Isolation and characterization of stem-like cells from a human ovarian cancer cell line
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Roman Mezencev
PROVIDER: E-GEOD-28799 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Increasing evidence supports the existence of a subpopulation of cancer cells capable of self-renewal and differentiation into diverse cell lineages. These cancer stem-like or cancer-initiating cells (CICs) also demonstrate resistance to chemo- and radiotherapy and may function as a primary source of cancer recurrence. We report here on the isolation and in vitro propagation of multicellular ovarian cancer spheroids from a well-established ovarian cancer cell line (OVCAR-3). The spheroid-derived cells (SDCs) display self-renewal potential, the ability to produce differentiated progeny, and increased expression of genes previously associated with CICs. SDCs also demonstrate higher invasiveness, migration potential, and enhanced resistance to standard anticancer agents relative to parental OVCAR-3 cells. Furthermore, SDCs display up-regulation of genes associated with epithelial-to-mesenchymal transition (EMT), anticancer drug resistance and/or decreased susceptibility to apoptosis, as well as, down-regulation of genes typically associated with the epithelial cell phenotype and pro-apoptotic genes. Pathway and biological process enrichment analyses indicate significant differences between the SDCs and precursor OVCAR-3 cells in TGF-beta-dependent induction of EMT, regulation of lipid metabolism, NOTCH and Hedgehog signaling. Collectively, our results indicate that these SDCs will be a useful model for the study of ovarian CICs and for the development of novel CIC-targeted therapies.
2011-04-23 | GSE28799 | GEO
Project description:Recently, multicellular spheroids were isolated from a well-established epithelial ovarian cancer cell line, OVCAR-3, and were propagated in vitro. These spheroid derived cells displayed numerous hallmarks of cancer stem cells, which are chemo and radioresistant cells thought to be a significant cause of cancer recurrence and resultant mortality. Gene set enrichment analysis of expression data from the OVCAR-3 cells and the spheroid-derived putative cancer stem cells identified several metabolic pathways enriched in differentially expressed genes. Before this, there had been little previous knowledge or investigation of systems-scale metabolic differences between cancer cells and cancer stem cells, and no knowledge of such differences in ovarian cancer stem cells. To determine if there were substantial metabolic changes corresponding with these transcriptional differences, we used two-dimensional gas chromatography coupled to mass spectrometry to measure the metabolite profiles of the two cell lines. These two cell lines exhibited significant metabolic differences in both intracellular and extracellular metabolite measurements. Principal components analysis, an unsupervised dimensional reduction technique, showed complete separation between the two cell types based on their metabolite profiles. Pathway analysis of intracellular metabolomics data revealed close overlap with metabolic pathways identified from gene expression data, with four out of six pathways found enriched in gene-level analysis also enriched in metabolite-level analysis. Some of those pathways contained multiple metabolites that were individually statistically significantly different between the two cell lines, with one of the most broadly and consistently different pathways, arginine and proline metabolism, suggesting an interesting hypothesis about cancerous and stem-like metabolic phenotypes in this pair of cell lines. Overall, we demonstrate for the first time that metabolism in an ovarian cancer stem cell line is distinct from that of more differentiated isogenic cancer cells, supporting the potential importance of metabolism in the differences between cancer cells and cancer stem cells.
2015-01-08 | MTBLS152 | MetaboLights
Project description:Developing effective model systems to evaluate new potential chemotherapeutic reagents is critical. Three-dimensional cell cultures, such as spheroids, aid in bridging the gap between commonly used monolayer cell cultures and significantly more complex and expensive animal models. Spheroid model systems contain pathophysiological and chemical gradients similar to an in vivo tumor, which ultimately form distinct cellular subpopulations. In the spatial SILAC model, labels are pulsed into the spheroid at discrete time windows during development. These media pulses result in labeled proteins in distinct regions of the spheroid, which allows for tracing of quantitative proteomic changes to be correlated to different cellular subregions. In this study, we use the Spatial SILAC model to evaluate the proteomic response of a colon carcinoma spheroid to the multikinase inhibitor Regorafenib. We determined regorafenib to be an effective kinase inhibitor which significantly altered whole spheroid proliferation and resulted in increased apoptosis when the signal was averaged for all cells in the culture. However, when regorafenib treatment is more closely examined among the cellular subpopulations, drastic differences are observed for how the drug impacted the proteome of the necrotic spheroid core and the proliferating outer regions. Whole spheroid and outer region analysis shows that regorafenib treatment inhibited critical pathways such as mTOR signaling, ERK/MAPK signaling, and colorectal cancer metastasis signaling. However, analysis of the core shows that regorafenib had an entirely different effect, including upregulation of MAPK1 and KRAS, possibly indicating drug resistance within these late apoptotic cells. Ultimately, these combinatory studies can be used to further understanding of drug metabolism is different cellular subpopulations and provide valuable information to ultimately improve the accuracy of therapeutic testing.
2024-08-09 | PXD038958 | Pride
Project description:Tumor angiogenesis is regarded as a promising target for limiting cancer progression because tumor-associated vasculature supplies blood and provides a path for metastasis. Thus, in vitro recapitulation of vascularized tumors is critical to understand the pathology of cancer and identify the mechanisms by which tumor cells proliferate, metastasize, and respond to drugs. In this study, we microengineered a vascularized tumor spheroid (VTS) model to reproduce the pathological features of solid tumors. We first generated tumor cell-EC hybrid spheroids with self-assembled intratumoral vessels, which enhanced the uniformity of the spheroids and peritumoral angiogenic capacity compared to spheroids comprised only with cancer cells. Notably, the hybrid spheroid also exhibited expression profiles associated with aggressive behavior. The blood vessels sprouting around the hybrid spheroids on the VTS chip were interconnected with intratumoral vessels and displayed the distinctive characteristics of leaky tumor vessels. With the VTS chip showing a progressive tumor phenotype, we validated the suppressive effects of axitinib on tumor growth and angiogenesis, which depended on exposure dose and time, highlighting the significance of tumor vascularization to predict the efficacy of anticancer drugs. Ultimately, we effectively induced both lymphangiogenesis and angiogenesis around the tumor spheroid by promoting interstitial flow. Thus, our VTS model is a valuable platform with which to investigate the interactions between tumor microenvironments and explore therapeutic strategies in cancer.
2022-09-07 | GSE212669 | GEO
Project description:Colorectal cancers have a rare population of cells that express specific cell surface markers, and are referred to as cancer stem cells (CSC). Targeting CSCs, implicated in tumor relapse, is a paradigm shifting approach for colorectal cancer. We used gene microarray to analyze gene expression in HT-29, a colon cancer cell line, grown as monolayer and spheroid and identify metabolic pathways that are upregulated. HT-29 monolayer cells were grown in DMEM/F12 media supplemented with 10% FBS, and 1X Antibiotic-Antimycotic liquid (AA); and spheroids were grown in stem cell media [DMEM:F12, 1X AA, 1X B27, epidermal growth factors, and fibroblast growth factor for five days. Total RNA was isolated using the mirVana™ miRNA Isolation Kit according to the manufacturer's protocol. RNA intergrity was confirmed on the Nanodrop ND-1000 and analyzed with Agilent 2100 bioanalyzer. The arrays were scanned with the Affymetrix 3000 7G plus scanner.
2015-04-01 | E-GEOD-65433 | biostudies-arrayexpress
Project description:Cancer progression and wound healing share some characteristics. Most colorectal cancers are differentiated adenocarcinomas that maintain three-dimensional structures to some extent. Hence, disruption of the architecture can provoke remodeling similar to the remodeling of normal intestinal epithelium. We used our recently developed three-dimensional culture system to investigate the response of cancer cell spheroids to mechanical disruption. Specifically, we developed a protocol for homogenous disruption of the spheroids that maintained the cell-cell contacts. After disruption, 9 spheroids from 9 patient samples reformed within a few hours, and 2 showed accelerated spheroid growth. Stemness increased after spheroid disruption, as assessed by marker expression, spheroid forming capacity, radiation sensitivity, and tumorigenesis. The spheroid-forming capacity increased in 6 of 11 spheroids. The disruption signature, as determined by gene expression profiling, supported the incidence of remodeling and predicted the prognosis of the colorectal cancer patients. WNT and HER3 signaling was increased in the reformed spheroids, and suppression of these signaling pathways attenuated the increases in growth and stemness after disruption. Thus, disorganized architecture in patient tumors might reflect the processes of disruption and subsequent remodeling and represent a cause rather than simply a consequence of malignancy progression. Gene expression in cancer tissue-originated spheroids (CTOSs) was measured at pre, 6 hr, 24 hr, and 4 days after mechanical disruption. One experiment was performed at each time point.
2015-12-10 | E-GEOD-75867 | biostudies-arrayexpress
Project description:Epithelial ovarian cancer (EOC) is a high-risk cancer presenting with heterogeneous tumors. The high incidence of EOC metastasis from primary tumors to nearby tissues and organs is a major driver of EOC lethality. We used cellular models of spheroid formation and re-adherence to investigate cellular signaling dynamics in each step toward EOC metastasis. In our system, adherent cells model primary tumors, spheroids formation represents the initiation of metastatic spread, while re-adherent spheroid cells represent secondary tumors. Proteomic and phosphoproteomic analyses show that spheroid cells are hypoxic and show markers for cell cycle arrest. Aurora kinase B (AURKB) abundance and downstream substrate phosphorylation are significantly reduced in spheroids and re-adherent cells, explaining their cell cycle arrest phenotype. The proteome of re-adherent cells is most similar to spheroids, yet greater changes in the phosphoproteome show that spheroid cells stimulate Rho-associated kinase 1 (ROCK1) mediated signaling, which controls cytoskeletal organization. In spheroids, we found significant phosphorylation of ROCK1 substrates that were reduced in both adherent and re-adherent cells. Application of the ROCK1-specific inhibitor Y-27632 to spheroids increased the rate of re-adherence and spheroid density. The data suggest ROCK1 inhibition increases EOC metastatic potential. We identified novel pathways controlled by AURKB and ROCK1 as major drivers of metastatic behavior in EOC cells. Our data show that phosphoproteomic reprogramming precedes proteomic changes that characterize spheroid re-adherence in EOC metastasis.
2024-01-26 | PXD043220 | Pride
Project description:The cancer initiating cells (CICs) act as a tumor initiation source. Recent studies have shown that CICs contribute to chemoresistance and radioresistance. The aims of this study were to investigate the relationship of CD133+ liver CICs and radiation resistance and to define a possible mechanism for radioresistance in hepatocellular carcinoma (HCC).
2013-07-20 | GSE22247 | GEO
Project description:Intro: Spheroid culture especially on MCF-7 cell has been used as in vitro CSC model for anti-CSC drug discovery. The role of miRNAs in the regulation of mRNA specifically looking at the self-renewal capacity and the drug resistance of the spheroid-enriched CSCs models has not been evaluated. Therefore, a comprehensive profiling of the miRNAs will provide an insight into the regulatory mechanisms associated with breast cancer and CSCs. Methods: Tumour spheroid of MCF-7 was generated and characterised for the biological features and CSCs characteristic comparing to the monolayer parental MCF-7 cells. Differential expression of miRNA between the spheroid and parental cells was evaluated using next generation sequencing (NGS). The differentially expressed miRNAs were then subjected to target genes predictions, followed pathway analyses to better understand the mechanism associated with spheroid-enriched CSCs. Results: Spheroids generated from MCF-7 cell line under serum-free condition were found to be enriched with CSCs characteristics. miRNA-NGS analysis revealed 119 differentially expressed miRNAs between the spheroids and parental, with 24 up-regulated and 94 down-regulated. The gene ontology (GO) analysis showed that the predicted genes of the differentially expressed miRNAs were enriched in various biological processes. Pathway analysis revealed that the differentially expressed miRNAs and their target genes were associated with a variety of KEGG pathways, which could explain the self-renewal capability, and the higher drug resistance in spheroids when compared to parental.
2016-05-10 | GSE68246 | GEO
Project description:Eight different human cancer cell lines were cocultured with human cancer associated fibroblasts (CAF) as spheroid cultures. In another setup UT-SCC-7 cutaneous squamous cell carcinoma cells, together with human skin fibroblasts (SFB) and UT-SCC-2 cells together with gingival fibroblasts (GFB) were cultured as spheroid cocultures. These spheroids were treated with PAD4 or with citrullination buffer only. All spheroids were hypotonically lysed, and the remaining insoluble material was digested to peptides and analysed by LC-MS/MS.
2022-11-21 | PXD033555 | Pride