title: [Pyr1]Apelin-13(1-12)Is a Biologically Active ACE2 Metabolite of the Endogenous Cardiovascular Peptide [Pyr1]Apelin-13.
abstract: Aims:Apelin is a predicted substrate for ACE2, a novel therapeutic target. Our aim was to demonstrate the endogenous presence of the putative ACE2 product [Pyr1]apelin-13(1-12)in human cardiovascular tissues and to confirm it retains significant biological activity for the apelin receptorin vitroandin vivo. The minimum active apelin fragment was also investigated.Methods and Results:[Pyr1]apelin-13 incubated with recombinant human ACE2 resulted in de novo generation of [Pyr1]apelin-13(1-12)identified by mass spectrometry. Endogenous [Pyr1]apelin-13(1-12)was detected by immunostaining in human heart and lung localized to the endothelium. Expression was undetectable in lung from patients with pulmonary arterial hypertension. In human heart [Pyr1]apelin-13(1-12)(pKi= 8.04 ± 0.06) and apelin-13(F13A) (pKi= 8.07 ± 0.24) competed with [125I]apelin-13 binding with nanomolar affinity, 4-fold lower than for [Pyr1]apelin-13 (pKi= 8.83 ± 0.06) whereas apelin-17 exhibited highest affinity (pKi= 9.63 ± 0.17). The rank order of potency of peptides to inhibit forskolin-stimulated cAMP was apelin-17 (pD2= 10.31 ± 0.28) > [Pyr1]apelin-13 (pD2= 9.67 ± 0.04) ≥ apelin-13(F13A) (pD2= 9.54 ± 0.05) > [Pyr1]apelin-13(1-12)(pD2= 9.30 ± 0.06). The truncated peptide apelin-13(R10M) retained nanomolar potency (pD2= 8.70 ± 0.04) but shorter fragments exhibited low micromolar potency. In a β-arrestin recruitment assay the rank order of potency was apelin-17 (pD2= 10.26 ± 0.09) >> [Pyr1]apelin-13 (pD2= 8.43 ± 0.08) > apelin-13(R10M) (pD2= 8.26 ± 0.17) > apelin-13(F13A) (pD2= 7.98 ± 0.04) ≥ [Pyr1]apelin-13(1-12)(pD2= 7.84 ± 0.06) >> shorter fragments (pD2< 6). [Pyr1]apelin-13(1-12)and apelin-13(F13A) contracted human saphenous vein with similar sub-nanomolar potencies and [Pyr1]apelin-13(1-12)was a potent inotrope in paced mouse right ventricle and human atria. [Pyr1]apelin-13(1-12)elicited a dose-dependent decrease in blood pressure in anesthetized rat and dose-dependent increase in forearm blood flow in human volunteers.Conclusions:We provide evidence that ACE2 cleaves [Pyr1]apelin-13 to [Pyr1]apelin-13(1-12)and this cleavage product is expressed in human cardiovascular tissues. We have demonstrated biological activity of [Pyr1]apelin-13(1-12)at the human and rodent apelin receptorin vitroandin vivo. Our data show that reported enhanced ACE2 activity in cardiovascular disease should not significantly compromise the beneficial effects of apelin based therapies for example in PAH.
authors.affiliation: Department of Medicine, Experimental Medicine and Immunotherapeutics, University of Cambridge Cambridge, UK.
authors.forename: Peiran
authors.initials: P
authors.lastname: Yang
authors.affiliation: Department of Medicine, Experimental Medicine and Immunotherapeutics, University of Cambridge Cambridge, UK.
authors.forename: Rhoda E
authors.initials: RE
authors.lastname: Kuc
authors.affiliation: Department of Medicine, Experimental Medicine and Immunotherapeutics, University of Cambridge Cambridge, UK.
authors.forename: Aimée L
authors.initials: AL
authors.lastname: Brame
authors.affiliation: Division of Medicine, Bloomsbury Institute of Intensive Care Medicine, University College London London, UK.
authors.forename: Alex
authors.initials: A
authors.lastname: Dyson
authors.affiliation: Division of Medicine, Bloomsbury Institute of Intensive Care Medicine, University College London London, UK.
authors.forename: Mervyn
authors.initials: M
authors.lastname: Singer
authors.affiliation: Department of Chemistry, Centre for Molecular Informatics, University of CambridgeCambridge, UK; Department of Surgery and Cancer, Biomolecular Medicine, Imperial College LondonLondon, UK.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Glen
authors.affiliation: Department of Medicine, Experimental Medicine and Immunotherapeutics, University of Cambridge Cambridge, UK.
authors.forename: Joseph
authors.initials: J
authors.lastname: Cheriyan
authors.affiliation: Department of Medicine, Experimental Medicine and Immunotherapeutics, University of Cambridge Cambridge, UK.
authors.forename: Ian B
authors.initials: IB
authors.lastname: Wilkinson
authors.affiliation: Department of Medicine, Experimental Medicine and Immunotherapeutics, University of Cambridge Cambridge, UK.
authors.forename: Anthony P
authors.initials: AP
authors.lastname: Davenport
authors.affiliation: Department of Medicine, Experimental Medicine and Immunotherapeutics, University of Cambridge Cambridge, UK.
authors.forename: Janet J
authors.initials: JJ
authors.lastname: Maguire
keywords: ACE2; [Pyr1]apelin-13(1–12); apelin; apelin receptor; biased signaling; forearm plethysmography; human heart; pulmonary arterial hypertension