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Hepatitis C VLPs delivered to dendritic cells by a TLR2 targeting lipopeptide results in enhanced antibody and cell-mediated responses.

Chua BY, Johnson D, Tan A, Earnest-Silveira L, Sekiya T, Chin R, Torresi J, Jackson DC - PLoS ONE (2012)

Bottom Line: Although many studies provide strong evidence supporting the development of HCV virus-like particle (VLP)-based vaccines, the fact that heterologous viral vectors and/or multiple dosing regimes are required to induce protective immunity indicates that it is necessary to improve their immunogenicity.While co-formulation of this lipopeptide with VLPs only resulted in marginal improvements in dendritic cell (DC) uptake, its ability to concomitantly induce DC maturation at very small doses is a feature not observed using VLPs alone or in the presence of an aluminium hydroxide-based adjuvant (Alum).These results suggest overall that the immunogenicity of HCV VLPs can be significantly improved by the addition of this novel adjuvant by targeting their delivery to DCs and could therefore constitute a viable vaccine strategy for the treatment of HCV.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, The University of Melbourne, Parkville, Victoria, Australia.

ABSTRACT
Although many studies provide strong evidence supporting the development of HCV virus-like particle (VLP)-based vaccines, the fact that heterologous viral vectors and/or multiple dosing regimes are required to induce protective immunity indicates that it is necessary to improve their immunogenicity. In this study, we have evaluated the use of an anionic self-adjuvanting lipopeptide containing the TLR2 agonist Pam(2)Cys (E(8)Pam(2)Cys) to enhance the immunogenicity of VLPs containing the HCV structural proteins (core, E1 and E2) of genotype 1a. While co-formulation of this lipopeptide with VLPs only resulted in marginal improvements in dendritic cell (DC) uptake, its ability to concomitantly induce DC maturation at very small doses is a feature not observed using VLPs alone or in the presence of an aluminium hydroxide-based adjuvant (Alum). Dramatically improved VLP and E2-specific antibody responses were observed in VLP+E(8)Pam(2)Cys vaccinated mice where up to 3 doses of non-adjuvanted or traditionally alum-adjuvanted VLPs was required to match the antibody titres obtained with a single dose of VLPs formulated with this lipopeptide. This result also correlated with significantly higher numbers of specific antibody secreting cells that was detected in the spleens of VLP+E(8)Pam(2)Cys vaccinated mice and greater ability of sera from these mice to neutralise the binding and uptake of VLPs by Huh7 cells. Moreover, vaccination of HLA-A2 transgenic mice with this formulation also induced better VLP-specific IFN-γ-mediated responses compared to non-adjuvanted VLPs but comparable levels to that achieved when coadministered with complete freund's adjuvant. These results suggest overall that the immunogenicity of HCV VLPs can be significantly improved by the addition of this novel adjuvant by targeting their delivery to DCs and could therefore constitute a viable vaccine strategy for the treatment of HCV.

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Cell-mediated responses elicited by vaccination.HLA-A2kb transgenic mice (n = 3 per group) were inoculated (100 µl) subcutaneously at the base of the tail on days 0 and 14 with 30 µg of VLPs alone, emulsified with an equal amount of complete freund’s adjuvant (CFA) or pre-mixed with 30 µg of E8Pam2Cys in saline. Splenocytes were obtained 28 days later and restimulated for 7 days in the presence of 10 µg VLPs or an irrelevant HCV-derived HLA-A2-restricted epitope not part of the VLP construct. The frequency of peptide-specific T cells producing IFN-γ was determined in an ELISPOT assay. Each bar represents the average number of IFN-γ producing T cells and standard deviation in each group after subtracting non-specific responses from corresponding samples stimulated with the irrelevant peptide.
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pone-0047492-g006: Cell-mediated responses elicited by vaccination.HLA-A2kb transgenic mice (n = 3 per group) were inoculated (100 µl) subcutaneously at the base of the tail on days 0 and 14 with 30 µg of VLPs alone, emulsified with an equal amount of complete freund’s adjuvant (CFA) or pre-mixed with 30 µg of E8Pam2Cys in saline. Splenocytes were obtained 28 days later and restimulated for 7 days in the presence of 10 µg VLPs or an irrelevant HCV-derived HLA-A2-restricted epitope not part of the VLP construct. The frequency of peptide-specific T cells producing IFN-γ was determined in an ELISPOT assay. Each bar represents the average number of IFN-γ producing T cells and standard deviation in each group after subtracting non-specific responses from corresponding samples stimulated with the irrelevant peptide.

Mentions: The ability of VLP formulations containing E8Pam2Cys to induce a cell-meditated immune response was examined by inoculating transgenic mice expressing the MHC class I (HLA-A2) allele but not endogenous H-2Db molecules [28]. Control transgenic animals were inoculated with VLPs alone or VLPs emulsified with an equal amount of complete Freund’s adjuvant (CFA). Splenocytes from vaccinated animals were obtained 28 days post-inoculation and re-stimulated with antigen in vitro. The results (Figure 6) of an ELISPOT assay carried out revealed significantly higher numbers of HCV VLP-specific IFN-γ producing cells in the spleens of mice inoculated with VLPs in the presence of E8Pam2Cys or VLPs emulsified in CFA than in those that received VLPs alone.


Hepatitis C VLPs delivered to dendritic cells by a TLR2 targeting lipopeptide results in enhanced antibody and cell-mediated responses.

Chua BY, Johnson D, Tan A, Earnest-Silveira L, Sekiya T, Chin R, Torresi J, Jackson DC - PLoS ONE (2012)

Cell-mediated responses elicited by vaccination.HLA-A2kb transgenic mice (n = 3 per group) were inoculated (100 µl) subcutaneously at the base of the tail on days 0 and 14 with 30 µg of VLPs alone, emulsified with an equal amount of complete freund’s adjuvant (CFA) or pre-mixed with 30 µg of E8Pam2Cys in saline. Splenocytes were obtained 28 days later and restimulated for 7 days in the presence of 10 µg VLPs or an irrelevant HCV-derived HLA-A2-restricted epitope not part of the VLP construct. The frequency of peptide-specific T cells producing IFN-γ was determined in an ELISPOT assay. Each bar represents the average number of IFN-γ producing T cells and standard deviation in each group after subtracting non-specific responses from corresponding samples stimulated with the irrelevant peptide.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3472981&req=5

pone-0047492-g006: Cell-mediated responses elicited by vaccination.HLA-A2kb transgenic mice (n = 3 per group) were inoculated (100 µl) subcutaneously at the base of the tail on days 0 and 14 with 30 µg of VLPs alone, emulsified with an equal amount of complete freund’s adjuvant (CFA) or pre-mixed with 30 µg of E8Pam2Cys in saline. Splenocytes were obtained 28 days later and restimulated for 7 days in the presence of 10 µg VLPs or an irrelevant HCV-derived HLA-A2-restricted epitope not part of the VLP construct. The frequency of peptide-specific T cells producing IFN-γ was determined in an ELISPOT assay. Each bar represents the average number of IFN-γ producing T cells and standard deviation in each group after subtracting non-specific responses from corresponding samples stimulated with the irrelevant peptide.
Mentions: The ability of VLP formulations containing E8Pam2Cys to induce a cell-meditated immune response was examined by inoculating transgenic mice expressing the MHC class I (HLA-A2) allele but not endogenous H-2Db molecules [28]. Control transgenic animals were inoculated with VLPs alone or VLPs emulsified with an equal amount of complete Freund’s adjuvant (CFA). Splenocytes from vaccinated animals were obtained 28 days post-inoculation and re-stimulated with antigen in vitro. The results (Figure 6) of an ELISPOT assay carried out revealed significantly higher numbers of HCV VLP-specific IFN-γ producing cells in the spleens of mice inoculated with VLPs in the presence of E8Pam2Cys or VLPs emulsified in CFA than in those that received VLPs alone.

Bottom Line: Although many studies provide strong evidence supporting the development of HCV virus-like particle (VLP)-based vaccines, the fact that heterologous viral vectors and/or multiple dosing regimes are required to induce protective immunity indicates that it is necessary to improve their immunogenicity.While co-formulation of this lipopeptide with VLPs only resulted in marginal improvements in dendritic cell (DC) uptake, its ability to concomitantly induce DC maturation at very small doses is a feature not observed using VLPs alone or in the presence of an aluminium hydroxide-based adjuvant (Alum).These results suggest overall that the immunogenicity of HCV VLPs can be significantly improved by the addition of this novel adjuvant by targeting their delivery to DCs and could therefore constitute a viable vaccine strategy for the treatment of HCV.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, The University of Melbourne, Parkville, Victoria, Australia.

ABSTRACT
Although many studies provide strong evidence supporting the development of HCV virus-like particle (VLP)-based vaccines, the fact that heterologous viral vectors and/or multiple dosing regimes are required to induce protective immunity indicates that it is necessary to improve their immunogenicity. In this study, we have evaluated the use of an anionic self-adjuvanting lipopeptide containing the TLR2 agonist Pam(2)Cys (E(8)Pam(2)Cys) to enhance the immunogenicity of VLPs containing the HCV structural proteins (core, E1 and E2) of genotype 1a. While co-formulation of this lipopeptide with VLPs only resulted in marginal improvements in dendritic cell (DC) uptake, its ability to concomitantly induce DC maturation at very small doses is a feature not observed using VLPs alone or in the presence of an aluminium hydroxide-based adjuvant (Alum). Dramatically improved VLP and E2-specific antibody responses were observed in VLP+E(8)Pam(2)Cys vaccinated mice where up to 3 doses of non-adjuvanted or traditionally alum-adjuvanted VLPs was required to match the antibody titres obtained with a single dose of VLPs formulated with this lipopeptide. This result also correlated with significantly higher numbers of specific antibody secreting cells that was detected in the spleens of VLP+E(8)Pam(2)Cys vaccinated mice and greater ability of sera from these mice to neutralise the binding and uptake of VLPs by Huh7 cells. Moreover, vaccination of HLA-A2 transgenic mice with this formulation also induced better VLP-specific IFN-γ-mediated responses compared to non-adjuvanted VLPs but comparable levels to that achieved when coadministered with complete freund's adjuvant. These results suggest overall that the immunogenicity of HCV VLPs can be significantly improved by the addition of this novel adjuvant by targeting their delivery to DCs and could therefore constitute a viable vaccine strategy for the treatment of HCV.

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