Several studies have found that GPR possesses a distinct pha
Several studies have found that GPR55 possesses a distinct pharmacology from that of CB1 and CB2, and that only certain cannabinoids act at this receptor. In transfected A 77-01 receptor expressing hGPR55, out of more than 20 synthetic and endogenous cannabinoids evaluated, only 2 compounds, SR141716A (Rimonabant) and AM251, both CB1 receptor antagonists/inverse agonists, activated GPR55 in four distinct signaling assays: protein kinase CβII activation, β-arrestin recruitment, receptor internalization and ERK1/2 activation (Kapur et al., 2009). LPI was also an agonist for GPR55 (Kapur et al., 2009). It is important to note that the high content imaging β-arrestin recruitment assay is unbiased with respect to G protein coupling (Milligan, 2011) and that this data has been reproduced by other laboratories (Henstridge et al., 2009, Henstridge et al., 2010). Interestingly, the synthetic potent CB1/CB2 receptor agonist, CP55, 940, acted as an antagonist in the β-arrestin, internalization and ERK1/2 assays and as a partial agonist in the protein kinase CβII assays (Kapur et al., 2009). Thus it is tempting to speculate that GPR55 may in a sense represent an “anti-cannabinoid” receptor as its pharmacology is somewhat opposite that of CB1. Continued characterization of GPR55 has shown that while it shares some ligands with CB1 and CB2 cannabinoid receptors, distinct chemical classes which do not recognize CB1 and CB2 are ligands for GPR55. In a library screen of 290,000 compounds (using the β-arrestin recruitment assay as a primary screen and the hGPR55 cell line) performed in the NIH Molecular Libraries program (by the Sanford-Burnham Center for Chemical Genomics), three new GPR55 agonist scaffolds were identified: ML184 (CID2440433) (3-[4-(2,3-dimethylphenyl)piperazine-1-carbonyl]-N,N-dimethyl-4-pyrrolidin-1-ylbenzenesulfon-amide); ML185 (CID1374043) (N-[4-(4-methoxyphenyl)-1,3-thiazol-2-yl]-2-[(4-methyl-[1,2,4]triazolo-[4,3-a]quinolin-1-yl)sulfanyl]acetamide) and ML186 (CID15945391) (N-(2-methoxy-5-morpholin-4-ylsulfonylphenyl)-4,5,6,7-tetrahydro-1-benzothiophene-2-carboxamide) (Heynen-Genel et al., 2010a). The EC50s of these compounds at GPR55 are as follows: ML184, 263 nM; ML185, 658 nM; and ML186, 305 nM. All compounds were counter-screened for both agonism and antagonism at GPR35, CB1, and CB2; they were found to have EC50s>32μM at these other targets. These compounds were confirmed as agonists in secondary assays (activation of mitogen activated protein kinase ERK1/2, in the Abood laboratory) (Heynen-Genel et al., 2010a). Interestingly, a similar structure to ML184 (CID2440433) was independently discovered in a screen conducted by Glaxo Smith Kline (GSK494581A, EC50 of 158 nM). The interaction of these compounds with GPR55 was examined in a recently derived molecular model of GPR55 (Kotsikorou et al., 2011). Interestingly, the agonist compounds resemble LPI and not cannabinoid ligands in their interaction with GPR55 in the molecular model (Kotsikorou et al., 2011). Mutational studies have confirmed that K2.60 is a crucial residue for agonist activation of GPR55; K2.60 was identified as a critical residue for binding interactions for agonists in the GPR55 active state model (Kotsikorou et al., 2011 and unpublished data). Several highly selective GPR55 antagonists have recently been identified; the three main scaffolds are: ML191 (CID23612552)(5-phenyl-3-(1-(1-(p-tolyl)cyclopropane-carbonyl)-piperidin-4-yl)-1,3,4-oxadiazol-2(3H)-one);> ML192 (CID1434953) (furan-2-yl(4-(2-methyl-5,6,7,8-tetrahydro-benzo-[4,5]thieno[2,3-d]pyri-midin-4-yl)piperazin-1-yl)methanone) and ML193 (CID1261822) (N-(4-(N-(3,4-dimethylisoxazol-5yl)sulfamoyl)-phenyl)-6,8-dimethyl-2-(pyridin-2-yl)quinoline-4-carboxamide) (Heynen-Genel et al., 2010b). ML191 has 1080nM potency for GPR55 and >100-fold selectivity against GPR35, CB1 and CB2. ML192 has 702nM potency for GPR55 and >45-fold antagonist and agonist selectivity against GPR35, CB1 and CB2. ML193 has a 221nM potency for GPR55 and >145-fold, >27-fold and >145-fold antagonist selectivity against GPR35, CB1 and CB2, respectively. However, to date, no low nanomolar potency ligands have been discovered for GPR55 nor are there radioligands developed to characterize binding at this receptor. The lack of such GPR55 ligands is a current barrier to progress in this field, as testes is not possible to determine ligand binding properties for this receptor.