Cancer Stem Cells Are Enriched In The Side-Population Cells In A Mouse Model Of Glioma
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE13490 | GEO | 2008/12/15
SECONDARY ACCESSION(S): PRJNA110169
REPOSITORIES: GEO
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Project description:The recent identification of cancer stem cells (CSCs) in multiple human cancers provides a new inroad to understanding tumorigenesis at the cellular level. CSCs are defined by their characteristics of self-renewal, multipotentiality, and tumor initiation upon transplantation. By testing for these defining characteristics, we provide evidence for the existence of CSCs in a transgenic mouse model of glioma, S100Ã-verbB;Trp53. In this glioma model, CSCs are enriched in the side-population (SP) cells. These SP cells have enhanced tumor-initiating capacity, self-renewal, and multipotentiality compared to non-SP cells from the same tumors. Furthermore, gene expression analysis comparing FACS-sorted cancer SP cells to non-SP cancer cells and normal neural SP cells identified 45 candidate genes that are differentially expressed in glioma stem cells. We validated the expression of two genes from this list (S100a4 and S100a6) in primary mouse gliomas and human glioma samples. Analyses of xenografted human GBM (glioblatoma multiforme) cell lines and primary human glioma tissues show that S100A4 and S100A6 are expressed in a small subset of cancer cells and that their abundance is positively correlated to tumor grade. In conclusion, this study shows that CSCs exist in a mouse glioma model, suggesting that this model can be used to study the molecular and cellular characteristics of CSCs in vivo and to further test the cancer stem cell hypothesis. Experiment Overall Design: This study features two factors, injected cell origin (either tumorsphere or neurosphere) and FACS cell population (either side population or non-side population cells). There were two different tumorspheres, labeled 3447 and 4346 that were isolated from brain tumors in S100beta-verbB;p53-/- or S100beta-verbB;p53+/- mice. The tumorspheres were injected separately into the brains of NOD.Cg-Prkdc<scid>Il2rg<tm1Wjl>/SzJ mice to generate biological triplicates of each primary tumor. Tumorspheres were isolated and cultured before FACS sorting to obtain side population and non-side population cells. As a control, untransformed neurospheres from three independent S100beta-verbB;p53-/- or S100beta-verbB;p53+/- mice were isolated, cultured, and FACS sorted to obtain side population and non-side population cells. Side population and non-side population cells cultured from three mice injected with the 3447 cultured tumorspheres were assayed for gene expression (six samples). Side-population stem cells cultured from three mice injected with the 4346 cultured tumorspheres were assayed for gene expression (three samples). Side-population and non-side population cells cultured from three mice injected with the neurospheres were assayed for gene expression (six samples).
2009-01-13 | E-GEOD-13490 | biostudies-arrayexpress
Project description:In this study, we describe a novel relationship between glioblastoma CSCs and the Notch pathway, which involves the constitutive activation of STAT3 and NF-κB signaling. We demonstrate that adherent glioma CSCs exhibit characteristics previously described for CSCs grown in suspension culture. The expression of CD133, Sox2 and Nestin increased when compared to glioma cells grown in monolayer, and the adherent CSCs were ~100 times more tumorigenic in vivo than monolayer cultured glioma cells. We also found that while the STAT3 and NF-κB signaling pathways are constitutively activated in glioma lines, these pathways are dramatically activated in glioma CSCs. Treatment with STAT3 inhibitors led to a loss of nuclear activation of STAT3 signaling and suppression of growth in both monolayer and CSC conditions. There was a markedly greater growth suppressive effect on glioma CSCs, suggesting that targeted therapy of these key pathways in glioma CSCs may be possible. To further investigate potential biomarkers in glioma CSCs, microarray analysis was performed and revealed deregulation of the Notch signaling pathway. This constitutive activation of STAT3, NF-κB, and Notch pathways in glioma CSCs helps identify novel therapeutic targets for the treatment of glioma. GBM6 cells were continuously maintained as subcutaneous xenografts in NSG mice, and monolayer and CSC cultures were derived from freshly harvested tumor tissue. A total of 6 samples were subjected to microarray analysis, with three biological replicates for each experimental condition.
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Project description:In this study, we describe a novel relationship between glioblastoma CSCs and the Notch pathway, which involves the constitutive activation of STAT3 and NF-κB signaling. We demonstrate that adherent glioma CSCs exhibit characteristics previously described for CSCs grown in suspension culture. The expression of CD133, Sox2 and Nestin increased when compared to glioma cells grown in monolayer, and the adherent CSCs were ~100 times more tumorigenic in vivo than monolayer cultured glioma cells. We also found that while the STAT3 and NF-κB signaling pathways are constitutively activated in glioma lines, these pathways are dramatically activated in glioma CSCs. Treatment with STAT3 inhibitors led to a loss of nuclear activation of STAT3 signaling and suppression of growth in both monolayer and CSC conditions. There was a markedly greater growth suppressive effect on glioma CSCs, suggesting that targeted therapy of these key pathways in glioma CSCs may be possible. To further investigate potential biomarkers in glioma CSCs, microarray analysis was performed and revealed deregulation of the Notch signaling pathway. This constitutive activation of STAT3, NF-κB, and Notch pathways in glioma CSCs helps identify novel therapeutic targets for the treatment of glioma.
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Project description:Novel therapies targeting cancer stem cells (CSCs), which play critical roles in chemo- and radio-resistance, metastasis, and possibly resistance against cancer immunotherapy including granulocyte-macrophage colony-stimulating factor (GM-CSF) gene-transduced tumor cell vaccines, may provide beneficial clinical outcomes. Here, we used syngeneic immunocompetent mice that allowed precise evaluation of the immunogenicity of the side population (SP) isolated from 4T1 murine breast carcinoma (4T1-SP) cells as putative CSCs. 4T1-SP cells showed various stem cell properties including high capacities for colony formation and tumorigenicity as well as high expression of phosphorylated signal transducer and activator of transcription-3 and vascular endothelial growth factor that are inductive of immune tolerance. Despite these progressive malignant characteristics of 4T1-SP cells, subcutaneous injection of non-transmissible Sendai virus-mediated GM-CSF gene-transduced 4T1-SP (4T1-SP/GM) cells remarkably impaired their tumorigenicity compared with that of the controls. This impairment of tumorigenicity was partially dependent on CD8+ T cells in concert with CD4+ T cells and natural killer cells. Notably, therapeutic vaccinations using irradiated 4T1-SP/GM cells markedly suppressed tumor development of subcutaneously transplanted 4T1-SP cells compared with that of the controls including irradiated 4T1-non-SP/GM cells. Tumor suppression was accompanied by robust accumulation of mature dendritic cells at vaccination sites and systemic Th1-based cellular immunity. Moreover, vaccinations comprising primary 4T1-SP cells isolated from transplanted 4T1-SP tumors elicited antitumor effects. cDNA microarray analysis showed that 4T1-SP cells predominantly expressed genes of cancer-related antigens including cancer/testis antigens. Collectively, we demonstrate that SP cell-based vaccinations induce effective antitumor immunity that may improve the efficacy of SP cell-based immunotherapy. Gene expression profiles were compared between sorted 4T1-SP and 4T1-NSP cells.
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Project description:Novel therapies targeting cancer stem cells (CSCs), which play critical roles in chemo- and radio-resistance, metastasis, and possibly resistance against cancer immunotherapy including granulocyte-macrophage colony-stimulating factor (GM-CSF) gene-transduced tumor cell vaccines, may provide beneficial clinical outcomes. Here, we used syngeneic immunocompetent mice that allowed precise evaluation of the immunogenicity of the side population (SP) isolated from 4T1 murine breast carcinoma (4T1-SP) cells as putative CSCs. 4T1-SP cells showed various stem cell properties including high capacities for colony formation and tumorigenicity as well as high expression of phosphorylated signal transducer and activator of transcription-3 and vascular endothelial growth factor that are inductive of immune tolerance. Despite these progressive malignant characteristics of 4T1-SP cells, subcutaneous injection of non-transmissible Sendai virus-mediated GM-CSF gene-transduced 4T1-SP (4T1-SP/GM) cells remarkably impaired their tumorigenicity compared with that of the controls. This impairment of tumorigenicity was partially dependent on CD8+ T cells in concert with CD4+ T cells and natural killer cells. Notably, therapeutic vaccinations using irradiated 4T1-SP/GM cells markedly suppressed tumor development of subcutaneously transplanted 4T1-SP cells compared with that of the controls including irradiated 4T1-non-SP/GM cells. Tumor suppression was accompanied by robust accumulation of mature dendritic cells at vaccination sites and systemic Th1-based cellular immunity. Moreover, vaccinations comprising primary 4T1-SP cells isolated from transplanted 4T1-SP tumors elicited antitumor effects. cDNA microarray analysis showed that 4T1-SP cells predominantly expressed genes of cancer-related antigens including cancer/testis antigens. Collectively, we demonstrate that SP cell-based vaccinations induce effective antitumor immunity that may improve the efficacy of SP cell-based immunotherapy.
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