Document Detail

Establishment of a novel cell-based assay for screening small molecule antagonists of human interleukin-6 receptor.
MedLine Citation:
PMID:  25345743     Owner:  NLM     Status:  Publisher    
Aim:Blockade of interleukin-6 (IL-6) or its receptor (IL-6R) is effective in preventing the progression of autoimmune diseases, such as systemic lupus erythematosus and rheumatoid arthritis. In the present study, we established a novel cell-based assay for identifying small molecule IL-6R antagonists.Methods:HEK293A cells were transfected with recombinant plasmids pTaglite-SNAP-IL6R and pABhFc-IL6 to obtain membrane-bound IL-6R and recombinant human IL-6 coupled with human Fc fragment (rhIL-6), respectively. A novel screening assay based on the interaction between IL-6R and rhIL-6 was established, optimized and validated. The stability of the assay was also assessed by calculating the Z'-factor.Results:RhIL-6 dose-dependently bound to IL-6R expressed at HEK293A cell surface. The IC50 value of the known antagonist ab47215 was 0.38±0.08 μg/mL, which was consistent with that obtained using the traditional method (0.36±0.14 μg/mL). The value of Z'-factor was 0.68, suggesting that the novel assay was stable for high throughput screening. A total of 474 compounds were screened using the novel screening assay, and 3 compounds exhibited antagonistic activities (IC50=8.73±0.28, 32.32±9.08, 57.83±4.24 μg/mL). Furthermore, the active compounds dose-dependently inhibited IL-6-induced proliferation of 7TD1 cells, and reduced IL-6-induced STAT3 phosphorylation in U937 cells.Conclusion:A novel cell-based screening assay for identifying small molecule IL-6R antagonists was established, which simplifies the procedures in traditional cellular ELISA screening and profiling and reduces the costs.
Yang-Yang He; Yu Yan; Chang Zhang; Peng-Yuan Li; Ping Wu; Peng Du; Da-di Zeng; Jian-Song Fang; Shuang Wang; Guan-Hua Du
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2014-10-27
Journal Detail:
Title:  Acta pharmacologica Sinica     Volume:  -     ISSN:  1745-7254     ISO Abbreviation:  Acta Pharmacol. Sin.     Publication Date:  2014 Oct 
Date Detail:
Created Date:  2014-10-27     Completed Date:  -     Revised Date:  2014-10-28    
Medline Journal Info:
Nlm Unique ID:  100956087     Medline TA:  Acta Pharmacol Sin     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
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