Pediatr Nephrol (2013) 28:711–720 DOI 10.1007/s00467-012-2316-4

EDUCATIONAL REVIEW

Endothelin antagonists in hypertension and kidney disease Kevin E. C. Meyers & Christine Sethna

Received: 12 July 2012 / Revised: 20 August 2012 / Accepted: 21 August 2012 / Published online: 16 October 2012 # IPNA 2012

Abstract The endothelin (ET) system seems to play a pivotal role in hypertension and in proteinuric kidney disease, including the micro- and macro-vascular complications of diabetes. Endothelin-1 (ET-1) is a multifunctional peptide that primarily acts as a potent vasoconstrictor with direct effects on systemic vasculature and the kidney. ET-1 and ET receptors are expressed in the vascular smooth muscle cells, endothelial cells, fibroblasts and macrophages in systemic vasculature and arterioles of the kidney, and are associated with collagen accumulation, inflammation, extracellular matrix remodeling, and renal fibrosis. Experimental evidence and recent clinical studies suggest that endothelin receptor blockade, in particular selective ETAR blockade, holds promise in the treatment of hypertension, proteinuria, and diabetes. Concomitant blockade of the ETB receptor is not usually beneficial and may lead to vasoconstriction and salt and water retention. The side-effect profile of ET receptor antagonists and relatively poor antagonist selectivity for ETA receptor are limitations that need to be addressed. This review will discuss what is currently known about the endothelin system, the role of ET-1 in the pathogenesis of hypertension and kidney disease, and summarize literature on the therapeutic potential of endothelin system antagonism.

K. E. C. Meyers (*) Nephrology Division, Department of Pediatrics, The Children’s Hospital of Philadelphia, 1st Floor Main Building, 34th and Civic Center Boulevard, Philadelphia, PA 19104, USA e-mail: [email protected] C. Sethna Division of Nephrology, Department of Pediatrics, Cohen Children’s Medical Center of New York, 269-01 76th Avenue, New Hyde Park, NY 11040, USA

Keywords Endothelin-1 . Endothelin-converting enzyme-1 . Endothelin A receptor . Endothelin B receptor . Chronic kidney disease

Introduction The main purpose of this review is to provide an overview of the endothelin system and its clinical relevance to hypertension and kidney disease. This review will approach the topic in two parts. In the basic science segment we will review the increasingly complex endothelin system with respect to the synthesis of endothelins, endothelin-converting enzymes, endothelin receptors, endothelin system antagonists, and the pathophysiology of endothelin in hypertension and renal disease. Of note, the major function of endothelin-1 (ET-1) is to act as a vasoconstrictor. However, it is much more than a vasoconstrictor, and as a multifunctional peptide with cytokine/hormone-like activity, ET-1 participates in a number of physiological activities, including mitogenesis, cell survival, angiogenesis, bone remodeling, epithelial to mesenchymal cell transition, and nociceptor stimulation; it also has a number of tumor-related activities. In the clinical section we will briefly review the use of endothelin receptor antagonists in pulmonary artery hypertension and will then focus on the potential use of ET-1 antagonists in the management of systemic hypertension and diabetic and nondiabetic proteinuric chronic kidney disease (CKD). Although therapeutic intervention with ET-1 antagonists has made a marked difference in the care of patients with severe pulmonary hypertension, their place in the management of systemic hypertension and use in CKD is not yet clear and continues to evolve over time.

712

Basic science: physiology and pathophysiology of endothelin-1 Endothelins Endothelin-1, one of the most potent vasoconstrictors recognized in humans to date, was initially identified and characterized from media in which there were cultured aortic endothelial cells [1]. Two additional isoforms termed endothelin-2 (ET-2) and endothelin-3 (ET-3) were subsequently identified from the venom of Actractapis engaddensis (mole viper) [2]. In humans the three major endothelin isoforms are transcribed from chromosome 6 (ET-1), chromosome 1 (ET-2) and chromosome 20 (ET-3) [3, 4]. The three ET isoforms are all 21 amino acid peptides. The differential expression of the three endothelin peptides in various tissues and cells is shown in Table 1. Of special note, ET-3, which is found in endothelial and intestinal cells, mediates the release of vasodilators, including NO and prostacyclin [5]. Transcription is considered the primary level of ET-1 regulation [6, 7]. The promoter region for transcription of the major isoform ET-1 is located upstream (5′) of the preproendothelin-1 gene. Basal transcription is regulated through a GATA binding site with gene transcription controlled by an Ap-1 nuclear factor and a hexonucleotide sequence, which in turn are regulated by chemical and mechanical means [8]. The chemical and mechanical factors known to enhance ET-1 production include angiotensin II, calcineurin inhibition, cortisol, epinephrine, hypoxia, transforming growth factor beta (TGFβ), insulin, thrombin, and vascular shear stress. In contrast, ET-1 production is inhibited by nitric oxide (NO), nitric oxide donor drugs, and vasodilator prostanoids via an increase in cGMP, and by natriuretic peptides via an increase in cAMP [8]. A peptide variant of ET-1, termed ET-11-31, also has vasopressor properties that are Table 1 Endothelin peptide expression

Pediatr Nephrol (2013) 28:711–720

reduced by endothelin receptor antagonists, abolished by endopeptidase inhibition and unaffected by the ECE-1 inhibitor CGS35066 [9]. Formation of the mature ET-1 peptide is generated from enzymatic cleavage of the initial gene product, termed preproendothelin-1 (Fig. 1). After removal of the secretory sequence the endopeptidase furin generates a 39-amino acid peptide termed big ET-1 (or proendothelin) [1]. The subsequent production of mature ET-1 is catalyzed by membranebound metalloprotease endothelin-converting enzyme-1 (ECE-1) by cleavage at a tryptophan21/valine22 site on big ET-1 [10]. Although other isoforms of ECE-1 are described in additional species, knockout studies suggest that ECE-1 is the major functional enzyme in the production of all three mature human ET peptide isoforms [11]. Of note, in response to the chemical and mechanical factors described above, mature ET-1 and ECE-1 can either be made de novo for subsequent release (through constitutive secretory vesicles) and/or they can effect immediate release of mature ET-1 and ECE-1 from stores in specialized regulatory granules that in endothelial cells are known as Weibel–Palade bodies [12]. Most ET-1 is secreted in the mature form, with about 10 % secreted in the big form. ET-1 has a half-life of less than 5 min in plasma, is primarily cleared by the lungs (80 %) and kidneys, and circulates at one tenth of the concentration of angiotensin II (AGII) and atrial natriuretic peptide (ANP), suggesting that for the most part ET-1 might have an autocrine/paracrine function [13, 14]. The implication of this is that the serum concentration of ET-1 in health and disease may be an unreliable indicator of vascular endothelin (re)activity [15]. In addition, urinary levels of ET-1 probably reflect local renal ET-1 activity, not systemic ET-1 function. Endothelin-converting enzyme-1 Endothelin-converting enzyme (ECE)-1 cleaves big endothelin(s), as well as bradykinin and beta-amyloid peptide. ECE-1 was originally identified from purified aortic endothelial cells and the gene was subsequently located on chromosome 1p36 [16]. ECE-1 belongs to the family of neutral metalloproteinase enzymes. Other proteins in this group include neutral endopeptidase (NEP) and the human Kell blood group protein. Interestingly, the Kell protein has homology with the neprilysin family of zinc endopeptidases, which are responsible for cleaving ET-3 [17]. Endothelin-converting enzyme-1 is a transmembrane dimer that is linked by a single disulphide bridge; the Nterminal intracellular portion of 56 amino acids is much shorter than the longer C-terminal segment. The longer Cterminal recognizes at least two cleavage sites on big ET-1. Through differential gene splicing, four isoforms of ECE-1 can be generated, termed ECE-1a, ECE-1b, ECE-1c, and ECE-1d, which differ only at the N-terminus. ECE-1a is

Pediatr Nephrol (2013) 28:711–720

713

Fig. 1 Formation of the mature ET-2 peptide is generated from enzymatic cleavage of the initial gene product, termed preproendothelin-1

located intracellularly with the enzymatically active Cterminus facing the Golgi apparatus; it is responsible for generating most functional ET-1. ECE-1a is constitutively expressed in endothelial cells. ECE-1b, on the other hand, spans the cell membrane of smooth muscle cells where it is able to cleave extracellular big-ET-1 to ET-1. ECE-1b presumably has a responder and regulator role in smooth muscle function. This responder/regulator role is also suggested by its promoter region, which has receptor sites for transcription regulators that allow for response modulation. Endothelin-converting enzyme-1c responds to high glucose levels, particularly in endothelial cells. ECE-1c may also play a permissive role for invasion in some forms cancer. In contrast to ECE-1b, ECE-1d is expressed at the cell surface, although less strongly than ECE-1a. In addition to full-length ECE-1 forms, alternatively spliced messenger RNAs (mRNAs) of ECE-1b, 1c, and 1d have been identified [18]. These splice variants (SVs) lack exon 3′, which codes for the transmembrane (TM) region that is present in full-length forms, and so are expected to remain cytosolic. These SVs are highly expressed in endothelial cells derived from both macrovascular and microvascular beds. ECE-1 SVs found in cellular compartments are different from the full-length forms of ECE-1 and may play a distinct physiological role [19]. Endothelin receptors The ET isoforms mediate their physiological effects through two seven-transmembrane G protein-coupled endothelin receptors, the endothelin A (ETA) receptor and endothelin B (ETB) receptor [20, 21]. The ET receptors are distinguished by their affinity for the three endothelin isoforms, where ETA is ET-1 selective and has an affinity that is ordered by ET-1>ET-2>ET-3, whereas ETB has the same

affinity for all three ET isoforms. The ET receptors have a varied distribution in tissues and cells where they are differentially expressed, establishing a multi-purpose endothelin system. ETA receptors are located on vascular smooth muscle cells (VSMCs) where their activation results in slow onset sustained vasoconstriction. In contrast, ETB receptors are located on VSMCs and on endothelial cells. Activation of ETB receptors on endothelial cells results in the release of vasoactive substances that cause VSMC relaxation and vasodilatation (NO). In addition, ETB receptors inhibit cell growth and vasoconstriction in the vascular system by clearing bound ET-1. Once engaged, the ETA receptor activates a number of G-proteins resulting in activation of protein kinase C, protein kinase A, and Gα12 to induce stress fiber formation. In contrast, the ETB receptor inhibits cyclic AMP formation, stimulates phosphoinositol hydrolysis, and binds Gα13 to induce stress fiber formation [22–26]. Endothelin system antagonists Selective ETA receptor antagonists should be beneficial as they do not reduce ETB receptor-mediated clearance of ET1- or NO-mediated vasodilatation. True selectivity of ET antagonists remains problematic though, as the currently available ETA receptor antagonists display ETB receptor antagonist activity, especially at higher doses. This explains in part the difficulties that have been experienced to date with human trials (see later). The endothelin system antagonists and their affinities are summarized in Table 2. Pathophysiology of ET in pulmonary artery hypertension Primary artery hypertension (PAH) is a condition in which there is increased vascular tone accompanied by vascular

714

Pediatr Nephrol (2013) 28:711–720

Table 2 Endothelin system antagonists ETA/ETB

ETA

ETB

ECE

TAK-044 Bosentana PD145065 L-744453 L-751281 L-754142 SB209670 Enrasentan

BQ-123 BQ-610 FR139317 IPI-725 A-127722.5 Darusentan Ambrisentan Atrasentan

BQ-788 RES −01.1 RO-468443 IRL-1620

CGS35066 Phosphoramidonc

Tezosentan Macitentan

Zibotentan Sitaxsentanb

ETA endothelin-1 A, ETB endothelin-1 receptor B, ECE endothelin converting enzyme inhibitor a

FDA approved for use in primary pulmonary hypertension

b

Withdrawn due to liver toxicity

c

Also inhibits neutral endopeptidase (NEP)

remodeling that leads to progressive right ventricular failure and eventual death. In patients with PAH, the ETA receptor, ETB receptor, and ET-1 mRNA expression are up-regulated in the lungs. Of interest, circulating levels of ET-1 do correlate with pulmonary vascular resistance, right atrial pressure, and oxygen saturation [27, 28]. In patients with PAH, selective and nonselective ETA receptor antagonists effectively reduce pulmonary artery pressure and prevent vascular remodeling. Pathophysiology of ET in systemic hypertension Involvement of ET-1 in systemic hypertension is suggested by increased ET-1 levels in the vascular wall of muscular arteries and from case reports of hemangioendothelioma patients who present with markedly elevated levels of plasma ET-1 and hypertension, but who show normalization of elevated ET-1 and BP levels after tumor removal [29]. ET-1 has growth-promoting activity in the vascular wall and both ET-1 and ET receptors are expressed in macrophages, vascular smooth muscle cells, and fibroblasts. In experimental hypertension, ETA receptor blockade prevents vascular hypertrophy and attenuates left ventricular hypertrophy [30, 31]. An increase in blood pressure is seen in ETB receptor knockout mice and with isolated ETB receptor blockade in humans [32–34]. There is changed sensitivity of the vasculature to endogenous ET-1 in patients with hypertension as there is often an exaggerated vasodilator response to ET receptor blockade that may be due in part to certain polymorphisms of the genes encoding both ET-1 and the ET receptors [35]. A common adenine insertion into the 5′–untranslated region of the ET-1 gene results in increased mRNa levels and is

associated with hypertension [36]. In contrast, it is important to note that an association with ET-1 polymorphisms is not uniformly linked to chronic hypertension [37]. There is also evidence that the antihypertensive effects and end-organ protection by ET receptor antagonists are more effective in patients with a high salt intake or who have increased angiotensin II levels [33, 38]. This is in keeping with the observation that an impaired natriuretic effect in spontaneously hypertensive rats may be, in part, related to impaired ETB and dopamine receptor interactions [39]. Pathophysiology of ET-1 in renal disease Both ETA and ETB receptors are present in the kidney, where the release of ET-1 has disparate effects on blood flow through the afferent (AA) and efferent arterioles (EA). Activation of the ETA receptor mediates vasoconstriction in the AA and contributes to vasoconstriction in the EA. In contrast, activation of the ETB receptor has no effect on the AA, but mediates basal NO release and vasodilatation in the EA. The greater vasoconstrictive effect of ET-1 on the AA supports observations of decreased glomerular filtration rate (GFR) to ET-1 release and indicates the potential contribution of ET-1 to the pathogenesis of kidney disease [40]. Infusion of ET-1 significantly decreases effective renal plasma flow, GFR, sodium excretion, and urine flow. However, the contribution of endogenous ET-1 and the ETA receptor in maintaining basal renal vascular tone in the human kidney is small. Of note, the selective ETA receptor antagonist ABT-627 is able to completely prevent all renal changes caused by ET-1 infusion [41]. Endothelin-1 has been implicated in the pathogenesis of collagen accumulation, extracellular matrix remodeling, and renal and cardiac fibrosis in diabetes. In vivo studies demonstrate that an autocrine signaling loop involving MCP-1 and IL-6 contributes to ET-1-induced collagen accumulation [42]. In vivo and in vitro studies suggest that the ETB receptor might play a role in the renal handling of salt and water, and thus in blood pressure control [43]. In the collecting duct the ETB receptor mediates natriuresis and diuresis through NO production. There is some evidence that both receptors may be needed for normal diuresis [44]. Patients with end stage renal disease (ESRD) may have an exaggerated pressor response after recombinant human erythropoietin (rhEPO) administration. The endothelin system may be involved in the pathogenesis of this “rhEPOinduced” hypertension as selective ETA, but not combined ETA/ETB receptor blockade, can prevent the exaggerated hypertensive effect of rhEPO given to rats with CKD [45]. Cyclosporin A use is a well-known cause of hypertension after transplantation. In anesthetized rats the acute hypertensive response to cyclosporine administration is not mediated through the ET system [46].

Pediatr Nephrol (2013) 28:711–720

Clinical information There is increasing evidence that the ET system plays an important role in hypertension and proteinuric renal disease, including diabetic nephropathy. Experimental evidence suggests that ETA receptor blockade might hold most promise in the treatment of proteinuric renal disease as well as in diabetic nephropathy, while the simultaneous blockade of both the ETA/ETB receptors is not desirable. This is because while ETA receptor blockade reduces blood pressure, salt and water retention, proteinuria, reactive oxygen species (ROS) production, inflammation, and fibrosis, blockade of the ETB receptor gives rise to undesirable effects, including reduced NO-mediated vasodilatation and reduced ET-1 clearance that leads directly to vasoconstriction and salt and water retention (Fig. 2). To date, clinical trials of ET receptor antagonists have been perturbed by issues associated with patient selection, questionable dosing regimens, and relatively poor antagonist selectivity for the ETA receptor. Relatively few clinical trials have been completed using ET-1 receptor antagonists, also because of the side-effect profile of this class of drugs. ET-1 antagonists like ARB and ACEi are contraindicated in pregnancy owing to teratogenic effects. They may also be hepatotoxic (Table 2), although this does not apply to all of the antagonists. In addition, ET receptor antagonists have caused testicular toxicity in some experimental animals and reduced sperm counts in some study subjects. Additional adverse events observed with ET receptor antagonist use may be linked to vasodilatation. These include nasal congestion, dizziness, edema, shortness of breath, and nausea. Hopefully, the

Fig. 2 The hemodynamic and nonhemodynamic effects of endothelin-1 receptor antagonism

715

fluid retention induced by dual ETA/ETB blockade will be eliminated and the side-effect profile minimized once highaffinity ETA blockers are available. This will, however, require identification of compounds that have at least a 1,000fold ETA:ETB selectivity in in vitro assays [47]. Currently the only clinical indications for which ET receptor antagonists are licensed in the USA are pulmonary artery hypertension (PAH) and the digital ulcers associated with scleroderma. Endothelin antagonists in PAH Endothelin receptor antagonists are now used as established therapy for the treatment of New York Heart Association (NYHA) class III and IV patients with idiopathic PAH who have had an inadequate response to vasodilator therapy. Bosentan is FDA (United States Food and Drug Administration) approved for use in these patients, as use of this agent has shown a survival advantage over placebo in clinical extension trials. Sitaxentan was recently withdrawn from the market owing to hepatotoxicity. No ET receptor antagonists are FDA (Federal Drug Administration) or EMA (European Medicines Agency) approved for use in children. The interested reader is referred to these references for further information [48–51]. Endothelin antagonists in systemic hypertension In patients with essential hypertension, nonselective ET receptor antagonists (TAK-044, bosentan) result in greater forearm vasodilatation when compared with normotensive controls. There is also evidence that salt-sensitive African–

716

Americans with essential hypertension have higher circulating ET-1 levels and increased ETA receptor-mediated vasoconstriction than normotensive controls [52]. Krum et al. showed in a 4-week treatment trial with bosentan, at a dose of 1 g given twice per day, that there was a fall in ambulatory diastolic (primary end-point) and systolic (secondary end-point) BP of approximately 10 mmHg, an effect similar to treatment with 20 mg of enalapril [53]. Recently, a phase III randomized trial using darusentan (selective ETA receptor antagonist) compared with placebo in resistant hypertension (DORADO) was conducted in 379 subjects. All subjects were on three or more antihypertensive agents, one of which was a diuretic. The study was conducted over a 14-week period and found a dose-dependent reduction in systolic (−18 mmHg) and diastolic (−11 mmHg) BP [54, 55]. In another randomized trial 72 normotensive subjects with multiple cardiovascular risk factors given atrasentan (ETA-selective antagonist) had their mean BP reduced by 12 mmHg after 6 months [56]. Compared with placebo, atrasentan decreased fasting glucose, glycosylated hemoglobin, triglyceride, lipoprotein A, and uric acid levels. Although reported sideeffects were minimal, testicular toxicity was not assessed. Thus, given the already many classes of antihypertensive agents, the clinical utility of ET receptor antagonists will at least for the present be limited to specific subsets of hypertensive patients in whom the benefits outweigh the risks. Patents who may benefit from this new class of drugs include those with resistant hypertension, metabolic syndrome, and atherosclerosis. There are no trials comparing ETA receptor antagonists with nonselective ET receptor antagonists in the above groups of patients. There are also no clinical trials in hypertensive patients using novel dual ETA receptor/ATII receptor antagonists. However, data to date suggest that ETA receptor antagonists hold therapeutic promise. Endothelin antagonists in kidney disease A number of studies have looked at the effects of ET receptor antagonists in CKD with and without associated hypertension, azotemia or proteinuria [57, 58]. The ETA receptor antagonists have the ability to reduce the BP substantially in hypertensive patients with CKD. This effect is synergistic to angiotensin-converting enzyme inhibitors (ACEi) and is abolished by significant concurrent ETB receptor blockade. Furthermore, especially in diabetic, but also in nondiabetic, renal disease patients, ET receptor antagonists often produce favorable renal hemodynamic changes that reduce proteinuria. There is also evidence in CKD that the left ventricular capillary/myocytes mismatch is prevented by use of selective ETA receptor antagonism (LU135252) [30]. Of possible clinical importance, although both ARB and ETA receptor antagonists reduce BP, vascular

Pediatr Nephrol (2013) 28:711–720

calcification is reduced by ETA receptor antagonists to a greater extent than angiotensin receptor blockade [59]. Consequently, ETA receptor antagonists hold hope of benefit in patients with CKD that extends beyond their BP-lowering effects. Acute kidney injury Endothelin-1 is a potent endogenous vasoconstrictor that through activation of ETA receptors impairs renal medullary blood flow, causing ischemia [60]. In a swine model of postcardiopulmonary bypass, acute kidney injury (AKI) was associated with endothelial dysfunction, regional tissue hypoxia, and proximal tubular epithelial cell stress, but not acute tubular necrosis. Use of the specific ETA receptor antagonist sitaxsentan reversed these changes suggesting that the ETA receptor may represent a therapeutic target for the prevention of post-cardiac surgery AKI [61]. Tezosentan reduces aortic ischemia–reperfusion AKI in rats by reducing oxidative stress, by inhibition of leukocyte infiltration into renal tissue, and through cytoprotection of renal tubular cells. This finding may be important in future prevention of postoperative AKI following abdominal aortic surgery [62]. Diabetic nephropathy The potential pathogenic role of ET-1 in renal injury in the context of insulin-resistant or -deficient states is of special importance. Insulin stimulates both the production and the action of ET-1 in experimental animals and in humans [63–65]. Insulin-stimulated ET-1 production occurs in part through phosphoinositide-3-kinase-dependent activation of glycogen synthase-3b [66]. Insulin potentiation of ET-1 action was recently shown to involve an interaction with the ATII receptor, as losartan blunted the enhanced ETA receptor-mediated vasoconstriction in insulin-treated diabetic rats [67, 68]. Such an interaction with the ATII receptor was highlighted in recent studies in diabetic rats, wherein renal damage actually regressed when avosentan and lisinopril were given together, but not separately [68]. These findings are further supported by the observation that endothelin antagonism reduced renal fibrosis in an animal model of severe angiotensin II-dependent hypertension [69]. There are experimental data indicating a specific role of ET in the pathogenesis and progression of diabetic nephropathy. ET receptor blockers have been shown to be nephroprotective in animal models of type 1 and type 2 diabetes mellitus with effects that are partly independent of lowering the blood pressure [57]. In patients with hypertension and diabetic nephropathy the data are less clear and depend on the stage of the disease and the drug used. Recently, a large international clinical study, “Avosentan on Doubling of Serum Creatinine, End Stage Renal Disease, and Death in

Pediatr Nephrol (2013) 28:711–720

Diabetic Nephropathy” (ASCEND) provided evidence for the beneficial effects of ETA receptor blockade, i.e., reduction in proteinuria. Unfortunately this trial was terminated as subjects taking avosentan experienced a high incidence of serious, sometimes life-threatening side-effects. These included complications of fluid overload, such as pulmonary edema, as well as congestive heart failure. In addition, there were more deaths in the groups taking avosentan (21 and 17) than in the group taking placebo [12]. Owing to the premature termination of the study, hard endpoints like death could not be assessed [70]. An additional study by this group involving 286 patients with diabetic nephropathy on ACEI/ARB therapy, using lower doses of avosentan, noted up to a 30 % decrease in albuminuria after 12 weeks of treatment [58]. An additional randomized, double-blind, placebo-controlled trial of subjects with diabetic nephropathy also provided evidence for a specific anti-proteinuric effect of atrasentan when added to an existing blockade of the renin–angiotensin–aldosterone system [71]. Chronic kidney disease other than diabetic nephropathy Endothelin-1 plasma concentrations increase in CKD as GFR decreases and this might contribute to hypertension [72]. Such an increase correlates with the thickening of the artery walls and left ventricular hypertrophy (LVH) [73]. These findings are consistent with (but not proof of) the pathogenic role of ET-1 in cardiac and vascular remodeling of renal failure. In addition, urinary excretion of ET-1 increases in CKD. Studies in patients with CKD suggest that ETA receptor antagonism results in up to a 10-mmHg drop in BP and that this antihypertensive effect is diminished when there is concomitant blockade of the ETB receptor [34]. In a recent double-blind, randomized crossover study using the ETA receptor antagonist BQ-123, nifedipine, and placebo in 22 subjects with proteinuric CKD, proteinuria and pulse wave velocity fell after BQ-123 exposure, but flow-mediated vasodilatation did not change. Nifedipine matched only the blood pressure and renal blood flow changes seen with BQ-123, but increased proteinuria. This suggests that selective ETA receptor antagonism in CKD patients might result in a reduction in proteinuria and arterial stiffness that is partly independent of blood pressure [74]. In keeping with the concept of an interaction with AII, a follow-up analysis by this group found that the magnitude of proteinuria reduction was greatest in patients on combined ACEI/ARB therapy [75]. Thus, although only proteinuria has been assessed to date, endothelin antagonism has exciting potential as a therapy in CKD and the use of selective ETA receptor antagonists may confer cardiovascular and renal benefits in patients with CKD. There are no longterm studies assessing the effect of endothelin antagonists on CKD progression.

717

Conclusions Endothelin-1 plays an important role in the generation and maintenance of hypertension and in CKD progression. There are a number of high-risk patients with hypertension who may benefit from this class of drugs, particularly in association with metabolic syndrome and/or atherosclerosis. There are numerous preclinical, as well as a few completed clinical, studies that suggest a substantial benefit to ETA receptor antagonism in these disorders. In order to move forward, though, there are several key issues that need to be addressed. First, the nature and mechanisms of endothelin antagonist-related adverse effects need elucidation. Second, appropriate doses of endothelin blockers need to be defined, including determination of the antagonists that are truly ETA-selective. Third, consideration must be given to which agents would best be co-administered with endothelin blockers. There are agents in development that show ETA receptor and ARBII receptor activity. Fourth, given the great promise of these drugs, the pharmaceutical industry and academia must work together to conduct longterm studies on cardiovascular and renal outcomes.

Key summary points & & &

The nature/mechanisms of ET antagonist-related adverse effects need elucidation. True ETA-selective antagonists need to be defined. Consideration must be given to which agents would best be co-administered with endothelin blockers.

Key research points & & &

Continued investigation into high-affinity ETA receptor blockers and dual receptor blockers (ETA/ATII). Further elucidation of the ET system. Pharmacia and academia must work together to conduct long-term studies on cardiovascular and renal outcomes in hypertensive patients treated with ETA receptor blockers.

Multiple choice questions (answers are provided following the reference list) 1. ET-2 production is deceased by which ONE of the following? a) b) c) d) e)

Insulin Brain natriuretic peptide Cortisol Epinephrine Angiotensin II

718

Pediatr Nephrol (2013) 28:711–720

2. Choose the ONE correct statement about the endothelin (ET) receptors ETA and ETB. a) ETA receptors are located only on vascular smooth muscle cells. b) ETB receptors are located only on vascular smooth muscle cells. c) ETA receptors are located only on endothelial cells. d) ETB receptors are located only on endothelial cells. e) ETA receptors are located on endothelial cells and vascular smooth muscle cells. 3. Most circulating ET-1 is cleared primarily by which ONE organ? a) Heart b) Kidneys c) Liver d) Lungs e) Bone marrow 4. Which ONE of the following is NOT decreased by ETA receptor blockade? a) Blood pressure b) Salt and water loss c) Proteinuria d) Inflammation e) Heart rate 5. Which ONE of the following is NOT a known sideeffect of ETA receptor blockers? a) b) c) d) e)

Hepatotoxicity Nephrotoxicity Edema formation Teratogenicity Reduced sperm count

References 1. Yanagisawa M, Kurihara H, Kimura S, Tomobe Y, Kobayashi M, Mitsui Y, Yazaki Y, Goto K, Masaki T (1988) A novel potent vasoconstrictor peptide produced by vascular endothelial cells. Nature 332:411–415 2. Kochva E, Bdolah A, Wollberg Z (1993) Sarafotoxins and endothelins: evolution, structure and function. Toxicon 31:541–568 3. Arinami T, Ishikawa M, Inoue A, Yanagisawa M, Masaki T, Yoshida M, Hamaguchi H (1991) Chromosomal assignments of the human endothelin family genes: the endothelin-1 gene (EDN1) to 6p23-p24, the endothelin-2 gene (EDN2) to 1p34, and the endothelin-3 gene (EDN3) to 20q13.2-q13.3. Am J Hum Genet 48:990–996 4. Rao V, Löffler C, Hansmann I (1991) The gene for the novel vasoactive peptide endothelin 3 (EDN3) is localized to human chromosome 20q13.2-qter. Genomics 10:840–841 5. Watanabe M, Izumi Y, Soma M, Watanabe Y, Fukuda N, Abe Y, Ito M, Kageura H, Nakayama T, Hatano M (1991) Effect of endothelin-3 (ET-3) on renal function in rat perfused kidney. Endocrinol Jpn 38:435–440

6. Todd-Turla KM, Zhu XL, Shu X, Chen M, Yu T, Smart A, Killen PD, Fejes-Toth G, Briggs JP, Schnermann JB (1996) Synthesis and secretion of endothelin in a cortical collecting duct cell line. Am J Physiol 271:F330–F339 7. Inoue A, Yanagisawa M, Takuwa Y, Mitsui Y, Kobayashi M, Masaki T (1989) The human preproendothelin-1 gene. Complete nucleotide sequence and regulation of expression. J Biol Chem 264:14954–14959 8. Stow L, Jacobs M, Wingo C, Cain B (2011) Endothelin-1 gene regulation. FASEB J 25:16–28 9. Simard E, Jin D, Takai S, Miyazaki M, Brochu I, D’Orleans-Juste P (2009) Chymase-dependent conversion of Big endothelin-1 in the mouse in vivo. J Pharmacol Exp Ther 328:540–548 10. Xu D, Emoto N, Giaid A, Slaughter C, Kaw S, deWit D, Yanagisawa M (1994) ECE-1: a membrane-bound metalloprotease that catalyzes the proteolytic activation of big endothelin-1. Cell 78:473–485 11. Yanagisawa H, Hammer RE, Richardson JA, Emoto N, Williams SC, Takeda S, Clouthier DE, Yanagisawa M (2000) Disruption of ECE-1 and ECE-2 reveals a role for endothelin-converting enzyme-2 in murine cardiac development. J Clin Invest 105: 1373–1382 12. Turner AJ, Murphy LJ (1996) Molecular pharmacology of endothelin converting enzymes. Biochem Pharmacol 26:91–102 13. Dupuis J, Stewart DJ, Cernacek P, Gosselin G (1996) Human pulmonary circulation is an important site for both clearance and production of endothelin-1. Circulation 94:1578–1584 14. Luscher TF (1990) Endothelium-derived vasoactive factors and regulation of vascular tone in human blood vessels. Lung 168:27–34 15. Goddard J, Webb DJ (1999) Endothelin receptor antagonists. Promising new agents in the management of cardiovascular disorders. Drugs R D 2:1–12 16. Valdenaire O, Rohrbacher E, Mattei MG (1995) Organization of the gene encoding the human endothelin-converting enzyme (ECE-1). J Biol Chem 270:29794–29798 17. Sahli S, Stump B, Welti T, Schweizer WB, Diederich F, BlumKaelin D, Aebi JD, Böhm HJ (2005) A New class of inhibitors for the metalloprotease neprilysin based on a Central Imidazole Scaffold. Helv Chim Acta 88:707–730 18. Meidan R, Klipper E, Gilboa T, Muller L, Levy N (2005) Endothelin-converting enzyme-1, abundance of isoforms a-d and identification of a novel alternatively spliced variant lacking a transmembrane domain. J Biol Chem 280:40867–40874 19. Klipper E, Levy N, Gilboa T, Muller L, Meidan R (2006) Identification of a novel alternatively spliced variant endothelin converting enzyme-1 lacking a transmembrane domain. Exp Biol Med (Maywood) 231:723–728 20. Arai H, Hori S, Aramori I, Ohkubo H, Nakanishi S (1990) Cloning and expression of a cDNA encoding an endothelin receptor. Nature 348:730–732 21. Sakurai T, Yanagisawa M, Takuwa Y, Miyazaki H, Kimura S, Goto K, Masaki T (1990) Cloning of a cDNA encoding a nonisopeptide-selective subtype of the endothelin receptor. Nature 348:732–735 22. Gohla A, Offermanns S, Wilkie TM, Schultz G (1999) Differential involvement of Galpha12 and Galpha13 in receptormediated stress fiber formation. J Biol Chem 274:17901– 17907 23. Masaki T, Miwa S, Sawamura T, Ninomiya H, Okamoto Y (1999) Subcellular mechanisms of endothelin action in vascular system. Eur J Pharmacol 375:133–138 24. Kawanabe Y, Okamoto Y, Nozaki K, Hashimoto N, Miwa S, Masaki T (2002) Molecular mechanism for endothelin-1-induced stress-fiber formation: analysis of G proteins using a mutant endothelin(A) receptor. Mol Pharmacol 61:277–284

Pediatr Nephrol (2013) 28:711–720 25. Kawanabe Y, Okamoto Y, Miwa S, Hashimoto N, Masaki T (2002) Molecular mechanisms for the activation of voltage-independent Ca2+ channels by endothelin-1 in Chinese hamster ovary cells stably expressing human endothelin(A) receptors. Mol Pharmacol 62:75–80 26. Kawanabe Y, Hashimoto N, Masaki T (2002) Effects of phosphoinositide 3-kinase on the endothelin-1-induced activation of voltage-independent Ca(2+) channels and mitogenesis in Chinese hamster ovary cells stably expressing endothelin(a) receptor. Mol Pharmacol 62:756–761 27. Nootens M, Kaufmann E, Rector T, Toher C, Judd D, Francis GS, Rich S (1995) Neurohormonal activation in patients with right ventricular failure from pulmonary hypertension: relation to hemodynamic variables and endothelin levels. J Am Coll Cardiol 26:1581–1585 28. Stewart DJ, Levy RD, Cernacek P, Langleben D (1991) Increased plasma endothelin-1 in pulmonary hypertension: marker or mediator of disease? Ann Intern Med 114:464–469 29. Yokokawa K, Tahara H, Kohno M, Murakawa K, Yasunari K, Nakagawa K, Hamada T, Otani S, Yanagisawa M, Takeda T (1991) Hypertension associated with endothelin-secreting malignant hemangioendothelioma. Ann Intern Med 114:213–215 30. Amann K, Munter K, Wessels S, Wagner J, Balajew V, Hergenroder S, Mall G, Ritz E (2000) Endothelin A receptor blockade prevents capillary/myocyte mismatch in the heart of uremic animals. J Am Soc Nephrol 11:1702–1711 31. Iglarz M, Clozel M (2010) At the heart of tissue: endothelin system and end-organ damage. Clin Sci (Lond) 119:453–463 32. Ge Y, Bagnall A, Stricklett PK, Strait K, Webb DJ, Kotelevtsev Y, Kohan DE (2006) Collecting duct-specific knockout of the endothelin B receptor causes hypertension and sodium retention. Am J Physiol Renal Physiol 291:F1274–F1280 33. Fink G, Li M, Lau Y, Osborn J, Watts S (2007) Chronic activation of endothelin B receptors: new model of experimental hypertension. Hypertension 50:512–518 34. Goddard J, Johnston NR, Hand MF, Cumming AD, Rabelink TJ, Rankin AJ, Webb DJ (2004) Endothelin-A receptor antagonism reduces blood pressure and increases renal blood flow in hypertensive patients with chronic renal failure: a comparison of selective and combined endothelin receptor blockade. Circulation 109:1186–1193 35. Agapitov AV, Haynes WG (2002) Role of endothelin in cardiovascular disease. J Renin Angiotensin Aldosterone Syst 3:1–15 36. Winker R, Garland EM, Rudiger HW, Diedrich A, Biaggioni I, Ponocny I, Cascorbi I, Robertson D (2005) Influence of an insertion variant in the 5′UTR of the endothelin-1 gene on orthostatic intolerance. Am J Med Sci 330:166–171 37. Sarzynski MA, Rankinen T, Sternfeld B, Fornage M, Sidney S, Bouchard C (2011) SNP-by-fitness and SNP-by-BMI interactions from seven candidate genes and incident hypertension after 20 years of follow-up: the CARDIA Fitness Study. J Hum Hypertens 25:509–518 38. Boesen EI, Pollock JS, Pollock DM (2010) Contrasting effects of intervention with ETA and ETB receptor antagonists in hypertension induced by angiotensin II and high-salt diet. Can J Physiol Pharmacol 88:802–807 39. Zhang Y, Fu C, Ren H, He D, Wang X, Asico LD, Jose PA, Zeng C (2011) Impaired stimulatory effect of ETB receptor on D receptor in immortalized renal proximal tubule cells of spontaneously hypertensive rats. Kidney Blood Press Res 34:75–82 40. Schildroth J, Rettig-Zimmermann J, Kalk P, Steege A, Fähling M, Sendeski M, Paliege A, Lai EY, Bachmann S, Persson PB, Hocher B, Patzak A (2011) Endothelin type A and B receptors in the control of afferent and efferent arterioles in mice. Nephrol Dial Transplant 26:779–789

719 41. Honing ML, Hijmering ML, Ballard DE, Yang YP, Padley RJ, Morrison PJ, Rabelink TJ (2000) Selective ET(A) receptor antagonism with ABT-627 attenuates all renal effects of endothelin in humans. J Am Soc Nephrol 11:1498–1504 42. Simonson MS, Ismail-Beigi F (2011) Endothelin-1 increases collagen accumulation in renal mesangial cells by stimulating a chemokine and cytokine autocrine signaling loop. J Biol Chem 286:11003–11008 43. Hoffman A, Abassi ZA, Brodsky S, Ramadan R, Winaver J (2000) Mechanisms of big endothelin-1-induced diuresis and natriuresis: role of ET(B) receptors. Hypertension 35:732–739 44. Boesen EI, Pollock DM (2010) Cooperative role of ETA and ETB receptors in mediating the diuretic response to intramedullary hyperosmotic NaCl infusion. Am J Physiol Renal Physiol 299: F1424–F1432 45. Brochu E, Lacasse S, Lariviere R, Kingma I, Grose JH, Lebel M (1999) Differential effects of endothelin-1 antagonists on erythropoietin-induced hypertension in renal failure. J Am Soc Nephrol 10:1440–1446 46. Davis LS, Haleen SJ, Doherty AM, Cody WL, Keiser JA (1994) Effects of selective endothelin antagonists on the hemodynamic response to cyclosporin A. J Am Soc Nephrol 4:1448–1454 47. Maguire JJ, Kuc RE, Davenport AP (2012) Defining the affinity and receptor sub-type selectivity of four classes of endothelin antagonists in clinically relevant human cardiovascular tissues. Life Sci. doi:10.1016/j.lfs.2012.05.008 48. Channick RN, Simonneau G, Sitbon O, Robbins IM, Frost A, Tapson VF, Badesch DB, Roux S, Rainisio M, Bodin F, Rubin LJ (2001) Effects of the dual endothelin-receptor antagonist bosentan in patients with pulmonary hypertension: a randomised placebo-controlled study. Lancet 358:1119–1123 49. Rubin LJ, Badesch DB, Barst RJ, Galie N, Black CM, Keogh A, Pulido T, Frost A, Roux S, Leconte I, Landzberg M, Simonneau G (2002) Bosentan therapy for pulmonary arterial hypertension. N Engl J Med 346:896–903 50. Barst RJ, Langleben D, Frost A, Horn EM, Oudiz R, Shapiro S, McLaughlin V, Hill N, Tapson VF, Robbins IM, Zwicke D, Duncan B, Dixon RA, Frumkin LR, STRIDE-1 Study Group (2004) Sitaxsentan therapy for pulmonary arterial hypertension. Am J Respir Crit Care Med 169:441–447 51. Trow TK, Taichman DB (2009) Endothelin receptor blockade in the management of pulmonary arterial hypertension: selective and dual antagonism. Respir Med 103:951–962 52. Ergul A (2000) Hypertension in black patients: an emerging role of the endothelin system in salt-sensitive hypertension. Hypertension 36:62–67 53. Krum H, Viskoper RJ, Lacourciere Y, Budde M, Charlon V (1998) The effect of an endothelin-receptor antagonist, bosentan, on blood pressure in patients with essential hypertension. Bosentan Hypertension Investigators. N Engl J Med 338:784–790 54. Weber MA, Black H, Bakris G, Krum H, Linas S, Weiss R, Linseman JV, Wiens BL, Warren MS, Lindholm LH (2009) A selective endothelin-receptor antagonist to reduce blood pressure in patients with treatment-resistant hypertension: a randomised, double-blind, placebo-controlled trial. Lancet 374: 1423–1431 55. Prasad VS, Palaniswamy C, Frishman WH (2009) Endothelin as a clinical target in the treatment of systemic hypertension. Cardiol Rev 17:181–191 56. Raichlin E, Prasad A, Mathew V, Kent B, Holmes DR Jr, Pumper GM, Nelson RE, Lerman LO, Lerman A (2008) Efficacy and safety of atrasentan in patients with cardiovascular risk and early atherosclerosis. Hypertension 52:522–528 57. Saleh MA, Pollock JS, Pollock DM (2011) Distinct actions of endothelin A-selective versus combined endothelin A/B receptor

720

58.

59.

60.

61.

62.

63.

64.

65.

66.

67.

68.

Pediatr Nephrol (2013) 28:711–720 antagonists in early diabetic kidney disease. J Pharmacol Exp Ther 338:263–270 Wenzel RR, Littke T, Kuranoff S, Jurgens C, Bruck H, Ritz E, Philipp T, Mitchell A, SPP301 (Avosentan) Endothelin Antagonist Evaluation in Diabetic Nephropathy Study Investigators (2009) Avosentan reduces albumin excretion in diabetics with macroalbuminuria. J Am Soc Nephrol 20:655–664 Rabkin SW (2011) Endothelin but not angiotensin II may mediate hypertension-induced coronary vascular calcification in chronic kidney disease. Int J Nephrol doi: 10.4061/2011/516237 Fenhammar J, Andersson A, Forestier J, Weitzberg E, Sollevi A, Hjelmqvist H, Frithiof R (2011) Endothelin receptor A antagonism attenuates renal medullary blood flow impairment in endotoxemic pigs. PLoS One 6:e21534 Patel NN, Toth T, Jones C, Lin H, Ray P, George SJ, Welsh G, Satchell SC, Sleeman P, Angelini GD, Murphy GJ (2011) Prevention of post-cardiopulmonary bypass acute kidney injury by endothelin A receptor blockade. Crit Care Med 39:793–802 Gulmen S, Kiris I, Narin C, Ceylan BG, Mermi B, Sutcu R, Meteoglu I (2009) Tezosentan reduces the renal injury induced by abdominal aortic ischemia-reperfusion in rats. J Surg Res 157: e7–e13 Abu-Saleh N, Ovcharenko E, Awad H, Goltsman I, Khamaisi M, Hoffman A, Heyman SN, Winaver J, Abassi Z (2012) Involvement of the endothelin and nitric oxide systems in the pathogenesis of renal ischemic damage in an experimental diabetic model. Life Sci. doi:10.1016/j.lfs.2012.02.002 Andress D, Coll B, Pritchett Y, Brennan J, Molitch M, Kohan D (2012) Clinical efficacy of the selective endothelin A receptor antagonist, atrasentan, in patients with diabetes and chronic kidney disease (CKD). Life Sci. doi:10.1016/j.lfs.2012.01.011 Weil BR, Westby CM, Van Guilder GP, Greiner JJ, Stauffer BL, DeSouza CA (2011) Enhanced endothelin-1 system activity with overweight and obesity. Am J Physiol Heart Circ Physiol 301:689– 695 Yang Z, Li JC (2008) Stimulation of endothelin-1 gene expression by insulin via phosphoinositide-3 kinase-glycogen synthase kinase-3beta signaling in endothelial cells. Life Sci 82:512–518 Kobayashi T, Nogami T, Taguchi K, Matsumoto T, Kamata K (2008) Diabetic state, high plasma insulin and angiotensin II combine to augment endothelin-1-induced vasoconstriction via ETA receptors and ERK. Br J Pharmacol 155:974–983 Matsumoto T, Ishida K, Taguchi K, Kobayashi T, Kamata K (2010) Short-term angiotensin-1 receptor antagonism in type 2

69.

70.

71.

72.

73.

74.

75.

diabetic Goto-Kakizaki rats normalizes endothelin-1-induced mesenteric artery contraction. Peptides 4:609–617 Seccia TM, Maniero C, Belloni AS, Guidolin D, Pothen P, Pessina AC, Rossi GP (2008) Role of angiotensin II, endothelin-1 and L-type calcium channel in the development of glomerular, tubulointerstitial and perivascular fibrosis. J Hypertens 26:2022–2029 Mann JF, Green D, Jamerson K, Ruilope LM, Kuranoff SJ, Littke T, Viberti G, ASCEND Study Group (2010) Avosentan for overt diabetic nephropathy. J Am Soc Nephrol 21:527–535 Kohan DE, Pritchett Y, Molitch M, Wen S, Garimella T, Audhya P, Andress DL (2011) Addition of atrasentan to renin-angiotensin system blockade reduces albuminuria in diabetic nephropathy. J Am Soc Nephrol 22:763–772 Wesson DE, Simoni J, Broglio K, Sheather S (2011) Acid retention accompanies reduced GFR in humans and increases plasma levels of endothelin and aldosterone. Am J Physiol Renal Physiol 300: F830–F837 Spieker LE, Noll G, Luscher TF (2001) Therapeutic potential for endothelin receptor antagonists in cardiovascular disorders. Am J Cardiovasc Drugs 1:293–303 Dhaun N, Macintyre IM, Melville V, Lilitkarntakul P, Johnston NR, Goddard J, Webb DJ (2009) Blood pressure-independent reduction in proteinuria and arterial stiffness after acute endothelin-a receptor antagonism in chronic kidney disease. Hypertension 54:113–119 Dhaun N, Macintyre IM, Melville V, Lilitkarntakul P, Johnston NR, Goddard J, Webb DJ (2009) Effects of endothelin receptor antagonism relate to the degree of renin-angiotensin system blockade in chronic proteinuric kidney disease. Hypertension 54:e19–e20

Answers 1. b. The natriuretic peptides decrease ET-1 production. 2. a. ETA receptors are located only on vascular smooth muscle cells (VSMC) where they mediate vasoconstriction. 3. d. Approximately 80 % of circulating ET-1 is cleared in the lungs 4. e. ETA receptor blockade decreases the following: blood pressure, salt and water retention, proteinuria, reactive oxygen species (ROS) production, inflammation, and fibrosis. Heart rate is not decreased by ETA receptor blockade. 5. b. Side-effects of ETA receptor blockade include hepatotoxicity, edema, teratogenicity, and reduced sperm count.