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Bolstering Components of the Immune Response Compromised by Prior Exposure to Adenovirus: Guided Formulation Development for a Nasal Ebola Vaccine.

Choi JH, Schafer SC, Freiberg AN, Croyle MA - Mol. Pharm. (2015)

Bottom Line: An amphiphilic polymer (F16, FW ∼39,000) significantly improved transgene expression in the presence of anti-Ad neutralizing antibodies (NAB) at concentrations of 5 times the 50% neutralizing dose (ND50).Multiparameter flow cytometry revealed that F16 induced significantly more polyfunctional antigen-specific CD8+ T cells simultaneously producing IFN-γ, IL-2, and TNF-α than other test formulations.These effects were not compromised by PEI.

View Article: PubMed Central - PubMed

Affiliation: †Division of Pharmaceutics, College of Pharmacy, The University of Texas at Austin, Austin, Texas 78712, United States.

ABSTRACT
The severity and longevity of the current Ebola outbreak highlight the need for a fast-acting yet long-lasting vaccine for at-risk populations (medical personnel and rural villagers) where repeated prime-boost regimens are not feasible. While recombinant adenovirus (rAd)-based vaccines have conferred full protection against multiple strains of Ebola after a single immunization, their efficacy is impaired by pre-existing immunity (PEI) to adenovirus. To address this important issue, a panel of formulations was evaluated by an in vitro assay for their ability to protect rAd from neutralization. An amphiphilic polymer (F16, FW ∼39,000) significantly improved transgene expression in the presence of anti-Ad neutralizing antibodies (NAB) at concentrations of 5 times the 50% neutralizing dose (ND50). In vivo performance of rAd in F16 was compared with unformulated virus, virus modified with poly(ethylene) glycol (PEG), and virus incorporated into poly(lactic-co-glycolic) acid (PLGA) polymeric beads. Histochemical analysis of lung tissue revealed that F16 promoted strong levels of transgene expression in naive mice and those that were exposed to adenovirus in the nasal cavity 28 days prior to immunization. Multiparameter flow cytometry revealed that F16 induced significantly more polyfunctional antigen-specific CD8+ T cells simultaneously producing IFN-γ, IL-2, and TNF-α than other test formulations. These effects were not compromised by PEI. Data from formulations that provided partial protection from challenge consistently identified specific immunological requirements necessary for protection. This approach may be useful for development of formulations for other vaccine platforms that also employ ubiquitous pathogens as carriers like the influenza virus.

No MeSH data available.


Related in: MedlinePlus

Formulated vaccines improve the anti-Ebolaglycoprotein antibodyresponse. Serum collected from individual mice 42 days after immunizationwas screened for total IgG and IgG isotypes by ELISA. (A) Antibodyprofile for naive mice. Naive B10.Br mice were given 1 × 108 particles of Ad-CAGoptZGP suspended in formulation or 4.6mg of PLGA microspheres containing the virus in KPBS by the intranasalroute. (B) Antibody profile for mice with pre-existing immunity toadenovirus. Pre-existing immunity was established by instillationof a dose of 5 × 1010 particles of AdNull in the nasalpassages of B10.Br mice 28 days prior to immunization with formulatedvaccines. In both panels, the average optical density read from samplesobtained from each treatment group are presented to serve as a measureof relative antibody concentration and data reported as average values± the standard error of the mean obtained from three separateexperiments each containing 5 mice per treatment. In each panel, theasterisk indicates a significant difference with respect to naive,immunized animals. *p < 0.05, **p < 0.01, ***p < 0.001, one-way ANOVA, Bonferroni/Dunnpost hoc analysis.
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fig4: Formulated vaccines improve the anti-Ebolaglycoprotein antibodyresponse. Serum collected from individual mice 42 days after immunizationwas screened for total IgG and IgG isotypes by ELISA. (A) Antibodyprofile for naive mice. Naive B10.Br mice were given 1 × 108 particles of Ad-CAGoptZGP suspended in formulation or 4.6mg of PLGA microspheres containing the virus in KPBS by the intranasalroute. (B) Antibody profile for mice with pre-existing immunity toadenovirus. Pre-existing immunity was established by instillationof a dose of 5 × 1010 particles of AdNull in the nasalpassages of B10.Br mice 28 days prior to immunization with formulatedvaccines. In both panels, the average optical density read from samplesobtained from each treatment group are presented to serve as a measureof relative antibody concentration and data reported as average values± the standard error of the mean obtained from three separateexperiments each containing 5 mice per treatment. In each panel, theasterisk indicates a significant difference with respect to naive,immunized animals. *p < 0.05, **p < 0.01, ***p < 0.001, one-way ANOVA, Bonferroni/Dunnpost hoc analysis.

Mentions: We have previouslyfound that prior exposure to adenovirus significantly reduced antibody-mediatedimmune response to Ebola glycoprotein in mice and guinea pigs.14 More specifically, we also found that a reductionin glycoprotein-specific IgG1 antibodies correlated with poor survivalafter challenge with rodent-adapted Ebola. Thus, we evaluated totalanti-Ebola glycoprotein-specific immunoglobulin (IgG) and IgG isotypesin serum to determine if each formulation could counterbalance theeffect of prior mucosal exposure to adenovirus on B cell mediatedimmune responses (Figure 4). Each formulationsignificantly increased the amount of each antibody isotype specificfor Ebola glycoprotein (GP) in naive mice (Figure 4A). The IgG2a level in mice given PLGA microspheres was theonly deviation from this trend as it was reduced by 29.9% with respectto unformulated virus (Figure 4A). Prior exposureto adenovirus significantly reduced each anti-Ebola GP-specific IgGisotype evaluated (Figure 4B). Although IgG2blevels in samples collected from mice immunized with the F3 formulationdoubled, IgG1 and IgG2a levels were not significantly different fromthose seen in animals given unformulated virus. IgG1 and IgG2b levelsin mice immunized with PLGA microspheres were 9.5 and 1.3 times thosefound in samples from mice given unformulated vaccine (Figure 4B). PEI to adenovirus reduced IgG2b levels by 45.9%in mice given PEGylated vaccine. IgG1 and IgG2a could not be detectedin serum of mice immunized with this preparation. Trace levels ofEbola GP-specific IgM antibodies were found in serum from mice giventhe PLGA and PEGylated preparations.


Bolstering Components of the Immune Response Compromised by Prior Exposure to Adenovirus: Guided Formulation Development for a Nasal Ebola Vaccine.

Choi JH, Schafer SC, Freiberg AN, Croyle MA - Mol. Pharm. (2015)

Formulated vaccines improve the anti-Ebolaglycoprotein antibodyresponse. Serum collected from individual mice 42 days after immunizationwas screened for total IgG and IgG isotypes by ELISA. (A) Antibodyprofile for naive mice. Naive B10.Br mice were given 1 × 108 particles of Ad-CAGoptZGP suspended in formulation or 4.6mg of PLGA microspheres containing the virus in KPBS by the intranasalroute. (B) Antibody profile for mice with pre-existing immunity toadenovirus. Pre-existing immunity was established by instillationof a dose of 5 × 1010 particles of AdNull in the nasalpassages of B10.Br mice 28 days prior to immunization with formulatedvaccines. In both panels, the average optical density read from samplesobtained from each treatment group are presented to serve as a measureof relative antibody concentration and data reported as average values± the standard error of the mean obtained from three separateexperiments each containing 5 mice per treatment. In each panel, theasterisk indicates a significant difference with respect to naive,immunized animals. *p < 0.05, **p < 0.01, ***p < 0.001, one-way ANOVA, Bonferroni/Dunnpost hoc analysis.
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fig4: Formulated vaccines improve the anti-Ebolaglycoprotein antibodyresponse. Serum collected from individual mice 42 days after immunizationwas screened for total IgG and IgG isotypes by ELISA. (A) Antibodyprofile for naive mice. Naive B10.Br mice were given 1 × 108 particles of Ad-CAGoptZGP suspended in formulation or 4.6mg of PLGA microspheres containing the virus in KPBS by the intranasalroute. (B) Antibody profile for mice with pre-existing immunity toadenovirus. Pre-existing immunity was established by instillationof a dose of 5 × 1010 particles of AdNull in the nasalpassages of B10.Br mice 28 days prior to immunization with formulatedvaccines. In both panels, the average optical density read from samplesobtained from each treatment group are presented to serve as a measureof relative antibody concentration and data reported as average values± the standard error of the mean obtained from three separateexperiments each containing 5 mice per treatment. In each panel, theasterisk indicates a significant difference with respect to naive,immunized animals. *p < 0.05, **p < 0.01, ***p < 0.001, one-way ANOVA, Bonferroni/Dunnpost hoc analysis.
Mentions: We have previouslyfound that prior exposure to adenovirus significantly reduced antibody-mediatedimmune response to Ebola glycoprotein in mice and guinea pigs.14 More specifically, we also found that a reductionin glycoprotein-specific IgG1 antibodies correlated with poor survivalafter challenge with rodent-adapted Ebola. Thus, we evaluated totalanti-Ebola glycoprotein-specific immunoglobulin (IgG) and IgG isotypesin serum to determine if each formulation could counterbalance theeffect of prior mucosal exposure to adenovirus on B cell mediatedimmune responses (Figure 4). Each formulationsignificantly increased the amount of each antibody isotype specificfor Ebola glycoprotein (GP) in naive mice (Figure 4A). The IgG2a level in mice given PLGA microspheres was theonly deviation from this trend as it was reduced by 29.9% with respectto unformulated virus (Figure 4A). Prior exposureto adenovirus significantly reduced each anti-Ebola GP-specific IgGisotype evaluated (Figure 4B). Although IgG2blevels in samples collected from mice immunized with the F3 formulationdoubled, IgG1 and IgG2a levels were not significantly different fromthose seen in animals given unformulated virus. IgG1 and IgG2b levelsin mice immunized with PLGA microspheres were 9.5 and 1.3 times thosefound in samples from mice given unformulated vaccine (Figure 4B). PEI to adenovirus reduced IgG2b levels by 45.9%in mice given PEGylated vaccine. IgG1 and IgG2a could not be detectedin serum of mice immunized with this preparation. Trace levels ofEbola GP-specific IgM antibodies were found in serum from mice giventhe PLGA and PEGylated preparations.

Bottom Line: An amphiphilic polymer (F16, FW ∼39,000) significantly improved transgene expression in the presence of anti-Ad neutralizing antibodies (NAB) at concentrations of 5 times the 50% neutralizing dose (ND50).Multiparameter flow cytometry revealed that F16 induced significantly more polyfunctional antigen-specific CD8+ T cells simultaneously producing IFN-γ, IL-2, and TNF-α than other test formulations.These effects were not compromised by PEI.

View Article: PubMed Central - PubMed

Affiliation: †Division of Pharmaceutics, College of Pharmacy, The University of Texas at Austin, Austin, Texas 78712, United States.

ABSTRACT
The severity and longevity of the current Ebola outbreak highlight the need for a fast-acting yet long-lasting vaccine for at-risk populations (medical personnel and rural villagers) where repeated prime-boost regimens are not feasible. While recombinant adenovirus (rAd)-based vaccines have conferred full protection against multiple strains of Ebola after a single immunization, their efficacy is impaired by pre-existing immunity (PEI) to adenovirus. To address this important issue, a panel of formulations was evaluated by an in vitro assay for their ability to protect rAd from neutralization. An amphiphilic polymer (F16, FW ∼39,000) significantly improved transgene expression in the presence of anti-Ad neutralizing antibodies (NAB) at concentrations of 5 times the 50% neutralizing dose (ND50). In vivo performance of rAd in F16 was compared with unformulated virus, virus modified with poly(ethylene) glycol (PEG), and virus incorporated into poly(lactic-co-glycolic) acid (PLGA) polymeric beads. Histochemical analysis of lung tissue revealed that F16 promoted strong levels of transgene expression in naive mice and those that were exposed to adenovirus in the nasal cavity 28 days prior to immunization. Multiparameter flow cytometry revealed that F16 induced significantly more polyfunctional antigen-specific CD8+ T cells simultaneously producing IFN-γ, IL-2, and TNF-α than other test formulations. These effects were not compromised by PEI. Data from formulations that provided partial protection from challenge consistently identified specific immunological requirements necessary for protection. This approach may be useful for development of formulations for other vaccine platforms that also employ ubiquitous pathogens as carriers like the influenza virus.

No MeSH data available.


Related in: MedlinePlus