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Identification of Mediators of T-cell Receptor Signaling via the Screening of Chemical Inhibitor Libraries
Published on: January 22, 2019
Signature-based small molecule screening identifies cytosine arabinoside as an EWS/FLI modulator in Ewing sarcoma
Kimberly Stegmaier1, Jenny S Wong, Kenneth N Ross
1Department of Pediatric Oncology, Dana-Farber Cancer Institute and Children's Hospital Boston, Harvard Medical School, Boston, Massachusetts, United States of America.
Background:
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 expression-based 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.
Methods And Findings:
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.
Conclusions:
We demonstrate that a gene expression-based 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.
Insights
Researchers identified a new way to find drugs targeting difficult cancer proteins like EWS/FLI in Ewing sarcoma. Cytosine arabinoside (ARA-C) effectively reduced tumor growth, showing promise for clinical trials.
Area of Science:
- Oncology
- Pharmacology
- Molecular Biology
Background:
- Targeting tumor-specific mutations, like the EWS/FLI oncoprotein in Ewing sarcoma, remains a challenge for conventional drug screening.
- Most cancer therapies still rely on nonspecific cytotoxic drugs, despite advances in identifying actionable mutations.
- The EWS/FLI oncoprotein, prevalent in Ewing sarcoma, has been intractable to traditional small-molecule drug discovery.
Purpose of the Study:
- To develop a gene expression-based screening approach to identify compounds modulating the EWS/FLI oncoprotein.
- To accelerate the clinical application of potential therapeutics by screening FDA-approved drugs.
- To find novel therapeutic strategies for Ewing sarcoma and other cancers with intractable oncoproteins.
Main Methods:
- Determined an EWS/FLI 'off-state' gene expression signature using microarray profiling and RNA interference in Ewing sarcoma cell lines.
- Screened a library of FDA-approved small molecules using a high-throughput ligation-mediated amplification assay with fluorescent bead detection.
- Evaluated the efficacy of identified compounds in vitro and in vivo using xenograft models.
Main Results:
- Identified cytosine arabinoside (ARA-C) as a modulator of EWS/FLI.
- ARA-C treatment reduced EWS/FLI protein levels, diminished cancer cell viability and transformation, and inhibited tumor growth in xenografts.
- Demonstrated the potential of ARA-C as a therapeutic agent for Ewing sarcoma.
Conclusions:
- A gene expression-based screening method can identify drugs targeting previously intractable oncoproteins for rapid clinical testing.
- This approach is applicable to various tumor-associated oncoproteins in both rare pediatric and common adult cancers.
- The findings support the clinical investigation of ARA-C for Ewing sarcoma patients, given its established safety profile and demonstrated efficacy.
