G Protein-Coupled Receptor GPR35 Suppresses Lipid Accumulation in Hepatocytes.

Although prevalent, nonalcoholic fatty liver disease is not currently treated effectively with medicines. Initially, using wild-type and genome-edited clones of the human hepatocyte cell line HepG2, we show that activation of the orphan G protein-coupled receptor GPR35 is both able and sufficient to block liver X-receptor-mediated lipid accumulation. Studies on hepatocytes isolated from both wild-type and GPR35 knock-out mice were consistent with a similar effect of GPR35 agonists in these cells, but because of marked differences in the pharmacology of GPR35 agonists and antagonists at the mouse and human orthologues, as well as elevated basal lipid levels in hepatocytes from the GPR35 knock-out mice, no definitive conclusion could be reached.

G-protein coupled receptor 35 (GPR35) regulates the colonic epithelial cell response to enterotoxigenic Bacteroides fragilis.

G protein-coupled receptor (GPR)35 is highly expressed in the gastro-intestinal tract, predominantly in colon epithelial cells (CEC), and has been associated with inflammatory bowel diseases (IBD), suggesting a role in gastrointestinal inflammation. The enterotoxigenic Bacteroides fragilis (ETBF) toxin (BFT) is an important virulence factor causing gut inflammation in humans and animal models. We identified that BFT signals through GPR35.

Combinatorial expression of GPCR isoforms affects signalling and drug responses.

G-protein-coupled receptors (GPCRs) are membrane proteins that modulate physiology across human tissues in response to extracellular signals. GPCR-mediated signalling can differ because of changes in the sequence or expression of the receptors, leading to signalling bias when comparing diverse physiological systems. An underexplored source of such bias is the generation of functionally diverse GPCR isoforms with different patterns of expression across different tissues.

Receptor selectivity between the G proteins Gα and Gα is defined by a single leucine-to-isoleucine variation.

Despite recent advances in structural definition of GPCR-G protein complexes, the basis of receptor selectivity between G proteins remains unclear. The Gα and Gα subtypes together form the least studied group of heterotrimeric G proteins. G protein-coupled receptor 35 (GPR35) has been suggested to couple efficiently to Gα but weakly to Gα.

Evidence for the Existence of a CXCL17 Receptor Distinct from GPR35.

The chemokine CXCL17 is associated with the innate response in mucosal tissues but is poorly characterized. Similarly, the G protein-coupled receptor GPR35, expressed by monocytes and mast cells, has been implicated in the immune response, although its precise role is ill-defined. A recent manuscript reported that GPR35 was able to signal in response to CXCL17, which we set out to confirm in this study.

G-Protein-Coupled Receptor 35 Mediates Human Saphenous Vein Vascular Smooth Muscle Cell Migration and Endothelial Cell Proliferation.

Vascular smooth muscle cell (VSMC) migration and proliferation is central to neointima formation in vein graft failure following coronary artery bypass. However, there are currently no pharmacological interventions that prevent vein graft failure through intimal occlusion. It is hence a therapeutic target.

Non-equivalence of Key Positively Charged Residues of the Free Fatty Acid 2 Receptor in the Recognition and Function of Agonist Versus Antagonist Ligands.

Short chain fatty acids (SCFAs) are produced in the gut by bacterial fermentation of poorly digested carbohydrates. A key mediator of their actions is the G protein-coupled free fatty acid 2 (FFA2) receptor, and this has been suggested as a therapeutic target for the treatment of both metabolic and inflammatory diseases. However, a lack of understanding of the molecular determinants dictating how ligands bind to this receptor has hindered development.

The emerging pharmacology and function of GPR35 in the nervous system.

G protein-coupled receptor 35 (GPR35) is an orphan G protein-coupled receptor (GPCR) that can be activated by kynurenic acid at high micromolar concentrations. A previously unappreciated mechanism of action of GPR35 has emerged as a Gα-coupled inhibitor of synaptic transmission, a finding that has significant implications for the accepted role of kynurenic acid as a broad-spectrum antagonist of the NMDA, AMPA/kainite and α7 nicotinic receptors. In conjunction with previous findings that link agonism of GPR35 with significant reduction in nociceptive pain, GPR35 has emerged as a potential effector of regulation of mechanical sensitivity and analgesia of the Ret tyrosine kinase, and as a receptor involved in the transmission of anti-inflammatory effects of aspirin- potentially through affecting leucocyte rolling, adhesion and extravasation.

G protein-coupled receptor 35: an emerging target in inflammatory and cardiovascular disease.

G protein-coupled receptor 35 (GPR35) is an orphan receptor, discovered in 1998, that has garnered interest as a potential therapeutic target through its association with a range of diseases. However, a lack of pharmacological tools and the absence of convincingly defined endogenous ligands have hampered the understanding of function necessary to exploit it therapeutically. Although several endogenous molecules can activate GPR35 none has yet been confirmed as the key endogenous ligand due to reasons that include lack of biological specificity, non-physiologically relevant potency and species ortholog selectivity.

The antiallergic mast cell stabilizers lodoxamide and bufrolin as the first high and equipotent agonists of human and rat GPR35.

Lack of high potency agonists has restricted analysis of the G protein-coupled receptor GPR35. Moreover, marked variation in potency and/or affinity of current ligands between human and rodent orthologs of GPR35 has limited their productive use in rodent models of physiology. Based on the reported modest potency of the antiasthma and antiallergic ligands cromolyn disodium and nedocromil sodium, we identified the related compounds lodoxamide and bufrolin as high potency agonists of human GPR35.

The pharmacology of TUG-891, a potent and selective agonist of the free fatty acid receptor 4 (FFA4/GPR120), demonstrates both potential opportunity and possible challenges to therapeutic agonism.

TUG-891 [3-(4-((4-fluoro-4'-methyl-[1,1'-biphenyl]-2-yl)methoxy)phenyl)propanoic acid] was recently described as a potent and selective agonist for the long chain free fatty acid (LCFA) receptor 4 (FFA4; previously G protein-coupled receptor 120, or GPR120). Herein, we have used TUG-891 to further define the function of FFA4 and used this compound in proof of principle studies to indicate the therapeutic potential of this receptor. TUG-891 displayed similar signaling properties to the LCFA α-linolenic acid at human FFA4 across various assay end points, including stimulation of Ca²⁺ mobilization, β-arrestin-1 and β-arrestin-2 recruitment, and extracellular signal-regulated kinase phosphorylation.

Defining the molecular basis for the first potent and selective orthosteric agonists of the FFA2 free fatty acid receptor.

FFA2 is a G protein-coupled receptor that responds to short chain fatty acids and has generated interest as a therapeutic target for metabolic and inflammatory conditions. However, definition of its functions has been slowed by a dearth of selective ligands that can distinguish it from the closely related FFA3. At present, the only selective ligands described for FFA2 suffer from poor potency, altered signaling due to allosteric modes of action, or a lack of function at non-human orthologs of the receptor.

High-throughput identification and characterization of novel, species-selective GPR35 agonists.

Drugs targeting the orphan receptor GPR35 have potential therapeutic application in a number of disease areas, including inflammation, metabolic disorders, nociception, and cardiovascular disease. Currently available surrogate GPR35 agonists identified from pharmacologically relevant compound libraries have limited utility due to the likelihood of off-target effects in vitro and in vivo and the variable potency that such ligands exhibit across species. We sought to identify and characterize novel GPR35 agonists to facilitate studies aimed at defining the physiologic role of GPR35.

Antagonists of GPR35 display high species ortholog selectivity and varying modes of action.

Variation in pharmacology and function of ligands at species orthologs can be a confounding feature in understanding the biology and role of poorly characterized receptors. Substantial selectivity in potency of a number of GPR35 agonists has previously been demonstrated between human and rat orthologs of this G protein-coupled receptor. Via a bioluminescence resonance energy transfer-based assay of induced interactions between GPR35 and β-arrestin-2, addition of the mouse ortholog to such studies indicated that, as for the rat ortholog, murine GPR35 displayed very low potency for pamoate, whereas potency for the reference GPR35 agonist zaprinast was intermediate between the rat and human orthologs.

An engineered zinc finger protein reveals a role for the insulin VNTR in the regulation of the insulin and adjacent IGF2 genes.

An engineered zinc finger protein (eZFP) was isolated from a library based on its ability to activate expression of the endogenous insulin gene in HEK-293 cells. Using a panel of insulin promoter constructs, the eZFP was shown to act through the variable number of tandem repeat (VNTR) region located 365 base pairs upstream of the transcription start site. The eZFP also activated expression of the IGF2 gene that lies close to INS on chromosome 11p15.