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SRP057101
Mus musculus
12 Downloadable Samples
Description
Smyd3 is a histone methyltransferase implicated in tumorigenesis. Here we show that Smyd3 expression in mice is required but not sufficient for chemically induced liver and colon cancer formation. In these organs Smyd3 is functioning in the nucleus as a direct transcriptional activator of several key genes involved in cell proliferation, epithelial-mesenchymal transition, JAK/Stat3 oncogenic pathways, as well as of the c-myc and b-catenin oncogenes. Smyd3 specifically interacts with H3K4Me3-modified histone tails and is recruited to the core promoter regions of many but not all active genes. Smyd3 binding density on target genes positively correlates with increased RNA Pol-II density and transcriptional outputs. The results suggest that Smyd3 is an essential transcriptional potentiator of a multitude of cancer-related genes. Overall design: Standard Smyd3-deficient (Smyd3-KO) mice were generated using gene-trap ES cell clones (AS0527 from International Gene Trap Consortium), in which a selection cassette, containing the splice acceptor site from mouse EN2 exon 2 followed by the beta-galactosidase and neomycin resistance gene fusion gene and the SV40 polyadenylation sequence was inserted into the 5th intron of the Smyd3 gene. The resulting mice were devoid of Smyd3 mRNA and protein in all tissues, including liver and colon. For the generation of Smyd3-Tg mice the open reading frame of the mouse Smyd3 cDNA, which contained 3 Flag epitopes at the 3’ end was inserted into the StuI site of the pTTR1-ExV3 plasmid (Yan et al, 1990). The 6.8 kb HindIII fragment containing the mouse transthyretin enhancer/promoter, intron 1, Smyd3 cDNA, three Flag epitopes and SV40 poly-A site was used to microinject C57Bl/6 fertilized oocytes. Founder animals were identified by Southern blotting and crossed with F1 mice to generate lines. Specific overexpression in the liver was tested by RT-PCR analysis in different tissues.
Publication Title
Smyd3 Is a Transcriptional Potentiator of Multiple Cancer-Promoting Genes and Required for Liver and Colon Cancer Development.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 8 Downloadable Samples
- IonTorrentProton
Description
The canonical Wnt pathway plays a central role in stem cell maintenance, differentiation and proliferation in the intestinal epithelium. Constitutive, aberrant activity of the TCF4/ß-catenin transcriptional complex is the primary transforming factor in colorectal cancer. Despite significant recent inroads, the full complement of Wnt target genes and the mechanisms of regulation remain incompletely understood. Here we identify a nuclear long non-coding RNA, termed WiNTRLINC1, as a direct target of TCF4/ß-catenin in colorectal cancer cells. WiNTRLINC1 positively regulates the expression of its close neighbor ASCL2, a transcription factor that controls intestinal stem cell fate. WiNTRLINC1 interacts with TCF4/ß-catenin to mediate the juxtaposition/physical contact of its own promoter with the regulatory regions of ASCL2. ASCL2, in turn, regulates WiNTRLINC1 expression. This feedforward regulatory loop controls stem cell-related gene expression and is highly amplified in colorectal cancer. Overall design: Derivatives of Ls174T colon cancer cells, overexpressing the Tet repressor were used for the construction of inducible overexpressing a shRNA against the WiNTRLINC1 long non coding RNA upon treatment with doxyxycline. siRNAs against WiNTRLINC1 were designed with the siDesign center tool from Dharmacon and their sequences were used for the construction of the shRNA stem loop structure as described in EMBO Rep. 2003 Jun;4(6):609-15. The modified pTER vector was used as a backbone for constructing the shRNA cassette as described in EMBO Rep. 2003 Jun;4(6):609-15. Positive cell clones were screened with RT-PCR in order to validate the efficiency of the knockdown of WiNTRLINC1. The Ls174T derivative cell line inducibly overexpressing a shRNA against ASCL2 has been described previously in Cell. 2009 Mar 6;136(5):903-12. RNA deep sequencing was performed in the WiNTRLINC1 KD and ASCL2 KD cells compared to controls cells in order to detect changes in gene expression due to the loss of either WiNTRLINC1 or ASCL2.
Publication Title
A Positive Regulatory Loop between a Wnt-Regulated Non-coding RNA and ASCL2 Controls Intestinal Stem Cell Fate.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 12 Downloadable Samples
Illumina HumanHT-12 V3.0 expression beadchip
Description
Three HL cell lines (HD-MyZ, L-540 and HDLM-2) were used to investigate the effects of perifosine and sorafenib using in vitro assays analyzing cell growth, cell cycle distribution, gene expression profiling (GEP), and apoptosis. Western blotting (WB) experiments were performed to determine whether the two-drug combination affected MAPK and PI3K/AKT pathways as well as apoptosis. Additionally, the antitumor efficacy and mechanism of action of perifosine/sorafenib combination were investigated in vivo in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice using tumor growth rates and survival as endpoints. RESULTS: While perifosine and sorafenib as single agents exerted a limited activity against HL cells, exposure of HD-MyZ and L-540 cell lines, but not HDLM-2 cells, to perifosine/sorafenib combination resulted in synergistic cell growth inhibition (40% to 80%) and cell cycle arrest. Upon perifosine/sorafenib exposure, L-540 cell line showed significant levels of apoptosis (up to 70%, P .0001) associated with severe mitochondrial dysfunction (cytochrome c, apoptosis-inducing factor release and marked conformational change of Bax accompanied by membrane translocation). Apoptosis induced by perifosine/sorafenib combination did not result in processing of caspase-8, -9, -3, or cleavage of PARP, and was not reversed by the pan-caspase inhibitor Z-VADfmk, supporting a caspase-independent mechanism of cell death. In responsive cell lines, WB analysis showed that antiproliferative and pro-apototic events were associated with dephosphorylation of MAPK and PI3K/Akt pathways. GEP analysis of HD-MyZ and L-540 cell lines, but not HDLM-2 cells indicated that perifosine/sorafenib treatment induced upregulation of genes involved in amino acid metabolism and downregulation of genes regulating cell cycle, DNA replication and cell death. In addition, in responsive cell lines, perifosine/sorafenib combination strikingly induced the expression of tribbles homologues 3 (TRIB3) both in vitro and in vivo. Silencing of TRIB3 prevented cell growth reduction induced by perifosine/sorafenib treatment. In vivo, the combined perifosine/sorafenib treatment significantly increased the median survival of NOD/SCID mice xenografted with HD-MyZ cell line as compared to controls (81 vs 45 days, P .0001) as well as mice receiving perifosine alone (49 days, P .03) or sorafenib alone (54 days, P .007). In mice bearing subcutaneous nodules generated by HD-MyZ and L-540 cell lines but not HDLM-2 cell line, perifosine/sorafenib treatment induced significantly increased levels of apoptosis (2- to 2.5-fold, P .0001) and necrosis (2- to 8-fold, P .0001), as compared to controls or treatment with single agents. In addition, perifosine/sorafenib treatment had no effect on HDLM-2 nodules, but significantly reduced L-540 nodules with 50% tumor growth inhibition, compared to controls. CONCLUSIONS: Perifosine/sorafenib combination resulted in strong anti-HL activity both in vitro and in vivo. These results warrant clinical evaluation of perifosine/sorafenib combined-treatment in HL patients.
Publication Title
Perifosine and sorafenib combination induces mitochondrial cell death and antitumor effects in NOD/SCID mice with Hodgkin lymphoma cell line xenografts.
Sample Metadata Fields
Specimen part, Cell line, Treatment
View Samples- Homo sapiens
- 12 Downloadable Samples
- Illumina HumanHT-12 V3.0 expression beadchip
Description
Three HL cell lines (HD-MyZ, L-540 and HDLM-2) were used to investigate the effects of perifosine and sorafenib using in vitro assays analyzing cell growth, cell cycle distribution, gene expression profiling (GEP), and apoptosis. Western blotting (WB) experiments were performed to determine whether the two-drug combination affected MAPK and PI3K/AKT pathways as well as apoptosis. Additionally, the antitumor efficacy and mechanism of action of perifosine/sorafenib combination were investigated in vivo in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice using tumor growth rates and survival as endpoints. RESULTS: While perifosine and sorafenib as single agents exerted a limited activity against HL cells, exposure of HD-MyZ and L-540 cell lines, but not HDLM-2 cells, to perifosine/sorafenib combination resulted in synergistic cell growth inhibition (40% to 80%) and cell cycle arrest. Upon perifosine/sorafenib exposure, L-540 cell line showed significant levels of apoptosis (up to 70%, P .0001) associated with severe mitochondrial dysfunction (cytochrome c, apoptosis-inducing factor release and marked conformational change of Bax accompanied by membrane translocation). Apoptosis induced by perifosine/sorafenib combination did not result in processing of caspase-8, -9, -3, or cleavage of PARP, and was not reversed by the pan-caspase inhibitor Z-VADfmk, supporting a caspase-independent mechanism of cell death. In responsive cell lines, WB analysis showed that antiproliferative and pro-apototic events were associated with dephosphorylation of MAPK and PI3K/Akt pathways. GEP analysis of HD-MyZ and L-540 cell lines, but not HDLM-2 cells indicated that perifosine/sorafenib treatment induced upregulation of genes involved in amino acid metabolism and downregulation of genes regulating cell cycle, DNA replication and cell death. In addition, in responsive cell lines, perifosine/sorafenib combination strikingly induced the expression of tribbles homologues 3 (TRIB3) both in vitro and in vivo. Silencing of TRIB3 prevented cell growth reduction induced by perifosine/sorafenib treatment. In vivo, the combined perifosine/sorafenib treatment significantly increased the median survival of NOD/SCID mice xenografted with HD-MyZ cell line as compared to controls (81 vs 45 days, P .0001) as well as mice receiving perifosine alone (49 days, P .03) or sorafenib alone (54 days, P .007). In mice bearing subcutaneous nodules generated by HD-MyZ and L-540 cell lines but not HDLM-2 cell line, perifosine/sorafenib treatment induced significantly increased levels of apoptosis (2- to 2.5-fold, P .0001) and necrosis (2- to 8-fold, P .0001), as compared to controls or treatment with single agents. In addition, perifosine/sorafenib treatment had no effect on HDLM-2 nodules, but significantly reduced L-540 nodules with 50% tumor growth inhibition, compared to controls. CONCLUSIONS: Perifosine/sorafenib combination resulted in strong anti-HL activity both in vitro and in vivo. These results warrant clinical evaluation of perifosine/sorafenib combined-treatment in HL patients.
Publication Title
Perifosine and sorafenib combination induces mitochondrial cell death and antitumor effects in NOD/SCID mice with Hodgkin lymphoma cell line xenografts.
Sample Metadata Fields
Specimen part, Cell line, Treatment
View Samples- Homo sapiens
- 12 Downloadable Samples
- Illumina HumanHT-12 V3.0 expression beadchip
Description
Three HL cell lines (HD-MyZ, L-540 and HDLM-2) were used to investigate the effects of perifosine and sorafenib using in vitro assays analyzing cell growth, cell cycle distribution, gene expression profiling (GEP), and apoptosis. Western blotting (WB) experiments were performed to determine whether the two-drug combination affected MAPK and PI3K/AKT pathways as well as apoptosis. Additionally, the antitumor efficacy and mechanism of action of perifosine/sorafenib combination were investigated in vivo in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. While perifosine and sorafenib as single agents exerted a limited activity against HL cells, exposure of HD-MyZ and L-540 cell lines, but not HDLM-2 cells, to perifosine/sorafenib combination resulted in synergistic cell growth inhibition (40% to 80%) and cell cycle arrest. Upon perifosine/sorafenib exposure, L-540 cell line showed significant levels of apoptosis (up to 70%, P .0001) associated with severe mitochondrial dysfunction (cytochrome c, apoptosis-inducing factor release and marked conformational change of Bax accompanied by membrane translocation). Apoptosis induced by perifosine/sorafenib combination did not result in processing of caspase-8, -9, -3, or cleavage of PARP, and was not reversed by the pan-caspase inhibitor Z-VADfmk, supporting a caspase-independent mechanism of cell death. In responsive cell lines, WB analysis showed that antiproliferative and pro-apototic events were associated with dephosphorylation of MAPK and PI3K/Akt pathways. GEP analysis of HD-MyZ and L-540 cell lines, but not HDLM-2 cells indicated that perifosine/sorafenib treatment induced upregulation of genes involved in amino acid metabolism and downregulation of genes regulating cell cycle, DNA replication and cell death. In addition, in responsive cell lines, perifosine/sorafenib combination strikingly induced the expression of tribbles homologues 3 (TRIB3) both in vitro and in vivo. Silencing of TRIB3 prevented cell growth reduction induced by perifosine/sorafenib treatment. In vivo, the combined perifosine/sorafenib treatment significantly increased the median survival of NOD/SCID mice xenografted with HD-MyZ cell line as compared to controls (81 vs 45 days, P .0001) as well as mice receiving perifosine alone (49 days, P .03) or sorafenib alone (54 days, P .007). In mice bearing subcutaneous nodules generated by HD-MyZ and L-540 cell lines but not HDLM-2 cell line, perifosine/sorafenib treatment induced significantly increased levels of apoptosis (2- to 2.5-fold, P .0001) and necrosis (2- to 8-fold, P .0001), as compared to controls or treatment with single agents. Perifosine/sorafenib combination resulted in strong anti-HL activity both in vitro and in vivo. These results warrant clinical evaluation of perifosine/sorafenib combined-treatment in HL patients.
Publication Title
Perifosine and sorafenib combination induces mitochondrial cell death and antitumor effects in NOD/SCID mice with Hodgkin lymphoma cell line xenografts.
Sample Metadata Fields
Specimen part, Cell line, Treatment
View Samples- Mus musculus
- 6 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Junction Adhesion Molecule-A (JAM-A) is present on leukocytes and platelets where it promotes cell adhesion and motility. We are interested in an interaction between JAM-A and tumor progression/metastases. To address this point, we mated JAM-A-/- mice and mouse mammary tumor model MMTV-PyMT mice which, which express polyoma middle T antigen under the control of mouse mammary tumor virus. MMTV-PyMT mice show 100% penetration of mammary tumor and highly metastases to lung. MMTV-PyMT mice without JAM-A show less primary tumor progression, therefore JAM-A enhance primary tumor progression. Then we are addressing the molecular mechanism of this phenomenon by in vivo. Furthermore, we would like to examine JAM-A deficient MMTV tumor signature.
Publication Title
Abrogation of junctional adhesion molecule-A expression induces cell apoptosis and reduces breast cancer progression.
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 4 Downloadable Samples
- Affymetrix Human Gene 1.0 ST Array (hugene10st)
Description
Demethyl fructiculin A is a diterpenoid quinone component of the exudates from Salvia corrugata (SCO-1) leafes. SCO-1 was recently reported to induce anoikis in mammalian cell lines via a molecular mechanism involving the presence of the membrane scavenging receptor CD36. However, experiments performed with cells lacking CD36, showed that SCO-1 was able to induce apoptosis also via alternate pathways. To contribute to a better characterization of the molecular mechanisms underlining the cytotoxic activity of SCO-1, we decided to pursue an unbiased pharmacogenomic approach by generating the gene expression profile of GBM TICs subjected to the administration of SCO-1 and comparing it with that of control cells exposed to the solvent. With this strategy we hypothesized to highlight those pathways and biological processes unlashed by SCO-1.
Publication Title
Demethyl fruticulin A (SCO-1) causes apoptosis by inducing reactive oxygen species in mitochondria.
Sample Metadata Fields
Time
View Samples- Mus musculus
- 6 Downloadable Samples
- Affymetrix Mouse Genome 430A 2.0 Array (mouse430a2)
Description
Astrocyte elevated gene-1 (AEG-1) as a positive inducer of hepatocellular carcinoma (HCC). Transgenic mice with hepatocyte-specific expression of AEG-1 were challenged with N-nitrosodiethylamine (DEN) and developed multinodular HCC with steatotic features. Thus, we have identified the follwoing AEG-1 functions: induction of steatosis, inhibition of senescence and activation of coagulation pathway to augment an aggressive hepatocarcinogenic phenotype.
Publication Title
Astrocyte elevated gene-1 promotes hepatocarcinogenesis: novel insights from a mouse model.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 18 Downloadable Samples
- Illumina HiSeq 2500
Description
AEG-1 is overexpressed in human hepatocellular carcinoma (HCC) and positively regulates development and progression of HCC A conditional hepatocyte-specific knockout mouse was generated by crossing floxed AEG-1 mouse with Alb/Cre mouse in C57BL/6 background Overall design: Livers were harvested from chow-fed 12 wks old AEG-1fl/fl and AEG-1DHEP mice. RNA was extracted and subjected to RNA-Seq.
Publication Title
A novel role of astrocyte elevated gene-1 (AEG-1) in regulating nonalcoholic steatohepatitis (NASH).
Sample Metadata Fields
Age, Specimen part, Subject
View Samples- Mus musculus
- 6 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Neural stem cells from different brain regions show differencies in gene expression patterns and physiological functions.
Publication Title
Innate neural stem cell heterogeneity determines the patterning of glioma formation in children.
Sample Metadata Fields
Sex, Specimen part
View Samples