Intranasal immunization of mice with CpG DNA induces strong systemic and mucosal responses that are influenced by other mucosal adjuvants and antigendistribution

Background: Synthetic oligodeoxynucleotides (ODN) containing immunostimulat ory cytosine-guanine phosphate-linked dinucleotide (CpG) motifs are potent systemic and mucosal adjuvants in mice that have synergistic action with nu merous other adjuvants, including alum and cholera toxin (CT). Herein, we e valuate CpG ODN with intranasal (IN) delivery of purified hepatitis B surfa ce antigen (HBsAg), relative to and in combination with CZ Escherichia coli heat labile enterotoxin (LT), the B subunit of CT (CTB), and a nontoxic de rivative of LT (LTK63). Materials and Methods: BALB/c mice were immunized by IN administration of H BsAg, alone or combined with CT, LT, CTB, or LTK63, and/or CpG ODN, or non- CpG control ODN. In addition, the effect of low-or high-volume administrati on was assessed, in order to target upper respiratory or entire respiratory tract, respectively. HBsAg-specific systemic (immunoglobulins: IgG, IgG1, IgG2a in plasma) and mucosal (IgA in fecal, lung, vaginal, saliva, and gut samples) humoral responses, as well as cell-mediated immune responses inclu ding T-cell proliferation and cytokines (interleukins: IL-4, IL-5; interfer on: IFN-gamma) were evaluated. Results: CpG ODN, CT, and LT augmented anti-HBs titers equally, and more so than did CTB or LTK63. CpG ODN acted synergistically with CT and LT,but no t CTB or LTK63 to enhance anti-HBs titers. Nevertheless, CpG ODN induced a more Th1-like response for all combinations, compared with the same formula tion without CpG. Strength of induced systemic and mucosal immune responses was better with IN delivery of a large volume. A small volume required mul tiple administrations and higher doses of antigen and adjuvant for equal re sults. This suggests that delivery of antigen to the lung and/or diges-tive system is superior to delivery to the nasal cavity. Conclusions: Our results suggest that the synergy between CpG ODN and nativ e toxins (CT, LT) may depend on their enzymatic activity and that the lack of synergy with nontoxic derivatives (LTB, LTK63) arises, since they do not have enzymatic activity. Because both CT and LT are too toxic for use in h umans, it is possible that CpG ODN may be combined with bacterial toxin mut ants that retain some enzymatic activity to optimize immune augmentation.