Publication Date:
2013-08-07
Description:
Interleukin-8 (CXCL8) exerts its role in inflammation by triggering neutrophils via its specific GPC receptors, CXCR1 and CXCR2, for which additional binding to endothelial heparan sulphate glycosaminoglycans (GAGs) is required. We present here a novel approach for blocking the CXCL8-related inflammatory cascade by generating dominant-negative CXCL8 mutants with improved GAG binding affinity and knocked-out CXCR1/CXCR2 activity. These non-signalling CXCL8 decoy proteins are able to displace wild type CXCL8 and to prevent CXCR1/CXCR2 signalling thereby interfering with the inflammatory response. We have designed 14 CXCL8 mutants which we sub-divided into three classes according to number and site of mutations. The decoys were characterised by isothermal fluorescence titrations and surface plasmon resonance to determine GAG affinity. Protein stability and structural changes were evaluated by far-UV circular dichroism spectroscopy and knocked-out GPC receptor response was shown by Boyden chamber and Ca 2+ release assays. From these experiments, CXCL8(D6F17KF21KE70KN71K) emerged with the most promising in vitro characteristics. This mutant was therefore further investigated in a murine model of mBSA-induced arthritis in mice where it showed strong anti-inflammatory activity. Based on these results, we propose that dominant-negative CXCL8 decoy proteins are a promising class of novel biopharmaceuticals with high therapeutic potential in inflammatory diseases.
Print ISSN:
0144-8463
Electronic ISSN:
1573-4935
Topics:
Biology
,
Chemistry and Pharmacology
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