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P41.13
Single and Combined Vaccination Modalities Result in Distinct Immunological Profiles in HIV-1 gp140-immunised Mice

Single and combined vaccination modalities result in distinct immunological profiles in HIV-1 gp140-immunised mice.

An effective HIV vaccine will likely require induction of both mucosal and systemic cellular and humoral immune responses.

Recent studies have indicated that co-immunisation of protein antigens and DNA can induce such responses.

We investigated whether simultaneous electroporated DNA administered by the intramuscular (IM) route and protein vaccinations by intranasal (IN) and IM routes could be combined to induce mucosal and systemic cellular and humoral immune responses to a model HIV-1 CN54 gp140 antigen in mice.

Seven groups of 8 Balb/c mice were immunised 3 times either by a single vaccination route, or co-immunised as described in table 1.

DNA = intramuscular injection with HIV-1 gp140-encoding plasmid with electroporation,

IM = Intramuscular injection with HIV-1 CN54 gp140 Env protein with MPLA,

IN = Intranasal delivery of HIV-1 CN54 gp140 Env protein with MPLA.  

Animals were sampled for serum and vaginal lavage one week after each vaccination, antigen specific and total IgG and IgA were measured in serum and vagina samples by ELISA.

IFNg responses were assessed by ELIspot (MABTech) in splenocytes one week after the final vaccination.  Cells were stimulated with gp140 peptide pools for 18 hours at 37°C.  Peptides were divided into two pools, pool 1 covering amino acids 1-78, and pool 2 covering amino acids 79-156 of HIV-1 CN54 gp140, and consisted of 15-mers, overlapping by 11 aa.

Antigen-specific cytokine responses were measured in splenocytes using Bio-Plex Pro mouse cytokine standard 23-plex, (Bio-Rad).  Freshly isolated splenocytes were stimulated with recombinant gp140 protein, supernatant was harvested after 3 days of culture and analysed for cytokine production, according to manufacturers instructions. 

Analysis of antigen specific IgG and IgA revealed that DNA+IN co-immunisation induced significantly higher antibody responses than either DNA alone or DNA + IM co-immunisation, (a, b).

IgG subclass analysis measuring antigen specific IgG1 and IgG2a, as a surrogate of Th2 and Th1 responses respectively, indicated DNA induced Th1 responses, whereasTh2 responses were induced by IN vaccination.  Co-immunisation with DNA + IN lead to a more balanced response (c, d). 

Induction of antigen-specific IgG in the vagina was observed in all protocols, with the highest levels seen in IN and triple vaccination groups (a).  Antigen-specific IgA was observed in protocols incorporating intranasal vaccination and was highest in IN and DNA+IN groups. 

DNA vaccination was the most effective at inducing IFNg+ T cells , whilst in protein only regimes (IM and IN + IM) vaccination resulted in low level cellular responses.

Co-immunisation protocols incorporating DNA and IN vaccination induced similar levels of IFNg+ T cells, but were significantly reduced compared to DNA alone in some cases. 

Cytokine secretion was measured in antigen stimulated splenocytes by Luminex.

DNA and DNA+IN vaccination induced IL-4 (a) and IL-13 (b), whilst IL-17 was observed in IN and DNA + IN groups (c).

The b-chemokines MIP-1a and MIP-1b, and MCP-1 were observed in DNA, IN, DNA+IN and IN+IM groups groups, whilst these were not induced by protocols incorporating IM vaccination. 

These data suggest that while IN vaccination alone is able to elicit both cellular and humoral responses, co-administration with vaccine-antigen homologous DNA can tailor this response toward a balanced humoral response, broadening the cellular inflammatory cytokine profile while eliciting high-levels of antigen-specific antibody. This work provides information on how to generate differential responses within the same vaccination visit, simplifying immunization schedules.