Phosphorothioate backbone modification modulates macrophage activation by CpG DNA

Macrophages respond to unmethylated CpG motifs present in nonmammalian DNA, Stabilized phosphorothioate-modified oligodeoxynucleotides (PS-ODN) contai ning CpG motifs form the basis of immunotherapeutic agents. In this study, we show that PS-ODN do not perfectly mimic native DNA in activation of macr ophages. CpG-containing PS-ODN were active at 10- to 100-fold lower concent rations than corresponding phosphodiester ODN in maintenance of cell viabil ity in the absence of CSF-1, in induction of NO production, and in activati on of the IL-12 promoter. These enhancing effects are attributable to both increased stability and rate of uptake of the PS-ODN, By contrast, PS-ODN w ere almost inactive in down-modulation of the CSF-1R from primary macrophag es and activation of the HIV-1 LTR, Delayed or poor activation of signaling components may contribute to this, as PS-ODN were slower and less effectiv e at inducing phosphorylation of the extracellular signal-related kinases 1 and 2. In addition, at high concentrations, non-CpG PS-ODN specifically in hibited responses to CpG DNA, whereas nonstimulatory phosphodiester ODN had no such effect. Although nonstimulatory PS-ODN caused some inhibition of O DN uptake, this did not adequately explain the levels of inhibition of acti vity. The results demonstrate that the phosphorothioate backbone has both e nhancing and inhibitory effects on macrophage responses to CPG DNA.