Target Identification of Compounds from a Cell Viability Phenotypic Screen Using a Bead/Lysate-Based Affinity Capture...

Area of Science:

• Drug Discovery and Development
• Chemical Biology
• Pharmacology

Background:

• The pharmaceutical industry faces challenges with limited target diversity in drug discovery.
• Phenotypic screening offers an alternative to target-centric approaches by identifying drug leads based on observed effects.
• Identifying the specific molecular targets of phenotypic screening hits is crucial for hit progression and optimization.

Purpose of the Study:

• To develop and validate a robust method for target deconvolution of phenotypic screening hits using affinity capture.
• To introduce a uniqueness index to improve the accuracy of target identification and reduce background noise.
• To demonstrate the platform's utility in identifying candidate targets for phenotypic screening hits.

Main Methods:

• A three-pronged affinity capture approach was developed, linking compounds to beads to isolate interacting proteins from cell homogenates.
• A uniqueness index was implemented to differentiate true targets from non-specific binders.
• The method was validated using known inhibitors and subsequently applied to phenotypic screening hits.

Main Results:

• The affinity capture platform, enhanced by the uniqueness index, successfully identified known and noncanonical targets.
• The approach was effective in deconvoluting targets for compounds with characterized mechanisms.
• Candidate targets were successfully identified for previously uncharacterized phenotypic screening hits.

Conclusions:

• The described affinity capture strategy provides a powerful tool for target deconvolution in phenotypic drug discovery.
• This method facilitates hit progression by enabling target identification, aiding in optimization and understanding mechanism-driven effects.
• The platform enhances the efficiency and success rate of identifying actionable targets from phenotypic screens.