Project description:We compared the expression changes in Ewing sarcoma cell lines following treatment with 2 known 20S proteasome inhibitors versus 2 novel compounds
Project description:Relapsed Ewing sarcoma is treated with TOP1 inhibitors. Our study confirmed prior studies showing that downregulation of TOP1 is a mechanism of resistance to camptothecin. We then screened a large library of clinically relevant compounds and found that TOP1-deficient Ewing sarcoma cells are more sensitive to the kinase inhibitor GNF-7. RNA-seq results reveal that GNF-7 modulates EWS/FLI1 gene signatures in both TOP1 wild type and TOP1-deficient cells.
Project description:Purpose: Our experimental results demonstrate an essential role for the lncRNA HOTAIR in Ewing sarcoma. We have repressed HOTAIR expression in three Ewing sarcoma cell lines and overexpressed HOTAIR, alone and with EWS-FLI1, in htert-immortalized human mesenchymal stem cells to evaluate its effects on gene expression in this cancer. Methods: RNA-Seq of Ewing sarcoma cell lines with HOTAIR represssed by GapmeR or treated with nonsilencing control, and hTERT-immortalized hMSCs with expression of control GFP, HOTAIR, or HOTAIR and EWS-FLI1, was used for gene expression analysis. Results: Defined gene expression signatures were defined as driven by HOTAIR in each cell line model, with a consensus set of gene identified in the Ewing sarcoma cell lines. Additionally, a set of genes regulated by HOTAIR independently of EWS-FLI1 was identified in the hTERT-hMSC models. Conclusions: HOTAIR expression regulates a set of genes critical to viability, cell adhesion, cell motility, and other markers of EMT and metastasis in Ewing sarcoma, independently of EWS-FLI1.
Project description:An increasing number of cancer-associated mutations have been identified. Unfortunately, little therapy today exploits these tumor-specific genetic lesions. Often, the resulting oncoproteins have been intractable to easy manipulation with current small molecule screening approaches. To overcome this impasse, we developed an expression-based approach to small molecule library screening. We applied this platform to the discovery of modulators of the activity of EWS/FLI, the Ewing sarcoma associated oncoprotein. Cytarabine (ARA-C) was identified as the top hit in a small molecule library screen. ARA-C modulates EWS/FLI by decreasing EWS/FLI protein level and has striking effects on cellular viability and transformation in in vitro and in vivo models of Ewing sarcoma. With poor outcomes for patients with relapsed Ewing sarcoma and the well established safety profile of ARA-C, clinical trials testing ARA-C in Ewing sarcoma are warranted. Expression data was created for A673 cells treated with ARA-C and two other compounds used to treat Ewing sarcoma (Puromycin and Doxorubicin) at two doses (EC50 and 2xEC50) and three time points (24 hours, 3 days, and 5 days). Experiment Overall Design: A673 cells were treated with ARA-C (at doses of EC50 and 2xEC50) or vehicle in triplicate and expression profiled at 24 hours, 3 days, and 5 days. To exclude the possibility that ARA-C's modulation of the EWS/FLI signature was simply a non-specific response to treatment with all cytotoxic agents, we asked whether other compounds known to kill Ewing sarcoma cells (Doxorubicin and Puromycin) would induce the EWS/FLI off genome-wide expression pattern. A673 cells were treated with Doxorubicin and Puromycin (at doses of EC50 and 2xEC50) and expression profiled at 24 hours, 3 days, and 5 days.
Project description:Ewing sarcoma is a pediatric bone cancer with poor survival in relapsed cases, highlighting the need for better preclinical models. Here, we developed a scalable 3D spheroid platform using SLA-printed microcavity arrays to generate uniform Ewing sarcoma spheroids. High-throughput screening of 11 compounds identified Torin 2, Talazoparib, and Trabectedin as potent inhibitors. Incorporating lung fibroblasts into hybrid spheroids revealed fibroblast-induced drug resistance, mimicking the metastatic tumor microenvironment. Transcriptomic analysis identified activation of NF-κB and TGF-β signaling pathways as key mediators of this resistance. This platform offers a cost-effective, biologically relevant system for drug testing and mechanistic studies in Ewing sarcoma and other stromal-influenced cancers.
Project description:This SuperSeries is composed of the following subset Series: GSE30513: MicroRNA expression profiling of Ewing sarcoma cancer stem cells GSE31144: MicroRNA expression profiling of Ewing sarcoma cell lines upon TARBP2 depletion GSE31145: MicroRNA expression profiling of Ewing sarcoma spheres vs. adherent cells Refer to individual Series
