Novel selective ligands for free fatty acid receptors GPR120 and GPR40

GPR120 and GPR40 are G-protein-coupled receptors whose endogenous ligands are medium- and long-chain free fatty acids, and they are thought to play an important physiological role in insulin release. Despite recent progress in understanding their roles, much still remains unclear about their pharmacology, and few specific ligands for GPR120 and GPR40 besides medium- to long-chain fatty acids have been reported so far. To identify new selective ligands for these receptors, more than 80 natural compounds were screened, together with a reference compound MEDICA16, which is known to activate GPR40, by monitoring the extracellular regulated kinase (ERK) and [Ca²⁺]_i responses in inducible and stable expression cell lines for GPR40 and GPR120, respectively. MEDICA16 selectively activated [Ca²⁺]_i response in GPR40-expressing cells but not in GPR120-expressing cells. Among the natural compounds tested, grifolin derivatives, grifolic acid and grifolic acid methyl ether, promoted ERK and [Ca²⁺]_i responses in GPR120-expressing cells, but not in GPR40-expressing cells, and inhibited the α-linolenic acid (LA)-induced ERK and [Ca²⁺]_i responses in GPR120-expressing cells. Interestingly, in accordance with the pharmacological profiles of these compounds, similar profiles of glucagon-like peptide-1 secretion were seen for mouse enteroendocrine cell line, STC-1 cells, which express GPR120 endogenously. Taken together, these studies identified a selective GPR40 agonist and several GPR120 partial agonists. These compounds would be useful probes to further investigate the physiological and pharmacological functions of GPR40 and GPR120.     

The orphan receptor GPR55 is a novel cannabinoid receptor

BACKGROUND: The endocannabinoid system functions through two well characterized receptor systems, the CB1 and CB2 receptors. Work by a number of groups in recent years has provided evidence that the system is more complicated and additional receptor types should exist to explain ligand activity in a number of physiological processes. EXPERIMENTAL APPROACH: Cells transfected with the human cDNA for GPR55 were tested for their ability to bind and to mediate GTPgammaS binding by cannabinoid ligands. Using an antibody and peptide blocking approach, the nature of the G-protein coupling was determined and further demonstrated by measuring activity of downstream signalling pathways. KEY RESULTS: We demonstrate that GPR55 binds to and is activated by the cannabinoid ligand CP55940. In addition endocannabinoids including anandamide and virodhamine activate GTPgammaS binding via GPR55 with nM potencies. Ligands such as cannabidiol and abnormal cannabidiol which exhibit no CB1 or CB2 activity and are believed to function at a novel cannabinoid receptor, also showed activity at GPR55. GPR55 couples to Galpha13 and can mediate activation of rhoA, cdc42 and rac1. CONCLUSIONS: These data suggest that GPR55 is a novel cannabinoid receptor, and its ligand profile with respect to CB1 and CB2 described here will permit delineation of its physiological function(s).     

GPR40 carboxylic acid receptor family and diabetes: a new drug target

Type-2 diabetes is strongly linked to visceral obesity and elevated levels of circulating free fatty acids. For years this correlation of obesity to diabetes has intrigued the minds of researchers and research in this direction has led to a possible solution to this question. Human Genome project has identified nearly 150 orphan GPCRs. The reverse pharmacology approaches have identified free fatty acids as ligands for the GPR40 family of orphan receptors. This review mainly emphasizes on the role of GPR40 carboxylic acid receptor family in the development of diabetes along with detailed coverage of each receptor of the family. GPR40 family has provided an insight into regulation of carbohydrate and lipid metabolism in vertebrates and has further provided targets for the development of therapeutic agents useful for treating or preventing disorders such as Type-2 diabetes. This review also suggests where further research and development could be beneficial.     

International Union of Basic and Clinical Pharmacology. LXXXII: Nomenclature and Classification of Hydroxy-carboxylic Acid Receptors (GPR81, GPR109A, and GPR109B)

The G-protein-coupled receptors GPR81, GPR109A, and GPR109B share significant sequence homology and form a small group of receptors, each of which is encoded by clustered genes. In recent years, endogenous ligands for all three receptors have been described. These endogenous ligands have in common that they are hydroxy-carboxylic acid metabolites, and we therefore have proposed that this receptor family be named hydroxy-carboxylic acid (HCA) receptors. The HCA(1) receptor (GPR81) is activated by 2-hydroxy-propanoic acid (lactate), the HCA(2) receptor (GPR109A) is a receptor for the ketone body 3-hydroxy-butyric acid, and the HCA(3) receptor (GPR109B) is activated by the β-oxidation intermediate 3-hydroxy-octanoic acid. HCA(1) and HCA(2) receptors are found in most mammalian species, whereas the HCA(3) receptor is present only in higher primates. The three receptors have in common that they are expressed in adipocytes and are coupled to G(i)-type G-proteins mediating antilipolytic effects in fat cells. HCA(2) and HCA(3) receptors are also expressed in a variety of immune cells. HCA(2) is a receptor for the antidyslipidemic drug nicotinic acid (niacin) and related compounds, and there is an increasing number of synthetic ligands mainly targeted at HCA(2) and HCA(3) receptors. The aim of this article is to give an overview on the discovery and pharmacological characterization of HCAs, and to introduce an International Union of Basic and Clinical Pharmacology (IUPHAR)-recommended nomenclature. We will also discuss open questions regarding this receptor family as well as their physiological role and therapeutic potential.     

Targeting GPR120 and other fatty acid-sensing GPCRs ameliorates insulin resistance and inflammatory diseases

The past decade has seen great progress in the understanding of the molecular pharmacology, physiological function and therapeutic potential of G-protein-coupled receptors (GPCRs). Free fatty acids (FFAs) have been demonstrated to act as ligands of several GPCRs including GPR40, GPR43, GPR84, GPR119 and GPR120. We have recently shown that GPR120 acts as a physiological receptor of ω3 fatty acids in macrophages and adipocytes, which mediate potent anti-inflammatory and insulin sensitizing effects. The important role GPR120 plays in the control of inflammation raises the possibility that targeting this receptor could have therapeutic potential in many inflammatory diseases including obesity and type 2 diabetes. In this review paper, we discuss lipid-sensing GPCRs and highlight potential outcomes of targeting such receptors in ameliorating disease.     

Novel cannabinoid receptors

Cannabinoids have numerous physiological effects. In the years since the molecular identification of the G protein-coupled receptors CB1 and CB2, the ion channel TRPV1, and their corresponding endogenous ligand systems, many cannabinoid-evoked actions have been shown conclusively to be mediated by one of these specific receptor targets. However, there remain several examples where these classical cannabinoid receptors do not explain observed pharmacology. Studies using mice genetically deleted for the known receptors have confirmed the existence of additional targets, which have come to be known collectively as non-CB1/CB2 receptors. Despite intense research efforts, the molecular identity of these non-CB1/CB2 receptors remains for the most part unclear. Two orphan G protein-coupled receptors have recently been implicated as novel cannabinoid receptors; these are GPR119, which has been proposed as a receptor for oleoylethanolamide, and GPR55 which has been proposed as a receptor activated by multiple different cannabinoid ligands. In this review I will present an introduction to non-CB1/CB2 pharmacology, summarize information on GPR55 and GPR119 currently available, and consider their phylogenetic origin and what aspects of non-CB1/CB2 pharmacology, if any, they help explain.     

Free fatty acid receptors and their physiological role in metabolic regulation

Free fatty acids (FFAs) are not only essential nutrient components, but they also function as signaling molecules in various physiological processes. In the progression of genomic analysis, many orphan G-protein coupled receptors (GPCRs) are found. Recently, GPCRs deorphanizing strategy successfully identified multiple receptors for FFAs. In these FFA receptors (FFARs), GPR40 (FFAR1) and GPR120 are activated by medium- to long- chain FFAs. GPR40 is expressed mainly in pancreatic β-cell and mediates insulin secretion, whereas GPR120 is expressed abundantly in the intestine and regulates the secretion of cholecystokinin (CCK) and glucagons-like peptide-1 (GLP-1), it promotes insulin secretion. Due to these biological activity, GPR40 and GPR120 are potential drug target for type 2 diabetes and selective ligands have been developed. In this review, we provide recent development in the field and discuss their physiological roles and their potential as drug targets.     

International Union of Basic and Clinical Pharmacology. LXXIX. Cannabinoid receptors and their ligands: beyond CB₁ and CB₂

There are at least two types of cannabinoid receptors (CB(1) and CB(2)). Ligands activating these G protein-coupled receptors (GPCRs) include the phytocannabinoid Δ(9)-tetrahydrocannabinol, numerous synthetic compounds, and endogenous compounds known as endocannabinoids. Cannabinoid receptor antagonists have also been developed. Some of these ligands activate or block one type of cannabinoid receptor more potently than the other type. This review summarizes current data indicating the extent to which cannabinoid receptor ligands undergo orthosteric or allosteric interactions with non-CB(1), non-CB(2) established GPCRs, deorphanized receptors such as GPR55, ligand-gated ion channels, transient receptor potential (TRP) channels, and other ion channels or peroxisome proliferator-activated nuclear receptors. From these data, it is clear that some ligands that interact similarly with CB(1) and/or CB(2) receptors are likely to display significantly different pharmacological profiles. The review also lists some criteria that any novel "CB(3)" cannabinoid receptor or channel should fulfil and concludes that these criteria are not currently met by any non-CB(1), non-CB(2) pharmacological receptor or channel. However, it does identify certain pharmacological targets that should be investigated further as potential CB(3) receptors or channels. These include TRP vanilloid 1, which possibly functions as an ionotropic cannabinoid receptor under physiological and/or pathological conditions, and some deorphanized GPCRs. Also discussed are 1) the ability of CB(1) receptors to form heteromeric complexes with certain other GPCRs, 2) phylogenetic relationships that exist between CB(1)/CB(2) receptors and other GPCRs, 3) evidence for the existence of several as-yet-uncharacterized non-CB(1), non-CB(2) cannabinoid receptors; and 4) current cannabinoid receptor nomenclature.     

Fatty acid receptors as new therapeutic targets for diabetes

G-protein-coupled receptors (GPCRs) are key regulators of several physiological functions. Their roles in cellular signal transduction have made them the target for majority of all currently prescribed drugs. Additionally, there are many orphan GPCRs that provide potential novel therapeutic targets. Several GPCRs are involved in metabolic regulation and glucose homeostasis such as GLP-1 receptor, glucagon receptor, adiponectin receptor and so on. Recently, free fatty acids (FFAs) have been demonstrated as ligands for orphan GPCRs and have been proposed to play a critical role in physiological glucose homeostasis. GPR40 and GPR120 are activated by medium and long-chain FFAs, whereas GPR41 and GPR43 can be activated by short-chain FFAs. GPR40, which is preferentially expressed in pancreatic β-cells, mediates the majority of the effects of FFAs on insulin secretion. In this review, these findings and also critical analysis of these GPCRs as novel targets for diabetes are discussed.     

Fatty acid receptors as new therapeutic targets for diabetes

G-protein-coupled receptors (GPCRs) are key regulators of several physiological functions. Their roles in cellular signal transduction have made them the target for majority of all currently prescribed drugs. Additionally, there are many orphan GPCRs that provide potential novel therapeutic targets. Several GPCRs are involved in metabolic regulation and glucose homeostasis such as GLP-1 receptor, glucagon receptor, adiponectin receptor and so on. Recently, free fatty acids (FFAs) have been demonstrated as ligands for orphan GPCRs and have been proposed to play a critical role in physiological glucose homeostasis. GPR40 and GPR120 are activated by medium and long-chain FFAs, whereas GPR41 and GPR43 can be activated by short-chain FFAs. GPR40, which is preferentially expressed in pancreatic beta-cells, mediates the majority of the effects of FFAs on insulin secretion. In this review, these findings and also critical analysis of these GPCRs as novel targets for diabetes are discussed.     

Characterisation and differential expression of two very closely related G-protein-coupled receptors, GPR139 and GPR142, in mouse tissue and during mouse development

By searching the human and mouse genomic databases we found two G-protein-coupled receptors, GPR139 and GPR142, with characteristic motifs of the rhodopsin family of receptors. The gene for GPR139 maps to chromosome 7F1 of mouse and 16p12.3 of human and that for GPR142 to 11E2 of mouse and 17q25.1 of human. We isolated GPR139 from a cDNA library of adult mouse brain and GPR142 from a cDNA library of brains from 15-day-old mouse embryos. GPR139 mRNA was predominantly expressed in specific areas of human and mouse brains, whereas GPR142 mRNA showed a more ubiquitous expression both in the brain and in various peripheral glands and organs. A 50% identity and a 67% homology at the amino-acid level between the two receptors and only 20-25% identity with other G-protein-coupled receptors established them as a new subbranch within the phylogenetic tree and hints at a common or similar ligand(s). Preliminary results suggest that the cognate ligand is present in brain extracts and is, most likely, a small peptide. GPR139 signal transduction in Chinese hamster ovary cells requires coupling to an inhibitory G-protein and is mediated by phospholipase C. Dimer formation may be necessary for proper function.     

Off-target cannabinoid effects mediated by GPR55

Given the vast therapeutic potential of the endocannabinoid system, the revelation of a novel cannabinoid-sensitive target was treated with great excitement. The orphan G-protein coupled receptor 55 (GPR55) was initially touted as a novel cannabinoid target in early industrial patent literature. Consequently, numerous studies have revealed GPR55 expression in a diverse array of cells and tissues, regulating various physiological and pathological processes. Although a confusing cannabinoid profile has prevented its classification as a cannabinoid receptor, the therapeutic potential of the receptor cannot be denied, with roles in cancer progression, bone resorption and analgesia. This commentary aims to summarize GPR55 expression data and speculate on potential therapeutic exploitation of this enigmatic orphan receptor.     

New Frontiers in Gut Nutrient Sensor Research: Free Fatty Acid Sensing in the Gastrointestinal Tract

Utilizing the human genome database, the recently developed G-protein–coupled receptors (GPCRs) deorphanizing strategy successfully identified multiple receptors of free fatty acids (FFAs). FFAs have been demonstrated to act as ligands of several GPCRs (FFAR1, FFAR2, FFAR3, and GPR120). These fatty acid receptors are proposed to play critical roles in various types of physiological homeostases. FFAR1 and GPR120 are activated by medium- and long-chain FFAs. In contrast, FFAR2 and FFAR3 are activated by short-chain FFAs. It has been elucidated that these four receptors are expressed in the gastrointestinal tract and have many essential roles as sensors of FFA. In this review, we summarize the physiological and pharmacological function of the receptors in the gastrointestinal tract.     

Fatty Acid Binding Receptors and Their Physiological Role in Type 2 Diabetes

G‐protein‐coupled receptors (GPCRs) respond to various physiological ligands such as photons, ions, and small molecules that include amines, fatty acids, and amino acids to peptides, proteins and steroids. Therefore, this family of proteins represents an attractive target for biopharmaceutical research [1]. The physiological role of fatty acids and other lipid molecules as important signal mediators is well studied in various metabolic pathways [2]. Acute administration of free fatty acids (FFAs) stimulates insulin release. Conversely, chronic exposure to high levels of free fatty acids leads to impairment of β cell function and lipotoxicity. However, the receptors through which these fatty acids and lipids act were unknown, until the identification of fatty acid binding receptors: GPR40, GPR41, GPR43, and GPR119. Based on their tissue‐expression profile, and pharmacologic analysis, the fatty acid binding receptors along with lipid binding receptor GPR119 are linked to diabetes and obesity. They play a critical role in the metabolic regulation of insulin release and glucose homeostasis. In this review, the mechanism of receptor activation, pharmacology, and the physiological functions of the fatty acid binding receptors will be discussed.     

In silico patent searching reveals a new cannabinoid receptor

Increasing evidence suggests that some cannabinoids mediate their effects independently of the known cannabinoid CB(1) and CB(2) receptors. Two recently published patents indicate that several cannabinoid receptor ligands also bind to the orphan G-protein-coupled receptor GPR55. This receptor is reported to be expressed in several tissues and might function in lipid or vascular biology. Thus, GPR55 might represent a new cannabinoid receptor.     

Recent advances in gut nutrient chemosensing

The field of gut nutrient chemosensing is evolving rapidly. Recent advances have uncovered the mechanism by which specific nutrient components evoke multiple metabolic responses. Deorphanization of G protein-coupled receptors (GPCRs) in the gut has helped identify previously unliganded receptors and their cognate ligands. In this review, we discuss nutrient receptors, their ligand preferences, and the evoked neurohormonal responses. Family A GPCRs includes receptor GPR93, which senses protein and proteolytic degradation products, and free fatty acid-sensing receptors. Short-chain free fatty acids are ligands for FFA2, previously GPR43, and FFA3, previously GPR41. FFA1, previously GPR40, is activated by long-chain fatty acids with GPR120 activated by medium- and long-chain fatty acids. The GPR119 agonist ethanolamide oleoylethanolamide (OEA) and bile acid GPR131 agonists have also been identified. Family C receptors ligand preferences include L-amino acids, carbohydrate, and tastants. The metabotropic glutamate receptor (mGluR), calcium-sensing receptor (CaR), and GPCR family C, group 6, subtype A receptor (GPRC6A) mediate L-amino acid-sensing. Taste receptors have a proposed role in intestinal chemosensing; sweet, bitter, and umami evoke responses in the gut via GPCRs. The mechanism of carbohydrate-sensing remains controversial: the heterodimeric taste receptor T1R2/T1R3 and sodium glucose cotransporter 1 (SGLT-1) expressed in L cells are the two leading candidates. Identification of specific nutrient receptors and their respective ligands can provide novel therapeutic targets for the treatment of diabetes, acid reflux, foregut mucosal injury, and obesity.     

GPR55 and the vascular receptors for cannabinoids

CB1 and CB2 receptors mediate most responses to cannabinoids but not some of the cardiovascular actions of endocannabinoids such as anandamide and virodhamine, or those of some synthetic agents, like abnormal cannabidiol (abn-cbd). These agents induce vasorelaxation which is antagonised by rimonabant but only at high concentrations relative to those required to block CB1 receptors. Vasorelaxation to anandamide is sensitive to Pertussis toxin (though that to abn-cbd is not), and so is thought to be mediated by a G protein-coupled receptor through Gi/o. An orphan receptor, GPR55, apparently a cannabinoid receptor, is activated by abn-cbd, but is not the receptor mediating vasorelaxation to this agent, as the response persists in vessels from GPR55 knockout mice. However, the activity of anandamide in GPR55 knockout mice is not yet reported and so the role of GPR55 as a cannabinoid receptor mediating vascular responses has yet to be finalised.     

FFA1-selective agonistic activity based on docking simulation using FFA1 and GPR120 homology models

BACKGROUND AND PURPOSE

The free fatty acid FFA1 receptor and GPR120 are GPCRs whose endogenous ligands are medium- and long-chain FFAs, and they are important in regulating insulin and GLP-1 secretion respectively. Given that the ligands of FFA1 receptor and GPR120 have similar properties, selective pharmacological tools are required to study their functions further.

EXPERIMENTAL APPROACH

We used a docking simulation approach using homology models for each receptor. Biological activity was assessed by phosphorylation of ERK and elevation of intracellular calcium ([Ca²⁺]_i) in cells transfected with FFA1 receptor or GPR120. Insulin secretion from murine pancreatic beta cells (MIN6) was also measured.

KEY RESULTS

Calculated hydrogen bonding energies between a series of synthetic carboxylic acid compounds and the homology models of the FFA1 receptor and GPR120, using docking simulations, correlated well with the effects of the compounds on ERK phosphorylation in transfected cells (R²= 0.65 for FFA1 receptor and 0.76 for GPR120). NCG75, the compound with the highest predicted selectivity for FFA1 receptors from this structure-activity relationship analysis, activated ERK and increased [Ca²⁺]_i as potently as the known FFA1 receptor-selective agonist, Compound 1. Site-directed mutagenesis analysis based on the docking simulation showed that different amino acid residues were important for the recognition and activation by FFA1 receptor agonists. Moreover, NCG75 strongly induced ERK and [Ca²⁺]_i responses, and promoted insulin secretion from MIN6 cells, which express endogenous FFA1 receptors.

CONCLUSION AND IMPLICATIONS

A docking simulation approach using FFA1 receptor and GPR120 homology models could be useful in predicting FFA1 receptor-selective agonists.     

Novel G-protein-coupled receptor genes expressed in the brain: continued discovery of important therapeutic targets

The rhodopsin family of G-protein-coupled receptors (GPCRs) is the largest known group of cell-surface mediators of signal transduction. The vast majority of these receptors were discovered by methods based upon shared sequence homologies found throughout this family. While such efforts identified a multitude of receptor subtypes for previously known ligands, numerous receptors have been discovered for which endogenous ligands were unknown. These receptors are commonly referred to as orphan receptors. One of the most important tasks of modern pharmacology lies in elucidating the functions of these receptors. Of particular interest are receptors with recognised expression in the central nervous system, given that many psychiatric and neurodegenerative disorders are mediated by unknown mechanisms. Hence, this collection of putative neurotransmitter and neuromodulator signal mediators represents a substantial and untapped resource for novel drug discovery. Recently, various methodologies have accelerated the discovery of novel ligands for these orphan receptors, identifying the basic components required for further physiological ligand/receptor system characterisation. Equipped with proven ligand identification strategies, the characterisation of all orphan GPCRs and the exploitation of their exciting potential as targets for the discovery of novel drugs is anticipated.