No abstract

GPR18 is a rhodopsin-like orphan G-protein-coupled receptor (GPCR) that is activated by the natural cannabinoid (CB) Δ9-tetrahydrocannabinol (THC). It is highly expressed in immune cells and represents a promising new drug target. However, THC is much more potent in activating CB receptors than GPR18, and several other proposed lipidic agonists for GPR18 have not been independently confirmed. Herein we describe the first non-lipid-like agonists for GPR18 based on a tricyclic xanthine-derived scaffold, along with initial structure–activity relationships. PSB-KD107 (5) and PSB-KD477 (16) displayed significantly higher potency and efficacy than THC, determined in a GPR18-dependent β-arrestin recruitment assay, and were found to be selective versus the CB-sensitive receptors CB1, CB2, and GPR55. Structure–activity relationships were steep, and indole substitution was crucial for biological activity. These first selective agonists, which are structurally distinct from the lipidic agonist(s), will allow target validation studies and may eventually contribute to the deorphanization of GPR18.

GPR18 is an orphan G protein-coupled receptor (GPCR) expressed in cells of the immune system. It is activated by the cannabinoid receptor (CB) agonist ∆9-tetrahydrocannabinol (THC). Several further lipids have been proposed to act as GPR18 agonists, but these results still require unambiguous confirmation. In the present study, we constructed a homology model of the human GPR18 based on an ensemble of three GPCR crystal structures to investigate the binding modes of the agonist THC and the recently reported antagonists which feature an imidazothiazinone core to which a (substituted) phenyl ring is connected via a lipophilic linker. Docking and molecular dynamics simulation studies were performed. As a result, a hydrophobic binding pocket is predicted to accommodate the imidazothiazinone core, while the terminal phenyl ring projects towards an aromatic pocket. Hydrophobic interaction of Cys251 with substituents on the phenyl ring could explain the high potency of the most potent derivatives. Molecular dynamics simulation studies suggest that the binding of imidazothiazinone antagonists stabilizes transmembrane regions TM1, TM6 and TM7 of the receptor through a salt bridge between Asp118 and Lys133. The agonist THC is presumed to bind differently to GPR18 than to the distantly related CB receptors. This study provides insights into the binding mode of GPR18 agonists and antagonists which will facilitate future drug design for this promising potential drug target.

The adenosine A2B receptor (A2BAR) belongs to the rhodopsin-like G protein-coupled receptor (GPCR) family. It is upregulated under hypoxic conditions, in inflammation and cancer. Previous studies indicated the coupling of the A2BAR to different G proteins, mainly Gs, but in some cases Gq/11 or Gi, depending on the cell type. We have now utilized novel technologies, (i) heterologous expression of individual members of the Gαq/11 protein family (Gαq, Gα11, Gα14, and Gα15) in Gαq/11 knockout cells, and (ii) the TRUPATH platform, allowing the direct observation of Gα protein activation for each of the Gα subunits by bioluminescence resonance energy transfer (BRET) measurements. Three structurally diverse A2BAR agonists were studied: the cognate agonist adenosine, its metabolically stable analog NECA, and the non-nucleosidic partial agonist BAY 60-6583. Adenosine and NECA activated most members of all four Gα protein families (Gαs, Gαq/11, Gαi, and Gα12/13). Significant differences in potencies and efficacies were observed; the highest efficacies were determined at the Gα15, Gαs, and Gα12 proteins, and for NECA additionally at the Gαi2 protein. In contrast, the partial agonist BAY 60-6583 only activated Gα15, Gαs, and Gα12 proteins. Adenosine deaminase, an allosteric modulator of ARs, selectively increased the potency and efficacy of NECA and BAY 60-6583 at the Gα15 protein, while it had no effect or decreased efficacy at the other Gα proteins. We conclude that the A2BAR is preferably coupled to the Gα15, Gαs, and Gα12 proteins. Upon upregulation of receptor or Gα protein expression, coupling to further Gα proteins likely occurs. Importantly, different agonists can display different activation profiles.

No abstract