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- Dahlén, Eva, et al.
(författare)
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Development of interleukin-1 receptor antagonist mutants with enhanced antagonistic activity in vitro and improved therapeutic efficacy in collagen-induced arthritis.
- 2008
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Ingår i: Journal of Immunotoxicology. - : Informa UK Limited. - 1547-6901 .- 1547-691X. ; 5:2, s. 189-199
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Tidskriftsartikel (refereegranskat)abstract
- Interleukin-1 receptor antagonist (IL-1Ra) is a naturally occurring inhibitor of the pro-inflammatory interleukin-1-mediated activation of the interleukin-1 receptor (IL-1R). Although wild-type IL-1Ra is used for treatment of inflammatory diseases, its effect is moderate and/or short-lived. The objective of this study was to generate IL-1Ra mutants with enhanced antagonistic activity for potential therapeutic use. Using a directed evolution approach in which libraries of IL-1Ra gene mutants were generated and screened in functional assays, mutants with desired properties were identified. Initially, diversity was introduced into the IL-1Ra using random mutagenesis. Mutations resulting in enhanced antagonistic activity were identified by screening in a reporter cell assay. To further enhance the antagonistic activity, selected mutations were recombined using the DNA recombination technology Fragment-INduced Diversity (FIND). Following three rounds of FIND recombination, several mutants with up to nine times enhanced antagonistic activity (mean IC50 +/- SEM value: 0.78 +/- 0.050 vs. 6.8 +/- 1.1 ng/ml for mutant and wild-type, respectively) were identified. Sequence analysis identified the mutations D47N, E52R and E90Y as being most important for this effect, however, the mutations P38Y, H54R, Q129L and M136N further enhanced the antagonistic function. Analysis of identified mutations in protein models based on the crystal structure of the IL-1Ra/IL-1R complex suggested that mutations found to enhance the antagonistic activity had a stabilizing effect on the IL-1Ra mutants or increased the affinity for the IL-1R. Finally, the therapeutic effect of one mutant was compared to that of wild-type IL-1Ra in collagen-induced arthritis in mice. Indeed, the enhanced antagonistic effect of the mutants observed in vitro was also seen in vivo. In conclusion, these results demonstrate that directed evolution of IL-1Ra is an effective means of generating highly potent therapeutic proteins.
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| 4. |
- Pedroletti, C, et al.
(författare)
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Clinical effects of purified air administered to the breathing zone in allergic asthma : A double-blind randomized cross-over trial
- 2009
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Ingår i: RESPIRATORY MEDICINE. - : Elsevier BV. - 0954-6111 .- 1532-3064. ; 103:9, s. 1313-1319
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Tidskriftsartikel (refereegranskat)abstract
- Background: Exposure to inhaled allergens is a pathogenetic factor in allergic asthma. However, most studies that previously looked at air cleaning devices have shown little or no effect on patients with perennial allergic asthma. Aims and objectives: We examined a novel treatment using temperature regulated laminar airflow with a very low particle concentration directed to the breathing zone of teenagers and young adults with mild to moderate allergic asthma during night steep. We hypothesised that the decreased allergen exposure during the night would have an effect on bronchial inflammation and quality of life. Method: Twenty-two patients (mean 18.8 years) were randomized to start with active or placebo treatment for 10 weeks. At( patients received both active and placebo treatment with unfiltered air, with a 2-week wash-out period in between treatments. Maintenance treatment with inhaled corticosteroids was unaltered during the trial period. Health related quality of life (miniAQLQ) was the primary effectiveness measure. Exhaled nitric oxide (FeNO) and spirometry were also investigated. Results: Active treatment resulted in an improved miniAQLQ compared to placebo (mean score 0.54, p andlt; 0.05, n = 20). An effect on bronchial, inflammation was also detected with significantly tower FeNO values during the active treatment period (mean -6.95 ppb, p andlt; 0.05, n = 22). Both effects were evident after 5 weeks. The change in lung function was not statistically significant. Conclusion: Clean air, administered directly to the breathing zone during steep, can have a positive effect on bronchial. inflammation and quality of life in patients with perennial allergic asthma.
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