Soluble multi-trimeric TNF superfamily ligand adjuvants enhance immune responses to a HIV-1 Gag DNA vaccine

Abstract

Background DNA vaccines remain an important component of HIV vaccination strategies, typically as part of a prime/boost vaccination strategy with viral vector or protein boost. A number of DNA prime/viral vector boost vaccines are currently being evaluated for both preclinical studies and in Phase I and Phase II clinical trials. These vaccines would benefit from molecular adjuvants that increase correlates of immunity during the DNA prime. While HIV vaccine immune correlates are still not well defined, there are a number of immune assays that have been shown to correlate with protection from viral challenge including CD8+ T cell avidity, antigen-specific proliferation, and polyfunctional cytokine secretion. Methodology and Principal Findings Recombinant DNA vaccine adjuvants composed of a fusion between Surfactant Protein D (SP-D) and either CD40 Ligand (CD40L) or GITR Ligand (GITRL) were previously shown to enhance HIV-1 Gag DNA vaccines. Here we show that similar fusion constructs composed of the TNF superfamily ligands (TNFSFL) 4-1BBL, OX40L, RANKL, LIGHT, CD70, and BAFF can also enhanced immune responses to a HIV-1 Gag DNA vaccine. BALB/c mice were vaccinated intramuscularly with plasmids expressing secreted Gag and SP-D-TNFSFL fusions. Initially, mice were analyzed 2 weeks or 7 weeks following vaccination to evaluate the relative efficacy of each SP-D-TNFSFL construct. All SP-D-TNFSFL constructs enhanced at least one Gag-specific immune response compared to the parent vaccine. Importantly, the constructs SP-D-4-1BBL, SP-D-OX40L, and SP-D-LIGHT enhanced CD8+ T cell avidity and CD8+/CD4+ T cell proliferation 7 weeks post vaccination. These avidity and proliferation data suggest that 4-1BBL, OX40L, and LIGHT fusion constructs may be particularly effective as vaccine adjuvants. Constructs SP-D-OX40L, SP-D-LIGHT, and SP-D-BAFF enhanced Gag-specific IL-2 secretion in memory T cells, suggesting these adjuvants can increase the number of self-renewing Gag-specific CD8+ and/or CD4+ T cells. Finally adjuvants SP-D-OX40L and SP-D-CD70 increased TH1 (IgG2a) but not TH2 (IgG1) antibody responses in the vaccinated animals. Surprisingly, the B cell-activating protein BAFF did not enhance anti-Gag antibody responses when given as an SP-D fusion adjuvant, but nonetheless enhanced CD4+ and CD8+ T cell responses. Conclusions We present evidence that various SP-D-TNFSFL fusion constructs can enhance immune responses following DNA vaccination with HIV-1 Gag expression plasmid. These data support the continued evaluation of SP-D-TNFSFL fusion proteins as molecular adjuvants for DNA and/or viral vector vaccines. Constructs of particular interest included SP-D-OX40L, SP-D-4-1BBL, SP-D-LIGHT, and SP-D-CD70. SP-D-BAFF was surprisingly effective at enhancing T cell responses, despite its inability to enhance anti-Gag antibody secretion.

DNA Vaccine Molecular Adjuvants SP-D-BAFF and SP-D-APRIL Enhance Anti-gp120 Immune Response and Increase HIV-1 Neutralizing Antibody Titers

“…Mean antibody titers were also increased with the combination of pgp140, pIL-12, and pSP-D-BAFF over those with gp140 antigen alone. Although we observed a trend toward higher antibody titers with the combination of the pIL-12 and pSP-D-BAFF adjuvants than with pIL-12 alone, the difference was not statistically significant, a result consistent with those of previous DNA vaccination studies (34,35).…”

“…In these studies, DNA vaccination with an SP-D-BAFF adjuvant enhanced cellular immune responses against an HIV-1 Gag antigen. However, we failed to observe a significant increase in antigenspecific antibody titers (34), possibly due to the requirement for additional costimulatory signals in the vaccine formulation.…”

“…Previous studies with DNA vaccines containing the pSP-D-BAFF adjuvant plasmid have shown only low levels of antibody titers relative to those with an antigen-only control vaccine (34). To explore methods of enhancing antibody titers from DNA vaccines, we performed a DNA vaccination experiment on mice with HIV-1 Gag DNA (as a model antigen) combined with plasmid pSP-D-BAFF or pIL-12, a single-chain IL12p70 construct expressing both IL-12p35 and IL-12p40 joined by a flexible linker (42).…”

“…The secreted HIV-1 Gag DNA plasmid (pGag; also called pScGag) has been described previously (37) and was provided by George N. Pavlakis. Plasmids coding for the 4-trimer soluble forms of murine SP-D-BAFF and SP-D-APRIL were generated in vectors pVAX1 and pcDNA3.1, respectively, as described previously (34). Constructs were cloned such that mouse SP-D from the N terminus to amino acid sequence ALFPDG was fused directly to the mouse TNF superfamily ligand (TNFSFL) extracellular domain, starting from the N-terminal amino acid sequences QLAALQ (BAFF) and MPASSP (APRIL).…”

“…These observations led us to hypothesize that soluble multitrimers of BAFF and APRIL, as opposed to single trimers, would induce optimal immune activation of B cells and enhance the overall immune response. We have shown previously that TNF superfamily ligands, including BAFF, APRIL, and CD40L, can be formed into soluble multitrimers by using the lung protein SP-D (surfactant protein D) as a scaffold (34)(35)(36). In these studies, DNA vaccination with an SP-D-BAFF adjuvant enhanced cellular immune responses against an HIV-1 Gag antigen.…”