Pyrimidinyl Biphenylureas Act as Allosteric Modulators to Activate Cannabinoid Receptor 1 and Initiateβ-Arrestin–Dependent Responses

Abstract

Cannabinoid receptor 1 (CB 1) is a G-protein-coupled receptor that is abundant in the central nervous system. It binds several compounds in its orthosteric site, including the endocannabinoids, arachidonoyl ethanolamide (anandamide) and 2-arachidonoyl glycerol, and the plant-derived D 9-tetrahydrocannabinol, one of the main psychoactive components of marijuana. It primarily couples to G i/o proteins to inhibit adenylate cyclase activity and typically induces downstream signaling that is G idependent. Since this receptor is implicated in several maladies, such as obesity, pain, and neurodegenerative disorders, there is interest in developing therapeutics that selectively target this receptor. Allosteric modulators of CB 1 offer one new approach that has tremendous therapeutic potential. Here, we reveal receptor-and cellular-level properties consistent with receptor activation by a series of pyrimidinyl biphenylureas (LDK1285, LDK1288, LDK1305, and PSNCBAM1), including promoting binding of the agonist CP55940 with positive cooperativity and inhibiting binding of the inverse agonist SR141716A with negative cooperativity, demonstrated via radioligand binding studies. Consistent with these findings, the allosteric modulators induced cellular internalization of the receptor and recruitment of b-arrestin 2 in human embryonic kidney cell line 293 cells monitored with confocal and total internal reflective fluorescence microscopy, respectively. These allosteric modulators, however, caused G-protein-independent but b-arrestin 1-dependent phosphorylation of the downstream kinases extracellular signal-regulated kinase 1/2, mitogen-activated protein kinase, and Src, shown by immunoblotting studies. These results are consistent with the involvement of b-arrestin and suggest that these allosteric modulators induce biased signaling.

Discovery of Soluble Epoxide Hydrolase Inhibitors from Chemical Synthesis and Natural Products

“…Here, we are mostly concentrating on the urea-based sEH inhibitors. Because urea function is also the central pharmacophore for compounds that inhibit proteases and kinases 170 and bind to canabinoid receptors 1, 171 it is possible that inhibitors for sEH could also alter the function of these other proteins. It is interesting that the Raf-1 kinase inhibitor sorafenib (80), used to treat some cancer, was found to inhibit sEH also.…”

Structural Insights into CB1 Receptor Biased Signaling

“…These new compounds positively modulate the binding of the CB1 orthosteric agonist CP55,940 while exhibiting an antagonism of G-protein coupling. Remarkably, the pyrimidinyl biphenylureas demonstrated ERK1/2 phosphorylation, mediated by β-arrestin1 [134,135]. Therefore, these allosteric modulators may help stabilize a CB1 active conformation that binds to β-arrestin1, while precluding G-protein coupling.…”

Utilization of Biased G Protein-Coupled Receptor Signaling towards Development of Safer and Personalized Therapeutics

“…In addition, allosteric modulators not only fine-tune receptor signaling but also attenuate risk of overdosing, which is relevant to saturation [25,73,74,75]. For instance, allosteric regulation of cannabinoid receptor 1 (CB 1 ) enables more precise control of downstream pathways [76]. Moreover, insurmountable character of allosteric ligands is also important, which is known as ability of allosteric ligand to decrease the potency and/or efficacy of the endogenous agonist even under high concentration [77].…”