Pharmacogenetics of 2-Agonists

Transcription

1 Allergology International. 2011;60: DOI: allergolint.11-rai-0317 REVIEW ARTICLE Pharmacogenetics of 2-Agonists Nobuyuki Hizawa 1 ABSTRACT Short-acting β2-agonists (SABAs) and long-acting β2-agonists (LABAs) are both important for treatment of asthma and chronic obstructive pulmonary disease (COPD) because of their bronchodilator and bronchoprotective effects. However, the use of these agonists, at least for asthma, has generated some controversy because of their association with increased mortality. Pharmacogenetics is the study of genetically determined variation in response to medications, which might prove useful for target therapies in highly responsive patients, especially for more expensive therapies or those with increased risk of side effects. Variation in response to both SABAs and LABAs has been observed in patients with polymorphisms in the β2 adrenoceptor gene (ADRB2). This review summarizes results from various studies on the possible relationship between ADRB2 polymorphisms and the bronchodilator or bronchoprotective effects of inhaled β2-agonists. By assessing the ADRB2 genotype, the hope is that it will be possible to predict the responsiveness to chronic administration of β2-agonists. Genetic testing, however, is of limited usefulness at this stage for ADRB2 because the common variants identified thus far account for only a small proportion of the variation observed for given responses. Carefully performed and adequately powered clinical trials continue to be important for achieving the goal of pharmacogenetic approaches to therapy. KEY WORDS asthma, beta-2 adrenergic receptor, chronic obstructive pulmonary disease (COPD), genetic polymorphism INTRODUCTION Beta-adrenergic receptors are important drug targets in asthma and COPD. Inhaled β-receptor agonists remain among the mostly commonly prescribed medications to treat airflow obstruction in adults. However, persistent stimulation of β2-adrenergic receptor (β2ar) pathways may have deleterious consequences, particularly in patients with asthma. Recently, long-term use of β-agonists has been linked to an increased rate of deaths due to asthma. 1 Data from a large placebo-controlled US study (the SMART study 2 ;) showed a small but significant increase in asthma-related deaths in patients receiving salmeterol (13 deaths out of patients treated for 28 weeks) versus placebo (3 deaths out of patients). As a result, the US Food and Drug Administration has recently issued recommendations on how long-acting β2-agonists (LABAs) should be used to treat asthma, placing a safety warning on LABAs used alone or in combination with inhaled corticosteroids (ICS). 3 Pharmacogenetics could be a useful means of optimizing bronchodilator therapy in patients with asthma or COPD. Much of the interest in genetic control of the airway responses to β-agonists has focused on the ADRB2 gene; some ADRB2 polymorphisms result in changes in the amino acid sequence of the β2ar, leading to alterations of its properties, possibly representing a risk factor for adverse responses to β-agonist therapy. 4-6 This review discusses the current understanding of the association of ADRB2 polymorphisms with bronchodilator and bronchoprotective responses to chronic inhaled β2- agonists. In the future, translation of this knowledge into clinical guidelines for β2-agonist prescription may contribute to improvement in the efficacy and safety of treatment for asthma and COPD. ADVERSE EFFECTS OF 2-AGONISTS AND THEIR POTENTIAL MECHANISMS Despite the ability of β-agonists to cause immediate reversal of airway narrowing, concern remains that regular use of these drugs may be associated with adverse outcomes especially when used to treat patients with asthma. In animal models of asthma, administra- 1Department of Pulmonary Medicine, Institute of Clinical Medicine, University of Tsukuba, Tsukuba, Japan. Correspondence: Nobuyuki Hizawa, MD, Department of Pulmonary Medicine, Institute of Clinical Medicine, University of Tsukuba, Tennodai Tsukuba, Ibaraki , Japan. nhizawa@md.tsukuba.ac.jp Received 17 March Japanese Society of Allergology Allergology International Vol 60, No3,

2 Hizawa N tion of β-agonists induced airway morphologic changes and bronchial hyperresponsiveness. 7,8 In human studies, regularly scheduled use of inhaled β- agonists has resulted in loss of asthma control, declines in morning peak flow, longer durations of asthma exacerbations, rebound airway hyperresponsiveness (AHR), 9-15 and airway inflammation. 16,17 Epidemiological studies have demonstrated a positive correlation between the chronic use of short-acting β2-agonists and asthma mortality. 18,19 Recently, a large trial with the long-acting β2-agonist, salmeterol, was stopped because of increased death rates and serious asthma related events among the African American subgroup on long-term salmeterol. 2 The potential mechanisms for higher risk of death and life-threatening adverse effects are unclear but may include masking increasing inflammation and delaying awareness of worsening asthma. 20 The common explanation for these observations is that the adverse effects of regular β-agonist therapy are related to desensitization of the β2ar, 10,11,13,21-23 a characteristic of many membrane-associated receptors after high-dose or repeated exposure to agonists. Endogenous downregulation is an issue. 24 Paradoxically, for reasons determined by genotype, the acute response to pharmacologic concentrations of β2-agonists in vivo may be greatest in patients harboring receptors that are theoretically resistant to downregulation with exposure to physiologic concentrations of endogenous catecholamine. 25 This may explain why acute responses in vivo are greater in individuals homozygous for Arg16, 26,27 whereas the same genotype predisposes to adverse outcomes, possibly resulting from enhanced pharmacologically induced downregulation during long-term β-agonist exposure In this regard, proinflammatory cytokines in the asthmatic condition known to cause heterologous desensitization of the β2ar 31 might also be of importance. Alternatively, emerging data suggest proinflammatory effects associated with persistent β2ar activation resulting in increased airway inflammation and hyperresponsiveness, cardinal features of asthma. Studies using transgenic mice overexpressing airway smooth muscle β2ar revealed that persistent high-level activation of the β2ar leads to increased expression of phospholipase C-β in airway smooth muscle and may augment the effects of bronchoconstrictors such as acetylcholine, histamine, and leukotrienes. 32 Another study provided evidence that prolonged exposure of cultural human airway smooth muscle cells to β2- agonists auguments procontractile signaling pathways elicited by thrombin, bradykinin and histamine. This study indicated repetitive β2-agonist use lead to sensitization of excitation-contraction signaling pathways as a result of reduced expression of Regulator of G protein signaling 5 (RGS5), which is an inhibitor of G-protein-coupled receptor (GPCR) activity. 33 Thus, the β2ar regulates constrictive signals, affecting bronchomotor reactivity by means other than direct dilatation. Chronic administration of β-blockers also attenuated airway responsiveness and eosinophilic inflammation both in a murine model of asthma 34 and in patients with asthma, 35 suggesting that persistent activity of the airway β2ar negatively impacts increased airway inflammation and remodeling in asthma. An animal study also showed that β2ar signaling is required for development of mucin production, airway hyperresponsiveness, and inflammatory cellular infiltrates in a murine model of asthma. 36 Continuous use of β-agonists as asthma monotherapy also has the potential to increase type 2 cytokinemediated airway inflammation. 37,38 β2-agonists also significantly enhanced the cytokine-induced TSLP production by primary human lung tissue cells. 39 Interestingly, an increased risk of severe exacerbation of respiratory symptoms as well as death in some patients using LABAs has been reported only for the treatment of asthma but not for COPD. This may indicate that β2ar agonism on airway hyperresponsiveness and allergic inflammation rather than agonistinduced receptor downregulation represents a very probable mechanism of the adverse effects associated with long-term β2-agonists therapy. ADRB2 GENE POLYMORPHISMS The ADRB2 gene is a small intronless gene on chromosome 5q31-q32, 40 a region that is genetically linked to asthma and related phenotypes. 41,42 Reihaus et al. 43 originally described 9 coding polymorphisms in ADRB2, 4 of which (Gly16Arg, Gln27Glu, Val34 Met, and Thr164Ile) create nonsynonymous changes in the amino acid sequence. Drysdale et al. 4 described the molecular haplotypic structure of the ADRB2 gene in several ethnic groups; 13 polymorphisms were organized into 12 haplotypes out of the theoretically possible 8192 combinations, and deep divergence in the distribution of some haplotypes was noted in white, black American, Asian, and Hispanic-Latino groups with differences of more than 20-fold among the frequencies of the 4 major haplotypes. Recently, comprehensive resequencing of a 5.3-kb region of ADRB2 in 669 African American and white participants has identified 49 polymorphisms. These polymorphisms are in the regulatory regions, including the promoter region and the 3 -UTR. They contribute to haplotypic structure and genetic association with asthma-related phenotypes. 44 Screening of the ADRB2 gene in Japanese participants has also revealed 15 single nucleotide polymorphisms (SNPs) within the 3-kb promoter and 1.2-kb structural regions. 45 FUNCTIONAL RELEVANCE OF ADRB2 GENE POLYMORPHISMS Functional responses in recombinant cell systems and primary cultured airway cell lines have revealed 240 Allergology International Vol 60, No3,

3 Pharmacogenetics of β2-agonists altered downregulation profiles for the Arg16Gly and Gln27Glu variants of this receptor and altered coupling with the Thr164Ile variant In addition, several more polymorphisms have been identified in the ADRB2 5 promoter region, one of which is localized to the 5 leader cistron (5 LC), a signaling peptide involved in regulation of β2ar translation. 49 In recombinant cells, 5 LC-Arg19Cys (allelic frequency about 60% in white individuals) leads to higher β2ar expression 49 and β2ar promoter-driven luciferase activity 50 than in wild type cells. Studies of the genetic effects of the ADRB2 polymorphisms on agonistinduced β2ar desensitization in airway smooth muscle cells 48,51 and mast cells 52 in vitro andinlymphocytes ex vivo 53,54 have yielded conflicting results: Arg 16Gly and Gln27Glu polymorphisms have been associated with both decreased and increased agonistinduced desensitization of the β2ar-mediated cyclic adenosine monophosphate (camp) response 48,51 or with downregulation of the receptor. 48,52-54 Alterations in function due to β2ar allelic variations might result from a combination of polymorphisms rather than from a single polymorphism. 4 Linkage disequilibrium may account for conflicting results regarding the functional effects of these polymorphisms when considered in isolation. 48,51-54 Haplotype may predict the clinical phenotype better, particularly if each polymorphism included has a functional role and when 1 polymorphism operates in a direction opposite to the other. 55 Liggett et al. recently constructed 8 common haplotypes derived from 26 polymorphisms. Whole-gene transfection was performed with COS-7 cells and revealed 4 haplotypes with increased cell surface β2ar protein expression as compared with the others. Agonistpromoted downregulation of β2ar protein expression was also haplotype-dependent, and was found to be increased for 2 haplotypes. 56 ADRB2 GENE POLYMORPHISMS AND RE- SPONSES TO -AGONISTS A) ACUTE BRONCHODILATOR RESPONSES The clinical response to β-agonists in the treatment of asthma shows a high degree of interindividual variation that is not adequately explained by ethnicity, age, 27,57 or baseline lung function. 1,58 A recent analysis estimated that roughly 60% of the population variance in the forced expiratory volume in 1 second (FEV1) response to albuterol was attributable to genetic variations, 59 which might be caused in part by variations in the ADRB2 gene polymorphisms. Martinez et al. 26 found that children homozygous for Arg16 were 5.3 times more likely to show reversibility to albuterol than were those homozygous for Gly16. Heterozygous Gly16 Arg16 children showed intermediate responses to albuterol, whereas variations at the Gln27Glu polymorphism had no significant effect. A reduced response to β-agonist therapy in participants homozygous for Gly16 has been replicated in other studies. 53,60,61 We studied a Ban-I RFLP and subsequently demonstrated that it coincided with SNP +523 in 58 individuals from 4 Japanese families 62 ; this variant appeared to be associated with bronchodilator responsiveness. The increases in FEV1 after inhalation of procaterol hydrochloride were also measured in 81 Japanese patients with moderate to severe asthma (mean age of the participants was 54 years, and 47% were smokers). 63 Multivariate linear regression analysis showed that increased responsiveness to procaterol was correlated with the number of Arg16 alleles, indicating that the ADRB2 Arg16Gly polymorphism was associated with bronchodilator responsiveness to β2-agonists even in elderly asthmatic patients and smokers. Drysdale et al. reported an effect of haplotype pairs on acute bronchodilator responses to albuterol. 4 Recent reports on patients with asthma, however, reported no significant relationship between haplotype pairs and bronchodilator response. 44,64 The study by Choudhry et al. 61 provides insight into potential ethnic group differences in pharmacogenetic effects (Genetics of Asthma in Latino Americans [GALA] Study). The investigators found that in a group of 274 Mexican and 393 Puerto Rican families recruited on the basis of having a child with asthma, the Arg16 allele was significantly associated with increased bronchial reversibility in the combined population; however, when stratified by ancestral country of origin, the association remained significant only in the Puerto Rican group. The investigators found that bronchodilator responses were greater in the group with a specific haplotype. The Childhood Asthma Management Program (CAMP), another large family-based study consisting primarily of participants with mild asthma, 5 included genotype data on 8 ADRB2 SNPs in 700 complete trios (mother, father, and child with asthma). The same haplotype was associated with lower bronchodilator response measures in both the CAMP participants and the GALA Puerto Rican participants. These data, however, contrast with the report by Drysdale et al., 4 in which the same haplotype was associated with lower measures of bronchodilator drug responsiveness. Furthermore, in the GALA study, no association was observed between Arg16 Gly genotypes and drug responsiveness among Mexicans with asthma. These discrepancies may be partly explained by different linkage disequilibrium patterns in the different study groups or by the interaction of socioeconomic and other environmental factors with genetic background. B) REGULAR USES The genetic effects of the ADRB2 gene seem to depend on the duration of therapy, and the genetic effects of long-term use cannot be predicted from the acute effects. In an Asthma Clinical Research Net- Allergology International Vol 60, No3,

4 Hizawa N Mean morning PEF (L/min) p = 0.99 p = p < p < Placebo Salmeterol p = 0.87 p < Arg/Arg Gly/Gly Study group PC20 (geometric mean; mg/ml) Arg/Arg Gly/Gly Study group Placebo Salmeterol Fig. 1 Dissociation between bronchodilator and bronchoprotective effect of ICS/LABA. (Left) Morning PEF in patients with asthma while assigned to receive treatment with ICS plus LABA or placebo, by genotype. Adapted from Reference 69. (Right) Methacholine responsiveness in participants while assigned to receive treatment with inhaled corticosteroid plus salmeterol or placebo, by genotype. Adapted from Reference 69. work; National Heart, Lung, and Blood Institute (ACRN) study, 28 participants with asthma who were homozygous for Arg16 and treated with regular, short-acting β-agonists experienced significant declines in PEFR compared with Gly16 homozygous participants. Taylor et al. observed similar effects as well as increased exacerbations in Arg16 homozygous participants on regular albuterol treatment. However, they did not report changes in Arg16 Arg16 participants receiving intermittent albuterol or the long-acting β-agonist salmeterol. 29 The first prospective, randomized clinical trial examining pharmacogenetic responses in asthma confirmed the poorer response of Arg16 Arg16 individuals to regular shortacting β2-agonist therapy. 30 In contrast, Gly16 Gly16 participants receiving regular albuterol experienced improvement in peak flow rates and fewer daily asthma symptoms. In that study, 30 the adverse effects of regular β-agonist use could be observed even weeks after their withdrawal, at a time when β2ar desensitization should be resolved. If regular use of β- agonists negatively impacts the balance of factors contributing to airway inflammation or airway remodeling in asthma, it could have long-term consequences. This contention is supported by the fact that in studies of regular use of β-agonists adverse effects have been reported only in participants homozygous for Arg16, which is rather resistant to desensitization in vitro. In addition, a report on the responses to salmeterol in 2 ACRN trials found that Arg16 Arg16 participants had decreased responses to salmeterol in the presence or absence of concurrent ICS. 65 The Arg16Gly polymorphism is common in the Japanese population, and we retrospectively analyzed the outpatient records of asthmatic patients. 66 Twenty-seven patients with the Arg16 Arg16 genotype and 35 patients with the Gly16 Gly16 genotype regularly used β2- agonists, whereas 37 patients with Arg16 Arg16 genotype and 29 patients with Gly16 Gly16 genotype used them as needed. During follow-up periods of at least 6 months at Hokkaido University Hospital, longterm bronchodilator responses were assessed using 3 indices: (a) improvement in FEV1 (DeltaFEV1 [ml]), (b) DeltaFEV1 FEV1 at the initial visit, and (c) DeltaFEV1 predicted FEV1. In patients with the Gly16 Gly16 genotype, those who regularly used β2- agonists had greater improvement in FEV1 in every index (p = ) than did those who used them as needed. In contrast, in patients with the Arg16 Arg16 genotype, regular usage of β2-agonists did not yield greater improvement in FEV1 when compared with as-needed β2-agonist usage. This finding suggests that Gly16 Gly16 and Arg16 Arg16 genotype responses to regular usage of β2-agonists may differ in Japanese patients with asthma. The relevance of ADRB2 haplotypes, rather than single polymorphisms, was not assessed in this study. In contrast, in a small retrospective analysis of clinical trial data, no genotype-related adverse effects were identified. 67 In a larger retrospective study 68 using 2650 participants with moderate asthma, of whom 430 had the Arg16 genotype, no association between treatment with either salmeterol or formoterol and clinical outcomes was identified after stratification by ADRB2 genotype or relevant haplotypes. However, the patients included in both studies were preselected to benefit from a long-acting bronchodilator in terms of baseline reversibility, thus perhaps exaggerating the effect of the long-acting β-agonist in these participants. In a prospective study in a genotype-stratified population in which salmeterol was added to inhaled corticosteroid therapy, mild-to-moderate asthmatic patients were stratified by Arg16Gly genotypes. The improvement in morning PEFR with salmeterol therapy was similar for Arg16 and Gly16 homozygous patients. Half of the Arg16 Arg16 patients, however, ex- 242 Allergology International Vol 60, No3,

5 Pharmacogenetics of β2-agonists p = p = 0.57 p = Placebo Salmeterol ΔFEV1 (L) p = 0.44 ΔFEV1 (L) p = Mean PEF (L/min) As needed Regular [N = 37] [N = 27] Arg/Arg 0.0 As needed Regular [N = 29] [N = 35] Gly/Gly 0 Arg/Arg Study group Gly/Gly Fig. 2 Influence of the ADRB2 genotype on bronchodilator responses to ICS/LABA both in Japanese and African American populations. (Left) Genetic infl uence of the Arg16Gly polymorphism on ΔFEV1 according to regular usage of β2-agonists in Japanese patients with asthma. Signifi cance of was shown for the interaction between genotypes and regular use of β2 agonist. Adapted from Reference 66. (Right) Morning PEF in African-American patients with asthma while assigned to receive treatment with ICS plus LABA or placebo, by genotype. Adapted from Reference 69. perienced worsening of methacholine responsiveness despite improvement in their airway function by addition of LABA to ICS (Fig. 1). This finding clearly indicated the dissociation between the bronchodilator and bronchoprotective effects of LABA, which may underlie the significant deterioration in asthma control caused by long-term usage of LABA. In the post hoc analysis, African American individuals with the Arg16 Arg16 genotype did not benefit from the addition of LABA to ICS, which was a finding identical to that of our previous report 66 (Fig. 2). Given that ADRB2 allele frequencies and genotype distributions in Japanese participants were similar to those of African Americans, 4 the LARGE trial may suggest that patients with Arg16 Arg16 are susceptible to an increased risk of serious asthma outcomes owing to long term use of LABA. LABA therapy might promote a high level of current control without improving stability and the risk of exacerbations in patients with particular genotypes. Bleecker and associates 70 recently demonstrated that asthmatic patients stratified by Arg16Gly genotype showed similar clinical responses to salmeterol alone or in combination with fluticasone. Meanwhile, they confirmed that the use of salmeterol significantly increases the risk of loss of asthma control, as suggested by the observation that 10 of 272 patients receiving salmeterol alone had an exacerbation, compared with only 1 of 272 patients receiving the ICS LABA combination, which was statistically significant. Furthermore, 1 asthma-related death occurred during the study in a patient receiving salmeterol alone. Bleecker et al. also found that patients homozygous for Arg16 had a significantly higher number of disease exacerbations during the run-in phases of the trial. C) COPD Chronic obstructive pulmonary disease is characterized by a persistent airflow limitation that is not fully reversible; however, reversibility of airflow limitations in response to a bronchodilator is an important component of COPD. The degree of reversibility of airflow obstruction shown by patients with COPD varies considerably. 71 In a recent study of 274 patients with COPD, 72 we showed that bronchodilator response to salbutamol therapy was associated with a continuous phenotype in patients, with a wide interindividual variation. We have reported preliminary evidence, in patients with COPD, that the Arg16Gly polymorphism has a significant physiologic role in regulating responses to β2-agonists. 73 By studying 246 patients with COPD who were participants in a longitudinal study of COPD (Hokkaido COPD cohort study), we compared short-term bronchodilator responses to salbutamol according to ADRB2 genotypes at codons 16 and 27. The presence of the Arg16 allele was associated with lower bronchodilator responses to β2- agonist inhalation. Log values (postbronchodilator FEV1-prebronchodilator FEV1) of individuals homozygous for Gly16 (n = 65), heterozygous for Arg16 Gly16 (n = 106), and homozygous for Arg16 (n = 75) Allergology International Vol 60, No3,

6 Hizawa N were2.19±0.43(mean±sd),2.09±0.42,and2.01± 0.42, respectively (p < 0.05). The genetic effects of the Arg16Gly polymorphism were independent of the severity of airflow limitation, age, and smoking status. The most common Arg16-Gln27 haplotype was also significantly associated with decreased bronchodilator responses to salbutamol (p < 0.01). Our finding in patients with COPD is in apparent contradiction with previous findings 26,63 showing that asthmatic patients carrying Arg16 had significantly greater β2-agonist responsiveness. It is possible that the mechanisms by which polymorphisms in the ADRB2 gene determine responses to β2-agonists in patients with asthma are different from those among patients with COPD, at least in part because asthma and COPD have mechanistic difference in their pathobiology, including different genetic and environmental backgrounds. CONCLUSIONS Although β2-agonists are effective and safe in most patients with asthma and COPD, they may be less effective or potentially harmful in some. Pharmacogenetic approaches hold the promise of matching individualized treatments to specific genotypes in a way that minimizes side effects while improving therapeutic outcomes. From the research reported to date, however, no consensus has been reached on the relationship between identified ADRB2 genetic variations and airway responses to β2-agonists, including bronchodilation and bronchoprotective effects. Interpretation of the findings of genetic association studies of the ADRB2 polymorphisms may be complicated by inadequate measurement of environmental exposures such as smoking and by differences in allele and haplotype frequencies of the ADRB2 gene. The complex genotype-response effects observed limit their clinical applications. Future large prospective studies should take account of ADRB2 haplotypic variations, possible gene-gene interactions, as well as geneenvironment interactions. More insight into how alterations in the expression or function of the gene affect clinical phenotypes is necessary to help us translate ADRB2 genetic information into application in clinical practice. ACKNOWLEDGEMENTS I m very grateful to Mrs. Flaminia Miyamasu, an associate professor at Institute of Clinical Medicine, University of Tsukuba, for proofreading this paper and for her comments. REFERENCES 1. Salpeter SR, Buckley NS, Ormiston TM, Salpeter EE. Meta-analysis: effect of long-acting beta-agonists on severe asthma exacerbations and asthma-related deaths. AnnInternMed2006;144: Nelson HS, Weiss ST, Bleecker ER, Yancey SW, Dorinsky PM; SMART Study Group. The salmeterol multicenter asthma research trial (SMART): a comparison of usual pharmacotherapy for asthma or usual pharmacotherapy plus salmeterol. 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