Non-Modified CpG Oligodeoxynucleotide Forming Guanine-Quadruplex Structure Complexes with e-Poly-L-Lysine Induce Antibody Production as Vaccine Adjuvants

Unmethylated cytosine-phosphate-guanosine oligodeoxynucleotides (CpG ODNs) induce inflammatory cytokines and type I interferons (IFNs) to activate the immune system. To apply CpG ODNs as vaccine adjuvants, the cellular uptake and stability of phosphodiester-based, non-modified ODNs require further improvement. Previously developed new CpG ODNs forming guanine-quadruplex (G4) structures showed higher nuclease resistance and cellular uptake than linear CpG ODNs; however, the complex formation of G4-CpG ODNs with antigen proteins is necessary for their application as vaccine adjuvants. In this study, we utilized a cationic polymer, epsilon-poly-(L)-lysine (epsilon-PLL), as a carrier for G4-CpG ODNs and antigen. The epsilon-PLL/G4-CpG ODN complex exhibited enhanced stability against nucleases. Cellular uptake of the epsilon-PLL/G4-CpG ODN complex positively correlated with the N/P ratio. In comparison to naked G4-CpG ODNs, the epsilon-PLL/G4-CpG ODN complex induced extremely high levels of interleukin (IL)-6, IL-12, and IFN-beta. Relative immune cytokine production was successfully tuned by N/P ratio modification. Mice with the epsilon-PLL/G4-CpG ODN/ovalbumin (OVA) complex showed increased OVA-specific immunoglobulin (Ig)G, IgG1, and IgG2c levels, whereas total IgE levels did not increase and weight gain rates were not affected. Therefore, epsilon-PLL can serve as a safe and effective phosphodiester-based, non-modified CpG ODN delivery system, and the epsilon-PLL/G4-CpG ODN/antigen complex is a highly promising candidate for vaccine adjuvants and can be further used in clinical research.

Automated summary

In this study, the ε-PLL/G4-CpG ODN complex was successfully synthesized as a carrier, providing better stability against nucleases and significantly improved cellular uptake. Modulation of the N/P ratio successfully tuned the levels of immune cytokine production.