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The OXE receptor: a new therapeutic approach for asthma?
Opinion
TRENDS in Molecular Medicine
Vol.11 No.6 June 2005
The OXE receptor: a new therapeutic approach for asthma? Carol E. Jones Respiratory Diseases Therapeutic Area, Novartis Institutes for Biomedical Research, Horsham, RH12 5AB, UK
The eicosanoid 5-oxo-6E,8Z,11Z,14Z-eicosatetraenoic acid (5-oxo-ETE) has recently been identified as the ligand for the oxoeicosanoid (OXE) receptor. In vitro and in vivo studies have suggested that 5-oxo-ETE has a role in the asthmatic inflammatory response and it has been shown to stimulate eosinophil migration to the airways. New data suggest that eosinophils have an important role in the pathogenesis of asthma, being required for mucus accumulation, airway hyperresponsiveness and remodelling of the airways. However, there are several mediators that can stimulate the recruitment of eosinophils to the airways and the development of antagonists against the OXE receptor is required to evaluate the potential of the OXE receptor as a new therapeutic approach for asthma.
Introduction Asthma is an inflammatory condition of the airways that is becoming increasingly prevalent, especially in children. Current estimates of prevalence are between 10 and 20% [1]. The disease is characterised by airway hyperresponsiveness, airway inflammation, excessive mucus production and structural changes to the airways (remodelling). The infiltration of leukocytes into lung tissue and airways is a key feature of asthma, with high numbers of eosinophils observed in the airways; the number of eosinophils correlates with disease severity [2]. The role of the eosinophil in asthma has been the subject of much debate. Although eosinophils in the airway are a characteristic feature of asthma, the precise role of the eosinophil in asthma pathophysiology remains controversial [3–5]. Early work in this area suggested that the eosinophil contributed to asthma pathogenesis through the release of granule-associated proteins, which cause tissue damage, lipid mediators, which cause bronchoconstriction, and the release of reactive oxygen species [5–7]. In addition, eosinophils can secrete Th2 cytokines and present antigens [8]. The role of the eosinophil in asthma became uncertain because clinical trials with anti-IL-5 antibodies failed to deliver the expected results. Although these antibodies were effective at depleting eosinophils, there was no significant clinical improvement [3,5]. However, recent in vivo data have suggested a role for the eosinophil in airway remodelling in asthma [9]. The targeted depletion of eosinophils using transgenic mice indicates Corresponding author: Jones, C.E. ([email protected]). Available online 10 May 2005
that eosinophils have a role in lung remodelling [9]. Eosinophils can also produce a range of molecules that are proposed to be involved in the remodelling process, providing further evidence suggesting a role for eosinophils in causing structural changes to the airways [3]. The accumulation of eosinophils into the airways is thought to be mediated by several factors. Following mobilisation from the bone marrow in response to cytokines, eosinophils migrate to the lung and other tissues in response to locally released chemoattractants. Certain lipid mediators, including products of arachidonic acid metabolism, and chemokines, such as eotaxin, are potent stimulators of this process [2,10,11]. An eosinophil chemoattractant that has recently attracted attention is 5-oxo-6E,8Z,11Z,14Z-eicosatetraenoic acid (5-oxo-ETE), a lipid that activates the oxoeicosanoid (OXE) receptor. With recent data shedding new insight into the role of the eosinophil in asthma pathogenesis, eosinophil chemoattractants should be revisited as potential targets for asthma. Here, 5-oxo-ETE and its recently identified receptor are discussed as a new therapeutic approach for asthma in light of recent developments in the understanding of the role of eosinophils in the disease process. The 5-LO pathway and 5-oxo-ETE Perhaps the most familiar products of the metabolism of arachidonic acid by the actions of 5-lipoxygenase (5-LO) are the cysteinyl leukotrienes (cysLTs), particularly leukotriene C4 (LTC4) and LTD4 (Figure 1). The cysLTs are potent mediators of contractile responses in the airways and antagonists of the cysLT1 receptor are marketed as therapies for asthma [10,12]. Another metabolite of arachidonic acid that is associated with the 5-LO pathway is 5-oxo-ETE. The major route of production of 5-oxo-ETE is believed to be from the precursor 5-hydroxy6,8,11,14 (E,Z,Z,Z)-eicosatetraenoic acid (5-HETE) by the actions of a specific dehydrogenase. The biosynthesis and metabolism of 5-oxo-ETE has recently been described [13,14]. Multiple cell types have been reported to produce 5-oxo-ETE, including monocytes, eosinophils and neutrophils, in response to nonphysiological stimuli, such as the calcium ionophore A23187 or the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA), but produce little 5-oxo-ETE without stimulation [15–17]. More recently, oxidative stress has been shown to stimulate 5-oxo-ETE synthesis by blood monocytes, lymphocytes and platelets but not neutrophils [18], which is of relevance to inflammatory respiratory diseases, such as
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Opinion
TRENDS in Molecular Medicine
Arachidonic acid
COX-1 COX-2
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Figure 1. Major pathways of arachidonic acid metabolism. Key products of arachidonic acid metabolism that are formed by the 5-LO, 12-LO, 15-LO, COX-1 and COX-2 pathways and are believed to have a role in the asthmatic response are shown (boxed). Metabolism of arachidonic acid on the 5-LO pathways results in the formation of the cysteinyl leukotrienes LTC4 and LTD4 in addition to 5-oxo-ETE. LXA4 is formed by the 5-LO, 12-LO and 15-LO pathways. The metabolism of arachidonic acid through the COX-1 and -2 pathways results in the formation of TXA2 and PGD2, which have been proposed to have a role in the asthmatic response. Abbreviations: COX, cyclooxygenase; 5-HETE, 5-hydroxy-6,8,11,14 (E,Z,Z,Z)-eicosatetraenoic acid; 5(S)-HPETE, 5(S)-hydroxyperoxy-6,8,11,14 (E,Z,Z,Z)-eicosatetraenoic acid; LO, lipoxygenase; LT, leukotriene; 5-oxo-ETE, 5-oxo6E,8Z,11Z,14Z-eicosatetraenoic acid; LX, lipoxin; PG, prostaglandin; TX, thromboxane.
asthma and chronic obstructive pulmonary diseases (COPDs). Most of the cell types described produce relatively small quantities of 5-oxo-ETE, which is in contrast to the higher levels reported from monocytederived dendritic cells [19].
Identification of the OXE receptor Although a receptor binding the lipid 5-oxo-ETE has been described in the literature for over ten years, its molecular identity remained unknown until recently. Two groups independently described a genome-datamining approach used for the molecular cloning of a G-protein coupled receptor (GPCR) activated by the ligand 5-oxo-ETE [20,21]. The two 5-oxo-ETE receptors identified, R527 [20]
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and TG1019 [21], are almost identical except for a 39 amino acid N-terminal extension in TG1019 and a single amino change towards the C-terminus. The receptor, recently renamed the OXE receptor by International Union of Pharmacology (IUPHAR) [13], is located on chromosome 2 and shows closest homology to the GPCR HM74, which has been reported to bind to nicotinic acid [22], and GPR81, for which the ligand has not yet been identified. The sequence identity at the amino acid level of the OXE receptor to HM74 and GPR81 is 40%. Two structurally related lipids, 5(S)-hydroxyperoxy6,8,11,14 (E,Z,Z,Z)-eicosatetraenoic acid [5(S)-HPETE] and 5(S)-HETE, were also found to be agonists of the OXE receptor, but are reported to be much less potent [20,21]. In primary cells, the OXE receptor has a restricted expression profile, with high levels of mRNA expression reported for eosinophils and neutrophils and lower levels in lung-derived macrophages [20]. In the same study, virtually no expression was detected in other cultured human cells, including lung epithelial cells, smooth muscle cells or lung fibroblasts. A more detailed study of the functions of the OXE receptor has been limited by the lack of good pharmacological tools. Four weak antagonists of the 5-oxo-ETE receptor have been described: 4Z,7Z,10Z,13Z,16Z,19Zdocosahexaenoic acid, 5Z,8Z,11Z,14Z,17Z-eicosapentaenoic acid, di-homo-g-linolenic acid and 11Z,14Z,17Zeicosatrienoic acid, with EC50 values of 1.6, 6.0, 3.7 and 5.1 mM, respectively, in a GTPgS assay [21]. The expression and pharmacological profiles of the OXE receptor suggest that it is the receptor described on human eosinophils, neutrophils and monocytes that is activated by 5-oxo-ETE, but not characterised at the molecular level [17,23,24]. The observed expression of the OXE receptor on human eosinophils, neutrophils and monocytes correlates with reported functional effects of 5-oxo-ETE on the same cell types. Many of the pharmacological responses reported for 5-oxo-ETE on human eosinophils and neutrophils, such as chemotaxis, are sensitive to pertussis toxin, indicating that the receptor is coupled to the G-protein Gai [25], which is also the case for the recombinant receptor [20]. E osinophil
Neutrophil Chemotaxis Ca2+ mobilisation CD11b expression Degranulation after priming
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5-oxo-ETE
Chemotaxis Ca2+ mobilisation CD11b expression L-selectin shedding Degranulation after priming Superoxide production
Chemotaxis GM-CSF production Monocyte TRENDS in Molecular Medicine
Figure 2. The actions of 5-oxo-ETE on primary cells. The eicosanoid 5-oxo-ETE has multiple actions on eosinophils, neutrophils and monocytes relevant to the inflammatory response in asthma. Examples of these effects of 5-oxo-ETE are given and include chemotaxis, calcium mobilisation, CD11b expression and degranulation [25–34,36,48]. www.sciencedirect.com
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5-oxo-ETE and inflammation Data from in vitro studies The eicosanoid 5-oxo-ETE has previously been suggested to have a role in the inflammatory response. A range of biological effects of 5-oxo-ETE that are relevant to the inflammatory response and asthma have been reported for eosinophils, neutrophils, monocytes and, more recently, for airway smooth muscle (Figure 2). These biological effects include cell migration, surface expression of the integrin CD11b, L-selectin shedding, calcium mobilisation, superoxide production and smooth muscle contraction. 5-oxo-ETE is a potent chemotactic factor, stimulating the migration of human eosinophils. A comparison of eosinophil chemoattractants indicates that 5-oxo-ETE is one of the most potent mediators known. Studies have shown that 5-oxo-ETE is more potent than plateletactivating factor (PAF) in eosinophil transmigration assays, inducing a slightly greater transmigration at 1 nM than does PAF at 1000 nM [26]. In fact, a low (1 nM) concentration of 5-oxo-ETE has been shown to potentiate the response of eosinophils to PAF in cellmigration assays [17], whereas the effects of 5-oxo-ETE and PAF were additive at a higher concentration (3 nM) of 5-oxo-ETE. Although reported to be w10-times less potent than eotaxin [27], in a comparison of a range of eosinophil chemoattractants, including PAF, eotaxin and prostaglandin D2 (PGD2), 5-oxo-ETE elicited the greatest chemotactic response [28]. 5-oxo-ETE also causes calcium mobilisation in eosinophils, a process that is thought to contribute to granulocyte migration, and EC50 values are reported to be in the range of 10 nM [29]. 5-oxo-ETE has also been demonstrated to cause CD11b expression and L-selectin shedding on human eosinophils, suggesting that it is important for the interaction of eosinophils with vascular endothelial cells and in transendothelial migration [30]. In addition, 5-oxo-ETE can stimulate the degranulation of eosinophils when the cells are primed with granulocyte macrophage-colony stimulating factor (GM-CSF), although it has little effect if the cells are not primed [31]. Studies have suggested that 5-oxo-ETE can synergise with other chemoattractants, such as eotaxin and RANTES (regulated on activation, normal T-cell expressed and secreted), in the eosinophilic chemotactic response [27,29]. This might be of relevance to the in vivo situations in which there are multiple mediators present, including the range of lipid mediators and chemokines that are released during pulmonary inflammation. Several similar responses to 5-oxo-ETE have been reported for human neutrophils, with effects on calcium mobilisation, cell migration, CD11b expression and degranulation after GM-CSF priming [25,32–34]. 5-oxoETE has been reported to bind to its receptor on human neutrophils with a Kd of 4 nM [35]. The chemotactic responses elicited by 5-oxo-ETE are less potent on neutrophils than those observed for eosinophils, paralleling the differences observed in relative receptor expression on these cell types [20]. Indeed, this eicosanoid is approximately five-times less potent at stimulating neutrophil than it is for eosinophil chemotaxis [17,23]. 5-oxo-ETE stimulates similar maximal responses in www.sciencedirect.com
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human neutrophils to the leukotriene LTB4, although LTB4 is a more potent mediator [14]. In contrast to the information available for the effects of 5-oxo-ETE on eosinophils and neutrophils, less is known about the effects on human monocytes. In human monocytes, 5-oxo-ETE induces a chemotactic response, although not as strong as that of monocyte-chemotactic protein-1 (MCP-1) or N-formyl-Met-Leu-Phe (fMLP) [24,14]. However, it was demonstrated that 5-oxo-ETE enhances the effects of MCP-1 in a chemotaxis assay. More recently, it has been shown that 5-oxo-ETE can stimulate the release of the eosinophil survival factor GM-CSF from human monocytes [36]. Effects of 5-oxo-ETE on airway smooth muscle have been reported, further supporting a potential link between 5-oxo-ETE and asthma. Guinea pig airway smooth muscle produced contractile responses to 5-oxo-ETE through the activation of the calcium pool and the responses were partially inhibited by a Rho-kinase pathway inhibitor [37]. However, the EC50 reported for 5-oxo-ETE in this system was 0.89 mM and therefore the physiological relevance is not clear. Although no detectable expression of the OXE receptor was reported in cultured human airway smooth muscle cells [20], it will be interesting to see if the contractile responses to 5-oxo-ETE extend to human tissue. Data from in vivo studies Data supporting a role for 5-oxo-ETE in the inflammatory response in vivo come from a rat model, in which 5-oxoETE delivered directly to the lung causes the recruitment of eosinophils to the airways [38,39]. Although a response was observed for 5-oxo-ETE in this rat model, a rodent homologue of the human OXE receptor has not yet been reported. In humans, 5-oxo-ETE has been recently reported to induce the infiltration of eosinophils, neutrophils and macrophages into the skin of healthy and asthmatic subjects [40]. No effects of 5-oxo-ETE on mast cells or lymphocytes were observed. Asthmatics were reported to be more sensitive to 5-oxo-ETE in this study, resulting in the infiltration of greater numbers of eosinophils into the skin [40]. In contrast to the results from the study in human skin, an in vitro study looking at the effects of 5-oxo-ETE on migration of human bloodderived eosinophils isolated from either asthmatic or healthy subjects showed no differences in cell migration between the two patient groups [26]. No reported studies have looked at differences in OXE-receptor expression between diseased and normal tissue. However, in a human skin model, 5-oxo-ETE caused a greater recruitment of eosinophils into the skin in subjects with asthma [40]. The OXE receptor – a promising asthma target? The eicosanoid 5-oxo-ETE has been reported to be involved in multiple processes associated with the inflammatory response. 5-oxo-ETE is produced under conditions of inflammatory stress [18], which commonly occur in respiratory conditions such as asthma and COPD. In vitro studies have demonstrated a role for 5-oxo-ETE in eosinophil and, to a lesser extent, neutrophil and monocyte cell migration [24–26]. In vivo data, both in rats and humans, support a role for 5-oxo-ETE in the recruitment
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of eosinophils to sites of inflammation [38,40]. Thus, these effects of 5-oxo-ETE on eosinophil migration suggest that it is involved in eosinophil-mediated allergic reactions. As previously described, when comparing the relative potencies of several mediators in the migration of eosinophils, whereas eotaxin is w10-fold more potent than is 5-oxoETE in its response [27], 5-oxo-ETE has been shown to cause the greatest maximal response [28]. Thus, the ability of 5-oxo-ETE to synergise with the chemokines RANTES and eotaxin in the eosinophil chemotactic response might be important at certain sites of inflammation. Other relevant reported effects of 5-oxo-ETE include cellular degranulation. Although 5-oxo-ETE alone has little effect on eosinophil and neutrophil degranulation, priming of the cells with GM-CSF enhances 5-oxo-ETE-induced degranulation of both cell types. In contrast to other chemotactic factors, such as eotaxin and PAF, 5-oxo-ETE can stimulate GM-CSF release from human monocytes and thereby not only stimulate the recruitment of eosinophils to the airways but also promote their survival [36]. GM-CSF itself causes degranulation and superoxide production and is an important survival factor once eosinophils reach the lung [41]. This is in contrast to the cytokine IL-5, which is important in the mobilisation of eosinophils from the bone marrow [2]. However, it is not known whether 5-oxo-ETE has a role in eosinophil mobilisation. Although the effects of 5-oxo-ETE on human neutrophils are less pronounced than those on eosinophils, a role for 5-oxo-ETE in persistent inflammation has been suggested in which neutrophils have become desensitized to LTB4 [14]. One concern when evaluating the role and relevance of 5-oxo-ETE in asthma is that there are currently no reports of 5-oxo-ETE measured in vivo either in human airways or in animals. However, this might reflect difficulties in accurately measuring 5-oxo-ETE. A sensitive quantitative method for the measurement of 5-oxo-ETE has recently been reported [42] and studies investigating levels of 5-oxo-ETE in asthmatic airway are awaited. Several other metabolites of arachidonic acid have been associated with asthma and these are illustrated in Figure 1. These include the leukotrienes LTC4 and LTD4 formed by the 5-LO pathway, which act at the cysLT1 receptor, are potent bronchoconstrictors and have effects on mucociliary clearance and eosinophilic inflammation [10,12,43]. Lipoxin A4 (LXA4) is another arachidonic acid metabolite formed by the actions of 15-LO and 12-LO [44] and stable analogues of LXA4 have been shown to have anti-inflammatory properties in mouse models of asthma [45]. A further proposed mediator of the asthmatic response is thromboxane A2 (TXA2), which is an arachidonic acid metabolite formed by the cyclooxygenase-2 pathway. TXA2 binds to the TXA2 (TP) receptor and causes bronchoconstriction of the airways [46]. More recently, the CRTH2 (chemoattractant-receptor-homologous molecule expressed on Th2 cells) receptor has been described, which binds to another arachidonic acid metabolite, PGD2. The actions of PGD2 acting through CRTH2 have been proposed to have an important role in allergic inflammation through effects on Th2 cells, eosinophils and www.sciencedirect.com
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basophils [47]. Indeed, taken together, the metabolites of arachidonic acid (LTC4, LTD4, LXA4, 5-oxo-ETE, PGD2 and TXA2) have been proposed to have a role in asthma but act on different combinations of cells types to produce different effects. Although antagonists against the cysLT1R are marketed therapies for asthma [12] and antagonists of the TXA2 response are marketed in Japan [46], selective antagonists of the OXE and CRTH2 receptors are not yet described. Without selective antagonists against all of the metabolites of arachidonic acid that have been proposed to have a role in asthma, a critical evaluation of the most important mediators cannot be accurately established. Concluding remarks In summary, 5-oxo-ETE is involved in several processes that are relevant to the inflammatory response in asthma, but many outstanding questions remain (Box 1). The recent molecular identification of a receptor that is activated by 5-oxo-ETE has provided a missing link to previous pharmacological studies looking at the responses of 5-oxo-ETE in human leukocytes. Studies to establish the role of 5-oxo-ETE and its receptor in inflammation have been limited by the lack of a potent antagonist of the OXE receptor. In addition, other metabolites of arachidonic acid have also been proposed to have a role in the inflammatory response in asthma, but without selective antagonists for each of the associated receptors, a critical evaluation of the relative contribution of each mediator is difficult. Indeed, targeting a single mediator, such as 5-oxo-ETE, might not be sufficient to block the inflammatory response in asthma, and dual antagonists of 5-oxoETE and, for example, the cysLT1 receptor might be required to see a significant effect. However, GPCRs have previously represented a successful class of drug targets and antagonists of lipid-binding GPCRs, such as the cysLT1 receptor, have proven successful targets. This would suggest that, at least technically, the development of 5-oxo-ETE-receptor antagonists is feasible. Further work to establish the precise role of the eosinophil in asthma is crucial to ascertain whether approaches to targeting the eosinophil in asthma will be prove to be successful therapies. Additionally, to fully understand and model the role of 5-oxo-ETE and its receptor in the Box 1. Outstanding questions 1. What is the contribution of 5-oxo-ETE to the asthmatic response relative to other arachidonic acid products, such as the cysteinyl leukotrienes, thromboxane A2 or prostaglandin D2? 2. Is 5-oxo-ETE produced in vivo and at what levels is the eicosanoid present in the airways? 3. Although much of the 5-lipoxygenase pathway for the conversion of arachidonic acid to the leukotrienes is conserved between humans and rodents, is the OXE receptor present in rodents? 4. Will antagonists of the OXE receptor have a significant therapeutic benefit over marketed therapies that target other parts of the 5-lipoxygenase pathway, such as the cysLT1-receptor antagonists? 5. Will targeting a single eosinophil chemoattractant, such a 5-oxoETE, be sufficient to block the eosinophilic inflammation observed in asthma? 6. What is the precise role of the eosinophil in the asthmatic response?
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asthmatic response and to assess the therapeutic potential of targeting the OXE receptor, the identification the rodent OXE receptor and the levels of 5-oxo-ETE in the airways remain important unaddressed issues. Acknowledgements Zarin Brown and Gino Van Heeke are thanked for their critical review of this manuscript.
References 1 The International Study of Asthma and Allergies in Childhood (ISAAC) Steering Committee. (1998) Worldwide variation in prevalence of symptoms of asthma, allergic rhinoconjunctivitis, and atopic eczema: ISAAC. Lancet 351, 1225–1232 2 Broide, D. and Sriramarao, P. (2001) Eosinophil trafficking to sites of allergic inflammation. Immunol. Rev. 179, 163–172 3 Kay, A.B. et al. (2004) A role for eosinophils in airway remodelling in asthma. Trends Immunol. 25, 477–482 4 Flood-Page, P.T. et al. (2003) Eosinophil’s role remains uncertain as anti-interleukin-5 only partially depletes numbers in asthmatic airway. Am. J. Respir. Crit. Care Med. 167, 199–204 5 Menzies-Gow, A. and Robinson, D.S. (2002) Eosinophils, eosinophilic cytokines (interleukin-5), and antieosinophilic therapy in asthma. Curr. Opin. Pulm. Med. 8, 33–38 6 Bandeira-Melo, C. and Weller, P.F. (2003) Eosinophils and cysteinyl leukotrienes. Prostaglandins Leukot. Essent. Fatty Acids 69, 135–143 7 Adamko, D.J. et al. (2005) The rise of the phoenix: the expanding role of the eosinophil in health and disease. Allergy 60, 13–22 8 Shi, H.Z. et al. (2000) Lymph node trafficking and antigen presentation by endobronchial eosinophils. J. Clin. Invest. 105, 945–953 9 Humbles, A.A. et al. (2004) A critical role for eosinophils in allergic airways remodeling. Science 305, 1776–1779 10 Holgate, S.T. et al. (2003) Roles of cysteinyl leukotrienes in airway inflammation, smooth muscle function, and remodelling. J. Allergy Clin. Immunol. 111, S18–S36 11 Teran, L.M. (2000) CCL chemokines and asthma. Immunol. Today 21, 235–242 12 Wenzel, S.E. (2003) The role of leukotrienes in asthma. Prostaglandins Leukot. Essent. Fatty Acids 69, 145–155 13 Brink, C. et al. (2004) International Union of Pharmacology XLIV. Nomenclature for the oxoeicosanoid receptor. Pharmacol. Rev. 56, 149–157 14 Powell, W.S. (2004) Biosynthesis and biological effects of 5-oxo-ETE and other oxoeicosatetraenoic acids. In The Eicosanoids (CurtisPrior, P., ed.), pp. 61–68, John-Wiley&Sons, Ltd 15 Powell, W.S. et al. (1994) Phorbol myristate acetate stimulates the formation of 5-oxo-6,8,11,14-eicosatetraenoic acid by human neutrophils by activating NADPH oxidase. J. Biol. Chem. 269, 25373–25380 16 Zhang, Y. et al. (1996) Synthesis of 5-oxo-6,8,11,14-eicosatetraenoic acid by human monocytes and lymphocytes. J. Leukoc. Biol. 59, 847–854 17 Powell, W.S. et al. (1995) 5-Oxo-6,8,11,14-eicosatetraenoic acid is a potent stimulator of human eosinophil migration. J. Immunol. 154, 4123–4132 18 Erlemann, K.R. et al. (2004) Oxidative stress stimulates the synthesis of the eosinophil chemoattractant 5-oxo-6,8,11,14-eicosatetraenoic acid by inflammatory cells. J. Biol. Chem. 279, 40376–40384 19 Zimpfer, U. et al. (2000) Human dendritic cells are a physiological source of the chemotactic arachidonic acid metabolite 5-oxo-eicosatetraenoic acid. Inflamm. Res. 49, 633–638 20 Jones, C.E. et al. (2003) Expression and characterization of a 5-oxo6E,8Z,11Z,14Z-eicosatetraenoic acid receptor highly expressed on human eosinophils and neutrophils. Mol. Pharmacol. 63, 471–477 21 Hosoi, T. et al. (2002) Identification of a novel human eicosanoid receptor coupled to Gi/o. J. Biol. Chem. 277, 31459–31465 22 Wise, A. et al. (2003) Molecular identification of high and low affinity receptors for nicotinic acid. J. Biol. Chem. 278, 9869–9874 23 Powell, W.S. et al. (1993) Stimulation of human neutrophils by 5-oxo6,8,11,14-eicosatetraenoic acid by a mechanism independent of the leukotriene B4 receptor. J. Biol. Chem. 268, 9280–9286
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24 Sozzani, S. et al. (1996) Stimulating properties of 5-oxo-eicosanoids for human monocytes: synergism with monocyte chemotactic protein-1 and -3. J. Immunol. 157, 4664–4671 25 O’Flaherty, J.T. et al. (2000) The coupling of 5-oxo-eicosanoid receptors to heterotrimeric G proteins. J. Immunol. 164, 3345–3352 26 Guilbert, M. et al. (1999) 5-Oxo-6,8,11,14-eicosatetraenoic acid induces important eosinophil transmigration through basement membrane components: comparison of normal and asthmatic eosinophils. Am. J. Respir. Cell Mol. Biol. 21, 97–104 27 Powell, W.S. et al. (2002) Interactions between 5-oxo-ETE and chemokines in stimulating eosinophils. Adv. Exp. Med. Biol. 507, 237–242 28 Powell, W.S. (2003) A novel PGD2 receptor expressed in eosinophils. Prostaglandins Leukot. Essent. Fatty Acids 69, 179–185 29 Powell, W.S. et al. (2001) Eotaxin and RANTES enhance 5-oxo6,8,11,14-eicosatetraenoic acid-induced eosinophil chemotaxis. J. Allergy Clin. Immunol. 107, 272–278 30 Powell, W.S. et al. (1999) 5-Oxo-6,8,11,14-eicosatetraenoic acid is a potent stimulator of L-selectin shedding, surface expression of CD11b, actin polymerization, and calcium mobilization in human eosinophils. Am. J. Respir. Cell Mol. Biol. 20, 163–170 31 O’Flaherty et al. (1996) 5-Oxo-eicosatetraenoate is a broadly active, eosinophil-selective stimulus for human granulocytes. J. Immunol. 157, 336–342 32 O’Flaherty et al. (1996) 5-Oxo-eicosanoids and hematopoietic cytokines cooperate in stimulating neutrophil function and the mitogenactivated protein kinase pathway. J. Biol. Chem. 271, 17821–17828 33 Powell, W.S. et al. (1997) Effects of 5-oxo-6,8,11,14-eicosatetraenoic acid on expression of CD11b, actin polymerization, and adherence in human neutrophils. J. Immunol. 159, 2952–2959 34 Powell, W.S. et al. (1996) Metabolism and biologic effects of 5-oxoeicosanoids on human neutrophils. J. Immunol. 156, 336–342 35 O’Flaherty, J.T. et al. (1998) Receptors for the 5-oxo class of eicosanoids in neutrophils. J. Biol. Chem. 273, 32535–32541 36 Stamatiou, P.B. et al. (2004) 5-Oxo-6,8,11,14-eicosatetraenoic acid stimulates the release of the eosinophil survival factor granulocyte/ macrophage colony-stimulating factor from monocytes. J. Biol. Chem. 279, 28159–28164 37 Mercier, F. et al. (2004) 5-oxo-ETE regulates tone of guinea pig airway smooth muscle via activation of Ca2C pools and Rho kinase pathway. Am. J. Physiol. Lung Cell Mol. Physiol. 287, L631–L640 38 Stamatiou, P. et al. (1998) 5-oxo-ETE induces pulmonary eosinophilia in an integrin-dependent manner in Brown Norway rats. J. Clin. Invest. 102, 2165–2172 39 Almishri, W. et al. (2004) Effects of prostaglandin D2, 15-deoxyD12,14-prostaglandin J2, and selective DP1 and DP2 receptor agonists on pulmonary infiltration of eosinophils in Brown Norway rats. J. Pharmacol. Exp. Ther. 313, 64–69 40 Muro, S. et al. (2003) 5-Oxo-6,8,11,14-eicosatetraenoic acid induces the infiltration of granulocytes into human skin. J. Allergy Clin. Immunol. 112, 768–774 41 Park, C.S. et al. (1998) Granulocyte macrophage colony-stimulating factor is the main cytokine enhancing survival of eosinophils in asthmatic airways. Eur. Respir. J. 12, 872–878 42 Powell, W.S. et al. (2001) Quantitative analysis of 5-oxo-6,8,11, 14-eicosatetraenoic acid by electrospray mass spectrometry using a deuterium-labeled internal standard. Anal. Biochem. 295, 262–266 43 Nicosia, S. et al. (2001) Leukotrienes as mediators of asthma. Pulm. Pharmacol. Ther. 14, 3–19 44 Norel, X. et al. (2004) The quest for new cysteinyl-leukotriene and lipoxin receptors: recent clues. Pharmacol. Ther. 103, 81–94 45 Levy et al. (2002) Multi-pronged inhibition of airway hyper-responsiveness and inflammation by lipoxin A4. Nat. Med. 8, 1018–1023 46 Dogne, J-M. et al. (2002) Therapeutic potential of thromboxane inhibitors in asthma. Expert Opin. Investig. Drugs 11, 275–281 47 Nagata, K. et al. (2003) The second PGD2 receptor CRTH2: structure, properties, and functions in leukocytes. Prostaglandins Leukot. Essent. Fatty Acids 69, 169–177 48 Czech, W. et al. (1997) Chemotactic 5-oxo-eicoatetraenoic acids induce oxygen radical production, Ca2C mobilisation and actin reorganisation in human eosinophils via a pertussis toxin-sensitive G-protein. J. Invest. Dermatol. 108, 108–112
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The OXE receptor: a new therapeutic approach for asthma? Carol E. Jones Respiratory Diseases Therapeutic Area, Novartis Institutes for Biomedical Research, Horsham, RH12 5AB, UK
The eicosanoid 5-oxo-6E,8Z,11Z,14Z-eicosatetraenoic acid (5-oxo-ETE) has recently been identified as the ligand for the oxoeicosanoid (OXE) receptor. In vitro and in vivo studies have suggested that 5-oxo-ETE has a role in the asthmatic inflammatory response and it has been shown to stimulate eosinophil migration to the airways. New data suggest that eosinophils have an important role in the pathogenesis of asthma, being required for mucus accumulation, airway hyperresponsiveness and remodelling of the airways. However, there are several mediators that can stimulate the recruitment of eosinophils to the airways and the development of antagonists against the OXE receptor is required to evaluate the potential of the OXE receptor as a new therapeutic approach for asthma.
Introduction Asthma is an inflammatory condition of the airways that is becoming increasingly prevalent, especially in children. Current estimates of prevalence are between 10 and 20% [1]. The disease is characterised by airway hyperresponsiveness, airway inflammation, excessive mucus production and structural changes to the airways (remodelling). The infiltration of leukocytes into lung tissue and airways is a key feature of asthma, with high numbers of eosinophils observed in the airways; the number of eosinophils correlates with disease severity [2]. The role of the eosinophil in asthma has been the subject of much debate. Although eosinophils in the airway are a characteristic feature of asthma, the precise role of the eosinophil in asthma pathophysiology remains controversial [3–5]. Early work in this area suggested that the eosinophil contributed to asthma pathogenesis through the release of granule-associated proteins, which cause tissue damage, lipid mediators, which cause bronchoconstriction, and the release of reactive oxygen species [5–7]. In addition, eosinophils can secrete Th2 cytokines and present antigens [8]. The role of the eosinophil in asthma became uncertain because clinical trials with anti-IL-5 antibodies failed to deliver the expected results. Although these antibodies were effective at depleting eosinophils, there was no significant clinical improvement [3,5]. However, recent in vivo data have suggested a role for the eosinophil in airway remodelling in asthma [9]. The targeted depletion of eosinophils using transgenic mice indicates Corresponding author: Jones, C.E. ([email protected]). Available online 10 May 2005
that eosinophils have a role in lung remodelling [9]. Eosinophils can also produce a range of molecules that are proposed to be involved in the remodelling process, providing further evidence suggesting a role for eosinophils in causing structural changes to the airways [3]. The accumulation of eosinophils into the airways is thought to be mediated by several factors. Following mobilisation from the bone marrow in response to cytokines, eosinophils migrate to the lung and other tissues in response to locally released chemoattractants. Certain lipid mediators, including products of arachidonic acid metabolism, and chemokines, such as eotaxin, are potent stimulators of this process [2,10,11]. An eosinophil chemoattractant that has recently attracted attention is 5-oxo-6E,8Z,11Z,14Z-eicosatetraenoic acid (5-oxo-ETE), a lipid that activates the oxoeicosanoid (OXE) receptor. With recent data shedding new insight into the role of the eosinophil in asthma pathogenesis, eosinophil chemoattractants should be revisited as potential targets for asthma. Here, 5-oxo-ETE and its recently identified receptor are discussed as a new therapeutic approach for asthma in light of recent developments in the understanding of the role of eosinophils in the disease process. The 5-LO pathway and 5-oxo-ETE Perhaps the most familiar products of the metabolism of arachidonic acid by the actions of 5-lipoxygenase (5-LO) are the cysteinyl leukotrienes (cysLTs), particularly leukotriene C4 (LTC4) and LTD4 (Figure 1). The cysLTs are potent mediators of contractile responses in the airways and antagonists of the cysLT1 receptor are marketed as therapies for asthma [10,12]. Another metabolite of arachidonic acid that is associated with the 5-LO pathway is 5-oxo-ETE. The major route of production of 5-oxo-ETE is believed to be from the precursor 5-hydroxy6,8,11,14 (E,Z,Z,Z)-eicosatetraenoic acid (5-HETE) by the actions of a specific dehydrogenase. The biosynthesis and metabolism of 5-oxo-ETE has recently been described [13,14]. Multiple cell types have been reported to produce 5-oxo-ETE, including monocytes, eosinophils and neutrophils, in response to nonphysiological stimuli, such as the calcium ionophore A23187 or the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA), but produce little 5-oxo-ETE without stimulation [15–17]. More recently, oxidative stress has been shown to stimulate 5-oxo-ETE synthesis by blood monocytes, lymphocytes and platelets but not neutrophils [18], which is of relevance to inflammatory respiratory diseases, such as
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Arachidonic acid
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Figure 1. Major pathways of arachidonic acid metabolism. Key products of arachidonic acid metabolism that are formed by the 5-LO, 12-LO, 15-LO, COX-1 and COX-2 pathways and are believed to have a role in the asthmatic response are shown (boxed). Metabolism of arachidonic acid on the 5-LO pathways results in the formation of the cysteinyl leukotrienes LTC4 and LTD4 in addition to 5-oxo-ETE. LXA4 is formed by the 5-LO, 12-LO and 15-LO pathways. The metabolism of arachidonic acid through the COX-1 and -2 pathways results in the formation of TXA2 and PGD2, which have been proposed to have a role in the asthmatic response. Abbreviations: COX, cyclooxygenase; 5-HETE, 5-hydroxy-6,8,11,14 (E,Z,Z,Z)-eicosatetraenoic acid; 5(S)-HPETE, 5(S)-hydroxyperoxy-6,8,11,14 (E,Z,Z,Z)-eicosatetraenoic acid; LO, lipoxygenase; LT, leukotriene; 5-oxo-ETE, 5-oxo6E,8Z,11Z,14Z-eicosatetraenoic acid; LX, lipoxin; PG, prostaglandin; TX, thromboxane.
asthma and chronic obstructive pulmonary diseases (COPDs). Most of the cell types described produce relatively small quantities of 5-oxo-ETE, which is in contrast to the higher levels reported from monocytederived dendritic cells [19].
Identification of the OXE receptor Although a receptor binding the lipid 5-oxo-ETE has been described in the literature for over ten years, its molecular identity remained unknown until recently. Two groups independently described a genome-datamining approach used for the molecular cloning of a G-protein coupled receptor (GPCR) activated by the ligand 5-oxo-ETE [20,21]. The two 5-oxo-ETE receptors identified, R527 [20]
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and TG1019 [21], are almost identical except for a 39 amino acid N-terminal extension in TG1019 and a single amino change towards the C-terminus. The receptor, recently renamed the OXE receptor by International Union of Pharmacology (IUPHAR) [13], is located on chromosome 2 and shows closest homology to the GPCR HM74, which has been reported to bind to nicotinic acid [22], and GPR81, for which the ligand has not yet been identified. The sequence identity at the amino acid level of the OXE receptor to HM74 and GPR81 is 40%. Two structurally related lipids, 5(S)-hydroxyperoxy6,8,11,14 (E,Z,Z,Z)-eicosatetraenoic acid [5(S)-HPETE] and 5(S)-HETE, were also found to be agonists of the OXE receptor, but are reported to be much less potent [20,21]. In primary cells, the OXE receptor has a restricted expression profile, with high levels of mRNA expression reported for eosinophils and neutrophils and lower levels in lung-derived macrophages [20]. In the same study, virtually no expression was detected in other cultured human cells, including lung epithelial cells, smooth muscle cells or lung fibroblasts. A more detailed study of the functions of the OXE receptor has been limited by the lack of good pharmacological tools. Four weak antagonists of the 5-oxo-ETE receptor have been described: 4Z,7Z,10Z,13Z,16Z,19Zdocosahexaenoic acid, 5Z,8Z,11Z,14Z,17Z-eicosapentaenoic acid, di-homo-g-linolenic acid and 11Z,14Z,17Zeicosatrienoic acid, with EC50 values of 1.6, 6.0, 3.7 and 5.1 mM, respectively, in a GTPgS assay [21]. The expression and pharmacological profiles of the OXE receptor suggest that it is the receptor described on human eosinophils, neutrophils and monocytes that is activated by 5-oxo-ETE, but not characterised at the molecular level [17,23,24]. The observed expression of the OXE receptor on human eosinophils, neutrophils and monocytes correlates with reported functional effects of 5-oxo-ETE on the same cell types. Many of the pharmacological responses reported for 5-oxo-ETE on human eosinophils and neutrophils, such as chemotaxis, are sensitive to pertussis toxin, indicating that the receptor is coupled to the G-protein Gai [25], which is also the case for the recombinant receptor [20]. E osinophil
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Figure 2. The actions of 5-oxo-ETE on primary cells. The eicosanoid 5-oxo-ETE has multiple actions on eosinophils, neutrophils and monocytes relevant to the inflammatory response in asthma. Examples of these effects of 5-oxo-ETE are given and include chemotaxis, calcium mobilisation, CD11b expression and degranulation [25–34,36,48]. www.sciencedirect.com
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5-oxo-ETE and inflammation Data from in vitro studies The eicosanoid 5-oxo-ETE has previously been suggested to have a role in the inflammatory response. A range of biological effects of 5-oxo-ETE that are relevant to the inflammatory response and asthma have been reported for eosinophils, neutrophils, monocytes and, more recently, for airway smooth muscle (Figure 2). These biological effects include cell migration, surface expression of the integrin CD11b, L-selectin shedding, calcium mobilisation, superoxide production and smooth muscle contraction. 5-oxo-ETE is a potent chemotactic factor, stimulating the migration of human eosinophils. A comparison of eosinophil chemoattractants indicates that 5-oxo-ETE is one of the most potent mediators known. Studies have shown that 5-oxo-ETE is more potent than plateletactivating factor (PAF) in eosinophil transmigration assays, inducing a slightly greater transmigration at 1 nM than does PAF at 1000 nM [26]. In fact, a low (1 nM) concentration of 5-oxo-ETE has been shown to potentiate the response of eosinophils to PAF in cellmigration assays [17], whereas the effects of 5-oxo-ETE and PAF were additive at a higher concentration (3 nM) of 5-oxo-ETE. Although reported to be w10-times less potent than eotaxin [27], in a comparison of a range of eosinophil chemoattractants, including PAF, eotaxin and prostaglandin D2 (PGD2), 5-oxo-ETE elicited the greatest chemotactic response [28]. 5-oxo-ETE also causes calcium mobilisation in eosinophils, a process that is thought to contribute to granulocyte migration, and EC50 values are reported to be in the range of 10 nM [29]. 5-oxo-ETE has also been demonstrated to cause CD11b expression and L-selectin shedding on human eosinophils, suggesting that it is important for the interaction of eosinophils with vascular endothelial cells and in transendothelial migration [30]. In addition, 5-oxo-ETE can stimulate the degranulation of eosinophils when the cells are primed with granulocyte macrophage-colony stimulating factor (GM-CSF), although it has little effect if the cells are not primed [31]. Studies have suggested that 5-oxo-ETE can synergise with other chemoattractants, such as eotaxin and RANTES (regulated on activation, normal T-cell expressed and secreted), in the eosinophilic chemotactic response [27,29]. This might be of relevance to the in vivo situations in which there are multiple mediators present, including the range of lipid mediators and chemokines that are released during pulmonary inflammation. Several similar responses to 5-oxo-ETE have been reported for human neutrophils, with effects on calcium mobilisation, cell migration, CD11b expression and degranulation after GM-CSF priming [25,32–34]. 5-oxoETE has been reported to bind to its receptor on human neutrophils with a Kd of 4 nM [35]. The chemotactic responses elicited by 5-oxo-ETE are less potent on neutrophils than those observed for eosinophils, paralleling the differences observed in relative receptor expression on these cell types [20]. Indeed, this eicosanoid is approximately five-times less potent at stimulating neutrophil than it is for eosinophil chemotaxis [17,23]. 5-oxo-ETE stimulates similar maximal responses in www.sciencedirect.com
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human neutrophils to the leukotriene LTB4, although LTB4 is a more potent mediator [14]. In contrast to the information available for the effects of 5-oxo-ETE on eosinophils and neutrophils, less is known about the effects on human monocytes. In human monocytes, 5-oxo-ETE induces a chemotactic response, although not as strong as that of monocyte-chemotactic protein-1 (MCP-1) or N-formyl-Met-Leu-Phe (fMLP) [24,14]. However, it was demonstrated that 5-oxo-ETE enhances the effects of MCP-1 in a chemotaxis assay. More recently, it has been shown that 5-oxo-ETE can stimulate the release of the eosinophil survival factor GM-CSF from human monocytes [36]. Effects of 5-oxo-ETE on airway smooth muscle have been reported, further supporting a potential link between 5-oxo-ETE and asthma. Guinea pig airway smooth muscle produced contractile responses to 5-oxo-ETE through the activation of the calcium pool and the responses were partially inhibited by a Rho-kinase pathway inhibitor [37]. However, the EC50 reported for 5-oxo-ETE in this system was 0.89 mM and therefore the physiological relevance is not clear. Although no detectable expression of the OXE receptor was reported in cultured human airway smooth muscle cells [20], it will be interesting to see if the contractile responses to 5-oxo-ETE extend to human tissue. Data from in vivo studies Data supporting a role for 5-oxo-ETE in the inflammatory response in vivo come from a rat model, in which 5-oxoETE delivered directly to the lung causes the recruitment of eosinophils to the airways [38,39]. Although a response was observed for 5-oxo-ETE in this rat model, a rodent homologue of the human OXE receptor has not yet been reported. In humans, 5-oxo-ETE has been recently reported to induce the infiltration of eosinophils, neutrophils and macrophages into the skin of healthy and asthmatic subjects [40]. No effects of 5-oxo-ETE on mast cells or lymphocytes were observed. Asthmatics were reported to be more sensitive to 5-oxo-ETE in this study, resulting in the infiltration of greater numbers of eosinophils into the skin [40]. In contrast to the results from the study in human skin, an in vitro study looking at the effects of 5-oxo-ETE on migration of human bloodderived eosinophils isolated from either asthmatic or healthy subjects showed no differences in cell migration between the two patient groups [26]. No reported studies have looked at differences in OXE-receptor expression between diseased and normal tissue. However, in a human skin model, 5-oxo-ETE caused a greater recruitment of eosinophils into the skin in subjects with asthma [40]. The OXE receptor – a promising asthma target? The eicosanoid 5-oxo-ETE has been reported to be involved in multiple processes associated with the inflammatory response. 5-oxo-ETE is produced under conditions of inflammatory stress [18], which commonly occur in respiratory conditions such as asthma and COPD. In vitro studies have demonstrated a role for 5-oxo-ETE in eosinophil and, to a lesser extent, neutrophil and monocyte cell migration [24–26]. In vivo data, both in rats and humans, support a role for 5-oxo-ETE in the recruitment
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of eosinophils to sites of inflammation [38,40]. Thus, these effects of 5-oxo-ETE on eosinophil migration suggest that it is involved in eosinophil-mediated allergic reactions. As previously described, when comparing the relative potencies of several mediators in the migration of eosinophils, whereas eotaxin is w10-fold more potent than is 5-oxoETE in its response [27], 5-oxo-ETE has been shown to cause the greatest maximal response [28]. Thus, the ability of 5-oxo-ETE to synergise with the chemokines RANTES and eotaxin in the eosinophil chemotactic response might be important at certain sites of inflammation. Other relevant reported effects of 5-oxo-ETE include cellular degranulation. Although 5-oxo-ETE alone has little effect on eosinophil and neutrophil degranulation, priming of the cells with GM-CSF enhances 5-oxo-ETE-induced degranulation of both cell types. In contrast to other chemotactic factors, such as eotaxin and PAF, 5-oxo-ETE can stimulate GM-CSF release from human monocytes and thereby not only stimulate the recruitment of eosinophils to the airways but also promote their survival [36]. GM-CSF itself causes degranulation and superoxide production and is an important survival factor once eosinophils reach the lung [41]. This is in contrast to the cytokine IL-5, which is important in the mobilisation of eosinophils from the bone marrow [2]. However, it is not known whether 5-oxo-ETE has a role in eosinophil mobilisation. Although the effects of 5-oxo-ETE on human neutrophils are less pronounced than those on eosinophils, a role for 5-oxo-ETE in persistent inflammation has been suggested in which neutrophils have become desensitized to LTB4 [14]. One concern when evaluating the role and relevance of 5-oxo-ETE in asthma is that there are currently no reports of 5-oxo-ETE measured in vivo either in human airways or in animals. However, this might reflect difficulties in accurately measuring 5-oxo-ETE. A sensitive quantitative method for the measurement of 5-oxo-ETE has recently been reported [42] and studies investigating levels of 5-oxo-ETE in asthmatic airway are awaited. Several other metabolites of arachidonic acid have been associated with asthma and these are illustrated in Figure 1. These include the leukotrienes LTC4 and LTD4 formed by the 5-LO pathway, which act at the cysLT1 receptor, are potent bronchoconstrictors and have effects on mucociliary clearance and eosinophilic inflammation [10,12,43]. Lipoxin A4 (LXA4) is another arachidonic acid metabolite formed by the actions of 15-LO and 12-LO [44] and stable analogues of LXA4 have been shown to have anti-inflammatory properties in mouse models of asthma [45]. A further proposed mediator of the asthmatic response is thromboxane A2 (TXA2), which is an arachidonic acid metabolite formed by the cyclooxygenase-2 pathway. TXA2 binds to the TXA2 (TP) receptor and causes bronchoconstriction of the airways [46]. More recently, the CRTH2 (chemoattractant-receptor-homologous molecule expressed on Th2 cells) receptor has been described, which binds to another arachidonic acid metabolite, PGD2. The actions of PGD2 acting through CRTH2 have been proposed to have an important role in allergic inflammation through effects on Th2 cells, eosinophils and www.sciencedirect.com
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basophils [47]. Indeed, taken together, the metabolites of arachidonic acid (LTC4, LTD4, LXA4, 5-oxo-ETE, PGD2 and TXA2) have been proposed to have a role in asthma but act on different combinations of cells types to produce different effects. Although antagonists against the cysLT1R are marketed therapies for asthma [12] and antagonists of the TXA2 response are marketed in Japan [46], selective antagonists of the OXE and CRTH2 receptors are not yet described. Without selective antagonists against all of the metabolites of arachidonic acid that have been proposed to have a role in asthma, a critical evaluation of the most important mediators cannot be accurately established. Concluding remarks In summary, 5-oxo-ETE is involved in several processes that are relevant to the inflammatory response in asthma, but many outstanding questions remain (Box 1). The recent molecular identification of a receptor that is activated by 5-oxo-ETE has provided a missing link to previous pharmacological studies looking at the responses of 5-oxo-ETE in human leukocytes. Studies to establish the role of 5-oxo-ETE and its receptor in inflammation have been limited by the lack of a potent antagonist of the OXE receptor. In addition, other metabolites of arachidonic acid have also been proposed to have a role in the inflammatory response in asthma, but without selective antagonists for each of the associated receptors, a critical evaluation of the relative contribution of each mediator is difficult. Indeed, targeting a single mediator, such as 5-oxo-ETE, might not be sufficient to block the inflammatory response in asthma, and dual antagonists of 5-oxoETE and, for example, the cysLT1 receptor might be required to see a significant effect. However, GPCRs have previously represented a successful class of drug targets and antagonists of lipid-binding GPCRs, such as the cysLT1 receptor, have proven successful targets. This would suggest that, at least technically, the development of 5-oxo-ETE-receptor antagonists is feasible. Further work to establish the precise role of the eosinophil in asthma is crucial to ascertain whether approaches to targeting the eosinophil in asthma will be prove to be successful therapies. Additionally, to fully understand and model the role of 5-oxo-ETE and its receptor in the Box 1. Outstanding questions 1. What is the contribution of 5-oxo-ETE to the asthmatic response relative to other arachidonic acid products, such as the cysteinyl leukotrienes, thromboxane A2 or prostaglandin D2? 2. Is 5-oxo-ETE produced in vivo and at what levels is the eicosanoid present in the airways? 3. Although much of the 5-lipoxygenase pathway for the conversion of arachidonic acid to the leukotrienes is conserved between humans and rodents, is the OXE receptor present in rodents? 4. Will antagonists of the OXE receptor have a significant therapeutic benefit over marketed therapies that target other parts of the 5-lipoxygenase pathway, such as the cysLT1-receptor antagonists? 5. Will targeting a single eosinophil chemoattractant, such a 5-oxoETE, be sufficient to block the eosinophilic inflammation observed in asthma? 6. What is the precise role of the eosinophil in the asthmatic response?
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asthmatic response and to assess the therapeutic potential of targeting the OXE receptor, the identification the rodent OXE receptor and the levels of 5-oxo-ETE in the airways remain important unaddressed issues. Acknowledgements Zarin Brown and Gino Van Heeke are thanked for their critical review of this manuscript.
References 1 The International Study of Asthma and Allergies in Childhood (ISAAC) Steering Committee. (1998) Worldwide variation in prevalence of symptoms of asthma, allergic rhinoconjunctivitis, and atopic eczema: ISAAC. Lancet 351, 1225–1232 2 Broide, D. and Sriramarao, P. (2001) Eosinophil trafficking to sites of allergic inflammation. Immunol. Rev. 179, 163–172 3 Kay, A.B. et al. (2004) A role for eosinophils in airway remodelling in asthma. Trends Immunol. 25, 477–482 4 Flood-Page, P.T. et al. (2003) Eosinophil’s role remains uncertain as anti-interleukin-5 only partially depletes numbers in asthmatic airway. Am. J. Respir. Crit. Care Med. 167, 199–204 5 Menzies-Gow, A. and Robinson, D.S. (2002) Eosinophils, eosinophilic cytokines (interleukin-5), and antieosinophilic therapy in asthma. Curr. Opin. Pulm. Med. 8, 33–38 6 Bandeira-Melo, C. and Weller, P.F. (2003) Eosinophils and cysteinyl leukotrienes. Prostaglandins Leukot. Essent. Fatty Acids 69, 135–143 7 Adamko, D.J. et al. (2005) The rise of the phoenix: the expanding role of the eosinophil in health and disease. Allergy 60, 13–22 8 Shi, H.Z. et al. (2000) Lymph node trafficking and antigen presentation by endobronchial eosinophils. J. Clin. Invest. 105, 945–953 9 Humbles, A.A. et al. (2004) A critical role for eosinophils in allergic airways remodeling. Science 305, 1776–1779 10 Holgate, S.T. et al. (2003) Roles of cysteinyl leukotrienes in airway inflammation, smooth muscle function, and remodelling. J. Allergy Clin. Immunol. 111, S18–S36 11 Teran, L.M. (2000) CCL chemokines and asthma. Immunol. Today 21, 235–242 12 Wenzel, S.E. (2003) The role of leukotrienes in asthma. Prostaglandins Leukot. Essent. Fatty Acids 69, 145–155 13 Brink, C. et al. (2004) International Union of Pharmacology XLIV. Nomenclature for the oxoeicosanoid receptor. Pharmacol. Rev. 56, 149–157 14 Powell, W.S. (2004) Biosynthesis and biological effects of 5-oxo-ETE and other oxoeicosatetraenoic acids. In The Eicosanoids (CurtisPrior, P., ed.), pp. 61–68, John-Wiley&Sons, Ltd 15 Powell, W.S. et al. (1994) Phorbol myristate acetate stimulates the formation of 5-oxo-6,8,11,14-eicosatetraenoic acid by human neutrophils by activating NADPH oxidase. J. Biol. Chem. 269, 25373–25380 16 Zhang, Y. et al. (1996) Synthesis of 5-oxo-6,8,11,14-eicosatetraenoic acid by human monocytes and lymphocytes. J. Leukoc. Biol. 59, 847–854 17 Powell, W.S. et al. (1995) 5-Oxo-6,8,11,14-eicosatetraenoic acid is a potent stimulator of human eosinophil migration. J. Immunol. 154, 4123–4132 18 Erlemann, K.R. et al. (2004) Oxidative stress stimulates the synthesis of the eosinophil chemoattractant 5-oxo-6,8,11,14-eicosatetraenoic acid by inflammatory cells. J. Biol. Chem. 279, 40376–40384 19 Zimpfer, U. et al. (2000) Human dendritic cells are a physiological source of the chemotactic arachidonic acid metabolite 5-oxo-eicosatetraenoic acid. Inflamm. Res. 49, 633–638 20 Jones, C.E. et al. (2003) Expression and characterization of a 5-oxo6E,8Z,11Z,14Z-eicosatetraenoic acid receptor highly expressed on human eosinophils and neutrophils. Mol. Pharmacol. 63, 471–477 21 Hosoi, T. et al. (2002) Identification of a novel human eicosanoid receptor coupled to Gi/o. J. Biol. Chem. 277, 31459–31465 22 Wise, A. et al. (2003) Molecular identification of high and low affinity receptors for nicotinic acid. J. Biol. Chem. 278, 9869–9874 23 Powell, W.S. et al. (1993) Stimulation of human neutrophils by 5-oxo6,8,11,14-eicosatetraenoic acid by a mechanism independent of the leukotriene B4 receptor. J. Biol. Chem. 268, 9280–9286
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24 Sozzani, S. et al. (1996) Stimulating properties of 5-oxo-eicosanoids for human monocytes: synergism with monocyte chemotactic protein-1 and -3. J. Immunol. 157, 4664–4671 25 O’Flaherty, J.T. et al. (2000) The coupling of 5-oxo-eicosanoid receptors to heterotrimeric G proteins. J. Immunol. 164, 3345–3352 26 Guilbert, M. et al. (1999) 5-Oxo-6,8,11,14-eicosatetraenoic acid induces important eosinophil transmigration through basement membrane components: comparison of normal and asthmatic eosinophils. Am. J. Respir. Cell Mol. Biol. 21, 97–104 27 Powell, W.S. et al. (2002) Interactions between 5-oxo-ETE and chemokines in stimulating eosinophils. Adv. Exp. Med. Biol. 507, 237–242 28 Powell, W.S. (2003) A novel PGD2 receptor expressed in eosinophils. Prostaglandins Leukot. Essent. Fatty Acids 69, 179–185 29 Powell, W.S. et al. (2001) Eotaxin and RANTES enhance 5-oxo6,8,11,14-eicosatetraenoic acid-induced eosinophil chemotaxis. J. Allergy Clin. Immunol. 107, 272–278 30 Powell, W.S. et al. (1999) 5-Oxo-6,8,11,14-eicosatetraenoic acid is a potent stimulator of L-selectin shedding, surface expression of CD11b, actin polymerization, and calcium mobilization in human eosinophils. Am. J. Respir. Cell Mol. Biol. 20, 163–170 31 O’Flaherty et al. (1996) 5-Oxo-eicosatetraenoate is a broadly active, eosinophil-selective stimulus for human granulocytes. J. Immunol. 157, 336–342 32 O’Flaherty et al. (1996) 5-Oxo-eicosanoids and hematopoietic cytokines cooperate in stimulating neutrophil function and the mitogenactivated protein kinase pathway. J. Biol. Chem. 271, 17821–17828 33 Powell, W.S. et al. (1997) Effects of 5-oxo-6,8,11,14-eicosatetraenoic acid on expression of CD11b, actin polymerization, and adherence in human neutrophils. J. Immunol. 159, 2952–2959 34 Powell, W.S. et al. (1996) Metabolism and biologic effects of 5-oxoeicosanoids on human neutrophils. J. Immunol. 156, 336–342 35 O’Flaherty, J.T. et al. (1998) Receptors for the 5-oxo class of eicosanoids in neutrophils. J. Biol. Chem. 273, 32535–32541 36 Stamatiou, P.B. et al. (2004) 5-Oxo-6,8,11,14-eicosatetraenoic acid stimulates the release of the eosinophil survival factor granulocyte/ macrophage colony-stimulating factor from monocytes. J. Biol. Chem. 279, 28159–28164 37 Mercier, F. et al. (2004) 5-oxo-ETE regulates tone of guinea pig airway smooth muscle via activation of Ca2C pools and Rho kinase pathway. Am. J. Physiol. Lung Cell Mol. Physiol. 287, L631–L640 38 Stamatiou, P. et al. (1998) 5-oxo-ETE induces pulmonary eosinophilia in an integrin-dependent manner in Brown Norway rats. J. Clin. Invest. 102, 2165–2172 39 Almishri, W. et al. (2004) Effects of prostaglandin D2, 15-deoxyD12,14-prostaglandin J2, and selective DP1 and DP2 receptor agonists on pulmonary infiltration of eosinophils in Brown Norway rats. J. Pharmacol. Exp. Ther. 313, 64–69 40 Muro, S. et al. (2003) 5-Oxo-6,8,11,14-eicosatetraenoic acid induces the infiltration of granulocytes into human skin. J. Allergy Clin. Immunol. 112, 768–774 41 Park, C.S. et al. (1998) Granulocyte macrophage colony-stimulating factor is the main cytokine enhancing survival of eosinophils in asthmatic airways. Eur. Respir. J. 12, 872–878 42 Powell, W.S. et al. (2001) Quantitative analysis of 5-oxo-6,8,11, 14-eicosatetraenoic acid by electrospray mass spectrometry using a deuterium-labeled internal standard. Anal. Biochem. 295, 262–266 43 Nicosia, S. et al. (2001) Leukotrienes as mediators of asthma. Pulm. Pharmacol. Ther. 14, 3–19 44 Norel, X. et al. (2004) The quest for new cysteinyl-leukotriene and lipoxin receptors: recent clues. Pharmacol. Ther. 103, 81–94 45 Levy et al. (2002) Multi-pronged inhibition of airway hyper-responsiveness and inflammation by lipoxin A4. Nat. Med. 8, 1018–1023 46 Dogne, J-M. et al. (2002) Therapeutic potential of thromboxane inhibitors in asthma. Expert Opin. Investig. Drugs 11, 275–281 47 Nagata, K. et al. (2003) The second PGD2 receptor CRTH2: structure, properties, and functions in leukocytes. Prostaglandins Leukot. Essent. Fatty Acids 69, 169–177 48 Czech, W. et al. (1997) Chemotactic 5-oxo-eicoatetraenoic acids induce oxygen radical production, Ca2C mobilisation and actin reorganisation in human eosinophils via a pertussis toxin-sensitive G-protein. J. Invest. Dermatol. 108, 108–112