Therapeutic Potential of Functional Selectivity in the Treatment of Heart Failure

heart function and prevent fibrosis in cardiomyopathy caused by lamin A/C gene mutation. Circulation 123:53– 61. Wu W, Shan J, Bonne G, et al: 2010. Pharmacological inhibition of c-Jun N-terminal kinase signaling prevents cardiomyopathy caused by mutation in LMNA gene. Biochim Biophys Acta 1802:632– 638.

Wydner KL, McNeil JA, Lin F, et al: 1996. Chromosomal assignment of human nuclear envelope protein genes LMNA, LMNB1, and LBR by fluorescence in situ hybridization. Genomics 32:474 – 478.

PII S1050-1738(11)00081-8

TCM

Therapeutic Potential of Functional Selectivity in the Treatment of Heart Failure Gitte Lund Christensena, Mark Aplinb, and Jakob Lerche Hansenc,d*

Adrenergic and angiotensin receptors are prominent targets in pharmacological alleviation of cardiac remodeling and heart failure, but their use is associated with cardiodepressant side effects. Recent advances in our understanding of seven transmembrane receptor signaling show that it is possible to design ligands with “functional selectivity,” acting as agonists on certain signaling pathways while antagonizing others. This represents a major pharmaceutical opportunity to separate desired from adverse effects governed by the same receptor. Accordingly, functionally selective ligands are currently pursued as next-generation drugs for superior treatment of heart failure. (Trends Cardiovasc Med 2010;20:221–227) © 2010 Elsevier Inc. All rights reserved. • Introduction Cardiac seven transmembrane receptors (7TMRs) or G protein-coupled receptors a Gitte Lund Christensen is at the Department of Clinical Biochemistry, Glostrup Research Institute, Glostrup Hospital, DK-2600 Glostrup, Denmark. b Mark Aplin is at the Department of Cardiology, Heart Centre, Copenhagen University Hospital, Rigshospitalet, DK-2100 Copenhagen O, Denmark. cJakob Lerche Hansen is at the Laboratory for Molecular Cardiology, The Danish National Research Foundation Centre for Cardiac Arrhythmia, Department of Biomedical Sciences, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3b, DK-2200 Copenhagen N, Denmark, and dNovo Nordisk A/S, Novo Nordisk Park, Diabetes NBEs and Obesity Biology, DK-2760 Måloev, Denmark. *Address correspondence to: Jakob Lerche Hansen, Novo Nordisk A/S, Novo Nordisk

TCM Vol. 20, No. 7, 2010

(GPCRs) such as the adrenergic and angiotensin receptors are profoundly involved in the development of cardiac hypertrophy and heart failure and are consequently among the most prominent pharmacological targets to treat this group of life-threatening diseases (Jessup et al. 2009). Clinicians are aware of the drawbacks of current ␤-adrenergic receptor blockers (␤-blockers) and angiotensin receptor blockers (ARBs) as cardiodepressant effects can greatly complicate their administration to patients in greatest need of intervention. Despite intensive research in the field, progression of maladaptive cardiac remodeling is not sufficiently prevented by Park, Diabetes NBEs and Obesity Biology, DK-2760 Måloev, Denmark. Tel.: (⫹45) 30791493; e-mail: [email protected]. © 2010 Elsevier Inc. All rights reserved. 1050-1738/$-see front matter

current treatments, and the 5-year survival rate for heart failure is as low as 42%, which is worse than that for most cancers (Askoxylakis et al. 2010, Stewart et al. 2001). These discouraging statistics underscore the need for development of new treatment strategies for heart failure. One promising concept emerging from basic research of the currently targeted cardiac receptors is the ability of receptor ligands to target subsets of receptor actions, referred to as “functional selectivity” or “biased agonism”. Accordingly, next-generation therapeutics may selectively inhibit deleterious receptor-mediated effects in the heart while simultaneously promoting alternative pathways activated by the same cardiac 7TMRs to increase cardiac output and confer cell protection. Here, we review the accumulating evidence that pharmacological dissection of G protein-dependent and -independent signaling downstream of the angiotensin II type I receptor (AT1R) and ␤-adrenergic receptors (␤-ARs) could have the potential to improve cardiac function and alleviate heart disease. • Functional Selectivity 7TMRs are the most successful pharmacological targets, accounting for more than 30% of registered prescription drugs on the market today (Allen and Roth 2011). The 7TMR targeting drugs currently applied in the clinic have largely been identified by using screening assays for G protein activity in the form of cAMP or inositol phosphate (IP) production. This methodology reflects the perception that all pathways and biological effects controlled by a given receptor are activated through their cognate G protein serving as a “central hub”. Based on this understanding, compounds have been classified as full agonists, partial agonists, inverse agonists, and neutral antagonists in relation to their intrinsic efficacy to stimulate or inhibit G protein activation and/or specific cellular phenotypes (Figure 1A). Intense basic research on 7TMR signaling has shown that multiple other proteins, such as ␤-arrestins, G proteincoupled receptor kinases, receptor tyrosine kinases, and small G proteins, interact directly with 7TMRs. ␤-Arrestins have been studied extensively with regard to their ability to cause receptor internalization, G protein uncoupling, 221

and initiation of “second-wave” signaling pathways. Studies of mutant AT1R and ␤-ARs have demonstrated that these alternative signaling pathways do not necessarily depend on prior G protein desensitizing mechanisms but, rather, may be elicited independently (Gaborik et al. 2003, Seta et al. 2002, Wei et al. 2003). These effects can also be separated by “biased” or “functionally selective” receptor ligands conferring different receptor conformations with subsequent differential coupling to central signal initiators. Thus, it is possible to pharmacologically induce, for example, ␤-arrestin-dependent signaling from native receptors without simultaneous G protein activation. The initial description of functional selectivity portrayed the ability of both AT1R mutants and selective ligands to activate ERK1/2 independently of G protein activation. This pioneering work showed that ERK1/2 can be activated by both G protein-dependent and ␤-arrestin-dependent pathways mediating distinct compartmentalization of active ERK1/2, followed by different cellular effects (Ahn et al. 2004, Aplin et al. 2007a, b, Gaborik et al. 2003, Holloway et al. 2002, Wei et al. 2003). Signaling through both G proteindependent and -independent mechanisms is becoming evident for an increasing number of 7TMRs. Ligands with different efficacy on disparate signaling pathways have been reported for a number of receptors, including AT1R, ␤1- and ␤2-adrenergic receptors, the serotonin receptor, the ␮-opioid receptor, the ␣2-adrenergic receptor, the V2 vasopressin receptor, the parathyroid hormone receptor, and dopamine receptors (Kendall and Luttrell 2009, Urban et al. 2007, Whalen et al. 2011). Because different cellular and physiological outcomes are associated with discrete signaling pathways arising from 7TMRs, functionally selective targeting may provide superior pharmacological profiles. • Pharmacological Description of Ligand Bias Our increased understanding of 7TMR signaling challenges the classical pharmacological term “efficacy,” which must be updated to specify the subset of receptor actions elicited by a given ligand. Such “multidimensional” efficacy and definition of ligand bias naturally necessitates consensus on what represents a 222

“balanced” response at a given receptor (Figure 1C). Furthermore, the ability of a receptor to assume a ligand-specific conformation and activate specific downstream pathways and actions may be heavily influenced by cellular settings such as expression of receptor interacting proteins (Kenakin 2009, Urban et al. 2007). To identify true pathway bias, it is essential to bear in mind that the receptor will activate individual pathways differently, and that signal transduction to two discrete pathways is generally not linear. Thus, a ligand may be interpreted as being biased because it displays different efficacies when evaluated with regard to two different measures of signaling activity (Figures 1B and 1C). Alternatively, bias may be dose dependent if a ligand elicits discrete cellular readouts with different potencies. Adequate pharmacological description of a ligand therefore necessitates an account of preference for various pathways. Methods for quantification of ligand bias have been proposed that involve the pathway-specific transduction constant, tau (␶), relevant to the particular cellular system in question (Black and Leff 1983, Evans et al. 2011, Kenakin 2009, McPherson et al. 2010, Rajagopal et al. 2011). Dynamic factors of receptor transduction may profoundly influence signaling quality, as illustrated by evident pathway interdependence. A ligand with limited G protein signaling will inevitably elicit less receptor phosphorylation, which would otherwise promote ␤-arrestin binding. Conversely, relative inability to recruit ␤-arrestins will naturally allow for prolonged G protein responses due to decreased desensitization. Therefore, in many cases, ligand bias reflects altered pathway balance in time or quantity, which leads to qualitative cellular differences. • The Angiotensin II Type I Receptor ARBs affect the heart both indirectly by lowering peripheral blood pressure and therefore the workload on the heart and directly by inhibition of hypertrophic growth and fibroblast infiltration (Ainscough et al. 2009). The blood pressurelowering effects of ARBs are associated with inhibition of G protein-mediated vasoconstrictory pathways (Harris et al. 2007, Violin et al. 2010). Cardiac cells such as myocytes and fibroblasts express

AT1Rs and respond to angiotensin II (Ang II) by hypertrophy or hyperplasia, respectively (Sadoshima and Izumo 1993). The effect of ARBs on the prevention of congestive heart failure has been shown to be better than expected based on their blood pressure-lowering effect alone, indicating that the direct effects on cardiac cells could be significant (Verdecchia et al. 2009). The negative aspect of the AT1Rdriven cardiac hypertrophic response has been associated with G␣q-dependent stimulation of cellular growth and fibrosis (Fan et al. 2005, Wettschureck et al. 2001). Transgenic mice overexpressing a constitutively active AT1R with decreased binding of ␤-arrestins develop increased cardiac fibrosis but not hypertrophy (Billet et al. 2007), whereas mice expressing a G␣q-uncoupled AT1R develop hypertrophy with decreased fibrosis (Zhai et al. 2005). Thus, major clinical benefits of ARB treatment are related to inhibition of G␣q-dependent signaling by the AT1R in peripheral vasculature and myocardium. In basic research, the AT1R has become a model receptor for the concept of functional selectivity. Selective receptor mutants and a strongly selective agonist have demonstrated the ability to separate G␣q and ␤-arrestin signaling in cellular model systems, organ preparations, and in vivo (Aplin et al. 2007a, b, Daniels et al. 2005, Holloway et al. 2002, Violin et al. 2010, Wei et al. 2003). These studies have associated ␤-arrestin signaling with cytoprotection through activation of anti-apoptotic and proliferative signaling molecules such as ERK1/2, RSK2, and AKT (Ahn et al. 2004, 2009, Aplin et al. 2007a) and with stimulation of migration through p38 (Hunton et al. 2005) (Figure 2). In addition, ␤-arrestin signaling has been shown to confer increased cardiomyocyte contraction (Boerrigter et al. 2011, Tilley et al. 2010, Violin et al. 2010). The pathway responsible for the ␤-arrestinmediated contractile response has not been identified, but ␤-arrestins are otherwise known to regulate activation of RhoA and following ROCK/MLCK signaling (Anthony et al. 2011, Godin and Ferguson 2010). Thus, it appears that ␤-arrestin signaling is able to stimulate adaptive growth and contractile capacity, possibly in part through activation of MNK-1 and EIF-4e in the translational system (DeWire et al. 2008), but inhibits TCM Vol. 20, No. 7, 2010

B

Classical perception of efficacy

Full agonist

80 60

Partial agonist

40 20

Neutral antagonist

0 -11

-10

-9

-8

-40

-7

-6

-5

Inverse agonist

30

B-arrestin recruitment

100

Ligand 1 Ligand 2 Ligand 3

80 60

G protein biased

B-arrestin biased

40 20 0

Depiction of ligand bias

C

G protein activation

120

100

-20

Multi-dimensional efficacy

S ig n alin g r e ad o u t (% of Max)

R e c e p t o r a c t iv it y

120

G p ro t e in s ig n a lin g (Ta u)

A

25

G protein biased

20 15 10

B-arrestin biased

5

B-arrestin biased -1 1

-1 0

-9

-8

[log ligand ]

-7

-6

0

-5

0

1

2

3

4

5

6

7

8

9

10

B -a rr e s tin s ig n a lin g (Ta u )

Figure 1. Pharmacological description of ligand efficacy. (A) The classical description of the efficacy of an agonist as determined measuring one receptor-mediated outcome (eg IP3, cAMP, or a physiological readout). The full agonist is defined as being able to reach the cellular maximum for the measured readout. (B) An agonist may show different efficacies for activation of distinct pathways (eg G protein- vs ␤-arrestin-dependent) and in this way possess functional selectivity. An agonist can be biased if it shows neutral agonism on one pathway and agonism on another pathway (green curve), if it is more potent in activation of one pathway versus another (red curve), or if it reaches maximum at one signaling readout and is a partial agonist of another (blue curve). (C) A quantitative measure of bias can only be estimated by taking into account the efficacy of an agonist for activation of each specific cellular readout. The efficacy can be expressed using the transducer constant tau (␶), which reflects the fraction of ligand-bound activated receptors required to reach half-maximum response of the cellular system.

induction of fibrosis. There is evidence that blockade of AT1R coupling to G␣q is clinically important, and there is increasing evidence that simultaneously permitting or stimulating ␤-arrestin-mediated effects of the same receptor could improve cardiac adaptation, although the role of the underlying signal pathways is still not fully understood. Although conventional ARBs are useful to dampen blood pressure and reduce cardiac remodeling and fibroblast infiltration, they may simultaneously inhibit ␤-arrestin-mediated myocyte contractile, cytoprotective, migratory, and regenerative potential, counteracting the physiologically stimulated healing and adaptation. New ␤-arrestin selective agonists at the AT1R may have equal efficacy as full ARBs in lowering blood pressure and ameliorating hypertrophy and fibrosis while simultaneously increasing cardiac output and conferring cytoprotection through alternative pathways. A pharmacological characterization of clinically applied ARBs showed no ␤-arrestin recruitment to the human AT1R, indicating that these are truly full blockers of both G protein activation and ␤-arrestin recruitment (Hansen et al. 2008). The first ␤-arrestin selective AT1R agonist, SII Ang II, was not suitable for in vivo studies due to a relatively low binding affinity. Novel ␤-arrestin selective ligands with higher affinities are currently being developed and tested for efficacy in animal models of heart failure. The first studies of such potential next-generation angiotensin receptor modulators are promising and report TCM Vol. 20, No. 7, 2010

AT1R Gαq

β-Arr

PLC-β

IP3

Desensitization of Gαq and internalization DAG

Ca2+

β-Arr MLCK

PKD

PKC ERK1/2

ERK1/2

p38

AKT

BAD

p Nucleus ELK-1

Hypertension Hypertrophy Contractility Cardiac remodeling

Cardioprotection Anti-apoptosis Regeneration

Figure 2. AT1R signaling pathways in cardiac myocytes. (Left) The AT1R couples primarily to G␣q and mediates activation of phospholipase C-␤ (PLC-␤). This in turn leads to accumulation of diacylglycerol (DAG) and inositol phosphates (IP3). Inositol phosphates bind to the ryanodine receptor on the sarcoplasmic reticulum, which facilitates release of calcium (Ca2⫹) to the cytoplasm. The increased Ca2⫹ concentration mediates activation of myosin light-chain kinase (MLCK) to facilitate myocyte contraction. In addition, Ca2⫹ and DAG in concert activate protein kinase C (PKC) and protein kinase D (PKD). Among other substrates, PKC promotes activation of the MAP kinase cascade, resulting in activation of ERK1/2. The activated ERK1/2 enters the nucleus and phosphorylates transcription factors such as ELK1, thus promoting transcription. Blocking ERK1/2 activation diminishes cardiac myocyte hypertrophy. Activation of PKD mediates phosphorylation of nuclear HDAC5 and initiation of the hypertropic gene program. Thus, overall, the activation of the G protein-dependent signaling pathways promotes both inotropy and unwanted cardiac effects such as hypertrophy and cardiac remodeling. (Right) Upon activation, AT1R is phosphorylated on its C-terminal tail, which increases its affinity for ␤-arrestin. ␤-Arrestin binds tightly to the receptor and facilitates its internalization through clatrin-coated pits. In addition to internalizing the receptor, ␤-arrestins function as scaffolds and create protein complexes for regulation of a second wave of signaling. Among the most well-described pathways known to be regulated ␤-arrestin-dependently are the MAP kinases (p38, ERK1/2, and JNK3). ERK1/2 activated by this pathway is restricted from entering the nucleus and therefore activates cytosolic substrates such as RSK2. RSK2 is involved in anti-apoptotic signaling through activation of the AKT/BAD pathway. The phenotypic consequence of restraining active ERK1/2 in the cytoplasm therefore tends to be protection against cardiotoxic or apoptotic stimuli. The functional differences between G protein-dependent and ␤-arrestin-dependent activation of PKD have not been uncovered.

223

enhanced cardiac performance – and simultaneously reduced peripheral vascular resistance and blood pressure (Boerrigter et al. 2011, Violin et al. 2010). This development holds promise for the use of ␤-arrestin selective ligands in both acute and chronic treatment of heart failure. Combined with the observed ability of ␤-arrestin-mediated AT1R signaling to confer cytoprotective effects and reduce fibrosis, these drugs may additionally reduce cardiotoxicity and remodeling. With the first compound currently in phase II clinical testing (by Trevena Inc.), it will be interesting to follow the development of ␤-arrestin selective ligands for the AT1R. • ␤-Adrenergic Receptors

␤-Blockers are central drugs in the treatment of cardiac remodeling and failure. Third-generation ␤-blockers with vasodilatory effects (␣-adrenergic receptor blocking or NO generating) are generally favored for afterload reduction in the treatment of heart failure. The effect of ␤-blockade in the heart is based on decreasing heart rate and contraction force, thereby lowering energy consumption and improving coronary supply in diastoly. In addition, inhibition of sympathetic drive may include a desirable shift in substrate utilization and attenuation of cardiomyocyte induction of the fetal gene program and apoptosis. The accompanying reduction in cardiac output seems counterintuitive and is often a challenge to titrate in patients with low output syndrome. Therefore, current pharmacological agents leave room for improvement that may be realized with novel concepts of pathway-selective receptor actions. The molecular mechanisms responsible for the effect of ␤-blockers in the treatment of heart failure are complex and involve multiple cardiac signaling mechanisms and a multifaceted interplay between the different ␤-adrenergic receptors. ␤1-AR is the dominant ␤-receptor expressed in the myocardium and is profoundly involved in the regulation of conduction and contractility through G␣s activation of adenylate cyclase (AC), production of cAMP, and calcium release. Upon prolonged catecholamine stimulation of the heart, ␤1-ARs become desensitized and uncouple from G␣s, thus decreasing cAMP production and associated contraction force (Lohse et 224

al. 1996). Therefore, prolonged ␤1-AR agonism was found incapable of increasing cardiac output. In addition to uncoupling of G␣s signaling, prolonged activation of ␤1-ARs leads to sustained activation of calmodulin-dependent protein kinase type II (CAMKII) entailing deleterious transcriptional regulation (Zhang et al. 2003). ␤1-AR activation of CAMKII can be mediated independently of PKA activation, raising the possibility that the inotropic and chronotropic AC/ cAMP/PKA pathway and the deleterious cAMP/EPAC/CAMKII pathway may be selectively activated (Figure 2). Activation of CAMKII is dependent on internalization of ␤1-AR and may occur in restricted microcompartments with high Ca2⫹ peaks (Saucerman and Bers 2008, Song et al. 2008). Interestingly, both CAMKII␦ and the cAMP binding protein EPAC have been shown to associate with ␤-arrestin, which is recruited to the tail of the activated ␤1-AR to create a specific microenvironment for activation of CAMKII (Mangmool et al. 2010). In the setting of congestive heart failure, negative inotropy and chronotrophy are often upsetting side effects of current ␤-blockers. Thus, a G␣s selective agonist at the cardiac ␤1-ARs might be desirable to preserve increased cardiac output and simultaneously inhibit the pathway responsible for CAMKII-mediated induction of the hypertrophic gene program. Although such a pharmacological profile does not match current use of ␤-blockers for acute reduction of energy consumption and control of heart rate, it may prove superior for the long-term prophylactic and prognostic indications associated with cardiac failure by counteracting cardiac apoptosis, remodeling, and hypertrophy. The ␤2-AR-mediated effects are also multifaceted because this receptor can signal through both G␣s and G␣i proteins. In contrast to ␤1-ARs, ␤2-ARs are not degraded upon prolonged catecholamine stimulation. Moreover, prolonged ␤2-AR stimulation promotes a switch from signaling through G␣s activation of AC to G␣i inhibition of AC (Daaka et al. 1997). In addition to inhibition of cAMP production, this shift to G␣i signaling promotes cytoprotective signaling through activation of ERK1/2, PI3K, and AKT pathways (Figure 3) (Daaka et al. 1997). Importantly, despite generation of

cAMP, ␤2-AR activation does not lead to EPAC binding to ␤-arrestin or activation of CAMKII (Mangmool et al. 2010, Wang et al. 2004, Yoo et al. 2009). Accordingly, signaling from the ␤2-AR may not be as deleterious to the heart as that arising from the ␤1-AR. Thus, the use of ␤1-AR-specific blockers for the treatment of heart failure seem to be a good choice. However, the nonspecific ␤-blocker carvedilol, which inhibits ␤1-AR, ␤2-AR, and ␣1-ARs, has been found to be superior to ␤1-AR-specific blockers with respect to adaptive remodeling and overall death (Metra et al. 2005, Poole-Wilson et al. 2003). The results obtained using carvedilol could not be explained by differences in ability to lower blood pressure. Other likely explanations are direct effects on cardiac cells in the form of cytoprotective effects conferred by anti-oxidative and antiapoptotic means. It is noteworthy that retrospective studies of its signaling properties show that carvedilol is not a full ␤2-AR/␤1-AR blocker but activates MAP kinases presumably through ␤-arrestin signaling; thus, it is a functionally selective agonist (Galandrin and Bouvier 2006, Kim et al. 2008, Wisler et al. 2007). Several studies have shown that ␤-arrestin signaling from other cardiac 7TMRs can be cardioprotective (Esposito et al. 2011, Noma et al. 2007). The cytoprotective effects of activating ␤-arrestin signaling have been linked to activation of ERK1/2 and AKT and transactivation of EGFR (Ahn et al. 2009, Esposito et al. 2011, Noma et al. 2007). Therefore, ␤-arrestin-dependent cytoprotective effects may in part explain the clinical superiority of carvedilol over other ␤-blockers in the treatment of heart failure. The ␤-blockers propranolol and alprenolol have a similar profile with regard to activation of MAP kinases, but the overall benefit of this property is difficult to determine because these drugs differ from carvedilol in that they do not inhibit ␣1-ARs, are less ␤1-AR selective, and are not generally recommended for treatment of heart failure (Frishman and Saunders, 2011, Gorre and Vandekerckhove 2010). Based on the opposing effects of individual signaling pathways from ␤-adrenergic receptors, the design of functionally selective ligands to target these important cardiac receptors holds potential to improve future treatment of TCM Vol. 20, No. 7, 2010

β2-AR

β1-AR

Gαs AC

β-Arr

Gαi

Desensitization of Gαs and internalization

Desensitization of Gαs and internalization

PI3K

EPAC

cAMP

cAMP

AKT

PKA

AKT

Recycling

PKA

Degradation

Hypertrophy Cardiotoxicity

β-Arr PI3K

CAMKII

Heart rate Contraction force Hypertrophy

β-Arr

AC

β-Arr cAMP

Gαs

Cardioprotection Anti-apoptosis

Heart rate Contraction force Hypertrophy

Cardioprotection Anti-apoptosis

Figure 3. Cardiac signaling pathways controlled by ␤-adrenergic receptors. Cardiac myocytes express both ␤1- and ␤2-adrenergic receptors, and the major signaling pathways and expected physiological endpoints controlled by each receptor are illustrated. (Left) The ␤1-adrenergic receptor couples primarily to G␣s following activation of AC, production of cAMP, and activation of protein kinase A (PKA). PKA and possibly downstream activation of calmodulin-dependent protein kinase type II (CAMKII) mediate cardiomyocyte contraction and increase heart rate. The ␤1-AR is internalized and thereby terminates further G␣s activation. The interaction with ␤-arrestin initiates a second wave of signaling and activates CAMKII through PKA-independent mechanisms. CAMKII activity is involved in the induction of the hypertrophic gene program. (Right) The ␤2-adrenergic receptor also initially signals through G␣s activation of AC, which leads to increased heart rate and cardiac contractility. In opposition, prolonged ␤2-AR signaling facilitates a shift from G␣s to G␣i signaling. G␣i signaling is associated with cardioprotective signaling through activation of PI3K and AKT. Moreover, ␤2-AR interacts differently with ␤-arrestin and does not promote maladaptive activation of CAMKII and is not degraded. Instead, ␤2-AR signaling through ␤-arrestin promotes the anti-apoptotic AKT signaling cascade.

cardiac diseases. Whereas ␤-arrestin signaling from ␤2-ARs appears to be advantageous for cardioprotective effects, analogous ␤-arrestin signaling from ␤1-AR is associated with cardiotoxic CAMKII signaling. We thus hypothesize that a compound capable of selectively blocking maladaptive ␤1-AR internalization and second-wave signaling could preserve G protein-dependent cardiac output while increasing cardioprotective signaling. Studies are needed to determine, for example, if the use of G protein biased ligands for ␤1-AR will help improve the discouraging statistics of heart failure. • Development of Future Functionally Selective Drugs With Add-On Benefits To realize the pharmacological potential of functional selectivity, several aspects of the phenomenon need further attention. First, in-depth understanding of the molecular mechanisms that allow ligand-specific receptor conformations with preference for individual signaling pathways may direct medicinal chemists in designing new compounds (Aplin et TCM Vol. 20, No. 7, 2010

al. 2009). Second, functionally selective agonists must separate pathways that mediate beneficial from adverse biological effects. To accomplish the ambitious goal of developing superior receptor modulators, the major challenge is to define which pathways to target and which to preserve for the individual receptor and disease. Individual studies of mechanisms and the biological importance of pathways that can be pharmacologically separated provide pieces to the complex puzzle. Simultaneously, more comprehensive understanding of signaling networks influenced by 7TMRs is important to direct future drug development. The complexity and nonlinearity of these networks have been highlighted by several large-scale mass spectrometry and antibody array studies of AT1R signaling comparing the “full” agonist Ang II and the functional selective agonist SII Ang II (Christensen et al. 2010, Xiao et al. 2010). Such studies provide comprehensive signalosome fingerprints of the individual agonist and allow global comparisons between ligand-induced effects downstream of a given receptor. Unraveling the biological outcomes of the differential signaling

observed in these signalosomes is a major task, but will hopefully be rewarding in the form of identifying new strategies for pharmacological treatment of various diseases in the cardiovascular system. Similar signalosomes for ␤-ARs are warranted to gain an overview of their complex signaling and guide development of superior drugs. The new line of thinking presented in this review is exemplified by the promising pharmacological profile of the ␤-arrestin selective AT1R ligand TRV120027, which combines amelioration of hypertension and increasing cardiac output (Boerrigter et al. 2011, Violin et al. 2010). As appreciation is mounting that receptors are not biological on/off switches but, rather, should be perceived as controllers of biological finetuning, so is confidence that functional selectivity can lead to improved treatment of cardiovascular diseases. • Acknowledgments The research projects that inspired this review were funded by the Danish Council for Independent Research | Medical Sciences, the Danish National Research 225

Foundation, the Købmand i Odense Johan og Hanne Weimann f. Seedorffs legat, and the Novo Nordisk Foundation.

References Ahn S, Kim J, Hara MR, et al: 2009. ␤-Arrestin-2 mediates anti-apoptotic signaling through regulation of BAD phosphorylation. J Biol Chem 284:8855– 8865. Ahn S, Shenoy SK, Wei H, & Lefkowitz RJ: 2004. Differential kinetic and spatial patterns of beta-arrestin and G proteinmediated ERK activation by the angiotensin II receptor. J Biol Chem 279:35518 – 35525. Ainscough JF, Drinkhill MJ, Sedo A, et al: 2009. Angiotensin II type-1 receptor activation in the adult heart causes blood pressure-independent hypertrophy and cardiac dysfunction. Cardiovasc Res 81:592– 600. Allen JA & Roth BL: 2011. Strategies to discover unexpected targets for drugs active at G protein-coupled receptors. Annu Rev Pharmacol Toxicol 51:117–144. Anthony DF, Sin YY, Vadrevu S, et al: 2011. ␤-Arrestin 1 inhibits the GTPase-activating protein function of ARHGAP21, promoting activation of RhoA following angiotensin II type 1A receptor stimulation. Mol Cell Biol 31:1066 –1075. Aplin M, Bonde MM, & Hansen JL: 2009. Molecular determinants of angiotensin II type 1 receptor functional selectivity. J Mol Cell Cardiol 46:15–24. Aplin M, Christensen GL, Schneider M, et al: 2007a. The angiotensin type 1 receptor activates extracellular signal-regulated kinases 1 and 2 by G protein-dependent and -independent pathways in cardiac myocytes and Langendorff-perfused hearts. Basic Clin Pharmacol Toxicol 100:289 –295. Aplin M, Christensen GL, Schneider M, et al: 2007b. Differential extracellular signal-regulated kinases 1 and 2 activation by the angiotensin type 1 receptor supports distinct phenotypes of cardiac myocytes. Basic Clin Pharmacol Toxicol 100:296 –301. Askoxylakis V, Thieke C, Pleger ST, et al: 2010. Long-term survival of cancer patients compared to heart failure and stroke: A systematic review. BMC Cancer 10:105. Billet S, Bardin S, Verp S, et al: 2007. Gainof-function mutant of angiotensin II receptor, type 1A, causes hypertension and cardiovascular fibrosis in mice. J Clin Invest 117:1914 –1925. Black JW & Leff P: 1983. Operational models of pharmacological agonism. Proc R Soc London B Biol Sci 220:141–162. Boerrigter G, Lark MW, Whalen EJ, et al: 2011. Cardiorenal actions of TRV120027, a novel, ␤-arrestin-biased ligand at the angiotensin II type I receptor, in healthy and heart failure canines: A novel therapeutic

226

strategy for acute heart failure. Circ Heart Fail 4:770 –778. Christensen GL, Kelstrup CD, Lyngso C, et al: 2010. Quantitative phosphoproteomics dissection of seven-transmembrane receptor signaling using full and biased agonists. Mol Cell Proteomics 9:1540 –1553. Daaka Y, Luttrell LM, & Lefkowitz RJ: 1997. Switching of the coupling of the beta2adrenergic receptor to different G proteins by protein kinase A. Nature 390:88 –91. Daniels D, Yee DK, Faulconbridge LF, & Fluharty SJ: 2005. Divergent behavioral roles of angiotensin receptor intracellular signaling cascades. Endocrinology 146: 5552–5560. DeWire SM, Kim J, Whalen EJ, et al: 2008. Beta-arrestin-mediated signaling regulates protein synthesis. J Biol Chem 283: 10611–10620. Esposito G, Perrino C, Cannavo A, et al: 2011. EGFR trans-activation by urotensin II receptor is mediated by beta-arrestin recruitment and confers cardioprotection in pressure overload-induced cardiac hypertrophy. Basic Res Cardiol 106:577–589. Evans BA, Broxton N, Merlin J, et al: 2011. Quantification of functional selectivity at the human alpha(1A)-adrenoceptor. Mol Pharmacol 79:298 –307. Fan G, Jiang YP, Lu Z, et al: 2005. A transgenic mouse model of heart failure using inducible G␣q. J Biol Chem 280:40337– 40346. Frishman WH & Saunders E: 2011. ␤-Adrenergic blockers. J Clin Hypertens 13:649 – 653. Gaborik Z, Jagadeesh G, Zhang M, et al: 2003. The role of a conserved region of the second intracellular loop in AT1 angiotensin receptor activation and signaling. Endocrinology 144:2220 –2228. Galandrin S & Bouvier M: 2006. Distinct signaling profiles of beta1 and beta2 adrenergic receptor ligands toward adenylyl cyclase and mitogen-activated protein kinase reveals the pluridimensionality of efficacy. Mol Pharmacol 70:1575–1584. Godin CM & Ferguson SS: 2010. The angiotensin II type 1 receptor induces membrane blebbing by coupling to Rho A, Rho kinase, and myosin light chain kinase. Mol Pharmacol 77:903–911. Gorre F & Vandekerckhove H: 2010. Betablockers: Focus on mechanism of action. Which beta-blocker, when and why? Acta Cardiol 65:565–570. Hansen JL, Aplin M, Hansen JT, et al: 2008. The human angiotensin AT(1) receptor supports G protein-independent extracellular signal-regulated kinase 1/2 activation and cellular proliferation. Eur J Pharmacol 590:255–263. Harris DM, Cohn HI, Pesant S, et al: 2007. Vascular smooth muscle G(q) signaling is involved in high blood pressure in both

induced renal and genetic vascular smooth muscle-derived models of hypertension. Am J Physiol Heart Circ Physiol 293:H3072–H3079. Holloway AC, Qian H, Pipolo L, et al: 2002. Side-chain substitutions within angiotensin II reveal different requirements for signaling, internalization, and phosphorylation of type 1A angiotensin receptors. Mol Pharmacol 61:768 –777. Hunton DL, Barnes WG, Kim J, et al: 2005. Beta-arrestin 2-dependent angiotensin II type 1A receptor-mediated pathway of chemotaxis. Mol Pharmacol 67:1229 –1236. Jessup M, Abraham WT, Casey DE, et al: 2009. 2009 focused update: ACCF/AHA guidelines for the diagnosis and management of heart failure in adults: A report of the American College of Cardiology Foundation/American Heart Association Task Force on Practice Guidelines: Developed in collaboration with the International Society for Heart and Lung Transplantation. Circulation 119:1977–2016. Kenakin TP: 2009. ’7TM receptor allostery: Putting numbers to shapeshifting proteins. Trends Pharmacol Sci 30:460 – 469. Kendall RT & Luttrell LM: 2009. Diversity in arrestin function. Cell Mol Life Sci 66: 2953–2973. Kim IM, Tilley DG, Chen J, et al: 2008. Betablockers alprenolol and carvedilol stimulate beta-arrestin-mediated EGFR transactivation. Proc Natl Acad Sci USA 105: 14555–14560. Lohse MJ, Engelhardt S, Danner S, & Bohm M: 1996. Mechanisms of beta-adrenergic receptor desensitization: From molecular biology to heart failure. Basic Res Cardiol 91(Suppl 2):29 –34. Mangmool S, Shukla AK, & Rockman HA: 2010. ␤-Arrestin-dependent activation of Ca(2⫹)/calmodulin kinase II after ␤(1)-adrenergic receptor stimulation. J Cell Biol 189:573–587. McPherson J, Rivero G, Baptist M, et al: 2010. ␮-Opioid receptors: Correlation of agonist efficacy for signalling with ability to activate internalization. Mol Pharmacol 78: 756 –766. Metra M, Torp-Pedersen C, Swedberg K, et al: 2005. Influence of heart rate, blood pressure, and beta-blocker dose on outcome and the differences in outcome between carvedilol and metoprolol tartrate in patients with chronic heart failure: Results from the COMET trial. Eur Heart J 26: 2259 –2268. Noma T, Lemaire A, Naga Prasad SV, et al: 2007. ␤-Arrestin-mediated ␤1-adrenergic receptor transactivation of the EGFR confers cardioprotection. J Clin Invest 117: 2445–2458. Poole-Wilson PA, Swedberg K, Cleland JG, et al: 2003. Comparison of carvedilol and metoprolol on clinical outcomes in patients with

TCM Vol. 20, No. 7, 2010

chronic heart failure in the Carvedilol or Metoprolol European Trial (COMET): Randomised controlled trial. Lancet 362:7–13. Rajagopal S, Ahn S, Rominger DH, et al: 2011. Quantifying ligand bias at seventransmembrane receptors. Mol Pharmacol 80:367–377. Sadoshima J & Izumo S: 1993. Molecular characterization of angiotensin II-induced hypertrophy of cardiac myocytes and hyperplasia of cardiac fibroblasts: Critical role of the AT1 receptor subtype. Circ Res 73:413– 423. Saucerman JJ & Bers DM: 2008. Calmodulin mediates differential sensitivity of CaMKII and calcineurin to local Ca2⫹ in cardiac myocytes. Biophys J 95:4597– 4612. Seta K, Nanamori M, Modrall JG, et al: 2002. AT1 receptor mutant lacking heterotrimeric G protein coupling activates the SrcRas-ERK pathway without nuclear translocation of ERKs. J Biol Chem 277:9268 – 9277. Song Q, Saucerman JJ, Bossuyt J, & Bers DM: 2008. Differential integration of Ca2⫹-calmodulin signal in intact ventricular myocytes at low and high affinity Ca2⫹-calmodulin targets. J Biol Chem 283:31531–31540. Stewart S, MacIntyre K, Hole DJ, et al: 2001. More “malignant” than cancer? Five-year survival following a first admission for heart failure. Eur J Heart Fail 3:315–322.

TCM Vol. 20, No. 7, 2010

Tilley DG, Nguyen AD, & Rockman HA: 2010. Troglitazone stimulates beta-arrestin-dependent cardiomyocyte contractility via the angiotensin II type 1A receptor. Biochem Biophys Res Commun 396:921–926. Urban JD, Clarke WP, von Zastrow M, et al: 2007. Functional selectivity and classical concepts of quantitative pharmacology. J Pharmacol Exp Ther 320:1–13. Verdecchia P, Angeli F, Cavallini C, et al: 2009. Blood pressure reduction and reninangiotensin system inhibition for prevention of congestive heart failure: A metaanalysis. Eur Heart J 30:679 – 688. Violin JD, DeWire SM, Yamashita D, et al: 2010. Selectively engaging beta-arrestins at the angiotensin II type 1 receptor reduces blood pressure and increases cardiac performance. J Pharmacol Exp Ther 335:572– 579. Wang W, Zhu W, Wang S, et al: 2004. Sustained ␤1-adrenergic stimulation modulates cardiac contractility by Ca2⫹/calmodulin kinase signaling pathway. Circ Res 95:798 – 806. Wei H, Ahn S, Shenoy SK, et al: 2003. Independent ␤-arrestin 2 and G protein-mediated pathways for angiotensin II activation of extracellular signal-regulated kinases 1 and 2. Proc Natl Acad Sci USA 100:10782– 10787. Wettschureck N, Rutten H, Zywietz A, et al: 2001. Absence of pressure overload induced myocardial hypertrophy after condi-

tional inactivation of G␣q/G␣11 in cardiomyocytes. Nat Med 7:1236 –1240. Whalen EJ, Rajagopal S, & Lefkowitz RJ: 2011. Therapeutic potential of ␤-arrestinand G protein-biased agonists. Trends Mol Med 17:126 –139. Wisler JW, DeWire SM, Whalen EJ, et al: 2007. A unique mechanism of beta-blocker action: Carvedilol stimulates beta-arrestin signaling. Proc Natl Acad Sci USA 104: 16657–16662. Xiao K, Sun J, Kim J, et al: 2010. Global phosphorylation analysis of beta-arrestinmediated signaling downstream of a seven transmembrane receptor (7TMR). Proc Natl Acad Sci USA 107:15299 –15304. Yoo B, Lemaire A, Mangmool S, et al: 2009. ␤1-Adrenergic receptors stimulate cardiac contractility and CaMKII activation in vivo and enhance cardiac dysfunction following myocardial infarction. Am J Physiol Heart Circ Physiol 297:H1377–H1386. Zhai P, Yamamoto M, Galeotti J, et al: 2005. Cardiac-specific overexpression of AT1 receptor mutant lacking G␣q/G␣i coupling causes hypertrophy and bradycardia in transgenic mice. J Clin Invest 115:3045– 3056. Zhang T, Maier LS, Dalton ND, et al: 2003. The deltaC isoform of CaMKII is activated in cardiac hypertrophy and induces dilated cardiomyopathy and heart failure. Circ Res 92:912–919. PII S1050-1738(11)00084-3

TCM

227