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Budesonide prevents Cytokine-induced Decrease of the Relaxant Response to β2-agonists in Mouse Trachea
S92 Abstracts
Budesonide prevents Cytokine-induced Decrease of the Relaxant Response to 2-agonists in Mouse Trachea M. Adner1, L. Cardell1, A. Miller-Larsson2; 1ENT, Lab Clin Exp Allergy Research, Malmö University Hospital, Malmö, SWEDEN, 2Medical Science, AstraZeneca R&D Lund, Lund, SWEDEN. RATIONALE: During asthma exacerbation periods, increased levels of pro-inflammatory cytokines in asthmatic airways may impair bronchodilating effects of 2-adrenoceptors agonists but this process may be counteracted by corticosteroid-treatment. The aim of this study was to investigate how the relaxant effects of 2-agonists in cultured mouse tracheal segments are affected by pro-inflammatory cytokines and concomitant treatment with corticosteroid. METHODS: BALB/cJ mice tracheal segments were cultured for 4 days in absence/presence of TNF (100ng/mL)+IL1 (10ng/mL) and a glucocorticoid, budesonide (1M). After the culture period, segments were moved to a myograph and pre-contracted with carbachol. Cumulative concentration-effect curves to formoterol, terbutalin and salmeterol were obtained with estimates of potency (logEC50) and maximal relaxation (Rmax; % decrease of pre-contraction). RESULTS: Formoterol relaxed tracheal segments with a higher potency and efficacy than both terbutalin and salmeterol; logEC50= -8.25±0.22, -6.15±0.23 and -7.60±0.17, and Rmax = 95.0±1.3%, 88.0±2.0% and 79.3±5.6% for formoterol, terbutalin and salmeterol, respectively (mean±SEM). Treatment with TNF+IL1 did not affect potency for any of the relaxant agonists but decreased Rmax by 14% for formoterol, by 33% for terbutalin and by 58% for salmeterol (p<0.05 versus formoterol). Budesonide completely prevented the cytokine-induced impairment of response to formoterol, terbutalin and salmeterol. CONCLUSIONS: Formoterol was more efficacious than both terbutalin and salmeterol to induce relaxation of trachea, especially in the presence of TNF+IL1. The decrease of the relaxant response to 2-agonists induced by cytokines was prevented by budesonide. These results highlight the value of combination treatment with budesonide and formoterol in maintaining optimal 2-adrenoceptor efficacy in periods with increased inflammation. Funding: AstraZeneca
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SUNDAY
Inhaled Corticosteroids and the Risk of Oropharyngeal Adverse Events: Results from a Meta-Analysis G. S. Rachelefsky1, H. Li2, N. Liao2, Y. Liao2, N. J. Combates2, R. Faruqi2, S. T. Varghese2; 1Allergy Research Foundation, Inc, Los Angeles, CA, 2Aventis Pharmaceuticals, a member of the sanofi-aventis Group, Bridgewater, NJ. RATIONALE: Oropharyngeal adverse events (AEs) are common among inhaled corticosteroid (ICS) users; however, the extent of their incidence is not clear. METHODS: A computerized MEDLINE (January 1966 to June 2004) and EMBASE (January 1974 to June 2004) search was conducted using indexed MedDRA terms for AEs and ICS available in 2004 (English only). Only studies emphasizing single entity or combination ICS for adolescents and adults with asthma were included. Studies were characterized by robust study designs that were randomized, double-blinded and placebo-controlled with clearly delineated study objectives, treatments and diagnostic criteria. Odds ratios (OR) were used to determine the AE incidence rate for ICS according to dose, device and propellant. RESULTS: A total of 23 studies (59 ICS arms and 23 placebo arms) were evaluated. The incidences of ICS-induced oral candidiasis (P0.0006), dysphonia (P0.0007) and pharyngitis (P0.0228) increased significantly with dose versus placebo (at all dose levels and combined, regardless of actuator device). For all ICS, doses and devices combined, ICS had a greater risk of oral candidiasis (OR=3.6), dysphonia (OR=5.2) and pharyngitis (OR=2.2) versus placebo (P<0.0001 for each event). When analyzing by device used, budesonide (BUD) metered dose inhaler (MDI; OR=19.29) and fluticasone propionate (FP) dry powder inhaler (DPI; OR=4.73) posed the greatest risk of oral candidiasis at all doses combined
358
J ALLERGY CLIN IMMUNOL FEBRUARY 2006
versus placebo. Likewise, BUD MDI (OR=11.45) and FP DPI (OR=5.70) had the greatest risk of dysphonia versus placebo, while BUD MDI (OR=5.08) and BUD DPI (OR=5.11) had the greatest risk of pharyngitis. CONCLUSIONS: Older ICS present a significant risk of oropharyngeal AEs. Funding: Aventis Pharmaceuticals, a member of the sanofi-aventis Group, and ALTANA Pharma Genome-Scale Analysis of Posttranscriptional Effects of Glucocorticoids (GC) J. Fan1, S. L. Curry1, U. X. Atasoy2, K. G. Becker3, M. Gorospe3, C. Stellato1; 1Division of Allergy and Clinical Immunology, Johns Hopkins University School of Medicine, Baltimore, MD, 2University of Missouri-Columbia, Columbia, MO, 3National Institute of Aging, NIH, Baltimore, MD. RATIONALE: Modulation of gene expression by GC is increasingly recognized to occur through post-transcriptional regulation (PTR). However, the impact of PTR in GC anti-inflammatory activity is poorly understood. We aim to define the proportion and nature of the genes regulated by GCs mainly through PTR. METHODS: We compared GC effects on the expression of total RNA versus newly transcribed RNAs isolated in parallel samples using a modified nuclear run-on protocol. RNA pools were identified by cDNA arrays of the human airway epithelial cell line BEAS-2B, challenged or not with TNF alpha, following treatment with the GC budesonide (BUD). We used an established scoring system to calculate the ratio between fold-changes (expressed as z-ratio) induced by GCs in the levels of total and newly generated transcripts in untreated and TNFalpha-stimulated cells. By revealing genes whose transcription is only partially affected, or uninfluenced, by GC despite parallel changes in steady-state RNA levels, this method uncovers the involvement of PTR. Genes were classified as: posttranscriptionally upregulated (mRNA-stability score (SS) > 1.0), posttranscriptionally downregulated (mRNA-SS <-1.0) or transcriptionally regulated (mRNA-SS between 1 and -1). RESULTS: Transcriptional regulation accounted for ~ 50% of GC-mediated changes in total mRNA levels. Among the genes affected by GC predominantly by PTR, we identified the chemokine CCL-2/MCP-1, which is both highly inducible by TNFalpha and strongly inhibited by GC through PTR. Validation experiments are ongoing. CONCLUSIONS: Changes in PTR account for a significant proportion of the effect of GCs on gene expression. Identification of the pathways involved could uncover novel targets of anti-inflammatory therapy. Funding: NIH
359
Control of Airway Inflammation in Asthma Patients Receiving Fluticasone Propionate (FP)/Salmeterol, FP Alone or FP Plus Montelukast N. Jarjour1, P. Dorinsky2, J. Stauffer2, D. Reilly2, S. Yancey2, L. Edwards2, L. Sutton2; 1University of Wisconsin School of Medicine, Madison, WI, 2GlaxoSmithKline, Research Triangle Park, NC. RATIONALE: To evaluate asthma control and control of airway inflammation after various treatments in patients symptomatic on ICS or shortacting beta2-agonists. METHODS: We combined data from two randomized, double-blind studies evaluating: (Study 1) FP/salmeterol (FSC) 100/50mcgBID (n=40) vs. FP 250mcgBID (n=48) or (Study 2) FP 100mcgBID vs. FP 100mcgBID (n=53) + montelukast (MON) 10mgQD (n=50). Subjects underwent biopsy/BAL prior to randomization and after 24 weeks of treatment (Study 1) or 12 weeks of treatment (Study 2). RESULTS: Measures of clinical efficacy were maintained or improved in subjects receiving FSC vs. FP250, whereas there were no improvements in clinical efficacy with FP+MON vs. FP100. Consistent with these clinical observations, no significant differences in bronchial mucosal eosinophils were found at 24 weeks for FSC vs. FP250 (95% CI: -0.64, 1.67) or at 12 weeks for FP100 vs. FP100+MON (95% CI: -6.61, 8.82) Similar results were noted for neutrophils, CD3+, CD4+, CD8+ and CD25+ lymphocytes in biopsies as well as BAL percentages of
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Budesonide prevents Cytokine-induced Decrease of the Relaxant Response to 2-agonists in Mouse Trachea M. Adner1, L. Cardell1, A. Miller-Larsson2; 1ENT, Lab Clin Exp Allergy Research, Malmö University Hospital, Malmö, SWEDEN, 2Medical Science, AstraZeneca R&D Lund, Lund, SWEDEN. RATIONALE: During asthma exacerbation periods, increased levels of pro-inflammatory cytokines in asthmatic airways may impair bronchodilating effects of 2-adrenoceptors agonists but this process may be counteracted by corticosteroid-treatment. The aim of this study was to investigate how the relaxant effects of 2-agonists in cultured mouse tracheal segments are affected by pro-inflammatory cytokines and concomitant treatment with corticosteroid. METHODS: BALB/cJ mice tracheal segments were cultured for 4 days in absence/presence of TNF (100ng/mL)+IL1 (10ng/mL) and a glucocorticoid, budesonide (1M). After the culture period, segments were moved to a myograph and pre-contracted with carbachol. Cumulative concentration-effect curves to formoterol, terbutalin and salmeterol were obtained with estimates of potency (logEC50) and maximal relaxation (Rmax; % decrease of pre-contraction). RESULTS: Formoterol relaxed tracheal segments with a higher potency and efficacy than both terbutalin and salmeterol; logEC50= -8.25±0.22, -6.15±0.23 and -7.60±0.17, and Rmax = 95.0±1.3%, 88.0±2.0% and 79.3±5.6% for formoterol, terbutalin and salmeterol, respectively (mean±SEM). Treatment with TNF+IL1 did not affect potency for any of the relaxant agonists but decreased Rmax by 14% for formoterol, by 33% for terbutalin and by 58% for salmeterol (p<0.05 versus formoterol). Budesonide completely prevented the cytokine-induced impairment of response to formoterol, terbutalin and salmeterol. CONCLUSIONS: Formoterol was more efficacious than both terbutalin and salmeterol to induce relaxation of trachea, especially in the presence of TNF+IL1. The decrease of the relaxant response to 2-agonists induced by cytokines was prevented by budesonide. These results highlight the value of combination treatment with budesonide and formoterol in maintaining optimal 2-adrenoceptor efficacy in periods with increased inflammation. Funding: AstraZeneca
357
SUNDAY
Inhaled Corticosteroids and the Risk of Oropharyngeal Adverse Events: Results from a Meta-Analysis G. S. Rachelefsky1, H. Li2, N. Liao2, Y. Liao2, N. J. Combates2, R. Faruqi2, S. T. Varghese2; 1Allergy Research Foundation, Inc, Los Angeles, CA, 2Aventis Pharmaceuticals, a member of the sanofi-aventis Group, Bridgewater, NJ. RATIONALE: Oropharyngeal adverse events (AEs) are common among inhaled corticosteroid (ICS) users; however, the extent of their incidence is not clear. METHODS: A computerized MEDLINE (January 1966 to June 2004) and EMBASE (January 1974 to June 2004) search was conducted using indexed MedDRA terms for AEs and ICS available in 2004 (English only). Only studies emphasizing single entity or combination ICS for adolescents and adults with asthma were included. Studies were characterized by robust study designs that were randomized, double-blinded and placebo-controlled with clearly delineated study objectives, treatments and diagnostic criteria. Odds ratios (OR) were used to determine the AE incidence rate for ICS according to dose, device and propellant. RESULTS: A total of 23 studies (59 ICS arms and 23 placebo arms) were evaluated. The incidences of ICS-induced oral candidiasis (P0.0006), dysphonia (P0.0007) and pharyngitis (P0.0228) increased significantly with dose versus placebo (at all dose levels and combined, regardless of actuator device). For all ICS, doses and devices combined, ICS had a greater risk of oral candidiasis (OR=3.6), dysphonia (OR=5.2) and pharyngitis (OR=2.2) versus placebo (P<0.0001 for each event). When analyzing by device used, budesonide (BUD) metered dose inhaler (MDI; OR=19.29) and fluticasone propionate (FP) dry powder inhaler (DPI; OR=4.73) posed the greatest risk of oral candidiasis at all doses combined
358
J ALLERGY CLIN IMMUNOL FEBRUARY 2006
versus placebo. Likewise, BUD MDI (OR=11.45) and FP DPI (OR=5.70) had the greatest risk of dysphonia versus placebo, while BUD MDI (OR=5.08) and BUD DPI (OR=5.11) had the greatest risk of pharyngitis. CONCLUSIONS: Older ICS present a significant risk of oropharyngeal AEs. Funding: Aventis Pharmaceuticals, a member of the sanofi-aventis Group, and ALTANA Pharma Genome-Scale Analysis of Posttranscriptional Effects of Glucocorticoids (GC) J. Fan1, S. L. Curry1, U. X. Atasoy2, K. G. Becker3, M. Gorospe3, C. Stellato1; 1Division of Allergy and Clinical Immunology, Johns Hopkins University School of Medicine, Baltimore, MD, 2University of Missouri-Columbia, Columbia, MO, 3National Institute of Aging, NIH, Baltimore, MD. RATIONALE: Modulation of gene expression by GC is increasingly recognized to occur through post-transcriptional regulation (PTR). However, the impact of PTR in GC anti-inflammatory activity is poorly understood. We aim to define the proportion and nature of the genes regulated by GCs mainly through PTR. METHODS: We compared GC effects on the expression of total RNA versus newly transcribed RNAs isolated in parallel samples using a modified nuclear run-on protocol. RNA pools were identified by cDNA arrays of the human airway epithelial cell line BEAS-2B, challenged or not with TNF alpha, following treatment with the GC budesonide (BUD). We used an established scoring system to calculate the ratio between fold-changes (expressed as z-ratio) induced by GCs in the levels of total and newly generated transcripts in untreated and TNFalpha-stimulated cells. By revealing genes whose transcription is only partially affected, or uninfluenced, by GC despite parallel changes in steady-state RNA levels, this method uncovers the involvement of PTR. Genes were classified as: posttranscriptionally upregulated (mRNA-stability score (SS) > 1.0), posttranscriptionally downregulated (mRNA-SS <-1.0) or transcriptionally regulated (mRNA-SS between 1 and -1). RESULTS: Transcriptional regulation accounted for ~ 50% of GC-mediated changes in total mRNA levels. Among the genes affected by GC predominantly by PTR, we identified the chemokine CCL-2/MCP-1, which is both highly inducible by TNFalpha and strongly inhibited by GC through PTR. Validation experiments are ongoing. CONCLUSIONS: Changes in PTR account for a significant proportion of the effect of GCs on gene expression. Identification of the pathways involved could uncover novel targets of anti-inflammatory therapy. Funding: NIH
359
Control of Airway Inflammation in Asthma Patients Receiving Fluticasone Propionate (FP)/Salmeterol, FP Alone or FP Plus Montelukast N. Jarjour1, P. Dorinsky2, J. Stauffer2, D. Reilly2, S. Yancey2, L. Edwards2, L. Sutton2; 1University of Wisconsin School of Medicine, Madison, WI, 2GlaxoSmithKline, Research Triangle Park, NC. RATIONALE: To evaluate asthma control and control of airway inflammation after various treatments in patients symptomatic on ICS or shortacting beta2-agonists. METHODS: We combined data from two randomized, double-blind studies evaluating: (Study 1) FP/salmeterol (FSC) 100/50mcgBID (n=40) vs. FP 250mcgBID (n=48) or (Study 2) FP 100mcgBID vs. FP 100mcgBID (n=53) + montelukast (MON) 10mgQD (n=50). Subjects underwent biopsy/BAL prior to randomization and after 24 weeks of treatment (Study 1) or 12 weeks of treatment (Study 2). RESULTS: Measures of clinical efficacy were maintained or improved in subjects receiving FSC vs. FP250, whereas there were no improvements in clinical efficacy with FP+MON vs. FP100. Consistent with these clinical observations, no significant differences in bronchial mucosal eosinophils were found at 24 weeks for FSC vs. FP250 (95% CI: -0.64, 1.67) or at 12 weeks for FP100 vs. FP100+MON (95% CI: -6.61, 8.82) Similar results were noted for neutrophils, CD3+, CD4+, CD8+ and CD25+ lymphocytes in biopsies as well as BAL percentages of
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