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Increasing the Vaccine Potential of Live M. bovis BCG by Coadministration with Plasmid DNA Encoding a Tuberculosis Prototype Antigen.

Bruffaerts N, Romano M, Denis O, Jurion F, Huygen K - Vaccines (Basel) (2014)

Bottom Line: Strongly increased T- and B-cell responses were observed with this protocol in C57BL/6 mice when compared to the administration of only BCG or in combination with an empty pDNA vector, as measured by Th1-type spleen cell cytokine secretion, specific IgG antibodies, as well as specific IFN-γ producing/cytolytic-CD8⁺ T-cells.Moreover, we observed a bystander activation induced by the coding plasmid, resulting in increased immune responses against other non-plasmid encoded, but BCG-expressed, antigens.In all, these results provide a proof of concept for a new TB vaccine, based on a BCG-plasmid DNA combination.

View Article: PubMed Central - PubMed

Affiliation: Scientific Institute of Public Health, Communicable and Infectious Diseases, Immunology, Brussels 1180, Belgium. nicolas.bruffaerts@wiv-isp.be.

ABSTRACT
The attenuated live M. bovis Bacille-Calmette-Guérin (BCG) is still the sole vaccine used against tuberculosis, but confers only variable efficacy against adult pulmonary tuberculosis (TB). Though no clear explanation for this limited efficacy has been given, different hypotheses have been advanced, such as the waning of memory T-cell responses, a reduced antigenic repertoire and the inability to induce effective CD8⁺ T-cell responses, which are known to be essential for latent tuberculosis control. In this study, a new BCG-based vaccination protocol was studied, in which BCG was formulated in combination with a plasmid DNA vaccine. As BCG is routinely administered to neonates, we have evaluated a more realistic approach of a simultaneous intradermal coadministration of BCG with pDNA encoding the prototype antigen, PPE44. Strongly increased T- and B-cell responses were observed with this protocol in C57BL/6 mice when compared to the administration of only BCG or in combination with an empty pDNA vector, as measured by Th1-type spleen cell cytokine secretion, specific IgG antibodies, as well as specific IFN-γ producing/cytolytic-CD8⁺ T-cells. Moreover, we observed a bystander activation induced by the coding plasmid, resulting in increased immune responses against other non-plasmid encoded, but BCG-expressed, antigens. In all, these results provide a proof of concept for a new TB vaccine, based on a BCG-plasmid DNA combination.

No MeSH data available.


Related in: MedlinePlus

Specific CD8+ T-cell responses induced by BCG/pDNA co-vaccination. (A) IFN-γ levels (pg/mL) were measured by ELISA in 72 h culture supernatant of splenocytes cultured with the PPE44 MHC Class I-restricted short peptide spanning amino acids 257–265 (PPE44[257–265]). RPMI, non-stimulated cells. Data are presented as the mean ± standard error of the mean and representative of two independent experiments, including six mice per group. * p < 0.05. (B) In vivo-specific cytotoxic T lymphocyte (CTL) activity. Unpulsed splenocytes (carboxyfluorescein diacetate (CFSE)low) and peptide-pulsed splenocytes (CFSEhigh) from naive mice were intravenously transferred to the immunized mice 18 h before flow cytometry analysis. Splenocytes were pulsed with the PPE44 MHC Class I-restrictedshort peptide spanning amino acids 257–265. The percentages of specific lysis in the immunized mice were compared to the ratio in transferred naive control mice. Data represent the mean ± standard error of the mean (n = 3–6/group). **p < 0.005. ID, intradermally. IM, intramuscularly.
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vaccines-02-00181-f003: Specific CD8+ T-cell responses induced by BCG/pDNA co-vaccination. (A) IFN-γ levels (pg/mL) were measured by ELISA in 72 h culture supernatant of splenocytes cultured with the PPE44 MHC Class I-restricted short peptide spanning amino acids 257–265 (PPE44[257–265]). RPMI, non-stimulated cells. Data are presented as the mean ± standard error of the mean and representative of two independent experiments, including six mice per group. * p < 0.05. (B) In vivo-specific cytotoxic T lymphocyte (CTL) activity. Unpulsed splenocytes (carboxyfluorescein diacetate (CFSE)low) and peptide-pulsed splenocytes (CFSEhigh) from naive mice were intravenously transferred to the immunized mice 18 h before flow cytometry analysis. Splenocytes were pulsed with the PPE44 MHC Class I-restrictedshort peptide spanning amino acids 257–265. The percentages of specific lysis in the immunized mice were compared to the ratio in transferred naive control mice. Data represent the mean ± standard error of the mean (n = 3–6/group). **p < 0.005. ID, intradermally. IM, intramuscularly.

Mentions: Specific IFN-γ was detected in spleen cell culture supernatants from mice vaccinated intramuscularly with pDNA-PPE44 and mice co-vaccinated with BCG/pDNA-PPE44 after stimulation with the synthetic PPE44257–265 peptide (Figure 3A). We have previously reported that PPE44257–265 spans a predicted Db-restricted epitope and on the in vivo cytotoxic T lymphocyte (CTL) activity against PPE44257–265 in C57BL/6 mice vaccinated with pDNA encoding PPE44 [27]. Hence, we verified whether BCG/pDNA-PPE44 co-vaccination could also induce effective CD8+ T-cells by assessing their in vivo cytolytic activity. PPE44257–265 peptide-pulsed, CFSE-labelled spleen cells from naive C57BL/6 mice were adoptively transferred as targets in vaccinated mice. As expected, a specific lysis of about 23% was measured by flow cytometry in mice vaccinated three times intramuscularly with pDNA-PPE44. In addition, a similar level of specific lysis was observed in mice intradermally coimmunized with BCG/pDNA-PPE44 (Figure 3B). In contrast, no PPE44-specific CTL activity could be detected in mice vaccinated with BCG alone or in mice vaccinated with BCG/empty vector, confirming the inability of the attenuated M. bovis BCG vaccine to induce detectable CD8+ T-cell responses.


Increasing the Vaccine Potential of Live M. bovis BCG by Coadministration with Plasmid DNA Encoding a Tuberculosis Prototype Antigen.

Bruffaerts N, Romano M, Denis O, Jurion F, Huygen K - Vaccines (Basel) (2014)

Specific CD8+ T-cell responses induced by BCG/pDNA co-vaccination. (A) IFN-γ levels (pg/mL) were measured by ELISA in 72 h culture supernatant of splenocytes cultured with the PPE44 MHC Class I-restricted short peptide spanning amino acids 257–265 (PPE44[257–265]). RPMI, non-stimulated cells. Data are presented as the mean ± standard error of the mean and representative of two independent experiments, including six mice per group. * p < 0.05. (B) In vivo-specific cytotoxic T lymphocyte (CTL) activity. Unpulsed splenocytes (carboxyfluorescein diacetate (CFSE)low) and peptide-pulsed splenocytes (CFSEhigh) from naive mice were intravenously transferred to the immunized mice 18 h before flow cytometry analysis. Splenocytes were pulsed with the PPE44 MHC Class I-restrictedshort peptide spanning amino acids 257–265. The percentages of specific lysis in the immunized mice were compared to the ratio in transferred naive control mice. Data represent the mean ± standard error of the mean (n = 3–6/group). **p < 0.005. ID, intradermally. IM, intramuscularly.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4494193&req=5

vaccines-02-00181-f003: Specific CD8+ T-cell responses induced by BCG/pDNA co-vaccination. (A) IFN-γ levels (pg/mL) were measured by ELISA in 72 h culture supernatant of splenocytes cultured with the PPE44 MHC Class I-restricted short peptide spanning amino acids 257–265 (PPE44[257–265]). RPMI, non-stimulated cells. Data are presented as the mean ± standard error of the mean and representative of two independent experiments, including six mice per group. * p < 0.05. (B) In vivo-specific cytotoxic T lymphocyte (CTL) activity. Unpulsed splenocytes (carboxyfluorescein diacetate (CFSE)low) and peptide-pulsed splenocytes (CFSEhigh) from naive mice were intravenously transferred to the immunized mice 18 h before flow cytometry analysis. Splenocytes were pulsed with the PPE44 MHC Class I-restrictedshort peptide spanning amino acids 257–265. The percentages of specific lysis in the immunized mice were compared to the ratio in transferred naive control mice. Data represent the mean ± standard error of the mean (n = 3–6/group). **p < 0.005. ID, intradermally. IM, intramuscularly.
Mentions: Specific IFN-γ was detected in spleen cell culture supernatants from mice vaccinated intramuscularly with pDNA-PPE44 and mice co-vaccinated with BCG/pDNA-PPE44 after stimulation with the synthetic PPE44257–265 peptide (Figure 3A). We have previously reported that PPE44257–265 spans a predicted Db-restricted epitope and on the in vivo cytotoxic T lymphocyte (CTL) activity against PPE44257–265 in C57BL/6 mice vaccinated with pDNA encoding PPE44 [27]. Hence, we verified whether BCG/pDNA-PPE44 co-vaccination could also induce effective CD8+ T-cells by assessing their in vivo cytolytic activity. PPE44257–265 peptide-pulsed, CFSE-labelled spleen cells from naive C57BL/6 mice were adoptively transferred as targets in vaccinated mice. As expected, a specific lysis of about 23% was measured by flow cytometry in mice vaccinated three times intramuscularly with pDNA-PPE44. In addition, a similar level of specific lysis was observed in mice intradermally coimmunized with BCG/pDNA-PPE44 (Figure 3B). In contrast, no PPE44-specific CTL activity could be detected in mice vaccinated with BCG alone or in mice vaccinated with BCG/empty vector, confirming the inability of the attenuated M. bovis BCG vaccine to induce detectable CD8+ T-cell responses.

Bottom Line: Strongly increased T- and B-cell responses were observed with this protocol in C57BL/6 mice when compared to the administration of only BCG or in combination with an empty pDNA vector, as measured by Th1-type spleen cell cytokine secretion, specific IgG antibodies, as well as specific IFN-γ producing/cytolytic-CD8⁺ T-cells.Moreover, we observed a bystander activation induced by the coding plasmid, resulting in increased immune responses against other non-plasmid encoded, but BCG-expressed, antigens.In all, these results provide a proof of concept for a new TB vaccine, based on a BCG-plasmid DNA combination.

View Article: PubMed Central - PubMed

Affiliation: Scientific Institute of Public Health, Communicable and Infectious Diseases, Immunology, Brussels 1180, Belgium. nicolas.bruffaerts@wiv-isp.be.

ABSTRACT
The attenuated live M. bovis Bacille-Calmette-Guérin (BCG) is still the sole vaccine used against tuberculosis, but confers only variable efficacy against adult pulmonary tuberculosis (TB). Though no clear explanation for this limited efficacy has been given, different hypotheses have been advanced, such as the waning of memory T-cell responses, a reduced antigenic repertoire and the inability to induce effective CD8⁺ T-cell responses, which are known to be essential for latent tuberculosis control. In this study, a new BCG-based vaccination protocol was studied, in which BCG was formulated in combination with a plasmid DNA vaccine. As BCG is routinely administered to neonates, we have evaluated a more realistic approach of a simultaneous intradermal coadministration of BCG with pDNA encoding the prototype antigen, PPE44. Strongly increased T- and B-cell responses were observed with this protocol in C57BL/6 mice when compared to the administration of only BCG or in combination with an empty pDNA vector, as measured by Th1-type spleen cell cytokine secretion, specific IgG antibodies, as well as specific IFN-γ producing/cytolytic-CD8⁺ T-cells. Moreover, we observed a bystander activation induced by the coding plasmid, resulting in increased immune responses against other non-plasmid encoded, but BCG-expressed, antigens. In all, these results provide a proof of concept for a new TB vaccine, based on a BCG-plasmid DNA combination.

No MeSH data available.


Related in: MedlinePlus