Adjuvant effect of DMXAA can improve potency of influenza SV vaccine and protect mice from a lethal challenge.

<p>C57BL/6 mice were immunized twice i.d. with 0.75 µg SV and 100 µg DMXAA, and their sera were assessed for anti-SV (A) tIgG, (B) IgG1 and (C) IgG2c antibodies. ** <i>P</i><0.01 by Students t-test. Results presented are pooled from two separate experiments. (D) C57BL/6 mice that had received two i.d. injections of PBS (open square), 100 µg DMXAA alone (open circle), 0.75 µg SV alone (grey-filled circle) or SV plus DMXAA (black filled circle) were challenged with a lethal dose of A/Puerto Rico/8/34 (PR) (H1N1) 7 days after the final immunization (n = 6 mice per experimental group). Results presented are pooled from two separate experiments. Survival rates were recorded daily and statistical analyses were performed using the log-rank (Mantel-Cox) test where ** and *** denotes p<0.01 and p<0.001 respectively (E) The rate of weight-loss by the challenged mice were monitored and presented as an average percentage of the initial base weight ± standard error. * denotes p<0.05 vs SV alone by Student's T-test.</p

Adjuvant effects of DMXAA require type-I-IFN responses induced by IRF3 activation.

<p>Anti-OVA (A) tIgG antibody responses of WT C57BL/6, <i>Ifnar^-/-</i> and <i>Irf3^-/-</i> mice immunized twice i.d. with 10 µg OVA plus 100 µg DMXAA. (B) tIgG, (C) IgG1 and (D) IgG2a antibody titers against OVA in WT BALB/c and <i>Il-33^-/-</i> mice immunized twice i.d. with 10 µg OVA plus 100 µg DMXAA. Results presented are pooled titers from two separate experiments. <i>In-vitro</i> cultured DCs derived from WT, <i>Ifnar^-/-</i> and <i>Irf3^-/-</i> mice were stimulated with DMXAA (2.5 µg/ml), LPS (1 µg/ml) or lipofectamine complexed c-di-GMP (10 µg/ml) for 6 h before the supernatant were collected and analysed for (E) IFNβ, (F) IL-6 and (G) TNFα secretion and CD11c⁺ cells were analysed for CD86 expression (H). Results presented are average of triplicate conditions ± SD and are representative of three separate experiments. *** <i>P</i><0.001 one-way ANOVA with Bonferroni's post-test.</p

DMXAA acts as a potent adjuvant.

<p>(A) Anti-OVA tIgG titers of C57BL/6 mice immunized with 100 µg OVA plus the indicated doses of DMXAA (µg). (B) Anti-OVA tIgG titers of C57BL/6 mice 21 days and 150 days after immunization with 100 µg OVA and 100 µg DMXAA. (C–E) C57BL/6 mice were immunized twice i.d. with 100 µg OVA plus DMXAA (100 µg), Alum (665 µg) or CpG DNA (25 µg) and the induction of (C) tIgG, (D) IgG1 and (E) IgG2c antibody responses against OVA were assessed. (F) IFN-γ secretion from splenocytes of immunized mice that were stimulated for 48 h with CD4 and CD8 OVA peptides and whole OVA protein. (G) Anti-OVA tIgG titers of C57BL/6 mice injected i.v. with 200 µg anti-CD4 (GK1.5) antibodies prior to immunization with 100 µg OVA and 100 µg DMXAA. Results presented are representatives of three separate experiments. *<0.05, ** <i>P</i><0.01, *** <i>P</i><0.001 by Students t-test when comparing between two groups and one-way ANOVA with Bonferroni's post-test when comparing three or more groups.</p