Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144

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Pulmonary Pharmacology & Therapeutics

journal homepage: www.elsevier.com/locate/ypupt

Emerging airway smooth muscle targets to treat asthma

Sana Siddiqui a,1, Naresh Singh Redhu b, Oluwaseun O. Ojo c, Bo Liu d, Nneka Irechukwu e,
Charlotte Billington d, Luke Janssen f, Lyn M. Moir g, h, *
a
Meakins-Christie Laboratories, Department of Medicine, McGill University, 3626 St Urbain, Montréal, Québec H2X 2P2, Canada
b
Department of Immunology, Faculty of Medicine, University of Manitoba, Winnipeg, MB R3E0T5, Canada
c
Department of Respiratory Physiology, University of Manitoba, Biology of Breathing Group, Manitoba Institute of Child Health, 745 Bannatyne Avenue, Winnipeg R3E 3P4, Canada
d
Division of Therapeutics and Molecular Medicine, The University of Nottingham, D Floor, South Block, Queen’s Medical Centre, Nottingham NG7 2UH, UK
e
Department of Asthma and Allergy, Kings College London, 5th Floor Tower Wing, Guy’s Hospital, London SE1 9RT, UK
f
Medicine, McMaster University, Firestone Institute for Respiratory Health, St. Joseph’s Hospital, Hamilton, Ontario L8N 4A6, Canada
g
Cell Biology, Woolcock Institute of Medical Research, Glebe, NSW 2037, Australia
h
Respiratory Research Group, Discipline of Pharmacology, The University of Sydney, Sydney, NSW 2006, Australia

a r t i c l e i n f o a b s t r a c t

Article history: Asthma is characterized in part by variable airflow obstruction and non-specific hyperresponsiveness to
Received 22 March 2012 a variety of bronchoconstrictors, both of which are mediated by the airway smooth muscle (ASM). The
Received in revised form ASM is also involved in the airway inflammation and airway wall remodeling observed in asthma. For all
28 July 2012
these reasons, the ASM provides an important target for the treatment of asthma. Several classes of drugs
Accepted 27 August 2012
were developed decades ago which targeted the ASM e including b-agonists, anti-cholinergics, anti-
histamines and anti-leukotrienes e but no substantially new class of drug has appeared recently. In this
Keywords:
review, we summarize the on-going work of several laboratories aimed at producing novel targets and/or
Asthma
Contraction
tools for the treatment of asthma. These range from receptors and ion channels on the ASM plasma-
Airway hyperreactivity lemma, to intracellular effectors (particularly those related to cyclic nucleotide signaling, calcium-
Kinases homeostasis and phosphorylation cascades), to anti-IgE therapy and outright destruction of the ASM
Calcium itself.
GPCRs Ó 2012 Elsevier Ltd. All rights reserved.
IgE
Bronchial thermoplasty

1. Introduction functions as an immunomodulatory cell and contributes to the

Asthma is a common respiratory disease that affects approxi-
mately 235 million people worldwide (WHO, Fact Sheet No 307
May 2011). It is characterized by airflow obstruction, airway
inflammation and airway remodeling. Classically, the airway
smooth muscle (ASM) cell was believed to contribute to the path-
ogenesis of asthma through its contractile properties: airway
hyperresponsiveness (AHR), one of the main characteristics in
asthma, refers to excessive narrowing of ASM induced by stimuli
which in otherwise normal individuals cause only limited airway
narrowing. However it is now widely accepted that ASM also
* Corresponding author. Cell Biology, Woolcock Institute of Medical Research, PO
Box M77, Missenden Road, Sydney, NSW 2050, Australia. Tel.: þ61 2 9114 0372;
fax: þ61 2 9114 0399.
E-mail address: lyn.moir@sydney.edu.au (L.M. Moir).
1
Sana Siddiqui is now at the Receptor Pharmacology Unit, National Institute on
Aging, National Institutes of Health, Baltimore, MD 21224, United States of America.
airway inflammation and structural alterations associated with the
disease. Thus, an increase in ASM bulk not only exacerbates airway
contraction resulting in increased airway narrowing but it may also
be a major driving force of disease progression. Since the ASM plays
such a central role it is reasonable to assume that targeting it may
be beneficial for the treatment of asthma.
Although current asthma therapies, namely glucocorticoids and
b2-adrenoceptor (b2AR) agonists, which abrogate airway inflam-
mation, reverse bronchoconstriction and improve quality of life,
are effective in controlling disease symptoms in the majority
of patients, a considerable population of patients with poorly
controlled asthma remains. Furthermore, these life-long therapies
only treat the symptoms and they have little or no effect on the
structural alterations associated with asthma. Taken together, these
points highlight the need for the development of new or improved
therapies. It is only with a greater understanding of the cellular and
molecular mechanisms that regulate ASM function that we can
begin to develop new treatment strategies for asthma.

1094-5539/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.pupt.2012.08.008
 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144 133

Abbreviations NFkB nuclear factor kappa B

Non-RTK non-receptor tyrosine kinase
AHR airway hyperresponsiveness PDE phosphodiesterase
ASM airway smooth muscle PDGFR platelet-derived growth factor receptor
b2AR b2-adrenoceptor PI3K phosphatidylinositol 3-kinase
BTR bitter taste receptor PKC protein kinase C
Ca2þ calcium PKG protein kinase G (cGMP-dependent protein kinase)
[Ca2þ]i intracellular calcium PM plasma membrane
cAMP cyclic 30 50 -adenosine monophosphate PMCA plasma membrane Ca2þ ATPase
CCL chemokine (CeC motif) ligand PTEN phosphatase and tensin homolog deleted on
cGMP cyclic 30 50 -guanosine monophosphate chromosome ten
COPD chronic obstructive pulmonary disease ROC receptor-operated channels
CPA cyclopiazonic acid ROCK Rho associated coiled coil-forming protein kinase
CTGF connective tissue growth factor RTK receptor tyrosine kinase
CXCL chemokine (CeXeC motif) ligand SERCA sarcoplasmic reticulum calcium-ATPase
DAG diacylglycerol SH2 Src homology domain 2
EGFR epidermal growth factor receptor SOCC store-operated calcium channels
ER endoplasmic reticulum SR sarcoplasmic reticulum
FPP farnesyl pyrophosphate STIM-1 stromal interaction molecule 1
GGPP geranylgeranyl pyrophosphate TGFb transforming growth factor beta
GPCR G protein-coupled receptors TNF tumor necrosis factor
HMG hydroxy-methylglutaryl TH2 T helper type 2
IL interleukin TRP transient receptor potential
JAK janus kinase TSLP thymic stromal lymphopoietin
MAPK mitogen-activated protein kinase VDCC voltage-dependent calcium channels
MMP matrix metalloproteinase VEGFR vascular endothelial growth factor receptor
NCX sodiumecalcium exchanger

In this review, we provide an update on existing ASM targets and growing appreciation of 3 important concepts: (1) GPCR desensi-
highlight new and emerging concepts for the treatment of asthma. tization, (2) constitutively active receptors and (3) biased agonism.
Space limitations do not allow us to be all inclusive or comprehen-
sive on this subject; instead, we focus particularly on the new
2.1. GPCR desensitization
directions being pursued by the attendees of the meeting. We begin
with stimulation of the ASM by its plasmalemmal receptors for
How GPCR signaling can be manipulated in human ASM to
external stimuli e the G protein-coupled receptors, or GPCRs e and
promote a pro-relaxant, anti-inflammatory, anti-proliferative
several pharmacological concepts related to their function (desen-
phenotype has long-been the subject of intensive research (see
sitization; constitutive activity; biased agonism). In addition,
reviews [1,2]). Pharmacological strategies to address this goal have
we briefly summarize one novel class of GPCR e the bitter taste
focused on the development of b2AR-selective ligands with opti-
receptor e given the considerable attention which this family has
mized efficacy and pharmacokinetic properties. b2AR signaling and
recently received in the airway field. Next, we consider several
functional efficacy is limited by desensitization mechanisms
diverse signaling events triggered by those GPCRs, including the
invoked upon exposure to b-agonist. Interestingly, data from Penn,
cyclic nucleotide/phosphodiesterase cascade, elevation of cytosolic
Walker, and colleagues reveals that directly targeting b2AR
calcium concentration via 4 different Ca2þ-influx pathways, and
desensitization mechanisms (via either GRK2/3 inhibition or
activation of various kinases. Finally, we move away from these
b-arrestin-2 inhibition/knockout) selectively enhances b2AR
strategies of treating asthma through control of an excited ASM, by
signaling in human ASM cells to and augments the relaxant effect of
instead now either controlling airway inflammation (anti-IgE
b-agonists on ASM [3,4]. Inhaled corticosteroids in combination
therapy), or by removing the ASM entirely (bronchial thermoplasty).
with b-agonists constitute the mainstay asthma therapy for many
patients and it is interesting to note that preincubation with
steroids both restores b-agonist-induced sensitivity and prevents
2. G protein-coupled receptors
b2-AR desensitization in human precision-cut lung slices [5]. In
a follow up study, Panettieri and colleagues revealed that the
G protein-coupled receptors (GPCR) on human ASM cells have
protective effect of the steroid was not observed for all b-agonists
been a major target for asthma therapy for decades and GPCR
studied and budesonide was found to prevent formoterol- but not
ligands still constitute the frontline treatment of asthma today. This
salmeterol-induced desensitization of human small airways [6].
widespread clinical use of GPCR ligands is a result of the expanded
body of knowledge regarding GPCR identity, localization, down-
stream signaling pathways, ligand specificity and duration as well 2.2. Constitutively active receptors
as improved methods of administration.
In addition to finding new GPCR targets, the current challenge is It is now appreciated that many GPCRs do not sit inactive
to advance the study of GPCR regulation and signaling in human awaiting stimulation but instead possess a measure of constitutive
ASM to create new ligands, modify existing ones or target modu- activity. Thus, in contrast to long-held assumptions, GPCRs are not
lators of GPCR function. These approaches are shaped by our necessarily on/off switches, but may be active in the unliganded
134 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144
state and tunable in different directions. With this in mind, Bond
and colleagues invite us to flip our idea of the type of b2AR ligand
that is useful in asthma therapy from the classically used b2AR-
agonists to b2AR inverse agonists (i.e., ligands capable of inhibiting
constitutive b2AR activity) [7]. The rationale for this arises from
reported adverse clinical effects associated with chronic use of
b2AR agonists (reviewed by [8]) and mirrors the shift in treatment,
from bAR agonists to b-blockers, in congestive heart failure (dis-
cussed in [9]). Data from Bond and colleagues shows that chronic
exposure to b2AR inverse agonist in mice induces a series of
anti-inflammatory endpoints, including reduced methacholine-
induced bronchoconstriction, reduced eosinophilia and also
increased density of b2ARs [10,11]. Interestingly they have also
recently reported that steroids complement the anti-inflammatory
indices promoted by b2AR inverse agonist exposure [10e14].
2.3. Biased agonism
Biased agonism encompasses the observations that (i) GPCRs
signal to more than one pathway and (ii) different ligands, or even
stereoisomers of the same ligand, may selectively (or with bias)
promote activation of one pathway over another (see review in
[15]). The goal here is clearly to select or produce a ligand capable of
inducing all of the favorable signaling events whilst bypassing
those which are potentially problematic.
In addition to the novel considerations discussed above which
are pertinent to all GPCR research, it is important to highlight new
perspectives regarding well-studied GPCRs as well as discuss the
new GPCRs as targets for asthma therapy.
Anti-cholinergics were used by the ancient Egyptians to alle-
viate symptoms of asthma and their ability to relieve broncho-
constriction via inhibition of muscarinic M3 receptors has been
established for decades. However, recent data from Meurs and
colleagues have revealed novel facets to this GPCR. The anti-
cholinergic drug tiotropium was observed to inhibit eosinophilic
and neutrophilic airway inflammation and indices of airway
remodeling in animal models of chronic allergic asthma and COPD
[16]. Interestingly it is also able to synergistically enhance the
bronchoprotective effect of the novel long-acting b2-agonist olo-
daterol in guinea-pigs.
In a similar vein, Satoru and colleagues have teased out previ-
ously unrealized aspects of PGE2 signaling in human ASM cells [17].
This work revealed that the inhibition of serum-induced human
ASM proliferation occurs predominantly via EP2 and EP4 receptor
activation and subsequent cAMP elevation. However, signaling via
the EP3 receptor increased [Ca2þ]i without affecting proliferation.
There is no evidence to date of functional EP1 receptor expression in
human ASM. Further analysis of the balance of signaling induced by
EP receptors will ascertain the likely effectiveness of targeting
specific isoforms of this GPCR family.
Sections 2.1e2.3 are summarized in Fig. 1.
2.4. Bitter taste receptors
Bitter taste receptors (BTR) have recently become the focus of
much discussion in the respiratory community. This family of
GPCRs comprises 25 members, 6 of which predominate on human
ASM (types 3, 4, 5, 10, 14 and 31) [18]. More importantly, BTR-
agonists have been found to produce bronchodilation superior to
that of the b-agonists [18]. This dilatory effect appears not to
involve the cyclic nucleotide signaling pathway (which is central to
b-agonist mediated responses). Instead, they purportedly act via
IP3-mediated release of Ca2þ from the SR, Kþ-channel activation
and membrane hyperpolarization. This is quite perplexing, since
release of Ca2þ is usually associated with contraction, and
hyperpolarization alone is not normally a powerful bronchodilatory
signal [19e21]. In contrast, that same study found that histamine
also elevated [Ca2þ]i in the same cells, but this resulted in
membrane depolarization and contraction [18]. The concentrations
of BTR-agonists needed to evoke these responses are relatively high
e in the high micromolar or even the millimolar range e which
raises several problems with respect to translation of these findings
to the clinical setting. For example, in addition to the unpleasant
sensation which inhaled BTR-agonists are sure to evoke, there is the
potential for the BTRs to undergo desensitization: a follow-up study
has indeed documented this to occur readily [22]. Also, another
more recent study produced evidence that some of the relaxant
action evoked by millimolar BTR-agonists may be due in part to
a cytotoxic effect [23]; it is worth pointing out that they made other
observations in their human second order bronchial preparations
which differed substantially from some of the key findings of the
original study which used murine fourth order bronchi [18].
Although it is unrealistic to propose applying or delivering such
high concentrations to the airways, a better understanding of how
they exert their inhibitory effect(s) may set the stage for a more
potent and clinically safe therapeutic tool.
3. Phosphodiesterases and cAMP
Phosphodiesterase (PDE) inhibitors have gained widespread
interest for their potential to treat a variety of diseases including
asthma and chronic obstructive pulmonary disease (COPD). Physi-
ological and pharmacological studies have highlighted the impor-
tant role of PDEs in the control of airway function, inflammation
and remodeling. Theophylline, a non-specific PDE inhibitor, has
been used in the treatment of asthma since the 1930s, as phar-
macological inhibition of PDEs augments the second messenger
cyclic adenosine monophosphate (cAMP) and promotes broncho-
dilation. However, although the use of theophylline as a treatment
for asthma has been declining, at least in part, due to its adverse
side-effects, a renewed interest in PDEs as pharmacological targets
has arisen due to more recent advancements in our understanding
of the PDE isoenzymes.
PDEs are a superfamily of enzymes ubiquitously expressed in
mammalian tissues that are essential for the control of numerous
physiological processes. They consist of 11 subfamilies (PDE1-11),
each with 1e4 gene products, and are characterized by their
structure, tissue distribution, substrate specificity and inhibitor
selectivity [24]. PDEs hydrolyze the second messenger cyclic
nucleotides cAMP and cyclic guanosine monophosphate (cGMP)
into inactive 50 AMP and 50 GMP respectively, thus terminating their
downstream signaling. PDEs-4, -7 and -8 are specific for cAMP,
whereas PDEs-5, -6 and -9 are cGMP-specific and PDEs-1, -2, -3, -10
and -11 can hydrolyze both cAMP and cGMP. Since PDEs regulate
the breakdown of cAMP and cGMP, inhibition of PDEs results in
elevation of intracellular cAMP and cGMP levels and thus regulates
ASM relaxation, proliferation and immunomodulatory functions
[25e27]. Recently, Trian and colleagues reported that ASM cells
derived from asthmatic subjects express decreased cAMP levels
and elevated PDE activity when compared with non-asthmatic-
derived ASM cells, suggesting dysregulation of the cAMP-PDE
pathway in disease [27].
ASM is the main cell in the regulation of bronchomotor tone and
inhibition of the specific isoenzymes PDE-3 and PDE-4 cause’s
relaxation in rodent and human airways. Pharmacological inhibi-
tion of PDE4 (using roflumilast) decreased ovalbumin-induced
contraction in guinea-pig trachea as well as large and small
airway contraction in ventilated rats and guinea-pigs whereas
inhibition of PDE-3 and -4 reduced inherent tone as well as allergen
and leukotriene C4-induced contraction in human airways [28e30].
 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144
ASM cells express a range of PDE isoforms, including PDEs-1, -2, -3,
-4 and -5; however, the majority of the cAMP-hydrolyzing activity
is attributed to the PDE-4 family [26,29,31,32]. PDE-4 is encoded by
four genes (4AeD) that generate multiple splice variants through
the activation of different promoters and alternative splicing. The
dominant PDE4 in human ASM cells is PDE4D [33], and this has
been implicated as an asthma susceptibility gene [34]. Further-
more, treatment of ASM cells with b2AR agonists induces PDE4D
mRNA and PDE4D5 has been identified in the control of b2AR-
induced cAMP signaling in human ASM cells from asthmatic and
non-asthmatic subjects [31,32].
In addition to their role in airway tone, PDEs are also important
in the regulation of the structural alterations associated with
asthma. Airway hyperplasia and altered extracellular matrix (ECM)
deposition contribute to the airway remodeling associated with
asthma, and proliferation of ASM cells from asthmatic subjects is
enhanced compared with non-asthma-derived cells [35,36].
Treatment with the PDE4 inhibitors cilomilast or roflumilast
inhibits ASM cell proliferation, thus suggesting that increasing
cAMP may abrogate cell growth [27]. In addition, treatment of ASM
cells with cAMP-mobilizing agents, such as PGE2, or the PDE4
inhibitor cilomilast inhibits migration [37]. Moreover, inhibition of
PDE4 also modulates ECM deposition, for example roflumilast
reduces transforming growth factor-b (TGFb)-induced fibronectin
secretion from ASM cells and TGFb-induced connective tissue
growth factor (CTGF), collagen I and fibronectin in bronchial ring
sections [38]. Furthermore, PDEs have also been implicated in the
regulation of inflammatory mediators from ASM cells for example,
rolipram and cilomilast attenuate TNFa-induced IL-8 and eotaxin
release; however, the PDE4 inhibitor roflumilast had no effect on
TGFb-induced IL-6 secretion from ASM cells [25,38].
The PDE4 inhibitor roflumilast is approved for the treatment of
COPD, where it has been shown to prevent exacerbations and
improve lung function [39e41]. Although preclinical and clinical
studies have shown some promising findings for the use of PDE4
inhibitors in asthma, such as inhibition of the late phase response,
improvements in FEV1 and reduction of inflammatory cell
numbers, they are not currently approved for the treatment of
asthma [42e46]. However, although corticosteroids are the main
treatment for the majority of asthmatic patients, there is a subset of
patients who have poorly controlled asthma despite the use of
current treatments and therefore, PDE4 inhibitors, either alone or
in combination with existing therapies may be beneficial in treating
these patients. In a recent in vitro study by Tannheimer et al. (2012)
the combination of roflumilast with a long acting beta agonist
(indacaterol) reduced inflammatory and fibrotic mediators associ-
ated with remodeling to a greater extent than either drug alone
[47], thus supporting the development of novel combination ther-
apies for asthma. However, despite the potential therapeutic use of
PDE inhibitors, there are currently clinical limitations due to
unwanted side effects, including nausea, headaches and diarrhea,
Fig. 1. Schematic representation of G-protein coupled receptor (GPCR) desensitization, constitutive activity and biased agonism.
135
of oral administration of these drugs. Therefore, strategies which
improve the risk to benefit ratio of PDE inhibitors will be important
if these drugs are to be used for the treatment of asthma. Targeting
PDE4 is summarized in Fig. 2.
4. Calcium
Calcium plays a key role in the initiation and maintenance of
ASM contraction and is closely coupled to AHR [48,49], given that it
activates myosin light chain kinase in particular, as well as other
kinases which participate in excitation-contraction coupling, such
as RhoA kinase [50]. [Ca2þ]i can be increased due to Ca2þ-influx
through Ca2þ channels in the plasma membrane (PM)dsuch as
voltage-dependent Ca2þ channels (VDCC), and store-operated Ca2þ
channels (SOCC)dand Ca2þ-release through Ca2þ-release channels
in the sarco/endoplasmic reticulumdsuch as inositol 1,4,5 tri-
sphosphate, (IP3) and ryanodine receptors.
4.1. Voltage-dependent Ca2þ channels
The role of VDCC in vascular contraction has been studied
extensively and VDCC blockers have been widely used to relax
vascular smooth muscle in diseases such as hypertension. Although
several studies have reported the existence of functional VDCC in
ASM both from humans and in a variety of animal models [51,52],
the VDCC blocker nifedipine clinically failed to induce bronchor-
elaxation in people with asthma [53,54]. The precise reason why
VDCC blockers failed to treat asthma is not clear. It might be related
to the adaptive phenotypes induced by asthma, including the
observed altered expression and function of Ca2þ channels [55].
Although VDCC blockers failed to relax ASM, VDCC have been re-
ported to play a very important role in the modulation of the
activation of TH2 cells. In a recent study, knock-down of Cav1
expression in TH2 cells impaired Ca2þ-signaling and cytokine
production, and suppressed airway hyperreactivity and inflam-
mation in a mouse model of asthma [56]. Further studies are
required to fully understand the role(s) of VDCCs in asthma.
4.2. Store-operated calcium channels
SOCC play a very important role in Ca2þ homeostasis of ASM.
Non-specific SOCC currents and Ca2þ-specific SOCC currents
(ICRAC) have been recorded from cultured human ASM [57] and
freshly-dissociated bovine ASM cells [58,59] treated with cyclo-
piazonic acid (CPA), a sarco/endoplasmic reticulum calcium
ATPase (SERCA) pump inhibitor that depletes internal stores. Both
groups showed activation of SOCC to be important in ASM
contraction. Peel et al., also report a tetra plasma membrane (PM)
protein-Orai-1 functions as a SOCC in the PM and that this is gated
through an interaction with the store-activated endoplasmic
reticulum (ER) Ca2þ sensor, stromal-interacting molecule 1
136 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144

Fig. 2. Phosphodiesterases (PDEs) and cAMP. cAMP is an important second messenger which mediates ASM relaxation and inhibits ASM proliferation. cAMP is regulated by
phosphodiesterases (PDEs) which hydrolyze it from its active form (cAMP) into inactive 50 AMP. In human ASM, PDE4 is the major PDE subtype involved in the degradation of cAMP.
Inhibition of PDE4, using the PDE4 specific inhibitors roflumilast and cilomilast, decreases the breakdown of cAMP and increases airway relaxation, as well as reducing ASM
proliferation, migration and fibronectin deposition, all of which contribute to asthma. AC ¼ adenylate cyclase.

(STIM-1, a single membrane spanning protein with an unpaired
Ca2þ binding EF hand) [57,60]. SOCC blockers were effective in
attenuating AHR in ovalbumin-sensitized guinea pigs [61].
Furthermore, a recent study showed that STIM-1/Orai-1-mediated
SOCC is involved in ASM proliferation [62].
Recently, CD38, a cell surface glycoprotein, was found to be
involved in the synthesis of cyclic ADP ribose, which potentiates the
release of Ca2þ from internal stores via ryanodine receptors in the
ASM cells [63]. Inflammatory cytokines upregulated CD38 and
cyclic ADP ribose, which is coupled with increased Ca2þ release
induced by agonists, suggesting it has a possible role in asthma [64].
CD38/cyclic ADP ribose pathway has been reported to play an
important role in airway hyperresponsiveness, since airway resis-
tance induced by methacholine was much smaller in CD38-
deficient mice than in the control mice [65]. Although the rela-
tionship between CD38/cyclic-ADP-ribose and STIM-1/Orai-1
pathway is still not clear, CD38/Cyclic-ADP-ribose modulated
SOCC possibly via interacting with TRP channels in the ASM.
4.3. Transient receptor potential channels and sarco/endoplasmic
reticulum calcium ATPase
Recently, two Ca2þ transporters e SERCA and transient receptor
potential (TRP) channels e have been shown to play an important
role in ASM physiology. TRP channels are non-selective cationic
channels that consist of several combinations of homo and hetero-
tetramers to form a functional pore unit [66]. Snetkov et al.,
described a non-selective cationic channel conductance of 25 pS,
synonymous to TRP channels in ASM cells [67]. Furthermore, TRPC,
TRPM and TRPV are expressed in ASM cells [68e71]. TRPC3 and
TRPC6 are induced by interleukin-(IL-) 13 and TGFb, with the
combination augmenting TRPC6 mRNA expression and [Ca2þ]i in
healthy ASM cells. Interestingly, coexpression of a fluorescently
tagged truncated TRPC6 with either untagged TRPC3 or TRPC6
subunit in HEK293 cells shows restoration of plasma membrane
trafficking compared to TRPC6 mutant alone. Similarly, a dominant
negative mutant of TRPC6 also selectively suppressed TRPC3 and
TRPC6 currents but not TRPC4 or TRPC5 [72]. Perhaps TRPC6 forms
a homo-multimer or hetero-multimer with TRPC3 to mediate
current activity. Using an in vivo model, one group has recently
shown increased TRPC3 expression in a murine allergic model of
asthma compared to naïve mice, and this increase in TRPC3
expression may be responsible for asthma evoked currents that led
to hyperresponsiveness [73]. Liao et al. have shown the formation
of Orai-TRP complexes, including inhibition of Ca2þ entry through
receptor-operated channels (ROC) and SOCC using a dominant
negative Orai mutant [74]. In this study, the Orai mutant prevented
diacylglycerol (DAG)-stimulated TRP channel activation.
As mentioned above, a similar transporter that plays an impor-
tant role in ASM Ca2þ-homeostasis is SERCA. SERCA-2 is the
predominant isoform in ASM. However, its expression levels are
reduced in asthma and correlate inversely with disease severity
[55]. Similar to TRP channels, inflammatory cytokines also modulate
the expression of SERCA-2 channels in cultured healthy ASM cells
[55,75]. Studies in these cultured healthy ASM cells show that
diminished SERCA-2 expression recapitulates an asthmatic pheno-
type with increased proliferation, cell spreading and IL-13-induced
eotaxin release [76]. This is possibly through a mechanism that
enhances SOCC that may involve TRP channels and Orai following
store depletion. Such signaling mechanisms between the sarco-
plasmic reticulum (SR) and SOCC will undoubtedly compromise
ASM Ca2þ-homeostasis and predispose the ASM response towards
an asthmatic phenotype.
Furthermore, SERCA-2 could be of high therapeutic value in
asthma, given that over-expression of SERCA-2 in animal models of
heart failure improves cardiac dynamics, reverses cardiac remod-
eling and improves cardiac relaxation [77e80]. If similar effects can
be achieved in asthma, it could improve lung mechanics, airway
remodeling and impaired relaxation.
With the advent of gene therapy, it may be possible to restore
SERCA and TRP channel expression back to normal levels and help
reverse the asthmatic ASM phenotype.
 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144 137

4.4. The sodiumecalcium exchanger inhibitors. Fig. 4 summarizes the role(s) of PI3-kinase, Rho kinase
and tyrosine kinases as potential targets in ASM.
Another source of Ca2þ that drives contraction is the sodiume
calcium exchanger (NCX). NCX was initially thought to only
5.1. PI-3-kinase
remove cytosolic Ca2þ, thereby restoring a relaxed or resting state.
However, it is now understood that it can equally facilitate Ca2þ
The PI3K signaling pathway plays a critical role in many cellular
entry into the cell if the driving forces acting on it are oriented
processes including contraction, proliferation, migration, cytokine/
correctly. In addition to responding to the Ca2þ gradient, NCX is also
chemokine secretion and cell survival. The family of PI3Ks consists
driven by the Naþ gradient and the membrane potential [81]. As
of 3 classes (I, II and III) of structurally related lipid kinases that
such, when cytosolic [Naþ] and/or the membrane potential rise e as
convert phosphatidylinositol 4,5 bisphosphate into phosphatidyli-
occurs during opening of non-selective cation and Cl channels
nositol 3,4,5 trisphosphate which then acts as a targeting site for
following stimulation with excitatory agonists [82] e the net
downstream signaling molecules such as AKT, resulting in altered
driving force on NCX is such that Ca2þ is translocated into the cell.
cellular function. The tumor suppressor phosphatase and tensin
In fact, electrophysiological calculations using the Nernst equation
homolog deleted on chromosome ten (PTEN) negatively regulates
show that a modest rise in [Naþ] (10e20 mM) is sufficient to cause
this pathway. Of the three classes of PI3K, class I (subdivided into
the NCX to respond to the electrical slow waves which are elicited
class IA and IB) is the most extensively studied. PI3Ks are hetero-
by all excitatory agonists [83], thus oscillating continuously
dimers consisting of a catalytic subunit (p110-a, -b, -d or -g) and
between the Ca2þ-efflux (forward) and Ca2þ-influx (reverse)
a regulatory subunit (p85a, p55a, p50a or p101). Human ASM cells
modes. Blockers of NCX-mediated Ca2þ-influx suppress agonist-
express the class I isoforms p110a, p110b and p110d but not p110g,
evoked contractions [84,85], attesting to the physiological impor-
as well as class II and class III isoforms [86e88].
tance of this pathway. Also, NCX expression is elevated in murine
The importance of the PI3K pathway in AHR and contractile
models of allergen-induced AHR [81], suggesting a possible role for
protein expression has been highlighted in many studies, through
NCX in asthma and/or usefulness for NCX-inhibitors in treatment of
the use of the pharmacological broad-spectrum PI3K inhibitors,
the same. The basic mechanisms of calcium homeostasis in ASM are
LY294002 and wortmannin and more recently through the use of
shown in Fig. 3.
novel isoform specific inhibitors, such as PIK-75 (inhibits p110a),
TGX-221 (p110b) and IC87114 (p110d). Inhibition of the PI3K
5. Kinases pathway using LY294002 decreased AHR in an ovalbumin challenge
mouse model of asthma [89] and wortmannin decreased IL-4 and
Phosphatidylinositol 3-kinase, Rho kinase and tyrosine kinases IL-13 augmented ATP-induced contraction of bovine tracheal
as novel therapeutic targets. smooth muscle cells in a collagen gel assay [90]. In support of the
ASM function is also regulated through the complex interaction role of the PI3K pathway in contraction, inhibition of PI3K also
between receptor signaling and the downstream kinases, phos- decreased the expression of the contractile proteins smooth
phatidylinositol 3-kinase (PI3K), Rho kinase and tyrosine kinases. muscle-myosin heavy chain (sm-MHC) and SM22 in serum-
Our understanding of the role(s) of these kinases in ASM function is deprived canine ASM cells in culture [91]. However, the use of
increasing with the availability of new and more targeted LY294002 and wortmannin has limitations: for example these

Fig. 3. Basic mechanisms of Ca2þ homeostasis in airway smooth muscle. A contractile agonist, such as acetylcholine, binds to a G-protein-coupled receptor and activates phos-
pholipase C (PLC). PLC enzymatic cleaves phosphatidylinositol (4,5)-bis phosphate (PIP2) to generate second messengers, inositol trisphosphate (IP3) and diacylglycerol (DAG). IP3
receptors (IP3Rs) on the sarcoplasmic reticulum (SR) can be activated by IP3 which causes an initial SR Ca2þ release that can induce further Ca2þ release from ryanodine receptors
(RyRs). On the other hand, DAG facilitates Naþ and Ca2þ entry by activating receptor-operated channels (ROCs). Agonist induced store depletion can be sensed by stromal-
interacting molecule 1 (STIM1), and these are evenly distributed within the SR. STIM forms at punctate sites and translocates to store-operated channels (SOCs). Consequently,
subplasmalemmal elevations in [Naþ] from non-selective ROCs or SOCs activation, also drive reverse-mode Naþ/Ca2þ exchanger (NCX), to facilitate Ca2þ entry. Ca2þ removal
mechanisms include the plasma membrane Ca2þ ATPase (PMCA) and normal NCX mode.
138 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144

Fig. 4. Schematic diagram highlighting the emerging roles of PI3-kinase, Rho associated coiled coil-forming protein kinase (ROCK) and tyrosine kinases in asthma. Airway smooth
muscle function is regulated through the complex interaction between receptor signaling and downstream kinases. The development of new and more targeted inhibitors has
highlighted (A) PI3-kinase, (B) Rho kinase and (C) tyrosine kinases as emerging therapeutic targets. (A) PI3-kinase. Airway smooth muscle cells express the class I PI3K isoforms
p110a, p110b and p110d. The use of novel isoform specific PI3K inhibitors including PIK-75 (p110a inhibitor), TGX-221 (p110b inhibitor) and IC87114 (p110d inhibitor) has enabled
researchers to begin to understand the role of the specific PI3K isoforms in the regulation of cellular functions. (B) Rho kinase. Inhibition of ROCK using Y27632 or fasudil reduces the
airway inflammation, airway remodeling and airway hyperresponsiveness, associated with asthma. (C) Tyrosine kinases. Receptor tyrosine kinase (RTK) inhibitors, such as tyr-
phostin AG1478, and non-RTK inhibitors, e.g. piceatannol, modulate AHR, ASM proliferation and contractility.

broad spectrum inhibitors may also have off-target effects. In
a study on rat ASM cells LY294002 reduced 5-HT-induced Ca2þ
release independent of PI3K signaling [92]. Furthermore, these
broad spectrum inhibitors do not allow identification of the specific
isoforms involved in PI3K signaling. More recent studies, using
novel isoform specific inhibitors, have begun to elucidate the
precise mechanisms involved. p110d has been identified in the
regulation of airway contraction as inhibition of p110d reduced IL-
13-induced contraction of mouse tracheal smooth muscle, as well
as AHR in mouse models of asthma [89,93,94]. In support of a role
for p110d in contraction, TGFb-induced calponin and a-actin
expression in human ASM cells [88] was inhibited by both the
broad spectrum inhibitor LY294002 and the p110d specific inhibitor
IC87114. Thus, these data support a role for targeting not only the
PI3K pathway in controlling AHR but more specifically the p110d
isoform.
In addition to its role in ASM contraction the PI3K pathway also
contributes to airway remodeling and airway inflammation. Initial
studies using LY294002 and wortmannin identified a role for PI3K
in ASM proliferation [95], with the PI3K pathway playing a more
prominent role in ASM cells derived from people with asthma
compared with those without [96] suggesting it may contribute to
the enhanced ASM bulk associated with asthma. PI3K also
contributes to migration, ECM deposition and cytokine secretion
from human ASM cells, with inhibition of the pathway decreasing
fibronectin, collagen I and connective tissue growth factor (CTGF)
deposition as well as IL-6 and vascular endothelial growth factor
(VEGF) secretion [87,88,97,98]. Recent studies using human ASM
cells in vitro have begun to investigate the roles of the specific
isoforms and have shown that different functional responses are
regulated by different isoforms with p110b and p110d regulating
VEGF and IL-6 but not fibronectin deposition [87,88]. Furthermore,
p110d has been implicated in the inflammatory response in vivo as
inhibition of p110d reduced eosinophilia, IL-4, IL-5, IL-13, eotaxin
and ICAM-1 expression in mouse models of asthma [89,93,94]. The
role of p110a is not yet fully understood; however, initial studies
have indicated it to be important in survival with homozygous
deletion of p110a being embryonic lethal and inhibition of p110a
decreasing ASM cell viability [87,99]. Although the prospect of
targeting the PI3K pathway to treat asthma seems promising, the
PI3K pathway is important for many normal cellular functions, thus
further studies are necessary to fully understand the roles of the
 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144
specific isoforms in health and disease. The roles of the specific
class I PI3K isoforms in ASM are shown in Fig. 4A.
5.2. Rho kinase
Rho associated coiled coil-forming protein kinase, is a ubiqui-
tous, serine/threonine protein kinase of which there are 2 isoforms;
ROCK 1 and ROCK 2, collectively referred to as ROCK [100]. ROCK is
the downstream target of the monomeric GTP-binding protein
RhoA [100]. Upon activation, ROCK phosphorylates and inactivates
the myosin binding subunit of myosin phosphatase, which
promotes the up-regulation of myosin light chain phosphorylation,
initiating multiple aspects of cell motility, including migration and
proliferation [101e103], and enhanced agonist-induced smooth
muscle contraction [104]. This phenomenon is termed Ca2þ sensi-
tization [105]. Although the precise mechanisms have not been
fully elucidated, it is thought that this pathway is mediated in part,
by the actions of ROCK [106,107].
5.2.1. Inhibitors of ROCK
Using the selective ROCK inhibitor [(þ)-(R)-trans-4-(1-
aminoethyl)-(4-pyridyl) cyclohexanecarboxamide] (Y27632), the
experimental inhibition of ROCK has been previously shown to
attenuate agonist-induced contractions of ASM [108e110]. In
ovalbumin-sensitized guinea pigs, inhaled Y27632 demonstrated
broncho-protective effects by preventing the development of
allergen-induced AHR [111]. Furthermore, Y27632 has also been
shown to potentiate the relaxant effects of b-adrenoceptor agonists
salbutamol and terbutaline in isolated bovine tracheal smooth
muscle, pre-constricted with methacholine [112]. This demon-
strates that the combination of b-adrenoceptor with Y27632 may
represent a more successful therapy than previous combinations
with other compounds such as methylxanthines [108].
HA-1077 (fasudil), is the only selective ROCK inhibitor currently
available for clinical use, approved for the treatment of cerebral
vasospasm [100]. However, fasudil does not have a great selectivity
profile and is a relatively potent inhibitor of other protein kinases
including protein kinase A (PKA), PKG and PKC. Furthermore,
fasudil and Y27632 are relatively weak inhibitors of ROCK [113].
More recently, azaindole-1, a novel, highly selective and orally
active ROCK inhibitor, with activity in the low nanomolar range has
been investigated. Although the effects of azaindole-1 inhibitors in
relation to airway disease have not been characterized, studies in
a variety of vascular smooth muscles demonstrate that oral
administration induces relaxant and vasodilator effects in a dose-
dependent manner [113,114].
In addition to its role in airway contraction, there is evidence of
a role for mediator dependent Rho activation in human ASM
proliferation, actin reorganization and migration [97,115], which
suggests a potential role for ROCK inhibitors in controlling the
proliferative response in ASM. Since the ROCK signaling pathway is
thought to play an important role in the pathogenesis of asthma,
the use of inhibitors in the treatment of asthma appears to repre-
sent a logical rationale (Fig. 4B). However, considering the wide-
spread expression of ROCK in virtually all tissues and the
importance of the ROCK signaling pathway in numerous cellular
functions, the inhibition of ROCK could have detrimental systemic
effects. Although fasudil is well tolerated in the clinic, there is still
a need to establish the practicality of the usefulness of ROCK as
a target for the treatment of asthma.
The mevalonate pathway also contributes to Rho/ROCK
signaling. Mevalonate is metabolized into the isoprenoid pyro-
phosphates farnesyl pyrophosphate (FPP) and geranylgeranyl
pyrophosphate (GGPP) which are substrates for the prenylation of
small GTP proteins including Rho. Inhibition of hydroxy-
139
methylglutaryl (HMG)-CoA reductase using statins decreases
AHR, ASM proliferation and ECM synthesis by inhibiting ger-
anylgeranylation of Rho [116e118]. Thus, targeting this pathway,
through the use of HMG-CoA reductase inhibitors or statins may
also be beneficial. Clinical studies examining the effects of statins in
asthma are in progress.
5.3. Tyrosine kinases
Receptor and non-receptor tyrosine kinases (RTKs and non-
RTKs respectively) are involved in the pathways associated with
AHR, airway inflammation and airway remodeling in asthma. More
specifically, mitogenic responses as well as contractile agonist-
evoked activation of ASM may involve tyrosine kinases [119].
Tyrosine kinases have the important role of transferring a phos-
phate group from ATP to a tyrosine residue of a target protein.
Activation of tyrosine kinases triggers multiple downstream
signaling pathways, including PI3K, mitogen-activated protein
kinase (MAPK), and nuclear factor-kB (NF-kB) leading to cell
differentiation, survival, proliferation, degranulation and chemo-
taxis. RTKs, of which there are 58 (as reviewed in [120]), include
epidermal growth factor receptor (EGFR) and platelet derived
growth factor receptor (PDGFR). Once activated, these receptors
undergo dimerization, tyrosine autophosphorylation and subse-
quent recruitment and activation of signaling molecules that
contain Src homology domain 2 (SH2) and phosphotyrosine
binding domains such as non-RTK Src, PI3K and phospholipase Cg,
resulting in ASM proliferation and mucus hypersecretion [as
reviewed in [121,122]]. Non-RTKs are enzymes involved in trans-
membrane signaling and are not on the cell surface but act
intracellularly.
Tyrosine kinase inhibitors, also known as tyrosine phorphor-
ylation inhibitors (tyrphostins) which target the RTKs, including
EGFR, c-kit (a stem cell factor receptor)/PDGFR, vascular endothelial
growth factor receptor (VEGFR), and the non-RTKs, such as Src,
spleen tyrosine kinase (Syk), and janus kinase (JAK) are available
and there have been numerous studies investigating the effects of
these inhibitors on the multi-faceted features observed in asthma.
However, in this review we focus specifically on the effects of
inhibitors on ASM function. Table 1 summarizes the effects of
several different tyrosine kinase inhibitors (as reviewed in
[123,124]). All in all, by way of tyrosine kinase inhibition, in vitro
and in vivo data thus far indicate a modulation in AHR, ASM
mediator release, cell proliferation and contractility (Fig. 4C).
Many tyrosine kinase inhibitors were developed initially as anti-
tumor and anti-leukemic agents, imatinib being one of them [125].
The biggest challenge in the development of drugs based upon
protein kinase inhibition has been to overcome the side effects
resulting from a lack of target selectivity [126]. Imatinib was orig-
inally developed against chronic myelogenous leukemia [125] and
was found to be well-tolerated [127]. It has favorable pharmaco-
kinetic properties; absorbed well after oral administration and
must be taken once daily [126]. Masitinib therapy in a human
clinical study for asthma was reported to be associated with more
transient skin rash and edema compared to the placebo-
administered subjects [128]. All other studies discussed in this
section were limited to either in vitro or in vivo data where no side-
effects were reported in the latter. Clinical trials investigating the
use of tyrosine kinase inhibitors for the treatment of asthma are
ongoing.
6. Anti-IgE therapy
Owing to a central role of IgE in allergic asthma, a recombinant,
humanized monoclonal anti-IgE antibody, omalizumab has been
140 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144

Table 1
RTK and non-RTK targets.

Target Effect Inhibitor Reference

Broad spectrum inhibitors Y AHR Genistein [155]
YCarbachol & serotonin induced ASM contractility (rat) [156]
YCarbachol & serotonin induced ASM contractility (rat) Tyrphostin A47 [157]
YThrombin induced ASM proliferation (guinea-pig)
EGFR tyrosine kinase Y ASM proliferation (human) Tyrphostin AG1478 [158-160]
Y AHR (mouse) [161,162]
Y ASM hyperplasia (rat) [163]
Y ASM proliferation (human) BIBX1522 [151]
YAHR & Y ASM thickening Erlotinib [164]
Y Ovalbumin-induced AHR Gefitinib [165]
c-Kit YIL-4, IL-13, CCL-2, CCL-5, CCL-6, (mouse) Imatinib [166]
[151]
Y AHR [167]
Y ASM thickening
Y AHR, Y IL-4, Y IL-13 (mouse) Sunitinib [168]
Improved asthma symptom score masitinib [128]
PDGFR Y ASM proliferation (human) Tyrphostin AG1296 [169]
Syk Y Histamine, Y ASM contraction (guinea-pig) Piceatannol [170]
Y Thrombin-induced ASM proliferation [157]
Y IgE-induced CCL11, CCL5, CXCL8, CXCL10 in ASM (human) Lentiviral shRNA silencing [142]
Y Transcriptional promoter activity of Th2-promoting TSLP R112 [144]
R406
Src YAngiotensin II-induced ASM hyperresponsiveness (rat) SU6656 [171]
Y Thrombin-induced ASM proliferation (guinea-pig) PP2 [157]
JAK Y AHR (mouse) P6 [172]
Y IL-13, eotaxin in BAL Tofactinib [173]

approved for the treatment of allergic asthma [129,130]. Omali-
zumab blocks the binding of IgE with its Fc receptors [131], and
acts by reducing serum IgE levels, FcεRI expression on inflamma-
tory cells, and by attenuating tissue mast cell function [130,132].
Although some studies were unable to detect FcεRI expression in
ASM cells [133e135], others suggest that ASM cells may indeed be
one of the FcεRI-bearing cells in airways [136]. In particular, ASM
cells were shown to express both of the functional high affinity
(FcεRI) [137e139] and the low affinity (FcεRII/CD23) receptors
[140]. The IgE-binding FcεRI-a-chain immunoreactivity was
detected on smooth muscle bundles from bronchial biopsies of
subjects with asthma. Cross-linking of FcεRI-bound IgE led to
a marked release of pro-asthmatic Th2 cytokines IL-4, IL-5, IL-13,
and eosinophil-attracting CCL11/eotaxin-1 chemokine; and
a rapid and transient increase in [Ca2þ]i mobilization that results
in smooth muscle contraction [138,141]. Blocking of IgE binding to
FcεRI attenuated the IgE cross-linking-induced mediator release
from ASM [138]. Moreover, both tumor necrosis factor (TNF) and
IL-4 were recently shown to upregulate the FcεRI expression, and
TNF presensitization amplified the IgE-induced ASM cell release of
CCL11/eotaxin-1, CCL5/RANTES, CXCL8/IL-8, and CXCL10/IP-10
chemokines (Fig. 5) [142]. Other mediators induced by FcεRI
activation in ASM include matrix metalloprotease 1 (MMP-1)
[143], and TSLP e a pro-allergic mediator known for its modula-
tory effects on dendritic cells which eventually instruct naïve T
cells to differentiate into Th2 cells [144]. In the only ASM specific
study, omalizumab treatment was shown to abrogate IgE-induced
mediators of asthma relevance such as IL-4, IL-6, CXCL8/IL-8, and
TNF from human ASM cells (Fig. 5) [139]. Although omalizumab
inhibits both the early- and late-phase asthmatic response to
inhaled allergens [145], and decreases nonspecific AHR in vitro
[146], it had little effects on AHR to methacholine in asthma
patients [145,147]; while its effects on airway remodeling remain
completely unknown. At least in a murine model of chronic
asthma, anti-IgE treatment decreased the thickness of ASM layer
and peribronchial fibrosis compared with the untreated mice;
suggesting that (i) IgE could perhaps be one of the factors inducing
ASM remodeling in vivo, and (ii) targeting IgE/FcεR network on
ASM with anti-IgE represents a novel approach in reducing airway
remodeling [148].
Further studies are required to evaluate the association between
FcεR expression in ASM tissue and allergic asthma severity in
a large population, and the outcome of IgE/FcεRI activation on ASM
contraction, proliferation and/or migration. As a relationship
between chronic inflammation and structural remodeling in
asthma remains controversial, it would be worthwhile to test
whether omalizumab therapy can improve airway remodeling.
7. Bronchial thermoplasty
Rather than trying to control pharmacologically the mechanical
output of the smooth muscle using inhibitors of excitatory
responses (anti-cholinergics; anti-histamines; anti-leukotrienes)
or agonists which promote relaxant responses (b-agonists; PDE
inhibitors), some have hypothesized that it might be possible to
remove the ASM entirely. Others have questioned the physiological
importance or even usefulness of the ASM [149] e in fact, referring
to it as the appendix of the lung e and gone on to show that it can
be selectively removed using thermal energy. Preclinical studies
showed that warming the airway wall to 65  C for 30 s results in an
immediate loss of contractile function which may be related to
denaturation of myosin itself [150]. Over the course of a few weeks,
there is ablation of the ASM, while the connective tissue, epithe-
lium and vasculature of the wall appear to be unaffected [151]. The
effects on the airway innervation have not yet been described. More
importantly, clinical studies in patients with mild to moderate
asthma [152] and moderate to severe asthma [153,154] also show
no major negative consequences, but an improvement in asthma
symptom scores. The mechanisms underlying the extreme thermal
sensitivity of the ASM and the cellular selectivity of such a blunt
intervention is not yet clear.
8. Future directions
Although there is some debate as to whether ASM plays an
important role at all in normal airway physiology [149], very few
 S. Siddiqui et al. / Pulmonary Pharmacology & Therapeutics 26 (2013) 132e144
Fig. 5. Anti-IgE (omalizumab) effects on IgE-induced ASM function. Both FcεRI and FcεRII/CD23 activation on ASM by IgE (allergen-dependent or -independent) leads to proin-
flammatory mediator and intracellular Ca2þ release, which may eventually contribute to airway inflammation, and AHR. Anti-IgE (omalizumab) therapy likely inhibits these effector
functions by blocking the interaction of IgE with FcεRs on ASM cells.
would contest that ASM is a major problem in asthma. Whether
that problem is due to altered function of the individual ASM cells
and/or hyperplasia of the whole airway wall tissue, bronchodilation
is the singular goal of rescue asthma therapy. Despite these facts,
there have been no substantially new pharmacological agents
introduced to the market to target the ASM. For the development of
future therapeutic interventions, we need to fully understand the
role of the ASM in the pathophysiology of asthma and have a better
understanding of the multifaceted action of potential drug targets.
These will include various cell surface receptors, intracellular
targets which play key roles in the signaling cascades triggered by
bronchoconstrictors, including various kinases (MLCK; PI3K, ROCK;
Src; tyrosine kinases) and Ca2þ itself.
Funding sources
N.S.R. was supported by graduate studentship from the Canada
Lung Association-Canadian Thoracic Society. S.S. was a recipient of
the Alexander McFee Studentship, Faculty of Medicine, McGill
University, Montréal, QC, Canada. O.O.O. is supported by the CIHR/
IMPACT Strategic Training Fellowship. L.M.M was supported by the
National Health and Medical Research Council, Australia and the
LAM Australasia Research Alliance.
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