A Selective Antagonist Reveals a Potential Role of G Protein–Coupled Receptor 55 in Platelet and Endothelial Cell Function

Abstract

The G protein-coupled receptor 55 (GPR55) is a lysophosphatidylinositol (LPI) receptor that is also responsive to certain cannabinoids. Although GPR55 has been implicated in several (patho)physiologic functions, its role remains enigmatic owing mainly to the lack of selective GPR55 antagonists. Here we show that the compound CID16020046 ((4-[4-(3-In yeast cells expressing human GPR55, CID16020046 antagonized agonistinduced receptor activation. In human embryonic kidney (HEK293) cells stably expressing human GPR55, the compound behaved as an antagonist on LPI-mediated Ca 21 release and extracellular signal-regulated kinases activation, but not in HEK293 cells expressing cannabinoid receptor 1 or 2 (CB 1 or CB 2 ). CID16020046 concentration dependently inhibited LPI-induced activation of nuclear factor of activated T-cells (NFAT), nuclear factor k of activated B cells (NF-kB) and serum response element, translocation of NFAT and NF-kB, and GPR55 internalization. It reduced LPI-induced wound healing in primary human lung microvascular endothelial cells and reversed LPI-inhibited platelet aggregation, suggesting a novel role for GPR55 in platelet and endothelial cell function. CID16020046 is therefore a valuable tool to study GPR55-mediated mechanisms in primary cells and tissues.

Structure-Activity Relationship of the GPR55 Antagonist, CID16020046

“…ML193 also blocks pro-nociceptive effects of LPI and LPI-evoked depolarisation of neurons from a brain region, the periaqueductal grey [15], reinforcing previous data linking GPR55 to pain perception [16,17]. CID16020046 has been used to understand the role of GPR55 in the cardiovascular system [12,18,19], in atherosclerosis and intestinal inflammation [20][21][22], and in brain [23]. CID16020046 has extended the association of GPR55 to cancer, blocking LPI-evoked angiogenesis in ovarian carcinoma and migration of breast cancer cells [24,25].…”

“…We previously described CID16020046 [12], which is identical to 6 except that the phenyl ring (position C) is replaced with 4-methylphenyl. CID16020046 was independently identified in an antagonist screen utilising U2OS (Human Osteosarcoma) cells stably expressing GFP-tagged β-arrestin and over-expressing hGPR55, measuring agonist-induced β-arrestin translocation by imaging [38].…”

“…The yeast gene-reporter assay and derivation of yeast strains were described previously [12]. In brief, yeast strain YIG150 contains human GPR55 (hGPR55) and Gpa1/G αi3 G-protein α-subunit, both integrated chromosomally.…”

“…Hence, an informative approach has been to study the effects of LPI on cells and tissues in combination with synthetic GPR55 antagonists identified from screening. ML193 and CID16020046 are antagonists from distinct structural classes that have found widespread use in studies of GPR55 function [11,12]. ML193 blocks LPI-evoked calcium transients in cultured neonatal cardiomyocytes [13].…”

GPR55: a new promising target for metabolism?

“…However, it has also been reported that Rimonabant is capable of blocking the GPR55 signaling responses induced by other agonist compounds such as JWH015 and AEA, which for instance trigger intracellular Ca 2+ transients (Lauckner et al 2008, WaldeckWeiermair et al 2008. CID-16020046 is a recently synthesized compound that selectively inhibits GPR55 without inducing any effect via CB1 or CB2, in both yeast and HEK293 cells expressing human GPR55 (Kargl et al 2013). In addition, a β2-adrenergic agonist, (R,R′)-40-methoxy-1-naphthylfenoterol (AMF), has been described as a potent GPR55 inhibitor, blocking both O-1602-and AM251-induced GPR55 signaling in cancer cell lines (Paul et al 2014) and reducing the chemoresistance of carcinogenic cells (Singh et al 2016).…”