Research ArticleIMMUNOLOGY

The adjuvant GLA-AF enhances human intradermal vaccine responses

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Science Advances  12 Sep 2018:
Vol. 4, no. 9, eaas9930
DOI: 10.1126/sciadv.aas9930
  • Fig. 1 Immunogenicity in mice.

    (A) Antibody endpoint titers. Following the boost immunization, the H5-specific total IgG, IgG1, and IgG2C titers were determined. As expected, the adjuvants enhanced titers across groups with high IgG2C titers, which are indicative of a TH1-type immune response, observed in groups containing the TLR4 agonist GLA. (B) Bone marrow enzyme-linked immunosorbent spots (ELISPOTs). Similarly, H5-specific antibody–secreting plasma cells were increased with H5-VLP + GLA-AF. ****P < 0.0001 for H5-VLP + GLA-AF versus other groups. ***P < 0.001 for H5-VLP + GLA-liposomes versus other groups not containing GLA. (C) HAI titers. HAI titers are the standard measurement for functional antibody in flu. The A/Indonesia/5/2005 virus strain was not available for testing; however, two different clades, either the “homologous” clade 2.3.4 A/Anhui/1/2005 or the “drifted” clade 2.1 A/Duck/Hunan/795/2002, were tested to determine whether these adjuvants could induce cross-reactive titers. ****P < 0.0001 for H5-VLP + GLA-AF versus all other groups; ***P < 0.001 for H5-VLP + GLA AF versus all other groups.

  • Fig. 2 Efficacy study in ferrets.

    Ferrets were immunized (either intradermally or intramuscularly, as indicated) once at day 0 and challenged with the heterologous A/Vietnam/1203/04 H5N1 strain of flu virus 3 weeks later. (A) Survival data. All ferrets that were immunized intradermally with H5-VLP/GLA-AF survived the heterologous challenge. All the ferrets succumbed to infection in the saline-treated group and in the group that received an intramuscular (IM) immunization with the unadjuvanted H5-VLP, whereas one ferret survived in the intradermally immunized group that received unadjuvanted H5-VLP. (B) Viral titers. Nasal swabs were performed to measure virus in the upper airways on the days indicated. Paralleling the survival data, the adjuvanted groups appeared to control the infection and rapidly demonstrated lower viral titers than those found in all the unadjuvanted groups. (C) Clinical scores. Clinical signs of morbidity including lethargy, body temperature, anorexia, and dyspnea were monitored daily. PFU, plaque-forming units.

  • Fig. 3 HAI titers from a human clinical study.

    The immunogenicity of the vaccine was evaluated by comparing HAI antibody responses of subjects in each treatment group. GMTs of HAI antibody responses were evaluated on days 0, 21, and 42, as shown below the x axis. Each group of three bars corresponds to these timepoints. Error bars indicate 95% confidence intervals. *P < 0.05 between indicated groups. Statistically significant differences were only achieved at the day 42 timepoint.

  • Fig. 4 Diversity of the immune response.

    HAI antibody responses of subjects in each treatment group were determined against a homologous virus, A/Indonesia/5/2005 (H5N1); a drifted virus within the homologous clade, A/Anhui/1/2005 (H5N1); and a drifted virus outside the clade, A/Vietnam/1203/2004 (H5N1). GMTs of HAI antibody responses were compared between days 0 and 42 to calculate geometric mean increases (GMI, top row), and the percentage of subjects seroconverting was plotted below. Adjuvanted injection groups are drawn in blue.

    Table 2 Seroconversion and seroprotection rates by serum HAI response in humans.

    Seroconversion was defined as the proportion of subjects in a given treatment group with either a ≥4-fold increase in HAI titers or a rise of an undetectable HAI titer (that is, <8) before vaccination to an HAI titer of ≥32 after vaccination. Seroprotection was defined as the proportion of subjects in a given treatment group attaining an HAI titer of ≥32 after vaccination. Statistical differences were found among the treatment groups at day 42 (P = 0.036 for seroconversion and P = 0.014 for seroprotection), but pairwise comparisons with multiplicity adjustments failed to reach significance.

    TimepointH5-VLP + GLA-AF (ID)H5-VLP + GLA-AF (IM)H5-VLP (ID)H5-VLP + alum (IM)Control vaccine
    Seroconversion
    Day 2130.0%19.0%30.0%20.0%21.1%
    Day 4265.0%84.2%52.6%83.3%43.8%
    Seroprotection
    Day 2135.0%23.8%30.0%25.0%26.3%
    Day 4270.0%89.5%52.6%88.9%50.0%
  • Table 1 Treatment assignments.

    The table depicts the number of subjects in each group and the treatment regimen, dose, and dose timing for each group. One hundred subjects were enrolled across three study sites and randomly assigned to one of five treatment groups for a total goal of 20 subjects in each of the five groups. An additional five subjects were screened and enrolled to replace five subjects who withdrew from the study for reasons other than dose-limiting toxicity before day 21. A total of 105 subjects were enrolled, as shown in the table.

    GroupTreatment
    assignment
    RouteVolumeTiming of
    injections
    N = 100*N = 105
    120 μg of H5-VLP +
    2.5 of μg GLA-AF
    ID0.2 mlDays 0 and 212020
    220 μg of H5-VLP +
    2.5 μg of GLA-AF
    IM0.2 mlDays 0 and 212023
    320 μg of H5-VLP
    alone
    ID0.2 mlDays 0 and 212022
    420 μg of H5-VLP +
    1 mg of alhydrogel
    IM0.2 mlDays 0 and 212020
    590 μg of influenza
    virus vaccine, H5N1
    (Sanofi Pasteur)
    IM1.0 mlDays 0 and 212020

    *Planned enrollment.

    †Total including replacements.

    ‡Al+3 content of 1 mg of alhydrogel = 0.5 mg.

    Supplementary Materials

    • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/4/9/eaas9930/DC1

      Section S1. Inclusion criteria

      Section S2. Exclusion criteria

      Fig. S1. Safety assessments in the guinea pig.

      Fig. S2. Assessment schedule.

      Table S1. Schedule of study visits and procedures.

      Table S2. Subjects with adverse events by study period, injection interval, severity, and relatedness to study injection (safety population).

      Table S3. Subjects with AEs occurring in ≥5 subjects in study period I (safety population).

    • Supplementary Materials

      This PDF file includes:

      • Section S1. Inclusion criteria
      • Section S2. Exclusion criteria
      • Fig. S1. Safety assessments in the guinea pig.
      • Fig. S2. Assessment schedule.
      • Table S1. Schedule of study visits and procedures.
      • Table S2. Subjects with adverse events by study period, injection interval, severity, and relatedness to study injection (safety population).
      • Table S3. Subjects with AEs occurring in ≥5 subjects in study period I (safety population).

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