Sanglifehrin A is a novel dendritic cell chemokine and migration inhibitor

Abstract

Objective: Sanglifehrin A (SFA) is an immunosuppressive drug. It is a representative of a class of macrolides produced by the actinomycetes strain Streptomyces A92-308110 that bind to cyclophilin A (CypA), the binding protein of Cyclosporine A (CsA). Although SFA has a higher affinity than CsA for cyclophilin, SFA does not inhibit the activity of calcineurin phosphatases. Different groups have reported that SFA exerts unique suppressive effects on human and mouse dendritic cells (DC). SFA effectively suppresses antigen uptake and bioactive IL-12 production of DC in vitro and in vivo but it does not inhibit DC differentiation and surface costimulatory molecule expression. The mechanism of action of SFA is still unknown. Our goal is to investigate the gene expression profile of SFA treated mature DCs with subsequent confirmation on the protein level and functional in vitro and in vivo assays. Material and methods: Human monocytes were isolated from buffy coats of healthy blood donors. CD14+ monocytes were cultured with IL-4 and GM-CSF to generate DCs. On day 5, SFA or vehicle was added to the culture for one hour. After 12 hours stimulation with Lipopolysaccharid, RNA was isolated and microarrays were performed. Expressions of proteins are studied by ELISA and FACS. In vitro migration was estimated with chemotaxis assays. FITC-skin-painting method was used to study the in vivo migration of DC towards the inguinal lymph node by C57BL/6NCrl mice. Results: Global gene expression analysis and subsequent protein level confirmation revealed that SFA suppressed CCL5, CCL17, CCL19, CXCL9 and CXCL10 expression in human monocyte-derived DC. Direct comparison with the related agent cyclosporine A (CsA) and the classical corticosteroid dexamethasone indicated that SFA uniquely suppresses DC chemokine expression. Given the emerging critical role of chemokines production and the important role for migration of DC to initiate innate and adaptive immune responses, we investigated the effect of SFA on DC migration in vitro and in vivo. In vitro, functional assays demonstrated impaired migratory activity of maturing, SFA-exposed DC against CCL19. Moreover, SFA suppressed expression of the ectoenzyme CD38 that was reported to regulate DC migration and cytokine production. In vivo analyses confirmed SFA's inhibitory effect on DC migration. Conclusion: This first systematic genomewide study revealed a novel anti-inflammatory mode of action of SFA being different from the related agent CsA. Our results identify SFA as a novel DC chemokine and migration inhibitor.