scholarly journals High-Level Expression of Plasmodium vivax Apical Membrane Antigen 1 (AMA-1) in Pichia pastoris: Strong Immunogenicity in Macaca mulatta Immunized with P. vivax AMA-1 and Adjuvant SBAS2

1999 ◽  
Vol 67 (1) ◽  
pp. 43-49 ◽  
Author(s):  
Clemens H. M. Kocken ◽  
Martin A. Dubbeld ◽  
Annemarie Van Der Wel ◽  
Jack T. Pronk ◽  
Andrew P. Waters ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Plasmodium Vivax ◽  
Apical Membrane ◽  
Gel Filtration ◽  
Methylotrophic Yeast ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen ◽  
High Level

ABSTRACT The apical membrane antigen 1 (AMA-1) family is a promising family of malaria blood-stage vaccine candidates that have induced protection in rodent and nonhuman primate models of malaria. Correct conformation of the protein appears to be essential for the induction of parasite-inhibitory responses, and these responses appear to be primarily antibody mediated. Here we describe for the first time high-level secreted expression (over 50 mg/liter) of thePlasmodium vivax AMA-1 (PV66/AMA-1) ectodomain by using the methylotrophic yeast Pichia pastoris. To prevent nonnative glycosylation, a conservatively mutagenized PV66/AMA-1 gene (PV66Δglyc) lacking N-glycosylation sites was also developed. Expression of the PV66Δglyc ectodomain yielded similar levels of a homogeneous product that was nonglycosylated and was readily purified by ion-exchange and gel filtration chromatographies. Recombinant PV66Δglyc43–487 was reactive with conformation-dependent monoclonal antibodies. With the SBAS2 adjuvant,Pichia-expressed PV66Δglyc43–487 was highly immunogenic in five rhesus monkeys, inducing immunoglobulin G enzyme-linked immunosorbent assay titers in excess of 1:200,000. This group of monkeys had a weak trend showing lower cumulative parasite loads following a Plasmodium cynomolgi infection than in the control group.

scholarly journals Invasion-Inhibitory Antibodies Elicited by Immunization with Plasmodium vivax Apical Membrane Antigen-1 Expressed in Pichia pastoris Yeast

2013 ◽  
Vol 82 (3) ◽  
pp. 1296-1307 ◽  
Author(s):  
Elaine C. Vicentin ◽  
Kátia S. Françoso ◽  
Mariana V. Rocha ◽  
Dmitri Iourtov ◽  
Fernanda L. dos Santos ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Recombinant Protein ◽  
Plasmodium Vivax ◽  
Apical Membrane ◽  
Content Type ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen ◽  
Monophosphoryl Lipid A ◽  
Antibody Titers ◽  
Inhibitory Antibodies

ABSTRACTIn a recent vaccine trial performed with African children, immunization with a recombinant protein based onPlasmodium falciparumapical membrane antigen 1 (AMA-1) conferred a significant degree of strain-specific resistance against malaria. To contribute to the efforts of generating a vaccine againstPlasmodium vivaxmalaria, we expressed the ectodomain ofP. vivaxAMA-1 (PvAMA-1) as a secreted soluble protein in the methylotrophic yeastPichia pastoris. Recognized by a high percentage of sera from individuals infected byP. vivax, this recombinant protein was found to have maintained its antigenicity. The immunogenicity of this protein was evaluated in mice using immunization protocols that included homologous and heterologous prime-boost strategies with plasmid DNA and recombinant protein. We used the following formulations containing different adjuvants: aluminum salts (Alum),Bordetella pertussismonophosphoryl lipid A (MPLA), flagellin FliC fromSalmonella entericaserovar Typhimurium, saponin Quil A, or incomplete Freund's adjuvant (IFA). The formulations containing the adjuvants Quil A or IFA elicited the highest IgG antibody titers. Significant antibody titers were also obtained using a formulation developed for human use containing MPLA or Alum plus MPLA. Recombinant PvAMA-1 produced under “conditions of good laboratory practice” provided a good yield, high purity, low endotoxin levels, and no microbial contaminants and reproduced the experimental immunizations. Most relevant for vaccine development was the fact that immunization with PvAMA-1 elicited invasion-inhibitory antibodies against different Asian isolates ofP. vivax. Our results show that AMA-1 expressed inP. pastorisis a promising antigen for use in future preclinical and clinical studies.

Analysis of the Plasmodium vivax apical membrane antigen-1 gene from re-emerging Korean isolates

2003 ◽  
Vol 90 (4) ◽  
pp. 325-329 ◽  
Author(s):  
Joon-Yong Chung ◽  
Eui-Hyun Chun ◽  
Jin-Ho Chun ◽  
Weon-Gyu Kho
Keyword(s):  
Plasmodium Vivax ◽  
Apical Membrane ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen

scholarly journals Natural Human Antibody Responses to Plasmodium vivax Apical Membrane Antigen 1 under Low Transmission and Unstable Malaria Conditions in Sri Lanka

2006 ◽  
Vol 74 (1) ◽  
pp. 798-801 ◽  
Author(s):  
Thilan Wickramarachchi ◽  
Prasad H. Premaratne ◽  
K. L. R. Lakshman Perera ◽  
Sumith Bandara ◽  
Clemens H. M. Kocken ◽  
...  
Keyword(s):  
Sri Lanka ◽  
Plasmodium Vivax ◽  
Malaria Vaccine ◽  
Apical Membrane ◽  
Human Antibody ◽  
Antibody Prevalence ◽  
Low Transmission ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen ◽  
Malaria Vaccine Candidate

ABSTRACT Plasmodium vivax apical membrane antigen 1, an important malaria vaccine candidate, was immunogenic during natural malaria infections in Sri Lanka, where low transmission and unstable malaria conditions prevail. Antibody prevalence increased with exposure in areas where malaria was or was not endemic. A marked isotype switch to cytophilic (immunoglobulin G1 [IgG1]/IgG3) antibodies was evident with increasing exposure exclusively in residents from areas of endemicity.

scholarly journals Identification of a Highly Antigenic Linear B Cell Epitope within Plasmodium vivax Apical Membrane Antigen 1 (AMA-1)

PLoS ONE ◽  
2011 ◽  
Vol 6 (6) ◽  
pp. e21289 ◽  
Author(s):  
Lilian Lacerda Bueno ◽  
Francisco Pereira Lobo ◽  
Cristiane Guimarães Morais ◽  
Luíza Carvalho Mourão ◽  
Ricardo Andrez Machado de Ávila ◽  
...  
Keyword(s):  
B Cell ◽  
Plasmodium Vivax ◽  
Apical Membrane ◽  
Cell Epitope ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen ◽  
B Cell Epitope ◽  
Linear B

Evaluation of the genetic diversity of domain II of Plasmodium vivax Apical Membrane Antigen 1 (PvAMA-1) and the ensuing strain-specific immune responses in patients from Sri Lanka

Vaccine ◽  
2011 ◽  
Vol 29 (43) ◽  
pp. 7491-7504 ◽  
Author(s):  
Sajani Dias ◽  
Maheshika Somarathna ◽  
Aresha Manamperi ◽  
Ananias A. Escalante ◽  
Anusha M. Gunasekera ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Sri Lanka ◽  
Immune Responses ◽  
Plasmodium Vivax ◽  
Apical Membrane ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen

scholarly journals Genetic Diversity and Selection at the Plasmodium vivax Apical Membrane Antigen-1 (PvAMA-1) Locus in a Sri Lankan Population

2007 ◽  
Vol 24 (4) ◽  
pp. 939-947 ◽  
Author(s):  
A. M. Gunasekera ◽  
T. Wickramarachchi ◽  
D. E. Neafsey ◽  
I. Ganguli ◽  
L. Perera ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Plasmodium Vivax ◽  
Apical Membrane ◽  
Sri Lankan ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen

scholarly journals A Diversity-Covering Approach to Immunization with Plasmodium falciparum Apical Membrane Antigen 1 Induces Broader Allelic Recognition and Growth Inhibition Responses in Rabbits

2008 ◽  
Vol 76 (6) ◽  
pp. 2660-2670 ◽  
Author(s):  
Edmond J. Remarque ◽  
Bart W. Faber ◽  
Clemens H. M. Kocken ◽  
Alan W. Thomas
Keyword(s):  
Plasmodium Falciparum ◽  
Amino Acid ◽  
Growth Inhibition ◽  
Apical Membrane ◽  
Recombinant Expression ◽  
Methylotrophic Yeast ◽  
Apical Membrane Antigen 1 ◽  
Apical Membrane Antigen ◽  
Amino Acid Variability

ABSTRACT Plasmodium falciparum apical membrane antigen 1 (PfAMA1), a candidate malaria vaccine, is polymorphic. This polymorphism is believed to be generated predominantly under immune selection pressure and, as a result, may compromise attempts at vaccination. Alignment of 355 PfAMA1 sequences shows that around 10% of the 622 amino acid residues can vary between alleles and that linkages between polymorphic residues occur. Using this analysis, we have designed three diversity-covering (DiCo) PfAMA1 sequences that take account of these linkages and, when taken together, on average incorporate 97% of amino acid variability observed. For each of the three DiCo sequences, a synthetic gene was constructed and used to transform the methylotrophic yeast Pichia pastoris, allowing recombinant expression. All three DiCo proteins were reactive with the reduction-sensitive monoclonal antibody 4G2, suggesting the DiCo sequences had conformations similar to those of naturally occurring PfAMA1. Rabbits were immunized with FVO strain PfAMA1 or with the DiCo proteins either individually or as a mixture. Antibody titers and the ability to inhibit parasite growth in vitro were determined. Animals immunized with the DiCo mix performed similarly to animals immunized with FVO AMA1 when measured against FCR3 strain parasites but outperformed animals immunized with FVO AMA1 when assessed against other strains. The levels of growth inhibition (∼70%) induced by the mix of three DiCo proteins were comparable for FVO, 3D7, and HB3, suggesting that a considerable degree of diversity in AMA1 is adequately covered. This suggests that vaccines based upon the DiCo mix approach provide a broader functional immunity than immunization with a single allele.

scholarly journals Prediction of Merozoite Surface Protein 1 and Apical Membrane Antigen 1 Vaccine Efficacies against Plasmodium chabaudi Malaria Based on Prechallenge Antibody Responses

2008 ◽  
Vol 16 (3) ◽  
pp. 293-302 ◽  
Author(s):  
Michelle M. Lynch ◽  
Amy Cernetich-Ott ◽  
William P. Weidanz ◽  
James M. Burns
Keyword(s):  
Apical Membrane ◽  
Protective Efficacy ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Antibody Responses ◽  
Enzyme Linked Immunosorbent Assay ◽  
Plasmodium Chabaudi ◽  
Apical Membrane Antigen 1 ◽  
Merozoite Surface Protein 1 ◽  
Apical Membrane Antigen

ABSTRACT For the development of blood-stage malaria vaccines, there is a clear need to establish in vitro measures of the antibody-mediated and the cell-mediated immune responses that correlate with protection. In this study, we focused on establishing correlates of antibody-mediated immunity induced by immunization with apical membrane antigen 1 (AMA1) and merozoite surface protein 142 (MSP142) subunit vaccines. To do so, we exploited the Plasmodium chabaudi rodent model, with which we can immunize animals with both protective and nonprotective vaccine formulations and allow the parasitemia in the challenged animals to peak. Vaccine formulations were varied with regard to the antigen dose, the antigen conformation, and the adjuvant used. Prechallenge antibody responses were evaluated by enzyme-linked immunosorbent assay and were tested for a correlation with protection against nonlethal P. chabaudi malaria, as measured by a reduction in the peak level of parasitemia. The analysis showed that neither the isotype profile nor the avidity of vaccine-induced antibodies correlated with protective efficacy. However, high titers of antibodies directed against conformation-independent epitopes were associated with poor vaccine performance and may limit the effectiveness of protective antibodies that recognize conformation-dependent epitopes. We were able to predict the efficacies of the P. chabaudi AMA1 (PcAMA1) and P. chabaudi MSP142 (PcMSP142) vaccines only when the prechallenge antibody titers to both refolded and reduced/alkylated antigens were considered in combination. The relative importance of these two measures of vaccine-induced responses as predictors of protection differed somewhat for the PcAMA1 and the PcMSP142 vaccines, a finding confirmed in our final immunization and challenge study. A similar approach to the evaluation of vaccine-induced antibody responses may be useful during clinical trials of Plasmodium falciparum AMA1 and MSP142 vaccines.

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