Design, characterization, and first-in-human study of the vascular actions of a novel biased apelin receptor agonist

Abstract

[Pyr(1)]apelin-13 is an endogenous vasodilator and inotrope but is downregulated in pulmonary hypertension and heart failure, making the apelin receptor an attractive therapeutic target. Agonists acting at the same G-protein-coupled receptor can be engineered to stabilize different conformational states and function as biased ligands, selectively stimulating either G-protein or β-arrestin pathways. We used molecular dynamics simulations of apelin/receptor interactions to design cyclic analogues and identified MM07 as a biased agonist. In β-arrestin and internalization assays (G-protein-independent), MM07 was 2 orders of magnitude less potent than [Pyr(1)]apelin-13. In a G-protein-dependent saphenous vein contraction assay, both peptides had comparable potency (pD2:[Pyr(1)]apelin-13 9.93±0.24; MM07 9.54±0.42) and maximum responses with a resulting bias for MM07 of ≈350- to 1300-fold for the G-protein pathway. In rats, systemic infusions of MM07 (10-100nmol) caused a dose-dependent increase in cardiac output that was significantly greater than the response to [Pyr(1)]apelin-13. Similarly, in human volunteers, MM07 produced a significant dose-dependent increase in forearm blood flow with a maximum dilatation double that is seen with [Pyr(1)]apelin-13. Additionally, repeated doses of MM07 produced reproducible increases in forearm blood flow. These responses are consistent with a more efficacious action of the biased agonist. In human hand vein, both peptides reversed an established norepinephrine constrictor response and significantly increased venous flow. Our results suggest that MM07 acting as a biased agonist at the apelin receptor can preferentially stimulate the G-protein pathway, which could translate to improved efficacy in the clinic by selectively stimulating vasodilatation and inotropic actions but avoiding activating detrimental β-arrestin-dependent pathways.

Keywords: [Pyr1]apelin-13; pulmonary arterial hypertension; β-arrestin G-protein coupled receptors.

Figure 1.

Representation of the overlap between the calculated poses of apelin-13 (red) and MM07 (blue) binding to the apelin receptor. The disulphide bond of MM07 is indicated in stick, the side chains in lines, and the backbone in cartoon representation. The transmembrane helices of the apelin receptor are indicated by roman numerals.

Figure 2.

Comparison of in vitro functional activity of [Pyr¹]apelin-13 () and MM07 (●) in G-protein-independent (A) β-arrestin recruitment and (B) receptor internalization assays and G-protein-dependent (C) saphenous vein contraction assay.

Figure 3.

Response to [Pyr¹]apelin-13 (, n=6) and MM07 (●, n=5) in anaesthetized rats showing changes in cardiac output (CO; A), change in blood pressure (BP) from baseline (B), heart rate (HR; C), and respiratory rate (RR; D). Echocardiograms showing an increase in peak velocity across the left ventricular outflow tract after bolus administration of [Pyr¹]apelin-13 (E) and MM07 (F). Horizontal line indicates baseline (BL) peak velocity. Significantly different from baseline *P<0.05, **P<0.01 or [Pyr¹]apelin-13 #P<0.05 (ANOVA).

Figure 4.

Absolute change in forearm blood flow (FBF) during intrabrachial infusion of [Pyr¹]apelin-13 (A) and MM07 (B). C, Repeated doses of MM07 produce a significant and reproducible response to 100 nmol/min (P<0.05). D, Concentration-dependent dilatation in dorsal hand veins after infusions of [Pyr¹]apelin-13, MM07, and saline control. Significantly different from baseline *P<0.05, **P<0.01 (ANOVA).