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Analysis of a Multi-component Multi-stage Malaria Vaccine Candidate--Tackling the Cocktail Challenge.

Boes A, Spiegel H, Voepel N, Edgue G, Beiss V, Kapelski S, Fendel R, Scheuermayer M, Pradel G, Bolscher JM, Behet MC, Dechering KJ, Hermsen CC, Sauerwein RW, Schillberg S, Reimann A, Fischer R - PLoS ONE (2015)

Bottom Line: To investigate the potential of such an approach we combined proteins and domains (11 in total) from the pre-erythrocytic, blood and sexual stages of P. falciparum into a cocktail of four different components recombinantly produced in plants.Using purified rabbit immune IgG we observed strong inhibition in functional in vitro assays addressing the pre-erythrocytic (up to 80%), blood (up to 90%) and sexual parasite stages (100%).While the results underline the feasibility of a multi-stage vaccine cocktail, the analysis of component-specific efficacy indicates significant differences in IC50 requirements for stage-specific antibody concentrations providing valuable insights into this complex scenario and will thereby improve future approaches towards malaria vaccine cocktail development regarding the selection of suitable antigens and the ratios of components, to fine tune overall and stage-specific efficacy.

View Article: PubMed Central - PubMed

Affiliation: Fraunhofer Institute for Molecular Biology and Applied Ecology (IME), Aachen, Germany.

ABSTRACT
Combining key antigens from the different stages of the P. falciparum life cycle in the context of a multi-stage-specific cocktail offers a promising approach towards the development of a malaria vaccine ideally capable of preventing initial infection, the clinical manifestation as well as the transmission of the disease. To investigate the potential of such an approach we combined proteins and domains (11 in total) from the pre-erythrocytic, blood and sexual stages of P. falciparum into a cocktail of four different components recombinantly produced in plants. After immunization of rabbits we determined the domain-specific antibody titers as well as component-specific antibody concentrations and correlated them with stage specific in vitro efficacy. Using purified rabbit immune IgG we observed strong inhibition in functional in vitro assays addressing the pre-erythrocytic (up to 80%), blood (up to 90%) and sexual parasite stages (100%). Based on the component-specific antibody concentrations we calculated the IC50 values for the pre-erythrocytic stage (17-25 μg/ml), the blood stage (40-60 μg/ml) and the sexual stage (1.75 μg/ml). While the results underline the feasibility of a multi-stage vaccine cocktail, the analysis of component-specific efficacy indicates significant differences in IC50 requirements for stage-specific antibody concentrations providing valuable insights into this complex scenario and will thereby improve future approaches towards malaria vaccine cocktail development regarding the selection of suitable antigens and the ratios of components, to fine tune overall and stage-specific efficacy.

No MeSH data available.


Related in: MedlinePlus

In vitro growth inhibition assay (GIA) of asexual P. falciparum 3D7 parasites with purified PlasmoMix-specific rabbit IgG.The rabbit IgGs were purified from the serum of three rabbits (R1, R2 and R3) collected on days 35, 63 and 91 post-immunization with PlasmoMix. Nomenclature of the sample first features the number of the rabbit (R1, R2 or R3) followed by the sampling day (35, 63 or 91). (A) Four serial 1/1 dilutions from 6–0.75 mg/ml of total IgGs were used to demonstrate the concentration dependency of the assay and to calculate the IC50 values (the total IgG concentrations needed for 50% inhibition). (B) The same GIA but instead of total IgG, the gAMA1-specifc antibody concentration (based on CFCA and calculated from total IgG, see methods section) was used and the IC50 values for gAMA1-specific antibodies were calculated. Each data point represents the mean of technical triplicates.
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pone.0131456.g004: In vitro growth inhibition assay (GIA) of asexual P. falciparum 3D7 parasites with purified PlasmoMix-specific rabbit IgG.The rabbit IgGs were purified from the serum of three rabbits (R1, R2 and R3) collected on days 35, 63 and 91 post-immunization with PlasmoMix. Nomenclature of the sample first features the number of the rabbit (R1, R2 or R3) followed by the sampling day (35, 63 or 91). (A) Four serial 1/1 dilutions from 6–0.75 mg/ml of total IgGs were used to demonstrate the concentration dependency of the assay and to calculate the IC50 values (the total IgG concentrations needed for 50% inhibition). (B) The same GIA but instead of total IgG, the gAMA1-specifc antibody concentration (based on CFCA and calculated from total IgG, see methods section) was used and the IC50 values for gAMA1-specific antibodies were calculated. Each data point represents the mean of technical triplicates.

Mentions: The efficacy of the blood-stage component of the vaccine cocktail (antigens gAMA1 and E3) was assessed using a classical GIA. Up to 90% inhibition was observed at a concentration of 6 mg/ml total IgG purified from rabbit immune sera (Fig 4A). Determination and comparison of IC50 values for antigen-specific IgGs indicated that the IC50 values for gAMA1-specific antibodies (Fig 4B) were in the same range (30–60 μg/ml) as reported in a previous study [30] after the immunization of rabbits with recombinant plant-derived gAMA1, indicating that the growth inhibition induced by the cocktail is mainly mediated by gAMA1-specific antibodies. To precisely quantify this contribution, a reversal of growth inhibition assay was performed by competitive neutralization of these antibodies with recombinant gAMA1 and E3 (Fig 5). Although growth inhibition was completely abolished by 120 μg/ml gAMA1, the neutralization of E3-specific IgGs using 24 μg/ml E3 did not affect growth inhibition.


Analysis of a Multi-component Multi-stage Malaria Vaccine Candidate--Tackling the Cocktail Challenge.

Boes A, Spiegel H, Voepel N, Edgue G, Beiss V, Kapelski S, Fendel R, Scheuermayer M, Pradel G, Bolscher JM, Behet MC, Dechering KJ, Hermsen CC, Sauerwein RW, Schillberg S, Reimann A, Fischer R - PLoS ONE (2015)

In vitro growth inhibition assay (GIA) of asexual P. falciparum 3D7 parasites with purified PlasmoMix-specific rabbit IgG.The rabbit IgGs were purified from the serum of three rabbits (R1, R2 and R3) collected on days 35, 63 and 91 post-immunization with PlasmoMix. Nomenclature of the sample first features the number of the rabbit (R1, R2 or R3) followed by the sampling day (35, 63 or 91). (A) Four serial 1/1 dilutions from 6–0.75 mg/ml of total IgGs were used to demonstrate the concentration dependency of the assay and to calculate the IC50 values (the total IgG concentrations needed for 50% inhibition). (B) The same GIA but instead of total IgG, the gAMA1-specifc antibody concentration (based on CFCA and calculated from total IgG, see methods section) was used and the IC50 values for gAMA1-specific antibodies were calculated. Each data point represents the mean of technical triplicates.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492585&req=5

pone.0131456.g004: In vitro growth inhibition assay (GIA) of asexual P. falciparum 3D7 parasites with purified PlasmoMix-specific rabbit IgG.The rabbit IgGs were purified from the serum of three rabbits (R1, R2 and R3) collected on days 35, 63 and 91 post-immunization with PlasmoMix. Nomenclature of the sample first features the number of the rabbit (R1, R2 or R3) followed by the sampling day (35, 63 or 91). (A) Four serial 1/1 dilutions from 6–0.75 mg/ml of total IgGs were used to demonstrate the concentration dependency of the assay and to calculate the IC50 values (the total IgG concentrations needed for 50% inhibition). (B) The same GIA but instead of total IgG, the gAMA1-specifc antibody concentration (based on CFCA and calculated from total IgG, see methods section) was used and the IC50 values for gAMA1-specific antibodies were calculated. Each data point represents the mean of technical triplicates.
Mentions: The efficacy of the blood-stage component of the vaccine cocktail (antigens gAMA1 and E3) was assessed using a classical GIA. Up to 90% inhibition was observed at a concentration of 6 mg/ml total IgG purified from rabbit immune sera (Fig 4A). Determination and comparison of IC50 values for antigen-specific IgGs indicated that the IC50 values for gAMA1-specific antibodies (Fig 4B) were in the same range (30–60 μg/ml) as reported in a previous study [30] after the immunization of rabbits with recombinant plant-derived gAMA1, indicating that the growth inhibition induced by the cocktail is mainly mediated by gAMA1-specific antibodies. To precisely quantify this contribution, a reversal of growth inhibition assay was performed by competitive neutralization of these antibodies with recombinant gAMA1 and E3 (Fig 5). Although growth inhibition was completely abolished by 120 μg/ml gAMA1, the neutralization of E3-specific IgGs using 24 μg/ml E3 did not affect growth inhibition.

Bottom Line: To investigate the potential of such an approach we combined proteins and domains (11 in total) from the pre-erythrocytic, blood and sexual stages of P. falciparum into a cocktail of four different components recombinantly produced in plants.Using purified rabbit immune IgG we observed strong inhibition in functional in vitro assays addressing the pre-erythrocytic (up to 80%), blood (up to 90%) and sexual parasite stages (100%).While the results underline the feasibility of a multi-stage vaccine cocktail, the analysis of component-specific efficacy indicates significant differences in IC50 requirements for stage-specific antibody concentrations providing valuable insights into this complex scenario and will thereby improve future approaches towards malaria vaccine cocktail development regarding the selection of suitable antigens and the ratios of components, to fine tune overall and stage-specific efficacy.

View Article: PubMed Central - PubMed

Affiliation: Fraunhofer Institute for Molecular Biology and Applied Ecology (IME), Aachen, Germany.

ABSTRACT
Combining key antigens from the different stages of the P. falciparum life cycle in the context of a multi-stage-specific cocktail offers a promising approach towards the development of a malaria vaccine ideally capable of preventing initial infection, the clinical manifestation as well as the transmission of the disease. To investigate the potential of such an approach we combined proteins and domains (11 in total) from the pre-erythrocytic, blood and sexual stages of P. falciparum into a cocktail of four different components recombinantly produced in plants. After immunization of rabbits we determined the domain-specific antibody titers as well as component-specific antibody concentrations and correlated them with stage specific in vitro efficacy. Using purified rabbit immune IgG we observed strong inhibition in functional in vitro assays addressing the pre-erythrocytic (up to 80%), blood (up to 90%) and sexual parasite stages (100%). Based on the component-specific antibody concentrations we calculated the IC50 values for the pre-erythrocytic stage (17-25 μg/ml), the blood stage (40-60 μg/ml) and the sexual stage (1.75 μg/ml). While the results underline the feasibility of a multi-stage vaccine cocktail, the analysis of component-specific efficacy indicates significant differences in IC50 requirements for stage-specific antibody concentrations providing valuable insights into this complex scenario and will thereby improve future approaches towards malaria vaccine cocktail development regarding the selection of suitable antigens and the ratios of components, to fine tune overall and stage-specific efficacy.

No MeSH data available.


Related in: MedlinePlus