Signature-Based Small Molecule Screening Identifies Cytosine Arabinoside as an EWS/FLI Modulator in Ewing Sarcoma

Stegmaier K, Wong JS, Ross KN, Chow KT, Peck D, et al. (2007) Signature-based small molecule screening identifies cytosine arabinoside as an EWS/FLI modulator in Ewing sarcoma. PLoS Med, Vol. 4, No. 4, e122 doi:10.1371/journal.pmed.0040122. Published: 2007.04.09

Kimberly Stegmaier, Jenny S. Wong, Kenneth N. Ross, Kwan T. Chow, David Peck, Renee D. Wright, Stephen L. Lessnick, Andrew L. Kung, Todd R. Golub

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The presence of tumor-specific mutations in the cancer genome represents a potential opportunity for pharmacologic intervention to therapeutic benefit. Unfortunately, many classes of oncoproteins (e.g., transcription factors) are not amenable to conventional small-molecule screening. Despite the identification of tumor-specific somatic mutations, most cancer therapy still utilizes nonspecific, cytotoxic drugs. One illustrative example is the treatment of Ewing sarcoma. Although the EWS/FLI oncoprotein, present in the vast majority of Ewing tumors, was characterized over ten years ago, it has never been exploited as a target of therapy. Previously, this target has been intractable to modulation with traditional small-molecule library screening approaches. Here we describe a gene expressionbased approach to identify compounds that induce a signature of EWS/FLI attenuation. We hypothesize that screening small-molecule libraries highly enriched for FDA-approved drugs will provide a more rapid path to clinical application. A gene expression signature for the EWS/FLI off state was determined with microarray expression profiling of Ewing sarcoma cell lines with EWS/FLI-directed RNA interference. A small-molecule library enriched for FDA-approved drugs was screened with a high-throughput, ligation-mediated amplification assay with a fluorescent, bead-based detection. Screening identified cytosine arabinoside (ARA-C) as a modulator of EWS/FLI. ARA-C reduced EWS/FLI protein abundance and accordingly diminished cell viability and transformation and abrogated tumor growth in a xenograft model. Given the poor outcomes of many patients with Ewing sarcoma and the well-established ARA-C safety profile, clinical trials testing ARA-C are warranted. We demonstrate that a gene expressionbased approach to small-molecule library screening can identify, for rapid clinical testing, candidate drugs that modulate previously intractable targets. Furthermore, this is a generic approach that can, in principle, be applied to the identification of modulators of any tumor-associated oncoprotein in the rare pediatric malignancies, but also in the more common adult cancers.

Keywords: AML Cytosine Arabinoside Gene Expression-Base High-Throughput Screening Leukemia

Supplemental Data

Description Link/Filename
Smith et al. paper
Smith et al. supplemental information and data
Manuscript 10.1371_journal.pmed.0040122-L.pdf
Microsoft Excel sheet with supplementary tables Stegmaier_Supplementary_Tables.xls
Data file with compound treated cell lines HTA_Ewings_CmpdTest_060510_ams.res
Untreated control sample CEL files
ARA-C treated samples CEL files
Doxorubicin treated samples CEL files
Puromycin treated samples CEL files
Supplementary Figure 1 Stegmaier_Supplementary_Figure1.pdf
Supplementary Figure 2 Stegmaier_Supplementary_Figure2.pdf
Supplementary Figure 3 Stegmaier_Supplementary_Figure3.pdf
Supplementary Figure 4 Stegmaier_Supplementary_Figure4.pdf
Supplementary Table 1 Stegmaier_Supplement.Table1.pdf
Supplementary Table 2 Stegmaier_Supplement.Table2.pdf
Supplementary Table 3 Stegmaier_Supplement.Table3.pdf
Supplementary Table 4 Stegmaier_Supplement.Table4.pdf
Supplementary Table 5 Stegmaier_Supplement.Table5.pdf