Recent advances in the treatment of pulmonary arterial hypertension

Drug Discovery Today: Disease Mechanisms

DRUG DISCOVERY

TODAY

Vol. 1, No. 1 2004

Editors-in-Chief Toren Finkel – National Heart, Lung and Blood Institute, National Institutes of Health, USA Tamas Bartfai – Harold L. Dorris Neurological Research Center and The Scripps Research Institute, USA

DISEASE Respiratory diseases MECHANISMS

Recent advances in the treatment of pulmonary arterial hypertension Ivan M. Robbins Department of Pulmonary Hypertension, Vanderbilt University School of Medicine Center, 1161 21st Avenue S., T-1217 MCN, Nashville, TN 37232-2650, USA

Over the past few years, the therapeutic options for the treatment of pulmonary arterial hypertension (PAH) have greatly expanded. Clinical trials have demonstrated efficacy with oral, inhaled, subcutaneous and intravenous therapies that target a variety of pathways involved in the disease process. New therapies that are commercially available include bosentan, an endothelin receptor antagonist and three prostacyclin analogues: treprostinil, beraprost and iloprost.

Introduction Pulmonary hypertension is the end-result of a multitude of processes, which are classified into five major categories (Box 1). This review will focus primarily on advanced therapy for pulmonary arterial hypertension (PAH), which can occur as an idiopathic illness (IPAH), formerly primary pulmonary hypertension, or in association with other disorders (Box 1). PAH is a rare disorder of unknown etiology, primarily affecting the small pre-capillary arteries (<100 mm in diameter). However, mutations in the type 2 receptor for bone morphogenic protein, a member of the transforming growth factor beta family, have recently been discovered as the underlying basis for familial PAH [1]. Pathological changes in PAH include smooth muscle medial hypertrophy, intimal proliferation, in situ thrombosis and the development of plexiform lesions. The incidence of PAH is unknown, but estimates for the incidence of PPH are 1–2 per 1,000,000. Patients can be divided into early and advanced disease, based on New York Heart Association (NYHA) functional E-mail address: [email protected]. 1740-6765/$ ß 2004 Elsevier Ltd. All rights reserved.

DOI: 10.1016/j.ddmec.2004.08.008

Section Editor: David Rodman—University of Colorado Health Sciences Center, Denver, CO, USA. Pulmonary arterial hypertension often presents at an advanced disease state and, although the changes that lead to disease are poorly understood, significant clinical improvement has been shown with several treatments. A variety of different pathways have been targeted for therapy, with promising results from prostacyclin analogues and endothelin receptor antagonists. The development of combination therapies targeting multiple pathways in pulmonary arterial hypertension provides a brighter outlook for the treatment of the disease. Ivan Robbins has been at the forefront of numerous clinical trials for the treatment of pulmonary arterial hypertension, and is ideally placed to provide an overview of the mechanisms involved and how they are targeted.

status, which ranges from Class I or no activity limitation to Class IV or dyspnea at rest. However, activity level does not correlate with the severity of pathological changes. No information is available about the initial stages of PAH. The pulmonary vascular disease is asymptomatic until the majority of vessels are affected. Most patients (80% or more), therefore, present with advanced disease and significant activity limitation (functional class 3 or 4) at the time of diagnosis.

Basic therapy for PAH Standard therapy for patients with early disease consists of diuretics, anticoagulation, digoxin and in the rare patients who present with significant vasoreactivity, calcium channel blockers. Despite the inability to differentiate the early and late stages of PAH other than clinically, investigators have demonstrated alterations in several vasoactive mediators and pathways, providing potential targets for therapeutic interventions for patients with advanced disease (Fig. 1). www.drugdiscoverytoday.com

129

Drug Discovery Today: Disease Mechanisms | Respiratory diseases

Box 1. Classification of pulmonary hypertensiona 1. Pulmonary arterial hypertension 2. Pulmonary venous hypertension 3. Pulmonary hypertension associated with disorders of the respiratory system and/or hypoxemia 4. Pulmonary hypertension due to chronic thromboembolic disease 5. Miscellaneous Classification of pulmonary arterial hypertension (PAH)a 1. Idiopathic (IPAH) 2. Familial (FPAH) 3. Associated with (APAH): a. Collagen vascular disease b. Congenital systemic-to-pulmonary shunts c. Portal hypertension d. HIV infection e. Drugs and toxins f. Other 4. Associated with significant venous or capillary involvement: a. Pulmonary veno-oclussive disease b. Pulmonary capillary hemangiomatosis a

Adapted from [21].

Recent efforts to treat PAH have focused less on vasodilation, and more on reversing the proliferative aspect of PAH which is now felt to play a much greater role in the disease process.

Vol. 1, No. 1 2004

Advanced therapy for PAH Prostaglandin therapy Prostacyclin is a potent vasodilator that exerts its effect by increasing intracellular cyclic adenosine monophosphate (cAMP) (Fig. 2). Prostacyclin also inhibits platelet aggregation and smooth muscle growth, has direct inotropic effects, and might inhibit oxidant stress, all of which could be more important in terms of its chronic effects in the treatment of PAH [2,3]. Prostacyclin production is decreased in patients with PAH, and decreased expression of prostacyclin synthase has been demonstrated in the pulmonary arteries of patients with PAH [2]. Continuous intravenous infusion of epoprostenol, the synthetic salt of prostacyclin, is the most effective treatment currently available for PAH. However, only two-thirds of patients with IPAH are alive three years after starting therapy [4]. Epoprostenol has a short in vivo half-life; therefore, it must be administered continuously through a permanent central venous catheter, and it can be associated with numerous side effects and complications. This has led to the development of three prostacyclin analogues that are administered subcutaneously (treprostinil), orally (beraprost) and by inhalation (iloprost). Randomized, controlled trials (RCTs) have been performed with all three analogues in patients with PAH and are summarized in Table 1. Treprostinil was evaluated in the largest

Figure 1. Genetic influences, risk factors and reported abnormalities that could be involved in initiating or maintaining the pathological changes that result in vascular remodeling in PAH.

130

www.drugdiscoverytoday.com

Vol. 1, No. 1 2004

Drug Discovery Today: Disease Mechanisms | Respiratory diseases

Figure 2. Effects of therapy on pulmonary vascular smooth muscle cell. Mediators synthesized in pulmonary endothelial cells – prostacyclin, nitric oxide and endothelin-1 either bind to receptors on smooth muscle cells or diffuse into the cytoplasm. Epoprostenol or its analogues bind to the prostacyclin receptor (IP) leading to an increase in intracellular cAMP. Nitric oxide leads to an increase in cGMP, and degradation of cGMP by phosphodiesterase 5 (PDE5) is inhibited by sildenafil. Increased levels of cGMP might also inhibit degradation of cAMP. In patients with PAH, endothelin B (ETB) receptors that function similar to endothelin A (ETA) receptors might be up-regulated on smooth muscle cells. Bosentan inhibits binding of endothelin-1 to both receptors whereas sitaxsentan and ambrisentan inhibit binding to the ETA receptor only.

study of patients with PAH and demonstrated mild improvement in exercise capacity and hemodynamics in those randomized to treprostinil, although improvement in exercise capacity was greater with higher doses [5]. Eighty-five percent of patients reported pain at the infusion site, leading to premature discontinuation from the study in 8% of patients receiving treprostinil and limiting dose escalation in the majority of patients. Inhaled iloprost was evaluated in patients with PAH as well as with chronic thromboembolic pulmonary hypertension (CTEPH), using a combined primary endpoint of improvement in NYHA functional class by at least one class and in 6-min walk distance by >10% [6]. A significantly greater number of patients receiving iloprost, compared to placebo, achieved the primary endpoint (16.8% versus 4.9%, P < 0.001). The major drawback to inhaled iloprost is the need for up to nine inhalations daily. Beraprost was evaluated in two phase III studies, with divergent results. The first study reported improvement in exercise capacity in patients receiving beraprost over a three-month period [7]. Conversely, in the second study, which evaluated patients for a longer period of time (up to 12 months), initial benefits were not sustained, and no improvement with beraprost was noted at 9 or 12 months [8].

Endothelin-1 Endothelin-1 (ET-1) is a potent endogenous vasoconstrictor that also has mitogenic properties and causes neurohumoral activation (Fig. 2). ET-1 binds to an A receptor (ETA), located

on vascular smooth muscle cells, and a B receptor (ETB), predominantly located on endothelial cells. Binding to the ETA receptor causes vasoconstriction and cellular proliferation. Conversely, binding to the ETB receptor leads to vasorelaxation through the release of prostacyclin and nitric oxide and is necessary for clearance of ET-1 as it passes through the pulmonary circulation [2]. The normal balance between production and clearance of ET-1 is lost in pulmonary hypertension. In animal models of PAH, there is evidence of up-regulation of ETB receptors on smooth muscle cells which function similarly to the ETA receptor and might amplify the vasoconstrictive and mitogenic effects of ET-1 [9]. Pulmonary arteries of patients with IPAH demonstrate much greater ET-1 immunoreactivity compared to control subjects. Plasma levels of ET-1 are also elevated and correlate with worse hemodynamics and outcome [3]. There is ongoing debate as to whether selective or non-selective ETA antagonists are of greater benefit in the treatment of PAH, and both have demonstrated benefit in PAH. Bosentan is a non-selective ET-1 receptor antagonist (ERA) and, presently, the only commercially available ERA for the treatment of PAH. A large RCT of patients with PAH showed improvement in exercise capacity with bosentan compared to placebo (Table 1) [10]. Bosentan also delayed time to clinical worsening. Sitaxsentan and ambrisentan, are selective ERAs that block the ETA receptor. Sitaxsentan was evaluated in a RCT, and exercise capacity and hemodynamics www.drugdiscoverytoday.com

131

Drug Discovery Today: Disease Mechanisms | Respiratory diseases

Vol. 1, No. 1 2004

Table 1. Placebo-controlled clinical phase 3 trials of therapy for PAH Bosentan

Treprostinil

Beraprost

Beraprost

Iloprost

Sitaxsentan

Category

ERA

Prostacyclin analogue

Prostacyclin analogue

Prostacyclin analogue

Prostacyclin analogue

ERA

Patients (n)

213

470

130

116

203

178

Population

PAH

PAH

PAH

PAH

PAH, CTEPH

PAH

Functional class

III–IV

II–IV

II–III

II–III

III–IV

N/A

Route of administration

Oral

Sub-cutaneous

Inhaled

Oral

a

Oral

Oral a

a

Dose

125 mg bid, 250 mg bid

9.3 ng/kg/min

80 mg qid

107 mg qid

5 m 6–9 times/day

100 mg/day, 300 mg/day

Study length (months)

4

3

3

12

3

3

Primary end-point

6-min walk

6-min walk

6-min walk

Disease progressionb

6-min walk + NYHA

Peak VO2

Positive treatment effect on primary end-point

Yes

Yes

Yes

No

Yes

Yesc

Treatment effect on mean change in 6-min walk distance (meters)

44

16d

25

None

36

21

Side effects/toxicity

Increased LFTs

Infusion site pain

H/A, jaw pain, nausea, diarrhea

H/A, jaw pain, diarrhea, palpitations

Flushing, syncope

Increased LFTs, INR

Abbreviations: ERA: endothelin receptor antagonist; INR: international normalized ratio; PAH: pulmonary arterial hypertension; CTEPH: chronic thromboembolic pulmonary hypertension; VO2: oxygen uptake with exercise testing; LFTs: liver function tests; H/A: headache. a Mean dose. b Disease progression = death, transplantation, epoprostenol rescue, or >25% decrease in peak VO2. c Improvement in peak VO2 only in 300 mg group. d Median change.

improved significantly with sitaxsentan compared to placebo [11]. A multicenter dose-ranging study reported improvement in exercise capacity and hemodynamics with ambrisentan [12]. Ambrisentan and sitaxsentan are currently in phase 3 RCTs. A class effect of ERAs is hepatic toxicity. Reversible liver function test abnormalities have been noted with all ERAs that have been studied in PAH, and have occurred in 6–21% of the patients during clinical testing. An important determinant of bosentan-induced liver injury appears to be inhibition of the hepatic bile salt export pump, causing intracellular accumulation of cytotoxic bile salts and hepatocyte damage [13]. Ambrisentan is metabolized via glucuronidation, and its mechanism of hepatic toxicity is uncertain. In addition to hepatic toxicity, sitaxsentan is a strong inhibitor of P450 cytochrome 2C9 activity, the enzyme through which warfarin is metabolized, and 24% of patients receiving the higher dose of 300 mg daily were noted to have an increase in international normalized ratio (INR) during the study [11].

Nitric oxide pathway Nitric oxide (NO) is a potent endogenous vasodilator and inhibitor of platelet aggregation that is important in maintaining normal vascular tone in the pulmonary circulation. NO synthase (NOS) is required to convert the precursor amino acid L-arginine to NO. Immunohistochemical analysis 132

www.drugdiscoverytoday.com

of lung samples from patients with IPAH show significantly decreased expression of endothelial NOS compared to control subjects [2]. Manipulation of the nitric oxide pathway is, therefore, an attractive therapeutic target in PAH. Improvement was demonstrated in exercise capacity and hemodynamics after one week of oral supplementation of L-arginine in 19 patients with PAH and CTEPH, suggesting that increasing the precursor for NO could be effective even with reduced eNOS [14]. No larger, long-term studies have yet been undertaken. A promising therapy for PAH is augmentation of cyclic guanosine monophosphate (cGMP). NO activates guanylate cyclase, which converts guanosine triphosphate to cGMP. This leads to the activation of protein kinase G, which initiates a protein phosphorylation cascade leading to decreased intracellular calcium and vascular smooth muscle relaxation. Cyclic GMP is hydrolyzed to its inactive form by phosphodiesterases (PDEs) 5, 6 and 11, and a significant amount of PDE5 is present in the lung. Sildenafil is a highly specific PDE5 inhibitor, and by inhibiting PDE5, cGMP is allowed to accumulate intracellularly (Fig. 2) [2]. There is growing data to support the efficacy of sildenafil in the treatment of patients with PAH; however, there are no published studies of large randomized, long-term trials. Several single-dose studies of sildenafil in patients with pulmonary hypertension (PH) have demonstrated acute

Vol. 1, No. 1 2004

Drug Discovery Today: Disease Mechanisms | Respiratory diseases

improvement in hemodynamics. Small case series of patients treated chronically with sildenafil, up to 100 mg tid, have reported conflicting results. The most commonly reported side effects include headache, flushing, dyspepsia and nasal congestion; systemic blood pressure effects are mild. A large RCT has recently been completed which will provide more definitive data regarding the efficacy of sildenafil in PAH.

Combination therapy The combination of medications targeting different pathological pathways is beginning to be evaluated in PAH. A small RCT that compared epoprostenol alone to the combination of epoprostenol and bosentan in 33 patients with PAH initiating epoprostenol therapy was performed (unpublished data). There was no statistical difference between the treatment arms with regard to improvement in exercise capacity or hemodynamics, although there was a trend towards greater hemodynamic improvement with combination therapy compared to epoprostenol alone. Two patients died in the combination therapy arm during the study; however, both deaths were felt to be related to disease progression and not to the addition of bosentan. A small open-label study additionally giving bosentan to 20 patients receiving either inhaled iloprost or oral beraprost showed a significant increase in exercise capacity after three months of combination therapy which was well tolerated in this cohort [15]. Studies of large cohorts are needed to determine whether the combination of prostaglandin therapy and bosentan is more efficacious. Elevated levels of cGMP can inhibit PDE3 hydrolysis of cAMP, thus allowing cAMP to accumulate, a phenomenon known as ‘‘cross-talk’’ (Fig. 2). Small, open-label studies have compared the short-term effects of the combination of iloprost and sildenafil to each agent alone and have shown that sildenafil plus iloprost is more effective in improving acute hemodynamics than either agent alone. A long-term, openlabel study of 14 patients with clinical deterioration, despite chronic treatment with inhaled iloprost, demonstrated rever-

sal of clinical deterioration and significant improvement in exercise capacity and hemodynamics after three months of combination therapy [16]. Improvement in 6-min walk distance was sustained for up to 12 months. Although two patients died of pneumonia, this was felt to be unrelated to combination therapy, which was well tolerated. Currently, a large RCT is underway to evaluate the long-term safety and efficacy of the combination of epoprostenol with sildenafil compared to epoprostenol alone.

Potential targets for future therapy Potential therapeutic targets are listed in Table 2. Increased expression of the serotonin transporter (SERT), leading to increased intracellular uptake of serotonin, has been demonstrated in patients with IPAH, and this correlates with increased growth of pulmonary artery smooth muscle cells in vitro compared to controls [17]. The ability of selective serotonin reuptake inhibitors to bind to the SERT and modulate its function suggests a new therapeutic approach for the treatment of PAH. Vasoactive intestinal peptide (VIP) has many properties similar to epoprostenol, such as vasodilation and inhibition of platelet aggregation and smooth muscle growth. A recent open-label study of eight patients with IPAH demonstrated improvement in exercise capacity and hemodynamics after three months of inhaled VIP [18]. Although promising, these results need to be duplicated in a large cohort. Abnormalities in voltage-gated potassium channels, causing increased intracellular calcium levels and vasoconstriction, have been shown in patients with IPAH [19]. Improving the function of these channels provides another potential target for treatment. Angiopoietin-1 (Ang-1) is a growth factor secreted by smooth muscle cells that is necessary for vasculogenesis but not normally found in mature human lungs. Increased Ang-1 mRNA has recently been demonstrated in lung samples from patients with PH due to multiple causes [20]. Modulation of this mediator is another potential target for treatment.

Table 2. Potential targets of therapy for PAH Therapy against target

Stage of development

Avantages and/or disadvantages

Who is working on the target?

Refs.

Phosphodiesterase 5 (PDE 5)

Sildenafil (inhibits PDE 5 activity and prevents breakdown of cGMP)

Phase 3 studies in progress

Oral therapy, well tolerated, good safety profile/3x per day dosing

Pfizer, Inc

[16]

Serotonin transporters

Selective serotonin reuptake inhibitors (inhibits binding of serotonin to receptor)

Not evaluated in PAH

Oral therapy, well tolerated, good safety profile, once daily dosing

N/A

[17]

Vasoactive intestinal peptide

Vasoactive intestinal peptide (increases cAMP and cGMP)

Phase 2 study completed

Inhaled therapy

N/A

[18]

Voltage-gated K+ channels

Dichloroacetate (increases expression or function of Kv channels)

Not evaluated in PAH

N/A

N/A

[19]

Angiopoietin

None

Not evaluated in humans

Unsure whether Angiopoietin is beneficial or determental

N/A

[20]

Abbreviations: cAMP, cyclic adenosine monophosphate; cGMP, cyclic guanosine monophosphate; Kv, potassium voltage-gated channels; PAH, pulmonary arterial hypertension. www.drugdiscoverytoday.com

133

Drug Discovery Today: Disease Mechanisms | Respiratory diseases

Vol. 1, No. 1 2004

Summary The initial events in the sequence of changes that eventually lead to advanced PAH remain unknown. The significant clinical improvement demonstrated with epoprostenol, and to a lesser extent with analogues of prostacyclin, supports an important role for a deficiency of prostacyclin in the pathogenesis of PAH. Increased ET-1and decreased eNOS in patients with PAH, in conjunction with the beneficial effects of ERAs and the preliminary reports of improvement with sildenafil, also support a role for alterations in the ET-1 and NO pathways in the pathogenesis of PAH. Presently, there are minimal long-term data regarding the effects of these therapies. Combination therapy in the treatment of PAH has only recently been attempted, but, mechanistically, it is logical to target more than one pathway in the treatment of this disorder. Identification of the role of bone morphogenetic protein receptor 2 in the pathogenesis of IPAH might provide important insight into the initiating events that ultimately lead to disease expression and progression, not only in IPAH but in PAH, and might provide additional pathways at which to aim therapy. Whether beneficial regression of vascular remodeling occurs with any therapy, alone or in combination, even in patients who demonstrate marked hemodynamic improvement, is uncertain. Unfortunately, there are currently no methods that can directly evaluate the pulmonary micro-vasculature in vivo to determine the effects of therapy. Lung specimens are available only from explanted lungs of patients who undergo lung transplantation, or from autopsy specimens in patients who have died. This is a very skewed group of patients, all of whom have failed therapy, and, not surprisingly, show no evidence of beneficial regression of vascular remodeling. Until an animal model or techniques to image the pulmonary microvasculature in vivo are available, it will not be possible to determine whether therapy reverses the pathological changes. These, and other limitations are impediments to the development of better medical therapy for PAH.

Conclusion During the last decade, the first effective treatments specifically for PAH have emerged. Improvement in exercise capacity has recently been demonstrated with prostacyclin analogues and ERAs. Information on long-term benefit is being accrued and should be available in the next few years as experience increases with newer agents. PDE5 inhibitors appear promising in the treatment of PAH, and the results from a large randomized study will also be available in the near future. Despite these important improvements in therapy, mortality and morbidity associated with PAH remains unacceptably high. Well designed RCTs, using combination therapy that target multiple pathways, are needed to further the progress that has already been made in the treatment of PAH. 134

www.drugdiscoverytoday.com

Related articles Giaid, A. et al. (1993) Expression of endothelin-1 in the lungs of patients with pulmonary hypertension. N. Engl. J. Med. 328, 1732–1739 Giaid, A. and Saleh, D. (1995) Reduced expression of endothelial nitric oxide synthase in the lungs of patients with pulmonary hypertension. N. Engl. J. Med. 333, 214–221 Gaine, S.P. et al. (1998) Primary pulmonary hypertension. Lancet 352, 719–725 Lane, K.B. et al. (2000) Heterozygous germline mutations in BMPR2, encoding a TGF-beta receptor, cause familial primary pulmonary hypertension. The International PPH Consortium. Nat. Genet. 26, 81–84 McLaughlin, V.V. et al. (2002) Survival in primary pulmonary hypertension: the impact of epoprostenol therapy. Circulation 106, 1477– 1482

Acknowledgement The author thank Tamara Lasakow for editorial assistance with the manuscript.

References 1 Lane, K.B. et al. (2000) Heterozygous germline mutations in BMPR2, encoding a TGF-beta receptor, cause familial primary pulmonary hypertension. The International PPH Consortium. Nat. Genet. 26, 81–84 2 Galie, N. et al. (2002) Emerging medical therapies for pulmonary arterial hypertension. Prog. Cardiovasc. Dis. 45, 213–224 3 Bowers, R. et al. (2004) Oxidative stress in severe pulmonary hypertension. Am. J. Respir. Crit. Care Med. 169, 764–769 4 McLaughlin, V.V. et al. (2002) Survival in primary pulmonary hypertension: the impact of epoprostenol therapy. Circulation 106, 1477–1482 5 Simonneau, G. et al. (2002) Continuous subcutaneous infusion of treprostinil, a prostacyclin analogue, in patients with pulmonary arterial hypertension: a double-blind, randomized, placebo-controlled trial. Am. J. Respir. Crit. Care Med. 165, 800–804 6 Olschewski, H. et al. (2002) Inhaled iloprost for severe pulmonary hypertension. N. Engl. J. Med. 347, 322–329 7 Galie, N. et al. (2002) Effects of beraprost sodium, an oral prostacyclin analogue, in patients with pulmonary arterial hypertension: a randomized, double-blind, placebo-controlled trial. J. Am. Coll. Cardiol. 39, 1496–1502 8 Barst, R.J. et al. (2003) Beraprost therapy for pulmonary arterial hypertension. J. Am. Coll Cardiol. 41, 2119–2125 9 Black, S.M. et al. (2003) Emergence of smooth muscle cell endothelin Bmediated vasoconstriction in lambs with experimental congenital heart disease and increased pulmonary blood flow. Circulation 108, 1646–1654 10 Rubin, L.J. et al. (2002) Bosentan therapy for pulmonary arterial hypertension. N. Engl. J. Med. 346, 896–903 11 Barst, R.J. et al. (2004) Sitaxsentan therapy for pulmonary arterial hypertension. Am. J. Respir. Crit. Care Med. 169, 441–447 12 Rubin, L. (2004) Ambrisentan improves exercise capacity and clinical measures in pulmonary arterial hypertension (PAH). Am J. Respir. Crit. Care Med. 169, A210 13 Fattinger, K. et al. (2001) The endothelin antagonist bosentan inhibits the canalicular bile salt export pump: a potential mechanism for hepatic adverse reactions. Clin. Pharmacol. Ther. 69, 223–231 14 Nagaya, N. et al. (2001) Short-term oral administration of L-arginine improves hemodynamics and exercise capacity in patients with precapillary pulmonary hypertension. Am. J. Respir. Crit. Care Med. 163, 887–891 15 Hoeper, M.M. et al. (2003) Bosentan treatment in patients with primary pulmonary hypertension receiving nonparenteral prostanoids. Eur. Respir. J. 22, 330–334

Vol. 1, No. 1 2004

16 Ghofrani, H.A. et al. (2003) Oral sildenafil as long-term adjunct therapy to inhaled iloprost in severe pulmonary arterial hypertension. J. Am. Coll. Cardiol. 158–164 17 Eddahibi, S. et al. (2001) Serotonin transporter overexpression is responsible for pulmonary artery smooth muscle hyperplasia in primary pulmonary hypertension. J. Clin. Invest. 108, 1141–1150 18 Petkov, V. et al. (2003) Vasoactive intestinal peptide as a new drug for treatment of primary pulmonary hypertension. J. Clin. Invest. 111, 1339–1346

Drug Discovery Today: Disease Mechanisms | Respiratory diseases

19 Yuan, J.X. et al. (1998) Dysfunctional voltage-gated K+ channels in pulmonary artery smooth muscle cells of patients with primary pulmonary hypertension. Circulation 98, 1400–1406 20 Du, L. et al. (2003) Signaling molecules in nonfamilial pulmonary hypertension. N. Engl. J. Med. 348, 500–509 21 Simonneau, G. et al. (2004) Clinical classification of pulmonary hypertension. J. Am. Coll. Cardiol. 12 (Suppl. 1), S5–S12

www.drugdiscoverytoday.com

135