title: Stable XIAP knockdown in HCT116 colon cancer cells

city: Edinburgh

first: Kathryn

middle: Clare

last: Connolly

address: Crewe Road South

department: Cancer Research Centre

institute: University of Edinburgh

summary: X-linked inhibitor of apoptosis (XIAP) is the most potent endogenous caspase inhibitor preventing cell death via caspase-9, -7 and -3 (initiator and executioner caspase pathways). Using short hairpin RNA (shRNA) against XIAP, stably expressed in a parent HCT116 human colon cancer cell line, a series of clones have been developed. XIAP mRNA levels were established by RT-PCR, the four X (XIAP knockdown) clonal cell lines show 82-93% reduction in XIAP mRNA when compared to the four L (luciferase control) cell lines. Immunoblot analysis showed a 67-89% reduction in XIAP protein in X cell lines compared to L. RNA was analysed by microarray and XIAP knockdown was confirmed in 7 probe sets, there was no significant compensation of other IAP family members. XIAP knockdown induced a 2-fold increase in the basal level of apoptosis without modification of caspase 3/7 activity. Finally, XIAP knockdown sensitises cells to radiotherapy by 20%, to recombinant TRAIL by a 3-fold factor, and to paclitaxel and docetaxel by >2 fold factor. Future work should focus on targeted agents such as rhTRAIL in combination with strategies to down regulate XIAP. XIAP antisense is now in clinical development in oncology.

overall_design: Human HCT116 colon cancer cell lines were transfected with either XIAP (X) or luciferase (L) shRNA.  The expression profiles of both were compared at early (p4) and late (p8) passage.

first: Kate

middle: C

first: Richard

last: Mitter

first: Morwenna

last: Muir

first: Duncan

middle: I

last: Jodwell

first: Sylvie

last: Guichard


title: mRNA Cancer Cell Line Profiles

status: Public on Dec 01 2008

city: X

first: Alex

middle: T.

last: Adai

state: X

address: X

department: Oncology Diagnostics

country: USA

institute: Genentech

summary:
mRNA cancer cell line profiles
Keywords: cancer, cell lines

overall_design: mRNA cancer cell line profiles with samples in either duplicate or triplicate.


title: Gene expression profiling in NCoR deficient mouse livers

status: Public on Dec 04 2008

city: Vienna

first: Martin

last: Bilban

country: Austria

institute: Medical University of Vienna

summary:
The thyroid hormone receptor (TR) has been proposed to regulate target genes in the absence of triiodothyronine (T3), through the recruitment of the corepressors, NCoR and SMRT. NCoR and SMRT may thus play a key role in both hypothyroidism and resistance to thyroid hormone, though this has never been tested in vivo. To accomplish this we developed mice that express in the liver a NCoR protein (L-NCoR∆ID) that cannot interact with the TR. L-NCoR∆ID mice develop normally, however when made hypothyroid the repression of many positively regulated T3-target genes is abrogated, demonstrating that NCoR plays a specific and sufficient role in repression by the unliganded TR. Remarkably, in the euthyroid state, expression of many T3-targets are also upregulated in L-NCoR∆ID mice, demonstrating that NCoR also determines the magnitude of the response to T3 in euthyroid animals. While positive T3 targets were upregulated in L-NCoR∆ID mice in the hypo and euthyroid state there was less effect seen on negatively regulated T3 target genes. Thus, NCoR is a specific regulator of T3-action in vivo and mediates the activity of the unliganded TR. Furthermore, NCoR may play a key role in determining the differences in individual responses to similar levels of circulating T3.
Keywords: NCoR, thyroid hormone signaling, mouse liver,  DNA Microarray

overall_design: To better assess the role of NCoR in positive and negative regulation we performed micorarray analysis of gene expression in the livers of euthyroid and hypothyroid control and L-NCoR∆ID mice.

first: Inna

last: Astapova

first: Tony

middle: H

last: Hollenberg


title: Effect of PTP inhibition on changes in gene expression after Cr(VI) exposure

status: Public on Dec 02 2008

city: Washington

first: DONG-SOON

last: BAE

state: DC

address: 2300 I Street, NW

department: Pharmacology & Physiology

institute: George Washinton Uni.

summary: Although the consequences of genotoxic injury include cell cycle arrest and apoptosis, cell survival responses after genotoxic injury can produce intrinsic death-resistance and contribute to the development of a transformed phenotype. Protein tyrosine phosphatases (PTPs) are integral components of key survival pathways, and are responsible for their inactivation, while PTP inhibition is are often associated with enhanced cell proliferation. Our aim was to elucidate signaling events that modulate cell survival after genotoxin exposure. Diploid human lung fibroblasts (HLF) were treated with Cr(VI) (as Na2CrO4), a well known human respiratory carcinogen that induces a wide spectrum of DNA damage, in the presence and absence of a broad-range PTP inhibitor, sodium orthovanadate. Notably, PTP inhibition abrogated Cr(VI)-induced clonogenic lethality. The enhanced survival of Cr(VI)-exposed cells after PTP inhibition was predominantly due to a bypass of cell cycle arrest and was not due to decreased Cr uptake as evidenced by unchanged Cr-DNA adduct burden. Additionally, the bypass of Cr-induced growth arrest by PTP inhibition, was accompanied by a decrease in Cr(VI)-induced expression of cell cycle inhibiting genes, and an increase in the Cr(VI)-induced expression of cell cycle promoting genes. Importantly, PTP inhibition resulted in an increase in forward mutations at the HPRT locus, supporting the hypothesis that PTP inhibition in the presence of DNA damage may lead to genomic instability, via bypass of cell cycle checkpoints.

overall_design:
Experimental factor was chemical treatment type (3 of them)
(1) No: HLF 1-1, HLF 1-2, HLF 1-3, HLF 1-4
(2) 1uM Cr(VI): HLF 3-1, HLF 3-2, HLF 3-3, HLF 3-4
(3) 10uM SOV + 1uM Cr(VI): HLF 4-1, HLF 4-2, HLF 4-3, HLF 4-4

Biological replicates: 4 different RNA extractions from 4 different cell cultures=quadruplicate per chemical treatment type

first: Dongsoon

last: Bae

first: Tura

last: Camilli

first: Gina

last: Chun

first: Madhu

last: Lal

first: Kristen

last: Wright

first: Travis

middle: J

last: O'Brien

first: Steven

middle: R

last: Patierno

first: Susan

last: Ceryak


title: Expression data from MCF7 wt xenografts

city: Houston

first: Chad

last: Creighton

state: TX

address: One Baylor Plaza, Mail Stop: BCM305

department: Biostatistics, Ducan Cancer Center

institute: Baylor College of Medicine

summary:
To investigate molecular mechanisms of resistance, we used two different in vivo xenograft models of estrogen receptor-positive (ER+) breast cancer, with or without HER2 over-expression (MCF7/HER2-18 and MCF7 wt, respectively). Mice with established tumors were assigned to the following treatment groups: continued estrogen supplementation (E2), estrogen deprivation (ED), ED plus tamoxifen (Tam), all with or without the EGFR tyrosine kinase inhibitor gefinitinib (G).  Another group received ED plus the antiestrogen fulvestrant (MCF7 wt only). Tumors with acquired or de novo resistance to these endocrine therapies were profiled for mRNA expression using Affymetrix Genechip arrays.
Keywords: multiple group comparison

overall_design: MCF7 xenografts were established in ovariectomized five to six week-old nu/nu athymic nude mice supplemented with 0.25 mg 21 day release estrogen pellets by inoculating subcutaneously (s.c.) 5E-6 cells. When tumors reached the size of 150-200 mm3 (3-5 weeks), the animals were randomly allocated to continued estrogen (E2), continued estrogen with gefitinib (E2+G; 100mg/kg, 5 days/week), estrogen withdrawal alone (ED; by removal of the estrogen pellets), and estrogen withdrawal plus tamoxifen citrate (Tam; 500 microg/animal s.c. in peanut oil, 5 days/week), with either gefitinib (Tam+G; 100mg/kg, 5 days/week) or vehicle (1% Tween 80) administered via gavage, as well as estrogen withdrawal plus fulvestrant (ICI 182,780) in the MCF7 wt model (Fulv; 5mg/mouse s.c. once weekly), and estrogen withdrawal with gefitinib (ED+G).  Tumors were harvested for molecular studies when they became resistant to treatment and reached the size of 1000 mm3 (n=7).

first: Suleiman

last: Massarweh

first: Rachel

last: Schiff

first: Kent

last: Osborne


title: Human Fetal Hemoglobin Expression is Regulated by the Developmental Stage-Specific Repressor BCL11A

status: Public on Dec 05 2008

city: Boston

first: Vijay

middle: G.

last: Sankaran

state: MA

address: 300 Longwood Avenue

department: Department of Medicine

institute: Children's Hospital Boston

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: Expression data from MCF7 wt and MCF7/HER2-18 xenografts

summary:
To investigate molecular mechanisms of resistance, we used two different in vivo xenograft models of estrogen receptor-positive (ER+) breast cancer, with or without HER2 over-expression (MCF7/HER2-18 and MCF7 wt, respectively). Mice with established tumors were assigned to the following treatment groups: continued estrogen supplementation (E2), estrogen deprivation (ED), ED plus tamoxifen (Tam), all with or without the EGFR tyrosine kinase inhibitor gefinitinib (G).  Another group received ED plus the antiestrogen fulvestrant (MCF7 wt only). Tumors with acquired or de novo resistance to these endocrine therapies were profiled for mRNA expression using Affymetrix Genechip arrays.

This SuperSeries is composed of the SubSeries listed below.

overall_design:
MCF7 xenografts were established in ovariectomized five to six week-old nu/nu athymic nude mice supplemented with 0.25 mg 21 day release estrogen pellets by inoculating subcutaneously (s.c.) 5E-6 cells. When tumors reached the size of 150-200 mm3 (3-5 weeks), the animals were randomly allocated to continued estrogen (E2), continued estrogen with gefitinib (E2+G; 100mg/kg, 5 days/week), estrogen withdrawal alone (ED; by removal of the estrogen pellets), and estrogen withdrawal plus tamoxifen citrate (Tam; 500 microg/animal s.c. in peanut oil, 5 days/week), with either gefitinib (Tam+G; 100mg/kg, 5 days/week) or vehicle (1% Tween 80) administered via gavage, as well as estrogen withdrawal plus fulvestrant (ICI 182,780) in the MCF7 wt model (Fulv; 5mg/mouse s.c. once weekly), and estrogen withdrawal with gefitinib (ED+G).  Tumors were harvested for molecular studies when they became resistant to treatment and reached the size of 1000 mm3 (n=7).
Refer to individual Series


title: Expression data from MCF7/HER2-18 xenografts


title: Gene expression profiling after RNA interference of CXCR4 in human ovarian cancer cell line IGROV-1.

status: Public on Nov 29 2008

city: London

first: Hagen

last: Kulbe

address: Charterhouse Square

department: Centre for Cancer and Inflammation

country: United Kingdom

institute: Institute of Cancer and the CR-UK Clinical Centre, Barts and The London School of Medicine and Dentistry

summary:
In the past three years the role of inflammatory cytokines and chemokines in tumour promotion and progression has been intensively studied. The chemokine receptor CXCR4 and its ligand CXCL12 are commonly expressed in malignant cells from primary tumours, metastases and also in malignant cell lines. To investigate the biological significance of this receptor/ligand pair, we knocked-down CXCR4 expression in ovarian cancer cell line IGROV-1 using shRNA, and established stable cell lines.
Using Affymetrix microarrays we compared in vitro gene expression in parental IGROV-1 and IGROV-Mock cells with two clones of IGROV-shCXCR4 cells. Gene Set Enrichment Analysis (GSEA) of those genes which were altered by RNA interference of CXCR4 revealed evidence for a cell autonomous signaling network involving CXCR4, TNF-a, IL6 and Notch pathways in ovarian cancer cells.

Keywords: Affymetrix GeneChip Human Genome U133Plus 2.0

overall_design: The Affymetrix GeneChip Human Genome U133Plus 2.0 arrays were used to define gene expression profiles in each cell line.

first: Joseph

last: Kwong


title: SJS blister cells

city: Taipei

first: Wen-Hung

last: Chung

address: 199, Tung-Hwa North Road

department: Dermatology

country: Taiwan

institute: Chang Gung Memorial Hospital

summary:
Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis
(TEN) are life-threatening adverse drug reactions characterized
by massive epidermal necrosis, in which the specific danger
signals involved remain unclear. Here we show that blister
cells from skin lesions of SJS-TEN primarily consist of cytotoxic
T lymphocytes (CTLs) and natural killer (NK) cells, and both
blister fluids and cells were cytotoxic. Gene expression profiling
identified granulysin as the most highly expressed cytotoxic
molecule, confirmed by quantitative PCR and immunohistochemistry.
Granulysin concentrations in the blister fluids
were two to four orders of magnitude higher than perforin,
granzyme B or soluble Fas ligand concentrations, and depleting
granulysin reduced the cytotoxicity. Granulysin in the blister
fluids was a 15-kDa secretory form, and injection of it into
mouse skin resulted in features mimicking SJS-TEN. Our
findings demonstrate that secretory granulysin is a key molecule
responsible for the disseminated keratinocyte death in SJS-TEN
and highlight a mechanism for CTL- or NK cell—mediated
cytotoxicity that does not require direct cellular contact.

overall_design: Blister cells on skin lesion from 5 different patients with SJS/TEN were collected and subjected to RNA extraction without any treatment. Total RNA was isolated using the RNeasy kit (Qiagen). The 28S and 18S ribosomal RNA peak ratios were determined using microfluidics technology (Agilent). RNA was subjected to reverse transcription using the Superscript II kit (Invitrogen), and the cleaned cRNA was then hybridized to an Affymetrix human genome U133 plus 2.0 array.

first: WenHung


title: Prediction of survival in diffuse large B cell lymphoma treated with chemotherapy plus Rituximab

status: Public on Nov 28 2008

city: Bethesda

first: Louis

middle: M.

last: Staudt

state: MD

address: 9000 Rockville Pike, Bldg 10, Rm 4N114

department: Lymphoid Malignancies Branch

institute: National Cancer Institute

summary:
Gene expression profiling of DLBCL patient samples was performed to investigate, whether molecular gene expression signatures retain their prognostic significance in patients treated with chemotherapy plus Rituximab. The lymphnode, germinal center signature and a new angiogenesis signature were combined to a final multivariate model which defined quartile groups among Rituximab-CHOP-treated patients with distinct 3-year overall survival rates.
Keywords: clinical history design

overall_design: The retrospective study included 181 clinical samples from CHOP-treated patients and 233 clinical samples from Rituximab-CHOP-treated patients.


title: Blister cells and PBMCs from patients with SJS-TEN


title: Breast cancer-associated fibroblasts confer AKT1-mediated epigenetic silencing of Cystatin M in epithelial cells.

city: Columbus

first: Sandya

last: Liyanarachchi

state: OH

address: Rm 830, 460, W 12th Avenue

department: Human Genetics

institute: OSU

summary:
Abstract
The interplay between histone modifications and promoter hypermethylation provides a causative explanation for epigenetic gene silencing in cancer. Less is known about the upstream initiators that direct this process. Here, we report that the Cystatin M (CST6) tumor suppressor gene is concurrently down-regulated with other loci in breast epithelial cells co-cultured with cancer-associated fibroblasts (CAFs). Promoter hypermethylation of CST6 is associated with aberrant AKT1 activation in epithelial cells, as well as the disabled INNP4B regulator resulted from the suppression by CAFs. Repressive chromatin, marked by trimethyl-H3K27 and dimethyl-H3K9, and de novo DNA methylation is established at the promoter. The findings suggest that microenvironmental stimuli are triggers in this epigenetic cascade, leading to the long-term silencing of CST6 in breast tumors. Our present findings implicate a causal mechanism defining how tumor stromal fibroblasts support neoplastic progression by manipulating the epigenome of mammary epithelial cells. The result also highlights the importance of direct cell-cell contract between epithelial cells and the surrounding fibroblasts that confer this epigenetic perturbation. Since this two-way interaction is anticipated, the described co-culture system can be used to determine the effect of epithelial factors on fibroblasts in future studies.

overall_design: MCF10A cells were cocultured with different breast fibroblasts that extracted from either breasttumors or normal breast tissues. After three weeks coculture, MCF10A cells were sorted and harvested for total RNA extraction. Affymetrix expression microarray was performed to analyze gene expression changes.  Neither replicates nor dye-swaps was carried out.

first: Huey-Jen

last: Lin

first: Tao

last: Zuo

first: Ching-Hung

first: Chieh

middle: T

last: Kuo

first: Shuying

last: Sun

first: Rulong

last: Shen

first: Daniel

middle: E

last: Deatherage

first: Dustin

last: Potter

first: Lisa

last: Asamoto

first: Shili

first: Pearlly

middle: S

last: Yan

first: Ann-Lii

last: Cheng

first: Michael

first: Tim

last: Huang


title: Genes regulated by AML1/ETO in U937 cells

city: Milan

first: Myriam

last: Alcalay

address: Via Adamello 16

department: Experimental Oncology

country: Italy

institute: European Institute of Oncology

summary:
Approximately 20% of Acute Myelogenous Leukemia (AML) cases carry the t(8;21) translocation, which involves the AML1 and ETO genes, and express the resulting AML1/ETO fusion protein that functions as a transcriptional repressor by recruiting NCoR/SMRT/HDAC complexes to DNA.
We used microarrays to identify genes differentially expressed in U937 cells expressing AML1/ETO compared to vector transfected U937 cells.
Keywords: Transcriptional regulation

overall_design: A U937 cell line that conditionally expresses HA-tagged AML1/ETO under the control of the mouse metallothionine promoter (U937-A1E) (Alcalay et al., J.Clin.Invest, 2003,112, 1751-1761) was used. A cell line caryying the empty vector was used as control Cells were treated for 8h with 100uM ZnSO4 to induce transgene expression. For each of the U937 cell lines (AML1/ETO and Mt), three independent RNA extractions were performed, and an equal quantity of each of the three RNA preparations was then mixed to generate an RNA pool for each sample.


title: Gene expression profile and DNA binding pattern of AML1/ETO in U937 cells

type: Expression profiling by genome tiling array

type: Genome binding/occupancy profiling by genome tiling array


title: CIS (multiple sclerosis) (case-control) (time-series)

status: Public on Nov 26 2008

city: San Francisco

first: Sergio

last: Baranzini

state: CA

address: 513 Parnassus Ave

department: Neurology

institute: University of California San Francisco

summary:
Clinically isolated syndrome (CIS) refers to the earliest clinical manifestation of multiple sclerosis (MS). Currently there are no prognostic biological markers that accurately predict conversion of CIS to clinically definite MS (CDMS). Furthermore, the earliest molecular events in MS are still unknown. We used microarrays to study gene expression in naïve CD4+ T cells from 37 CIS patients at time of diagnosis and after one year. Supervised machine-learning methods were used to build predictive models of disease conversion. We identified 975 genes whose expression segregated CIS patients into 4 distinct subgroups. A subset of 108 genes further discriminated patients from one of these (group#1) from other CIS patients. Remarkably, 92% of patients from group #1 converted to CDMS within 9 months. Consistent downregulation of TOB1, a critical regulator of cell proliferation, was characteristic of group #1 patients. Decreased TOB1 expression at the RNA and protein levels was also confirmed in experimental autoimmune encephalomyelitis (EAE). Finally, a genetic association was observed between TOB1 variation and MS progression in an independent cohort. These results indicate that CIS patients at high risk of conversion have impaired regulation of T cell quiescence resulting in earlier activation of pathogenic CD4+ cells.

Keywords: Disease state: CIS (Clinically Isolated Syndrome - Multiple Sclerosis) versus control. Time series: some patients and controls were resampled approximately one year later

overall_design: Expression data was taken from 37 CIS patients and 28 healthy controls at baseline. 34 CIS patients and 10 healthy controls were resampled at a second time point, approximately one year later. Patients were followed clinically for up to two years to determine the TTC (time to conversion to MS)

first: Corvol

last: Jean-Christophe

first: Baranzini

last: Sergio

first: Oksenberg

last: Jorge


title: Chorionic villus sampling (CVS) microarray in preeclampsia

status: Public on Nov 25 2008

city: Pittsburgh

first: Sandra

middle: Anne

last: Founds

state: PA

address: 448 Victoria Building, 3500 Victoria St.

department: School of Nursing

institute: University of Pittsburgh

summary:
4 chorionic villus sampling specimens in pregnancies destined for preeclampsia and 8 matched controls were analyzed
36 genes were differentially expressed using J5 score and prediction modeling in caGEDA computer program

Keywords: snap frozen, banked CVS specimens with known pregnancy outcomes revealed genes differentially expressed in preeclampsia pregnancies

overall_design: case-control comparisons

middle: A

first: James

middle: F

last: Lyons-Weiler

first: Yvette

middle: P

last: Conley

first: Arundathi

last: Jeyabalan

first: W

last: Hogge

first: Kirk

last: Conrad


title: Expression data from mammary epithelial cells from BRCA1 mutation carriers and non BRCA1 mutation carriers

status: Public on Nov 20 2008

first: Panagiotis

last: Konstantinopoulos

address: 3 Blackfan Circle

department: Hematology/Oncology

institute: Beth Israel Deaconess Medical Center

summary:
Female BRCA1 mutation carriers have a nearly 80% probability of developing breast cancer during their life-time. We hypothesized that the breast epithelium at risk in BRCA1 mutation carriers harbors mammary epithelial cells (MECs) with altered proliferation and differentiation properties.
Microarray studies revealed that PMEC colonies from BRCA1 mutation carriers anticipate expression profiles found in BRCA1-related tumors, and that the EGFR pathway is upregulated in BRCA1 mutation carriers compared ton non BRCA1 mutation carriers.

Keywords: Class comparison and pathway analysis

overall_design: 10 colonies were collected and RNA was isolated using the Absolutely RNA Nanoprep kit, Stratagene. The arrays included duplicates from four normal controls and from two BRCA1 mutation carriers and single arrays from another two BRCA1 mutation carriers.

first: Gerburg

middle: M

last: Wulf


title: Genome wide analysis of gene expression of rat ES cells, rat embryonic fibroblast cells and mouse ES cells

city: Los Angeles

first: Qilong

last: Ying

address: 1501 San Pablo St, ZNI 507

department: Broad Center for Regenerative Medicine and Stem Cell Research

institute: USC

summary:
Rat ES cells were derived using 3I medium from E4.5 blastocysts. Rat embryonic fibroblast cells were derived form E14.5 embryos. To analyze the mechanism under the  selfrenewal of rat ES cells, microarrays were used for the genome wide analysis of gene expressoin profiles in rat ES cells. Rat embryonic fibroblast cells and mouse ES cells were tested at same time as control. Our results from clustering analysis demonstrated that the gene expression profile of rat ES cells resembles mouse ES cells, but not REFs.

Keyword: 3I medium; rat embryonic stem cells; mouse ES cells; rat embryonic fibroblast cells

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus

overall_design: Rat ES cells were cultured in 3I medium; rat embryonic fibroblast cells were derived and cultured GMEM/10% FBS; mouse ES cells (C57/BL6)were cultured in GMEM/10% FBS added LIF and feeder cells were removed before RNA extraction. Three replicates each.

first: Ping

last: Li


title: MCF-7 6A-SA5 and WT-WB1 cells untreated

status: Public on Nov 17 2008

city: Chicago

first: Boris

last: Pasche

state: IL

address: 303 E Superior Lurie 6-220

department: Medicine

institute: Northwestern University

summary:
	TGFBR1*6A is a common hypomorphic variant of the type 1 Transforming Growth Factor Beta Receptor (TGFBR1), which has been associated with increased cancer risk in some studies. Although TGFBR1*6A is capable of switching TGF-β growth inhibitory signals into growth stimulatory signals when stably transfected into MCF-7 breast cancer cells, TGFBR1*6A biological effects are largely unknown. To broadly explore TGFBR1*6A potential oncogenic properties, we assessed its impact on the migration and invasion of MCF-7 cells.  We found that TGFBR1*6A significantly enhances MCF-7 cell migration and invasion in a TGF-β signaling independent manner.  We set up and performed a gene array using the conditions mimicking the cell migration experiments to determine which genes in the migratory pathway were differentially regulated between the MCF-7*6A cells and the MCF-7*9A (wild type transfected) cells.  The gene array identified two downregulated genes in *6A compared to *9A that are involved in cell migration and invasion: ARHGAP5, encoding ARHGAP5, and FN1, encoding fibronectin-1 (FN1).  We were subsequently able to use this information in further studies in the lab.
Keywords: cell type comparison

overall_design: MCF-7 cells were stably transfected with pIRES-TGFBR1-HA-FLAG or pIRES-TGFBR1*6A-HA-FLAG.  Clones were picked up and named 6A-SA5 and WT-WB1 referring to the *6A and TGFBR1 clones, respectively. We have found that the *6A mutation causes an increase in migration and invasion of MCF-7 cells which is independent of TGF-beta.  We used the gene array to identify genes that were differentially regulated between the two cell lines that could lead to this phenotype. We collected RNA from samples that were serum-deprived overnight prior to being fed with complete media to mimic the conditions in the migration experiment.  No exogenous growth factors were added to the media besides the normal 10% heat inactivated FBS.  Samples were collected and run in triplicate (referred to in this data set as sample 1, sample 2, and sample 3).   The “untreated” refers to the fact that samples were grown in complete media alone with no exogenously added growth factors.

first: Diana

last: Rosman

first: Chiang-Ching


title: MKL1 promotes megakaryocytic differentiation in HEL cells

status: Public on Dec 10 2008

city: New Haven

first: Vince

last: Schulz

state: CT

address: 333 Cedar St. LMP 4085

department: Department of Pediatrics

institute: Yale University

summary: Megakaryoblastic Leukemia 1 (MKL1) was identified as part of the t(1;22) translocation specific to acute megakaryoblastic leukemia, but nothing is known regarding its role in hematopoiesis. Here we show that overexpression of MKL1 enhances megakaryocytic differentiation of the Human Erythroleukemia cell line (HEL). Microarray analysis reveals that MKL1 promotes expression of megakaryocyte-specific genes such as glycoprotein V (GP5), as well as cytoskeletal and adhesion molecule genes relevant to megakaryocyte differentiation and proplatelet formation. MKL1 is a transcriptional coactivator of Serum Response Factor. In this study, MKL1 also upregulates known SRF targets. Results provide insight into the role of MKL1 in megakaryocytopoiesis.

overall_design: Analysis of megakaryocytic human erythroleukemia (HEL) cells after 24 hours of TPA induced differentiation with MKL1 overexpression. Gene expression data for control/HEL cells and MKL1/HEL cells, which expresses high levels of MKL1 protein upon doxycycline addition, were determined. TPA, which was added to the samples to induce differentiation, was added together with doxycycline for 24 hours. The TPA differentiated MKL1/HEL samples with or without doxycyline induced MKL overexpression were done in duplicate and were used for both Affymetrix and Illumina platforms.

first: Ee-chun

last: Cheng

first: Qing

last: Luo

first: Emanuela

last: Bruscia

first: Matthew

last: Renda

last: Troy

first: Stephanie

last: Massaro

first: David

last: Tuck

first: Vincent

first: Shrikant

last: Mane

first: Nancy

last: Berliner

first: Yi

last: Sun

first: Stephan

middle: W

last: Morris

first: Caihong

last: Qiu

first: Diane

last: Krause


title: PBMS cells from SJS/TEN

first: Wen-Hung

last: Chung

overall_design: Blood samples were obtained from 5 different patients with SJS/TEN. The peripheral blood mononuclear cells (PBMCs) were isolated from the whole blood samples by Ficoll-Isopaque (Pharmacia Fine Chemicals) density gradient centrifugation. Total RNA from PBMC was isolated using the RNeasy kit (Qiagen). The 28S and 18S ribosomal RNA peak ratios were determined using microfluidics technology (Agilent). RNA was subjected to reverse transcription using the Superscript II kit (Invitrogen), and the cleaned cRNA was then hybridized to an Affymetrix human genome U133 plus 2.0 array.

first: WenHung


title: Gene expression profiling of AML

status: Public on Nov 18 2008

first: Martin

last: Bilban

summary:
AML/MDS patients carrying 11q amplifications involving the mixed lineage leukemia gene (MLL) locus are characterized by a later onset, a complex aberrant karyotype (CAK) frequently including deletions within 5q, 17p and 7q, as well as fast progression of the disease with extremely poor prognosis. We and other have shown that the MLL gene is over expressed in amplified cases, however, in most of the cases the amplified region is not restricted to the MLL locus. In the present study we investigated 19 patients with AML/MDS and MLL gain/amplification [15 AML (two secondary, following MDS and PV, and three therapy related) and 4 MDS cases (two therapy related)]. By means of array CGH performed in 12 patients (GSE9928) we were able to delineate the minimal deleted regions within 5q, 17p and 7q and identified three independent regions 11q/I-III that were amplified in all cases. Gene expression profiles established in 15 cases were used to define the candidate genes within these regions. Interestingly, analysis of our data suggests an interdependency of genes influenced by losses of 5q and 17p and expression of genes present in 11q23-25. Additionally, we demonstrate that the gene expression signature can be used to discriminate AML/MDS with MLL amplification from all other types of AML, thus, indicating specific pathogenesis present in this entity.
Keywords: AML patients, tumor stratification

overall_design:
In this study, gene expression profiling performed for 15 AML patients.
aCGH analysis performed on 12 DNA samples derived from patients with AML, preselected for the presence of MLL amplifications, were analysed on a submegabase resolution BAC array. No replicates, no dye swap was done (GSE9928).

first: Andrea

last: Zatkova

first: Christa

last: Fonatsch


title: Effect of CORM-2 on E. coli transcriptome

status: Public on Dec 17 2008

city: Oeiras

first: Lígia

middle: Santos

last: Nobre

address: Av. da Republica (EAN)

country: Portugal

institute: Instituto de Tecnologia Quimica e Biologica

summary: We recently reported that carbon monoxide (CO) has bactericidal activity. To understand its mode of action we analysed the gene expression changes occurring when Escherichia coli, grown aerobically and anaerobically, is treated with the carbon monoxide releasing molecule, CORM-2. The E. coli microarray analysis shows that E. coli CORM-2 response is multifaceted with a high number of differentially regulated genes spread through several functional categories, namely genes involved in inorganic ion transport and metabolism, regulators, and genes implicated in posttranslational modification, such as chaperones. CORM-2 has higher impact in E. coli cells grown anaerobically, as judged by the existence of repressed genes belonging to eight functional classes which are absent in aerobically CORM-2 treated cells. In spite of the relatively stable nature of the CO molecule, our results show that CO is able to trigger a significant alteration in the transcriptome of E. coli which necessarily has effects in several key metabolic pathways.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Total RNA from two independent cultures of cells grown either aerobically or anaerobically and treated with 250 μM CORM-2 during 15 min or left untreated were processed for use in the Affymetrix GeneChip E. coli Genome 2.0 Arrays, according to the manufacturer’s instructions. Arrays were scanned in an Affymetrix GeneChip scanner 2500 and analysed first with Affymetrix MAS 5.0 software to obtain Absent/Present calls and to assure that all quality parameters were within the recommended range. Hybridization, scanning and detection procedures were done at the Genomics Unit of Instituto Gulbenkian de Ciência (Portugal).


title: Ad-HER-wt and Ad-HER2-ki infected HMECs

status: Public on Nov 15 2008

city: Durham

first: Zachary

middle: Conrad

last: Hartman

state: NC

address: Research Drive MSRB rm 414

department: Surgery

institute: Duke University

summary: As an oncogene, use of HER2 vaccines in humans requires the development of HER2 immunotherapies with maximal immunologic potential, but minimal oncologic potential.  To address these issues, we developed a recombinant adenoviral vector expressing a mutated HER2 inactivated for kinase function (Ad-HER2-ki).  Ad-HER2-ki was highly expressed, but non-phosphorylated and elicited minimal transcription dysregulation in primary cells.  In contrast, Ad-HER2-wt elicited a strong oncogenic signature associated with tumorigenesis.

overall_design: Early Passage Human Mammary Epithelial cells (HMECs) were serum starved for 36hrs. and infected at a MOI of 150 with either Ad-GFP, Ad-HER2-wt, or Ad-HER2-ki vectors.  At 18 hpi, RNA was extracted and transcriptomes evaluated by microarray.  Five samples were infected per virus treatment, each a completely separate biologic replicate.

middle: C

first: Timothy

last: Clay

first: Herbert

middle: K

last: Lyerly


title: Skeletal muscle transcriptome in ICU patients suffering from sepsis induced multiple organ failure

status: Public on Nov 14 2008

city: Stirling

first: Iain

last: Gallagher

address: Room 4B133 Cottrell Building, University of Stirling, Airthrey Road

department: Faculty of Health Sciences and Sport

institute: University of Stirling

summary:
Septic patients treated in the intensive care unit (ICU) often develop multiple organ failure including persistent skeletal muscle dysfunction which results in the patient’s protracted recovery process. We have demonstrated that muscle mitochondrial enzyme activities are impaired in septic ICU patients resulting in decreased cellular energy which will interfere with muscle function and metabolism. Here we use detailed phenotyping and genomics to elucidate mechanisms leading to these impairments.  Methodology/Principle Findings  Utilising biopsy material from seventeen patients and ten age-matched controls we demonstrate that neither mitochondrial in vivo protein synthesis nor expression of mitochondrial genes are compromised. Indeed, there was partial activation of the mitochondrial biogenesis pathway involving NRF2?/GABP and its target genes TFAM, TFB1M and TFB2M yet clearly this failed to maintain mitochondrial function. We therefore utilised transcript profiling and pathway analysis of ICU patient skeletal muscle to generate insight into the molecular defects driving loss of muscle function and metabolic homeostasis. Gene ontology analysis of Affymetrix analysis demonstrated substantial loss of muscle specific genes, a global oxidative stress response related to most probably cytokine signalling, altered insulin related signalling and a substantial overlap between patients and muscle wasting/inflammatory animal models. MicroRNA 21 processing appeared defective suggesting that post-transcriptional protein synthesis regulation is altered by disruption of tissue microRNA expression. Finally, we were able to demonstrate that the phenotype of skeletal muscle in ICU patients is not merely one of inactivity, it appears to be an actively remodelling tissue, influenced by several mediators, all of which may be open to manipulation with the aim to improve clinical outcome.   Conclusions/Significance  This first combined protein and transcriptome based analysis of human skeletal muscle obtained from septic patients demonstrated that losses of mitochondria and muscle mass are accompanied by sustained protein synthesis (anabolic process) while dysregulation of transcription programmes appears to fail to compensate for increased damage and proteolysis. Our analysis identified both validated and novel clinically tractable targets to manipulate these failing processes and pursuit of these could lead to new potential treatments.

Keywords: Disease state analysis

overall_design: 13 septic samples, 8 controls

first: James

last: Timmons

first: Olav

last: Rooyackers


title: Gene Expression Analysis of ARC (NSC 188491) Treated  MCF7 cells

status: Public on Nov 11 2008

city: Frederick

first: Luke

middle: Hunter

last: Stockwin

address: Ft. Detrick

department: DTP

institute: NCI Frederick

summary: ARC (NSC 188491, SMA-491), 4-amino-6-hydrazino-7-beta-d-ribofuranosyl-7H-pyrrolo-(2,3-d)-pyrimidine-5-carboxamide, is a nucleoside analog with profound in vitro anti-cancer activity. First identified in a high-throughput screen for inhibitors of p21 mRNA expression, subsequent experiments showed that ARC also repressed expression of hdm2 and survivin, leading to its classification as a global inhibitor of transcription 1. The following Hu U133 plus 2.0 arrays represent single time point (24 hour) gene expression analysis of transcripts altered by ARC treatment. Arrays for the other compounds (sangivamycin and doxorubicin) are included as comparators.

overall_design: MCF7 cells treated with ARC, Sangivamycin or Doxorubicin for 24 hours.

middle: H

first: Dianne

middle: L

last: Newton


title: Transcriptome comparison of NKp80+ and NKp80- CD8+ CCR7- TCR alpha beta + T cells

status: Public on Nov 12 2008

city: Tübingen

first: Sabrina

last: Kuttruff

address: Auf der Morgenstelle 15

department: of Immunology

country: Germany

institute: University of Tübingen

summary:
We have performed whole genome expression arrays covering over 47000 transcripts comparing the transcriptional profile of NKp80+ to NKp80- CD8+ CCR7- alpha beta T cells. A highly similar global gene expression profile was observed between both memory phenotype T cell subsets. Interestingly, the majority of differentially expressed genes are immune-associated. NKp80+ cells contained markedly increased levels of transcripts encoding for MHC class I and II molecules and for numerous members of the KIR family. Also other NK-related transcripts were more abundantly expressed in the NKp80+ subset. With regards to cytokines, chemokines and their receptors, transcripts important for homeostasis and proliferation are expressed differently. Also transcripts encoding for adhesion molecules are present at different levels in both T cell subsets. Further cytotoxic effector molecules are expressed differently.
Keywords: cell type comparsion

overall_design: We used CD4, CD14 depleted PBMCs pooled from six healthy donors. Following FACS sorting of NKp80+ CD8+ CCR7- alpha beta T cells and NKp80- cells, respectively, transcript levels of both subsets have been compared on human whole genome expression arrays.

first: Alexander

last: Steinle


title: Gene expression profiling in skeletal muscle of PCOS after pioglitazone therapy

status: Public on Nov 10 2008

city: Roskilde

first: Vibe

last: Skov

address: Koegevej 7-13

department: Hematology

country: Denmark

institute: Roskilde Hospital

summary:
Insulin resistance is a common metabolic abnormality in women with PCOS and leads to an elevated risk of type 2 diabetes. Studies have shown that thiazolidinediones (TZD) improve metabolic disturbances in PCOS patients. We hypothesized that the effect of TZD in PCOS is in part mediated by changes in the transcriptional profile of muscle favoring insulin sensitivity.
Using Affymetrix microarrays, we examined the effect of pioglitazone (30 mg/day for 16 weeks) on gene expression in skeletal muscle of 10 obese women with PCOS metabolically characterized by a euglycemic-hyperinsulinemic clamp. Moreover, we explored gene expression changes between these PCOS patients before treatment and 13 healthy control women. Treatment with pioglitazone improved insulin-stimulated total, oxidative and non-oxidative glucose metabolism, and reduced fasting serum insulin (all p < 0.05). Global pathway analysis using Gene Map Annotator and Pathway Profiler (GenMAPP 2.1) and Gene Set Enrichment Analysis (GSEA 2.0.1) revealed a significant upregulation of genes involved in mitochondrial oxidative phosphorylation (OXPHOS), ribosomal proteins, mRNA processing reactome, translation factors, and proteasome complexes in PCOS patients after pioglitazone therapy. Quantitative real-time PCR suggested that upregulation of OXPHOS genes was mediated by an increase in PGC-1α expression (p < 0.05). Expression of genes involved in ribosomal proteins and OXPHOS was down-regulated in PCOS patients before treatment compared to matched healthy women using GenMAPP 2.1 and GSEA 2.1.
These data indicate that pioglitazone therapy restores insulin sensitivity in part by a coordinated upregulation of genes involved in mitochondrial oxidative metabolism and protein biosynthesis in skeletal muscle of PCOS. These transcriptional effects of pioglitazone therapy may contribute to prevent the onset of type 2 diabetes in these women.
Keywords: PCOS, microarray, global pathway analysis, insulin resistance, pioglitazone, protein metabolism, mitochondrial oxidative metabolism

overall_design: Ten obese women of reproductive age with PCOS participated in the study to test the effect of pioglitazone therapy (data set 1). To test if pioglitazone ameliorate existing defects in PCOS patients, the expression profile of the 10 PCOS patients before treatment were compared to the same cohort of 13 control subjects (data set 2).

first: Dorte

last: Glintborg

first: Steen

last: Knudsen

first: Qihua

last: Tan

first: Thomas

last: Jensen

first: Torben

last: Kruse

first: Henning

last: Beck-Nielsen

first: Kurt

last: Højlund


title: FOXF2-regulated genes in human primary prostate stromal cells

status: Public on Nov 01 2008

city: Rotterdam

first: Leonie

last: van der Heul

address: Dr. Molewaterplein 50

country: Netherlands

institute: Erasmus MC

summary: To identify the genes and pathways regulated by FOXF2, we investigated potential FOXF2 gene targets by microarray analyses of primary prostate stromal cells (PrSC) in which FOXF2 was knocked down by siRNA. 190 differentially expressed genes were selected, of which 104 genes were more highly expressed in PrSC cells treated with FOXF2 siRNA and 86 were more highly expressed in PRSC cells treated with negative control siRNA.

overall_design: In each experiment, we compared gene expression of PrSC cells treated with FOXF2 siRNA versus PrSC cells treated with negative control siRNA, in a total of 6 affymetrix arrays. 190 differentially expressed genes were selected (ratio negative control siRNA/siRNA ≥ 2log |0.8| as average in all arrays).

last: van der Heul-Nieuwenhuijsen

first: Guido

last: Jenster


title: Rhinovirus-induced modulation of gene expression in bronchial epithelial cells from subjects with asthma

city: Madison

first: Yury

last: Bochkov

state: WI

address: 600 Highland Ave, K4/945 CSC

department: Pediatrics

institute: University Of Wisconsin-Madison

summary:
Human Rhinovirus (HRV) infection can trigger exacerbations of asthma. Understanding of the mechanisms provoking airway inflammation and remodeling in asthma, as well as the pathogenic mechanisms of HRV infection and its association with asthma exacerbations, may offer significant opportunities for improved disease management. Genome-wide expression analysis of HRV type 1A-infected primary bronchial epithelial (PBE) cells from normal and asthmatic donors was performed to determine whether asthma is associated with a unique pattern of gene expression after HRV infection in vitro.

Keywords: response to rhinovirus infection

overall_design:
Frozen stocks of human PBE cells obtained from the bronchial brushings of six normal and five asthmatic individuals were grown in vitro in Bronchial Epithelial Growth Media (BEGM, Lonza). Subconfluent monolayers of PBE cells were infected with purified and concentrated minor group HRV serotype 1A at multiplicity of infection of 10 plaque forming units (pfu) per cell or mock-infected with growth media alone. Cells were lysed 16 hours post infection (p.i.).
Isolated total RNAs were analyzed using the Affymetrix Human Genome U133 Plus 2.0 GeneChip arrays (Samples S7-S18) or Affymetrix Human HG-Focus arrays samples (Samples S1-S6).
Samples S1-S6 and Samples S7-S18 were normalized by RMA using different methods.

first: Krista

last: Hanson

first: Sunduz

last: Keles

first: Rebecca

last: Brockman-Schneider

last: Gern


title: Differential gene expression profiles are dependent upon method of peripheral blood RNA isolation (direct, heparin)

status: Public on Oct 30 2008

city: Baltimore

first: Zhong

last: Gao

address: Ross 844 720 Rutland Avenue

institute: Johns Hopkins University

summary:
RNA isolation and purification steps greatly influence the results of gene expression profiling. There are two commercially available products for whole blood RNA collection, PAXgene and Tempus blood collection tubes, and each comes with their own RNA purification method. We examined the impact of RNA isolation methods on gene expression profiles.
We demonstrated that peripheral blood RNA isoaltion can critically impact differential expression results, particularly in the clinical setting where fold-change differences are typically small and there is inherent variability within biological corhorts

Keywords: micorarray quality assurance

overall_design: Comparison of 5 healthy control samples drawn directly in PAXgene or Tempus tubes vs. the same 5 healthy control samples drawn in Li Heparin tubes with no PHA stimulation.

first: Adam

last: Asare

first: Svetlana

last: Kolchinsky

first: Richard

last: Wang

first: Khadir

last: Raddassi

first: Katarzyna

last: Bourcier

first: Vicki

last: Seyfert-Margolis


title: Expression data of Hepatocellular Carcinoma

city: Singapore

first: Kun

last: Yu

department: Cellular & Molecular Research

country: Singapore

institute: National Cancer Centre

summary:
Expression profiling of Xenografts of Hepatocellular Carcinoma
Keywords: Human Cancer

overall_design: Briefly, primary HCCs obtained during liver resection were minced into fine fragments capable of passing through an 18-gauge needle, mixed in a 1:1 (v/v) ratio with Matrigel (Collaborative Research, Bedford, MA) and injected in the subcutaneous flanks of 8-week-old male severe combined immune deficient (SCID/SCID, The Jackson Laboratory, Harbor, ME) mice. Six to eight mice were injected for each primary tumor. Growth of established tumor xenografts was monitored twice weekly by vernier caliper measurement of tumor length (a) and width (b), and tumor volume was calculated as (a x b2) / 2. Serial passages of xenograft lines were obtained by dissecting tumors from sacrificed animals and reinjecting the dissociated tumor cells into successive generations of SCID mice as described above. Seven xenograft lines were studied in this report (2-1318, 5-1318(1), 5-1318(3), 2006, 26-1004, 26-1004(cirr), and 30-1004). Xenografts 2-1318 and 5-1318(1) were derived from Hep B-positive patients.

first: Patrick

last: Tan


title: Differential gene expression profiles are dependent upon method of peripheral blood RNA isolation

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

overall_design: Refer to individual Series


title: Differential gene expression profiles are dependent upon method of peripheral blood RNA isolation (PHA)

summary:
RNA isolation and purification steps greatly influence the results of gene expression profiling. There are two commercially available products for whole blood RNA collection, PAXgene and Tempus blood collection tubes, and each comes with their own RNA purification method. We examined the impact of RNA isolation methods on gene expression profiles.
We demonstrated that peripheral blood RNA isoaltion can critically impact differential expression results, particularly in the clinical setting where fold-change differences are typically small and there is inherent variability within biological corhorts

Keywords: micorarray quality assurance, phytohemagglutinin (PHA)

overall_design: Whole blood from 7 healthy individuals was collected using Li Heparin tubes, and was incubated for 3 hours at room temperature with or without addition of 25 ug/mL of PHA. After incubation, samples were subsequently transferred to either Tempus or PAXgene tubes.


title: Identification of genes responsive to hyperthermia in human leukemia U937 cells

city: Toyama

first: Yoshiaki

last: Tabuchi

state: Toyama

address: 2630 Sugitani

department: Life Science Research Center

country: Japan

institute: University of Toyama

summary: Hyperthermia (41˚C <) is widely used to treat patients with various cancers. Here, the effects of hyperthermia (42˚C for 90 min) on the gene expression in human lymphoma U937 cells were investigated using an Affymetrix GeneChip system. The cells were treated with hyperthermia (42°C for 90 min) and followed by incubation for 0, 1, 3 or 6 h at 37°C. The percentage of DNA fragmentation was 7.5 ± 0.9 (mean ± SD), 10.1 ± 0.2, and 17.3 ± 2.3 at the incubation periods of 1, 3, and 6 h, respectively. Of approximately 47,000 probe sets analyzed, the hyperthermia down-regulated 4,214 probe sets and up-regulated 1,334 by a factor 2.0 or greater.

overall_design: U937 cells, a human lymphoma cell line, were treated with hyperthermia (42°C for 90 min) and followed by incubation for 0, 1, 3 or 6 h at 37°C. Non-treated cells were served as control. Total RNA samples were prepared from the cells. Gene expression was analyzed by an Affymetrix GeneChip® system with a Human Genome U133-plus 2.0 array for analysis of over 47,000 transcripts. Sample preparation for array hybridization was carried out as described in the manufacturer’s instructions.

first: Ichiro

last: Takasaki

first: Yukihiro

last: Furusawa

first: Takashi

last: Kondo


title: In vivo study of ADH-1 in combination with melphalan for treatment of melanoma

first: Christina

last: Augustine

address: VAMC RmE4001 508 Fulton St.

institute: Duke University Medical Center

summary:
To identify patterns of change in gene expression that correlated with drug treatment (saline control, ADH-1 alone, melphalan alone and ADH-1 plus melphalan) in our animal model of regional therapy for extremity melanoma we evaluated gene expression using microarray genechips and a Pearson correlation analysis.   Genes were ranked according to the degree to which expression correlated with systemic ADH-1 treatment alone or in combination with regional melphalan.  Tissue samples were harvested at 4, 24 and 48 hours after treatment with melphalan.
Specific cellular pathways associated with drug treatment were identified using gene set enrichment analysis (GSEA).
Keywords: time course; chemotherapy; melanoma; melphalan; N-cadherin; ADH-1

overall_design: Sample size was 2 rats for each drug treatment group at both the 4 and 24 hour time points.  At the 48 hour time point sample size was 1 rat for each drug treatment group.  ADH-1 treatment preceded melphalan by 1 hour.  ADH-1 treatment was repeated at 24 and 48 hrs.  Tissue was harvested for RNA analysis at 4, 24 and 48 hours after completion of isolated limb infusion with melphalan (and 1 hour after an ADH-1 dose).

first: Douglas

last: Tyler


title: Zalypsis: A novel marine-derived compound with potent antimyeloma activity

status: Public on Nov 30 2008

city: Salamanca

first: Enrique

last: Ocio

address: Pº San Vicente, 58-182

country: Spain

institute: Hospital Universitario de Salamanca

summary: Multiple Myeloma (MM) remains incurable, and new drugs with novel mechanisms of action are still needed. In this report, we have analyzed the action of Zalypsis, an alkaloid analogous to certain natural marine compounds, in MM. Zalypsis turned out to be the most potent antimyeloma agent we have tested so far, with IC50s from picomolar to low nanomolar ranges. It also showed remarkable ex vivo potency in plasma cells from patients and in MM cells in vivo xenografted in mice. Besides the induction of apoptosis and cell cycle arrest, Zalypsis provoked DNA double-strand-breaks (DSB), evidenced by an increase in phospho-Histone-H2AX and phospho-CHK2, followed by a striking overexpression of p53 in p53-wild type cell lines. In addition, in those cell lines in which p53 was mutated, Zalypsis also provoked DSB and induced cell death, although higher concentrations were required. Immunohistochemical studies in tumours also demonstrated Histone-H2AX phosphorylation and p53 overexpression. Gene expression profile studies were concordant with these results, revealing an important deregulation of genes involved in DNA-damage response. The potent in vitro and in vivo antimyeloma activity of Zalypsis uncovers the high sensitivity of tumour plasma cells to DSB, and strongly supports the use of this compound in MM patients.

overall_design: MM cells treated in vitro with Zalypsis (5 nM) were harvested at the beginning of induction of cell death (15-20% cell death as assessed by Annexin V-FITC staining). Total RNA was extracted using Trizol reagent (Life Technologies, MD, USA) and purified with RNAeasy Mini Kit (Qiagen, CA, USA). RNA integrity was verified with the Agilent 2100 Bioanalyzer (Agilent, CA, USA). Double-stranded cDNA and biotinylated cRNA were synthesized with T7-polyT primer and the BioArray RNA labeling kit (Enzo, NY, USA), respectively. The labelled RNA was then fragmented and hybridized to HG-U133 Plus 2.0 oligonucleotide arrays (Affymetrix, CA, USA), which were scanned in a Gene Array Scanner and analyzed using the DNA-Chip Analyzer software (DChip). Changes in gene expression greater than two-fold were considered significant.

first: Patricia

last: Maiso

first: Xi

last: Chen

first: Mercedes

last: Garayoa

first: Stela

last: Álvarez-Fernández

first: Laura

last: San-Segundo

first: David

last: Vilanova

first: Lucía

last: López-Corral

first: Juan

last: Montero

first: Teresa

last: Hernández-Iglesias

last: Álava

first: Carlos

last: Galmarini

first: Pablo

last: Avilés

first: Carmen

last: Cuevas

first: Jesús

last: San-Miguel

first: Atanasio

last: Pandiella


title: Gene expression profiling of pulmonary MALT lymphoma

first: Wee

middle: Joo

last: Chng

address: 5 Lower Kent Ridge Road

department: Haematology-Oncology

institute: National University Cancer Institute, Singapore

summary:
Molecular pathways activated in MALT lymphoma are not well defined.
In this study, we perform gene expression profiling on 35 pulmonary MALT lymphoma using fresh frozen biopsy samples

Keywords: Cell type analysis and disease state analysis

overall_design: Total RNA was extracted from fresh frozen tisues of 35 newly diagnosed Pulmonary MALT, as defined by WHO classification, and hybridized to Affymetrix U133plus2 chip. Gene expression profile from other B and T cell malignancies and normal cellular and tissue counterpart were obtained from published dataset for comparison.

first: Ellen

last: Remstein

first: Ahmet

last: Dogan


title: HepG2/C3A cells cultured for 42 h in complete or leucine-devoid medium

city: Ithaca

first: Wei

last: Wang

state: NY

address: T2-003 Vet Res Tower

institute: Cornell University

overall_design: Total of 6 Samples (Leu vs noLeu)

first: Martha

last: Stipanuk

first: Angelos

last: Sikalidis


title: Gene Expression Profiling in A549 Lung Cancer Cell Line Following siRNA Mediated Knock-down of ALDH1A1 and ALDH3A1

status: Public on Oct 29 2008

city: Gainesville

first: M Cecilia

last: Lopez

state: FL

address: 1376 Mowry Road

department: Molecular Genetics and Microbiology

institute: University of Florida

summary:
Aldehyde dehydrogenase isozymes ALDH1A1 and ALDH3A1 are highly expressed in non small cell cell lung cancer. Neither the mechanism nor the biological significance  for such over expression have  been studied.
We used microarrays to analyze changes in A549 lung cancer cell line in which ALDH activity was reduced using  lentiviral mediated expression of siRNA  against both isozymes (Lenti 1+3)
Keywords: Gene Profiling after ALDH Knock Down

overall_design: A549 lung cancer cell lines were transduced with lentiviral vectors containing specific siRNA sequences against ALDH1A1, ALDH3A1, both vectors  (Lenti 1+3 cells), and against the green flourescent protein (GFP) gene (GFP cells, used as control).

first: Jan

last: Moreb

first: Henry

middle: V

last: Baker

first: Lung-Ji

last: Chang

first: Cecilia

last: López

first: Blanca

last: Ostmark

first: Wayne

last: Chou


title: TGF-beta-induced gene expression data and Smad2/3 binding sites of HaCaT keratinocytes

city: Tokyo

first: Daizo

last: Koinuma

address: Hongo 7-3-1, Bunkyo-ku

department: Pathology

institute: University of Tokyo

summary:
Smad2/3 are transcription factors that engage in TGF-beta-induced transcription. Here we analyzed the effect of identified Smad2/3 binding sites to transcription.  We used expression microarrays to compare the Smad2/3 binding sites identified by ChIP-chip to TGF-beta-induced gene expressions. Keywords: time course
We also examined the effect of either ETS1/TFAP2A/SMAD2/SMAD3 siRNAs on TGF-beta-induced gene expression change.

overall_design: HaCaT cells were treated with TGF-beta3 for Smad2/3 ChIP-chip or RNA extraction and hybridization on Affymetrix microarrays.

first: Shuichi

last: Tsutsumi


title: In-transit extremity melanoma I

first: Christina

last: Augustine

summary:
An evaluation of multifocal lesions from patients with in-transit extremity melanoma to determine if all lesions from a patient harbor homogeneous patterns of gene expression

Gene expression profiling studies can help guide treatment for cancer patients by providing tools in the form of gene-expression signatures to characterize a tumor in terms of underlying biology, predicted response to therapy, metastatic progression and/or recurrence.  The utility of gene signatures for defining therapeutic strategies in the treatment of extremity in-transit melanoma will be dependent on the genetic relationship between the multifocal lesions typically present in this disease and the extent to which a single lesion is representative of residual tumor burden.  Using microarray-based gene expression profiling we examined 43 in-transit melanoma lesions across 17 patients with multifocal disease to determine whether one lesion could accurately characterize the underlying biology and genetic profile of a patient's tumor.  Principal component analysis, unsupervised hierarchical clustering, one-way analysis of variance (ANOVA) and gene signatures predictive of chemosensitivity and oncogenic pathway activation showed gene expression patterns to be highly similar (p-values: <0.006; average r = 0.979) between lesions from a single patient but to be significantly different across patients (p<0.05).  These findings demonstrate that individual melanoma tumor nodules in patients with multifocal disease are genetically similar and a single lesion can be used to predict response to chemotherapy, evaluate the activation status of oncogenic signaling pathways and characterize other aspects of the biology of an individual patient's disease.  These results will facilitate the utilization of gene expression profiling in clinical trials of targeted therapy in melanoma allowing for more rational identification of candidates for specific therapies.
Keywords: Disease state analysis

overall_design: Gene expression profiles were obtained from 43 lesions across 17 patients and evaluated using several different algorithms to determine the degree of correlation across multifocal lesions and to evaluate patterns of gene expression that are different across patients

first: Douglas

last: Tyler


title: Expression data from human melanocyte BRAF kinase activated

status: Public on Dec 05 2008

city: Las Vegas

first: Byungwoo

last: Ryu

state: NV

address: 1 Breakthrough Way

institute: Nevada Cancer Institute

summary:
Both benign nevi and melanomas exhibit high rates of activating mutations in BRAF kinase, suggesting that this event is important for melanocyte proliferation and melanoma initiation in vivo; however, the precise mechanism of BRAF kinase action in these lesions remains to be elucidated. We have used comprehensive gene expression profiling to evaluate the downstream effectors of activated BRAFV600E kinase in primary human melanocytes and identified a dominant proliferative target gene signature for BRAF kinase in these cells.
We used microarrays to detail the global gene expression pattern change induced by the activation of RAS/BRAF/MEK/ERK signaling pathway and identified distinct classes of up-regulated genes during this process.

Keywords: Genetic modification

overall_design: Primary human melanocytes were isolated from neonatal foreskins.  Melanocytes isolated from five individual donors were pooled together in order to reduce genetic variations and transduced by BRAFV600E expressing lentivirus and control virus with MOI 10 respectively. Cells were harvested 5 days following infection and total RNA was extracted and purified by RNeasy kit by following manufacturer’s protocol. RNA quality check, double strand complementry DNA synthesis, hybridization with Human Genome U133 Plus 2.0 Array Chips (Affymetrix Inc. Santa Clara, CA), initial data extraction, normalization using RMAExpress software.

first: Rhoda

last: Alani

first: Dave

last: Kim


title: Expression data from human embryonic stem cell differentiation

status: Public on Oct 25 2008

city: Taipei

first: Sung-Liang

last: Yu

address: Jen Ai Road Section1

department: Clinical Laboratory Sciences and Medical Biotechnology

institute: National Taiwan University

summary:
MicroRNAs (miRNAs) are noncoding RNAs of approximately 22 nucleotides in length that usually suppress the translation of target messenger RNAs (mRNAs) through partial complementarity to the 3¡¦ untranslated region (3¡¦ UTR) of protein-coding mRNAs in animals. However, there is increasing evidence that miRNAs can also reduce the steady-state levels of their target mRNAs in animals. In this investigation, both miRNA and mRNA profiles from the undifferentiated human embryonic stem cell line hES-T3 (T3ES), hES-T3 derived embryoid bodies (T3EB) and hES-T3 differentiated fibroblast-like cells (T3DF) were quantitatively determined. Three common target genes of three highly expressed hES cell-specific miRNAs were identified by inverse expression levels of miRNAs to their target mRNAs.
Keywords: cell type comparison

overall_design: Three types of human embryonic stem cells and their derived cells (the undifferentiated human embryonic stem cell line hES-T3 (T3ES), hES-T3 derived embryoid bodies (T3EB) and hES-T3 differentiated fibroblast-like cells (T3DF)) were selected for RNA extraction. The genome-wide mRNA expression of T3ES, T3EB and T3DF cells was determined using Affymetrix human genome U133 plus 2.0 GeneChip. The abundantly expressed genes (10-fold above overall mean) in T3ES, T3EB and T3DF cells are selected to compare with the predicted gene targets of differentially expressed miRNAs in the same set of cells.

first: Steven

last: Li

first: Li-Pin

last: Kao

first: Zong-Yun

last: Tsai

first: Sher

last: Singh

first: Bo-Zhi

last: Chen

first: Pan-Chyr

last: Yang


title: Highly efficient generation of induced pluripotent stem cells from human keratinocytes

status: Public on Oct 27 2008

city: Barcelona

first: Stephanie

last: Boue

address: Dr Aiguader 88

institute: CMRB

summary: The generation of induced pluripotent stem (iPS) cells 1-4 has spawned unprecedented opportunities for investigating the molecular logic that underlies cellular pluripotency and reprogramming, as well as for obtaining patient-specific cells for future clinical applications. However, both prospects are hampered by the low efficiency of the reprogramming process. Here, we show that juvenile human primary keratinocytes can be efficiently reprogrammed to pluripotency by retroviral transduction with Oct4, Sox2, Klf4 and c-Myc. Keratinocyte-derived iPS (KiPS) cells appear indistinguishable from human embryonic stem (hES) cells in colony morphology, growth properties, expression of pluripotency-associated transcription factors and surface markers, as well as in vitro and in vivo differentiation potential. Notably, keratinocyte reprogramming to pluripotency is, at least, 100-fold more efficient and 2-fold faster than that of fibroblasts. This increase in reprogramming efficiency allowed us to expand the practicability of the technology and to generate KiPS cells from single plucked hairs from adult individuals.

overall_design: RNA samples from hES cells, human fibroblasts, human keratinocytes, kiPS cells from human keratinocytes, obtained after infection with 3 or 4 factors. Gene expression of those cells were analyzed.

first: Trond

last: Aasen

first: Angel

last: Raya

first: Maria

middle: J

last: Barrero

first: Elena

last: Garreta

first: Antonella

last: Consiglio

first: Federico

last: Gonzalez

first: Rita

last: Vassena

first: Josipa

last: Bilic

first: Vlad

last: Pekarik

first: Gustavo

last: Tiscornia

first: Michael

last: Edel

first: Stéphanie

last: Boué

first: Juan

last: Izpisua


title: Brief bout of exercise alters gene expression in peripheral blood mononuclear cells of early- and late-pubertal males

status: Public on Oct 31 2008

city: Irvine

first: Shlomit

last: Radom-Aizik

address: 101 Academy

institute: University of California, Irvine

summary:
We compared PBMC genomic response to exercise in both early (EB) and late-pubertal boys (LB)

Keywords: stress response

overall_design: Twenty healthy males (age range 8-18 y/o) participated in this study. A baseline blood sample was taken before the onset of exercise and immediately after 30-min exercise bout. PBMCs were isolated using OptiPrep. Density Gradient Medium (SIGMA). Total RNA was extracted using TRIzol. cRNA hybridized onto Affymetrix U133+2 arrays (total of 40 chips).

first: Szu-Yun

last: Leu

first: Frank

last: Zaldivar

first: Dan

last: Cooper


title: Expression data from Human Saliva Exosomes

status: Public on Nov 07 2008

city: Los Angeles

first: Viswanathan

last: Palanisamy

address: 10833 Le Conte Ave

department: Dental Research Institute

institute: UCLA Dentistry

summary:
Exosomes are molecular entities derived from membrane vesicles of endocytic origin secreted by most cell types. These vesicles are implicated in cell-to-cell communication, deliver proteins and mRNA molecules between cells. Recent studies have shown that exosomes are found in body fluids such as saliva, blood, urine, amniotic fluid, malignant ascites, bronchoalveolar lavage fluid, synovial fluids and breast milk. Exosomes secreted through human saliva contain mRNA may potentially be useful for diagnostic purposes. Although the exact protective mechanism of saliva RNA is a topic of debate, the consensus is that the enrichment of mRNAs in these nano-vesicles in one of the features of the biomarker discoveries. Our aim was to determine if exosomes are present in human saliva and to nano-characterize their transcriptomic content.  Exosomes were purified by differential ultracentrifugation, identified by immunoelectron microscopy, flow cytometry and western blot using a CD-63 antibody. Atomic force microscopy studies revealed ultra structural analysis of both size and density of exosomes. Microarray analysis revealed the presence of 590 mRNA core transcripts are relatively stable inside the exosomes, which can be of saliva mRNA biomarkers.  Exosomal mRNA stability was determined by detergent lyses with treatment of RNase.  Under in vitro conditions fluorescent dye labeled saliva exosomes were able to communicate between human oral keratinocytes studied by using fluorescence microscopy. The RNA from saliva exosomes can transfer their genetic information to human oral keratinocytes and alters gene expression in the new location.  Together, these results suggest that saliva is involved in mRNA trafficking via exosomes, and provides a mechanism for cargoing passenger mRNAs. Our findings are consistent with proposal that exosomes can shuttle RNAs between cells and mRNA is protected inside these vesicles may be a possible resource for biomarker discovery.

Keywords: Human saliva, exosomes, mRNA profiling, gene expression, disease diagnosis

overall_design: Human saliva exosomes were purified through differential centrifugation followed by RNA extraction and hybridization on Affymetrix microarrays. We were able to obtain normal human subjects saliva which are pooled and subjected to ultracentrifugation. The protocol was approved by UCLA Institutional review board. 1 ml of saliva exosomes were used to extract RNA followed by two rounds of amplification by Actorus Amp kit. The amplified RNA was biotin labled and hybridized with Affymetrix protocol.

first: Visu

first: Hui

last: Zhou


title: LSD1 knock down in SY5Y Cells

status: Public on Oct 22 2008

city: Essen

first: Johannes

middle: Hubertus

last: Schulte

state: NRW

department: Peadiatric Hematology and Oncology

institute: University Children's Hospital Essen

summary: To analyze the functional relevance of LSD1 in neuroblastic tumors, SH-SY5Y cells were transiently transfected with siRNA directed against LSD1 or with a scrambled control siRNA. Microarray analysis revealed changes in expression that were consistent with these observations 72 hours after LSD1 knock-down. At this time, 28 genes were significantly induced at least 1.5-fold and 29 genes were significantly repressed at least 1.5-fold. Among the 28 induced genes, 4 are markers of cytoskeletal remodelling (TNS1, TPM1, DNM2, DNAL4), indicating differentiation, and 3 (TPM1, DNM2 and SHANK2) are functionally linked to neurite dynamics and synaptic trafficking. TaqMan quantitative RT-PCR confirmed the expression changes detected via microarray analysis for LSD1, DNAL4, DNM2, TNS1 and TPM1

overall_design: SH-SY5Y cells were transfected with siRNA directed against LSD1 or control siRNA. 2 independent transfection experiments were performed each. Reverse transcription, labelling of total RNA and subsequent hybridization to Affymetrix U133v2 chips were performed according to the manufacturer´s protocols

first: Jutta

last: Krifel

first: Ludger

last: Klein-Hitpaß


title: HL cell line associated gene expression changes compared with CD10+ GC B cells

status: Public on Oct 10 2008

city: Durham

first: Wenbin

last: Wei

address: Stockton Road

department: Biosciences

institute: Durham University

summary:
Gene expression was compared between four B-cell derived HL cell lines (L428, L1236, L591, KMH2) and GC B cells from three different patients.
Keywords: Cell type comparison

first: Martina

last: Vockerodt

first: Susan

last: Morgan

last: Kuo

first: Marilyn

middle: B

last: Chukwuma

first: John

middle: R

last: Arrand

first: Dieter

last: Kube

last: Gordon

first: Lawrence

middle: S

last: Young

first: Ciaran

last: Woodman

first: Paul

middle: G

last: Murray


title: GBM Xenograft response to 1 hour and 8 hour Cilengitide exposure

city: Phoenix

first: Brock

last: Armstrong

state: AZ

address: 445 N. 5th Street

department: Cancer and Cell Biology

institute: TGen

summary: Goal of this experiment is the identify differentially expressed genes in GBM zenografts that have been exposed to Cilengitide for 1 or 8 hours.  A control with no cilengitide is also included.  None of the tumors recieved radiation.

overall_design: 4 replicates of mice brains with tumor 103 are used as a control for expression with no cilengtide exposure.  4 replicates of mice brains with tumor 104 were exposed to cilengitide for 1 hour.  4 replicates of mice brains with tumor 105 were exposed to cilengitide for 8 hours.

first: Tom

last: Mikkelsen

first: Michael

middle: E

last: Berens

middle: A


title: Identification of candidate neuroblastoma genes by combining genomic and expression microarrays: expression data

status: Public on Oct 09 2008

city: Newcastle upon Tyne

middle: Stewart

last: Jackson

address: Central Parkway

department: Institute of Genetic Medicine

institute: Newcastle University

summary:
Gene expression analysis was performed on 30 Neuroblastomas to identify genes whose transcription is significantly altered by recurrent chromosomal alterations. Genomic copy number losses and gains had been delineated in the tumours using FISH and SNP arrays. We have identified genes significantly altered by 7 recurrent alterations: 1p, 3p, 4p, 10q and 11q loss, 2p and 17q gain, and genes co-amplified and over-expressed as a result of MYCN amplification.
Identifying genes whose expression is consistently altered by chromosomal gains or losses is an important step in defining genes of biological relevance in a wide variety of tumour types. However, additional criteria are needed to discriminate further among the large number of candidate genes identified. This is particularly true for neuroblastoma, where multiple genomic copy number changes of proven prognostic value exist. We have used Affymetrix microarrays and a combination of fluorescent in-situ hybridisation and single nucleotide polymorphism (SNP) microarrays to establish expression profiles and delineate copy number alterations in 30 primary neuroblastomas.

Keywords: Disease State Analysis

overall_design: 30 neuroblastomas were obtained from patients of all stages (10 patients - stage 1, 2, 3 or 4s disease, 20 patients - stage 4 disease). RNA samples were extracted and analysed using Affymetrix Human Genome U133 Plus 2.0 Arrays. Patient were treated according to the United Kingdom Children’s Cancer Study Group [UKCCSG], European Neuroblastoma Study Group and Localised Neuroblastoma European Study Group protocols.

first: Maria

last: Lastowska


title: Identification of candidate neuroblastoma genes by combining genomic and expression microarrays

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

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

overall_design: Refer to individual Series


title: Solexa / Illumina deep sequencing of the hippocampal transcriptome of wild-type and DLCK transgenic mice

status: Public on Oct 17 2008

city: Leiden

first: Peter

last: 't Hoen

address: PO Box 9600

department: Center for Human and Clinical Genetics

institute: Leiden University Medical Center

type: Expression profiling by high throughput sequencing

summary:
The hippocampal expression profiles of wild-type mice and mice transgenic for deltaC-doublecortin-like kinase were compared with Solexa/Illumina deep sequencing technology and five different microarray platforms. With Illumina's digital gene expression assay, we obtained approximately 2.4 million sequence tags per sample, their abundance spanning four orders of magnitude. Results were highly reproducible, even across laboratories. With a dedicated Bayesian model, we found differential expression of 3179 transcripts with an estimated false-discovery rate of 8.5%. This is a much higher figure than found for microarrays. The overlap in differentially expressed transcripts found with deep sequencing and microarrays was most significant for Affymetrix. The changes in expression observed by deep sequencing were larger than observed by microarrays or quantitative PCR. Relevant processes such as calmodulin-dependent protein kinase activity and vesicle transport along microtubules were found affected by deep sequencing but not by microarrays. While undetectable by microarrays, antisense transcription was found for 51% of all genes and alternative polyadenylation for 47%. We conclude that deep sequencing provides a major advance in robustness, comparability and richness of expression profiling data and is expected to boost collaborative, comparative and integrative genomics studies.
Keywords: SAGE, 17mer, NlaIII, mRNA expression, transgenic mice

overall_design: 3' SAGE tags of four individual mice per group were sequenced.

first: Yavuz

last: Ariyurek

first: Johan

middle: T

last: den Dunnen


title: LMP1 mediated gene expression changes in transfected CD10+ GC B cells

last: Wei

summary:
In this study, we have investigated the effect of LMP1 on gene expression in normal human GC B cells using a non-viral vector based system
Keywords: transfection of viral oncogene in normal human B cells

overall_design: Gene expression was compared between LMP1-transfected and control vector-transfected GC B cells from three patients. RNA from the FACS-sorted transfected GC B cells was amplified. 10ug of fragmented cRNA was hybridized to HG-U133 Plus 2.0 microarrays. Differentially expressed genes were identified using significance analysis of microarrays (SAM) with a 1.5 fold change threshold and the q-value threshold set to 5%.

last: Vockerodt

last: Morgan


title: Gene expression in human CD4+ T-lymphocytes infected with VSVG-pseudotyped HIV-1 viruses lacking Env, Vpr, and Nef

status: Public on Oct 02 2008

city: Boston

first: Alicja

last: Dabrowska

address: 300 Longwood Ave.

department: Infectious Diseases

institute: Children's Hospital Boston

summary:
The high mutation rate of HIV is linked to the generation of viruses expressing proteins with altered function whose impact on disease progression is unknown. We investigated the effects of HIV-1 viruses lacking Env, Vpr and Nef on CD4+ T cell gene expression using high-density DNA microarray analysis and functional assays.

Keywords: three independent donors

overall_design: Human activated CD4+ T-lymphocytes from three independent donors were infected  with HIV-1 viruses that lack Env and Nef (pNL4-3.eGFP.R+E- or HIVD2GFP) or Env, Vpr and Nef. (pNL4-3.eGFP.R-E- or HIVD3GFP) were pseudotyped with VSVG envelope.  As a control, CD4+ T-lymphocytes were infected with VSVG-pseudotyped eGFP.  CD4+ T-cells were sorted 48 hours after infection using GFP as a marker of infectivity. RNA was isolated 10 hours after sorting, labeled, and prepared for microarray analysis.

first: Nayoung

last: Kim

first: Anna

last: Aldovini


title: Analysis of the MSX1 downstream signaling network in neuroblastoma

status: Public on Oct 15 2008

city: Amsterdam

first: Richard

last: Volckmann

state: Noord-Holland

address: van der Boechorststraat 6a

department: Laboratory of Experimental Oncology and Radiobiology

institute: msterdam UMC and University of Amsterdam

summary:
Neuroblastoma is an embryonal tumour of the peripheral sympathetic nervous system 	(SNS). One of the master regulator genes for peripheral SNS differentiation, the homeobox transcription factor PHOX2B, is mutated in familiar and sporadic neuroblastomas. Here we report that inducible expression of PHOX2B in the neuroblastoma cell line SJNB-8 down-regulates MSX1, a homeobox gene important for embryonic neural crest development. Inducible expression of MSX1 in SJNB-8 caused inhibition of both cell proliferation and colony formation in soft agar. Affymetrix micro-		array and Northern blot analysis demonstrated that MSX1 strongly up-regulated the Delta-Notch pathway. These experiments describe for the first time regulation of the Delta-Notch pathway by MSX1, and connect these genes to the PHOX2B oncogene, indicative of a role in neuroblastoma biology.
Keywords: Delta, Notch, MSX1, neuroblastoma, PHOX2B

overall_design: Time course experiments of two independent clones (K2 and K5), with 2 or 4 	time points, respectively.

first: Ingrid

last: Revet

first: Gerda

last: Huizenga

first: Alvin

last: Chan

first: Jan

last: Koster

middle: v

last: Sluis

last: Øra

first: Rogier

last: Versteeg

first: Dirk

last: Geerts


title: Expression data from short-time cultured primary cells derived from colorectal adenocarcinoma

status: Public on Oct 07 2008

city: Hamburg

first: Annika

last: Sprüssel

address: Orchideenstieg 14

institute: Indivumed GmbH

summary:
Time course comparison to tissue origin and with control cell line HT29 derived from colorectal adenocarcinoma.

Status of expression pattern is different in adenocarcinoma of each patient. Human tumor cells extensively changes their gene- and protein expression patterns during their cultivation, clonal selection and expansion, thereby loosing many of the characteristics of their primary origin. In this study we analyzed if these expression changes could be circumvented by using short-term primary cell culture models derived from colorectal cancer patients.
We compared several primary cells from tumor tissues using a standardized protocol which yielded similar cell populations. For monitoring the gene expression changes induced by cell preparation and cultivation we collected the tissues immediately after resection and isolated cells before seeding, and after 24 and 72 hours of cultivation from each patient.

Keywords: time course

overall_design: Reference tissue, viable primary cells, and cells of cell line HT29 were selected at indicated times, total RNA's were preparated and amplified to biotinylated cRNA for microarray analysis using high-density HG-U133plus 2.0 GeneChips® from Affymetrix.


title: Genome-wide expression analysis of a human pDC cell line

city: New York

first: Babacar

last: Cisse

address: West 168th St HHSC 609

department: Microbiology

institute: Columbia Uinversity

summary:
Analysis of expression profiles of human pDC cell line (CAL1) compared to an immature T cell line (MOLT4)

Keywords: Genome-wide expression analysis

overall_design: Duplicates of both CAL1 and MOLT4 cells used


title: In vitro monocyte-differentiated dendritic cells (moDCs) obtained from APECED patients and healthy controls.

status: Public on Oct 01 2008

city: Tampere

first: Niko

middle: Kristian

last: Sillanpaa

address: Papinkatu 14 A 10

department: Molecular immunology

country: Finland

institute: National Institute of Health, Finland

summary:
Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is a monogenic autoimmune disease caused by mutations in the AIRE gene. AIRE functions as a transcriptional regulator and it has a central role in the development of immunological tolerance. AIRE regulates the expression of ectopic antigens in epithelial cells of the thymic medulla and has been shown to participate in the development of peripheral tolerance. However, the mechanism of action of AIRE has remained elusive. To further investigate the role of AIRE in host immune functions, using microarray technology, we studied the properties and transcript profiles in in vitro monocyte-differentiated dendritic cells (moDCs) obtained from APECED patients and healthy controls.
Keywords: patient vs. healthy control comparative analysis

overall_design: This study includes three female APECED patients between the ages of 26 and 51 years and all patients carried the homozygote FinnMajor mutation (R257X) in the AIRE gene. As controls, we used buffy coats from helathy voluntary blood donors obtained from the Finnish Red Cross Transfusion Service (Helsinki, Finland). The controls were sex- and age-matched. Peripheral blood mononuclear cells (PBMC) were isolated from these subjects. The PBMCs were stimulated to differentiate into mature moDCs. RNA harvested from these cells was labeled and hybridized into microarrays. Part of the cells were treated with candida albicans. RNA from these cells was similarly labeled and studied with mocroarrays.

first: Nora

last: Pontynen

first: Mari

last: Strengell

first: Juha

last: Saharinen

first: Ilkka

last: Julkunen

first: Leena

last: Peltonen


title: Topoisomerase II inhibition involves characteristic chromosomal expression patterns: Trovafloxacin study

city: Hannover

first: Jürgen

last: Borlak

address: Nikolai-Fuchs-Strasse 1

institute: Fraunhofer Institut

summary:
Trovafloxacin is a broad spectrum antibiotic that inhibits the uncoiling of supercoiled DNA in various bacteria by blocking the activity of DNA gyrase and topoisomerase IV. Specific members of this drug family display high activity against eukaryotic type II topoisomerase, as well as cultured mammalian cells and in vivo tumor models. Trovafloxacin seems to have a higher affinity for eukaryotic polymerase II system than the other quinolone agents tested. This effect coupled with other factors, such as an inflammatory response, might result in a hepatotoxic reaction seen with drug.
In this study we found genes regulated by trovafloxacin - induced and repressed - to be located much closer to each other than genes distributed randomly all over the genome (< 100 kbp).

Keywords: Trovafloxacin-treated human hepatocytes versus non-treated human hepatocytes

overall_design: We calculated specifically the probability by which particular genes which were deregulated by the treatment of human hepatocytes with trovafloxacin will occur within DNA windows of different sizes as compared to the probability of all known mapped genes (RefSeq transcripts) of the human genome (NCBI RefSeq 19,360; build 36.2) to occur within the same DNA windows.

first: Susanne

last: Reymann


title: Topoisomerase II inhibition involves characteristic chromosomal expression patterns


title: Topoisomerase II inhibition involves characteristic chromosomal expression patterns: Doxorubicin study

summary:
A variety of important anticancer drugs kill cells by increasing cellular levels of topoisomerase II-DNA cleavage complex. The anthracycline anticancer drug doxorubicin forms a stable ternary complex with DNA and topoisomerase IIa, thereby inhibiting the normal function of the enzyme.
In this study we found genes regulated by doxorubicin - induced and repressed - to be located much closer to each other than genes distributed randomly all over the genome (< 100 kbp).

Keywords: Doxorubicin-treated human hepatocytes versus non-treated human hepatocytes

overall_design: We calculated specifically the probability by which particular genes which were deregulated by the treatment of human hepatocytes with doxorubicin will occur within DNA windows of different sizes as compared to the probability of all known mapped genes (RefSeq transcripts) of the human genome (NCBI RefSeq 19,360; build 36.2) to occur within the same DNA windows.


title: Reversible commitment to differentiation by human multipotent stromal cells in single-cell-derived colonies

city: Temple

first: Joni

last: Ylostalo

state: TX

address: 5701 Airport Rd., Module C

department: Institute for Regenerative Medicine

institute: Texas A&M

summary: Human multipotent stromal cells readily form single-cell-derived colonies when plated at clonal densities. However, the colonies are heterogeneous because cells from a colony form new colonies that vary in size and differentiation potential when replated at clonal densities. The experiments here tested the hypothesis that cells in the inner regions of colonies are partially differentiated, but the differentiation is reversible. Cells were separately isolated from the dense inner (IN) regions and less-dense outer regions (OUT) of single-cell-derived colonies. Cells were then compared by assays of their transcriptomes and proteins, and for clonogenicity and differentiation. IN cells expressed fewer cell-cycle genes and higher levels of genes for extracellular matrix than the OUT cells. When transferred to differentiation medium, differentiation of the colonies occurred primarily in the IN regions. However, the IN cells were indistinguishable from OUT cells when replated at clonal densities and assayed for rates of propagation and clonogenicity. Also, colonies formed by IN cells were similar to colonies formed by OUT cells because they had distinct IN and OUT regions. Cultures of IN and OUT cells remained indistinguishable through multiple passages (30-75 population doublings), and both cells formed colonies that were looser and less dense as they were expanded. The results demonstrated that cells in the IN region of single-cell-derived colonies are partially differentiated, but the differentiation can be reversed by replating the cells at clonal densities.

overall_design: In this study heterogeneity of human multipotent stromal cells in single-cell-derived colonies was examined. Cells were grown as colonies on slides for 12 days and cells from inside (IN) and outside (OUT) of colonies were isolated with laser capture microdissection. Cells from two colonies were pooled for each sample. This study consists of 3 biological replicates for each colony region (3 for IN and 3 for OUT). All 6 samples were run on Affymetrix microarrays after amplification and labeling using Nugen Ovation technology.

first: Nikolay

last: Bazhanov

first: Darwin

last: Prockop


title: Expression data from human PBMCs treated for 6h with the staphylococcal superantigens SEB and SEI

city: Greifswald

first: Sandra

last: Scharf

address: F.-L.-Jahn-Str. 15a

department: Department of Functional Genomics

institute: Ernst Moritz Arndt University Greifswald, Institute for Genetics and Functional Genomics

summary:
The species Staphylococcus (S.) aureus harbors 19 superantigen gene loci, six of which are located in the enterotoxin gene cluster (egc). While these egc superantigens are far more prevalent in clinical S. aureus isolates than non-egc superantigens, they are not a prominent cause of toxic shock. Moreover, neutralizing antibodies against egc superantigens are very rare, even among carriers of egc-positive S. aureus strains. In search of an explanation we have tested two non-exclusive hypotheses: 1) egc and non-egc superantigens have unique intrinsic properties and drive the immune system into different directions; 2) egc and non-egc-superantigens are released by S. aureus under different conditions, which shape the immune response.
A comparison of three egc (SEI, SElM and SElO) and three non-egc superantigens (SEB, SElQ, TSST-1) revealed that both induced proliferation of human PBMC with comparable potency and elicited similar Th1/Th2-cytokine signatures. This was supported by gene expression analysis of PBMC stimulated with one representative superantigen from each group (SEI and SEB). They induced very similar transcriptional changes, especially of inflammation-associated gene networks, corresponding to a very strong Th1- and Th17-dominated immune response.
In contrast, the regulation of superantigen release differed markedly between both superantigen groups. Egc-encoded proteins were secreted by S. aureus during exponential growth, while non-egc superantigens were released in the stationary phase.
We conclude that the distinct biological behavior of egc and non-egc superantigens is not due to their intrinsic properties, which are very similar, but caused by their differential release by S. aureus.

overall_design: PBMCs from 3 healthy blood donors were purified and treated with Medium (control) and recombinant staphylococcal superantigens SEB and SEI. After 6 hours of treatment, total RNA was extracted and hybridized to Affymetrix GeneChip Arrays.

first: Dorothee

last: Grumann

first: Silva

last: Holtfreter

first: Christian

last: Kohler

first: Leif

last: Steil

last: Engelmann

first: Michael

last: Hecker

first: Uwe

last: Völker

first: Barbara

middle: M

last: Bröker


title: Transcriptional signature of IFN-alfa in the side population of ovarian cancer cells

city: Padova

first: Alberto

last: Corradin

state: PD

address: via Tiepolo

department: Dipertimento di Ingegneria dell'Informazione

institute: Università degli Studi di Padova

summary:
The side population (SP), recently identified in several normal tissues and in a variety of tumors, may comprise cells endowed with stem cell features. In this study, we investigated the presence of SP in epithelial ovarian cancer (EOC) and found it in 4 out of 6 primary cultures from xenotransplants, as well as in 9 out of 25 clinical samples analyzed. SP cells from one xenograft bearing a large SP fraction were characterized in detail and they were capable of recreate the full repertoire of cancer cell populations observed in the parent tumor. Moreover, SP cells had higher proliferation rates, were much less apoptotic compared to non-SP cells, and generated tumors more rapidly than non-SP cells.
We also investigated the effects of interferon-alfa (IFN-alpha), a cytokine which has been widely used to treat solid tumors, on EOC cells and observed that IFN-alpha exerts marked anti-proliferative and pro-apoptotic effects on primary cultures containing SP cells. IFN-alfa-treatment invariably caused a dramatic reduction in SP size in tumor cell lines of different origin and in normal bone-marrow SP cells, associated with a distinctive transcriptional profile. Gene therapy with human IFN-alpha resulted in regression of established tumors bearing a large SP fraction, which was not observed when tumors bearing low SP levels were treated. These findings could have relevant  clinical implications since they imply that tumors bearing large SP numbers - albeit rare - could be sensitive to IFN-alpha treatment.

Keywords: response to IFN-alfa

overall_design:
4 Affymetrix microarrays:
2 technical replicates: Ovarian cancer, untreated Side Population, rep1
                              Ovarian cancer, untreated Side Population, rep2
                                      +
2 technical replicates: Ovarian cancer, IFN-alpha treated Side Population, rep1
                               Ovarian cancer, IFN-alpha treated Side Population, rep2

last: Amadori

first: Stefano

last: Indraccolo

first: Lidia

last: Moserle


title: Identification of oxygen-sensitive transcriptional programs in human embryonic stem cells.

city: Columbia

first: Toshihiko

last: Ezashi

state: MO

address: 240a CS Bond Life Sciences Center

department: Animal Sciences

institute: University of Missouri-Columbia

summary:
To realize the full potential of human embryonic stem cells (hESC), it is important to develop culture conditions that maintain hESC in a pluripotent, undifferentiated state. A low O2 atmosphere (~4% O2), for example, prevents spontaneous differentiation and supports self-renewal of hESC. To identify genes whose expression is sensitive to O2 conditions, microarray analysis was performed on RNA from hESC that had been maintained under either 4% or 20% O2.  Of 149 genes differentially expressed, 42 were up-regulated and 107 down-regulated under 20% O2. Several of the down-regulated genes are most likely under the control of hypoxia-inducing factors and include genes encoding enzymes involved in carbohydrate catabolism and cellular redox state. Although genes associated with pluripotency, including OCT4, SOX2 and NANOG were generally unaffected, some genes controlled by these transcription factors, including LEFTY2, showed lowered expression under 20% O2, while a few genes implicated in lineage specification were up-regulated. Although the differences between O2 conditions were generally subtle, they were observed in two different hESC lines and at different passage numbers. The data are consistent with the hypothesis that 4% O2 favors the molecular mechanisms required for the maintenance of pluripotency.
This report emphasizes the importance of employing physiological concentrations of O2 when culturing hESC. The transcript profiles of cultures under 20 % O2 suggest that the cells are more poised to differentiate than when they are under the lower 4 % O2 conditions and that the down-regulation of LEFTY2 under 20 % O2 may destabilize the network of genes maintaining ESC pluripotency. Finally, the association of HIF2A with undifferentiated but not differentiating cells is consistent with a particular role for that transcription factor in control of pluripotency.
Keywords: different oxygen concentrations in hESC culture condition

overall_design: The H1 and H9 hESC lines had been maintained in a physiological O2 (3-5% O2) atmosphere since passage 26 and 23, respectively. The both H1 and H9 cells were propagated under 4% O2 for seven passages on MEF feeder layer, thereafter the both cells were switched to Matrigel coated culture wells for at least three passages prior to RNA collection. RNA extraction was performed within 3 min following removal of the culture plates from respective O2 conditions and well before dissolved O2 in the culture medium showed any detectable changes. Total six RNA samples were collected from H1 cells at two different passage numbers (H1p37 and H1p50) and H9 cells at passage 32 (H9p32) under 4% and 20% O2 conditions, respectively. The cells maintained under 4% O2 were split into both 4% and 20% O2 conditions and cultured for 7 days before of RNA collection.

first: Suzanne

middle: D

last: Westfall