Long-Acting Beta Agonists Enhance Allergic Airway Disease

<div><p>Asthma is one of the most common of medical illnesses and is treated in part by drugs that activate the beta-2-adrenoceptor (β₂-AR) to dilate obstructed airways. Such drugs include long acting beta agonists (LABAs) that are paradoxically linked to excess asthma-related mortality. Here we show that LABAs such as salmeterol and structurally related β₂-AR drugs such as formoterol and carvedilol, but not short-acting agonists (SABAs) such as albuterol, promote exaggerated asthma-like allergic airway disease and enhanced airway constriction in mice. We demonstrate that salmeterol aberrantly promotes activation of the allergic disease-related transcription factor signal transducer and activator of transcription 6 (STAT6) in multiple mouse and human cells. A novel inhibitor of STAT6, PM-242H, inhibited initiation of allergic disease induced by airway fungal challenge, reversed established allergic airway disease in mice, and blocked salmeterol-dependent enhanced allergic airway disease. Thus, structurally related β₂-AR ligands aberrantly activate STAT6 and promote allergic airway disease. This untoward pharmacological property likely explains adverse outcomes observed with LABAs, which may be overcome by agents that antagonize STAT6.</p></div

PM-242H inhibits development of allergic lung disease in Balb/c mice.

<p>Mice were treated every other day for two weeks with vehicle (DLPC) or one of two doses of PM-242H (242H; 5 mg or 50 μg i.n.) and challenged with <i>A</i>. <i>niger</i> conidia (AN) or PBS i.n. after which the allergic airway disease phenotype was assessed as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0142212#pone.0142212.g003" target="_blank">Fig 3</a>. (<b>a</b>) Airway responsiveness (*: P < 0.05 determined by ANOVA), (<b>b</b>) bronchoalveolar lavage fluid inflammatory cells, (<b>c</b>) total lung IL-4-secreting cells, and (<b>d</b>) fungal colony forming units (CFU) recovered from the lungs of infected mice are shown. *: P < 0.05 determined by Kruskal-Wallis test (n = 4 mice/treatment group). Data are from one of 4 independent and comparable biological experiments.</p

Long acting beta agonists promote allergic airway disease.

<p>(<b>a</b>) Wild type mice were challenged every other day intranasally (i.n.) over 10 days with the spores of <i>A</i>. <i>niger</i> (AN) and every day as indicated with the drugs salmeterol (SX), formoterol (FX), carvedilol (CV), albuterol (Alb) or salmeterol or the combination of salmeterol and fluticasone (SX/FT) and compared to mice challenged with vehicles (PBS and dilauroylphosphatidylcholine (DLPC) liposomes) and (<b>b, c</b>) the effect on airway hyperresponsiveness (AHR) was determined (all data from C57BL/6 mice). *: P < 0.05 determined by ANOVA. (<b>d</b>) Genotype matched wild type and β₂-AR- and ßarr2-deficient mice were challenged with salmeterol or DLPC alone i.n. and assessed for airway hyperresponsiveness. *: P < 0.05 determined by ANOVA. The effect of salmeterol on lung (<b>e</b>) IL-4- and (<b>f</b>) IL-17A-secreting cells, (<b>g</b>) total bronchoalveolar lavage fluid (BALF) inflammatory cells (eos: eosinophils; mac: macrophages; lym: lymphocytes; neut: neutrophils) and (<b>h</b>) airway goblet cell metaplasia was determined after <i>A</i>. <i>niger</i> challenge i.n. *: P < 0.05 determined by Mann Whitney test. Data are from one of 3 or more independent and comparable biological experiments with n = 4 mice/treatment group.</p

PM-242H reverses established airway hyperresponsiveness.

<p>(<b>a</b>) Mice were intranasally challenged with <i>A</i>. <i>niger</i> spores daily i.n. for two weeks after which airway responsiveness to acetylcholine was determined. PM-242H was then given i.n. every other day as spore challenges were continued. Airway responsiveness was determined 7 and 14 days after the initial determination and airway inflammation, lung cytokines and pathology were examined at the end of the experiment. (<b>b</b>) Airway responsiveness of vehicle (DLPC) and PM-242H treated animals. *: P < 0.05 determined by ANOVA. (<b>c</b>) Total BALF inflammatory cells. (<b>d, e</b>) Total lung IL-4 and IL-17A-secreting cells. (<b>f</b>) Total recovered lung fungal CFU. (<b>g</b>) Representative lung bronchovascular bundles depicting airway epithelial goblet cell metaplasia (periodic acid-Schiff stain; bar represents 100 μm). *: P < 0.05 determined by Mann-Whitney test (n = 4 or 5 mice/treatment group as indicated). Data are from one of 4 independent and comparable biological experiments.</p

PM-242H reverses salmeterol-dependent exaggerated allergic airway disease.

<p>(<b>a</b>) Mice were treated daily with DLPC, 50 μg of salmeterol (SX), or salmeterol and PM-242H (50 μg) i.n. and challenged intranasally every other day for 10 days with PBS or <i>A</i>. <i>niger</i> (AN) i.n. The effects of combined therapy on (<b>b</b>) airway hyperresponsiveness (*: P < 0.05 determined by ANOVA), (<b>c</b>) total bronchoalveolar lavage fluid (BALF) inflammatory cells (eos: eosinophils; mac: macrophages; lym: lymphocytes; neut: neutrophils), lung (<b>d</b>) IL-4- and (<b>e</b>) IL-17A-secreting cells, and (<b>f</b>) <i>Muc5AC</i> expression was determined. (<b>g</b>) Total fungal colony forming units (CFU) recovered from lungs of challenged mice. *: P < 0.05 determined by Kruskal-Wallis test (n = 4 or 5 mice/treatment group). Data are from one of 4 independent and comparable biological experiments.</p

PM242H inhibits development of allergic airway disease.

<p>(<b>a</b>) Mice (C57BL/6) were treated every other day for two weeks with vehicle (DLPC) or one of two doses of PM-242H (5 or 50 μg i.n.) while also challenged every other day with 400 × 10³<i>A</i>. <i>niger</i> conidia (AN) or PBS i.n. after which the allergic airway disease phenotype was assessed. (<b>b</b>) Airway resistance (*: P < 0.05 determined by ANOVA), (<b>c</b>) bronchoalveolar lavage fluid inflammatory cells, (<b>d</b>) total lung IL-4, (<b>e</b>) IL-17A, and (<b>f</b>) IFN-γ-secreting cells, and (<b>g</b>) goblet cell metaplasia from representative bronchovascular bundles are shown. *: P < 0.05 determined by Kruskal-Wallis test (n = 4 mice/treatment group). Data are from one of 4 independent and comparable biological experiments.</p