Current and future strategies for targeting the endothelin pathway in cardiovascular disease

The first endothelin (ET)-1 receptor antagonist was approved for clinical use over 20 years ago, but to date this class of compounds has been limited to treating pulmonary arterial hypertension, a rare disease. Translational research over the last 5 years has reignited interest in the ET system as a therapeutic target across the spectrum of cardiovascular diseases including resistant hypertension, microvascular angina and post-coronavirus disease 2019 conditions. Notable developments include approval of a new ETA receptor antagonist and, intriguingly, combining the actions of ETA and an angiotensin II type 1 receptor antagonist within the same novel small molecule. Combinations of ET receptor blockers with other drugs, including phosphodiesterase-5 inhibitors and sodium–glucose co-transporter-2 antagonists, may drive synergistic benefits with the prospect of alleviating side effects. These new therapeutic strategies have the potential to dramatically widen the scope of indications targeting the ET-1 pathway. Abraham and colleagues review the recent developments and future strategies to therapeutically target the endothelin pathway for a broad spectrum of cardiovascular diseases.

The first endothelin (ET)-1 receptor antagonist was approved for clinical use over 20 years ago, but to date this class of compounds has been limited to treating pulmonary arterial hypertension, a rare disease.Translational research over the last 5 years has reignited interest in the ET system as a therapeutic target across the spectrum of cardiovascular diseases including resistant hypertension, microvascular angina and post-coronavirus disease 2019 conditions.Notable developments include approval of a new ET A receptor antagonist and, intriguingly, combining the actions of ET A and an angiotensin II type 1 receptor antagonist within the same novel small molecule.Combinations of ET receptor blockers with other drugs, including phosphodiesterase-5 inhibitors and sodium-glucose co-transporter-2 antagonists, may drive synergistic benefits with the prospect of alleviating side effects.These new therapeutic strategies have the potential to dramatically widen the scope of indications targeting the ET-1 pathway.
The first-in-class ET-1 receptor antagonist bosentan was approved for clinical use in 2002, 14 years after the endothelium-derived peptide was first identified and shown to be one of the most potent vasoconstrictors ever described 1 (Figs.1-3).Multiple other biological effects of ET-1 are recognized including immune regulation, vascular remodeling and fibrosis, while dysregulation of ET-1 and its related pathways have been causally identified as an important mediator in many diseases 2 .Three further small-molecule antagonists have been approved: ambrisentan, macitentan and sitaxentan, which was subsequently withdrawn in 2010.In the ensuing years, use of ET receptor antagonists in the clinic has been limited to treating pulmonary arterial hypertension (PAH), a rare but devastating condition.
This Review focuses on the therapeutic targeting of the ET receptors over the past 5 years, which has undergone a remarkable renaissance, with a series of discoveries, clinical trials and key papers reigniting the field (Tables 1 and 2 and Fig. 4).Developments include the 2022 approval of the first new ET A antagonist in over 10 years 3 and a phase 3 clinical trial demonstrating efficacy in treating resistant hypertension 4 , heralding a new era that may see ET receptor antagonists enter the clinical mainstream.New chemical entities, such as the combination of an ET and angiotensin receptor antagonist within the same small molecule 5 , are emerging with the potential to cause a paradigm shift for a broad range of cardiovascular indications.More conventionally, the combination of ET antagonists with other drugs may help to drive synergistic benefits such as new treatments for portal hypertension (NCT05516498) and severe COVID-19 infection (NCT04393246; Box 1).In parallel, studies in human genetics are linking ET signaling with disease susceptibility variant genes, highlighting new mechanistic insights.Genome-wide association studies (GWAS) have linked angina and other cardiovascular diseases to a common functional single nucleotide polymorphism (SNP) in an intron of the PHACTR1 gene that regulates expression of the EDN1 gene 6,7 .This SNP

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https://doi.org/10.1038/s44161-023-00347-2with new mechanisms of action, combinations with other agents and the potential for precision medicine with emerging genetic variants in the ET pathway.Recent reviews cover the wider biological actions of ET-1 (ref.2), ET antagonist clinical trials 8 and pharmacology of ET peptides and receptors 9 .is being exploited in a precision medicine trial to identify patients with the variant and enrich the study population with participants most likely to benefit from ET receptor antagonism.
The aim of this Review is to focus on recent clinical trials of ET antagonists in cardiovascular disease to highlight therapeutic agents Big ET-1  ET-converting enzymes (yellow arrow; mainly ECE-1 at neutral pH but also by the isoform ECE-2 at acidic pH) to yield the biologically active peptide and an inactive C-terminal fragment 9 .Three further peptides have been identified as products of processing of ppET-1 within endothelial cells that correlate with ET-1 synthesis: N-terminal-proET-1 (ppET-11 8-50 ), ELDP (ppET-1 93-166 ) and C-terminal-proET-1 (ppET-1 169-212 ).The other two structurally related 21-amino-acid isoforms, ET-2 and ET-3, are also present in humans but with a more limited distribution and are less extensively studied.ET-2 has a similar affinity for the ET A receptor, whereas ET-3 is lower.All three isoforms have a similar affinity at the ET B receptor 9 .ECE-1, endothelin-converting enzyme-1; ECE-2, endothelin-converting enzyme-2; ELDP, endothelin-like domain peptide; NT-proET-1, N-terminal proendothelin-1; ppET-1, preproendothelin-1.ET-1 binding to ET A receptors, the principal subtype on smooth muscle, causes long-lasting vasoconstriction that can be fully reversed by ET A antagonists, suggesting little contribution to vasoconstriction by the low density of smooth muscle ET B receptors.ET-1-ET A complex is internalized to the endosome and the subtype may be recycled to the cell surface.Vasoconstriction is beneficially regulated by ET-1 acting in an autocrine/paracrine manner to bind endothelial cell ET B receptors to release vasodilators.Endothelial ET B receptors abundant in the lungs, kidney and liver remove the peptide as an ET-1-ET B complex that is internalized and degraded to the lysosome.In the kidney, collecting duct ET B receptors also beneficially promote Na + excretion via the epithelial sodium channel 9 .ET antagonists are used in combination with PDE5 inhibitors (to increase levels of cGMP to promote vasodilatation) or SGLT2 inhibitors (to lower plasma glucose levels by blocking reabsorption of filtered glucose).Green and red indicate approved and investigational ET antagonists, respectively.cGMP, cyclic guanosine monophosphate; ENaC, epithelial sodium channel.

Therapeutic targeting of the ET pathway
Compared with all other mRNA-encoding vasoactive peptides present in human endothelial cells, EDN1 mRNA is the most abundant by an order of magnitude (Fig. 3a).The peptide is expressed by endothelial cells of all human vascular beds studied, including small resistance vessels, from where it is locally released and contributes, unlike angiotensin II, to maintenance of normal vascular tone.In the human vasculature, vasoconstriction is mediated principally by ET A receptors (Fig. 3b,c) on the underlying smooth muscle where a small proportion of ET B receptors may also be present.These responses are typically long lasting (Fig. 3b) and the magnitude of response (for example, in human coronary arteries) is greater than that with angiotensin II or other vasoactive agents (Fig. 3c) 1 .Crucially, ET A receptor antagonists can fully reverse ET-1-mediated vasoconstriction in isolated human vessels 9 (Fig. 3d).In normotensive volunteers, ET A receptor-selective antagonists, such as BQ-123, cause vasodilatation consistent with ET-1, contributing to blood pressure maintenance 10 .Thus far, clinical benefit of ET receptor antagonists has been best established in idiopathic      11 .
Locally released ET-1 also acts in an autocrine/paracrine manner to activate endothelial ET B receptor cells to release vasodilators 12 and limit vasoconstriction.Endothelial ET B receptors, particularly in the lungs but also in the kidney and liver, undergo endocytosis to beneficially remove the peptide from circulation 9 .This is an important mechanism to inactivate ET-1 in addition to metabolism by neutral endopeptidases.
ET-1 produced in nephrons of the kidney under physiological conditions regulates sodium and water excretion, likely contributing to the principal limiting adverse effect of ET receptor antagonists: fluid retention.Peripheral edema has been reported in 17% of patients with PAH treated with bosentan 13 .Earlier attempts to use ET receptor antagonists for heart failure were associated with increased risk of worsening heart failure 14 ; ET receptor antagonists are not indicated for the treatment of heart failure 8 .Defining a dominant mechanism for ET receptor antagonist-induced fluid retention is complex, owing to the numerous biological actions of ET-1 in the kidney.For example, in the glomerulus, ET-1 may induce afferent arteriolar vasoconstriction and efferent arteriolar vasodilatation, cellular injury to podocytes and mesangial cell contraction; these effects provide the rationale for testing ET receptor antagonists in chronic kidney disease (CKD).Much preclinical evidence exists for ET B receptor activation inhibiting sodium and water retention, thus beneficially promoting natriuresis, especially in the collecting duct 15 .A key study showed collecting-ductspecific ET B receptor knockout mice had elevated blood pressure and diminished sodium excretion in response to salt loading compared with controls 16 .However, this effect translates only partially to clinical relevance, not least given the observation that highly selective ET A receptor antagonists, such as zibotentan, with no detectable binding to ET B receptors also give rise to symptomatic fluid overload 17 , perhaps even with higher frequency than nonselective ET receptor antagonists for similar indications 18 .Nephron and collecting-ductspecific deletion of ET A receptors abolished fluid retention observed with ambrisentan and atrasentan, supporting an additional natriuretic effect of ET A receptors in the collecting duct 19 .It has been suggested that in the presence of selective ET A receptor blockade, edema results from overstimulation of ET B receptors, causing systemic vasodilation, subsequent activation of norepinephrine, arginine vasopressin and aldosterone, and increased vascular permeability in a rat model 18 .A reason for caution is that fluid overload symptoms remained high for a relevant number of patients treated with the selective ET A antagonist darusentan, despite co-therapy with other interacting heart failure medications 20,21 .For example, angiotensin-converting enzyme (ACE) inhibitor therapy reduces peripheral ET-1 levels 22 , whereas angiotensin II promotes EDN1 gene transcription 23 .A critical interplay also exists between the sympathetic nervous system and ET-1.Central infusion of ET-1 in the cerebrospinal fluid of rats resulted in a significant vasopressor response; however, this was abolished by peripheral treatment with the alpha-adrenergic blocker prazosin 24 .Treatment with beta blockers in heart failure patients resulted in reductions in the plasma ET-1 concentration 25 .As such, additional clinical benefits of ET receptor blockade in heart failure may have been diminished in patients already on maximal medical therapy.The benefits of selectively blocking ET A receptors versus blocking both subtypes continue to be debated and are likely to vary based on the indication (Box 2).For example, a beneficial inotropic effect of ET-1 agonism for ET A (hence a deleterious effect of ET A antagonists) has been reported in patients with preserved left ventricular function 26 .Emerging strategies combining ET A selective antagonists with other classes of drugs to mitigate unwanted side effects are discussed below.
Despite the early identification of endothelin-converting enzymes (ECEs) specifically catalyzing the synthesis of the active peptide from the inactive precursor, big ET-1, no inhibitors of ECEs have been developed for clinical use.A combined ECE/neutral endopeptidase inhibitor (daglutril, SLV-306) has been tested in clinical studies but has not progressed 27,28 .Apart from ET-1 and big ET-1, three further peptides derived from their common precursor, pre-proET-1, have been identified unequivocally by specific immunoassays and mass spectrometry in conditioned media from endothelial cells 29 .Although N-terminal proendothelin-1 was not subsequently detected in human plasma, levels of endothelin-like domain peptide (ELDP) and C-terminal proendothelin-1 (CT-proET-1) were significantly higher in patients with

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https://doi.org/10.1038/s44161-023-00347-2chronic heart failure compared with pre-hypertension or mild hypertension.Interestingly, in rat isolated mesenteric resistance arteries, ELDP enhanced ET-1 vasoconstriction and prolonged the increase in blood pressure in anesthetized rats, although it was not active on its own in these preparations.Whether ELDP has similar actions in human isolated vessels and could therefore potentiate ET-1 responses in vivo remains to be studied.Of the three peptides, CT-proET-1 had the slowest systemic clearance in rats 29 .A specific assay for CT-proET-1 (ref.30) has been developed.Plasma levels of CT-proET-1 are an order of magnitude higher than ET-1 and have been used as a surrogate biomarker for ET-1 release in disease states such as PAH 31 .To date, whether CT-proET-1 has affinity for ET receptors and has any pharmacological activity has not been explored.However, sitaxentan, an ET A selective receptor antagonist, modestly increased plasma ELDP and CT-proET-1 in patients with CKD, although the biological effect of these circulating peptides is unknown 32 .

Precision medicine in cardiovascular disease
Associations of known interindividual genetic variants with deep phenotypic characteristics have now been documented comprehensively in large open resource datasets such as the UK Biobank project 33 .These data allow validation of new variants, most commonly SNPs, identified Box 1

ET-1, COVID-19 and post-COVID-19 conditions ('long COVID')
The COVID-19 pandemic continues to place strain on healthcare services worldwide, and there remains an ongoing requirement for new therapies.Where statistics are available in the United Kingdom from the period of widespread population testing from April 2020 to July 2022, hospitalization rates for all COVID-19 infections ranged from 0.5% to 5%; among those hospitalized during this period, rates of mechanical ventilation bed occupancy have ranged from 1% to 10% (https://coronavirus.data.gov.uk/details/healthcare?areaTyp e=nation&areaName=England), reflecting the significant burden of progressive severe COVID-19 disease.In this subset of patients, virus-induced endothelial damage resulting in a systemic syndrome of excessive vasoconstriction, inflammation and thrombosis has been suggested as an important pathological mechanism 126,127 .Severe acute respiratory syndrome coronavirus 2, the causative agent of COVID-19, infects host cells by exploiting ACE2, a membrane receptor expressed by epithelial cells and the vascular endothelium 128 .Viral inclusions and lymphocytic infiltration of apoptotic endothelial cells have been identified from histological examination of tissues obtained from individuals with COVID-19 with acute lung 129 , kidney 130 and myocardial injury 131 .In plasma, multiple biomarkers for endothelial injury are elevated in individuals hospitalized with COVID- 19 (refs.132,133).Consistent with these observations, elevated ET-1 plasma levels during the acute phase of illness have also been reported 134 and shown to be associated with multiorgan complications 135 .Higher levels of CT-proET-1, frequently measured as a surrogate marker of ET-1 pathway activation in observational studies (owing to its higher plasma concentration), have also shown association with COVID-19 disease severity 136 , although its use as a prognostic biomarker independent of confounding variables has shown conflicting results 137,138 .
Increased circulating ET-1 in severe COVID-19 infection (Fig. 5) is likely a consequence of multiple factors causing excess ET-1 to spill over into plasma, despite the predominant abluminal release of ET-1 (ref.139).ET-1 bioavailability is principally governed at the transcriptional level of the EDN1 gene to pre-proET-1 mRNA.Acutephase reactant regulatory elements responsive to inflammatory cytokines such as TNF, IFNγ and IL-1β, as well as other extracellular signals including hypoxia and thrombin, have been characterized 140 and demonstrated to mediate upregulated transcription within minutes 141 .Alongside release of stored ET-1 peptides from damaged endothelial cells, the systemic inflammatory response to infection additionally provides multiple stimuli for the regulated pathway of ET-1 release via promotion of Weibel-Palade body exocytosis (Fig. 5).A randomized, double-blind placebo-controlled trial with bosentan was effective in patients with acute COVID-19, supporting a pathological role of ET-1 (ref.142).Concurrently, the TACTIC-E (Therapeutic study in pre-ICU patients admitted with COVID-19: experimental drugs and mechanisms) trial 143 will investigate ET receptor blockade as a therapeutic strategy for hospitalized patients with COVID-19 infection, with one treatment arm randomized to receive a combination of 5 mg ambrisentan and 10 mg dapagliflozin.Detailed analysis of serial plasma biomarkers and clinical features in this group is anticipated to test the hypothesized mechanisms for ET-1 dysregulation discussed here, as well as the functional effects of excess ET-1.
Persisting impairments after COVID-19 infection are common, and there is increasing recognition of how this may result in a longterm, population-wide legacy of multisystem illness.For example, a study has reported approximately one in eight patients hospitalized with COVID-19 have evidence of myocarditis 28-60 days after discharge on cardiac magnetic resonance imaging; this finding was associated with subsequent pulmonary fibrosis during longer-term follow-up 144 .Consensus diagnostic criteria for the post-COVID-19 condition, commonly known as 'long COVID' are now available and can be summarized as symptoms persisting 3 months from the onset of infection, most commonly exertional fatigue and malaise (World Health Organization (WHO) clinical case definition of post-COVID-19 condition: https://www.who.int/europe/news-room/fact-sheets/item/post-covid-19-condition/).The pathophysiology of post-COVID-19 condition remains to be fully characterized; however, increased ET-1 levels and peripheral endothelial dysfunction have been suggested as important features 134,145 .The causal link between viral infection and persistently raised ET-1 levels is not well defined.One theory relates to vascular damage and ongoing activation of the intrinsic coagulation pathway generating thrombin, which is permissive for ET-1 production and release 134 .Autoantibodies directed at the ET A receptor and angiotensin II type I (AT 1 ) receptor have been well characterized in kidney and heart transplant recipients as a risk factor for rejection and vasculopathy.These autoantibodies have also been associated with poor outcomes among patients hospitalized with COVID-19 (ref.146) and identified in a minority of patients within a small case series of post-COVID-19 condition 147 .ET A receptor and AT 1 receptor autoantibodies are known to act as functional agonists with resultant deleterious effects including excessive vasoconstriction and proinflammatory endothelial signaling 148 .Mechanisms for the generation and persistence of these autoantibodies remain to be defined; however, their association with severe COVID-19 and post-COVID-19 condition add further rationale for ET receptor antagonism as a future therapeutic option.

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https://doi.org/10.1038/s44161-023-00347-2by GWAS of specific diseases.Coronary artery disease (CAD) is the leading cause of mortality worldwide and cardiovascular risk modification represents a generational global health priority.There is an emerging focus on identifying heritable risk to further optimize disease prevention and treatment strategies in the era of precision medicine.Thus far, >250 genetic loci have been associated with CAD, with the vast majority residing in noncoding sequences of the genome 34 .Of all CAD-associated loci, a number have demonstrated functional roles in gene expression, being designated as cis-acting (with an effect on a closely residing gene) or trans-acting (where the effect is on a distant gene, including on another chromosome) regulatory regions through expression quantitative trait loci mapping 35 .Numerous expression quantitative trait loci relationships may exist for each variant; thus, identification of a true causal mechanism is elusive.Common variants in vasomotor pathway signaling such as nitric oxide (NO) have been associated previously with the genetic risk of CAD, highlighting the importance of genetic regulation of vascular tone in disease pathogenesis 36 .
In the ET-1 pathway, multiple SNPs associated by proximity to the EDNRA gene locus in linkage disequilibrium have reported associations with CAD in GWAS and the UK Biobank (Table 3).Of note, while CAD risk is generally associated with the minor alleles of the EDNRA SNPs, the major alleles of the upstream gene variants-rs1878406, rs6842241 and the intronic variant rs6841473-are associated with increased risk of intracranial aneurysm 37 .The effect allele G at rs6841581 demonstrated increased binding of nuclear proteins and reduced transcriptional activity in reporter assays, suggesting this variant may functionally reduce EDNRA transcription 37 .The pathological mechanism of intracranial aneurysm formation related to genetic dysregulation of EDNRA and the implied converse relationship with atherosclerotic CAD remains incompletely defined.Further work is similarly required to establish the pathological mechanism of a missense variant at rs5370 located in the CT-proET region in the EDN1 gene associated with elevated circulating levels of CT-proET 38 .In smallscale studies, this variant has been suggested as a potential risk factor for premature CAD 39 , hypertension complicating pregnancy 40 and obesity 41 .Increased circulating ET-1 peptides have also been reported with other SNPs including minor variants involving KLKB1 (rs4253238), which encodes the proteolytic enzyme kallikrein; GRK6 (rs2731672), which encodes a G-protein-coupled receptor kinase 38 ; and ADIPOQ (rs17366743), which encodes the adipokine hormone adiponectin 42 .A rare missense variant in ABCC9 (rs61688134) mapping to a region encoding a nucleotide binding domain of SUR2, a subunit of the endothelial K ATP channel, has also been associated with increased circulating ET-1 as well as susceptibility to myocardial infarction 43 .A synonymous variant in GNB3 (rs5443), which encodes the G-protein β3 subunit associated with hypertension, has been suggested to increase ET-1-induced venoconstriction, unrelated to circulating ET-1 levels, tantalizing the possibility of a functional variant 44 .The associations of these ET-1 pathway genetic variants with disease remain coincidental.Further elaboration at the molecular level may provide mechanisms of ET-1 dysregulation, which could then be validated for a causal effect by Mendelian randomization studies.Thus far, the most securely established mechanism for ET-1 genetic dysregulation in CAD comes from a series of key investigations by Gupta et al. focused on the lead variant rs9349379 at locus 6p24 in an intron of the PHACTR1 gene 6 .PHACTR1 itself has no effect on vascular physiology.Hence, the effect of this variant is most likely related to its influence on EDN1 (refs.45,46).The minor G allele (population frequency, 40%) of this SNP is associated with increased risk of atherosclerotic CAD and reduced risk of coronary and cervical artery dissection, hypertension, migraine and fibromuscular dysplasia.Conversely, the major A allele is associated with a higher incidence of fibromuscular dysplasia and spontaneous coronary artery dissection 47 .
Further mining of detailed phenotypic data revealed association of the G allele with endothelial dysfunction and discordantly reduced arterial stiffness.The latter finding has been recently validated using an artificial intelligence technique to analyze the arterial waveform obtained using finger plethysmography 48 .In aortic tissues, the locus at rs9349379 demonstrated local chromatin signatures (acetylated histone H3 Lys27) highly associated with enhancer activity.Targeted deletion of this region and the GG homozygous minor allelic series in stem-cell-derived endothelial cells demonstrated significantly increased EDN1 expression and big ET-1 peptide production.Structural analysis of chromatin structure localized the enhancer effect to a long-range chromatin loop in the intergenic region between EDN1 and PHACTR1.Finally, increased peripheral venous big ET-1 was measured in healthy individuals possessing the minor G allele compared with the AA homozygous genotype 6 .Extending these findings in a cohort of patients with nonobstructive coronary disease enrolled in the CorMicA (coronary microvascular angina) study, the G allele of rs9349379 was also associated with higher circulating ET-1.Additionally, the overall frequency of the G allele was higher in this patient group (46%) and associated with coronary microvascular dysfunction on invasive testing, perfusion defects on stress perfusion cardiac magnetic resonance imaging and reduced exercise tolerance due to angina, with the highest risk observed for patients with the homozygous GG genotype 7 .These results inform the ongoing PRIZE (Precision medicine with zibotentan in microvascular angina) trial, a randomized, double-blind crossover study of the highly ET A selective antagonist zibotentan, in a population of patients with microvascular angina selectively enriched for 'responders' possessing the rs9349379 G allele 49 .Subgroup analysis of the PRIZE trial may extend the known GWAS associations of this SNP for atherosclerosis to an important subgroup of patients without significant coronary atherosclerosis, whose angina is attributable to coronary microvascular dysfunction.

Defining endothelin receptor selectivity
ET receptor antagonists are described as either selective for one receptor subtype or alternatively as nonselective antagonists that block both ET A and ET B receptors (Fig. 6 and Table 1).The classification is made by the pharmaceutical company or academic group discovering the compound and is initially based on the binding affinity of the compound for each of the receptor subtypes.There is no agreed degree of selectivity that determines whether a compound is described as nonselective or receptor subtype selective 149 .Selectivity data may be obtained from radioligand binding and functional experiments generating a range of values that are dependent on experimental design.At least 100-fold selectivity is required for a subtype selective compound to theoretically have little receptor occupancy of the lower-affinity receptor at concentrations occupying a significant proportion of the higher-affinity receptor.We propose that ET A selective compounds should display more than 100-fold, and preferably 1,000-fold, selectivity for the ET A subtype and those with less than 100-fold ET A selectivity should be classified as nonselective antagonists.Within this range, marginally selective ET A antagonists displaying 200-fold selectivity, such as ambrisentan, have been reported to increase plasma ET-1 levels at clinically used doses (Table 1), suggesting ET B engagement.Similarly, ET B agonists and antagonists should display at least 100-fold selectivity to be useful clinical agents or research compounds.

Monoclonal antibody ET A antagonists
Monoclonal antibodies are a major class of therapeutic biopharmaceutical agents with over 120 approved for clinical use and over 200 in development, mainly for the treatment of cancer and rheumatoid arthritis.Therapeutic monoclonal antibodies have potential advantages over small molecules, principally higher specificity with fewer off-target effects and increased patient compliance with longer plasma half-lives.Only one monoclonal antibody against a class A G-protein-coupled receptor (GPCR), mogamulizumab, that inhibits the CCR4 receptor used in cancer, and one in class B, erenumab, used for the treatment of migraine, have clinical approval (https://www.guidetopharmacology. org/).This reflects the difficulty of generating monoclonal antibodies given the limited extracellular epitopes of GPCRs and low density of receptor expression compared with other successful protein targets.
Getagozumab (GMA301) 50,51 is a humanized monoclonal antibody raised against the ET A receptor, with a high affinity (equilibrium dissociation constant ~8.7 nM) and potency (half-maximal inhibitory concentration 38 nM) measured in a functional assay (calcium influx)

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https://doi.org/10.1038/s44161-023-00347-2 in human ET A -expressing CHO cells.The monoclonal antibody has no effect on ET B receptors in these cells.In cynomolgus monkeys, the half-life of getagozumab was 7-10 days based on a single intravenous dose up to 750 mg per kilogram of body weight.This compares with a half-life of 15 h for the small-molecule ET A antagonist ambrisentan.
In the repeated dose toxicity study of up to a maximum of 250 mg per kilogram of body weight, no obvious toxicities were observed (on heart rate, electrocardiogram, body temperature and body weight).Interestingly, there was no reduction in blood pressure.Overall, getagozumab was well tolerated with no observed adverse effects such as fluid retention or liver toxicity 50 .Further assessment of a 6-week biweekly course of injections of 5 mg per kilogram of body weight getagozumab in cynomolgus monkey models of PAH (using the plant alkaloid monocrotaline) demonstrated significant reductions in left ventricular hypertrophy and right ventricular systolic pressure.Efficacy was similar or better than that with a daily dose of ambrisentan (1 mg per kg body weight, intragastric).First-in-human studies for safety and tolerability with getagozumab (NCT04505137) were favorable and a long plasma half-life of ~21 days was reported 52 .Getagozumab (75-1,000 mg) was well tolerated; minor side effects included headache, nausea and pharyngitis.Ongoing trials will clarify whether the monoclonal antibody induces fluid retention including a randomized, placebo-controlled, doubleblind, dose-escalation multicenter study to assess patients with PAH in WHO functional class II and III taking standard-of-care medications, except for ET receptor antagonists (NCT04503733).Demonstration of efficacy in subsequent clinical trials would provide proof of principle in using monoclonal antibodies to target GPCRs, not only for cardiovascular disease, but also for other clinical conditions where ET-1 has been shown to contribute to pathophysiology.A long-acting antagonist blocking ET A receptors over weeks or months may have a therapeutic advantage.

Recent key clinical trials of ET receptor agonists and antagonists
Hypertension represents a major global public health priority with an estimated population prevalence among adults of 46% 53 .The condition is highly pathologically important-even though atherosclerosis has an indolent course, it is often left undiagnosed until late in its natural history.Consequently, hypertension confers an elevated risk of lifethreatening cardiovascular events such as myocardial infarction and stroke.Additionally, hypertensive injurious forces transmitted to the brain, heart and kidney accumulate over time, resulting in chronic organ dysfunction.High blood pressure can be mitigated by current standard-of-care medications; however, in around 15% of patients, blood pressure remains above recommended targets for cardiovascular risk modification even after treatment with a combination of drugs targeting several pathways, referred to as 'resistant hypertension'.Individuals with this diagnosis, as a subgroup of hypertensive patients, have significantly higher rates of adverse cardiovascular events and progression to severe CKD 54 .Resistant hypertension is a heterogeneous phenomenon frequently associated with increased blood pressure, sensitivity to exogenous salt (salt-sensitive hypertension), chronic sodium retention and relative suppression of renin 55,56 .New therapeutic options, especially those with effects independent of the renin-angiotensin axis, are therefore desirable.
Preclinical evidence supports using ET antagonists for resistant hypertension.The ET-1 system is upregulated in several animal models of salt-sensitive hypertension, and ET antagonists have been shown to reduce high blood pressure and mitigate end-organ damage in these animals 57,58 .In human observational studies, ET-1 levels are elevated specifically among hypertensive patients with low renin 59 and other individuals at risk of resistant hypertension such as African Americans or individuals with obesity, obstructive sleep apnea, diabetes and CKD 56 .Despite this, previous clinical trials using the ET A selective receptor antagonist darusentan to treat resistant hypertension have demonstrated inconclusive results 60,61 .

Aprocitentan in resistant arterial hypertension
In this context, the positive results of the PRECISION (Parallel-group, phase 3 study with aprocitentan in subjects with resistant hypertension) trial testing aprocitentan, the active metabolite of macitentan 4 , marks a renaissance for ET receptor antagonists in this space.Although further evaluation of aprocitentan is required to establish whether the degree of blood pressure reduction translates to significant clinical benefit over longer-term follow-up, the critical discovery is that despite pharmacological blockade of three distinct pathways, the untreated ET-1 pathway persists to raise blood pressure.The PRECISION trial initially randomized 730 patients to placebo, aprocitentan 12.5 mg or aprocitentan 25 mg.During a 12-week screening phase before randomization, patients had been established on a maximally tolerated polypill comprising amlodipine (calcium channel antagonist), valsartan (AT 1 receptor antagonist) and hydrochlorothiazide (diuretic).The key findings were that systolic blood pressure was significantly reduced in the treatment arm after a 4-week double-blind phase.This was sustained during a 32-week non-placebo-controlled phase using 25 mg aprocitentan and reversed after a 12-week withdrawal phase, where patients were re-randomized to 25 mg aprocitentan or placebo.Noncompliance with prescribed therapy is a confounder for the diagnosis of resistant hypertension; therefore, the long-term success of aprocitentan will depend on minimizing nonadherence to combination therapy, and adequate diuretic therapy, especially given the high adverse event rate (most frequently related to symptomatic fluid retention) of 9-18% reported.
The pharmacological mechanism of aprocitentan at the doses used in PRECISION remains to be defined and will be crucial to understanding the benefit of this compound compared with other ET receptor antagonists.The relative contribution of ET A to ET B blockade is undefined for the doses used in the trial.ET B receptors function to clear circulating ET-1; therefore, significantly increased plasma ET-1 levels during treatment are a surrogate for ET B blockade and could be used to confirm engagement of this receptor subtype 9 .In healthy volunteers, plasma ET-1 concentrations did increase following treatment with aprocitentan at doses ≥25 mg 62 ; however, it is unknown if this occurs at lower doses.In previous functional assays comparing macitentan with its metabolite aprocitentan, aprocitentan demonstrated less potent antagonism (pA 2 for ET A = 6.7 versus 7.6) 63 ; however, the benefit of aprocitentan over the other clinically approved ET receptor antagonists may be underpinned by a roughly three times longer terminal plasma half-life, in the region of 45 h 9 .

Atrasentan and diabetic CKD
Diabetic nephropathy develops in 25-30% of patients with diabetes and represents the most common cause of end-stage renal failure.The principal pathology is glomerulosclerosis, resulting in albuminuria and reduced glomerular filtration rate.Glomerular ET A receptor activation causing podocyte and mesangial cell dysfunction has been shown to be an important mechanism in preclinical studies 64 .Atrasentan is a selective ET A antagonist developed as a treatment for diabetic nephropathy.The pivotal SONAR (Study of diabetic nephropathy with atrasentan) trial invoked an innovative design using a 6-week run-in period to identify responders and nonresponders 65 .Early responders demonstrating ≥30% albuminuria lowering enriched the study population versus nonresponders (~2.5:1), while patients who showed indications of fluid retention (increases in brain natriuretic peptide and body weight) were excluded.The study was terminated when it became apparent that an insufficient number of outcome-related events would be collected.However, a significant favorable hazard ratio of 0.65 was seen for the composite renal endpoint on the available data, while a nonsignificant increase in hospitalization (hazard ratio = 1.33) for heart failure was reported.Notably, the results did not differ based

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https://doi.org/10.1038/s44161-023-00347-2 on whether the patients were responders or nonresponders in the initial run-in period.These findings demonstrate the potential of ET receptor antagonists to improve outcomes for patients with CKD, supporting further development of the class, but also highlight the need to optimize both dosing and other measures to minimize fluid retention adverse effects.Atrasentan is currently being tested in the more niched indication of IgA nephropathy (Berger's disease) using the same dose used that was tested in SONAR, with a predicted study completion date of late 2025 (NCT04573478).

Clazosentan for cerebral vasospasm after aneurysmal SAH
A notable milestone for the ET-1 pathway in clinical therapeutics was the approval of the intravenous drug clazosentan, a highly selective ET A receptor antagonist for the prevention of cerebral vasospasm following aneurysmal subarachnoid hemorrhage (SAH) in Japanese patients 66 .The term SAH describes bleeding into the subarachnoid space surrounding the brain.SAH is the culprit pathology in 5% of strokes, with an estimated incidence of 2-20 per 100,000 patients per year (https:// www.nice.org.uk/guidance/ng228/) and is associated with a high risk of death and long-term morbidity.The commonest cause of nontraumatic SAH is rupture of a cerebral aneurysm, a localized vulnerable area of dilatation in an artery of the cerebral circulation.Surgical or endovascular repair (clipping/coiling) of the aneurysm is the principal therapeutic intervention.A feared complication of SAH, even after aneurysmal repair, is cerebral vasospasm resulting in delayed cerebral ischemia (DCI) and clinical deterioration, which affects around 25% of patients 4-8 days after the primary event.Medical options to prevent DCI are currently limited to nimodipine, a calcium channel antagonist, along with careful management of fluid status to maintain cerebral perfusion 67 .
Early evidence for ET-1 as a mediator of cerebral vasospasm includes biomarker case series reporting elevated cerebrospinal fluid ET-1 in positive cases 68 , as well as from animal models of SAH.For example, intra-cisternal injection of BQ-123 (experimental ET A antagonist) 69 and phosphoramidon (ECE inhibitor) 70 showed beneficial effects on cerebral vasoconstriction following experimental induction of SAH.The landmark clinical investigations using clazosentan for aneurysmal SAH are the suite of CONSCIOUS (Clazosentan to overcome neurological ischemia and infarct occurring after SAH) trials, and more recently, two Japanese phase 3 trials: JapicCTI-163368 and JapicCTI-163369.In the CONSCIOUS-1 trial 71 , 413 patients with aneurysmal SAH were randomized to placebo or clazosentan at 1, 5 or 15 mg per hour of continuous intravenous infusions within 3 days of the event and continued for up to 14 days.Compared with placebo, there was a dose-dependent relative risk reduction of moderate-to-severe angiographic vasospasm in the clazosentan-treated arm; however, there were no statistically significant differences in vasospasm-related morbidity and mortality between treatment arms.In a post hoc analysis of patients treated with neurosurgical clipping, treatment with clazosentan at 5 mg per hour was associated with a reduction in adverse clinical outcomes.This was investigated further in the larger CONSCIOUS-2 trial 72 , where 1,157 patients with aneurysmal SAH and diffuse clot on the admission computed tomography scan who were treated by neurosurgical clipping were randomized at a 2:1 ratio to clazosentan at 5 mg per hour or

Review article
https://doi.org/10.1038/s44161-023-00347-2placebo infusion.The trial reported no overall difference in rates of adverse clinical events, although there was a numerical trend toward lower vasospasm-related morbidity (driven by reduced use of rescue therapy, such as endovascular balloon angioplasty for worsening neurological status).The follow-up study, the CONSCIOUS-3 trial 73 , was terminated prematurely following the review of CONSCIOUS-2 by the data and safety monitoring board, although analysis of results obtained from the 571 already enrolled patients did suggest important risk reductions of DCI using clazosentan at the 15 mg per hour infusion rate.These positive findings were replicated in the pivotal studies testing clazosentan at the 10 mg per hour infusion rate in patients with aneurysmal SAH secured by endovascular coiling ( JapicCTI-163369) and neurosurgical clipping ( JapicCTI-163368) 3 .Both studies had identical protocols and patient numbers (n = 220) and reported significant risk reduction of the composite primary endpoint (6-week all-cause mortality and vasospasm-associated morbidity) for clazosentan versus placebo.These data underpin the approval of clazosentan in Japan.Outstanding issues requiring resolution for the neurosurgical and stroke community in other countries include a resolution of the dosing strategy (10 mg per hour versus 15 mg per hour), appraisal of the risks of serious adverse events (for example, 16.5% of clazosentan-treated patients developed pleural effusion and 11.9% developed pulmonary edema in pooled data from the Japanese studies) and confirmation of clinical benefit using longer-term functional status endpoints.GWAS have suggested EDNRA gene (which encodes the ET A receptor) variants are relevant to risk of aneurysmal SAH in Japanese people 37 .This is particularly pertinent when considering the higher incidence of SAH in Japan and perhaps raises the possibility of differential treatment response in a Western population sample.Further clarification is expected in the highly anticipated multinational REACT (Prevention and treatment of vasospasm with clazosentan) trial, which has recently completed recruitment 74 .
An alternative-use case for clazosentan has been suggested by the open-label proof-of-concept phase 2 REVERSE (Reversal of vasospasm with clazosentan post-aneurysmal subarachnoid hemorrhage) study where intravenous clazosentan infused at 15 mg per hour was shown to treat established moderate/severe cerebral vasospasm in both large cerebral vessels 75 and the distal vasculature 76 .These results highlight the potential of clazosentan as an intravenous bail-out therapy for refractory vasospasm and alternative to current invasive endovascular therapies.Complementary evidence for ET A receptor blockade as a treatment for systemic vasospasm has been suggested from ex vivo studies of peripheral resistance arterioles from patients with vasospastic angina 77 .

Sovateltide for acute ischemic stroke
Sovateltide, a modified peptide analog of ET-1 (N-succinyl-(Glu9, Ala11,15) endothelin-1), acts as an agonist with high affinity and selectivity for the ET B receptor.In humans, like many other therapeutic peptides, it has a short half-life 78 (Table 1).ET B agonist has been to target the high densities of this subtype in the human brain that have been implicated in preclinical studies in a range of neurological conditions, where there is currently an unmet need 79 .A notable milestone in 2023 (Fig. 4) was approval of sovateltide for cerebral ischemic stroke within 24 h of stroke onset 80 .This is the first-in-class approved drug selectively targeting the ET B receptor, opening an exciting frontier in ET research.A new mechanism of action is proposed, whereby sovateltide promotes differentiation of neuronal progenitors that have the capacity to differentiate into certain neuronal and glial types.In the ischemic brain, following a stroke, these differentiated cells are thought to promote neural regeneration and repair 79 .

PDE5 inhibitors and soluble guanylate cyclase stimulators
In PAH (WHO group 1 pulmonary hypertension), the therapeutic arsenal encompasses three main therapeutic targets: ET-1, prostacyclin and NO.For NO, phosphodiesterase 5 (PDE5) inhibitors prolong the action of NO in vascular smooth muscle by inhibiting the enzyme's degradation of cyclic guanosine monophosphate (cGMP).Tadalafil, a PDE5 inhibitor, has been shown to increase exercise capacity in PAH.Initial combination therapy utilizing two mechanisms with little interaction has been hypothesized as conferring additional benefits beyond the traditional approach of sequential add-on therapy.In adults with PAH, this strategy was first tested in the AMBITION (Ambrisentan and tadalafil in patients with pulmonary arterial hypertension) trial 81 .Here, 500 patients were randomized in a 2:1:1 ratio to a combination strategy of daily ambrisentan (10 mg) and tadalafil (40 mg) versus monotherapy with each agent plus placebo.The combination strategy resulted in fewer clinical failure events (a composite endpoint including death, hospitalization for worsening PAH and disease progression) over the observed period up to around 144 weeks.The results of the AMBITION trial informed an updated class I recommendation for initial combination therapy with ambrisentan and tadalafil, from the European Society of Cardiology and European Respiratory Society guidelines for PAH treatment in WHO functional class II and III patients 81,82 .
Soluble guanylate cyclase stimulators are another class of medications used to treat PAH.Similarly to PDE5 inhibitors, soluble guanylate cyclase stimulators also act on the NO pathway by increasing intracellular cGMP.A small, single-arm open-label study 83 tested the upfront combination of riociguat with ambrisentan in 20 functional class III patients with PAH; however, a high treatment discontinuation rate for one or both treatments (6-8 out of 20) in this cohort limits any meaningful assessment of the benefit of this combination over ambrisentan/ tadalafil and suggests limited tolerability.

Sodium-glucose co-transporter-2 inhibitors
ET receptor antagonists are also under investigation in combination with sodium-glucose co-transporter-2 inhibitors (SGLT2i).The primary mechanism of SGLT2i involves inhibition of the SGLT2 channel in the renal proximal convoluted tubule resulting in increased glucose excretion favoring improved glycemic control in patients with type 2 diabetes

Review article
https://doi.org/10.1038/s44161-023-00347-2mellitus (T2DM).In recent years, a wealth of evidence has accumulated for SGLT2i and dramatic decreases in cardiovascular adverse event rates, independent of glycemic status, resulting in landmark trials in heart failure 84 .Intriguingly, a post hoc analysis of the DAPA-HF (Dapagliflozin in patients with heart failure and reduced ejection fraction) trial showed that SGLT2i therapy conferred significant benefits independent of patients' baseline ET-1 levels, suggesting a rationale for combination therapy with ET A antagonists 85 .A variety of additional pleiotropic mechanisms of action for SGLT2i continue to emerge, including in the heart, where there is minimal expression of the SGLT2 receptor.SGLT2i have been shown to improve myocardial sodium homeostasis and energy metabolism, and reduce myocardial fibrosis and volume overload 86 .In the kidney, additional beneficial effects on renal hemodynamics and a notable diuretic effect involving reductions in both the intravascular and extravascular compartment have been proposed to highly complement concomitant ET receptor antagonist therapy.
A small post hoc analysis 87 in 14 patients from the SONAR trial who started an SGLT2i at the time of initiating atrasentan demonstrated a 27.1% greater reduction in albuminuria at 6 weeks than a comparator group of patients treated with atrasentan alone.At the same time, the atrasentan monotherapy group increased body weight by more than 1 kg, indicating fluid retention, while the combination group displayed a small decrease in body weight.A preclinical salt-fed rat study demonstrated that treatment with the ET antagonist zibotentan resulted in a hemodilution and increase in body weight that was prevented by co-administration of the SGLT2i dapagliflozin 88 .Together, these data suggest a combination strategy may mitigate fluid retention while providing synergistic efficacy.A phase 2b study of the highly selective ET A receptor antagonist zibotentan, in combination with dapagliflozin, an SGLT2i, is underway in CKD (NCT04724837).The same combination is also under investigation to treat liver cirrhosis with features of portal hypertension in another phase 2b study (NCT05516498).Several lines of evidence have suggested that elevated levels of ET-1 in patients with cirrhosis contribute to the development and severity of portal hypertension, which in turn results in complications leading to hospitalization, transplantation or death 89,90 .Selective ET A antagonists have been found to decrease portal pressure while maintaining liver blood flow in preclinical studies 91,92 .Additionally, rats that underwent bile duct occlusion and were treated with an ET A receptor antagonist demonstrated reduced liver collagen accumulation, indicative of reduced liver fibrogenesis, by blocking the actions of ET 93 .A number of studies have displayed positive effects of SGLT2i in patients with liver disease, likely due to their multiple metabolic actions on liver parameters 93 .A series of case studies have also suggested that SGLT2i-mediated osmotic diuresis can favorably impact ascites treatment, including electrolyte balance, by facilitating removal of accumulated extravascular fluid while circulatory volumes are maintained similar to those proposed in heart failure 94,95 .Pertinent beneficial actions of SGLT2i in liver cirrhosis include improved endothelial function, decreased inflammation and improved fluid balance, which are likely to complement the actions of ET receptor antagonists.

Dual endothelin ET A /AT 1 receptor antagonists
Combining antagonist actions against two G-protein-coupled receptors in the same molecule is a strategy that has had varied success to date given the challenge of titrating efficacy at each receptor while also achieving selectivity.However, there is a compelling rationale for simultaneously targeting the major vascular homeostasis systems: ET-1 and the renin-angiotensin-aldosterone system.Both ET-1 and angiotensin II are injurious to podocytes 64 and therefore of pathological relevance for proteinuric kidney disease.Sparsentan, a dualacting selective ET A and AT 1 receptor antagonist, was approved in 2023 for the treatment of patients with IgA nephropathy and it is being tested for other proteinuric kidney diseases, such as focal segmental glomerulosclerosis (FSGS) 96 .In the phase 2 DUET (randomized, double-blind, active-control, dose-escalation) study in patients with biopsy-demonstrated primary (idiopathic) FSGS 5 , pooled data from all dose arms demonstrated a 44.8% reduction in urine protein-tocreatinine ratio versus 18.5% for the AT 1 receptor antagonist irbesartan, while achieving an FSGS partial remission score in 28.1% versus 9.4% of patients, respectively.The US Food and Drug Administration (FDA) has requested that sparsentan is the subject of an updated risk evaluation and mitigation strategy for liver monitoring (https://ir.travere.com/news-releases/news-release-details/travere-therapeutics-announcesfda-accelerated-approval).This may be related to the high lipophilicity and high dose of sparsentan, which has been shown to be associated with drug-induced liver injury 97 .

Conclusions and future directions: pipes not the pump?
The introduction of the first-in-class nonselective ET receptor antagonist bosentan in 2002 (Fig. 4) revolutionized the treatment of PAH, with an orally active drug replacing intravenous infusions of prostacyclin receptor agonists.ET A selective antagonists (ambrisentan and sitaxentan) were subsequently found to be equally effective.Why were ET antagonists successful in PAH? ET-1 is the most efficacious and abundantly expressed constrictor (Fig. 3a,c) and promotes inflammation and cell proliferation.PAH is quintessentially a small-vessel disease.
ET-1 levels are increased in the plasma and lungs of patients with PAH, as well as in animal models, contributing to increased vasoconstriction and vascular remodeling.Despite demonstrated efficacy in this rare condition, in the ensuing decade ET antagonists did not permeate more common cardiovascular disease, resulting from a combination of factors.Although ET levels are raised in heart failure and nonselective antagonists had favorable hemodynamic effects, they failed to show any benefit in major clinical studies (early 2000s) in these patients.The precise reasons are unclear, as not all studies were fully published, but use of antagonists with greater ET A selectivity has been proposed as potentially effective in heart failure and should be tested 98 .
The recent FDA approval of an ET antagonist in aneurysmal SAH and a positive phase 3 study in resistant hypertension has led to a renaissance in ET therapeutics and reinvigorated the field.Over 20 active phase 1-4 clinical trials in cardiovascular disease are due to be reported in the next 4 years, 11 of these in phase 3.These encompass exciting new chemical entities and mechanisms of action: long-acting monoclonal antibodies to ET A receptors in PAH, the first ET B peptide agonist in cerebral ischemic stroke and dual ET A /AT 1 antagonism in antineutrophil cytoplasmic antibody vasculitis.An eclectic range of new clinical indications where there is a major unmet need will test ET A antagonist efficacy in proteinuric glomerular diseases, microvascular angina and Eisenmenger syndrome.ET A antagonists combined with SGLT2i will be evaluated in liver cirrhosis, T2DM with albuminuria and CKD.Finally, the burgeoning genetic information about variants in the ET-1 signaling pathway linked to cardiovascular disease will identify patient subgroups that are likely to benefit most from current and emerging therapeutic strategies.

1 Fig. 1 |
Fig. 1 | Synthesis of endogenous ET peptides.After removal of the signal peptide, ppET-1 is cleaved at paired basic amino acids (KR) by furin endopeptidases (blue arrows) to yield big ET-1, an inactive precursor.Big ET-1 is further cleaved by unusual proteolytic processing between W 73 and V 74 by ET-converting enzymes (yellow arrow; mainly ECE-1 at neutral pH but also by the isoform ECE-2 at acidic pH) to yield the biologically active peptide and an inactive C-terminal fragment 9 .Three further peptides have been identified as products of processing of ppET-1 within endothelial cells that correlate with ET-1 synthesis: N-terminal-proET-1 (ppET-11 8-50 ), ELDP (ppET-1 93-166 ) and C-terminal-proET-1

Fig. 2 |
Fig. 2 | Therapeutic targets of approved and investigational ET antagonistsin the human vasculature.ET-1 synthesized within vesicles of the constitutive pathway is continuously released in an abluminal manner from endothelial cells.ET-1 levels may be augmented by release from larger Weibel-Palade bodies, sentinel organelles that respond to external proinflammatory stimuli.ET-1 binding to ET A receptors, the principal subtype on smooth muscle, causes long-lasting vasoconstriction that can be fully reversed by ET A antagonists, suggesting little contribution to vasoconstriction by the low density of smooth muscle ET B receptors.ET-1-ET A complex is internalized to the endosome and the subtype may be recycled to the cell surface.Vasoconstriction is beneficially regulated by ET-1 acting in an autocrine/paracrine manner to bind endothelial Figure adapted from ref. 9 under a Creative Commons license CC BY 4.0.

Fig. 4 |
Fig. 4 | Timeline of key discoveries in the ET-1 signaling pathway and identification of selective therapeutic agents.Approved (green) and investigational (orange) antagonists; investigational ET B agonist (purple).

Fig. 5 |
Fig. 5 | Hypothesized mechanism for increased plasma ET-1 levels observed in severe COVID-19 infection.Viral entry into endothelial cells induces cell damage and release of stored ET-1 and precursors including CT-proET-1 into

Table 2 (continued) | Important clinical trials of ET antagonists with recently published results and others, currently ongoing (limited to those registered since 2017), selected for relevance to adult cardiovascular disease Review article
https://doi.org/10.1038/s44161-023-00347-2and connective tissue-associated PAH (WHO group 1 classification), a rare but debilitating disease defined by elevated pulmonary vascular resistance and progressive right heart failure.Beneficial actions in the lung are attributed to ET A receptor blockade causing vasodilatation and mitigating adverse vascular remodeling

Table 3 | ET-1-associated SNPs with significant CV disease associations identified using NHGRI GWAS catalog and/or those with reported functional effects on the ET-1 pathway SNP (nearest gene) Risk allele Variant description RAF Highest UK Biobank PheWAS Other reported CV associations (for risk allele unless specified)
The 'approved drugs' refer to selective ETA or mixed ET A /ET B antagonists.Note that following approval, sitaxentan was subsequently voluntarily withdrawn from markets globally.The ET B agonist sovateltide and dual ET A /AT 1 blocker sparsentan have both received first approval this year.AT 1 , angiotensin II type I.