Transcription profiling by array of human SK-MEL-2 cells infected with Ad-E2F-1 after treatment with doxorubicin
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Hao H
PROVIDER: S-ECPF-GEOD-2715 | biostudies-other | 2006 May-Jun
REPOSITORIES: biostudies-other
Similar Datasets
Project description:SK-MEL-2 cells treated with Ad-E2F-1 (MOI 2), Ad-E2F-1 (MOI 2)plus doxorubicin 0.1uM, or Ad-LacZ (MOI 2) plus doxorubicin 0.1uM Keywords: ordered
2005-06-03 | GSE2715 | GEO
Project description:Melanoma tumor antigen p97 or melanotransferrin (MTf) is an iron (Fe)-binding protein with high homology to serum transferrin. MTf is expressed at very low levels in normal tissues and in high amounts in melanoma cells. The over-expression of MTf in tumor cells was hypothesized to assist rapidly proliferating neoplastic cells with their increased Fe requirements. However, our recent characterization of the MTf knockout (MTf -/-) mouse demonstrated that MTf did not have an essential role in Fe metabolism. To understand the function of MTf, we utilized whole-genome microarray analysis to examine the gene expression profile of five models after modulating MTf expression. These models included two new stably transfected MTf hyper-expression models (SK-N-MC neuroepithelioma and LMTK- fibroblasts) and one cell type (SK-Mel-28 melanoma) where MTf was down-regulated by post-transcriptional gene silencing. These findings were compared to alterations in gene expression identified using the MTf -/- mouse. In addition, the changes identified from the gene array data were also assessed in a new model of MTf down-regulation in SK-Mel-2 melanoma cells. In the cell line models, MTf hyper-expression led to increased cellular proliferation, while MTf down-regulation resulted in decreased proliferation. Across all five models of MTf down- and up-regulation, we identified three genes modulated by MTf expression. These included ATP-binding cassette sub-family B member 5 (Abcb5), whose change in expression mirrored MTf down- or up-regulation. In addition, thiamine triphosphatase (Thtpa) and transcription factor 4 (Tcf4) were inversely expressed relative to MTf levels across all five models. The products of these three genes are involved in membrane transport, thiamine phosphorylation and cell proliferation/survival, respectively. This study identifies novel molecular targets directly or indirectly regulated by MTf and potential pathways involved in its function. These molecular targets could be involved, at least in part, to the role of MTf in modulating proliferation. Experiment Overall Design: To prepare a construct capable of generating hairpin siRNA specific for human MTf mRNA, the expression vector, pSilencer™ 3.1-H1 neo (Ambion, Texas, USA) was used. The vector was used to clone transgene of 66-bp that transcribe 19-mer double-stranded hairpin RNAs of the target gene. The transgene were specifically targeted to positions 2031-2049-bp in the MTf gene (Genbank Accession: NM_005929). Transfections of pS-MTf transgenes or pS-scrambled vectors into human SK-Mel-28 melanoma cells were performed with LipofectamineTM 2000 reagent (Invitrogen, Melbourne, Australia). Total RNA was isolated from the cells using TRIzol Reagent® (Sigma-Aldrich) according to the manufacturer’s protocol. Total RNA from the stably-transfected SK-Mel-28 melanoma cell lines were prepared and hybridized onto Human Genome U133 Plus 2.0 Array. The human GeneChip® U133 Plus 2.0 consists of greater than 47,000 transcripts and variants from over 38,500 well characterized human genes (Affymetrix, Santa Clara, CA).
| S-ECPF-GEOD-6817 | biostudies-other
Project description:We previously identified the induction of growth arrest with phenotypic characteristics of senescence in melanoma cell lines sensitive to diterpene esters, indicating a therapeutic potential. Here we compared the cytostatic effects of two diterpene esters namely TPA (12-O-tetradecanoylphorbol-13-acetate) and PEP008 (20-O-acetyl-ingenol-3-angelate) in sensitive and resistant cell lines derived from melanoma, breast cancer and colon cancer. We showed the diterpene esters to induce senescence-like growth arrest in the sensitive cells at 100-1000 ng/ml. Use of the pan-PKC inhibitor bisindolylmaleimide-l demonstrated that activation of PKC was required for growth arrest. Full genome expression profiling revealed that pivotal genes involved in DNA synthesis and cell cycle control were down-regulated by treatment in all three sensitive solid tumor models. At the protein level, prolonged down-regulation of E2F-1 and proliferating cell nuclear antigen (PCNA), sustained expression of p21WAF1/CIP1 and dephosphorylation of retinoblastoma (Rb) occurred in the sensitive cells. Although activation of extracellular signal-related kinase (ERK) 1/2 by the diterpene esters occurred in both sensitive and resistant cell lines, the HRASLS3 type II tumor suppressor, which appears to have a role in MAPK pathway suppression, was constitutively elevated in the resistant cell lines compared to their sensitive counterparts. Together, these results demonstrate the ability of the PKC activating drugs TPA and PEP008 to induce growth arrest with characteristics of senescence in solid tumor cell lines derived from a variety of tissue types through a similar mechanism. PKC-activating diterpene esters may therefore have therapeutic potential in a range of solid tumors. Experiment Overall Design: We analyzed the transcriptional profiles of the diterpene ester sensitive cell lines MCF7, COLO-205 and SK-MEL-5 following treatment with PEP008 using full genome expression profiling (Affymetrix, U133 Plus 2.0). Cells were treated for 24 h and 24 h plus 72 h recovery with 1000 ng/ml of the drug, before harvesting RNA for analysis. From the cell growth assays, all three cell lines demonstrated permanent growth arrest with diterpene ester treatments at the 1000 ng/ml dose. Mock controls were treated with solvent alone for 24 h. SK-MEL-5 cells were also treated with 1000 ng/ml TPA for 24 h.
| S-ECPF-GEOD-8742 | biostudies-other
Project description:More than two thirds of breast cancers express the estrogen receptor (ER) and depend on estrogen for growth and survival. Therapies targeting ER function including aromatase inhibitors that block the production of estrogens and ER antagonists that alter ER transcriptional activity play a central role in the treatment of ER+ breast cancers of all stages. In contrast to ER- breast cancers, which frequently harbor mutations in the p53 tumor suppressor, ER+ breast cancers are predominantly wild type for p53. Despite harboring wild type p53, ER+ breast cancer cells are resistant to chemotherapy-induced apoptosis in the presence of estrogen. Using genome-wide approaches we have addressed the mechanism by which ER antagonizes the pro-apoptotic function of p53. Interestingly both ER agonists such as estradiol and selective ER modulators (SERM) such as tamoxifen promote p53 antagonism. In contrast the full ER antagonist fulvestrant blocks the ability of ER to inhibit p53-mediated cell death. This suggests an improved strategy for the treatment of ER+ breast cancer utilizing antagonists that completely block ER action together with drugs that activate p53-mediated cell death. MCF7 cells were hormone-depleted for 3 days and then treated with 10 uM doxorubicin for 12 hours
| S-ECPF-GEOD-39870 | biostudies-other
Project description:Summary: Cancer imparts an increased risk for venous thromboembolism (VTE). Direct mechanisms linking VTE with malignancy remain unknown. Our characterization of Doxorubicin (DOX)-resistant MCF7 breast cancer cells revealed links with antithrombotic pathways, as an anticoagulant transcriptional profile was observed. Specifically, the intracellular anticoagulant Tissue Factor Pathway Inhibitor 1 (TFPI1) protein was elevated and functional in all resistant models tested; TFPI1 accumulation coincided with reduced thrombin. TFPI1 overexpression in unselected cells elevated pro-malignant gene markers and increased DOX resistance. Furthermore, TFPI1 and Breast Cancer Resistant Protein (BCRP) levels increased early and remained sustained during selection. Finally, consistent with antithrombin-mediated hypoxic metastasis, Hypoxia-Inducible Factor 1 alpha (HIF1alpha) was elevated in drug resistant and TFPI1 overexpressing cells. Thus, increased TFPI1 may create an intratumoral hypoxic environment, potentially leading to drug resistance. MCF7 cells were selected for DOX resistance by first treating the cells with 1 microM DOX for 48 hours. The cells were then exposed to 100 nM DOX for 2 weeks. After this period, the surviving cells were resistant to further DOX treatment.
| S-ECPF-MTAB-1643 | biostudies-other
Project description:SK-MEL-2 cells treated with Ad-E2F-1 (MOI 2), Ad-E2F-1 (MOI 2)plus doxorubicin 0.1uM, or Ad-LacZ (MOI 2) plus doxorubicin 0.1uM. The combination of E2F-1 gene therapy and chemotherapy produces a synergistic effect on melanoma cell apoptosis. However, the molecular mechanisms have not been fully elucidated. The purpose of this study was to identify novel genes or pathways that may play key roles in apoptosis when E2F-1 gene therapy is combined with doxorubicin chemotherapy. MATERIALS AND METHODS: SK-MEL-2 melanoma cells were infected with Ad-E2F-1 alone, Ad-E2F-1 plus doxorubicin, or Ad-LacZ plus doxorubicin. After 16 hours of treatment, the total RNA was extracted from these cells and subjected to microarray analysis. Quantitative real-time PCR was performed to confirm the microarray data. RESULTS: Our results showed that the combination treatment of Ad-E2F-1 and doxorubicin affected the expression of cytokines, transcription factors, as well as genes involved in signal transduction, cell cycle regulation and apoptosis. CONCLUSION: Our findings have identified, for the first time, novel molecular targets and pathways that led to apoptosis in melanoma cells when Ad-E2F-1 was combined with doxorubicin. The molecular information provided here will enhance further mechanistic studies.
2007-10-27 | E-GEOD-2715 | biostudies-arrayexpress
Project description:"Bmi-1 and Mel-18 are close structural homologues that belong to the polycomb group (PcG) of transcriptional regulators of homeotic gene expression in development. They are believed to stably maintain repression of gene expression by altering the state of chromatin at specific promoters. A number of clinical and experimental observations have also implicated Bmi-1 in tumorigenesis and stem cell maintenance. Bmi-1 overexpression or amplification has been observed in a number of human malignancies, particularly in B-cell lymphomas, medulloblastomas and breast cancer. We report here that shRNA-mediated knock-down of either Bmi-1 or Mel-18 in human medulloblastoma DAOY cells results in the inhibition of proliferation, loss of clonogenic survival and anchorage-independent growth, and suppression of xenograft tumor formation in nude mice. Furthermore, overexpression of both Bmi-1 and Mel-18 significantly increased clonogenic survival of Rat1 fibroblasts. In contrast, stable downregulation of Bmi-1 or Mel-18 alone did not affect the growth of SK-OV-3 or U2OS cancer cell lines or normal human WI38 fibroblasts. Gene expression analysis of shRNA-expressing DAOY cells has demonstrated a significant overlap in the Bmi-1- and Mel-18-regulated genes and revealed novel gene targets under their control. Taken together, these results suggest that Bmi-1 and Mel-18 might have overlapping functions in human tumorigenesis. Experiment Overall Design: DAOY cells were transduced with the indicated lentiviral shRNA and stable cell lines were generated by selection in puromycin (1 µg/ml). 2 x 105 cells of each stable cell line were plated onto 60 mm dishes in triplicates and harvested 4 days later. Triplicate total RNA samples from shRNA-expressing and mock-transduced DAOY cells were isolated using affinity resin (Qiagen RNeasy Mini Kit, Qiagen AG). RNA Integrity and purity were assessed with the RNA 6000 Nano LabChip system on Bioanalyzer 2100 (Agilent Technologies). Gene array analysis was conducted at the Genomics Factory at Novartis PHARMA AG, Basel, Switzerland using Gene Chip Human Genome 133 2.0 Plus Expression Array (Affymetrix Inc.)."
| S-ECPF-GEOD-7578 | biostudies-other
Project description:Melanoma is a highly aggressive type of skin cancer produced through the malignant transformation of melanocytes, and it is usually associated with a poor prognosis. Clinically, melanoma has several stages associated with migration and invasion of the cells through the skin's layers, the rapid spreading of cells and the formation of tumors in multiple organs. The main problem is the emergence of resistance in melanoma to the applied methods of treatment; thus, it is of primary importance to find more crucial signaling pathways that control the progression of this type of cancer and could be targeted to prevent melanoma spreading. Here, we uncover novel aspects of the role of the mechanosensitive ion channel Piezo1 in melanoma tumor formation. Using a combinative approach, we showed the functional expression of mechanosensitive Piezo1 channels in the aggressive human melanoma SK-MEL-2 cell line. We found that chemical activation of Piezo1 by its agonist, Yoda1, prevents melanoma spheroid formation; thus, Piezo1 could be a potential target for selective modulation aimed at the prevention of melanoma development.
| S-EPMC10648948 | biostudies-literature
Project description:Melanoma tumor antigen p97 or melanotransferrin (MTf) is an iron (Fe)-binding protein with high homology to serum transferrin. MTf is expressed at very low levels in normal tissues and in high amounts in melanoma cells. The over-expression of MTf in tumor cells was hypothesized to assist rapidly proliferating neoplastic cells with their increased Fe requirements. However, our recent characterization of the MTf knockout (MTf -/-) mouse demonstrated that MTf did not have an essential role in Fe metabolism. To understand the function of MTf, we utilized whole-genome microarray analysis to examine the gene expression profile of five models after modulating MTf expression. These models included two new stably transfected MTf hyper-expression models (SK-N-MC neuroepithelioma and LMTK- fibroblasts) and one cell type (SK-Mel-28 melanoma) where MTf was down-regulated by post-transcriptional gene silencing. These findings were compared to alterations in gene expression identified using the MTf -/- mouse. In addition, the changes identified from the gene array data were also assessed in a new model of MTf down-regulation in SK-Mel-2 melanoma cells. In the cell line models, MTf hyper-expression led to increased cellular proliferation, while MTf down-regulation resulted in decreased proliferation. Across all five models of MTf down- and up-regulation, we identified three genes modulated by MTf expression. These included ATP-binding cassette sub-family B member 5 (Abcb5), whose change in expression mirrored MTf down- or up-regulation. In addition, thiamine triphosphatase (Thtpa) and transcription factor 4 (Tcf4) were inversely expressed relative to MTf levels across all five models. The products of these three genes are involved in membrane transport, thiamine phosphorylation and cell proliferation/survival, respectively. This study identifies novel molecular targets directly or indirectly regulated by MTf and potential pathways involved in its function. These molecular targets could be involved, at least in part, to the role of MTf in modulating proliferation. Experiment Overall Design: To prepare a construct capable of generating hairpin siRNA specific for human MTf mRNA, the expression vector, pSilencer⢠3.1-H1 neo (Ambion, Texas, USA) was used. The vector was used to clone transgene of 66-bp that transcribe 19-mer double-stranded hairpin RNAs of the target gene. The transgene were specifically targeted to positions 2031-2049-bp in the MTf gene (Genbank Accession: NM_005929). Transfections of pS-MTf transgenes or pS-scrambled vectors into human SK-Mel-28 melanoma cells were performed with LipofectamineTM 2000 reagent (Invitrogen, Melbourne, Australia). Total RNA was isolated from the cells using TRIzol Reagent® (Sigma-Aldrich) according to the manufacturerâs protocol. Total RNA from the stably-transfected SK-Mel-28 melanoma cell lines were prepared and hybridized onto Human Genome U133 Plus 2.0 Array. The human GeneChip® U133 Plus 2.0 consists of greater than 47,000 transcripts and variants from over 38,500 well characterized human genes (Affymetrix, Santa Clara, CA).
2008-06-15 | E-GEOD-6817 | biostudies-arrayexpress
Project description:A total of 79 breast tumors (71 malignant tumors, 6 fibroadenomas, and 2 DCIS) were studied and compared to the reduction mammoplastic specimens of 12 healthy women as well as adjacent cancer-free tissue for 23 of the malignant tumors. Polycomb Group (PcG) proteins Bmi-1 and Mel-18 were then studied more closely first on mRNA expression level using microarray data from two of our ongoing projects and then on protein expression level using immunohistochemistry.
| S-ECPF-MTAB-271 | biostudies-other