Yeast-based screening platform for rapid identification of GPCR agonists

This invention is a yeast-based screening platform that uses tethered peptide libraries to identify human GPCR agonists quickly and cost-effectively, yielding receptor-specific ligands and valuable insights into GPCR drug discovery.

Background

The field of receptor pharmacology and drug discovery continues to seek improved methods for reliably identifying and characterizing small molecules or peptides that interact with G protein-coupled receptors (GPCRs). With GPCRs representing one of the most critical targets for therapeutic intervention, there is an ever-increasing demand for cost-effective, high-throughput screening platforms that can streamline the discovery process.

Traditional methods often use mammalian cell-based systems, which can be hindered by complex signaling pathways, receptor crosstalk, and variable expression levels that impact reproducibility and efficiency. However, current approaches face significant challenges that impede rapid progress in understanding receptor-ligand interactions.

Many conventional screening techniques require intricate engineering of cell lines and are limited by low-throughput capabilities and high operational costs. Additionally, the intricate nature of mammalian signaling often introduces background noise, making it difficult to isolate specific receptor responses. These shortcomings underscore the need for alternative strategies that can offer simplified, robust, and scalable solutions to address the inherent limitations in studying both well-characterized and understudied GPCR targets.

Technology description

The technology employs a yeast-based screening platform designed to identify agonists of human G protein-coupled receptors using Saccharomyces cerevisiae. It leverages a tethered agonism strategy in which peptide libraries are displayed on the yeast cell surface, with peptides anchored in a way that facilitates effective receptor interaction. A reporter gene, typically GFP under the control of a pheromone response pathway, is activated when a ligand binds the receptor, allowing for quantifiable detection via flow cytometry.

The system has been validated on opioid receptors, using peptide libraries engineered from a dynorphin A 1-8 scaffold, and includes technical optimizations such as C-terminal peptide tethering, a glycine-serine linker for flexibility, and the use of ergosterol-producing yeast strains for enhanced transformation efficiency.

The platform is differentiated by its cost effectiveness, simplified genetic background, and high-throughput capability, enabling simultaneous screening of up to 10⁶ peptide variants. Its design significantly reduces receptor cross-talk and streamlines the discovery process, making it particularly advantageous for exploring challenging or “dark” receptors. This innovative approach offers valuable insights into receptor-ligand interactions and structural determinants of specificity, positioning it as a powerful tool for accelerating GPCR-targeted drug discovery.

Benefits

Cost-effective screening using Saccharomyces cerevisiae reduces reliance on expensive mammalian cell systems
Simplified signaling pathways and minimized receptor cross-talk facilitate clearer analysis of GPCR-ligand interactions
High-throughput screening capability allows simultaneous evaluation of up to 10⁶ peptide variants
The tethered agonism strategy ensures proper orientation of peptides, enhancing specificity and structural insights
Flexible and genetically manipulable yeast platform offers broader applicability to study challenging “dark” and other GPCR families

Commercial applications

GPCR-targeted drug discovery
High-throughput peptide screening
Novel GPCR agonist identification
Cost-effective receptor assays
Yeast display screening technology

Additional information

This advanced yeast-based screening platform identifies agonists of human GPCRs through tethered peptide libraries displayed on Saccharomyces cerevisiae. Engineered yeast expresses target GPCRs while a GFP reporter driven by pheromone response signaling quantifies receptor activation. Optimized linkers and anchoring facilitate precise peptide-receptor interactions. Validated using opioid receptors, the method enables high-throughput, cost-effective analysis of receptor-ligand dynamics, providing detailed insights into receptor specificity and activation mechanisms.