Extracellular adenosine is produced in a coordinated manner from cells following cellular challenge or tissue injury. Once produced, it serves as an autocrine- and paracrine-signaling molecule through its interactions with seven-membrane-spanning G-protein-coupled adenosine receptors. These signaling pathways have widespread physiological and pathophysiological functions. Immune cells express adenosine receptors and respond to adenosine or adenosine agonists in diverse manners. Extensive in vitro and in vivo studies have identified potent anti-inflammatory functions for all of the adenosine receptors on many different inflammatory cells and in various inflammatory disease processes. In addition, specific proinflammatory functions have also been ascribed to adenosine receptor activation. The potent effects of adenosine signaling on the regulation of inflammation suggest that targeting specific adenosine receptor activation or inactivation using selective agonists and antagonists could have important therapeutic implications in numerous diseases. This review is designed to summarize the current status of adenosine receptor signaling in various inflammatory cells and in models of inflammation, with an emphasis on the advancement of adenosine-based therapeutics to treat inflammatory disorders.

The pathophysiological processes underlying respiratory diseases like asthma are complex, resulting in an overwhelming choice of potential targets for the novel treatment of this disease. Despite this complexity, asthmatic subjects are uniquely sensitive to a range of substances like adenosine, thought to act indirectly to evoke changes in respiratory mechanics and in the underlying pathology, and thereby to offer novel insights into the pathophysiology of this disease. Adenosine is of particular interest because this substance is produced endogenously by many cells during hypoxia, stress, allergic stimulation, and exercise. Extracellular adenosine can be measured in significant concentrations within the airways; can be shown to activate adenosine receptor (AR) subtypes on lung resident cells and migrating inflammatory cells, thereby altering their function, and could therefore play a significant role in this disease. Many preclinical in vitro and in vivo studies have documented the roles of the various AR subtypes in regulating cell function and how they might have a beneficial impact in disease models. Agonists and antagonists of some of these receptor subtypes have been developed and have progressed to clinical studies in order to evaluate their potential as novel antiasthma drugs. In this chapter, we will highlight the roles of adenosine and AR subtypes in many of the characteristic features of asthma: airway obstruction, inflammation, bronchial hyperresponsiveness and remodeling. We will also discuss the merit of targeting each receptor subtype in the development of novel antiasthma drugs.

Adenosine, an important signaling molecule in asthma, produces bronchoconstriction in asthmatics. Adenosine produces bronchoconstriction in allergic rabbits, primates, and humans by activating A 1 adenosine receptors (ARs). Effects of L-97-1 [3-[2-(4-aminophenyl)-ethyl]-8-benzyl-7-{2-ethyl-(2-hydroxyethyl)-amino]-ethyl}-1-propyl-3,7-dihydro-purine-2,6-dione] a water-soluble, small molecule A 1 AR antagonist were investigated on early and late phase allergic responses (EAR and LAR) in a hyper-responsive rabbit model of asthma. Rabbits were made allergic by intraperitoneal injections of house dust mite [HDM; 312 allergen units (AU)] extract within 24 h of their birth. Booster HDM injections were given weekly for 1 month, biweekly for 4 months, and continued monthly thereafter. Hyperresponsiveness was monitored by measuring lung dynamic compliance (Cdyn), after histamine or adenosine aerosol challenge in allergic rabbits. Hyper-responsive rabbits were subjected to aerosol of HDM (2500 AU), 1 h after intragastric administration of L-97-1 (10 mg/kg) solution or an equivalent volume of saline. Cdyn was significantly higher after treatment with L-97-1 compared with untreated controls (p Ͻ 0.05 n ϭ 5). Histamine PC 30 was significantly higher (p Ͻ 0.05; n ϭ 5) after L-97-1 at 24 h compared with histamine PC 30 at 24 h after HDM. Adenosine PC 30 was significantly higher at 15 min and 6 h after L-97-1 compared with control (p Ͻ 0.05; n ϭ 5). L-97-1 showed strong affinity for human A 1 ARs in radioligand binding studies and no inhibition toward human phosphodiesterase II, III, IV, and V enzymes. These data suggest that L-97-1 produces a significant reduction of histamine or adenosineinduced hyper-responsiveness and HDM-induced EAR and LAR in allergic rabbits by blocking A1 ARs and may be beneficial as an oral therapy for human asthma.Adenosine is an endogenous nucleoside-signaling molecule and acts on adenosine receptors (ARs) to produce a number of physiological effects in humans, including bronchoconstriction and lung inflammation. Moreover, it is becoming increasingly apparent that adenosine is an important signaling molecule in human asthma. When administered by inhalation, adenosine produces concentration-dependent bronchoconstriction in patients with asthma, but not in normal subjects (Cushley et al., 1983;Polosa, 2002;Rorke and Holgate, 2002). Adenosine levels are increased in the bronchoalveolar fluid of asthmatics and also in the plasma of patients with exercise-induced asthma (Driver et al., 1993;Vizi et al., 2002). There is also an association between allergen exposure and adenosine monophosphate (AMP) responsiveness in asthmatics (Currie et al., 2003). In asthma, the airway re-

According to an executive summary of the GINA dissemination committee report, it is now estimated that approximately 300 million people (5% of the global population or 1 in 20 persons) have asthma. Despite the scientific progress made over the past several decades toward improving our understanding of the pathophysiology of asthma, there is still a great need for improved therapies, particularly oral therapies that enhance patient compliance and that target new mechanisms of action. Adenosine is an important signalling molecule in human asthma. By acting on extracellular G-protein-coupled ARs on a number of different cell types important in the pathophysiology of human asthma, adenosine affects bronchial reactivity, inflammation and airway remodelling. Four AR subtypes (A 1 , A 2a , A 2b and A 3 ) have been cloned in humans, are expressed in the lung, and are all targets for drug development for human asthma. This review summarizes what is known about these AR subtypes and their function in human asthma as well as the pros and cons of therapeutic approaches to these AR targets. A number of molecules with high affinity and high selectivity for the human AR subtypes have entered clinical trials or are poised to enter clinical trials as anti-asthma treatments. With the availability of these molecules for testing in humans, the function of ARs in human asthma, as well as the safety and efficacy of approaches to the different AR targets, can now be determined.British Journal of Pharmacology (2008) 155, 475-486; doi:10.1038/bjp.2008 Keywords: adenosine; ARs; asthma; A 1 AR; A 2a AR; A 2b AR; A 3 AR Abbreviations: ADA, adenosine deaminase; AMP, adenosine monophosphate; APCs, antigen presenting cells; AR, adenosine receptor; BAL, bronchoalveolar lavage; CFTR, cystic fibrosis transmembrane conductance regulator; CPA, N 6 -cyclopentyladenosine; DPCPX, 1,3-dipropyl-8-cyclopentylxanthine; FEV 1 , forced expiratory volume in one second; GINA, Global Initiative for Asthma; HBECs, human bronchial epithelial cells; HBSMCs, human bronchial smooth muscle cells; ICSs, inhaled corticosteroids; IP(3), inositol trisphosphate; KO, knockout; LABAs, long acting beta 2 agonists; LAMAs, long acting muscarinic antagonists; LTRAs, leukotriene receptor antagonists; MCP, monocyte chemotactic protein; NECA, 5 0 -N-ethylcarboxamidoadenosine; PC 20 for AMP, provocative concentration (PC) of AMP required to reduce FEV 1 by 20%; PDE-IV, phosphodiesterase-IV; RT-PCR, reverse transcription-polymerase chain reaction IntroductionUnmet medical need for new drugs that improve patient compliance and prevent and treat airway remodelling According to an executive summary of the GINA dissemination committee report, it is now estimated that approximately 300 million people (5% of the global population or 1 in 20 persons) have asthma (Masoli et al., 2004). Moreover, according to the World Health Organization, 255 000 people died of asthma in 2005 and asthma is the most common chronic disease among children. Despite the scientific progress made over the past s...

Previously, it was reported that A(1) adenosine receptor antagonists prevent endotoxin-induced acute lung injury and pulmonary arterial endothelial cell damage. In competition radioligand binding experiments in membranes prepared from human pulmonary artery endothelial cells (PAECs), lipopolysaccharides (LPSs) of Escherichia coli, Salmonella typhimurium, Klebsiella pneumoniae, and Pseudomonas aeruginosa displaced the binding of a selective A(1) adenosine receptor antagonist [(125)I]-BWA844U (IC(50) values: 195 ng/ml, 290 ng/ml, 602 ng/ml, and 693 ng/ml, respectively) in a dose-dependent, competitive manner. There was no displacement of this radioligand by enterotoxin (< or = 10 microg/ml), diphosphoryl lipid A (< or = 10 microg/ml), and glycolipids, monosialoganglioside (< or = 1 microg/ml), lactocerebroside (< or = 100 microg/ml), or NBD galactocerebroside (< or = 100 microg/ml). Based on calculated IC(50) values, LPS (E. coli, IC(50) 111 ng/ml) displaced the selective A(1) adenosine receptor agonist, [(3)H]-2-chloro, N(6)-cyclopentyladenosine (CCPA) in human PAECs with a potency profile, CCPA > LPS > 2-phenylaminoadenosine (CV 1808), a selective A(2) adenosine receptor agonist. The potency profile for displacement of the selective A(2a) adenosine receptor agonist [(3)H]-CGS 21680 was CV 1808 > CCPA. LPS (E. coli 0.1 pg/ml-10 microg/ml) did not displace [(3)H]-CGS 21680 binding. In human PAECs, IL-6 and TXA(2) release induced by LPS (0-1 microg/ml) or CCPA (0-1 microM) at high doses was significantly reduced by the selective A(1) adenosine receptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; 1 microM). These data suggest that LPS binds to and activates A(1) adenosine receptors on human PAECs to induce the release of IL-6 and TXA(2). Activation of A(1) adenosine receptors on human PAECs by LPS, may contribute to the pathophysiology of acute lung injury associated with Gram-negative septicemia and endotoxemia.

Adenosine produces bronchoconstriction in allergic rabbits, primates, and humans by activating adenosine A 1 receptors. Previously, it is reported that a high dose of L-97-1, a water-soluble, small molecule adenosine A 1 receptor antagonist, blocks early and late allergic responses, and bronchial hyper-responsiveness to histamine in a hyper-responsive rabbit model of allergic asthma. Effects of a lower dose of L-97-1 are compared to montelukast, a cysteinyl leukotriene-1 receptor antagonist on early allergic response, late allergic response, bronchial hyper-responsiveness, and inflammatory cells in bronchoalveolar lavage (BAL) fluid following house dust mite administration. Rabbits received intraperitoneal injections of house dust mite extract within 24 h of birth followed by booster house dust mite injections. Hyper-responsive rabbits received aerosolized house dust mite (2500 allergen units), 1 h after intragastric administration of L-97-1 (1 mg/kg) or montelukast (0.15 mg/ kg) and lung dynamic compliance was measured for 6 h. Lung dynamic compliance was significantly higher following L-97-1 at all time points and with montelukast at 60-300 min following house dust mite (P < 0.05). L-97-1 blocks both early and late allergic responses. Montelukast blocks only the late allergic response. Both L-97-1 and montelukast significantly blocked bronchial hyperresponsiveness at 24 h (P < 0.05). Both L-97-1 and montelukast significantly reduced BAL eosinophils at 6 h and neutrophils at 6 and 24 h (P < 0.05). L-97-1 significantly reduced BAL lymphocytes at 6 and 24 h (P < 0.05). Montelukast significantly reduced BAL macrophages at 6 and 24 h (P < 0.05). By blocking both bronchoconstriction and airway inflammation, L-97-1 may be an effective oral anti-asthma treatment.

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Yersinia pestis, a Gram-negative bacillus causing plague and Centers for Disease Control and Prevention (CDC) classified Category A pathogen, has high potential as a bioweapon. Lipopolysaccharide, a virulence factor for Y. pestis, binds to and activates A1 adenosine receptor (AR)s and, in animals, A1AR antagonists block induced acute lung injury (ALI) and increase survival following cecal ligation and perforation. In this study, rats were infected intratracheally with viable Y. pestis [CO99 (pCD1+/Δpgm) 1 × 108 CFU/animal] and treated daily for 3 d with ciprofloxacin (cipro), the A1AR antagonist L-97-1, or cipro plus L-97-1 starting at 0, 6, 24, 48, or 72 h post-Y. pestis. At 72 h post-Y. pestis, cipro plus L-97-1 significantly improved 6-d survival to 60–70% vs 28% for cipro plus H2O and 33% for untreated Y. pestis controls (P = 0.02, logrank test). Lung edema, hemorrhage and leukocyte infiltration index (LII) were evaluated histologically to produce ALI scores. Cipro plus L-97-1 significantly reduced lung edema, as well as aggregate lung injury scores vs controls or cipro plus H2O, and LII vs controls (P < 0.05, Student's unpaired t test). These results support efficacy for L-97-1 as a post-exposure medical countermeasure, adjunctive therapy to antibiotics for Y. pestis.