Programmable G-protein-coupled receptors redefine therapeutic applications.

By Dr. Chinta SidharthanReviewed by Lexie CornerDec 12 2024

A recent study published in Nature by a research team from the United States and China introduced a new class of engineered G-protein-coupled receptors (GPCRs) called programmable antigen-gated engineered receptors (PAGERs). These receptors are designed to integrate antigen detection with drug activation. The study demonstrated that PAGERs could be used to control a range of biological activities, including immune cell modulation, therapeutic protein production, and neuronal activity regulation.

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Background

Cell-surface receptors are essential for sensing and responding to extracellular signals, and they mediate processes crucial for cellular behavior and organismal function. Synthetic receptors, such as chimeric antigen receptors, have advanced therapeutic applications but are limited by factors such as restricted output diversity and antigen specificity.

GPCRs are versatile cell-surface receptors that detect various extracellular signals, including hormones, neurotransmitters, and light, and trigger intracellular responses. Studies in synthetic biology have sought to harness GPCRs for programmable cell behaviors, but their structural complexity has posed significant challenges.

Current receptor engineering methods, such as mutagenesis and directed evolution, remain labor-intensive and limit the adaptability of synthetic receptors. While platforms like chimeric antigen receptors and synthetic Notch receptors offer novel capabilities, they often lack versatility, particularly in responding to both soluble and cell-surface antigens. These systems are also constrained by their activation mechanisms and output options.

The current study

The present study addresses these gaps by utilizing the modularity of GPCR scaffolds to create programmable, drug-controllable receptors that enable diverse applications in biological research and medicine. The researchers developed a novel synthetic receptor platform, PAGERs, using engineered GPCR scaffolds to sense antigens and control cellular functions.

They created an auto-inhibitory mechanism by attaching a nanobody to the GPCR scaffold. This mechanism inhibits receptor activation until antigen binding occurs. Auto-inhibition is removed when the receptor detects an antigen, which then enables receptor activation by a specific small-molecule drug. This dual-input system allows for controlled and precise activation of the receptor.

The researchers optimized the design using the κ-opioid receptor-based DREADD or Designer Receptors Exclusively Activated by Designer Drugs system, which is activated by bioorthogonal drugs and insensitive to native ligands. Bioorthogonal drugs can perform chemical reactions within living organisms without disrupting their natural biochemical processes.

The study also screened various peptide antagonists to assess their ability to inhibit the receptor and their reversibility upon antigen binding. Functional testing demonstrated that PAGERs could be tuned to respond to a range of soluble and surface antigens, such as cytokines, growth factors, and viral proteins, by swapping the nanobody domains.

Additionally, PAGERs were designed to support three main functions: driving transgene expression (PAGERTF), activating native G-protein pathways (PAGERG), and enabling real-time fluorescence for antigen detection (PAGERFL).

Different receptor configurations were tested to determine their ability to induce specific cellular responses, including macrophage polarization, T-cell activation, therapeutic antibody secretion, and modulation of neuronal activity. Advanced techniques, such as light-gated protease systems and reporter gene assays, were also used to validate the functionality and reversibility of the receptors.

Key findings

The results showed that PAGERs enabled precise, programmable control over cellular behavior by integrating antigen detection with drug activation. These synthetic receptors demonstrated modularity and functionality across various biological systems.

The researchers demonstrated that PAGERs could detect both soluble and cell-surface antigens with high specificity. By replacing the nanobody in the receptor scaffold, they were able to detect a variety of antigens, including vascular endothelial growth factor, tumor necrosis factor, interleukin-17, and the SARS-CoV-2 spike protein. The receptors elicited robust cellular responses, such as transgene expression and fluorescence signaling, confirming their effectiveness.

Additionally, PAGERTF constructs were used to link antigen detection with the secretion of bioactive molecules. For instance, PAGERs detected tumor antigens and induced macrophage differentiation into pro-inflammatory states or activated T cell-mediated tumor cell killing by secreting therapeutic antibodies. These results highlighted the potential applications of PAGERs in immune modulation and cancer therapy.

For real-time applications, PAGERFL constructs were developed to fluoresce upon antigen detection, providing rapid readouts suitable for monitoring dynamic processes. These fluorescent receptors demonstrated modular adaptability and quickly responded to a range of antigens.

PAGERG receptors were shown to control endogenous G-protein signaling pathways. In neuronal systems, PAGERG constructs mediated antigen-dependent activation or inhibition, indicating potential for spatially targeted therapies in the nervous system.

Conclusions

The findings showed that PAGERs demonstrated broad utility, high modularity, and precise control, providing a foundation for advancements in synthetic biology, therapeutic design, and basic research. PAGERs combine antigen specificity, modular adaptability, and drug-controlled activation, offering potential improvements in synthetic receptor technology.

The researchers suggest that PAGERs, with their ability to dynamically and precisely reprogram cellular behavior, address several limitations of existing receptor systems, presenting diverse applications in cell biology, therapeutics, and neuroscience.

Journal reference

Kalogriopoulos, NA., Tei, R., Yan, Y., Klein, PM., Ravalin, M., Cai, B., Soltesz, I., Li, Y., Ting, A. (2024). Synthetic GPCRs for programmable sensing and control of cell behaviour. Nature. DOI:10.1038/s41586024082823, https://www.nature.com/articles/s41586-024-08282-3