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Assay for the Positive Selection of Protein Destabilizers

A new assay for the identification of novel protein targets that control protein stability and chemicals that destabilize proteins of interest.

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Project Overview

  • Reporter assays are used routinely in the pharmaceutical and biotechnology industries to identify lead compounds that affect protein function by measuring the activity of a protein of interest (POI) subject to a test agent.
  • Historically, most assays screening agents for downregulation of a POI were “down” assays, meaning a decrease in a reporter protein or such signal was measured. Down assays, however, can be a source of artifacts as there are typically more spurious or trivial ways to downregulate than upregulate in biological assays. 
  • Dana-Farber scientists have created a unique “up” assay for identifying compounds that can destabilize a protein of interest.
  • The assay technology is available for a non-exclusive license.

Technology

Dr. William Kaelin’s lab has developed a novel “up” screening assay for identifying chemical or genetic perturbants that can destabilize a protein of interest.

This assay uses, for example, cell survival and/or cell viability as a phenotypic identifier to positively select for destabilizing agents. For example, cells expressing fusion constructs having an enzyme, which converts an exogenous substrate that is not cytotoxic into a product that is fused to a POI may be cultured in the presence of a test agent. If the test agent destabilizes the POI, a corresponding decrease in the cytotoxic element fused to the POI promotes cell survival and/or viability. In this assay, for example, a protein of interest can be fused to a modified deoxycytidine kinase (dCK) gene capable of converting bromovinyldeoxyuridine (BVdU) to a toxic metabolite.

This method provides a powerful new way of identifying protein destabilizing agents as it provides enhanced sensitivity including better signal-to-noise ratio and fewer false positives compared to prior assays, while enabling performance in pooled, arrayed, multi-well, high-throughput formats. 

Further details:

Koduri V, Duplaquet L, et al. Targeting oncoproteins with a positive selection assay for protein degraders. Science Advances. 2021 Feb; 7(6): eabd6263.

Applications

Identification of novel protein targets that control protein stability and chemicals that destabilize proteins of interests. 

Opportunities

Dana-Farber is seeking a Non-Exclusive License for this technology. 

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