G-quadruplex-based CpG oligodeoxynucleotide/DOTAP complex strongly stimulates immunity in CpG motif-specific and loop-length-dependent manners

Guanine-quadruplex (G4) oligodeoxynucleotides (ODNs) that contain unmethylated cytosine-phosphate-guanine motifs (G4 CpG ODN) with phosphodiester backbones are safer than the phosphorothioate (PT)-modified CpG ODNs recently used as vaccine adjuvants. However, cellular uptake and the nuclease stability of G4 CpG ODNs are still insufficient, resulting in lower immunostimulatory activity than PT-modified CpG ODNs. We aimed to enhance the immunostimulatory properties of G4 CpG ODNs by complexing with the cationic liposome 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP). The complex acquired nuclease resistance and improved cellular uptake. The immunostimulatory activity of the G4 CpG ODN-DOTAP lipoplexes was enhanced to a level comparable to that of PT-modified ODNs. In addition, the lipoplexes based on unmodified G4 CpG ODNs demonstrated CpG motif-specific immunostimulant activity, although PT-modified ODNs lacking the CpG motif could activate human immune cells. Interestingly, G4 CpG ODN-DOTAP lipoplexes induced interferon-α production in a loop-length dependent manner.