Abstract
Signaling through the apelin receptor is beneficial for a number of diseases including pulmonary arterial hypertension. The endogenous small peptides, apelin and elabela/toddler, are downregulated in pulmonary arterial hypertension but are not suitable for exogenous administration owing to a lack of bioavailability, proteolytic instability and susceptibility to renal clearance. CMF-019, a small molecule apelin agonist that displays strong bias towards G protein signaling over β-arrestin (∼400 fold), may be more suitable. This study demonstrates that in addition to being a positive inotrope, CMF-019 caused dose-dependent vasodilatation in vivo (50 nmol 4.16 ± 1.18 mmHg, **p < 0.01; 500 nmol 6.62 ± 1.85 mmHg, **p < 0.01), without receptor desensitization. Furthermore, CMF-019 rescues human pulmonary artery endothelial cells from apoptosis induced by tumor necrosis factor α and cycloheximide (5.66 ± 0.97%, **p < 0.01) by approximately 50% of that observable with rhVEGF (11.59 ± 1.85%, **p < 0.01), suggesting it has disease-modifying potential in vitro. CMF-019 displays remarkable bias at the apelin receptor for a small molecule and importantly recapitulates all aspects of the cardiovascular responses to the endogenous ligand, [Pyr1]apelin-13, in vivo. Additionally, it is able to protect human pulmonary artery endothelial cells from apoptosis, suggesting that the beneficial effects observed with apelin agonists extend beyond hemodynamic alleviation and address disease etiology itself. These findings support CMF-019 as a G protein biased small molecule apelin agonist in vitro and in vivo that could form the basis for the design of novel therapeutic agents in chronic diseases, such as, pulmonary arterial hypertension.
Highlights
The apelin receptor, a class A G protein coupled receptor (O’Dowd et al, 1993) has two endogenous peptide ligands, apelin (Tatemoto et al, 1998) and elabela/toddler (ELA) (Chng et al, 2013; Pauli et al, 2014)
In this study we have investigated whether CMF-019 alters apoptosis in human pulmonary arterial endothelial cells (PAECs), a driver of early disease phase (Wilson et al, 1992; Rabinovitch 2012)
tumor necrosis factor α (TNFα)/ CHX significantly increased the percentage of annexin+/PI− cells (19.54 ± 1.76%, ####p < 0.0001) compared to the endothelial basal medium 2 (EBM-2) 2% FBS control and recombinant human vascular endothelial growth factor (rhVEGF) was able to partially rescue this (11.59 ± 1.85%, **p < 0.01), as was CMF-019 at 1 μM (5.66 ± 0.97%, **p < 0.01)
Summary
The apelin receptor, a class A G protein coupled receptor (O’Dowd et al, 1993) has two endogenous peptide ligands, apelin (Tatemoto et al, 1998) and elabela/toddler (ELA) (Chng et al, 2013; Pauli et al, 2014). The apelin system is a therapeutic target in diseases (Read et al, 2019), such as diabetes (Castan-Laurell et al, 2012), fibrosis (Huang et al, 2016), heart failure (Berry et al, 2004; Jia et al, 2006; Atluri et al, 2007; Koguchi et al, 2012; Pang et al, 2014) and pulmonary arterial hypertension (PAH) (Falcão-Pires et al, 2009; Yang et al, 2015). CMF-019 has nanomolar affinity for the apelin receptor in both human and rat heart and whereas this compound inhibits Gαi mediated cAMP accumulation with sub-nanomolar potency comparable to [Pyr1]apelin-13 it is over two orders of magnitude less efficient in recruiting β-arrestin or inducing apelin receptor internalization compared to the endogenous agonist (Read et al, 2016). We aimed to confirm that a G protein biased compound produces vasodilatation in vivo, (the main mechanism of action for most current PAH therapies) without desensitization, as recent studies have suggested that apelin-mediated vasodilatation may occur via β-arrestin signaling (El Messari et al, 2004; Iturrioz et al, 2010; Ceraudo et al, 2014)
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