Abstract by Boris Trapkov

G protein-coupled receptors (GPCRs) are central to cellular signaling and human physiology, yet many remain poorly characterized, including orphan receptors such as GPR139. Although identified over two decades ago, GPR139 has not been conclusively linked to cognate endogenous ligands or clearly defined physiological functions. Nevertheless, the receptor is highly enriched in specific brain regions, including the habenula and striatum, and is highly conserved across vertebrates, suggesting an important role in neurophysiology. Preclinical studies have implicated GPR139 in motivation and reward processing, nociception, and motor activity, among others. Consistent with this profile, a GPR139 agonist (TAK-041) has advanced to Phase I/II clinical trials for neuropsychiatric indications, underscoring continued interest in GPR139 as a potential target for brain disorders.

The overarching aim of this PhD thesis is to address critical gaps in the current understanding of GPR139 by molecularly characterizing the receptor to i) define the structural basis of its function, ii) develop novel tool-compounds for functional interrogation, and iii) to delineate its cellular signaling properties.

In Manuscript I, we combined cryo-electron microscopy, molecular docking, molecular dynamics simulations, and in vitro pharmacology to achieve a detailed structural characterization of GPR139. We resolved multiple receptor structures and elucidated key mechanisms and structural determinants underlying ligand binding, receptor activation and G protein coupling, thereby providing a structural framework for understanding GPR139 pharmacology.

In Manuscript II, we employed an integrative approach that bridges structural biology, computational chemistry, and pharmacology to discover novel GPR139 agonists. These compounds exhibited robust pharmacological activity in vitro, while one compound was shown to also have pharmacological activity in vivo and to exert behavioral effects in mice, thus representing valuable new tool-compounds. This work moreover establishes a framework for ligand discovery and optimization and demonstrates how structural insights can be leveraged to guide rational drug discovery.

In Manuscript III, we investigated the signaling properties of GPR139 and uncovered previously unrecognized aspects of its signaling behavior. Using a systematic strategy that combined multiple cell-based assays, molecular inhibitors, and genetically engineered knockout cell lines, we revealed an uncommonly broad G protein coupling profile, including hidden coupling events, context-dependent signaling and ligand-specific signaling fingerprints.

In summary, this PhD thesis advances the understanding of GPR139 biology by elucidating its molecular structure, pharmacology, and cellular signaling mechanisms. Collectively, these findings provide a foundation for future studies aimed at ligand development, probing function, and informing translational efforts.