Current Medicinal Chemistry - Volume 17, Issue 14, 2010

The plant Cannabis sativa produces ∼80 terpeno-phenol compounds, termed “phytocannabinoids”, among which Δ9- tetrahydrocannabinol (THC) is the most psychotropic component [1]. THC binds to specific G-protein-coupled receptors (GPCRs), named type-1 (CB1) and type-2 (CB2) cannabinoid receptors, that are also activated by endogenous ligands called “endocannabinoids” (eCBs). Among these lipid substances anandamide (N-arachidonoylethanolamine, AEA) was discovered as the first endogenous ligand of CB1 receptors, the most abundant GPCRs in mammalian brain. AEA is a prototype member of fatty acid amides, whereas 2-arachidonoylglycerol (2-AG) is the most prominent representative of monoacylglycerols, another group of eCBs [2]. Since the discovery of AEA and 2-AG, eCBs have received ever increasing attention, and their anti-cancer, anti-ischemic, anti-inflammatory, anti-depressant, anxiolytic, anorectic and bone-stimulant actions (just to list a few) are now widely recognized. This background has boosted pharmacological research, aimed at exploiting the therapeutic potential of eCBs-oriented drugs for the treatment of several diseases, both centrally and peripherally [3, 4]. AEA, 2-AG and other congeners interact with members of at least three of the four major classes of receptor proteins: GPCRs, ion channels (including ligand-gated ion channels), and nuclear receptors. In this context, it seems noteworthy that eCBs are able to interact with their receptors at binding sites exposed either extracellularly or intracellularly, and located within lipid rafts or in non-rafts microdomains [5]. Such a wealth of possibilities seems to underpin the critical role of eCBs-binding receptors in intra- and inter-cellular signalling [6, 7]. On this basis, eCBs-binding receptors are the focus of this Hot Topic issue of Current Medicinal Chemistry, where an update on CB1 [8] and CB2 [9] receptors will be presented, along with an overview of the other well-established target of AEA, the ion channel TRPV1 (transient receptor potential vanilloid-1) [10]. Furthermore, the actions of eCBs that engage nuclear receptors like peroxisome proliferator-activated receptors [11], and the pros and cons of the “orphan” G-protein-coupled receptor GPR55 as a true “type-3” (CB3) cannabinoid receptor will be discussed [12]. Overall, a complete survey of the eCBs-binding receptors best characterized to date will be presented. In addition, it should be stressed that even more candidates are emerging as targets of eCBs, like the transient receptor potential of melastatin-type 8 (TRPM8), that is antagonized by AEA [13], or the GPR119, which can recognize N-oleoylethanolamine and Npalmitoylethanolamine that are not ligands at classical CB receptors [14]. It seems evident that such a variety of potential triggers of endocannabinoid signalling has been disclosed by means of selective synthetic agonists or antagonists [15], and implies per se a variety of natural ligands in different cell types [16]. Furthermore, the determinants of ligand recognition by different receptors will be addressed in this Hot Topic issue [17], as will be the role of the membrane environment in ligand-receptor interaction [18]. Overall, I hope that this book can represent a useful instrument for the broad readership of Current Medicinal Chemistry, in order to get acquainted with one of the most exciting classes of molecules discovered at the end of the last millennium: the endocannabinoids. I expect that reading the chapters written by leading experts might foster novel ideas and boost new collaborations within the scientific community. I wish to dedicate this book to Claudia, Giuseppe and Gianna.

In this review we describe recent advances in the chemistry of novel CB1/CB2 agonists, CB1 antagonists, selective CB2 agonists, fatty acid amide hydrolase inibitors, monoglyceride (MGL) and diglyceride (DAGL) inhibitors and cannabinoid-type agonists and antagonists of non CB1/CB2 receptors. In view of recent interest in the activities of fatty acid amides of amino acids (N-acyl amino acids) a list of this type of compounds was compiled and is presented as a Table. We conclude that further synthetic work based on both the plant cannabinoids and on the endocannabinoids may lead to novel therapeutics and that the identification and the elucidation of the biological profile of the myriad of endogenous N-acyl amino acids and related compounds may enhance the already wide spectrum of lipidomics.

It is widely accepted that non-endogenous compounds that target CB1 and/or CB2 receptors possess therapeutic potential for the clinical management of an ever growing number of disorders. Just a few of these disorders are already treated with Δ9-tetrahydrocannabinol or nabilone, both CB1/CB2 receptor agonists, and there is now considerable interest in expanding the clinical applications of such agonists and also in exploiting CB2-selective agonists, peripherally restricted CB1/CB2 receptor agonists and CB1/CB2 antagonists and inverse agonists as medicines. Already, numerous cannabinoid receptor ligands have been developed and their interactions with CB1 and CB2 receptors well characterized. This review describes what is currently known about the ability of such compounds to bind to, activate, inhibit or block non-CB1, non- CB2 G protein-coupled receptors such as GPR55, transmitter gated channels, ion channels and nuclear receptors in an orthosteric or allosteric manner. It begins with a brief description of how each of these ligands interacts with CB1 and/or CB2 receptors.

CB1 receptors are G-protein coupled receptors (GPCRs) abundant in neurons, in which they modulate neurotransmission. The CB1 receptor influence on memory and learning is well recognized, and disease states associated with CB1 receptors are observed in addiction disorders, motor dysfunction, schizophrenia, and in bipolar, depression, and anxiety disorders. Beyond the brain, CB1 receptors also function in liver and adipose tissues, vascular as well as cardiac tissue, reproductive tissues and bone. Signal transduction by CB1 receptors occurs through interaction with Gi/o proteins to inhibit adenylyl cyclase, activate mitogen-activated protein kinases (MAPK), inhibit voltage-gated Ca2+ channels, activate K+ currents (Kir), and influence nitric oxide (NO) signaling. CB1 receptors are observed in internal organelles as well as plasma membrane. β-Arrestins, adaptor protein AP-3, and G-protein receptor-associated sorting protein 1 (GASP1) modulate cellular trafficking. Cannabinoid Receptor Interacting Protein1a (CRIP1a) is an accessory protein whose function has not been delineated. Factor Associated with Neutral sphingomyelinase (FAN) regulates ceramide signaling. Such diversity in cellular signaling and modulation by interacting proteins suggests that agonists and allosteric modulators could be developed to specifically regulate unique, cell type-specific responses.

Marijuana has been used for thousands of years to affect human health. Dissecting the peripheral effects from the central psychotropic effects has revealed a complex interplay between cannabinoids, endocannabinoids and their receptors. This review examines recent advances in understanding the expression, regulation and utilization of the CB2 receptor. Here we highlight the molecular aspects of the CB2 receptor, CB2 receptor signaling and new ligands for this receptor. We focus in the rest of the review on recent findings in the immune system, the gastrointestinal tract and liver, the brain and the cardiovascular system and airways as examples of areas where new developments in our understanding of the CB2 receptor have occurred. Early studies focused on expression of this receptor under baseline physiologic conditions; however, perturbations such as those that occur during inflammation, ischemia/reperfusion injury and cancer are revealing a critical role for the CB2 receptor in regulating these disease processes amongst others. As a result, the CB2 receptor is an appealing therapeutic target as well as a useful tool for shedding new light on physiological regulatory processes throughout the body.

In the last decade, accumulated evidence highlighted that GPR55 might be activated by several classical cannabinoid ligands, making this orphan receptor the main candidate to be considered as the “third” cannabinoid receptor. The investigation of its pharmacology has often provided divergent and more intricate results, that have complicated the understanding of the physiological role of GPR55. Nevertheless, the patent analysis regarding GPR55 outlines the fair interest of big pharmaceutical companies, especially in the first years of this decade. This investigation provides a brief overview of the current “state of the art” of our knowledge of GPR55, giving particular emphasis to its functional selectivity. This property could account for controversial roles of GPR55, whose pharmacology and downstream signaling is known to vary significantly both in ligand- and system-dependent manners. In addition, we gain insights into the challenging aspect of finding out novel GPR55 modulators, by analyzing conserved structural and functional motifs that, together with future studies, could help to elucidate its mechanism of action and to design more selective and potent small-molecules directed towards GPR55. Preliminary data highlight remarkable differences, but also intriguing commonalities, between GPR55 and other members of class A G-protein-coupled receptors. It is anticipated that, in the next future, novel lead candidates targeting GPR55 could represent new tools to better understand GPR55-mediated human diseases and, hopefully, generate an innovative class of effective next-generation therapeutics.

In the late 1990's, a series of experiments carried out independently in two laboratories led to establish an important connection between the function of the endocannabinoids, which, as exemplified in this special issue, is per se very complex and ubiquitous in animals, and that of the transient receptor potential (TRP) channels, a large family of plasma membrane cation channels involved in several mammalian and non-mammalian physiological and pathological conditions, overlapping only in part with those in which the cannabinoid receptors participate. These experiments were initially based on the observation that the endocannabinoid anandamide and the xenobiotic ligand of TRP channels of V1 type (TRPV1), capsaicin, are somehow chemically similar, both compounds being fatty acid amides, as are also synthetic activators of these channels and inhibitors of anandamide cellular re-uptake. As discussed in this article, the same type of “chemical thoughts” led to the discovery of N-arachidonoyl-dopamine, an endogenous ligand of TRPV1 channels that behaves also an endocannabinoid. The overlap between the ligand recognition properties of some TRP channels and proteins of the endocannabinoid system, namely the cannabinoid receptors and the proteins and enzymes catalyzing anandamide cellular re-uptake and hydrolysis, is being actively explored through the rational design and synthesis of new endocannabinoid- based drugs with multiple mechanisms of action. These aspects are discussed in this review article, together with the possible functional and pharmacological consequences of endocannabinoid-TRP channel interactions.

Peroxisome proliferator-activated receptors (PPARs) have long been known as mediators of several physiological functions, among which the best characterized are lipid metabolism, energy balance and anti-inflammation. Their rather large and promiscuous ligand binding site has been recently discovered to accommodate, among a plethora of lipid molecules and metabolic intermediates, endocannabinoids and their cognate compounds, specifically belonging to the Nacylethanolamine group. In fact, oleoylethanolamide, palmitoylethanolamide and probably anandamide bind with relatively high affinity to PPARs and have now been included among their endogenous ligands. Through activation of PPARs these molecules exert a variety of physiological processes. Particularly, both long-term effects via genomic mechanisms and rapid non-genomic actions have been described, which in several instances are opposite to those evoked by activation of “classical” surface cannabinoid receptors. In this review, we describe how these effects are relevant under diverse physiological and pathophysiological circumstances, such as lipid metabolism and feeding behaviour, neuroprotection and epilepsy, circadian rhythms, addiction and cognition. A picture is emerging where nuclear receptors are involved in anorexiant, anti-inflammatory, neuroprotective, anti-epileptic, wakefulness- and cognitive-enhancing, and anti-addicting properties of endocannabinoid-like molecules. Further studies are necessary to fully understand cellular mechanisms underlying the interactions between endocannabinoids and PPARs, but also between their surface and nuclear receptors, and to exploit their potential therapeutic applications.

The cannabinoid CB1 and CB2 receptors are Class A G protein-coupled receptors (GPCRs). While many Class A GPCRs have endogenous ligands that are hydrophilic cations (e.g., the serotonin and dopamine receptors), the cannabinoid receptors have neutral, highly lipophilic ligands derived from the fatty acid, arachidonic acid. The most well-studied of these are N-arachidonoylethanolamine (anandamide, AEA) and sn-2-arachidonoylglycerol (2-AG). This review focuses on the experimental and computational studies that have been used to probe the nature of endocannabinoid interaction with the cannabinoid receptors. These studies include mutation, SAR and NMR studies, as well as, QSAR, docking and molecular dynamics simulations. Gaps in our knowledge are identified. The review begins more generally, however, by discussing the entire endocannabinoid system, of which the cannabinoid receptors are part. For in order to understand endocannabinoid action, one needs an appreciation for the environments for which these ligands have been designed and the conformational changes these ligands must undergo in order to act on the cannabinoid receptors.

Cellular signaling is regulated by several biochemical reactions, whose dynamics depends on changes in the fluxes of specific ligands through the containment barriers that are the biological membranes. The regulation of this complex dynamic equilibrium is mainly due to the activity of border proteins, that must be able to interact simultaneously with the lipid bilayer and the extracellular milieu. Endocannabinoid receptors, that include type-1 and type-2 cannabinoid receptors, the transient vanilloid potential receptors and the peroxisome proliferator-activated receptors, represent one of the most intriguing examples of “border” proteins. They have also been identified as important drug discovery targets with potential therapeutic applications, including antiemesis, appetite enhancement, analgesia, glaucoma treatment, and immune suppression. However, as yet the molecular details of endocannabinoid receptor regulation remain elusive. In this review we summarize the most relevant aspects of the structural/functional characterization of these receptors, with a focus on the active role played by biological membranes (in particular lipid rafts) in the modulation of their accessibility and mode of ligand binding. Based on available evidence, we propose that endocannabinoid receptors can be regulated by the rate of interlayer exchange and lateral diffusion of endocannabinoid/cholesterol complexes within lipid bilayers, thus suggesting innovative approaches for the therapeutic exploitation of the membrane component of endocannabinoid signaling.