitulation of embryonic kidney by WT components and strongly suggest that the WNT signaling pathway plays a crucial role in Wilms tumorigenesis.
Keywords: Wilms tumor, cDNA microarray, Histological components, WNT signaling pathway

overall_design: We performed gene expression analyses by customized cDNA array using dye swap with reference design, of unique histological components of Wilms tumors (6 blastemal, 6 epithelial and 3 stromal components) from different patients and compared to 3 normal mature and a pool of fetal kidneys (triplicate). These differentially expressed lists were compared with available data of differentially expressed genes during kidney development.

first: Mariana

last: Maschietto

first: Beatriz

last: de Camargo

first: Helena

last: Brentani

first: Diogo

last: Patrão F

first: Louise

middle: D

last: Mota

first: Isabela

last: Werneck

first: Simone

last: Sredni

first: Cecília

middle: L

last: Costa M

first: Daniel

middle: F

last: Simão

first: Fernando

last: Soares

first: Ricardo

first: Dirce

middle: M

last: Carraro


status: Public on Jul 27 2006

city: Bellinzona

first: Francesco

last: Bertoni

state: Ticino

address: Via V. Vela 6

department: Lymphoma & Genomics Research Program

institute: IOR

summary:
A cyanobacterial LPS antagonist prevents endotoxin shock and blocks sustained TLR4 stimulation required for cytokine expression.  We report the identification and biologic characterization of an LPS-like molecule extracted from the cyanobacterium Oscillatoria Planktothrix FP1 (CyP).
Keywords: different ligands

overall_design: 2 untreated (CTR), 2 treated with LPS at 3h, 2 treated with CyP at 1h,  2 treated with CyP at 3h, 2 treated with LPS + CyP at 3h

first: Annalisa

last: Maccagno

first: Molteni

last: Monica

first: Andrea

last: Rinaldi

first: Antonio

last: Lanzavecchia

first: Rossetti

last: Carlo

first: Federica

last: Sallusto


title: Survey of Intra- and Inter-Tumour Heterogeneity of Gene Expression in Cervical Cancer

status: Public on Oct 01 2006

city: Toronto

first: Paul

last: Boutros

address: 101 College Street, Suite 800

institute: Ontario Institute for Cancer Research

summary:
Purpose: To explore intratumor heterogeneity in gene expression profiles from patients with cervical cancer.

Experimental Design: A total of 33 biopsies were obtained from 11 patients, sampling between two and five different areas for each tumor.The extracted RNAwas hybridized onto the Affymetrix U133 Plus 2.0 oligonucleotide chip. The variance of expression within a patient (W), between patients (B) and the total variance (T =W + B) were calculated for each ProbeSet, and the ratio W/T was used as a measure of intratumor heterogeneity. Gene Ontology functional analysis was done to assess the function of genes that had high W/T (top 10%) and low W/T (bottom 10%) values.

Results: In total, 448 ProbeSets (2.2% of the total) had W/T < 0.10, indicating low intratumor heterogeneity, and 537 ProbeSets (2.7% of the total) had W/T > 0.90, indicating high intratumor heterogeneity. In total 14,473 ProbeSets (72.4%) had higher intertumor than intratumor heterogeneity (W/T < 0.5). Genes with low intratumor heterogeneity were characterized by a statistically significant enrichment of immune-related functions (P < 0.0001). Genes with high intratumor heterogeneity were characterized by a significant tendency towards nuclear localization and nucleic acid binding (both P < 0.0001). For genes with W/T > 0.5, more than six biopsies would be required to minimize the intratumoral heterogeneity to <0.15; if W/T is 0.3 to 0.4, four biopsies are required; and for low W/T of 0.16 to 0.3, only two to three biopsies would be needed.

Conclusion: Although the intratumor heterogeneity was low for the majority of the tested ProbeSets, for many genes, multiple biopsies are required to obtain a reliable estimate of gene expression.

Keywords: disease state comparison

overall_design: Sixty-six flash-frozen punch biopsies were obtained from 16 patients with cervical cancer who were undergoing examination under general anesthesia as part of their pretreatment evaluation for cervical cancer.

first: Barbara

last: Bachtiary

first: Melania

last: Pintilie

first: Willa

last: Shi

first: Carlo

last: Bastianutto

first: Jian

last: Li

first: Joerg

last: Schwock

first: Wendy

last: Zhang

first: Linda

middle: Z

last: Penn

first: Igor

last: Jurisica

first: Anthony

last: Fyles

first: Fei

last: Liu


title: Gene Expression and Functional Evidence of Epithelial-to-Mesenchymal Transition in Papillary Thyroid Cancer Invasion

city: Columbus

first: Sandya

last: Liyanarachchi

state: OH

address: Rm 830, 460, W 12th Avenue

department: Human Genetics

institute: OSU

summary:
Papillary thyroid cancers (PTC) that invade into local structures are associated with a poor prognosis, but the mechanisms for PTC invasion are incompletely defined limiting the development of new therapies. To characterize biological processes involved in PTC invasion, we analyzed the gene expression profiles of microscopically dissected intratumoral samples from central and invasive regions of seven widely invasive PTCs and normal thyroid tissue by oligonucleotide microarray and performed confirmatory expression and functional studies. In comparison to the central regions of primary PTCs, the invasive fronts overexpressed TGFbeta, NFkappaB and integrin pathway members, and regulators of small G-proteins and CDC42. Moreover, reduced levels of mRNAs encoding proteins involved in cell-cell adhesion and communication were identified, consistent with epithelial-to-mesenchymal transition (EMT). To confirm that aggressive PTCs were characterized by EMT, 35 additional PTCs were examined for expression of vimentin, a hallmark of EMT.  Overexpression of vimentin was associated with PTC invasion and nodal metastasis. Functional, in vitro studies demonstrated that vimentin was required for the development and maintenance of both a mesenchymal morphology and invasiveness in thyroid cancer cells. We conclude that EMT is a common mechanism of PTC invasion and that vimentin regulates thyroid cancer EMT in vitro.
Keywords: Genetic modification

overall_design: Total RNA was obtained from all 7 central and invasion regions, as well as from 4 of 7 normal tissues. In addition, the comparison of central vs. normal tissues were compared to 9 paired central and normal samples from The Ohio State University tumor bank simultaneously analyzed using the same methods

first: Vasily

last: Vasko

first: Allan

last: Espinosa

first: William

last: Scouten

first: Huiling

last: He

first: Herbert

last: Auer

first: Alexander

last: Larin

first: Victoria

last: Savchenko

first: Gary

last: Francis

first: Albert

last: de la Chapelle

first: Motoyasu

last: Saji

first: Matthew

last: Ringel


title: Gene expression profiling of CD4+ T-cells and GM6990 lymphoblastoid cell lines

status: Public on Jul 20 2006

city: Bethesda

first: Sean

last: Davis

address: 37 Convent Drive, Room 6138

institute: National Cancer Institute

summary:
CD4+ T-cells isolated from three normal individuals and GM6990 cell lines (three biological replicates) are compared
Keywords: gene expression analysis

overall_design: 8 arrays comparing CD4+ T-cells and GM6990 lymphoblastoid cell lines

first: Gregory

middle: W

last: Crawford

first: Francis

last: Collins

first: Paul

last: Meltzer


title: Stratagene Universal Human RNA Hybridized to Affymetrix U133 Plus 2.0 and U133 A Chips

status: Public on Jul 25 2006

city: Tampa

last: Yoder

state: FL

address: 12902 Magnolia Dr.

department: Molecular Genomics Core Facility

institute: Moffitt Cancer Center & Research Institute

summary:
Stratagene universal human RNA hybridized to both U133 Plus 2.0 and U133 A arrays.
Keywords: Affymetrix platform comparison

overall_design: Stratagene universal human RNA was processed using 5 aliquots of 2 ug each.  The samples were hybridized to both 133 Plus 2.0 and 133 A arrays.

first: Steven

last: Enkemann

first: Lan

last: Zhang


title: Epigenetics of gene expression in human hepatoma cells

city: Indianapolis

first: Howard

last: Edenberg

state: IN

institute: Indiana University School of Medicine

summary:
Expression profiling the response to inhibition of DNA methylation and histone deacetylation.
Comparison of expression in HepG2 cells treated with 5-aza-dC, Trichostatin A, both, or none (control) to change methylation and acetylation status.
Background:DNA methylation and histone deacetylation are epigenetic mechanisms that play major roles in eukaryotic gene regulation. We hypothesize that many genes in the human hepatoma cell line HepG2 are regulated by DNA methylation and histone deacetylation. Treatment with 5-aza-2'-deoxycytidine (5-aza-dC) to inhibit DNA methylation with and/or Trichostatin A (TSA) to inhibit histone deacetylation should allow us to identify genes that are regulated epigenetically in hepatoma cells.
Results:5-aza-dC had a much larger effect on gene expression in HepG2 cells than did TSA, as measured using Affymetrix® HG-U133A Plus 2.0 microarrays. The expression of 1504 probe sets was affected by 5-aza-dC (at p < 0.01), 535 probe sets by TSA, and 1929 probe sets by the combination of 5-aza-dC and TSA. 5-aza-dC treatment turned on the expression of 211 probe sets that were not detectably expressed in its absence. Expression of imprinted genes regulated by DNA methylation, such as H19 and NNAT, was turned on or greatly increased in response to 5-aza-dC. Genes involved in liver processes such as xenobiotic metabolism (CYP3A4, CYP3A5, and CYP3A7) and steroid biosynthesis (CYP17A1 and CYP19A1), and CCAAT element-binding proteins (CEBPA, CEBPB,  and CEBPG) were affected by 5-aza-dC or the combination. Many of the genes that fall within these groups are also expressed in the developing fetal liver. Quantitative real-time RT-PCR assays confirmed selected gene expression changes seen in microarray analyses.
Conclusions:Epigenetics play a role in regulating the expression of several genes involved in essential liver processes such as xenobiotic metabolism and steroid biosynthesis in HepG2 cells. Many genes whose expression is normally silenced in these hepatoma cells were re-expressed by 5-aza-dC treatment. Many genes that are expressed in the fetal liver are up-regulated by demethylation, indicating that DNA methylation is a major factor in restricting the expression of fetal genes during liver development.
Keywords: comparison of treatments

overall_design: 4 cell culture replicates per treatment group.

first: Luke

middle: O

last: Dannenberg


title: Human bronchial epithelial cell differentiation time course

status: Public on Jul 08 2006

city: Research Triangle Park

first: Andrea

last: Ross

state: NC

address: NHEERL/HSD/CRB

institute: US EPA

summary:
Microarray analysis was performed to identify transcriptional changes that occur during mucociliary differentiation of human primary bronchial epithelial cells cultured at an air-liquid interface (ALI).
Keywords: time course

overall_design: Cells from three different donors were cultured and collected at 11 different time points from day 0 to day 28 of ALI culture.

first: Robert

last: Devlin


title: Affymetrix U133 Plus 2.0 Arrays Hybridized With Varying Amounts of Starting Material

summary:
Affymetrix U133 Plus 2.0 arrays were hybridized with varying amounts of starting material to determine whether different measurements of gene expression were observed.
Keywords: Affymetrix starting material study

overall_design: 1 ug, 2, ug, 5, and 10 ug of Stratagene universal human reference RNA was used to setup a series of reactions which were subsequently hybridized to Affymetrix U133 Plus 2.0 arrays.  1 ug samples were performed in triplicate and the entire experiement was repeated.


title: Nef transfection of Jurkat T cells

city: Davis

first: Michael

last: George

address: 451 Health Sciences Drive

department: Medical Microbiology and Immunology

institute: University of California, Davis

summary:
Jurkat T cell line was transfected with SIV Nef controlled by an unducible promoter system. Nef was induced by addition of 10 micromolar pronasterone A and gene expression values were determined after 24 hrs. Control Jurkat cells were untransfected and treated with 10 micromolar pronasterone A and gene expressio values were determined after 24 hrs.
Keywords: SIV Nef, Jurkat, pronasterone A

overall_design:
Experimental conditions: 24 hr SIV Nef expression at physiological concentrations in T cells.

Controls: Non-transfected. 24 hr.


title: The Insecticides Cyfluthrin & Chlorpyrifos Alter Expression of Genes with Diverse Functions in Primary Human Astrocytes

city: New York

first: Sarah

last: Mense

address: 722 West 168th Street

department: Environmental Health Sciences

institute: Columbia University

summary:
Given the widespread use of insecticides in the environment, it is important to perform studies evaluating their potential effects on humans.  Organophosphate insecticides, such as chlorpyrifos, are being phased out; however, the use of pyrethroids in household pest control is increasing.  While chlorpyrifos is relatively well studied, much less is known about the potential neurotoxicity of cyfluthrin and other pyrethroids.  To gain insights into the neurotoxicity of cyfluthrin, we compared and evaluated the toxicity profiles of chlorpyrifos and cyfluthrin in primary human fetal astrocytes.  We found that at the same concentrations, cyfluthrin exerts as great as, or greater toxic effects on the growth, survival, and proper functioning of human astrocytes.  By using microarray gene expression profiling, we systematically identified and compared the potential molecular targets of chlorpyrifos and cyfluthrin, at a genome-wide scale.  We found that chlorpyrifos and cyfluthrin targeted a similar number of transcripts.  These targets include chaperones, signal transducers, transcriptional regulators, transporters, and those involved in behavior and development.  Further computational and biochemical analyses suggest that cyfluthrin and chlorpyrifos up-regulated certain targets of the interferon-gamma and insulin signaling pathways, and that they increased the protein levels of activated ERK1/2, a key component of insulin signaling; IL-6, a key inflammatory mediator; and GFAP, a marker of inflammatory astrocyte activation.  These results suggest that inflammatory activation of astrocytes might be an important mechanism underlying neurotoxicity of both chlorpyrifos and cyfluthrin.
Keywords: treatment comparison

overall_design: Primary human fetal astrocytes treated with 25 uM cyfluthrin, 25 uM chlorpyrifos, or no insecticide for 7 days were collected.  Total RNA was extracted by using TRIzol reagent (GIBCOBRL Life Technologies).  The synthesis of cDNAs and biotin-labeled cRNAs were carried out exactly as described in the Affymetrix GeneChip Expression Analysis Technical Manual (2000).  The human genome U133 plus 2.0 arrays were purchased from Affymetrix, Inc.  Probe hybridization and data collection was carried out by the Columbia University Affymetrix GeneChip processing center.  Specifically, the Affymetrix GeneChip Hybridization Oven 640 and the next generation GeneChip Fluidics Station 450 were used for hybridization and chip processing.  Chip scanning was performed by using the GeneChip scanner 3000.  Initial data analysis was performed by using the Affymetrix GCOS1.2 software.

first: Li


title: Comparative gene expression profile of PPARg and RARa ligand treated human dendritic cells

status: Public on Sep 19 2006

city: Debrecen

first: Istvan

last: Szatmari

address: Egyetem tér 1.

department: Department of Biochemistry and Molecular Biology

institute: University of Debrecen

summary:
Our data indicated that activation of the PPARg nuclear receptor induces a retinoid response in human dendritic cells. In order to assess the contribution of retinoid signaling to the PPARg response we decided to use a combination of pharmacological activators and inhibitors of these pathways. Cells were treated with the synthetic PPARg ligand rosiglitazone (RSG), or with RSG along with the RARa antagonist (AGN193109) to block RARa mediated gene expression, or the RARa specific agonists (AM580) alone.  This design allows one to determine if retinoid signaling is a downstream event of PPARg activation and what portion of PPARg regulated genes are regulated via induced retinoid signaling.
Keywords: ligand response

overall_design: Monocytes were cultured for 5 days with 500 U/ml IL-4 and 800 U/ml GM-CSF, cytokine treatment was repeated at day 3. Ligands were added at the beginning of differentiation but AGN193109 (AGN) treatment was repeated at day 3. Cells were obtained from three healthy individuals (three biological replicates) and cells were treated with vehicle (DC), 2.5 uM rosiglitazone (DC RSG), 100 nM AM580 (DC AM). RSG treatment was also combined with 1 uM AGN193109 (DC RSG AGN) in this case two biological replicates was used (RNA was obtained from 2. and 3. individuals).

first: Laszlo

last: Nagy


title: Global activation of embryonic colon gene expression in murine colon tumor models and human colorectal cancer

city: Cincinnati

first: Bruce

last: Aronow

institute: Cincinnati Children's Hospital Medical Center

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: E6 in HOK/Bmi-1 cells

status: Public on Oct 20 2006

city: Los Angeles

first: Reuben

middle: Han-Kyu

last: Kim

address: 10083 Le Conte Ave.

department: Dentistry

institute: UCLA

summary:
The goal of this study is to identify genes that are differentially expressed by E6 in oral epithelial cells that are stably expressing Bmi-1.
Keywords: oncogenesis.

overall_design: Primary Normal human oral keratinocytes (NHOK) is explanted from normal human gingival tissue.  The cells were infected with retroviruses capable of expressing Bmi-1 and being selected.  Actively dividing HOK/Bm-1 cells were then superinfected with LXSN-based retorviruses capable of expressing E6 and E6(delta)118-122, and selected.  Actively proliferating cells are then harvested to isolate total RNA which was subsequently subjected to microarray analysis.

first: Mo

last: Kang

first: Ki-Hyuk

last: Shin

first: No-Hee

last: Park


title: Monocyte Derived Dendritic Cell Maturation

status: Public on Aug 02 2006

first: Benhur

last: Lee

address: 609 Charles E Young Dr East

department: MIMG

summary:
Human monocyte derived dendritic cells matured via galectin-1 or LPS.
Keywords: dendritic cell maturation

overall_design: 4 conditions with 3 replicates of each include control or intreated immature MDDCS, galectin-1 treated MDDCs, LPS treated MDDCs, and vehicle control MDDCs. (each replicate is from a distinct DONOR)

first: Jennifer

last: Fulcher

first: Sara

last: Hashimi

first: Ernest

last: Levroney

first: Mabel

last: Pang

first: Kevin

middle: B

last: Gurney

first: Linda

middle: G

last: Baum


title: Gene expression profiling of CD34+ cells from MDS patients and normal controls

status: Public on Jul 01 2006

city: Oxford

first: Andrea

last: Pellagatti

address: John Radcliffe Hospital

department: NDCLS, RDM

country: United Kingdom

institute: University of Oxford

summary:
In order to gain insight into the poorly understood pathophysiology of the myelodysplastic syndromes (MDS), we have determined the gene expression profiles of the CD34+ cells of 55 MDS patients using the Affymetrix GeneChip U133 Plus2.0 platform
Keywords: Disease v normal

overall_design: 55 patients with MDS and 11 healthy controls were included in the study. At the time of investigation, 18 patients had RA, 19 RARS and 18 RAEB. The RAEB group was further subdivided into RAEB1 and RAEB2 for a selected part of the analysis. Bone marrow samples were obtained and CD34+ cells isolated from MDS patients and healthy controls using MACS magnetic cell separation columns. Total RNA was extracted using TRIZOL following the protocol supplied by the manufacturer. RNA was amplified using the Two-Cycle cDNA Synthesis and the Two-Cycle Target Labelling and Control Reagent packages (Affymetrix). Samples were hybridized to GeneChip Human Genome U133 Plus 2.0 arrays. Affymetrix CEL files were pre-processed using Robust MultiChip Analysis (RMA).

first: James

middle: S

last: Wainscoat

first: Jacqueline

last: Boultwood


title: hTERT effects in CD8 T Lymphocytes from two healthy donors

status: Public on Jun 24 2006

city: Lausanne

first: Mauro

last: Delorenzi

address: Génopode Building, Quartier Sorge

institute: Swiss Institute of Bioinformatics

summary:
Expression in GFP vs. GFP/hTERT transduced CD8 T Lymphocytes from Healty Donors (HD) 1 and 2 at early and late passages. Using CD8+ T lymphocyte clones over-expressing telomerase we investigated the molecular mechanisms that regulate T cell proliferation. Transduction and subcloning procedures were performed on CD8 + naive T-cell clones isolated from two different healthy individuals aged between 30 to 35 years (HD1 and HD2). T-cell cloneswere transduced to express hTERT/GFP or GFP alone.
Keywords: Cell Line Comparison.

overall_design: triplicates of 4 conditions for each of 2 donors, 1 chip (hTERT_YOUNG_HD1_REP2) of the first donor excluded because of low quality hybridization, one condition (GFP YOUNG)lost from second donor because of problems in cell culture.

first: Nathalie

last: Rufer

first: Olivier

last: Menzel

first: Marco

last: Migliaccio

first: Darlene

last: Goldstein

first: Sophie

last: Dahoun


title: Large-scale deployment of embryonic gene programming in human and murine colon cancer: a new target for intervention.

first: Bruce

middle: J

last: Aronow

summary:
Goal of the experiment: To identify and understand the overall transcriptional programming of human colorectal cancer tumors by evaluating gene expression profiles of tumors from four murine models, and comparing and contrasting these to the developing stages of the mouse embryonic colon.
Experiment description: Colorectal cancer results from multiple genetic and epigenetic events that produce variable histologies and clinical outcomes. To identify gene regulatory programs that underlie colon tumorigenesis, we profiled gene expression in 39 mouse colon tumors from four independent mouse models and compared this to mouse colon embryonic development, as well as with 100 human colon carcinomas. Here, we report a striking recapitulation of embryonic patterns of gene expression in both mouse and human colon tumors.  All four of the mouse colon tumor models exhibited large-scale activation of embryonic gene expression signatures.  The two nuclear beta-catenin-positive mouse tumors (azoxymethane-treated [AOM] and ApcMin/+), exhibited strong activation of genes characteristic of those expressed in the earliest embryonic stages, while tumors from two other models (Smad3-/- and Tgfb1-/- x Rag2-/-) exhibited lower activation of early stage-specific genes but substantial expression of general embryonic colon genes.  Human colon cancer cases over-expressed genes characteristic of both early and late embryonic stages.  Examining tumor gene expression through the lens of development has revealed an extensive network of therapeutic targets for cancer control.
Keywords: Colorectal cancer, tumor, animal models, development, comparative genomics, orthologenome

overall_design:
Transcriptional signatures across human colorectal carcinomas and normal human colon tissue samples were evaluated by referencing all the samples to the median gene expression value across all the samples. A second complementary strategy compared the gene expression levels of the tumors to those of normal adult colon.
Values were transformed from a log base 2 to linear values.  Each measurement was divided by the 50.0th percentile of all measurements in that sample.     Each gene was divided by the median of its measurements in all samples. If the median of the raw values was below 10 then each measurement for that gene was divided by 10 if the numerator was above 10, otherwise the measurement was thrown out.


title: Gene copy number polymorphism, a major source of genetic variation between humans

first: David

middle: Lawrence

last: Newsom

address: 700 Children's Drive

department: Microbial Pathogenesis

institute: Nationwide Children's Hospital

summary:
We have performed a genome wide analysis of gene copy number polymorphisms and report here for the first time that the human genome contains thousands of well-characterized genes at copy numbers different from one maternal and one paternal allele; and that, furthermore, the copy numbers of hundreds of well-characterized genes can vary between two normal healthy humans making this a major source of genetic variation. Groups of genes affected by CNPs include genes involved in signal transduction, oncogenesis, cell adhesion activity and several types of immune response. In contrast to SNPs, which preferentially affect non-coding regions of the genome, copy number polymorphisms of well-characterized and actively expressed genes are very likely to have important biological consequences.
Keywords: Gene Copy Number Variation

overall_design: We utilized an expression microarray to analyze gene copy number variations (GCNVs).

first: Herbert

last: Auer

first: Karl

last: Kornacker


title: Evaluation of RNA binding specificity of aminoglycosides with DNA microarrays

status: Public on Jun 22 2006

city: La Jolla

first: William

middle: A

last: Greenberg

address: 10550 N Torrey Pines Rd

department: Chemistry

institute: The Scripps Research Institite

summary:
We have developed methods for using DNA array technology to probe the entire transcriptome to determine RNA-binding specificity of ligands.  Two methods were investigated.  In the first, the RNA-binding aminoglycoside antibiotic tobramycin was covalently linked to magnetic beads.  The beads were bound to human liver mRNA and washed, and specifically bound RNA was eluted, amplified, and analyzed with DNA array technology. A small number of genes were found to bind specifically to the tobramycin beads.  In the second method, the aminoglycoside ligand was added directly to the array hybridization reaction, and the signal compared to a control experiment in the absence of ligand. The aminoglycosides were found to interfere with a small percentage of all hybridization events. These methods differ from traditional DNA array experiments in that the readout is a direct measure of the interaction between mRNA and a ligand, rather than an indirect measure of effect on expression.  We expect the results will lead to discovery of new aminoglycoside-binding RNA motifs, and may also have relevance toward understanding and overcoming side effects observed with these antibiotics in the clinic.
Keywords: RNA binding analysis

overall_design: 5 samples including controls were analyzed for effect of RNA binding ligands on hybridization of mRNA to DNA microarrays.

first: Fu-Sen

last: Liang

first: Jennifer

last: Hammond

first: Julia

last: Hoffmann

first: Steven

last: Head

first: Chi-Huey

last: Wong


title: Genomic profile of the estrogen induced neoplastic transformation of the human breast epithelial cell MCF10F

status: Public on Jun 20 2006

city: Charlottesville

first: Yong

last: Huang

address: 1340 Jefferson Park Ave

institute: University of Viginia

summary:
Estrogen is a risk factor for breast cancer, yet its mechanism in the initiation of this disease is not clear. Here we use a model of estrogen-mediated malignant transformation of MCF-10F cells to identify the temporal acquisition of changes in genome structure and gene expression that correspond to the progressive transformed phenotype culminating in tumorigenesis.  Genomic DNA and total RNA were isolated from MCF10F, trMCF (MCF10F transformed by 70 nM 17-beta estradiol), bsMCF (trMCF selected by Boyden chamber, tumorigenic), bcMCF(clones of bsMCF), and caMCF (cells from tumors of bsMCF grown in SCID mice). The Affymetrix 100k SNP and HG-U133_Plus_2 chips were used for genotyping and gene expression analyses. Changes in chromosomal copy number and loss of heterozygosity were progressive.  Gross changes were rarely observed in the trMCF10F cells; the earliest was a gain in chromosome 1p, 1p36.12-1p36.21.  In the bsMCF and their sub-clones bcMCF, additional gains were seen in chromosomes 1p, 1p36.12-1pter, 5q, 5q21.1-5q35.3, and 13q, 13q21.32-13q34; losses were detected in chromosomes 4 and 8p, 8p11.1-8p23.1.  In the caMCF, additional losses were seen in chromosomes 3p, 3p12.1-3p14.1, 9p, 9p22.1-9p24.3, and 18q, 18q11.2-18q23.  Using microarray analysis we were able, in some cases, to narrow the regions of interest and their associated candidate cancer genes.  The observation of progressive genomic changes, along with the tumor phenotype of a poorly differentiated adenocarcinoma, indicate an underlying mechanism common to breast cancer development in humans, and adds to the evidence indicating that estrogens are mutagenic and contribute early to the process of breast cancer.
Keywords: Cell type comparison

overall_design: Human MCF-10F cells were transformed by 70nM 17-beta-estradiol. The transformed cells were selected by Boyden chamber. The selected cells were injected into SCID mice and also cloned and expanded. The thus grown tumors were harvested for cell culture. Genomic DNA and total RNA were isolated from cell lines in each step for 100K SNP and gene expression microarray assay.

first: Thomas

middle: R

last: Sutter

first: Sandra

middle: V

last: Fernadez

first: Patricia

last: Russo

first: Shirlean

last: Goodwin

first: Irma

middle: H

first: Jose


title: Transcriptional profile of aging human muscle

status: Public on Jun 16 2006

city: Stanford

first: Jacob

last: Zahn

address: 279 Campus Dr.

department: Developmental Biology

institute: Stanford University

summary:
We analyzed expression of 81 normal muscle samples from humans of varying ages, and have identified a molecular profile for aging consisting of 250 age-regulated genes. This molecular profile correlates not only with chronological age but also with a measure of physiological age. We compared the transcriptional profile of muscle aging to previous transcriptional profiles of aging in kidney and brain, and found a common signature for aging in these diverse human tissues. The common aging signature consists of six genetic pathways; four pathways increase expression with age (genes in the extracellular matrix, genes involved in cell growth, genes encoding factors involved in complement activation, and genes encoding components of the cytosolic ribosome), while two pathways decrease expression with age (genes involved in chloride transport and genes encoding subunits of the mitochondrial electron transport chain). We also compared transcriptional profiles of aging in human to those of the mouse and fly, and found that the electron transport chain pathway decreases expression with age in all three organisms, suggesting that this may be a public marker for aging across species.
Keywords: time course

overall_design: Human skeletal muscle samples of various ages (spanning 16-89 years) were surgically removed from patients and gene expression changes with age profiled using Affymetrix HU-133 2.0 arrays.  62 muscles were taken from the rectus abdominis; 19 were taken from various other regions of the anatomy.

first: Rebecca

last: Sonu

first: Stuart

last: Kim


title: Genome-wide expression profile of human trabecular meshwork cultured cells, non-glaucomatous and POAG tissue

status: Public on Jun 21 2006

city: Durham

first: Pedro

last: Gonzalez

address: Erwin Road Box 3802

department: Ophthalmology

institute: Duke University

summary:
The goal of this study was to contrast genome-wide gene expression profiles of cultured human trabecular meshwork (HTM) cells, to that of control and primary open angle glaucoma (POAG) HTM tissues.

Total RNA from cultured HTM cells and HTM tissue dissected from control and POAG anterior segments fixed in RNA later™ was linearly amplified with the OvationTM Biotin RNA Amplification and Labeling System and individually hybridized to Affymetrix Human Genome U133 Plus 2.0 high density microarrays. Data analysis was performed using the GeneSpring Software 7.0. Selected genes showing significant differential expression were validated by Quantitative real-time PCR in nonamplified RNA.

Cultured HTM cells retained the expression of some genes characteristic of the HTM tissue, including chitinase 3-like 1 and matrix Gla protein, but demonstrated downregulation of physiologically important genes such as myocilin. POAG HTM tissue showed relatively small changes compared to that of control donors. These changes included the statistically significant upregulation of several genes associated with inflammation and acute-phase response, including selectin-E (ELAM-I), as well as the downregulation of the antioxidants, paraoxonase 3 and ceruloplasmin.

Downregulation in cultured HTM cells of genes potentially relevant for outflow pathway function highlights the importance of developing new conditions for the culture of TM cells capable of preserving the characteristics of TM cells in vivo. Comparative analysis between control and POAG tissues suggests that the upregulation of inflammation-associated genes might be involved in the progression of glaucoma.
Keywords: Cell type comparison

overall_design: Total RNA from three cultured HTM cells and HTM tissue dissected from three pairs of control and two pairs of POAG anterior segments fixed in RNA later™ was linearly amplified with the OvationTM Biotin RNA Amplification and Labeling System and individually hybridized to Affymetrix Human Genome U133 Plus 2.0 high density microarrays.

first: Paloma

last: Liton

first: Coralia

last: Luna

first: Pratap

last: Challa

first: David

last: Epstein


title: Novel allele of crybb2 in the mouse and its expression in the brain

city: Neuherberg

first: Martin

last: Irmler

state: Bayern

address: Ingolstaedter Landstrasse 1

department: Institute of Experimental Genetics

country: Germany

institute: Helmholtz Zentrum München GmbH

summary:
O377 mutant mouseline with progressive lens opacity. Mutation mapped to distal chromosome 5 and finally characterized by an A - T bp substiution at the end of intro 5 of Crybb2. Crybb2 expression also found in olfactory bulb, pyramidal cells of the cortex, hippocampus, Purkinje and stellate cells of cerebellum.
Keywords: other

overall_design: 15 dual-color DNA-chip hybridizations including 8 dye swap experiments of cDNAs were made of brains

first: Koustav

last: Ganguly

first: Graw

last: Jochen

first: Marion

last: Horsch

first: Johannes

last: Beckers


title: Transcriptome of GIP- and ACTH-dependent Cushing's syndrome.

city: Montréal

first: Antoine

last: Lampron

state: Qc

address: 3840 St-Urbain

department: Research Centre

institute: CHUM-Hotel Dieu

summary:
Abstract submitted to the Journal of clinical encodcrinology and metabolism:

The molecular mechanisms responsible for the ectopic expression of the GIP receptor in the adrenal cortex of patients with GIP-dependent Cushing’s syndrome (CS) are unknown. Chronic adrenal stimulation by ACTH in Cushing’s disease (CD) or by GIP in GIP-dependent AIMAH both lead to induction of a set of genes which stimulate adrenal proliferation and steroidogenesis. The objective of this study was to compare the whole genome expression profile of adrenal glands of five cases of GIP-dependent bilateral macronodular adrenal hyperplasia with CS, one case of GIP-dependent adenoma, compared to five cases of ACTH-dependant hyperplasias (CD) and a pool of adrenals from 62 normal individuals. We used the genome-spanning Affymetrix U133 plus 2.0 microarray oligochips to identify genes differentially expressed specifically in GIP-dependent CS and which would be candidate genes implicated in the ectopic expression of the GIP receptor in the adrenal cortex. After data normalization and filtering, genes with differential expression were identified with a multi-step statistical analysis involving a student’s t-test, a filter on flags and a SAM analysis. A total of 721 probesets were thus isolated with intensity levels robustly related to the presence of a GIP-dependent hyperplasia. We performed a functional classification to further define potentially important biological processes and signaling mechanism for the formation of GIP-dependent AIMAH. Various probesets were related to metabolic processes, cell-surface and intracellular signaling, tumorigenesis, transport and transcription factors. The most relevant genes had their expression profile confirmed by real-time RT-PCR. This study reports an extensive series of potentially novel targets in the identification of the molecular mechanisms of ectopic expression of the GIP-receptor in this pathology.
Keywords: Comparative genomic analysis

overall_design: The 5 GIP-dependent AIMAH (G1 to G5) and 5 ACTH-dependent hyperplasias (H1 to H5) were grouped to perform statistical filtering of the data.

first: André

last: Lacroix

first: Isabelle

last: Bourdeau

first: Johanne

last: Tremblay

first: Pavel

last: Hamet


title: hTERT effects in CD8 T Lymphocytes HD2

first: Mauro

last: Delorenzi

summary:
Using CD8+ T lymphocyte clones over-expressing telomerase weinvestigated the molecular mechanisms that regulate T cell proliferation.  Transduction and subcloning procedures were performed on CD8 + naive T-cell clones isolated from two different healthy individuals aged between 30 to 35 years (HD1 and HD2). T-cell cloneswere transduced to express hTERT/GFP or GFP alone. HD1 was profiled on U133A and submitted as a separate GEO series.
Keywords: Cell Line Comparison.

overall_design: triplicates of 4 conditions, 1 condition (GFP YOUNG)  lost because of problems in cell culture.

first: Nathalie

last: Rufer

first: Olivier

last: Menzel

first: Marco

last: Migliaccio

first: Darlene

last: Goldstein

first: Sophie

last: Dahoun


title: Polyamides alleviate transcription inhibition associated with long GAA•TTC repeats in Friedreich’s ataxia

status: Public on Jun 14 2006

first: Ryan

middle: Stephen

last: Burnett

address: 10550 N. Torrey Pines Road, MB-27

department: Molecular Biology

institute: The Scripps Research Institute

summary:
Lymphoblast cells from a patient with Freidriech's Ataxia were incubated with pyrrole-imidazole polyamides targeted to the GAA triplet repeat in the intron 1.  The polyamides were shown in cell culture to increase levels of endogenous frataxin mRNA.  A normal sibling derived lymphoblast cell line was used as a control.
Keywords: human lymphoblast cells

overall_design: Normal (GM15851) and patient (GM15850) cell lines were incubated in the presence of match polyamide FA1 at 1uM, 2uM or mismatch polyamide FA2 at 2uM for 7days prior to RNA purification and microarray analysis.

first: Christian

last: Melander

first: James

last: Puckett

first: Peter

last: Dervan

first: Joel

last: Gottesfeld


title: Greene-5P01NS017771-220003

status: Public on Jun 06 2006

city: Phoenix

first: Winnie

last: Liang

state: AZ

address: 445 N. Fifth Street

institute: Translational Genomics

summary:
Parkinson's disease is a prevalent neurodegenerative disorder for which there is no cure.  The cause of PD symptoms is loss of dopamine neurons in the midbrain, but it is not known why these neurons die.  Pesticide exposure is epidemiologically associated with PD, and administration of the organic pesticide rotenone to rats recapitulates most of the behavioral, neurochemical, and neuropathological findings in PD, including specific death of dopamine neurons.  We have developed an in vitro model of rotenone toxicity using a dopaminergic cell line (SK-N-MC neuroblastoma cells) that mimics many of the cellular changes seen with in vivo rotenone toxicity and with PD, such as alpha-synuclein aggregation and oxidative damage.  We are currently using this simple model to explore mechanisms of dopaminergic neurodegeneration, with our ultimate goal being the discovery of novel mechanisms for dopaminergic neuroprotection in PD.
We will examine gene expression profiles of cultured SK-N-MC cells at several time points during rotenone exposure to determine pathways involved in rotenone toxicity and dopaminergic degeneration.  We will compare these profiles to baseline profiles of rodent dopaminergic neurons that we have already obtained, as well as to profiles of dopamine neurons from rotenone-treated rats that we will obtain in the near future.  We will also compare these data to published results from SN neurons from human PD patients. This technique will not only help us to detect gene expression changes relevant to dopaminergic neurodegeneration in PD, but it will allow us to determine if the SK-N-MC system can be reliably used to screen for neuroprotective therapies for PD.  We anticipate that SK-N-MC cells will show a relevant subset of the gene changes seen in dopamine neurons in vivo and that this will guide us in the sorts of mechanisms and drugs that can be screened in this system.
Chronic exposure to low levels of rotenone causes changes in gene expression in SK-N-MC cells that sensitize the cells to toxic insults.  We also hypothesize that there are several compensatory protective pathways that are stimulated by chronic rotenone, although these pathways are ultimately ineffective at preventing damage.  We anticipate that gene expression profiling of rotenone-treated cells over time will suggest several novel strategies for neuroprotective intervention.
SK-N-MC cells will be grown in three different media: media only, vehicle (EtOH), and rotenone (5 nM).  All current experimental evidence in our lab indicates that vehicle-treated cells are indistinguishable from media-only cells. Rotenone-treated cellls have a stereotypical response in culture.  At one week, the only noticed change is an increase in alph-synuclein aggregation.  At two weeks, evidence of increased oxidative stress appears (increased protein carbonyls and lipid peroxidation).  At four weeks, the cells are markedly sensitized to oxidative challenge with H2O2.  Therefore, we will examine gene expression at baseline, and during 1, 2, and 4 weeks of rotenone treatment. Three experiments will be performed, each lasting 4 weeks.  For each experiment, three separate dishes of vehicle-treated, and rotenone-treated cells will be harvested at 1, 2, and 4 weeks (18 independent samples).  Untreated, media-only cells will be harvested after 1 week in vitro to serve as baseline cells.  Total RNA will be isolated. An equal amount of RNA from one dish per experiment per group will be used to compose the final samples.  Therefore, each independent sample will consist of RNA from 3 separate experiments.  This will allow us to take advantage of a pooling strategy, yet not sacrifice  technical and biological replication. 21 samples will be sent to the Consortium.  Three will be from untreated cells.  Nine will be vehicle-treated at 1, 2, and 4 weeks (3 each).  Nine will be rotenone-treated at 1, 2 , and 4 weeks (3 each).  Each sample will be labeled and hybridized to one Affymetrix Human Genome U133 Plus 2.0 Gene Chip. With assistance of the consortium, we will analyze the data using the Signifiance Analysis of Microarrays (SAM) program and self-organizing map algorithms.
Keywords: time-course

last: Greene


title: MicroArray Quality Control (MAQC) Project

status: Public on Sep 08 2006

city: Shanghai

first: Leming

last: Shi

address: 2005 Songhu Road

department: School of Life Sciences

country: China

institute: Fudan University

type:
Expression profiling by array
Expression profiling by RT-PCR

summary:
Microarray technology has had a profound impact on gene expression research.  Some studies have questioned whether similar expression results are obtained when the same RNA samples are analyzed on different platforms.

The MicroArray Quality Control (MAQC) project was initiated to address these concerns, as well as other performance and analysis issues.  We demonstrate the consistency of results within a platform across test sites as well as the high level of cross-platform concordance in terms of genes identified as differentially expressed.  The MAQC study provides a rich resource that will help build consensus on the use of microarrays in research, clinical and regulatory settings.  Manuscripts related to the MAQC project have been published in Nature Biotechnology, 24(9), September, 2006.  More information about the MAQC project can be found at http://edkb.fda.gov/MAQC/.
Keywords: Cross-platform comparison

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design:
Expression data from two distinct reference RNA samples (A and B) in four titration pools were generated at multiple test sites using a variety of microarray-based and alternative technology platforms.  Sample A = Stratagene Universal Human Reference RNA (UHRR, Catalog #740000), Sample B = Ambion Human Brain Reference RNA (HBRR, Catalog #6050), Sample C = Samples A and B mixed at 75%:25% ratio (A:B); and Sample D = Samples A and B mixed at 25%:75% ratio (A:B).  In general, each microarray platform was tested at three sites and each sample was tested in five replicates at each test site.  Samples (hybridizations) were named according to the following convention: Platform_Testsite_SampleRelicate.  For example, AFX_2_B1 represents the hybridization (array) from platform AFX processed by test site 2 for the first replicate of sample B.

Assignment of platform code: ABI = Applied Biosystems (microarray); AFX = Affymetrix; AG1 = Agilent one-color; AGL = Agilent two-color; GEH = GE Healthcare; ILM = Illumina; NCI = NCI two-color (Operon oligos); EPP = Eppendorf; TAQ = TaqMan (Applied Biosystems); QGN = QuantiGene (Panomics); GEX = StaRT-PCR (Gene Express); H25K = TeleChem two-color; H25K1 = TeleChem one-color; BIO = CapitalBio two-color (Operon oligos); BIO1 = CapitalBio one-color (Operon oligos); OPN = Operon two-color (Operon oligos); NMC = Norwegian Microarray Consortium two-color (Operon oligos).

first: Laura

last: Reid

first: Wendell

middle: D

last: Jones

first: Richard

last: Shippy

first: Janet

last: Warrington

first: William

last: Slikker Jr


title: Expression data from p63 siRNA in squamous cell lines

city: Nashville

first: Jennifer

last: Pietenpol

state: TN

address: 23rd Ave South at Pierce

department: Biochemistry

institute: Vanderbilt University Medical Center

summary:
p63, a homologue of the tumor suppressor p53, is critical for the development and maintenance of squamous epithelia.  p63 is specifically expressed in the basal layers of stratified epithelial tissues, and is considered to be a specific marker for cells of this type.  The role of p63 in tumorigenesis remains poorly defined.  Numerous studies have highlighted the oncogenic potential of the predominant p63 isoform, ΔNp63α; however, data suggests that other p63 proteins can act as tumor suppressors or alter the metastatic potential of tumors.  ΔNp63α can act as a transcriptional repressor, but the link between the transcriptional functions of p63 and its biological role is still unclear.  In this study, we used a loss-of-function approach to investigate the transcriptional programs controlled by p63.
Keywords: cell line panel

overall_design: A panel of squamous cell lines, including transformed and non-transformed cells, were transfected with hygromycin selectable vectors encoding p63 siRNA or control, then were selected with Hygromycin B and harvested.

first: Christopher

middle: E

last: Barbieri


title: Identifying targets of MeCP2 during neuronal maturational differentiation

status: Public on Jun 15 2006

first: Sailaja

last: Peddada

address: 3426 Tupper Hall, One Shields Avenue

institute: University of California, Davis

summary:
Rett syndrome (RTT) is an X-linked dominant neurodevelopmental disorder caused by mutations in MECP2, encoding methyl-CpG binding protein 2. MeCP2 is a transcriptional repressor elevated in mature neurons and is predicted to be required for neuronal maturation by regulating multiple target genes. Identifying primary gene targets in either Mecp2-deficient mice or human RTT brain has proven to be difficult, perhaps because of the transient requirement for MeCP2 during neuronal maturation. In order to experimentally control the timing of MeCP2 expression and deficiency during neuronal maturation, human SH-SY5Y cells undergoing mature neuronal differentiation were transfected with methylated MeCP2 oligonucleotide decoy to disrupt the binding of MeCP2 to endogenous targets. Genome-wide expression microarray analysis identified all four known members of the inhibitors of differentiation or inhibitors of DNA binding (ID1, ID2, ID3 and ID4) subfamily of helix-loop-helix (HLH) genes as novel neuronal targets of MeCP2. Chromatin immunoprecipitation analysis confirmed binding of MeCP2 near or within the promoters of ID1, ID2 and ID3, and quantitative RT-PCR confirmed increased expression of all four Id genes in Mecp2-deficient mouse brain. All four ID proteins were significantly increased in Mecp2-deficient mouse and human RTT brain using immunofluorescence and laser scanning cytometric analyses. Because of their involvement in cell differentiation and neural development, ID genes are ideal primary targets for MeCP2 regulation of neuronal maturation that may explain the molecular pathogenesis of RTT.
Keywords: Neuronal Differentiation, Targets of MeCP2, Rett syndrome.

overall_design:
The series GSE4600 submission consists of 12 samples representing data from three biological replicates with four treatments (UD, D-UT, D-MD, and D-CD) per one replicate experiment. This study was conducted to identify the neuronal targets of MeCP2 during PMA induced SH-SY5Y neuronal maturational differentiation. Data from each sample consisted of dChip derived PM-only model gene expression index from four SH-SY5Y treatments:
1. Undifferentiated SH-SY5Y cells, i.e. before PMA induced differentiation (UD) (Sample ID - GSM102825, GSM102842 and GSM102870).
2. 48 h differentiated and untransfected cells (D-UT) (Sample ID - GSM102875, GSM102876 and GSM102877).
3. 48 h differentiated and MeCP2 decoy transfected cells (D-MD) (Sample ID – GSM102878, GSM102879 and GSM102880).
 4. 48 h differentiated and Control decoy transfected cells (D-CD) (Sample ID – GSM102881, GSM102882 and GSM102883).
Affymetrix HG U133 plus 2.0 genechips were used. Data analysis was performed using dChip analysis software and significant differences between different cell treatments were identified. List of genes regulated by MeCP2 were obtained by comparing gene lists with fold changes ≥ 2 and significant P values ≤ 0.05 between undifferentiated and MeCP2 decoy transfected but not undifferentiated and control decoy (UD vs. D-MD NOT D-CD) during neuronal maturational differentiation. Short listed candidate gene targets were validated by quantitative real time RT-PCR. Additional functional studies were also conducted on interesting and relevant neuronal targets of MeCP2.

first: Dag

last: Yasui

first: Janine

middle: M

last: LaSalle


title: Gene Expression Profiles of Multiple Myeloma (N=414) Before Treatment

status: Public on Jun 01 2006

city: Little Rock

first: Shaughnessy

middle: Jr.

last: John

state: AR

address: 4301 West Markham St., Slot 776

institute: Myeloma Institute for Research and Therapy

summary:
Samples in this series are pre-treatment bone marrow aspirates from multiple myeloma patients
Keywords: other

overall_design:
Samples in this series are: CD-138-selected plasma cells from bone marrow of newly diagnosed multiple myeloma patients susequently treated with Total Therapy

CEL files are available upon request via a Materials Transfer Agreement. Please contact Dr. John Shaughnessy  for details.

first: John

last: Shaughnessy


title: Beta-catenin/LEF-1 induced epithelial-mesenchymal transition in DLD1 colon carcinoma cells

city: Boston

first: Damian

middle: Darren

last: Medici

state: MA

address: 188 Longwood Ave. REB 413

department: Cell Biology

institute: Harvard Medical School

summary:
Our goal was to assess gene expression changes that occur when Lymphoid Enhancer Factor-1 (LEF-1) promotes epithelial-mesenchymal transition (EMT), the primary mechanism of tumor metastasis. To observe this phenomenon without interference from other signaling pathways, we selected DLD1 colon carcinoma cells (ATCC) which contain a mutation in APC. APC is a necessary component of a ubiquitin protein complex (including GSK-3beta, Axin, etc.) that is responsible for degrading cytoplasmic beta-catenin. Therefore, sufficient levels of LEF-1 can be easily activated by forming complexes with the abundant beta-catenin located in the cytoplasm of DLD1 cells. These complexes can then promote transcription of genes that stimulate EMT.

We treated DLD1 cells with an adenoviral LEF-1 expression construct, which induced EMT within 48 hours. RNA was then extracted from these cells along with untreated DLD1 cells, then subjected to microarray analysis. From this analysis, we acquired several gene expression profiles by which epithelial colon carcinoma cells transform to an invasive, mesenchymal phenotype to initiate metastasis.
Keywords: epithelial-mesenchymal transition, tumor metastasis, cancer progression, epithelial cell plasticity

overall_design: DLD1 cells were treated with an adenoviral LEF-1 expression construct as described by Kim et al. (2002). Total RNA was extracted from both untreated and treated DLD1 cells using the RNeasy mini extraction kit (Qaigen). RNA amplification, biotin labeling, microarray hybridization, and fluidics were performed following the eukaryotic sample and array processing protocol (Affymetrix). Chips were scanned using an Affymetrix Gene Array Scanner (Hewlett-Packard). Raw data was compiled using Microarray Suite 5.0 software (Affymetrix).

first: Elizabeth

last: Hay

first: Bjorn

last: Olsen


title: Gene Expression Profiles of Multiple Myeloma Before Treatment

summary:
Samples in this series are pre-treatment bone marrow aspirates from multiple myeloma patients

Keywords = CKS1B in Multiple Myeloma, Bone Marrow, Pre-Treatment
Keywords: other

overall_design:
Samples in this series are:

1. CD-138-selected plasma cells from bone marrow of newly diagnosed multiple myeloma patients susequently treated with Total Therapy 2 (pre-treatment TT2)

2. CD-138-selected plasma cells from bone marrow of newly diagnosed multiple myeloma patients susequently treated with Total Therapy 3 (pre treatment TT3).

Overall Design:

Baseline gene expression signatures were used to: 1) develop unsupervised hierarchical cluster classes; 2) establish links with outcome following stem cell transplantation

CEL files are available upon request via a Materials Transfer Agreement. Please contact Dr. John Shaughnessy  for details.


title: Expression profiling of leukemia cell lines

status: Public on May 31 2006

city: Regensburg

first: Michael

last: Rehli

address: F.-J.-Strauss-Allee 11

department: Internal Med III

institute: University Hospital Regensburg

type:
Expression profiling by array
Methylation profiling by array

summary:
Methylation of CpG islands is associated with transcriptional repression and, in cancer, leads to the abnormal silencing of tumor-suppressor genes. Genome wide methylation profiling of myeloid leukemia cell lines identified a large number of genes with aberrantly methylated CpG islands. Comparative mRNA expression analysis suggests that more than half of these genes show extremely low or absent expression in normal cells, suggesting that hypermethylation in cancer may be independent of the transcriptional status of the affected gene.
Keywords: comparison of expression status and CpG methylation status

overall_design: The expression profiles of the three leukemia cell lines KG-1, U937 and THP-1 was compared to normal human blood monocytes (reference). The set includes a single hybridization for each sample.

first: Claudia

last: Gebhard

first: Lucia

last: Schwarzfischer


title: Expression data from whole blood

status: Public on May 26 2006

city: Helsinki

first: Antti

last: Kokko

address: POBOX 63 (Haartmaninkatu 8)

department: Dept of Medical Genetics

country: Finland

institute: University of Helsinki

summary:
While identification of genes mutated in high penetrance tumor predisposition syndromes has been a success story, much less progress has been made in characterizing the genetic basis of low penetrance tumor susceptibility. Combining recently introduced chip-based technologies with traditional genealogy work we have identified inactivating germline mutations in patients with pituitary adenoma predisposition (PAP).
The experiment consisted of a collection of blood samples from identified families where PAP was observed and analysis of gene expression data used together with SNP genoptyping and linkage analysis. The findings were further studied using direct screening and other supporting methods.
Keywords: affected and obligatory carriers vs controls

overall_design: To identify the PAP locus whole genome SNP genotyping and linkage analysis was combined with gene expression profiling from 16 individuals (9 affected/obligatory carriers: A2, A6, A8, A14, A16, A18, A20, A21, A22, and 7 controls). Statistical analysis was performed on probe sets mapped to the linked region.

first: Pia

last: Vahteristo

first: Virpi

last: Launonen

first: Auli

last: Karhu

first: Lauri

last: Aaltonen


title: Scheck-3U24NS043571-01S1-2

first: Winnie

last: Liang

summary:
Meningiomas are typically considered a benign tumor that can be cured by complete surgical resection; however, a percentage of patients have recurrent disease, even after apparently complete resections. These patients require additional surgeries, radiation therapy, chemotherapy, or a combination of all three. The ability to recognize these patients prior to recurrence would promote earlier use of adjuvant therapy, thus improving overall patient outcome. Unfortunately, identification of meningiomas with this more aggressive phenotype is difficult, and standard histopathological techniques rarely suffice. The identification of genetic and molecular parameters that can help to define these more aggressive tumors would improve prognostication and treatment planning for patients with meningiomas.
1. Establish gene profiles for benign (grade 1) and aggressive (grades 2 and 3) meningiomas. 2. Determine if there are particular expression profiles that can help differentiate between benign and aggressive meningiomas. 3. Determine if there is/are specific gene(s) whose expression is/are altered in benign vs aggressive tumors. 4. Determine if there is a correlation between specific genetic abnormalities in these tumors (as analyzed by fluorescent in situ hybridization; FISH) and gene expression profiles.
Our overall hypothesis is that there are molecular and biochemical changes that can be used to identify meningiomas that will have a more aggressive clinical course.
Specific Aims 1 and 2: RNA from flash frozen or RNA-later preserved tissue (from all three grades of meningiomas) has been used for RNA isolation using standard protocols. RNA quantity has been determined using a RiboGreen RNA quantitation Kit (Molecular Probes), and RNA quality has been demonstrated using standard formaldehyde gels. These samples will be sent to the NINDS/NIMH microarray consortium for Affymetrix microarray analyses. Data analysis will be done using GeneSpring software (Silicon Genetics, Inc.) with assistance from consortium personnel. Specific Aim 3: Differentially expressed genes identified through microarray analyses will be analyzed using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Real time qRT-PCR is a standard technique used in our laboratory for gene expression analysis. Specific Aim 4: FISH analyses of paraffin-embedded tissue has been completed for 77 tumors. We have frozen tissue from a number of these patients. RNA from these samples will be used for microarray analyses (Specific Aims 1-3). The results of Speicifc Aims 1 and 2 will affect how we perform our correlation analyses. This will be done with the assistance of contracted statistical personnel
Keywords: other

first: Adrienne

middle: C

last: Scheck


title: Alzheimer’s disease: neurofibrillary tangles (Rogers-3U24NS043571-01S1)

status: Public on May 02 2006

summary:
Alzheimer's Disease (AD) is a devastating neurodegenerative disorder affecting approximately 4 million people in the U.S. alone.  AD is characterized by the presence of senile plaques and neurofibrillary tangles in cortical regions of the brain.  These pathological markers are thought to be responsible for the massive cortical neurodegeneration and concomitant loss of memory, reasoning, and often aberrant behaviors that are seen in patients with AD.  Understanding the molecular mechanisms whereby these histopathological markers develop will greatly enhance our understanding of AD development and progression.  A clearer understanding of the mechanisms underlying neurofibrillary tangle formation specifically may help to clarify the basis for dementia of AD as well as the dementias associated with other diseases that are collectively referred to as "tauopathies."
To expression profile both neurons containing neurofibrillary tangles and normal neurons from the entorhinal cortex of 10 mid-stage AD cases.
The gene expression profile of neurons that contain neurofibrillary tangles will differ from the expression profile of histopathologically normal neurons from the same patient and from the same brain region.  Some of these differences will be informative as to the mechanisms of tangle formation.
Keywords: disease-state analysis

overall_design: First use laser capture microdissection to select 1000 neurons bearing neurofibrillary tangles and 1000 normal neurons from the layer 2 stellate island neurons of the entorhinal cortex of 10 mid-stage AD cases.  Second, to isolate the RNA from these captured neurons, perform double round linear amplification and hybridize the labeled cRNA to affymetrix U133A arrays.  Third, to use paired, permutational t-tests to analyze the microarray data to select tangle-specific differences in gene expression.  For the purposes of submitting this proposal, a value of 1 ug of RNA was entered due to web site constraints on values placed in that field.  However, all samples are from LCM captured cells and the actual RNA yield is likely closer to 100 pg.

first: Joseph

last: Rogers


title: Diferential gene expression in Trophoblast cell cultures

status: Public on May 10 2006

city: Rochester

first: Jesus

last: Gonzalez Bosquet

state: MN

address: 200 First Street, SW

department: Gynecologic Oncology

institute: Mayo Clinic

summary:
The syncytiotrophoblast develops as a result of the fusion of placental villous cytotrophoblast cells. These multinuclear cells are essential for fetal-maternal exchange and production of pregnancy-related hormones. Placental deficiency is considered a major risk factor associated with intrauterine growth restriction and preeclampsia. Better understanding of placental patho-physiology requires knowledge on gene expression regulation involved in trophoblast differentiation. Using the well-established in vitro trophoblast differentiation model, we have performed a microarray analysis on mRNA expression in trophoblast and syncytiotrophoblast cell cultures. Dramatic changes in gene expression patterns were detected during trophoblast differentiation. As many as 3524 novel and known genes are found to be up- or down regulated for more than 2-fold. Real-time PCR analysis of a group of genes confirmed the reliability of the microarray data. Overall, this study provided a global view on the molecular events underlying trophoblast differentiation. Subsequent characterization on regulation and function of the identified genes may lead to discovery of new pathways important for placental differentiation and function.
Keywords: Trophoblast culture cells

overall_design: Affymetrix GeneChipTM Human Genome U133 Plus 2.0 microarrays were used for mRNA profiling. RNA samples were subject to Agilent analysis for quality controls. cDNA was prepared from 10 µg of RNA, quantified by spectrometry, and used as a template for the synthesis of biotinylated cRNA using RNA transcript labeling reagent. The quality of the cRNA probe was verified by gel electrophoresis as well as a pilot hybridization using the Test-3 arrays. Hybridization solution containing fragmented cRNA probes and control cRNA was supplemented with herring sperm DNA and bovine serum albumin. The probe solution was heated at 99° C for 5 min followed by incubation at 45° C for 5 min before use. Hybridization was carried out at 45° C for 16 hr with constant rotation at 60 rpm. The arrays were washed and stained with streptavidin-phycoerythrin. The GeneChip 5.0 (Affymetrix) program was used to scan and quantitatively document the hybridization signals. Compilation of candidate gene and calculation of changes were performed on SpotFire and Microsoft Excel programs. To minimize the false-positive conclusion, only genes satisfying the following two criteria were considered candidate regulation targets for further analysis: 1) The gene’s hybridization signal reached an absolute level that was significantly higher than that of the background (P<0.05); 2) The changes is more than 2-fold when compared to control group.


title: Tumor stem cells more closely mirror the phenotype and genotype of primary human tumors than do cancer cell lines

city: Bethesda

first: Howard

last: Fine

state: MD

address: 9030 Old Georgetown Rd

department: Neurological Disorders and Stroke

institute: National Cancer Institute

summary:
The concept of tumor stem cells (TSCs) provides a new paradigm for understanding tumor biology, although it remains unclear whether TSCs will prove to be a more robust model than traditional cancer cell lines. We demonstrate marked phenotypic and genotypic differences between primary human tumor-derived TSCs and their matched glioma cell lines. TSCs derived directly from primary glioblastomas harbor extensive similarities to normal NSC and recapitulate the genotype, gene expression patterns and in vivo biology of human glioblastomas. By contrast, the matched, traditionally grown tumor cell lines do not secondary to in vitro genomic alterations. These findings suggest that TSCs may be a more reliable model than many commonly utilized cancer cell lines for understanding the biology of primary human tumors. Analysis of gene expression data is described in Lee et al., Cancer Cell, 2006.
Keywords: Gene expression profiling of primary GBM tumors and their derivative cells

overall_design: To further understand the differences between NBE- and serum-cultured GBM cells, we generated gene expression profiles of NBE-cells, serum-cells, their derived xenograft tumors, and the original GBMs taken directly from the patients. NBE- and serum-cultured GBM cells from different passages were also included in the analyses in order to evaluate the potential effects of in vitro passage number on gene expression.

first: Jeongwu

last: Lee

first: Svetlana

last: Kotliarova

first: Yuri

last: Kotliarov

first: Aiguo

last: Li

first: Qin

last: Su

first: Nicholas

last: Donin

first: Sandra

last: Pastorino

first: Benjamin

middle: W

last: Purow

first: Neil

last: Christopher

first: Wei

last: Zhang

middle: K

last: Park


title: Molecular phenotyping of human endometrium

status: Public on May 24 2006

city: San Francisco

first: Said

last: Talbi

state: CA

address: 500 Parnassus

institute: UCSF

summary:
To examine the possibility that biochemical or molecular signatures of endometrium may prove to be more useful, we have investigated whole genome molecular phenotyping (54,600 genes/ESTs) of this tissue sampled across the cycle in 28 normo-ovulatory women, using high-density oligonucleotide microarrays. The results demonstrate that endometrial samples obtained by two different sampling techniques (biopsy and curetting hysterectomy specimens) from subjects who are as normal as possible in a human study and 4 including those with unknown histology, can be classified by their molecular signatures and correspond to known phases of the menstrual cycle with identical results using two independent analytical methods. Also, the results enable global identification of biological processes and molecular mechanisms that occur dynamically in the endometrium in the changing steroid hormone milieu across the menstrual cycle in normo-ovulatory women. The results underscore the potential of gene expression profiling for developing molecular diagnostics of endometrial normalcy and abnormalities and identifying molecular targets for therapeutic purposes in endometrial disorders.
Keywords: Gene expression arrays human endometrium

overall_design: Unsupervised principal component analysis (PCA) of all samples revealed that samples selfcluster into four groups consistent with histologic phenotypes of proliferative (PE), early secretory (ESE), mid-secretory (MSE), and late secretory (LSE) endometrium. Independent hierarchical clustering analysis revealed equivalent results, with 2 major dendrogram branches corresponding to PE/ESE and MSE/LSE and sub-branching into the 4 respective phases with heterogeneity among samples within each sub-branch. K-means clustering of genes revealed 4 major patterns of gene expression (high in PE, high in ESE, high in MSE, and high in LSE), and gene ontology analysis of these clusters demonstrated cycle-phase specific biological processes and molecular functions. Six samples with ambiguous histology were identically assignable to a cycle phase by both PCA and hierarchical clustering. Additionally, pair-wise comparisons of relative gene expression across the cycle revealed genes/families that clearly distinguish the transitions of PE to ESE, ESE to MSE, and MSE to LSE, including receptomes and signaling pathways. Select genes were validated by quantitative RT-PCR.

first: Amy

last: Hamilton

first: Kim

last: Vo

first: Suzana

last: TulaC

first: Michael

middle: T

last: Overgaard

first: Christy

last: Dosiou

first: N.

last: Le Shay

first: Nezhat

first: R.

last: Kimpson

first: Bruce

last: Lessey

first: Nihar

last: Nayak

first: Linda

last: Giudice


title: MCF10A cells in 3-D culture after treatment with BRCA1-RNAi or CtIP-RNAi

city: Irvine

first: Wen-Hwa

last: Lee

address: 140 Sprague Hall

department: Biological Chemistry

institute: University of California, Irvine

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

overall_design: Refer to individual Series


title: RSV gene expression

status: Public on May 01 2006

first: Yuh-Chin

middle: Tony

last: Huang

address: Erwin Rd

department: Medicine

institute: Duke University Medical Center

summary:
Human bronchial epithelial cells (BEAS-2B cells) were treated with RSV at 1.0 MOI for 4 and 24 hours or control (vehicle-treated for 4 hours). Global gene expression was measured by Affy Hu133 plus 2.0 microarray chips.
Keywords: Inflammation, microtubules, RSV, time course

overall_design:
4 controls (vehicle-treated for 4 hours)
4 RSV-treated for 4 hours
4 RSV-treated for 24 hours

first: Zhuowei

last: Li

first: Joleen

last: Soukup


title: Gene Expression Profiling of matched Ductal Carcinomas in Situ and Invasive Breast Tumors

city: Tübingen

first: Kay

last: Nieselt

state: DE

address: Sand 14

department: Department for Computer Science

institute: Institute for Bioinformatics and Medical Informatics

summary:
This is a matched-pair analysis of ductal carcinoma in situ (DCIS) and invasive component (IDC) of nine breast ductal carcinoma to identify novel molecular markers characterizing the transition from DCIS to IDC for a better understanding of its molecular biology.
Keywords: Affymetrix-based Microarrays in Mamma carcinoma

overall_design: Absolute, one-channel, oligo-based microarrays used for whole-genome expression profiling of human mamma carcinoma

first: Christina

middle: S

last: Schütz

first: Hans

last: Neubauer


title: Gene expression profiling of human inflammatory Cardiomyopathy

city: Berlin

first: Henning

last: Witt

address: Ihnestrasse 65

department: Hans Lehrach

institute: Max Planck Institute for Molecular Genetics

summary:
Since targeting of specific pathogenic pathways may be more efficient than current unspecific heart failure treatment, we obtained genomewide expression profiles of a DCM subtype characterized by cardiac inflammation (DCMi) in association with parvovirus B19. This study was entirely based on RNA isolated from endomyocardial biopsies so far only rarely used for genomic expression profiling.
Keywords: disease state analysis

overall_design: Samples derived from 8 DCMi and 4 healthy control patients were hybridised onto Affymetrix U133 Plus arrays

first: Frank

last: Wittchen

first: Lennart

last: Suckau

first: Ute

last: Ungethuem

first: Heinz-Peter

last: Schultheiss

first: Wolfgang

last: Poller


title: MCF10A cells in 3-D culture for 15 h after treatment with CtIP-RNAi vs Luc-RNAi adenovirus

summary:
BRCA1 exerts transcriptional repression through interaction with CtIP in the C-terminal BRCT domain and ZBRK1 in the central domain. A dozen of genes including angiopoietin-1 (ANG1), a secreted angiogenic factor, are co-repressed by BRCA1 and CtIP based on microarray analysis of mammary epithelial cells in 3-D culture. BRCA1, CtIP and ZBRK1 form a complex that coordinately represses ANG1 expression via a ZBRK1 recognition site in ANG1 promoter. Impairment of this complex upregulates ANG1, which stabilizes endothelial cells forming capillary-like network structure. Consistently, Brca1-deficient mouse mammary tumors exhibit accelerated growth, pronounced vascularization and overexpressed ANG1. These results suggest, besides its role in maintaining genomic stability, BRCA1 directly regulates the expression of angiogenic factors to modulate the tumor microenvironment.
Keywords: MCF10A, human mammary epithelial cells, 3-D Matrigel, 15 h, CtIP-RNAi

overall_design: MCF10A cells seeded at 5x105 cells/60mm plate were infected with adenoviral luciferase- or CtIP-RNAi in duplicate at 20 MOI for 24h. Infected cells were re-seeded at 5x105 cells in a 60mm plate pre-coated with Growth Factor Reduced Matrigel and covered with the growth medium containing 2% Matrigel at 37°C for 15h. RNA was extracted and quality assessed. cDNA synthesized from the harvested RNA was biotin-labeled, hybridized onto Affymetrix HG U133 PLUS 2.0 array (54,676 genes) and stained with streptavidin-phycoerythrin. The hybridized array was analyzed using GeneChip® Scanner 3000 and GCOS 1.2 software (Affymetrix) at the UCI Microarray Core service.

first: Saori

last: Furuta

first: Ju-Ming

last: Wang

first: Shuanzeng

last: Wei

first: Yung-Ming

last: Jeng

first: Xianzhi

last: Jiang

first: Bingnan

last: Gu

first: Phang-Lang

last: Chen

first: Eva

middle: Y