Targeted therapies in Pulmonary arterial Hypertension (PAH)
Targeted Therapies In Pulmonary Arterial Hypertension
David Montani1,2,3, MD, PhD, Marie-Camille Chaumais1,2,3,4, PharmD, PhD, Christophe Guignabert, PhD1,2,3, Sven Gunther, MD1,2,3, Barbara Girerd, PhD1,2,3, Xavier Jaïs, MD, Vincent Algalarrondo, MD, PhD, Laura C. Price5, MD, PhD, Olivier Sitbon1,2,3, MD, PhD, Gérald Simonneau1,2,3, MD, Marc Humbert 1,2,3, MD, PhD.
1 Université Paris-Sud, Faculté de Médecine, Kremlin-Bicêtre, France.
2 AP-HP, Centre de Référence de l’Hypertension Pulmonaire Sévère, Service de Pneumologie et Réanimation Respiratoire, DHU Thorax Innovation, Hôpital de Bicêtre, Le Kremlin-Bicêtre, France.
3 INSERM UMR 999, LabEx LERMIT, Centre Chirurgical Marie Lannelongue, Le Plessis Robinson, France.
4 AP-HP, Service de Pharmacie, DHU Thorax Innovation, Hôpital Antoine Béclère, Clamart, France
5 National Heart and Lung Institute, Imperial College London, Royal Brompton Hospital, Dovehouse Street, London SW3 6LY, UK
Corresponding author:
Marc Humbert, MD, PhD
Centre de Référence de l’Hypertension Pulmonaire Sévère, Service de Pneumologie
Hôpital de Bicêtre, Assistance Publique Hôpitaux de Paris, Université Paris-Sud,
78, rue du Général Leclerc
94270 Le Kremlin-Bicêtre, France.
Tel: (33) 1 45 21 79 72; Fax: (33) 1 45 21 79 71;
e-mail: marc.humbert@bct.aphp.fr
CONTENTS
1. Introduction
2. Molecular basis of PAH and molecular targets
2.1 Nitric oxide (NO) pathway
2.2 Prostacyclin (PGI2) pathway
2.3 Endothelin (ET-1) pathway
2.4 Receptor- (RTKs) and non-receptor tyrosine kinase (NRTKs)
2.5The RhoA/Rho kinase (ROCK) pathway
2.6 Inflammation & Autoimmunity
2.7 Cellular metabolism
2.8 Oxidative stress (OS)
2.9 Genetic Predisposition
3. Conventional therapy
4. Approved Specific PAH therapies
4.1 Calcium channels blockers
4.2 Currently available PAH therapies target endothelial dysfunction
4.2.1 Therapies targeting PGI2 pathway: epoprostenol and derivatives
4.2.2 Therapies targeting ET-1 pathway: Endothelin receptor antagonists
4.2.3 Therapies targeting NO pathway: phosphodiesterases type 5 inhibitors
4.3 Optimization of route administration and pharmakocinetic
4.3.1 New formulation of epoprostenol Oral
4.3.2 Tissue targeting ET-1 receptor antagonist: macitentan
4.3.3 Soluble guanylate cyclase stimulator: riociguat
4.3.4 Oral PGI2 receptor agonists: selexipag
5. NEW THERAPEUTIC TARGETS
5.1 Tyrosine kinase inhibitors
5.1.1 Efficacy of kinase inhibitors in experimental models of PH
5.1.2 Evaluation of kinase inhibitors in human PAH
5.1.3 Cardiotoxicity of kinase inhibitors
5.1.4 PH induced by dasatinib
5.2 Other
5.2.1 Vascular intestinal peptide (VIP)
5.2.2 Statin
5.2.3 Rho Kinases inhibitors
5.2.4 Progenitor cells
6. ERS/ESC guidelines for the treatment of pulmonary hypertension
ABBREVIATIONS
ACVRL1: activin A receptor type II-like kinase-1
BMPR2: bone morphogenetic protein receptor type 2
cAMP: cyclic adenosine monophosphate
cGMP: cyclic guanosine monophosphate
CO: cardiac output
CYP: cytochrome P450
EC: endothelial cell
EGF: epidermal growth factor
ENG: endoglin
ET: endothelin
ERA: endothelin receptor antagonists
FGF2: fibroblast growth factor 2
iPAH: idiopathic PAH
IPr: PGI2 receptor
Kv: voltage-gated potassium channels
LVEF: left ventricular ejection fraction
MCT: monocrotaline
mPAP: mean pulmonary arterial pressure
NYHA: New York Heart Association
OS: oxidative stress
PH: pulmonary hypertension
PAH: pulmonary arterial hypertension
PASMC: pulmonary arteries smooth muscle cells
PDE-5: phosphodiesterase type 5
PDGF: platelet-derived growth factor
Ppcw: pulmonary capillary wedge pressure
PGI2: prostacycline
PVR: pulmonary vascular resistance
RTK: receptor tyrosine kinase
ROCK: RhoA/Rho kinase
ROS: reactive oxygen species
sGc: soluble gyanylyl cyclase
SOD: superoxyde dismutase
SMC: smooth muscle cell
TGF: transforming growth factor
TKI: tyrosine kinase inhibitor
TXA2: thromboxane
VIP: vasoactive intestinal polypeptide
VPAC: vasoactive intestinal polypeptide receptor
ABSTRACT
Pulmonary arterial hypertension (PAH) is a rare disorder characterized by progressive obliteration of small pulmonary arteries that leads to elevated pulmonary arterial pressure and right heart failure. During the last decades, the better understanding of the pathophysiology of the disease has resulted in the development of effective therapies targeting endothelial dysfunction (epoprostenol and derivatives, endothelin receptor antagonists and phosphodiesterase type 5 inhibitors). These drugs allow clinical, functional and hemodynamic improvement, but to date, no cure exists for PAH and prognosis remains poor. Recently, several additional pathways have been suggested to be involved in the pathogenesis of PAH, and may represent innovative therapies.
In this Review, we reviewed conventional therapy, the pharmacological agents currently available for the treatment of PAH and the benefit/risk ratio of potential future therapies.
Key words: endothelial dysfunction, pulmonary hypertension, prostacyclin, endothelin receptor antagonist, phoshodiesterases type 5 inhibitors, kinase inhibitors.
1. Introduction
Pulmonary hypertension (PH) is defined as an increase in mean pulmonary arterial pressure (mPAP) ≥ 25 mmHg at rest as assessed by right heart catheterization [pic](Galie, Hoeper et al. 2009). According to pulmonary capillary wedge pressure (Ppcw), pulmonary vascular resistance (PVR) and cardiac output (CO), different haemodynamic definitions of PH exist as follows. Pre-capillary PH includes the clinical groups 1, 3, 4 and 5 while post-capillary PH includes the clinical group 2 (Table 1)(Oudiz 2007). PAH or ‘group 1 PH’ comprises PAH due to many different etiologies, despite which patients with PAH share clinical features and pathological changes to the pulmonary circulation including endothelial dysfunction, endothelial and smooth muscle cell (SMC) proliferation, vasoconstriction and in situ thrombosis. In addition, sub-groups of PAH share similarities in term of management [pic](Simonneau, Robbins et al. 2009). However, despite many new available therapies over the last two decades, PAH remains an incurable disease process, which if not interrupted, subsequently leads to right heart failure and death (Chin and Rubin 2008).
2. MOLECULAR BASIS OF PAH AND MOLECULAR TARGETS
The imbalance in the production of endothelium-derived vasodilator and constrictor factors is a pivotal element in the development and progression of the disease (Figure 1). Over the past two decades, this observation has led to the development of all current approved specific therapies for PAH.
2.1 Nitric oxide (NO) signaling pathway and Phosphodiesterases (PDEs)
NO is a gaseous lipophilic free radical generated by three distinct isoforms of nitric oxide synthases (NOS): neuronal (nNOS), inducible (iNOS) and endothelial NOS (eNOS). NO dilates blood vessels, inhibits leukocyte adhesion, platelet aggregation, thrombus formation, and vascular proliferation and modulates many other physiological processes including cell metabolism and intracellular signaling pathways [pic](Mayer and Hemmens 1997; Lincoln, Dey et al. 1998). Reduced NO bioavailability has been reported to be associated with the development of many different vascular diseases including PAH. There are different molecular mechanisms explaining the decreased NO bioavailability in PAH including increased levels of endogenous competitive inhibitor of eNOS (such as asymmetric dimethylarginine or ADMA), eNOS “uncoupling”, decreased L-arginine levels, increased NO scavenging by haemoglobin and reactive oxygen species (ROS) [pic](Steinhorn 2008; Zuckerbraun, George et al. 2011). It has been well documented that exogenous and endogenous NO inhibit vascular SMC proliferation and migration [pic](Sarkar, Meinberg et al. 1996; Zuckerbraun, Stoyanovsky et al. 2007; Tsihlis, Oustwani et al. 2011). Most of the effects of NO, on smooth muscle cells, platelets and cardiac myocytes, are mediated through its activation of soluble guanylate cyclase and amplifying the production of cyclic guanosine monophosphate (cGMP) (Rybalkin, Yan et al. 2003) (Figure 1).
Phosphodiesterases (PDEs) are a superfamily of enzymes that inactivate cAMP and cGMP and have different tissue distributions. Among all PDEs, PDE-5 is abundantly expressed in lung tissue and is upregulated in PAH, contributing to endothelial dysfunction by inactivating cGMP. In addition, a PDE-1C upregulation in hyperproliferative pulmonary arteries SMC (PASMCs) has been noted in patients with PAH [pic](Schermuly, Pullamsetti et al. 2007). PDE-5 inhibitors facilitates the antiproliferative and vasodilating effects of endogenous NO and represent the rationale for the use of PDE inhibitors in PAH (Moncada and Higgs 1993; Rabe, Tenor et al. 1994; Giordano, De Stefano et al. 2001; Michelakis, Tymchak et al. 2002; Corbin, Beasley et al. 2005; Tantini, Manes et al. 2005; Wharton, Strange et al. 2005).
2.2 Prostacyclin (PGI2) signaling pathway
With its very short half-life (t½), the major active metabolite of arachidonic acid (AA) PGI2 is a critical endogenous regulator of vascular homeostasis. PGI2 is produced in vascular endothelial cells (ECs) and acts on neighbouring vascular SMCs as well as circulating platelets and cells (Vane and Corin 2003). PGI2 is an agonist of adenylate cyclase and a potent vasodilator with antithrombotic properties (Gryglewski, Bunting et al. 1976; Vane and Corin 2003). An imbalance in AA metabolism with decreased PGI2 and increased thromboxane (TXA2) urinary metabolites have been demonstrated in idiopathic PAH (iPAH) patients (Christman, McPherson et al. 1992). In addition, expression of the key enzyme for PGI2 synthesis PGI2 synthase (PGIS) is reduced in pulmonary arteries of PAH patients (Tuder, Cool et al. 1999) (Figure 1).
The actions of PGI2 are mediated by binding to cell surface PGI2 receptors (IPr) or by binding to nuclear peroxisome proliferator-activated receptors (PPARs) (Lim and Dey 2002; Vane and Corin 2003; Wise 2003; Falcetti, Flavell et al. 2007; Midgett, Stitham et al. 2011; Stitham, Midgett et al. 2011). Binding of PGI2 to IPr induces cell-specific signaling that lead to elevation of intracellular cAMP through Gs-protein coupling to adenylate cyclase [pic](Hashimoto, Negishi et al. 1990; Coleman, Smith et al. 1994; Chaumais, Jobard et al. 2010). Administration of PGI2 or its analogues in experimental pulmonary hypertension leads to a decrease in pulmonary arterial pressure and pulmonary vascular resistance [pic](Leffler and Hessler 1979; Archer, Chesler et al. 1986; Yuki, Sato et al. 1994; Miyata, Ueno et al. 1996; Max, Kuhlen et al. 1999).
2.3 Endothelin (ET-1) signaling pathway
ET-1 is a 21 amino acid peptide that is produced by the vascular endothelium from a 39 amino acid precursor, big ET-1, through the actions of an ET-1 converting enzyme (ECE) found on the EC membrane. In addition to ECs, PASMCs (Markewitz, Farrukh et al. 2001) and lung fibroblasts (Shi-Wen, Chen et al. 2004) can produce small amount of ET-1. The ET-1 biosynthesis is stimulated by different stimuli: including hypoxia, growth factors, cytokines, shear stress, thrombin and angiotensin II. ET-1 acts through two receptors, ET-A and ET-B. Both of these receptors are coupled to a Gq-protein and the formation of IP3. Increased inositol triphosphate (IP3) causes calcium release by the sarcoplasmic reticulum, which causes smooth muscle contraction. In addition to be a potent vasoconstrictor, ET-1 stimulates PASMC proliferation (Giaid, Yanagisawa et al. 1993). Binding of ET-1 to ETA and ETB receptors on PASMCs promotes vasoconstriction, whereas activation of ETB receptors on EC causes vasodilatation through increased PGI2 and NO levels (Hirata, Emori et al. 1993; Seo, Oemar et al. 1994) (Figure 1). Several studies in animal models document that both selective and non selective ET receptors antagonists (ERAs) prevent or attenuate pulmonary hypertension by vasodilatation, decrease of vascular remodelling and right heart hypertrophy [pic](Chen, Chen et al. 1995; Chen, Chen et al. 1997; Kim, Yung et al. 2000; Tilton, Munsch et al. 2000).
2.4 Receptor- (RTKs) and Non-Receptor Tyrosine Kinase (NRTKs)
RTKs have critical functions in several cellular processes including regulating cell proliferation, migration, metabolic changes, differentiation and survival. They are transmembrane glycoproteins that act as receptors for growth factors, neurotrophic factors, and other extracellular signaling molecules. Upon ligand binding, intracellular signal transduction pathways are activated within the cell by phosphorylating tyrosine residues on the receptors themselves (autophosphorylation) and on downstream effector molecules. The NRTKs are integral components of the signaling cascades triggered by RTKs and by other cell surface receptors such as G protein-coupled receptors and receptors of the immune system. The NRTKs are divided into ten families: Src, Abl, Jak, Ack, Csk, Fak, Fes, Frk, Tec, and Syk [pic](Blume-Jensen and Hunter 2001). In contrast to RTKs, the NRTKs lack the extracellular ligand-binding domain and transmembrane spanning region and are located in the cytoplasm or anchored to cell membrane through amino terminal modification such as myristoylation or palmitoylation. However, NRTKs possess domain that mediate protein-protein, protein-lipid and/ or protein-DNA interactions, in which the most common found protein-protein interaction domains are the Src homology 2 (SH2) and SH3 domain.
In PAH, special attention has been devoted to the pathogenic role of platelet-derived growth factor (PDGF)-, epidermal growth factor (EGF)-, fibroblast growth factor (FGF)- and c-kit-receptors. Indeed, altered expression and/ or increased activity of these four RTK signaling pathways and their contribution to the excessive proliferation and migration of SMCs and ECs have been demonstrated in human and experimental models of PH (Figure 2). In addition, their inhibition by specific inhibitors (TKIs) have been shown to exert beneficial effects in rodent models (Merklinger, Jones et al. 2005; Schermuly, Dony et al. 2005; Perros, Montani et al. 2008; Izikki, Guignabert et al. 2009; Tu, Dewachter et al. 2011; Tu, De Man et al. 2012). c-Src is abundant and plays a critical role in the integrity and function of the pulmonary vascular bed [pic](Oda, Renaux et al. 1999). Several studies have obtained evidence that Src and its signaling is abnormally activated in PAH, thereby indirectly promote cell growth and migration [pic](Courboulin, Paulin et al. 2011; de Man, Tu et al. 2012; Tu, De Man et al. 2012).
2.5 The RhoA/Rho kinase (ROCK) pathway
The small GTPase RhoA and its downstream effectors, ROCK1 and ROCK2, regulate many essential cellular processes such as contraction, cell migration, proliferation, survival, and differentiation. RhoA, which belongs to the Rho subfamily, responds to cell surface receptors for various growth factors, cytokines, adhesion molecules, and G-protein-coupled receptors by cycling between an inactive GDP-bound and an active GTP-bound conformation. The activation of RhoA subsequently activates ROCKs, which is known to phosphorylate several substrates, including myosin phosphatase target subunit 1 (MYPT-1), phosphatase and tensin homolog on chromosome 10 (PTEN), myosin light chain, ezrin/radixin/moesin (ERM) family, adducin, and LIM-kinases.
Since the RhoA/ROCK pathway plays a crucial in vasoconstriction and vascular remodeling and because it is activated by upstream key molecules for pulmonary vascular remodeling in PH such as PDGF, ET-1, serotonin, angiotensin II, several studies have been undertaken to evaluate its pathogenic role in the disease. Guilluy and collaborators (Guilluy, Eddahibi et al. 2009) have reported a twofold increase in Rho kinase and RhoA activity in lungs from patients with idiopathic PAH as compared to control patients. Consistent with this observation, Do and colleagues (Do e, Fukumoto et al. 2009) have noted close relationships between the activity of ROCK and the disease duration. In addition, beneficial effects of Rho-kinase inhibition have been observed in different cardiovascular disorders (Shimokawa and Rashid 2007) as well as in different animal models of PH i.e. in the monocrotaline-induced PH (Abe, Shimokawa et al. 2004; Nagaoka, Fagan et al. 2005; Tawara, Fukumoto et al. 2007; Mouchaers, Schalij et al. 2010), in the chronic hypoxia-induced PH (Fagan, Oka et al. 2004; Guilluy, Sauzeau et al. 2005; Abe, Tawara et al. 2006), and in the high flow-induced PH (Li, Xia et al. 2007).
2.6 Inflammation & Autoimmunity
Although the exact pathophysiology remains unknown, there is increasing evidence to suggest an important role for inflammation in the onset and progression of human and experimental PH (Perros, Montani et al. 2011). Circulating levels of several key cytokines and chemokines (such as interleukin (IL)-6, IL-1(, TNF( and MCP-1) are elevated in iPAH patients and may correlate with a worse clinical outcome (Humbert, Monti et al. 1995; Balabanian, Foussat et al. 2002; Sanchez, Marcos et al. 2007; Soon, Holmes et al. 2010). The pulmonary vascular lesions observed in lungs of iPAH patients are also sites of intense chemokine production associated with inflammatory cell recruitment and accumulation (Dorfmuller, Perros et al. 2003). Furthermore, a recent study has reported presence of pulmonary lymphoid neogenesis, revealing chronic inflammation in iPAH (Perros, Dorfmuller et al. 2012). In addition, circulating anti-EC and anti-fibroblast autoantibodies have been reported in 10 to 40 % of iPAH patients (Tamby, Chanseaud et al. 2005; Terrier, Tamby et al. 2008), suggesting a possible role of autoimmunity in the pathogenesis of PAH pulmonary vascular lesions. Consistent with this notion, a recent study has reported excessive production and release of the cytokine-like hormone leptin in PAH that act as a key modulator of the hyporesponsiveness of regulatory T cells (Treg) (Huertas, Tu et al. 2012).
The importance of inflammatory mechanisms in PAH pathophysiology has also been highlighted by the kinetics of inflammatory patterns in standard experimental models, such as monocrotaline (MCT)-induced and hypoxia-induced PAH in rats. In these models, it has been clearly shown that inflammation precedes vascular remodeling and PAH. It has also been demonstrated, particularly in MCT-induced PH, that immunosuppressive therapies prevent PH development and totally or partially reverse PH lesions (Voelkel, Tuder et al. 1994; Ikeda, Yonemitsu et al. 2002; Price, Montani et al. 2011). Finally, immune mechanisms are obviously implicated in the etiology of PAH associated with autoimmune diseases or with HIV infection (Dorfmuller, Perros et al. 2003), in which PH develops in a clear inflammatory context. In some cases, immunosuppressive or anti-inflammatory treatments significantly improve hemodynamic and clinical parameters (Jais, Launay et al. 2008), highlighting the role of immune mechanisms in PAH pathogenesis or progression.
2.7 Cellular metabolism
The Warburg effect is defined as an increased dependence on glycolysis for ATP synthesis, even in the presence of abundant oxygen. The Warburg effect has been found in a wide spectrum of human cancers as well as in PAH [pic](Rehman and Archer 2010). Restitution of oxidative metabolism with the use of dichloroacetate has been shown to be efficient in several animal models of PH [pic](Michelakis, McMurtry et al. 2002; McMurtry, Bonnet et al. 2004; Guignabert, Tu et al. 2009). Similarly, inhibition of fatty acid oxidation also prevents this metabolic shift and limits the proliferative and anti-apoptotic cell phenotype observed in PH [pic](Sutendra, Bonnet et al. 2010).
2.8 Oxidative stress (OS)
Free radicals are defined as molecules having an unpaired electron in the outer orbit. They are generally unstable and very reactive. The pathogenic role of reactive oxygen (or ROS: such as superoxide (O2- .), hydroperoxyl (HO2 .), peroxyl (RO2 .), alkoxyl (RO .), hydroxyl (OH- .), and hydroperoxyl (HO2 .) radicals) and nitrogen species (or NOS: such as nitric oxide (NO .), nitrogen dioxide (NO2 .), nitrogen trioxide (N2O3), and peroxynitrite (ONOO−)) in PH has been reviewed recently by Voelkel NF and colleagues (Voelkel, Bogaard et al. 2012) and Tabima TM and colleagues (Tabima, Frizzell et al. 2012). Indeed, it is well recognized that OS contributes to the development and/or progression of many systemic vascular disease including PAH modulating inflammation, energy metabolism, cellular differentiation, proliferation and apoptosis. Excessive OS appears following an imbalance between antioxidant and oxidant molecules or enzymes, leading to overproduction of ROS and NOS. Indeed, eNOS uncoupling, nicotinamide adenine dinucleotide phosphate (NADPH) oxidases, xanthine oxidoreductase (XOR) and mitochondria have been identified as the main contributors of this imbalance in PH. In by 2001, Geraci and al., showed an increase in pulmonary expression of oxidant genes in PAH patients’ (Geraci, Moore et al. 2001), urinary isoprostanes levels were also increased in PAH patients (Cracowski, Souvignet et al. 2001), which correlated with worse survival (Cracowski, Degano et al. 2012). Studies that followed have confirmed the presence of OS in situ in lungs of PAH patients with oxidative stress markers (8-hydroxyguanosine and nitrotyrosine) and NOX-4, an isoform of NADPH-oxidase in the media and adventitial fibroblasts of diseased vessels (Bowers, Cool et al. 2004; Mittal, Roth et al. 2007; Li, Tabar et al. 2008). A recent study reported that the transfection of a BMPR2 mutation to human PASMCs increased production of OS in vascular human SMCs, and in mice BMPR2-mutated transgenic mice, a significant increase in isoprostanes was seen in lung tissue with a specific location of vascular bed, with mitochondria being a likely source (Lane, Talati et al. 2011). Similar results were seen in mice with ALK-1 mutations with the presence of OS associated with decoupling eNOS (Jerkic, Kabir et al. 2011). Many experimental models of PH have emphasized the role of OS in pulmonary artery lesions. In MCT-induced PH in rats, levels of 8-isoprostanes and NOX-4 were increased, in association with a decrease of eNOS (Mathew, Yuan et al. 2002; Farahmand, Hill et al. 2004; Kamezaki, Tasaki et al. 2008; Csiszar, Labinskyy et al. 2009; Seta, Rahmani et al. 2011). In the rat MCT/pneumonectomy model, elevation of OS and specifically NOX-4 was also reported, and correlated with inflammation and vascular remodeling (Dorfmuller, Chaumais et al. 2011). Moreover, development of right heart failure in experimental PH was also associated with increased OS production in the right ventricle (Redout, Wagner et al. 2007). Treatment with antioxidants including superoxide dismutase (SOD), EUK-134 (a SOD and catalase mimetic) and resveratrol improved PH in MCT experimental models (Farahmand, Hill et al. 2004; Csiszar, Labinskyy et al. 2009; Redout, van der Toorn et al. 2010). In mice exposed to chronic hypoxia, an increase in OS was also noted with an overproduction of superoxide anions in pulmonary arteries; treatment with the antioxidant N-acetylcysteine significantly improved PH (Hoshikawa, Ono et al. 2001). In addition, the antioxidant edorstein in rats exposed to chronic hypoxia (Uzun, Balbay et al. 2006). More recently, in the Sugen/chronic hypoxia model, treatment with protadim (an inducer of antioxidant enzymes such as heme-oxygenase-1 and SOD) protected rats from developing right ventricular failure (Voelkel, Bogaard et al. 2012).
2.9 Genetic Predispositions
The transforming growth factor (TGF) family can be divided into two groups: 1) the bone morphogenetic proteins (BMPs) and certain “growth and differentiation factors”, which act through Smad 1, 5 and 8; 2) the TGF(s, activins, nodal and myostatin, which act through Smad 2 and 3. TGF-β signaling is initiated by the binding of TGF-β to its serine and threonine kinase receptors, the type II (TβRII) and type I (TβRI) receptors on the cell membrane. Several exonic mutations in the bone morphogenetic protein receptor type 2 (BMPR2) gene (Newman, Phillips et al. 2008), in the activin A receptor type II-like kinase-1 (ACVRL1) [pic](Girerd, Montani et al.) and to a lesser extent in the endoglin (ENG) gene have been reported in patients with heritable (70%) and idiopathic (3.5 to 40%) PAH, or in PAH associated with appetite suppressants, or in PAH and hereditary hemorrhagic telangiectasia with co-existent PAH (Deng, Morse et al. 2000; Lane, Machado et al. 2000; Trembath, Thomson et al. 2001; Humbert, Deng et al. 2002). BMPR2 gene mutations confer a reduction in BMPR-II signaling activity. However BMPR-II expression is also substantially reduced in patients with various form of PH without a mutation (Atkinson, Stewart et al. 2002), as well as in experimental animal models (Takahashi, Goto et al. 2006; Reynolds, Holmes et al. 2012). PAH patients carrying a BMPR2 mutation present approximately 10 years earlier the disease than non-carriers and have a more severe hemodynamic compromise at diagnosis (Sztrymf, Coulet et al. 2008). ACVRL1 mutation carriers are also significantly younger at PAH diagnosis, as compared with BMPR2 mutation carriers and non-carriers, suggesting a more rapid disease evolution (Girerd, Montani et al. 2010). These patients are also less likely to respond to vasodilator therapy (Rosenzweig, Morse et al. 2008). No differences in survival and time to death or lung transplantation were found in male and female PAH patients carrying a BMPR2 mutation (Girerd, Montani et al. 2010).
3. Conventional therapy
An appropriate level of physical activity is required for patients with PAH avoiding breathlessness, which can precipitate PH worsening and extreme effort limitation (Rubin 1997). In addition, situations associated with hypobaric hypoxia should be avoided, such as high altitude travel. The prevention of infections, which may provoke deteriorations in PH, is important, thus routine immunizations against influenza and pneumococcal infections are advised (Montani, Sitbon et al. 2005; Galie, Hoeper et al. 2009). Diuretics in PAH decrease right ventricular overload and improve symptoms. Careful dose adjustment is needed on the basis of clinical, echocardiographic and hemodynamic findings (Montani, Jais et al. 2005). Although there is no controlled trial examining the effect of anticoagulation in PAH, three open label studies suggest a survival benefit with oral anticoagulants (Fuster, Steele et al. 1984; Rich, Kaufmann et al. 1992; Frank, Mlczoch et al. 1997). This benefit is partially attributable to a decrease of in situ thrombosis in PAH patients characterized by right-heart failure and immobility. Vitamin K antagonists are therefore prescribed for PAH patients (INR: 1.5 to 2.5), unless there is a contraindication (Montani, Jais et al. 2005). In PAH patients with severe hypoxia (PaO2 < 60mmHg), oxygen therapy (for at 15 hours a day) can improve quality of life by improving dyspnea and exercise capacity (Montani, Jais et al. 2005; Barst, Gibbs et al. 2009), although no controlled trials have been performed. Pregnancy is associated with 30% to 50% mortality in patients with PAH and is a formal contraindication in pulmonary hypertension (Bonnin, Mercier et al. 2005). A safe and effective method of contraception is always advised in PAH women of childbearing age. However the best contraception is not yet specified (Barst, Gibbs et al. 2009; Frachon, Gaudin et al.).
4. Approved Specific PAH therapies
4.1 Calcium channels blockers (CCBs)
CCBs decrease calcium influx into SMCs of arterial wall and myocardium cells by inhibition of L-type voltage-dependant calcium channels. All CCBs are peripheral arterial dilators leading to vasodilatation. Historically, many CCBs treatments were proposed in PAH management. However, only a small proportion of patients respond to this treatment. Therefore, vasoreactivity testing is performed at the time of PAH diagnosis, for example using inhaled NO, to identify patients who may benefit from long-term therapy with CCB. A response to vasoreactivity testing is defined as a fall in mPAP by a least 10 mmHg to reach an absolute level of < 40 mmHg, with stable or increased cardiac output (Galie, Hoeper et al. 2009). Sustained benefit with CCBs is defined by the improvement of dyspnea to NYHA class II or less in concert with sustained hemodynamic improvement after a year of treatment (Galie, Torbicki et al. 2004; Sitbon, Humbert et al. 2005). The current approach is to initiate CCBs if the patient has a positive vasodilator test, adequate systemic blood pressure (above 90 mmHg) and a stable dyspnea in NYHA class I-IV prior to initiation of therapy (Barst, Gibbs et al. 2009). The agents commonly recommended are long-acting nifedipine, diltiazem or amlodipine (Barst, Gibbs et al. 2009). The choice between nifedipine and diltiazem is guided by heart rate at rest: diltiazem is only used if the value is > 80/min (Rich, Kaufmann et al. 1992). Due to its prominent negative inotropic effect, verapamil is not recommended (Packer, Medina et al. 1984).
4.2 Currently available PAH therapies target endothelial dysfunction
For many years PAH treatment was extremely limited and the prognosis was poor, with a mean survival time of less than 3 years after diagnosis (D'Alonzo, Barst et al. 1991; Badesch, Champion et al. 2009). The introduction of current therapies over the last quarter of the century has significantly improved PAH management and patients’ survival (Humbert, Sitbon et al. 2010). However, PAH remains a progressive disease with unacceptably high mortality, and no long-term exists [pic](Galie, Hoeper et al. 2009; Simonneau, Robbins et al. 2009). All current therapeutic options predominantly act to restore the imbalance in vascular tone of PAH patients.
4.2.1 Therapies targeting prostacyclin (PGI2) pathway: Epoprostenol and PGI2 derivatives
Epoprostenol
Therapeutic use of prostaglandin has a convincing rationale in PAH patients. Epoprostenol is the synthetic form (sodium salt) of PGI2, and was the first PGI2 analogue approved by the FDA in 1995. Epoprostenol binds to the IP receptor (IPr) and has two major pharmacologic actions in PAH: direct vasodilation of pulmonary and systemic arterial vascular bed, and inhibition of platelet aggregation (Friedman, Mears et al. 1997). Like native PGI2, the structure of the epoprostenol molecule is unstable in solution, prone to rapid degradation. Therefore, epoprostenol must be delivered by continuous intravenous route through a permanent indwelling central venous catheter via a portable infusion pump system. Treatment with epoprostenol therefore has limitations based on its pharmacological characteristics. Complications of chronic intravenous therapy with epoprostenol include catheter associated venous thrombosis and line-related infections (which range from exit site reactions, tunnel infections and cellulitis, to bacteremia or sepsis) (Barst, Rubin et al. 1996; Badesch, Tapson et al. 2000; Kallen, Lederman et al. 2008). Other serious drug-related complications include mild ascites and thrombocytopenia (Rubin, Mendoza et al. 1990; Chin, Channick et al. 2009). Thrombocytopenia was reported in 34 % of patients treated with epoprostenol compared with 15 % of patients receiving specific oral therapy for PAH (Chin, Channick et al. 2009). Moreover, interruption of epoprostenol delivery, either as a result of pump malfunction or catheter obstruction or damage, can result in potentially fatal rebound pulmonary hypertension. Common side effects of epoprostenol therapy include flushing, headache, jaw pain, diarrhea, nausea, photosensitivity, erythroderma, anxiety and musculoskeletal aches and pain, predominantly involving legs and feet (Barst, Rubin et al. 1996; Magnani and Galie 1996; Ahearn, Selim et al. 2002; McLaughlin, Shillington et al. 2002; Sitbon, Humbert et al. 2002). Metabolism perturbations as hyperglycemia or hyperthyroidism are also reported (Szczeklik, Pieton et al. 1980; Rubin, Mendoza et al. 1990). These side effects tend to be dose-dependent and they often respond to a cautious dose reduction. To date, epoprostenol remains the only PAH therapy to have been associated with a mortality benefit in a randomized clinical trial and, on this basis, was granted regulatory approval for treatment of patients with NYHA functional class III or IV symptoms.
Trepostinil
Other prostanoids are more chemically stable in solution and their plasma t½ is much longer than epoprostenol. Treprostinil, a tricyclic benzidine analogue of epoprostenol, is chemically stable at room temperature (for up to 72 h) and at neutral pH. These characteristics allow administration of the compound by the intravenous, subcutaneous, inhaled or oral routes. The intravenous and subcutaneous forms of treprostinil are considered to be bioequivalent at steady state at reference dose of 10 ng/kg/min (Laliberte, Arneson et al. 2004). At steady-state, the elimination t½ of treprostinil is 4.4 and 4.6 h following intravenous and subcutaneous administration, respectively (Mubarak 2010). Intravenous treprostinil provides similar beneficial hemodynamic results to epoprostenol, with an acute decrease in pulmonary vascular resistance (22% with epoprostenol and 20% reduction with treprostinil) despite a higher dose for intravenous treprostinil [pic](McLaughlin, Gaine et al. 2003; Oudiz and Farber 2009). Approval of subcutaneous treprostinil was granted in 2002 in the USA for patients with NYHA functional class II–IV symptoms. In Europe, approval was granted in 2005, but only for PAH with NYHA class III symptoms. The major limitation of this formulation is infusion site pain, which occurs in the majority of patients. A recent study reported that if up-titration beyond 6-months is tolerated, effective doses of treprostinil are reached and outcomes of patients are good with a survival rate of 57% at 9 years (Sadushi-Kolici, Skoro-Sajer et al. 2012). Intravenous treprostinil is only licensed in USA with the advantage of less frequent need for drug reservoir replacement (every 48h compared with every 12-24h for epoprostenol) (Laliberte, Arneson et al. 2004). Switching between epoprostenol and treprostinil is feasible in cases of clinical worsening or intolerance to epoprostenol (Gomberg-Maitland, Tapson et al. 2005; Reisbig, Coffman et al. 2005).
Inhaled treprostinil was approved by the FDA in 2009 for PAH patients with NYHA functional class III symptoms, after beneficial results obtained in the TRIUMPH-1 trial regarding 6-MWD and quality of life, despite no improvements in time to clinical worsening, Borg dyspnea scores or NYHA functional class (McLaughlin, Benza et al. 2010). The efficacy and safety of oral treprostinil with concomitant ERA and/or iPDE5 was studied in the FREEDOM-C study. Additional oral treatment with treprostinil did not significantly improved the primary end point of 6-minute walk distance after 16 weeks, although patients able to achieve higher treatment doses showed greatest improvement in exercise capacity (Tapson, Torres et al. 2012).
Iloprost
Iloprost is a carbacyclin analog of PGI2 available for intravenous and inhaled administration (Hoeper, Schwarze et al. 2000). Inhaled iloprost rapidly enters into the systemic circulation, with peak concentrations immediately observed after cessation of the inhalation. Iloprost has a short serum t½ (20-25 min) and it undergoes β-oxidation to an inactive metabolite (Krause and Krais 1986), therefore doses are administered 6 to 9 times a day. In the AIR study, 17% of inhaled iloprost-treated patients reached the combined primary end point of improvement in functional class at 12 weeks, compared with 5% of placebo-treated patients, and 10% increase in 6MWD in the absence of clinical deterioration or death (Olschewski, Simonneau et al. 2002). Following the AIR study, the European authorities and the FDA granted regulatory approval for inhaled iloprost in 2003 and in late 2004, respectively (Olschewski, Simonneau et al. 2002). Data from non-randomized studies suggested that intravenous iloprost has an efficacy profile similar to that of intravenous epoprostenol, with a similar improvement in hemodynamics and in exercise tolerance in PAH patients (Higenbottam, Butt et al. 1998; Opitz, Wensel et al. 2003). Intravenous iloprost can be considered for patients with advanced PAH, although it has not been formally tested in a randomized, placebo-controlled trial. Moreover, a retrospective analysis of 79 patients with PAH treated with intravenous iloprost (introduced primarily because of clinical worsening despite initial inhaled iloprost) revealed a high rate of treatment failure and poor survival (Hoeper, Gall et al. 2009). New Zealand remains the only country to have granted regulatory approval for intravenous iloprost.
Due to their similar biological profile, intravenous treprostinil and iloprost have similar side effects as epoprostenol. Dose limiting adverse events are similar for epoprostenol and intravenous treprostinil including headache, nausea, chest pain, jaw pain and restlessness [pic](McLaughlin, Gaine et al. 2003). The safety profiles of epoprostenol and intravenous treprostinil during short-term therapy appear comparable. Similarly, intravenous iloprost is well tolerated with headaches, diarrhea and abdominal pain (Oudiz and Farber 2009).
Beraprost
Oral beraprost is rapidly absorbed during fasting, peak concentration is reached after 30 min and the elimination t½ is obtained 35-40 min after oral administration (Galie, Manes et al. 2002). However, the clinical improvement in exercise capacity persists only up to 3 -to- 6 months. In addition, there is no hemodynamic benefit compared to PGI2 analogues, whereas the side effects are similar. Therefore, beraprost is neither approved in Europe nor in the US (Galie, Humbert et al. 2002; Barst, McGoon et al. 2003).
4.2.2 Therapies targeting endothelin pathway: endothelin receptor antagonists (ERAs)
Dual ERAs
Bosentan is a non-peptide pyrimidine derivative that competitively antagonizes the binding of ET-1 to both ETA and ETB receptor subtypes, and irreversibly blocks their activities (Clozel, Breu et al. 1994). Bosentan decreases pulmonary and systemic vascular resistances and increases CO without increasing heart rate. In addition to its hemodynamic effects, bosentan has been reported to significantly improve pulmonary fibrosis, and to inhibit cell proliferation of PASMCs and inflammatory response in pulmonary tissue to injection of ET-1 in guinea-pig and mouse lung (Filep, Fournier et al. 1995; Clozel and Roux 2000; Guimaraes, Da-Silva et al. 2000). Bosentan has also been reported to substantially improve the primary endpoint 6MWD, dyspnea score and delayed time to clinical worsening in the clinical trial BREATHE-1 (Rubin, Badesch et al. 2002).
Following its approval by the US Food and Drug administration (FDA) in 2001, bosentan is now available in many parts of the world. For the time being, the authorities have approved the drug only for patients with PAH of functional class III (Europe) or III/IV (USA and Canada) of the NYHA. Bosentan is associated with reversible and in most cases asymptomatic elevation of transaminases (Benedict et al., 2007; Hoeper, 2005), possibly as a consequence of the cellular accumulation of bile acids due to impaired canalicular excretion (Fattinger, Funk et al. 2001). In order to obtain further safety data, European authorities required the introduction of a post-marketing surveillance system. Within 30 months, this system has assembled data from 4,994 patients, representing 79% of those exposed to bosentan in Europe during that time period (Humbert, Segal et al. 2007). The reported annual rate of aminotransferase level elevation was 10.1%, and 3.2% of patients had to discontinue the drug for this reason. Aminotransferase level elevation was reversible in all cases, and there was no permanent liver injury. Monthly monitoring of liver transaminases is mandatory in patients treated with bosentan. Anaemia is another biologic side effect that could appear during bosentan treatment. Due to its vasodilator properties, bosentan induces potential headache and oedema. Teratogenic effects were also reported with bosentan, thus pregnancy is a formally contraindication (Dhillon and Keating 2009). However, contraception only with hormonal strategy, whatever its administration route, is not reliable due to powerful enzymatic induction of bosentan on CYP2C9 and CYAP3A4. Generally, multiple drug interactions with bosentan are reported due to its property of enzymatic induction such as cyclosporin A and glyburide (contraindicating) or simvastatin, lopinavir/ritonavir, and rifampicin (dose adjustments requiring) (Venitz, Zack et al. 2011).
Selective ET-A receptors antagonists
Selective ET-A receptor inhibition may provide benefits by preserving vasodilator and clearance functions specific to ET-B receptors, while preventing vasoconstriction and cellular proliferation mediated by ET-A receptors (Wilkins 2004). Ambrisentan is an orally active highly selective ET-A receptor antagonist approved for treatment of PAH patients in the ARIES study (Galie, Olschewski et al. 2008) although patients with certain subclasses of PAH (including portopulmonary hypertension (PoPH) and congenital pulmonary-to-systemic shunts) were excluded. Of note, patients with PoPH have also demonstrated beneficial outcomes (Cartin-Ceba, Swanson et al. 2011). Ambrisentan has low potential for drug-drug interactions and requires only once daily administration compared to bosentan (Frampton). Furthermore, ambrisentan has not been shown to increase the risk of liver dysfunction over placebo (McGoon, Frost et al. 2009). However, there is a higher incidence of peripheral edema observed in patients with PAH treated with selective ET-A receptor antagonists, such as ambrisentan, compared with dual ERAs (Trow and Taichman 2009) which is paradoxical as ETB receptors are known to promote natriuresis and diuresis. One possible explanation for different rates of edema with selective versus dual ERAs may reflect differences in affinities to the ETA receptor (Trow and Taichman 2009). Another potential explanation may involve the renin/angiotensin system (RAS). Selective ETA receptor blockade during early congestive heart failure caused sustained sodium retention by activating the RAS, resulting in edema (Schirger, Chen et al. 2004). Finally, a study in rats compared bosentan with the ERA sitaxsentan, and suggested that ERA-induced fluid retention was occurring by activation of the vasopressin system via secondary stimulation by endothelin of the uninhibited ETB receptors (Vercauteren, Strasser et al. 2012). A final consideration is the occurrence of pulmonary edema associated with circulating ET-1 via activation of the ET-B receptor that can be prevented by ET-B blockade (Comellas, Briva et al. 2009). Together these results, at least regarding peripheral and pulmonary edema, propose the safer profile of bosentan.
Sitaxsentan was another selective ET-A receptor antagonist available in Europe, Australia and Canada (Dupuis and Hoeper 2008). However, it was taken off the market after two cases of fatal liver toxicity (Lavelle, Sugrue et al. 2009; Affsaps 2010; Lee, Kirkham et al. 2011). After this market withdrawal a study assessing the efficacy and safety of lower dosages of sitaxsentan in PAH was conducted [pic](Sandoval, Torbicki et al.). In this study PAH patients mostly in WHO functional class II and receiving sitaxsentan 100 mg retained or improved their WHO functional class status and none experienced a clinical worsening event. There was no significant effect of sitaxsentan 100 mg on 6MWD; sitaxsentan 50 mg appeared subtherapeutic. Sitaxsentan was generally well tolerated. The 100-mg dose of sitaxsentan was associated with a low incidence of liver enzyme abnormalities (liver transaminases >3 _ ULN) that was similar to that observed with placebo treatment [pic](Sandoval, Torbicki et al.). This study may have been relevant if no other ERA was available on the market for PAH treatment. However, bosentan and ambrisentan are available and proved their efficacy and safety in PAH.
4.2.3 Therapies targeting NO pathway: phosphodiesterase (PDE) type 5 inhibitors
When compared with expression of PDE-5 in other tissues such as the penile corpus cavernosum and the myocardium, the expression and activity of PDE-5 is considerably higher in lung and in pulmonary vascular smooth muscle cells (Corbin, Beasley et al. 2005). Therefore, the three PDE-5 inhibitors initially approved for the treatment of erectile dysfunction (sildenafil, tadalafil, and vardenafil) were evaluated in PAH. Among them, sildenafil and tadalafil are currently approved for the treatment of PAH in NYHA functional class II and III. Sildenafil, tadalafil, and vardenafil cause significant pulmonary vasodilatation in patients with maximum effects observed after 60, 75 to 90 minutes, and 40 to 45 minutes, respectively (Ghofrani, Voswinckel et al. 2004). Sildenafil and vardenafil have very similar molecular structures, derived from cGMP, whereas tadalafil has a different chemical structure (Montani, Chaumais et al. 2009). These structural differences are reflected in the pharmacokinetic properties and selectivity for the PDE isoenzyme (Wright 2006). Both Sildenafil and vardenafil have a terminal half-life of approximately 4 hours, and tadalafil has a half-life of 17.5 hours. Due to its half-life, tadalafil is administered once per day compared to sildenafil, which have to be taken thrice per day. All three PDE-5 inhibitors are eliminated by hepatic metabolism (Wright 2006). Sildenafil is predominantly metabolized by CYP3A4 into a N-desmethyl metabolite that also has some PDE-5 activity. This metabolite is thought to account for approximately one fifth of the drug’s activity. As bosentan induces CYP3A4, this leads to a pharmacokinetic interaction whereby sildenafil plasma levels are reduced and bosentan plasma levels are increased if the two drugs are co-administered (Burgess, Hoogkamer et al. 2008). However, clinical studies reported no significant transaminases elevation as well as no clinical inefficiency of sildenafil when both drugs are coadministered (Humbert, Segal et al. 2007). Vardenafil also has an active metabolite that accounts for approximately 7% of total pharmacological activity. The activity of tadalafil is solely through the parent drug (Wright 2006). The adverse event profiles of the different PDE-5 inhibitors are generally similar. PDE-5 inhibitor class-specific side effects include headache, flushing, nasal congestion, digestive disorders (mainly dyspepsia and nausea), and myalgia, which are a reflection of their vasodilatory effects (Wright 2006; Simonneau, Rubin et al. 2008; Galie, Brundage et al. 2009; Jing, Jiang et al. 2009). Most adverse events were transient and reported as mild or moderate and are dose-dependent. Sildenafil inhibits also PDE analogs retinal PDE6 and could therefore alter the vision (Galie, Ghofrani et al. 2005).
In controlled trials, sildenafil and tadalafil monotherapy have been shown to improve clinical status, exercise capacity and hemodynamic parameters. The pivotal study of sildenafil given at 20 mg, 40 mg, or 80 mg three times a day versus placebo in more than 200 patients suffering from PAH demonstrated a significant increase in 6MWD, improvement in WHO functional class and decrease in mean PAP after 12 weeks (Galie, Ghofrani et al. 2005). Although sildenafil approval in Europe is limited to 20 mg three times daily, no data is currently available on the long-term efficacy of this lower dosage and uptitration beyond this dosage (mainly 40 mg to 80 mg three times a day) may be needed in clinical practice. In combination therapy to bosentan or epoprostenol, sildenafil improved clinical status, 6MWD, time to clinical worsening and quality of life (Hoeper, Faulenbach et al. 2004; Simonneau, Rubin et al. 2008). Efficacy and safety of tadalafil was studied in The PHIRST (Pulmonary Arterial Hypertension and ReSponse to Tadalafil) trial double blind, placebo-controlled study followed by a long-term extension phase (Galie, Brundage et al. 2009). In this 16-week study, 405 PAH patients either treatment-naïve or receiving bosentan, were randomly assigned to receive placebo or tadalafil (2.5, 10, 20 or 40 mg once daily). Tadalafil increased the 6MWD in a dose-dependent manner but only the highest dose (40 mg) reached a level of significance as well in time to clinical worsening and quality of life (Galie, Brundage et al. 2009). In clinical practice, tadalafil is given at 40 mg (2x20mg) once a day.
All of these current therapeutic options mainly act at restoring an imbalance in vascular tone by prostanoid replacement, ET-1 antagonism or PDE-5 inhibition. However, it is now well recognized that all these therapeutic agents exhibit also antiproliferative (Wharton, Davie et al. 2000; Clapp, Finney et al. 2002; Tantini, Manes et al. 2005; Wharton, Strange et al. 2005) and immunomodulatory effects (Raychaudhuri, Malur et al. 2002; Toward, Smith et al. 2004; Jaffar, Ferrini et al. 2007; Zhou, Hashimoto et al. 2007; Karavolias, Georgiadou et al. 2010), and that they act against oxidative stress (Ferrari, Cargnoni et al. 1989; Shinmura, Tamaki et al. 2005; Elgebaly, Portik-Dobos et al. 2007; Gur, Sikka et al. 2008; Shukla, Rossoni et al. 2009). Unfortunately, at this time there is no cure for PAH. Mastering of chemistry leads to the development of new promising drugs in term of kinetic and side effects, increasing our chances of having a new, better-tolerated and more powerful therapeutic tools. In addition, our improved understanding of additional pathways in this condition will presumably lead to the identification of novel molecular targets and the development of novel therapeutic strategies in the near future.
4.3 Optimization of route administration and pharmacokinetic improvements
4.3.1 Novel formulation of epoprostenol
A novel formulation of epoprostenol with a greater chemical stability is currently under investigation (EPITOME-1). It is reconstituted with sterile water for injection or sodium chloride 0.9% injection (Lambert, Bandilla et al. 2012). The reconstituted solution has a pH of > 11. This new formulation is stable for 24 h at room temperature when diluted at concentrations of 15,000 ng/ml compared to 12 h for the first formulation, reducing the frequency of use of the drug delivery pump system (Witchey-Lakshmanan L, Palepu N et al. 2010). If this new formulation has the same efficacy in PAH to the previous epoprostenol, the quality of life of patients would be likely to improve. In United States, the FDA has approved the novel formulation of epoprostenol Veletri® (Actelion Pharmaceuticals Ltd, Allschwil, Switzerland) in 2008.
4.3.2 Oral PGI2 receptor (IPr) agonists: selexipag
Clinical efficacy and tolerability of current PGI2 analogs may be compromised by concomitant activation of other prostanoid receptors. Therefore, the selectivity of PGI2 replacement therapies for the IPr has emerged as an important consideration in the management of PAH. For example, vasorelaxant effect of iloprost and beraprost is attenuated in pulmonary artery from rats with monocrotaline (MCT)-induced pulmonary hypertension (Kuwano et al, 2008) via a mechanism that involves co-activation of the contractile EP3 receptor. A promising therapeutic approach under investigation is the use of a non-prostanoid agonist to directly activate the IPr. Selexipag is a first-in-class orally active prodrug metabolized to the highly selective IPr agonist ACT-333679 (previously known as MRE-269), which has a half-life of over 6 h (Asaki, Hamamoto et al. 2007). With its high selectivity for the IPr over other prostanoid receptors (at least 130-fold selectivity), selexipag can be distinguished from beraprost or iloprost currently used in the management of PAH (Kuwano, Hashino et al. 2007). With no affinity for the prostaglandin E receptor 3 (EP3), selexipag exerts similar vasodilatory activity on both large and small pulmonary arterial branches [pic](Kuwano, Hashino et al. 2008) and its relaxant efficacy is not modified under conditions associated with PAH, whereas relaxation to treprostinil may be limited in the presence of mediators of disease such as ET-1 (Morrison, Studer et al. 2012). Preclinical study results showed that twice-daily administration of selexipag attenuates right ventricular hypertrophy, improves pulmonary hemodynamics, and significantly increases survival in MCT-treated rats [pic](Kuwano, Hashino et al. 2008). In a microdosing study using 100 μg of selexipag in healthy, white, male volunteers, headache was the most commonly reported adverse event (Kuwano, Hashino et al. 2007). A 2010 report from a phase II study, involving 43 patients with PAH, showed that treatment with selexipag for 17 weeks conferred significant improvements in PVR (-30.3% versus placebo) values compared with placebo (Simonneau, Torbicki et al. 2012). A numerical improvement in 6-MWD was also observed (+24.7 m and +0.4 m in the selexipag and placebo groups, respectively). Treatment with selexipag was well tolerated by most patients in this study. Adverse events were consistent with the known side effect profile of IPr agonism such as headache, pain in extremity, pain in jaw, nausea, and diarrhea. These side effects decreased over time in patients treated with selexipag (Simonneau, Torbicki et al. 2012). It is likely that they were related to the rapid up-titration to maximal tolerated dosage, which was relatively aggressive. Gastric side-effects including cramping, nausea, vomiting, disruption of gastric contractility and development of emesis could be related to activation of gastric smooth muscle via stimulation of EP3 receptors (Morrison, Ernst et al. 2010) (Pal, Brasseur et al. 2007) (Kan, Jones et al. 2002; Forrest, Hennig et al. 2009). In the rat stomach, selexipag does not evoke gastric contraction, nor disrupts gastric function (Morrison, Ernst et al. 2010). A role for EP3 receptors has also been invoked in the development of peripheral pain reported in patients receiving treatment with PGI2 analogs (Minami et al, 2001; Southall and Vasko, 2001). Thus, the development of a selective IPr agonist that is devoid of off-target effects may provide improved efficacy and tolerability in patients with PAH. A phase III randomized trial GRIPHON (NIH 2011) to examine the effect of selexipag on morbidity and mortality in PAH is underway and will afford more information regarding efficacy and safety of selexipag. If positive results are obtained in this clinical trial, an orally active PGI2 analog will be available for the first time in Europe and in US.
4.3.3 Tissue targeting ET-1 receptors: macitentan
Macitentan, also called ACT-064992 [N-[5-(4-bromophenyl)- 6-(2-(5-bromopyrimidin-2-yloxy)ethoxy)-pyrimidin-4-yl]-N′- propylaminosulfamide], is a novel, highly potent, tissue-targeting both ET-1 receptors and characterized by high lipophilicity [pic](Iglarz, Binkert et al. 2008). Twenty-six-weeks of multiple oral dosing (10 mg/Kg/day) of macitentan in rats leads to a maximal plasma concentration to 1.5 µg/µL and a four to five higher level of its major and pharmacology active metabolite ACT-132577. ACT-132577 is formed by oxidative depropylation of macitentan and has a longer half-life than the parent compound (8.4 h versus 2h respectively) [pic](Iglarz, Binkert et al. 2008). By remaining active in local tissue environments, macitentan and its metabolite have the advantage of extended functional in vivo half-life targeting ET-1 receptors in tissues. Moreover, macitentan increased binding to receptors than existing ERAs indicating a more potent pharmacological activity in vivo (Iglarz, Binkert et al. 2008; Kummer, Haschke et al. 2009). In rats with pulmonary hypertension, macitentan prevented both the increase of pulmonary pressure and the right ventricular hypertrophy, and it markedly improved survival, without effect on systemic arterial blood pressure (Iglarz 2008). Moreover, macitentan does not inhibit canalicular bile acid transport in rats, which could lead to better liver safety profile than bosentan (Bolly et al., J Med Chem 2012). In a phase II clinical trial, macitentan was slowly absorbed and, at a dose of 300 mg, the t½ was about 17.5 h (Sidharta, van Giersbergen et al. 2011). Formation of the ACT-132577 active metabolite is slow with maximum plasma concentrations attained at least 30 h after dosing and a t½ around 65.6 h, in accordance with a once-a-day dosing regimen. Macitentan is well tolerated: Headache, nausea and vomiting were the dose-limiting adverse events (Sidharta, van Giersbergen et al. 2011). During the time of the study, administration of macitentan had no effect on serum total bile salt concentrations, providing some support that, in contrast to other ERAs, macitentan may be devoid of liver toxic effects (Sidharta, van Giersbergen et al. 2011). However, as elevations in liver aminotransferases induced by ERAs are typically seen after repeated dosing and sometimes only after months of treatment (Humbert, Segal et al. 2007), trials with longer duration of treatment are needed to truly demonstrate a lack of liver injury associated with macitentan which was found in this phase II study. In plasma, only macitentan and the pharmacologically active oxidative depropyl metabolite, ACT-132577, were found whereas in urine two minor metabolites were detected. Another study regarding the metabolic profile of macitentan in humans by radioactivity reported that in addition to macitentan and the ACT-132577 metabolite, a carboxylic acid metabolite ACT-373898 was also identified in plasma (Bruderer, Hopfgartner et al. 2012). Moreover, in urine, four entities were identified, with the hydrolysis product of ACT-373898 as the most abundant one. In faeces, five entities were identified, macitentan, ACT-132577, ACT-373898, its hydrolysis product (M 323u), and ACT-080803 (Bruderer, Hopfgartner et al. 2012). The urinary excretion is the most important route of elimination of drug-related material than faeces in humans. Concomitant treatment with rifampin, a strong inducer of CYP3A4, reduced significantly the exposure to macitentan and its metabolite ACT-373898 at steady-state but did not affect the exposure to the active metabolite ACT-132577 to a clinically relevant extent (Bruderer, Aanismaa et al. 2012). In the phase III placebo-controlled SERAPHIN trial (NIH 2010), the efficacy and safety of macitentan will be tested in approximately 750 patients with PAH. The primary objective in this study is to establish whether treatment with macitentan prolongs time to the first morbidity or mortality event. Results are expected in the end of 2012.
4.3.4 Soluble guanylate cyclase stimulator: riociguat
Endothelium-derived NO regulates vascular homeostasis through PASMC relaxation via the activation of the second messenger cGMP. The clinical benefits associated with the PDE-5 inhibitor class has led to interest in testing whether other agents that modulate NO signaling might be similarly beneficial in PAH. Riociguat is a first-in-class drug that augments cGMP biosynthesis through direct stimulation of the enzyme soluble guanylate cyclase (sGC) promoting vasodilatation by direct stimulation of sGC in an NO-independent fashion, and by sensitization of sGC to low endogenous NO levels (Ghofrani and Grimminger 2009). Since the therapeutic effect of PDE-5 inhibitors is dependent on baseline NO expression (levels of which are typically reduced in PAH) (Evgenov, Pacher et al. 2006), treatments that act directly on sGC could potentially have a greater efficacy than PDE-5 inhibitors. sGC forms heterodimers consisting of a larger alpha-subunit and a smaller hem-binding beta-subunit (Zabel, Hausler et al. 1999). Binding of NO to the heme group leads to an approximately 200-fold increase in the conversion of GTP to cGMP (Friebe and Koesling 2003). First sGC activators such as YC-1 or BAY 41-2272 require the presence of a reduced hem group in order to activate the enzyme and can be blocked by drugs oxidizing the hem-group as the sGC inhibitor ODQ (Stasch, Becker et al. 2001). Then, a new class of NO-independent sGC-activators has been discovered, notably with the BAY 58-2667, also called cinaciguat (Stasch et al., 2002). Cinaciguat can activate the enzyme without requiring the presence of a reduced hem group and provides synergistic effects when combined with NO donors (evgenov 2006). Due to its effects in lowering systemic blood pressure, cinaciguat have led its development as treatment for acute decompensated heart failure. Further research led to riociguat (BAY 63-2521), which is structurally similar to BAY 41-2272, with improved pharmacokinetic and pharmacological properties (Mittendorf, Weigand et al. 2009). Riociguat directly stimulates sGC activity independent of NO and also acts in synergy with NO to produce anti-aggregatory, anti-proliferative, and vasodilatory effects. An initial study found that oral riociguat attenuated acute hypoxic pulmonary vasoconstriction in mice in a dose-dependent fashion and improved established MCT-induced pulmonary hypertension in rats (Schermuly, Stasch et al. 2008), with a greater effect than sildenafil in PH induced by hypoxia and SU5416 in rats (Lang, Kojonazarov et al. 2012). A phase I randomized placebo-controlled study in 58 healthy male subjects were given riociguat orally was designed to test the safety profile, pharmacokinetics and pharmacodynamics of single oral doses of riociguat (0.25–5 mg). Doses of riociguat were increased stepwise, and riociguat was well tolerated up to 2.5 mg (Frey, Muck et al. 2008). Plasma concentrations reached maximal levels 0.5 - 1.5 h after administration and declined thereafter with a terminal half-life of 5 - 10 h (Frey, Muck et al. 2008). A proof-of-concept study was conducted to investigate oral riociguat in patients with moderate to severe PH in a two-part, non-randomized, open-label, single center trial (Grimminger, Weimann et al. 2009). Riociguat was well tolerated in doses up to 2.5 mg, whereas 5.0 mg caused asymptomatic hypotension in one patient. Therefore the 2.5 mg dose (n = 10) was used in the second part of the trial to demonstrate efficacy. Riociguat significantly reduced mPAP, PVR and systemic vascular resistance and increased cardiac index CI (Grimminger, Weimann et al. 2009). Systolic blood pressure was significantly reduced but this effect was not associated with systemic hypotension. Results from a multicenter, open-label, uncontrolled phase II trial involving 75 patients with PAH (n = 33) and chronic thromboembolic pulmonary hypertension (n = 42) showed that 12 weeks of oral riociguat given 3 times daily conferred improvements in symptoms, NYHA functional class, exercise capacity, NT-proBNP level, and pulmonary hemodynamics [pic](Ghofrani, Hoeper et al.). Treatment was well tolerated overall up to the highest planned 2.5 mg dose. A decrease in systemic arterial diastolic pressure was the only significant side effect reported, with none associated with symptoms leading to a permanent discontinuation of riociguat (Ghofrani, Hoeper et al. 2010). Riociguat is also under investigation in other form of PH as PH associated with chronic obstructive pulmonary disease, with interstitial lung disease or with left ventricular dysfunction (Schermuly, Janssen et al. 2011; Ghio, Bonderman et al. 2012; Hoeper, Halank et al. 2012). An ongoing multicentre randomized, double-blind placebo-controlled phase III study of riociguat (PATENT-1) (NIH 2011) assessed the effects of riociguat on 462 patients with PAH. The primary endpoint is 6MWD; secondary endpoints include changes in PVR, NT-pro-BNP levels, WHO FC, Borg dyspnoea score, time to clinical worsening and quality of life. Results from PATENT-1will be available at the end of 2012. PATENT-2 (NCT00863681) is a non-randomized, open-label, multicenter, long-term extension study involving patients who complete PATENT-1. Another Phase III clinical study, CHEST-1 (NCT00855465), is a multicenter randomized, double blind placebo involves patients with CTEPH that is either inoperable or with persistent or recurrent PH after surgery for CTEPH. The primary and secondary endpoints are similar to those of PATENT-1. As PATENT-2, CHEST-2 (NCT00910429) is a nonrandomized, open-label, multicenter, long-term extension study involving patients who complete CHEST-1. Results from PATENT-1 and CHEST-1 are expected for 2013.
5. NEW THERAPEUTIC TARGETS
5.1 The tyrosine kinase inhibitors
5.1.1. Efficacy of kinase inhibitors in experimental models of pulmonary hypertension
Imatinib
It has been demonstrated that imatinib could reverse PAH in animal models (monocrotaline-induced or hypoxic PH) (Schermuly, Dony et al. 2005). These authors demonstrated that the development of PAH in rats exposed to monocrotaline is associated with an increase in PDGF receptor expression, and particularly in its activated (phosphorylated) form. In animal model, imatinib was able to restore a normal expression of this receptor. Moreover, a 2-week treatment by imatinib of rats exposed to monocrotaline corrected hemodynamic characteristics, reversed vascular remodelling and significantly improved survival (Schermuly, Dony et al. 2005).
Sorafenib
Sorafenib, a multikinase inhibitor (MKI) targeting c-kit, VEGFR, Raf, has been evaluated in several animal models of pulmonary hypertension (Klein, Schermuly et al. 2008; Moreno-Vinasco, Gomberg-Maitland et al. 2008). Klein et al demonstrated that sorafenib could reverse vascular remodelling of rats with monocrotaline-induced PH (Klein, Schermuly et al. 2008). Moreover, Klein et al. showed that this improvement was associated with a decrease of proliferation and an increase of apoptosis in PAH lesions. These results were confirmed in two other animal models (a conventional hypoxia-induced PH model and a model of severe PH combining dual VEGFR-1 and -2 inhibition and hypoxia). In these models, sorafenib prevents pulmonary vascular remodelling and the onset of PAH in these animal models.
Nilotinib
As imatinib, nilotinib inhibits PDGFR, c-kit and c-Abl, with a stronger effect on c-Abl than imatinib. Nilotinib has been approved for the treatment of patients with chronic myeloid leukemia in the context of imatinib resistance or intolerance. Preliminary studies have reported a benefit effect with nilotinib in experimental pulmonary hypertension. Nilotinib is under investigation in human PAH with a phase II clinical trial ( Identifier NCT01179737).
5.1.2 Evaluation of kinase inhibitors in human PAH
Imatinib
After highlighting the importance of the PDGF pathway in the pathophysiology of PAH and the possibility to reverse PAH in experimental models, several isolated cases of clinical, functional and hemodynamic have been reported (Ghofrani, Seeger et al. 2005)(Souza, Sitbon et al. 2006). Following these encouraging results, a randomized (imatinib vs placebo), double-blind, 24 weeks Phase II study was realized in 59 PAH patients in NYHA functional class II to IV receiving specific PAH therapies [pic](Ghofrani, Morrell et al.). Change from baseline in the 6MWD after 24 weeks, was the primary endpoint and was not different between the 2 groups (+22m in imatinib group as compared to placebo, p=0.21) [pic](Ghofrani, Morrell et al.). Only secondary endpoints, including cardiac output and pulmonary vascular resistance (PVR), were significantly improved. Post-hoc subgroup analyses suggested a greater improvement in patients with severe hemodynamic impairment. Serious adverse events occurred in 11 imatinib recipients (39%) and 7 placebo recipients (23%). Three deaths occurred in each group. This preliminary study does not allow concluding on the potential benefits of imatinib in PAH, but led to development of a clinical trial phase III (IMPRES) evaluating imatinib in a randomized and double-blind trial of 24-week, in severe PAH patients (PVR> 800 dynes.sec.cm-5) treated with at least two PAH specific treatments. After 24 weeks, it was observed a significantly improvement of the primary endpoint which was the 6MWD (+32m in imatinib group vs placebo) and an improvement of secondary endpoints such as hemodynamic characteristics (PVR and cardiac output). However, it was no observed improvement in NYHA functional class or in time before clinical deterioration.
This 24-week randomized trial was followed by a 3-year open-label extension phase including 143 patients, all of them receiving the imatinib treatment. An interim analysis of safety was recently reported (Hoeper, oral presentation, ATS 2012). In this study, 112 patients developed adverse events, mainly nausea (30.8%), vomiting (18.2%), peripheral edema (16.8%), ENT symptoms (15.4%), periorbital edema (15.4%), headache (14.7%) and diarrhea (12.6%). Moreover, 2 cases of subdural hematoma (SDH) were reported in the double-blind period of 6 months (both in the group treated with imatinib) and 7 supplemental cases of SDH were observed in the open-label trial, including one death directly related to this complication. Estimation of the SDH incidence >0.01 SDH /patient-year in patients treated by imatinib. The precise mechanism of SDH in these patients is not elucidated and the high incidence of SDH observed in PAH may be partly favoured by anticoagulation recommended for PAH patients. Netherless, SDH seems to be a specific complication of imatinib and SDH have been reported in other indications, including oncology or pulmonary fibrosis. Indeed, this complication never reported at this level in previous clinical trial evaluated PAH therapies.
Sorafenib
Sorafenib (MKI) was evaluated in a 16-week, phase Ib, single-center, open-label trial in PAH patients receiving parenteral prostanoids in NYHA functional class I to III. The aim of this study was to evaluate the safety of sorafenib; eleven PAH patients were treated and only 2 patients reached the maximum dose of 400mg twice daily. The most common adverse events were moderate skin reactions on the hands and feet, alopecia and diarrhea (only one adverse event (skin reaction) led to discontinuation of sorafenib) [pic](Gomberg-Maitland, Maitland et al.). No clinical improvement (6MWD +23 m, p=0.33) were observed and hemodynamic evaluation arise concerns about safety of this drug in PAH patients with a decrease of cardiac output in all the eleven patients (see above) [pic](Gomberg-Maitland, Maitland et al.).
5.1.3 Cardiotoxicity of kinase inhibitors
Although TKI and MKI represent potential innovative therapeuties in the setting of PAH, their use in clinical practice may raise safety concerns. Indeed, experimental data and human reports suggested that most of these molecules may induce cardiotoxicity. Although this risk seems rare in their common use (chronic myeloid leukemia, cancer), the existence of an underlying heart disease appears to be the main risk factor for development of cardiotoxicity. Thus, prevalence of cardiotoxicity of TKI and MKI could be higher when use in PAH patients because of the right heart failure classically associated with PAH. To date, the mechanism of this cardiotoxicity is not completely elucidated. Indeed, each molecule inhibits a large number of targets, and cardiotoxicity is probably not specific to a single target and may be shared by several MKI or TKI. Clinical trials evaluating imatinib as a treatment for chronic myeloid leukemia, have reported a high incidence of peripheral and pulmonary edema (52-68%) and 5 -16% of patients complain of dyspnea (Cohen, Williams et al. 2002).
In 2006, Kerkela et al. reported 10 patients displaying signs of cardiac dysfunction despite a normal cardiac evaluation prior to treatment with imatinib (Kerkela, Grazette et al. 2006). These patients presented a decrease in left ventricular ejection fraction (LVEF) associated with mild left ventricular dilation. Transmission electron micrographs of cardiac biopsies showed prominent membrane whorls in the myocytes, in cardiomyocyte mitochondria and in the sarcoplasmic reticulum, and glycogen accumulation. The authors demonstrated that mice treated with high-dose of imatinib may develop similar lesions in cardiomyocyte mitochondria and sarcoplasmic reticulum, associated with a decrease in LVEF (Kerkela, Grazette et al. 2006). In addition, the authors demonstrated that the main mechanism of cardiotoxicity of imatinib was associated with mitochondrial toxicity, leading to a decrease in mitochondrial membrane potential, a release of cytochrome c into the cytosol, activation of caspases 3 and 7, and cardiomyocytes apoptosis (Kerkela, Grazette et al. 2006). Interestingly, cardiomyocytes transfected with a gene encoding for a c-Abl receptor resistant to imatinib, were at least in part protected of toxicity induced by imatinib, suggesting a possible implication of this tyrosine kinase in the mechanism of imatinib induced cardiotoxicity (Kerkela, Grazette et al. 2006).
Moreover, Atallah et al. conducted a meta-analysis of patients treated with imatinib in published clinical trials (Atallah, Durand et al. 2007). Among 1276 patients enrolled, 22 (1.7%) patients were identified as having symptoms that could be attributed to systolic heart failure, 8 out of them (0.6%) were considered possibly or probably related to imatinib (Atallah, Durand et al. 2007). The two factors that seemed to favour the occurrence of cardiotoxicity were age and preexisting cardiovascular disease: congestive heart failure, diabetes, coronary artery disease, hypertension, arrhythmia, cardiomyopathy. This observation underlines the potential risk of imatinib in patients with pre-existing cardiac failure, such as PAH.
Sunitinib, a MKI used in metastatic renal-cell carcinoma and gastrointestinal stromal tumours, blocks several kinases, including PDGFR, VEGFR, c-kit, RET, FLT3 but not c-Abl. Chu et al assessed the cardiovascular risk associated with sunitinib in patients with metastatic gastrointestinal stromal tumours enrolled in a phase I/II trial (Chu, Rupnick et al. 2007). These authors showed that 11% of patients treated by sunitinib had cardiovascular events, mainly due to left heart failure (8%) (Chu, Rupnick et al. 2007). As demonstrated for imatinib, the main risk factor of developing cardiotoxicity was the presence of an underlying cardiac pathology (Chu, Rupnick et al. 2007). Echocardiography analysis of 36 patients receiving 4 weeks of sunitinib showed an absolute LVEF reduction >15% or more in seven (19%) patients (Chu, Rupnick et al. 2007). This LVEF decrease was proportional to the cumulative dose of sunitinib (Chu, Rupnick et al. 2007). Endomyocardial biopsies were obtained in two patients showing abnormalities evocative of mitochondrial toxicity. Indeed, these authors confirmed in animal model, that sunitinib may induce mitochondrial damage, release of cytochrome c and apoptosis of cardiomyocytes (Chu, Rupnick et al. 2007).
Sorafenib, another MKI used in kidney cancer and hepatocellular carcinoma, share a broadly similar profile of targets with sunitinib (inhibition of PDGFR, c-kit, VEGFR, Raf, FLT3). In an open-label trial evaluated the safety of sorafenib in 12 PAH patients, no beneficial effect was observed of the treatment but a decrease in cardiac output in all patients after 16 weeks of treatment (-0,6 L/min, p300 times in vitro) [Lombardo, 2004]. Dasatinib was initially proposed as a second-line therapy in chronic myeloid leukemia. Of note, pulmonary complications, and specifically pleural effusions, have been reported more frequently with dasatinib use compared with other TKIs [Bergeron, 2007; Masiello, 2009; Quintas-Cardama, 2007]. Recently, we reported a series of 9 cases of severe precapillary PAH confirmed by right heart catheterization, occurring in patients treated with dasatinib in France [Montani, circ 2012]. Median delay between initiation of dasatinib and PAH diagnosis was 34 months. Clinical, functional, or hemodynamic improvements were usually observed after dasatinib discontinuation. However, no patients demonstrate complete clinical and hemodynamic recovery even after long-term follow-up. Interestingly, all patients had received imatinib prior dasatinib and 6 patients received nilotinib after discontinuation of dasatinib without any recurrence of PAH, suggesting that PAH may be a specific complication of dasatinib. Paradoxically, dasatinib inhibit PDGFR, c-kit and c-Abl, and should have been a potential candidate in PAH treatment. However, dasatinib has a very wide spectrum of targets including Src kinase family, suggesting that one of these targets may be responsible of the PAH occurrence. The lowest estimate of incident PH occurring in patients exposed to dasatinib in France was 0.45% (Montani, Circ 2012). Of note, it has been recently demonstrated that first-line dasatinib may induce significantly higher and faster rates of complete cytogenetic response in newly diagnosed CML in comparison with imatinib [Kantarjian, 2010]. Therefore, this agent is likely to become increasingly prescribed in the future, and the number of cases of dasatinib-induced PAH may increase, particularly with extended duration of therapy.
5.2 Other
5.2.1 Vascular intestinal peptide (VIP)
Vascular intestinal peptide (VIP) causes PASMC relaxation (Saga and Said 1984; Nandiwada, Kadowitz et al. 1985), neutralizes a variety of pulmonary vasoconstrictors, including endothelin and hypoxia (Hamidi, Lin et al.), inhibits airway and PVSMCs proliferation (Maruno, Absood et al. 1995; Petkov, Mosgoeller et al. 2003), and has broad anti-inflammatory actions (Petkov, Mosgoeller et al. 2003). VIP acts by biding to its associated receptors, vasoactive intestinal polypeptide receptor 1 (VPAC1) and VPAC2, downstream activation of the cAMP and cGMP second messenger systems occurs, leading to modulation of vascular tone (Couvineau and Laburthe 2012). VIP was therefore considered a potent new target therapy in PAH management. Experimental studies reported involvement of VIP deficiency in the pathophysiology of PH (Said, Hamidi et al. 2007; Said 2008). Patients with PAH showed reduced lung and serum VIP levels and upregulated pulmonary artery expression of both VIP receptor subtypes (Petkov, Mosgoeller et al. 2003), suggesting that targeting VIP might be a useful therapeutic approach. VIP has been studied in clinical trials with conflicting results. Aviptadil improved exercise capacity, dyspnea, and pulmonary hemodynamics among eight treatment-naive patients with IPAH. A subsequent study that evaluated 20 patients with pulmonary hypertension of various etiologies demonstrated that inhalation of 100 μg of aviptadil was associated with transient pulmonary vasodilatation, increased stroke volume and mixed venous oxygen saturation (Leuchte, Baezner et al. 2008). A modest increase in CO was also observed. However, results from an as-yet unpublished double-blind, placebo-controlled, dose-finding phase II study involving 56 patients with PAH already treated with endothelin receptor antagonists, PDE-5 inhibitors, or both showed no significant effects on exercise capacity or pulmonary hemodynamics after addition of inhaled aviptadil (Galiè 2010). In line with these clinical results, PAH treatment with ViP appears to be compromised at least for the next few years.
5.2.2 Statins
Statins are well known inhibitors of endogenous cholesterol synthesis, acting on the 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase. Different statins were shown to improve biomarkers, hemodynamic, pulmonary vascular and right cardiac hypertrophy in animal models of PH with an additive effect in combination with specific therapeutics as tadalafil (Girgis, Li et al. 2003; Girgis, Mozammel et al. 2007); (Nishimura, Faul et al. 2002; Nishimura, Vaszar et al. 2003; Zhang, Liu et al. 2012). Human studies are limited, with one observational study showing benefits (Kao 2005), but results from small, short-term randomized trials have been conflicting. A study with pravastatin showed general benefits on both biomarkers and lung physiology (Lee, Chen et al. 2009), whereas the simvastatin as a treatment for PH trial with simvastatin added to PDE-5 inhibitors and endothelin-1 antagonists showed transient improvement in NT-proBNP levels and right ventricular mass at 6 months but not over 1 year (Wilkins, Ali et al. 2010). Similarly, rosuvastatin showed benefits on biomarkers t not on physiological function at 6 months (Barreto, Maeda et al. 2008). Statins seem to improve biomarkers in human PAH, but as yet studies are too preliminary to indicate whether these effects are clinically relevant (Katsiki, Wierzbicki et al. 2011). Recently, a randomized clinical trial of aspirin and simvastatin for pulmonary hypertension failed (Kawut, Bagiella et al. 2011).
5.2.3 Rho Kinases inhibitors
Rho Kinases inhibitors are a new class of agents which may be beneficial in the treatment of PAH and two different approaches exist to inhibit to RhoA-ROCK axis in PAH: The direct ROCK inhibition with fasudil and aminofurazan derivatives, and an indirect ROCK inhibition with statins.
Fasudil (HA-1077) is the main specific ROCK inhibitor (Nagumo, Sasaki et al. 2000). In small-caliber arteries exposed to hypoxia, fasudil inhibited the MLC phosphorylation produced by the ROCK activation and produced the relaxation of the PASMCs in a dose-dependent manner (Wang, Jin et al. 2001). Fasudil has also been reported to inhibit in vitro PDGF produced by PASMC from humans (Liu, Ling et al. 2011). In PH animal models, fasudil decreases mPAP, PVR, pulmonary vascular remodeling and right cardiac hypertrophy (Nagaoka, Fagan et al. 2005; Abe, Tawara et al. 2006; Mouchaers, Schalij et al. 2010). In monocrotaline-induced PH in rats, fasudil improved PH to a greater degree than did bosentan and sildenafil, with no synergistic effect when fasudil is combined with bosentan or sildenafil (Mouchaers, Schalij et al. 2010), whereas combination with prostacyclin has been shown a better improvement than fasudil alone (Tawara, Fukumoto et al. 2007). Fasudil was studied in PAH patients in its intravenous and inhaled form (Fujita, Fukumoto et al. 2010; Fukumoto, Matoba et al. 2005). The first clinical trial showed that intravenous fasudil exerts an acute pulmonary vasodilator effects in patients with severe PAH with a decrease of PVR by 17 % (Fukumoto, Matoba et al. 2005). However, despite a decreased PAP and increased cardiac index in these patients, there were no significant differences. A further trial of intravenous fasudil in 8 PAH patients allowed to obtain a decrease of mPAP but induced also and decrease of systemic vascular resistance and systolic arterial pressure (Rhodes, Davidson et al. 2009). These systemic effects could be avoided by the inhalation route. In 15 PAH patients, inhalation of fasudil significantly reduced mPAP and tended to decrease pulmonary vascular resistance (Fujita, Fukumoto et al. 2010). Moreover, fasudil in inhalation form selectively affects lung tissues (Fujita, Fukumoto et al. 2010). Recently, inhalation of fasudil leads to a decrease of systolic PAP and PVR in 19 patients with high-altitude PH (Kojonazarov, Myrzaakhmatova et al. 2012).
Another specific ROCK inhibitor, azaindole-1, was recently characterized, with a higher specificity than fasudil. However, despite benefit effects on PAP in PH animal models, azaindole-1 induces also a strong decrease of systemic arterial pressures (Pankey, Byun et al. 2012).
SB-772077-B is a potent aminofurazan derivative which was recently characterized and demonstrated to inhibit the ROCK activity. A treatment with in SB-772077-B rats with monocrotaline-induced PH reported a decrease of PAP but also systemic arterial pressure (Dhaliwal, Badejo et al. 2009).
In addition to its inhibition on HMG-CoA reductase, statins have also been found to inhibit synthesis of isoprenoids such as geranylgeranylpyrophosphate and farnesylpyrophosphate, which are involved in the activation of various signaling molecules such as RhoA by facilitating the mediator interaction with the cell membrane, and its coupling to GTP (Rhodes, Davidson et al. 2009). By inhibiting the addition of geranylgeranylpyrophosphate to RhoA, statins inhibit the RhoA-ROCK activation (Xing, Gan et al. 2007). These properties explain in part the benefits observed in PH animal models with statins associated to their antioxidant, anti-inflammatory and antithrombotic actions.
5.2.4 Endothelial progenitor cells
Endothelial progenitor cells (EPCs) have the property to proliferate and migrate in response to angiogenic growth factors and subsequently differentiate into mature cells in situ for blood vessel formation. In PAH patients, a deficiency of bone-marrow derived EPCs might contribute to endothelial dysfunction and the progression of the disease (Diller, van Eijl et al. 2008). Introduction of EPCs in the circulation of PAH patients could therefore repair and regenerate blood vessels improving endothelial dysfunction and damaged in pulmonary microvasculature. Administration of EPCs in rats with monocrotaline-induced PH led to prevention of PH with a decrease of RVSP and a restoring of microvasculature structure (Zhao, Courtman et al. 2005). Pulmonary hypertension and Cell Therapy (PHACeT) is a clinical trial ongoing in Canada in order to assess safety of administrating autologous mononuclear cells transduced with eNOS in idiopathic PAH patients (NIH. 2010). Associated to this safety study, efficacy of EPCs infusion have been reported in adult and children patients with idiopathic PAH with improvement on exercise capacity and pulmonary hemodynamics (Wang, Zhang et al. 2007; Zhu, Wang et al. 2008).
6. guidelines for the treatment of pulmonary hypertension AND FUTURE DIRECTIONS
In 2009, the Task Force for the Diagnosis and Treatment of Pulmonary Hypertension of the European Society of Cardiology (ESC) and the European Respiratory Society (ERS) proposed an algorithm for the treatment of pulmonary arterial hypertension. This algorithm is presented in Figure 3. First, this algorithm underlines the importance to test vasoreactivity during right heart catheterization to screen potential long-term responders to CCB.
In the absence of acute vasodilator response or absence of long-term response to CCB, specific PAH therapies targeting endothelial dysfunction (epoprostenol and prostacyclin derivatives, endothelin receptor antagonists or PDE5i) are indicated. The choice of the drugs and route of administration depends on the clinical severity based on NYHA functional class. Of note, intravenous epoprostenol is recommended as a first line therapy for all the most severe patients in NYHA functional class IV. All approved drugs (AREs, PDE5i, inhaled iloprost, i.v epoprostenol) may be proposed as first-line therapy for patients in NYHA functional class III. Few data are currently available in patients in NYHA functional class II and ERS/ESC guidelines proposed oral specific PAH therapy (AREs, PDE5i) as first-line therapy. These guidelines proposed sequential combination therapy in absence of adequate response. This combination therapy may include the three classes of drugs currently available, and no specific recommendation on the best strategy of association have been proposed. Preliminary reports suggest that a more “aggressive” therapeutic strategy with first-line triple combination therapy (prostacyclin, ARE, PDE5i) may be more effective to improve hemodynamics in severe PAH patients. The 5th World Symposium on Pulmonary Hypertension will be held in Nice in 2013 and new algorithm of treatment should be discussed including recent advances in the management of PAH.
CONCLUSION
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FIGURES
[pic]
Figure 1. Pathological pathways in pulmonary arterial hypertension and current and emerging therapeutic target therapies.
Pathogenesis of pulmonary arterial hypertension corresponds to an imbalance between excessive vasoconstrictors (endothéline-1) and lack of vasodilators (nitric oxide and prostacyclin) leading to vasoconstriction and proliferation. Pulmonary artery smooth muscle cell therapeutic targets are representing as licensed (full line blue boxes) or investigational (dotted line brown boxes) treatment approaches for pulmonary arterial hypertension. Arrows represent receptor stimulation, whereas terminated lines show receptor blockade. Abbreviations: AC, adenylate cyclase; cAMP, cyclic AMP; cGMP, cyclic GMP; ECE-1, endothelin converting enzyme 1; eNOS, endothelial nitric oxide synthase ; ET-A, endothelin receptor type A; ET-B, endothelin receptor type B; ETRA, endothelin receptor antagonists; IP, prostaglandin I2; PDE-5, phosphodiesterase type 5; PGIS, prostaglandin I synthase; sGC, soluble guanylate cyclase.
[pic]
Figure 2. Receptor tyrosine kinases involved in pathogenesis of pulmonary arterial hypertension
Binding of growth factors to their receptor tyrosine kinase induce proliferation, migration and survival of smooth muscle cells (pink), endothelial cells (blue) and activation of inflammatory cells (purple). These cells act on both leading to a self-maintained phenomenon and vascular remodeling. Abbreviations: SCF: stem cell factor; PDGF: platelet derives growth factor, PDGF-R: PDGF receptor; FGF-2: fibroblast growth factor 2: FGF-R: FGF receptor, EGF: epidermal growth factor; EGF-R: EGF receptor.
[pic]
Figure 3. Evidence-based treatment algorithm for pulmonary arterial hypertension patients (adapted from [pic]Galie, Hoeper et al. 2009).
APAH: associated pulmonary arterial hypertension; BAS: balloon atrial septostomy; CCB: calcium channel blocker; ERA: endothelin receptor antagonist; IPAH: idiopathic pulmonary arterial hypertension; PAH: pulmonary arterial hypertension; PDE-5 I: phosphodiesterase type-5 inhibitor; s.c.: subcutaneously; WHO-FC: World Health Organization functional class.
# : to maintain arterial blood O2pressure >8 kPa (60 mmHg).
Of note, sitaxsentan was taken off the market after two cases of fatal liver toxicity.
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