Characterization of suppressive oligodeoxynucleotides that inhibit Toll‐like receptor‐9‐mediated activation of innate immunity

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Abstract
Summary: Synthetic oligodeoxynucleotides containing unmethylated CpG sequences (CpG‐ODNs) stimulate Toll‐like receptor‐9 (TLR‐9), thereby activating innate immunity. Stimulatory CpG‐ODNs have been shown to be valuable in modifying immune responses in allergy, infection and cancer. Recently, it has been reported that the stimulation of TLR‐9 by endogenous DNA might contribute to the pathogenesis of autoimmune diseases. We here report the identification of a suppressive, guanosine‐rich ODN (G‐ODN) that inhibited the activation of TLR‐9 by stimulatory CpG‐ODNs. The G‐ODN was suppressive in murine macrophages and dendritic cells as well as in human plasmacytoid dendritic cells in vitro. G‐ODN blocked the secretion of tumour necrosis factor‐α (TNF‐α) and interleukin‐12p40 and interfered with the up‐regulation of major histocompatibility complex (MHC) class II and costimulatory molecules. G‐ODN was inhibitory even at a molar ratio of 1 : 10 (G‐ODN:CpG‐ODN) and when administered up to 7 hr after stimulation with CpG. G‐ODN specifically inhibited TLR‐9 but not other TLRs. Inhibition was dependent on a string of five guanosines. G‐ODN was also inhibitory in an in vivo model of CpG/galactosamin (GalN) lethal shock. G‐ODN interfered with upstream TLR‐9 signalling. However, by extensive analysis we can exclude that G‐ODN acts at the stage of cellular uptake. G‐ODN therefore represents a class of suppressive ODNs that could be of therapeutic use in situations with pathologic TLR‐9 activation, as has been proposed for certain autoimmune diseases.