- Email: [email protected]
Detailed sequence and haplotype analysis of the beta-2 adrenergic receptor gene in Caucasians and African Americans
S148 Abstracts
Variability in Asthma Severity in Pediatric Asthma Patients Previously Receiving Short-Acting Beta2-Agonists B. Chipps1, J. Spahn1, C. Sorkness2, L. Sutton3, A. Emmett3, P. Dorinsky3; 1Capital Allergy and Respiratory Disease Center, Sacramento, CA, 2University of Wisconsin, Madison, WI, 3GlaxoSmithKline, Research Triangle Park, NC. RATIONALE: To date, few studies have evaluated the degree of variability in asthma severity in pediatric patients. METHODS: An analysis of 5 previously conducted studies was performed to evaluate asthma variability in pediatric subjects, previously receiving short-acting beta2-agonists (SABAs) alone, who were subsequently randomized to continue this regimen during 12-week clinical trials. Subjects were stratified based upon % reversibility and FEV1 % predicted at baseline; changes in % predicted AMPEF were analyzed as an index of asthma severity. RESULTS: Untreated pediatric subjects receiving SABAs only exhibited marked fluctuations in asthma severity. In subjects with <20% reversibility at baseline, >35% of subjects had >15 changes in their asthma severity classification based on AMPEF. In subjects with 20-30% or ≥30% reversibility at baseline, over 25% and 40%, respectively, had >15 changes in their asthma severity classification. Furthermore, in subjects with FEV1 <60% predicted at baseline, 49% of subjects had >15 changes in their asthma severity classification. For subjects with baseline FEV1 values of 60-80% predicted or ≥80% predicted, 30% and 28% of subjects, respectively, had >15 changes in their asthma severity classification. CONCLUSION: This analysis clearly demonstrates that asthma severity is highly variable in untreated pediatric asthma patients and patients frequently move between severity categories over time. However, neither baseline reversibility nor FEV1 % predicted were reliable predictors of which patients exhibited marked variability. These data highlight the importance of ensuring that pediatric patients with asthma are carefully monitored and receive appropriate maintenance therapy to eliminate fluctuations in asthma control. Funding: GlaxoSmithKline
589
MONDAY
Improved Asthma Symptom Control and Asthma-Related Quality of Life With Fluticasone Propionate and Salmeterol (FS) in a Single Inhaler R. Manjunath1, A. S. Gilmore2, R. D. O’Connor3, R. H. Stanford1, A. P. Legorreta4, P. M. Jhingran1; 1GlaxoSmithKline, Rtp, NC, 2Health Benchmarks, Inc, Woodland Hills, CA, 3University of California at San Diego, San Diego, CA, 4California Department of Health Services, School of Public Health, University of California, Los Angeles, CA. RATIONALE: The purpose of this study was to evaluate symptom control and quality of life in patients with persistent asthma receiving fluticasone propionate/salmeterol (FS) from a single inhaler. METHODS: Prospective observational 12-month asthma study conducted with 224 physicians participating throughout the US. Patients (≥15 years) with a physician-confirmed diagnosis of asthma with no other concurrent respiratory disease prescribed FS for the treatment of persistent asthma were included. Patients prescribed FS or any single controller agents other than ICS, prior to the start of the study, were excluded. Patients were surveyed at baseline (index date) and 12 months. The Asthma Control Questionnaire (ACQ), a validated disease-specific instrument, scored on a scale of 0-6, with 6 indicating poorest asthma control was used to measure asthma control. The Asthma Quality of Life Questionnaire (AQLQ), a validated disease-specific instrument, scored on a scale from 1 to 7, with 7 indicating best quality of life, was used to measure quality of life. RESULTS: 390 subjects were included in the analysis. A mean decrease in overall ACQ score from baseline of 1.05 (p<0.00001) was observed, while the mean change in AQLQ score from baseline was 1.09 (p<0.00001). This change in quality of life is clinically meaningful (change of at least 0.5). CONCLUSIONS: These findings suggest that treatment with FS resulted in clinically meaningful and statistically significant improvement in asthma related quality of life as well as significant improvements in asth-
590
J ALLERGY CLIN IMMUNOL FEBRUARY 2005
ma control from baseline over 12 months in a population of asthma patients being treated in a general practice setting. Funding: GSK Linear Growth in Prepubertal Asthmatic Children Treated With Montelukast, Beclomethasone, or Placebo: A 56-Week, Randomized, Double-Blind Study B. A. Knorr1, O. Kuznetsova1, T. F. Reiss1, E. Beckford1, O. Johnson1, S. B. Dass1, A. B. Becker2; 1Merck & Co. Inc., Rahway, NJ, 2Department of Pediatrics and Child Health, University of Manitoba, Winnipeg, MB, CANADA. RATIONALE: Montelukast, a cysteinyl leukotriene receptor antagonist, and inhaled corticosteroids are widely used asthma controller agents. Some reports have shown that inhaled corticosteroids decrease pediatric linear growth. This study reports for the first time the effect of an antileukotriene on linear growth in asthmatic children. METHODS: Patients were 6.4-9.4 yr-old boys and 6.4-8.4 yr-old girls at Tanner Stage I, with mild, persistent asthma. After a placebo run-in, 360 patients were randomized to montelukast 5 mg, beclomethasone 200 g twice daily, or placebo. The primary endpoint was linear growth velocity over the 56-week, double-blind, treatment period. Height was measured (3 replicates) by trained personnel at each visit using a stadiometer. RESULTS: Linear growth velocity in the beclomethasone group was significantly less than in the placebo group (difference = -0.78 cm/year (95% CI -1.06, -0.49), (p<0.001)) or the montelukast group (0.81 cm/year (0.53, 1.09), p<0.001)). The growth rate was similar between montelukast and placebo: difference = 0.03 cm/year (-0.26, 0.31). The percent-days without -agonist use was significantly greater in the montelukast and beclomethasone groups versus the placebo group: medians were 89.5% (95% CI 86.8, 92.1) and 93.4% (90.9, 95.8) vs. 85.4% (80.7, 90.2). Numerically more patients used oral corticosteroid rescue in the placebo group (34.7% of patients) than with montelukast (25.0%) or beclomethasone (23.5%). CONCLUSION: In this study of pre-pubertal asthmatic children, montelukast did not affect linear growth, while growth with beclomethasone was significantly decreased during one year of treatment. Funding: Merck Research Laboratories
591
Detailed Sequence and Haplotype Analysis of the Beta-2 Adrenergic Receptor Gene in Caucasians and African Americans D. A. Meyers1, G. A. Hawkins1, K. Tantisira2, E. J. Ampleford1, M. Hernandez2, B. Richter2, S. P. Peters1, S. B. Liggett3, S. T. Weiss2, E. R. Bleecker1; 1Center for Human Genomics, Wake Forest University School of Medicine, Winston-Salem, NC, 2Channing Laboratory, Brigham and Women’s Hospital and Harvard Medical School, Boston, MA, 3Department of Medicine, University of Cincinnati College of Medicine, Cincinnati, OH. RATIONALE: Since 2 adrenergic receptor (2AR) variations appear to alter an asthma patient’s response to beta-agonist therapy, it is important to fully characterize 2AR gene variations and haplotypes in different ethnic groups. Clinical studies show that individuals homozygous for Arg16 have an unsatisfactory beta-agonist response versus individuals homozygous for Gly16. METHODS: A region -3470 bp 5’ of the ATG start site to +1886 bp in the 3’ UTR of the 2AR gene was re-sequenced in 429 Caucasian and 240 African American asthma cases and controls. RESULTS: Fifty-one validated polymorphisms were identified. Haplotypes were constructed using polymorphisms with frequencies ≥0.04 and LD measured. LD was strong across the 2AR gene, except for the 3’UTR region. Five Caucasian haplotypes containing Arg16 were differentiated exclusively by 3’ UTR variations. Ten African American haplotypes containing Arg16 were differentiated by 3’ UTR and promoter variations. Three novel promoter insertion/deletions were found in African Americans. The Thr164Ile variant was found exclusively in Caucasians while the Ser220Cys variant was found exclusively in African Americans. A potentially important 3’UTR polymorphism consists of an interrupted poly(C)
592
Abstracts S149
J ALLERGY CLIN IMMUNOL VOLUME 115, NUMBER 2
Airways Responsiveness and Airway Remodeling After Chronic Exposure to Procaterol and Fenoterol in Guinea Pigs In Vivo H. Nishimura, K. Tokuyama, H. Arakawa, Y. Ohki, H. Mochizuki, A. Morikawa; Pediatrics, Gunma University, Maebashi, JAPAN. BACKGROUND: Chronic exposure to fenoterol (FEN) is shown to induce both airways hyperresponsiveness and airway remodeling in experimental animals. OBJECTIVE: We wanted to know the effects of chronic exposure to procaterol (PRO), because this agent is widely used as a bronchodilator in Japan. METHODS: Aerosolised PRO (0.1 or 1 mg/ml), FEN (1mg/ml) or vehicle (0.9 % NaCl) were given to guinea pigs 3 times a day for 6 weeks. At 72 hrs after the last inhalation of PRO, FEN or vehicle, bronchial responsiveness to acetylcholine (ACh) was measured. After measuring RL, histological changes in noncartilaginous airway dimensions were evaluated. RESULTS: The amount of sublaryngeal deposition of 0.1 mg/ml PRO in the present study was speculated to be 100 times more than that of therapeutical use. Bronchial responsiveness to ACh was not different among 4 groups. In the smaller membranous airways (<0.4mm in diameter), but not the larger ones, thickening of adventitial areas was significantly greater in animals treated with 2-AR agonists than control animals (23 and 25 %, and 96 % higher in 0.1 and 1 mg/ml PRO, and in 1 mg/ml FENtreated animals, respectively). CONCLUSION: Chronic exposure to neither PRO nor FEN induced airways hyperresponsiveness. The degree of thickening in adventitial areas caused by PRO was mild when comparing with that of FEN. Further studies would be required as for the pathophysiological role of the thickening in this areas caused by 2-AR agonists. Funding: none
593
The Effect of Y-27632, a Rho/ Rho Kinase Inhibitor, on Leukotriene D4- and Histamine-Induced Airway Microvascular Leage in Guinea Pigs K. Tokuyama1, H. Nishimura1, K. Iizuka2, M. Kato1, H. Arakawa1, H. Mochizuki1, A. Morikawa1; 1Department of Pediatrics and Developmental Medicine, Gunma University Graduate School of Medicine, Maebashi, JAPAN, 2First Department of Internal Medicine, Gunma University Graduate School of Medicine, Maebashi, JAPAN. The Rho/Rho kinase pathway is shown to be involved in the mechanism of not only airway smooth muscle contraction but also vascular endothelial permeability. Thus, Rho/Rho kinase inhibitors may be useful to reduce airway microvascular leakage (MVL) which is seen in asthma. Therefore, we examined the effect of Y-27632, a selective Rho kinase inhibitor, on MVL caused by leukotriene D4 (LTD4) and histamine by comparing its effect against airflow obstruction. For comparison, the effects of procaterol, a beta2-adrenoceptor agonist, were also studied. Tracheostomized guinea pigs were given either aerosolized Y-27632 (3 or 15 mmol/l), procaterol (6 mol/l) or vehicle for 5 min under spontaneous breathing. After being mechanically ventilated, the animals were given intravenous Evans blue dye (EBD) 15 min after the end of inhalation. One minute later, either 2 nmol/kg LTD4, 300 nmol/kg histamine or vehicle was administered intravenously. After measurements of lung resistance (RL) for 6 min, the lungs of animals were taken out, and the amount of extravasated EBD was examined. Inhaled Y-27632 dose-dependently attenuated increases in RL caused by LTD4 and histamine. The degree of inhibition was similar between 15 mmol/l Y-27632 and 6 mol/l procaterol. By contrast, only 15 mmol/l Y-27632 partially reduced LTD4-induced leakage. Histamineinduced EBD extravasation was not inhibited by 15 mmol/l Y-27632.
594
Procaterol significantly inhibited the dye extravasation caused by either LTD4 or histamine. These results suggest that Y-27632 is not a useful agent in attenuating MVL which is seen in asthma, although it is potent in inhibiting airflow obstruction. In Vitro Comparison of the Performance of Salmeterol/Fluticasone Propionate HFA Metered Dose Inhaler (MDI) With and Without MDI Counter J. J. West, O. O’Brien, J. C. Ellis, C. A. O’Brien; GlaxoSmithKline, Ware, UNITED KINGDOM. RATIONALE: An MDI Counter has been developed for salmeterol/fluticasone propionate (SFC) HFA MDI (Seretide™ ) to indicate the number of actuations remaining for use in the canister. The counter is attached directly to the neck of the canister and is used in conjunction with a slightly modified actuator. This study was conducted to confirm that the performance of the MDI is unaffected by the presence of the MDI counter and associated modifications of the actuator. METHODS: Mean emitted dose (ex-actuator content per actuation) of SFC HFA MDI, was determined throughout the nominal number of actuations. Fine particle mass (FPM), corresponding to a particle size range of 1.1 to 4.7 m, was determined using the Andersen Cascade Impactor. RESULTS: For both the salmeterol (S) and fluticasone propionate (FP) components, product tested with/without MDI counter gave similar results for mean content per actuation (n=10 cans) and mean FPM (n=3 cans) for each of the product strengths: SFC 25 g/50 g: mean content per actuation 22/22g (S), 47/47g (FP); mean FPM 11/11g (S), 22/24g(FP). SFC 25 g /125 g: mean content per actuation 23/23g (S), 120/121g (FP); mean FPM 10/11g (S), 49/54g (FP). SFC 25 g /250 g: mean content per actuation 23/22g (S), 234/231g (FP); mean FPM 9/9g (S), 86/86g (FP). CONCLUSIONS: The results for the mean content per actuation and mean FPM for both salmeterol and fluticasone propionate demonstrate that the performance of each strength of SFC HFA MDI fitted with the MDI Counter is comparable with that of each strength without the MDI Counter. Funding: GlaxoSmithKline
595
Ventolin HFA MDI Safety in Children 24-<48 Months Old With Symptoms of Bronchospasm A. Davis-Allen, W. Lincourt, R. Trivedi, A. Ellsworth, C. Crim; GlaxoSmithKline, Research Triangle Park, NC. RATIONALE: Ventolin is indicated for quick relief of bronchospasm symptoms in children four years of age and older and is used in even younger children for relief of respiratory symptoms. Limited safety data exists in this young population. METHODS: Data from a multicenter, randomized, double-blind, parallel group, placebo-controlled study, evaluating Ventolin 90mcg or 180mcg versus placebo(PLB) in 24-<48 months old children, were analyzed. Ventolin and PLB treatments were administered via a MDI with Valved Holding Chamber and facemask TID for 4 weeks in children experiencing symptoms. Twenty-three children each received PLB and Ventolin 180mcg, and 24 received Ventolin 90mcg. Adverse events (AEs), tremor, ECGs and clinical laboratory tests were assessed. RESULTS: The overall incidence of AEs during treatment was in favor of Ventolin 90mcg (35%) compared with Ventolin 180mcg (52%) and placebo (42%). The most frequently reported AEs were ECG QT prolongation (12%in Ventolin 180mcg only); pyrexia (8% each -PLB and Ventolin 90mcg and 4%-Ventolin 180mcg); vomiting (8% PLB and 4% in Ventolin 90mcg only) and diarrhea (8% in Ventolin 180mcg only). Tremor was reported (using 4point scale of none-severe) in 3 subjects in Ventolin 90mcg group, 2 in Ventolin 180mcg and 1 subject in the PLB group, all reporting mild tremor. No cases of drug related, tachycardia, abnormal blood glucose or serum potassium were reported in the Ventolin treatment groups. CONCLUSIONS: This study demonstrates that 4 weeks of treatment with Ventolin HFA 90 or 180mcg TID delivered via MDI-spacer facemask to symptomatic children aged 24-<48 months, has a safety profile comparable to PLB. Funding: GlaxoSmithKline
596
MONDAY
repeat varying in size and frequency. This poly(C) region lies adjacent to an AU rich element, which is known to affect 2AR mRNA stability and translation in animal models. It is not clear what affect the hyper-variable 3’UTR region may have on regulation of human 2AR expression. CONCLUSION: These new observations provide opportunity to perform detailed comparison to beta-agonist responses and genotype and haplotype differences in the 2AR gene. Funding: NIH U0165899, NIH HL01-012
Variability in Asthma Severity in Pediatric Asthma Patients Previously Receiving Short-Acting Beta2-Agonists B. Chipps1, J. Spahn1, C. Sorkness2, L. Sutton3, A. Emmett3, P. Dorinsky3; 1Capital Allergy and Respiratory Disease Center, Sacramento, CA, 2University of Wisconsin, Madison, WI, 3GlaxoSmithKline, Research Triangle Park, NC. RATIONALE: To date, few studies have evaluated the degree of variability in asthma severity in pediatric patients. METHODS: An analysis of 5 previously conducted studies was performed to evaluate asthma variability in pediatric subjects, previously receiving short-acting beta2-agonists (SABAs) alone, who were subsequently randomized to continue this regimen during 12-week clinical trials. Subjects were stratified based upon % reversibility and FEV1 % predicted at baseline; changes in % predicted AMPEF were analyzed as an index of asthma severity. RESULTS: Untreated pediatric subjects receiving SABAs only exhibited marked fluctuations in asthma severity. In subjects with <20% reversibility at baseline, >35% of subjects had >15 changes in their asthma severity classification based on AMPEF. In subjects with 20-30% or ≥30% reversibility at baseline, over 25% and 40%, respectively, had >15 changes in their asthma severity classification. Furthermore, in subjects with FEV1 <60% predicted at baseline, 49% of subjects had >15 changes in their asthma severity classification. For subjects with baseline FEV1 values of 60-80% predicted or ≥80% predicted, 30% and 28% of subjects, respectively, had >15 changes in their asthma severity classification. CONCLUSION: This analysis clearly demonstrates that asthma severity is highly variable in untreated pediatric asthma patients and patients frequently move between severity categories over time. However, neither baseline reversibility nor FEV1 % predicted were reliable predictors of which patients exhibited marked variability. These data highlight the importance of ensuring that pediatric patients with asthma are carefully monitored and receive appropriate maintenance therapy to eliminate fluctuations in asthma control. Funding: GlaxoSmithKline
589
MONDAY
Improved Asthma Symptom Control and Asthma-Related Quality of Life With Fluticasone Propionate and Salmeterol (FS) in a Single Inhaler R. Manjunath1, A. S. Gilmore2, R. D. O’Connor3, R. H. Stanford1, A. P. Legorreta4, P. M. Jhingran1; 1GlaxoSmithKline, Rtp, NC, 2Health Benchmarks, Inc, Woodland Hills, CA, 3University of California at San Diego, San Diego, CA, 4California Department of Health Services, School of Public Health, University of California, Los Angeles, CA. RATIONALE: The purpose of this study was to evaluate symptom control and quality of life in patients with persistent asthma receiving fluticasone propionate/salmeterol (FS) from a single inhaler. METHODS: Prospective observational 12-month asthma study conducted with 224 physicians participating throughout the US. Patients (≥15 years) with a physician-confirmed diagnosis of asthma with no other concurrent respiratory disease prescribed FS for the treatment of persistent asthma were included. Patients prescribed FS or any single controller agents other than ICS, prior to the start of the study, were excluded. Patients were surveyed at baseline (index date) and 12 months. The Asthma Control Questionnaire (ACQ), a validated disease-specific instrument, scored on a scale of 0-6, with 6 indicating poorest asthma control was used to measure asthma control. The Asthma Quality of Life Questionnaire (AQLQ), a validated disease-specific instrument, scored on a scale from 1 to 7, with 7 indicating best quality of life, was used to measure quality of life. RESULTS: 390 subjects were included in the analysis. A mean decrease in overall ACQ score from baseline of 1.05 (p<0.00001) was observed, while the mean change in AQLQ score from baseline was 1.09 (p<0.00001). This change in quality of life is clinically meaningful (change of at least 0.5). CONCLUSIONS: These findings suggest that treatment with FS resulted in clinically meaningful and statistically significant improvement in asthma related quality of life as well as significant improvements in asth-
590
J ALLERGY CLIN IMMUNOL FEBRUARY 2005
ma control from baseline over 12 months in a population of asthma patients being treated in a general practice setting. Funding: GSK Linear Growth in Prepubertal Asthmatic Children Treated With Montelukast, Beclomethasone, or Placebo: A 56-Week, Randomized, Double-Blind Study B. A. Knorr1, O. Kuznetsova1, T. F. Reiss1, E. Beckford1, O. Johnson1, S. B. Dass1, A. B. Becker2; 1Merck & Co. Inc., Rahway, NJ, 2Department of Pediatrics and Child Health, University of Manitoba, Winnipeg, MB, CANADA. RATIONALE: Montelukast, a cysteinyl leukotriene receptor antagonist, and inhaled corticosteroids are widely used asthma controller agents. Some reports have shown that inhaled corticosteroids decrease pediatric linear growth. This study reports for the first time the effect of an antileukotriene on linear growth in asthmatic children. METHODS: Patients were 6.4-9.4 yr-old boys and 6.4-8.4 yr-old girls at Tanner Stage I, with mild, persistent asthma. After a placebo run-in, 360 patients were randomized to montelukast 5 mg, beclomethasone 200 g twice daily, or placebo. The primary endpoint was linear growth velocity over the 56-week, double-blind, treatment period. Height was measured (3 replicates) by trained personnel at each visit using a stadiometer. RESULTS: Linear growth velocity in the beclomethasone group was significantly less than in the placebo group (difference = -0.78 cm/year (95% CI -1.06, -0.49), (p<0.001)) or the montelukast group (0.81 cm/year (0.53, 1.09), p<0.001)). The growth rate was similar between montelukast and placebo: difference = 0.03 cm/year (-0.26, 0.31). The percent-days without -agonist use was significantly greater in the montelukast and beclomethasone groups versus the placebo group: medians were 89.5% (95% CI 86.8, 92.1) and 93.4% (90.9, 95.8) vs. 85.4% (80.7, 90.2). Numerically more patients used oral corticosteroid rescue in the placebo group (34.7% of patients) than with montelukast (25.0%) or beclomethasone (23.5%). CONCLUSION: In this study of pre-pubertal asthmatic children, montelukast did not affect linear growth, while growth with beclomethasone was significantly decreased during one year of treatment. Funding: Merck Research Laboratories
591
Detailed Sequence and Haplotype Analysis of the Beta-2 Adrenergic Receptor Gene in Caucasians and African Americans D. A. Meyers1, G. A. Hawkins1, K. Tantisira2, E. J. Ampleford1, M. Hernandez2, B. Richter2, S. P. Peters1, S. B. Liggett3, S. T. Weiss2, E. R. Bleecker1; 1Center for Human Genomics, Wake Forest University School of Medicine, Winston-Salem, NC, 2Channing Laboratory, Brigham and Women’s Hospital and Harvard Medical School, Boston, MA, 3Department of Medicine, University of Cincinnati College of Medicine, Cincinnati, OH. RATIONALE: Since 2 adrenergic receptor (2AR) variations appear to alter an asthma patient’s response to beta-agonist therapy, it is important to fully characterize 2AR gene variations and haplotypes in different ethnic groups. Clinical studies show that individuals homozygous for Arg16 have an unsatisfactory beta-agonist response versus individuals homozygous for Gly16. METHODS: A region -3470 bp 5’ of the ATG start site to +1886 bp in the 3’ UTR of the 2AR gene was re-sequenced in 429 Caucasian and 240 African American asthma cases and controls. RESULTS: Fifty-one validated polymorphisms were identified. Haplotypes were constructed using polymorphisms with frequencies ≥0.04 and LD measured. LD was strong across the 2AR gene, except for the 3’UTR region. Five Caucasian haplotypes containing Arg16 were differentiated exclusively by 3’ UTR variations. Ten African American haplotypes containing Arg16 were differentiated by 3’ UTR and promoter variations. Three novel promoter insertion/deletions were found in African Americans. The Thr164Ile variant was found exclusively in Caucasians while the Ser220Cys variant was found exclusively in African Americans. A potentially important 3’UTR polymorphism consists of an interrupted poly(C)
592
Abstracts S149
J ALLERGY CLIN IMMUNOL VOLUME 115, NUMBER 2
Airways Responsiveness and Airway Remodeling After Chronic Exposure to Procaterol and Fenoterol in Guinea Pigs In Vivo H. Nishimura, K. Tokuyama, H. Arakawa, Y. Ohki, H. Mochizuki, A. Morikawa; Pediatrics, Gunma University, Maebashi, JAPAN. BACKGROUND: Chronic exposure to fenoterol (FEN) is shown to induce both airways hyperresponsiveness and airway remodeling in experimental animals. OBJECTIVE: We wanted to know the effects of chronic exposure to procaterol (PRO), because this agent is widely used as a bronchodilator in Japan. METHODS: Aerosolised PRO (0.1 or 1 mg/ml), FEN (1mg/ml) or vehicle (0.9 % NaCl) were given to guinea pigs 3 times a day for 6 weeks. At 72 hrs after the last inhalation of PRO, FEN or vehicle, bronchial responsiveness to acetylcholine (ACh) was measured. After measuring RL, histological changes in noncartilaginous airway dimensions were evaluated. RESULTS: The amount of sublaryngeal deposition of 0.1 mg/ml PRO in the present study was speculated to be 100 times more than that of therapeutical use. Bronchial responsiveness to ACh was not different among 4 groups. In the smaller membranous airways (<0.4mm in diameter), but not the larger ones, thickening of adventitial areas was significantly greater in animals treated with 2-AR agonists than control animals (23 and 25 %, and 96 % higher in 0.1 and 1 mg/ml PRO, and in 1 mg/ml FENtreated animals, respectively). CONCLUSION: Chronic exposure to neither PRO nor FEN induced airways hyperresponsiveness. The degree of thickening in adventitial areas caused by PRO was mild when comparing with that of FEN. Further studies would be required as for the pathophysiological role of the thickening in this areas caused by 2-AR agonists. Funding: none
593
The Effect of Y-27632, a Rho/ Rho Kinase Inhibitor, on Leukotriene D4- and Histamine-Induced Airway Microvascular Leage in Guinea Pigs K. Tokuyama1, H. Nishimura1, K. Iizuka2, M. Kato1, H. Arakawa1, H. Mochizuki1, A. Morikawa1; 1Department of Pediatrics and Developmental Medicine, Gunma University Graduate School of Medicine, Maebashi, JAPAN, 2First Department of Internal Medicine, Gunma University Graduate School of Medicine, Maebashi, JAPAN. The Rho/Rho kinase pathway is shown to be involved in the mechanism of not only airway smooth muscle contraction but also vascular endothelial permeability. Thus, Rho/Rho kinase inhibitors may be useful to reduce airway microvascular leakage (MVL) which is seen in asthma. Therefore, we examined the effect of Y-27632, a selective Rho kinase inhibitor, on MVL caused by leukotriene D4 (LTD4) and histamine by comparing its effect against airflow obstruction. For comparison, the effects of procaterol, a beta2-adrenoceptor agonist, were also studied. Tracheostomized guinea pigs were given either aerosolized Y-27632 (3 or 15 mmol/l), procaterol (6 mol/l) or vehicle for 5 min under spontaneous breathing. After being mechanically ventilated, the animals were given intravenous Evans blue dye (EBD) 15 min after the end of inhalation. One minute later, either 2 nmol/kg LTD4, 300 nmol/kg histamine or vehicle was administered intravenously. After measurements of lung resistance (RL) for 6 min, the lungs of animals were taken out, and the amount of extravasated EBD was examined. Inhaled Y-27632 dose-dependently attenuated increases in RL caused by LTD4 and histamine. The degree of inhibition was similar between 15 mmol/l Y-27632 and 6 mol/l procaterol. By contrast, only 15 mmol/l Y-27632 partially reduced LTD4-induced leakage. Histamineinduced EBD extravasation was not inhibited by 15 mmol/l Y-27632.
594
Procaterol significantly inhibited the dye extravasation caused by either LTD4 or histamine. These results suggest that Y-27632 is not a useful agent in attenuating MVL which is seen in asthma, although it is potent in inhibiting airflow obstruction. In Vitro Comparison of the Performance of Salmeterol/Fluticasone Propionate HFA Metered Dose Inhaler (MDI) With and Without MDI Counter J. J. West, O. O’Brien, J. C. Ellis, C. A. O’Brien; GlaxoSmithKline, Ware, UNITED KINGDOM. RATIONALE: An MDI Counter has been developed for salmeterol/fluticasone propionate (SFC) HFA MDI (Seretide™ ) to indicate the number of actuations remaining for use in the canister. The counter is attached directly to the neck of the canister and is used in conjunction with a slightly modified actuator. This study was conducted to confirm that the performance of the MDI is unaffected by the presence of the MDI counter and associated modifications of the actuator. METHODS: Mean emitted dose (ex-actuator content per actuation) of SFC HFA MDI, was determined throughout the nominal number of actuations. Fine particle mass (FPM), corresponding to a particle size range of 1.1 to 4.7 m, was determined using the Andersen Cascade Impactor. RESULTS: For both the salmeterol (S) and fluticasone propionate (FP) components, product tested with/without MDI counter gave similar results for mean content per actuation (n=10 cans) and mean FPM (n=3 cans) for each of the product strengths: SFC 25 g/50 g: mean content per actuation 22/22g (S), 47/47g (FP); mean FPM 11/11g (S), 22/24g(FP). SFC 25 g /125 g: mean content per actuation 23/23g (S), 120/121g (FP); mean FPM 10/11g (S), 49/54g (FP). SFC 25 g /250 g: mean content per actuation 23/22g (S), 234/231g (FP); mean FPM 9/9g (S), 86/86g (FP). CONCLUSIONS: The results for the mean content per actuation and mean FPM for both salmeterol and fluticasone propionate demonstrate that the performance of each strength of SFC HFA MDI fitted with the MDI Counter is comparable with that of each strength without the MDI Counter. Funding: GlaxoSmithKline
595
Ventolin HFA MDI Safety in Children 24-<48 Months Old With Symptoms of Bronchospasm A. Davis-Allen, W. Lincourt, R. Trivedi, A. Ellsworth, C. Crim; GlaxoSmithKline, Research Triangle Park, NC. RATIONALE: Ventolin is indicated for quick relief of bronchospasm symptoms in children four years of age and older and is used in even younger children for relief of respiratory symptoms. Limited safety data exists in this young population. METHODS: Data from a multicenter, randomized, double-blind, parallel group, placebo-controlled study, evaluating Ventolin 90mcg or 180mcg versus placebo(PLB) in 24-<48 months old children, were analyzed. Ventolin and PLB treatments were administered via a MDI with Valved Holding Chamber and facemask TID for 4 weeks in children experiencing symptoms. Twenty-three children each received PLB and Ventolin 180mcg, and 24 received Ventolin 90mcg. Adverse events (AEs), tremor, ECGs and clinical laboratory tests were assessed. RESULTS: The overall incidence of AEs during treatment was in favor of Ventolin 90mcg (35%) compared with Ventolin 180mcg (52%) and placebo (42%). The most frequently reported AEs were ECG QT prolongation (12%in Ventolin 180mcg only); pyrexia (8% each -PLB and Ventolin 90mcg and 4%-Ventolin 180mcg); vomiting (8% PLB and 4% in Ventolin 90mcg only) and diarrhea (8% in Ventolin 180mcg only). Tremor was reported (using 4point scale of none-severe) in 3 subjects in Ventolin 90mcg group, 2 in Ventolin 180mcg and 1 subject in the PLB group, all reporting mild tremor. No cases of drug related, tachycardia, abnormal blood glucose or serum potassium were reported in the Ventolin treatment groups. CONCLUSIONS: This study demonstrates that 4 weeks of treatment with Ventolin HFA 90 or 180mcg TID delivered via MDI-spacer facemask to symptomatic children aged 24-<48 months, has a safety profile comparable to PLB. Funding: GlaxoSmithKline
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repeat varying in size and frequency. This poly(C) region lies adjacent to an AU rich element, which is known to affect 2AR mRNA stability and translation in animal models. It is not clear what affect the hyper-variable 3’UTR region may have on regulation of human 2AR expression. CONCLUSION: These new observations provide opportunity to perform detailed comparison to beta-agonist responses and genotype and haplotype differences in the 2AR gene. Funding: NIH U0165899, NIH HL01-012