Plasmid chemokines and colony-stimulating factors enhance the immunogenicity of DNA priming-viral vector boosting human immunodeficiency virus type 1 vaccines

Abstract

Heterologous "prime-boost" regimens that involve priming with plasmid DNA vaccines and boosting with recombinant viral vectors have been shown to elicit potent virus-specific cytotoxic T-lymphocyte responses. Increasing evidence, however, suggests that the utility of recombinant viral vectors in human populations will be significantly limited by preexisting antivector immunity. Here we demonstrate that the coadministration of plasmid chemokines and colony-stimulating factors with plasmid DNA vaccines markedly increases the immunogenicity of DNA prime-recombinant adenovirus serotype 5 (rAd5) boost and DNA prime-recombinant vaccinia virus (rVac) boost vaccine regimens in BALB/c mice. In mice with preexisting anti-Ad5 immunity, priming with the DNA vaccine alone followed by rAd5 boosting elicited only marginal immune responses. In contrast, cytokine-augmented DNA vaccine priming followed by rAd5 vector boosting was able to generate potent immune responses in mice with preexisting anti-Ad5 immunity. These data demonstrate that plasmid cytokines can markedly improve the immunogenicity of DNA prime-viral vector boost vaccine strategies and can partially compensate for antivector immunity.

FIG. 1.

Magnitude, kinetics, and variability of DNA vaccine-elicited CD8⁺ T-lymphocyte responses. Groups of mice (n = 20 per group) were immunized once with either the gp120 DNA vaccine or the gp120/GM-CSF/MIP-1α DNA vaccine. Vaccine-elicited cellular immune responses were measured by tetramer binding to CD8⁺ T lymphocytes. Means and standard errors of responses for each group are shown.

FIG. 2.

Plasmid cytokines augment CD8⁺ T-lymphocyte responses elicited by DNA prime-rAd5 boost regimens. (A) Groups of mice were primed with the gp120 DNA vaccine and then boosted with a dose titration of rAd5-gp140ΔCFI. (B) Groups of mice (n = 4 per group) were primed with sham plasmid, the gp120 DNA vaccine alone, or the gp120 DNA vaccine with the plasmid cytokine adjuvant GM-CSF, MIP-1α, GM-CSF plus MIP-1α, MCP-1, or GM-CSF plus MCP-1 and then boosted on day 45 with 10⁶ particles of rAd5. Vaccine-elicited cellular immune responses were measured by tetramer binding to CD8⁺ T lymphocytes. Arrows indicate immunizations.

FIG. 3.

Plasmid cytokines augment CD8⁺ T-lymphocyte responses elicited by DNA prime-rVac boost regimens. (A) Groups of mice were primed with the gp120 DNA vaccine and then boosted with a dose titration of rVac-gp160. (B) Groups of mice (n = 4 per group) were primed with sham plasmid, the gp120 DNA vaccine alone, or the gp120 DNA vaccine with the plasmid cytokine adjuvant GM-CSF, MIP-1α, GM-CSF plus MIP-1α, MCP-1, or GM-CSF plus MCP-1 and then boosted on day 45 with 10⁴ PFU of rVac-gp160. Vaccine-elicited cellular immune responses were measured by tetramer binding to CD8⁺ T lymphocytes. Arrows indicate immunizations.

FIG.4.

Functional analysis of vaccine-elicited immune responses. Groups of mice (n = 4 per group) were primed with sham plasmid, the gp120 DNA vaccine alone, or the gp120 DNA vaccine with the plasmid cytokine adjuvant GM-CSF, MIP-1α, or GM-CSF plus MIP-1α. These mice were then boosted at week 8 with the gp120 DNA vaccine aloneand at week 24 with 10⁶ particles of rAd5-gp140ΔCFI. Vaccine-elicited cellular immune responses were assessed by (A) tetramer binding to CD8⁺ T lymphocytes, (B) fresh ex vivo cytotoxicity assays, and pooled peptide and P18-specific IFN-γ ELISPOT assays using either unfractionated (C), CD4-depleted (D), or CD8-depleted (E) splenocytes. SFC, spot-forming cells. (F) Vaccine-elicited humoral immune responses were assessed by a gp120 ELISA prior to immunization (white bars) and 28 days after the rAd5-gp140ΔCFI boost (black bars).

FIG. 5.

Preexisting anti-Ad5 immunity suppresses DNA/rAd5 immune responses. Groups of mice (n = 4 per group) were preimmunized with either saline (A) or 5 × 10⁹ particles of empty Ad5 (B and C) to induce anti-Ad5 immunity. Mice were primed with the gp120 DNA vaccine (A and B) or the gp120/GM-CSF/MIP-1α DNA vaccine (C) and then boosted with various doses of rAd5-gp140ΔCFI. Vaccine-elicited cellular immune responses were measured by tetramer binding to CD8⁺ T lymphocytes following the rAd5 boost. (D) Anti-Ad5 antibody responses were measured by ELISA prior to (white bars) and 28 days after (black bars) preimmunization with saline or Ad5. (E) Vaccine-elicited humoral immune responses were assessed by a gp120 ELISA. ND, not done.