A potential therapeutic role for angiotensin-converting enzyme 2 in human pulmonary arterial hypertension

Abstract

Pulmonary arterial hypertension (PAH) is a deadly disease with no cure. Alternate conversion of angiotensin II (AngII) to angiotensin-(1-7) (Ang-(1-7)) by angiotensin-converting enzyme 2 (ACE2) resulting in Mas receptor (Mas1) activation improves rodent models of PAH. Effects of recombinant human (rh) ACE2 in human PAH are unknown. Our objective was to determine the effects of rhACE2 in PAH.We defined the molecular effects of Mas1 activation using porcine pulmonary arteries, measured AngII/Ang-(1-7) levels in human PAH and conducted a phase IIa, open-label pilot study of a single infusion of rhACE2 (GSK2586881, 0.2 or 0.4 mg·kg-1 intravenously).Superoxide dismutase 2 (SOD2) and inflammatory gene expression were identified as markers of Mas1 activation. After confirming reduced plasma ACE2 activity in human PAH, five patients were enrolled in the trial. GSK2586881 was well tolerated with significant improvement in cardiac output and pulmonary vascular resistance. GSK2586881 infusion was associated with reduced plasma markers of inflammation within 2-4 h and increased SOD2 plasma protein at 2 weeks.PAH is characterised by reduced ACE2 activity. Augmentation of ACE2 in a pilot study was well tolerated, associated with improved pulmonary haemodynamics and reduced markers of oxidant and inflammatory mediators. Targeting this pathway may be beneficial in human PAH.

Trial registration: ClinicalTrials.gov NCT01884051.

Conflict of interest statement

Conflict of interest: A.R. Hemnes reports grants from the National Institutes of Health (NIH) and the Cardiovascular Medical Research and Education Fund, personal fees from Actelion, Bayer and United Therapeutics, and personal fees and nonfinancial support (drug supplies to perform experiments) from GlaxoSmithKline, during the conduct of the study. In addition, she has a patent Annamometer issued (noninvasive diagnosis of pulmonary vascular disease). E.A. Austin has received grants from the NIH. He has served as a consultant to Acceleron. E.L. Brittain has received grants from the NIH. He has served as a consultant to Hovine Pharmaceuticals. E.J. Carrier has received grants from the NIH. She has intellectual property pending with Cumberland Pharmaceuticals and has received a grant from this entity. J.P. Fessel has received grant support from the NIH, Gilead, Actelion, the LAM foundation and the Parker B. Francis Foundation. C.D. Fike is a co-inventor on patent applications filed on behalf of Vanderbilt University in the USA and abroad and licensed to Asklepion Pharmaceuticals for the use of intravenous citrulline in lung conditions, without current financial compensation. J.H. Newman has received grants from the NIH. M.E. Pugh has received grant funding from the NIH and has served as a consultant to Gilead. T.W. Rice has received grant funding from the NIH. He has served as a consultant to Avisa Pharma, ACI Clinical and Nestle. He is the Director of Medical Affairs at Cumberland Pharmaceuticals. I.M. Robbins has received funding from the NIH. J.E. Loyd has received grants from the NIH. J. West has received grants from the NIH and Cumberland Pharmaceuticals, and holds intellectual property without financial gain at Cumberland Pharmaceuticals.

Copyright ©ERS 2018.

Figures

Figure 1.. Evidence of insufficient ACE2 activity in human PAH

Plasma Angiotensin II (AngII) and Angiotensin-(1–7) levels were measured in 11 PAH patients and 8 healthy controls as a marker of ACE2 activity. Log transformed values are normally distributed by Shapiro-Wilk W Test for each variable and the ratio, allowing parametric testing. AngII was increased in PAH (*p

Figure 2.. Acute molecular effects of Mas1 activation in a porcine pulmonary hypertension model

AVEO991, a direct Mas receptor agonist, was administered to pig arteries pre-constricted with ET-1. A. There was a dose-dependent increase in percent dilation with AVE0991 administration. No dilation was seen with exposure to a biologically irrelevant control compound. B. RNA was isolated from pig arteries with and without AVE0991 exposure and RNASequencing performed. Gene ontology analysis demonstrated significant differences in gene expression in several pathways including inflammatory responses (p<1×10−4), cell and leukocyte migration (p<1×10−4) and pressure regulation (p<1×10−5). C. When absolute change in gene expression with drug exposure was plotted as dependent on expression level in control arteries, SOD2 expression was strongly up-regulated in response to Mas1 activation. N=8 vessels.

Figure 3.. Pulmonary and systemic hemodynamic effects of intravenous GSK2586881

A. A pulmonary artery catheter was placed and hemodynamics measured (−1H). Just prior to GSK2586881 exposure, pulmonary hemodynamics were recorded again (0) and then 1 hour (1H), 2 hours (2H) and 4 hours (4H) after drug administration, n=5. There was no significant change in mean pulmonary arterial pressure. B. There was a significant increase in cardiac output 2 hours after drug administration (*p=0.008). C-F. Individual raw data on pulmonary vascular resistance, systolic blood pressure, diastolic blood pressure and heart rate. G. Percent (%) change in pulmonary vascular resistance (PVR) showed a significant decrease when comparing time points before drug administration (−1H and 0) with those after (1H, 2H and 4H), # = p<0.05. H. Angiotensin II (AngII) and Angiotensin-(1–7) (Ang-(1–7)) levels were measured just prior to drug (0) and 2 hours (2H) and 4 hours (4H) after drug exposure. In the 3 patients with elevation in this ratio at baseline, there was a significant decrease in the ratio, suggesting effective ACE2 augmentation with administration of GSK2586881 (p=0.009 and 0.008 vs. 0 at 2H and 4H).

Figure 4.. Effect of GSK2586881 on markers of oxidant stress

A. Plasma SOD2 protein levels were measured by ELISA assay at just prior to drug administration (0), and 2 hour (2 hr), 4 hour (4 hr), 24 hours (24 hr) and 2 weeks (2 wk) after drug administration, n=5. There is a significant increase in plasma SOD2 levels at two weeks (*p=0.009). B. We measured plasma isoprostanes and isofurans as markers of oxidant stress. Isofurans are lipid peroxidation products similar to isoprostanes, but are formed in times of oxygen availability. There was a decrease in the isoprostanes:isofuran ratio (*p<0.05), suggesting improved oxygenation. C. Nitrotyrosine dot blot assay from the plasma of 5 healthy controls and study enrollees (PAH) and quantification of densitometry D. In PAH patients, there was a decrease in plasma nitrotyrosine levels to that of controls by four hours (p=0.004) that was sustained at 24 hours (p=0.02). Please see Online Supplement Figure 1 for full original blot.

Figure 5.. Schematic of study findings.

PAH: pulmonary arterial hypertension; AngII: angiotensin II; Ang-(1–7): angiotensin-(1–7); SOD2: superoxide dismutase 2; IP: isoprostane; IF: isofuran; IL: interleukin; TNF: tumour necrosis factor.