The adjuvant GLA-AF enhances human intradermal vaccine responses

Abstract

Adjuvants are key to shaping the immune response to vaccination, but to date, no adjuvant suitable for human use has been developed for intradermal vaccines. These vaccines could be self-administered and sent through the mail as they do not require long needles or technical expertise in immunization. In the event of a pandemic outbreak, this approach could alleviate the congregation of patients in health centers and thus reduce the potential of these centers to enhance the spread of lethal infection. A reliable and potent vaccine system for self-administration would provide an effective countermeasure for delivery through existing product distribution infrastructure. We report results from preclinical and clinical trials that demonstrate the feasibility of an adjuvanted, intradermal vaccine that induced single shot protection in ferrets and seroprotection in humans against one of the more lethal strains of pandemic flu, Indonesia H5N1. In the human trial, the vaccine was safe and clinical responses were above approvable endpoints for a protective flu vaccine. Inclusion of a modern TLR4 (Toll-like receptor 4) agonist-based adjuvant was critical to the development of the response in the intradermal groups. In humans, this is the first report of a safe and effective intradermal adjuvant, GLA-AF (aqueous formulation of glucopyranosyl lipid adjuvant), and provides a future path for developing a vaccine-device combination for distribution by mail and self-administration in case of a pandemic.

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 T_H1-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.