In:
Arteriosclerosis, Thrombosis, and Vascular Biology, Ovid Technologies (Wolters Kluwer Health), Vol. 33, No. suppl_1 ( 2013-05)
Abstract:
The co-stimulatory CD40-CD40L dyad is well known for its pro-inflammatory role in atherosclerosis. Inhibition of CD40(L) in hyperlipidemic mice drastically reduces atherosclerosis. However, long term inhibition of CD40(L) results in immunosuppression and/or thromboembolic events. Therefore more targeted inhibition of the CD40L-CD40 dyad is required. To elicit intracellular signalling upon activation, CD40 needs to recruit adaptor proteins: the tumour necrosis factor receptor-associated factors (TRAFs). Using mice with a mutation in the CD40-TRAF binding site, we previously showed that CD40-TRAF6, and not CD40-TRAF2/3/5, are pivotal in atherosclerosis. To discover small drug-like molecules that inhibit the CD40-TRAF6 interaction an in silico approach of virtual ligand screening was used; this resulted in the identification of 40.000 compounds. Analysis of the top 800 compounds identified 7 compounds that dose-dependently reduced NFκB activation and IL1β and IL6 expression in RAW cells and CD40-stimulated bone marrow-derived macrophages, respectively. Surface plasmon resonance experiments confirmed direct binding of the compounds to the TRAF6 C-domain. To analyze the effects of the top two compounds on atherosclerosis, Apoe -/- mice were daily treated with compounds 6877002 or 6860766 for 6 weeks. Compound treatment reduced atherosclerosis in the aortic arch by 47.1% (6877002) and 66.8% (6860766). The number of plaque monocytes/macrophages was decreased by 41.4% (6877002) and 53.0% (6860766), and granulocytes by 62.5% (6877002) and 41.5% (6860766). Intravital microscopy showed that the compounds reduced monocyte recruitment to the endothelium by 40.1% (6877002) and 51.2% (6860766), neutrophil adhesion was reduced by 40.2% (6877002) and 51.2% (6860766). In accordance, expression of CCL2-CCR2 and CCL5-CCR5 was remarkably reduced in compound treated macrophages. Moreover, compound treatment abolished CD40-induced expression of TNFα, IL1β, IL6, IL10, and IL12. These results establish the importance and possibilities of long-term therapeutic inhibition of CD40-TRAF6 interactions in atherosclerosis and other inflammatory diseases and harbor the potential to overcome the current limitations of CD40(L) blocking therapies.
Type of Medium:
Online Resource
ISSN:
1079-5642
,
1524-4636
DOI:
10.1161/atvb.33.suppl_1.A14
Language:
English
Publisher:
Ovid Technologies (Wolters Kluwer Health)
Publication Date:
2013
detail.hit.zdb_id:
1494427-3
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