Transgenic Parasites Stably Expressing Full-Length Plasmodium falciparum Circumsporozoite Protein as a Model for Vaccine Down-Selection in Mice Using Sterile Protection as an Endpoint

ABSTRACTCircumsporozoite protein (CSP) ofPlasmodium falciparumis a protective human malaria vaccine candidate. There is an urgent need for models that can rapidly down-select novel CSP-based vaccine candidates. In the present study, the mouse-mosquito transmission cycle of a transgenicPlasmodium bergheimalaria parasite stably expressing a functional full-lengthP. falciparumCSP was optimized to consistently produce infective sporozoites for protection studies. A minimal sporozoite challenge dose was established, and protection was defined as the absence of blood-stage parasites 14 days after intravenous challenge. The specificity of protection was confirmed by vaccinating mice with multiple CSP constructs of differing lengths and compositions. Constructs that induced high NANP repeat-specific antibody titers in enzyme-linked immunosorbent assays were protective, and the degree of protection was dependent on the antigen dose. There was a positive correlation between antibody avidity and protection. The antibodies in the protected mice recognized the native CSP on the parasites and showed sporozoite invasion inhibitory activity. Passive transfer of anti-CSP antibodies into naive mice also induced protection. Thus, we have demonstrated the utility of a mouse efficacy model to down-select human CSP-based vaccine formulations.

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A Full-Length Plasmodium falciparum Recombinant Circumsporozoite Protein Expressed by Pseudomonas fluorescens Platform as a Malaria Vaccine Candidate

PLoS ONE ◽

10.1371/journal.pone.0107764 ◽

2014 ◽

Vol 9 (9) ◽

pp. e107764 ◽

Cited By ~ 24

Author(s):

Amy R. Noe ◽

Diego Espinosa ◽

Xiangming Li ◽

Jordana G. A. Coelho-dos-Reis ◽

Ryota Funakoshi ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Pseudomonas Fluorescens ◽

Malaria Vaccine ◽

Vaccine Candidate ◽

Circumsporozoite Protein ◽

Full Length ◽

Malaria Vaccine Candidate

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A Plasmodium falciparum malaria vaccine candidate which contains epitopes from the circumsporozoite protein and a blood stage antigen, 5.1

Molecular and Biochemical Parasitology ◽

10.1016/0166-6851(91)90173-4 ◽

1991 ◽

Vol 47 (2) ◽

pp. 143-150 ◽

Cited By ~ 14

Author(s):

Patrick Caspers ◽

Howard Etlinger ◽

Hugues Matile ◽

J.Richard Pink ◽

Dietrich Stüber ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Falciparum Malaria ◽

Malaria Vaccine ◽

Vaccine Candidate ◽

Plasmodium Falciparum Malaria ◽

Circumsporozoite Protein ◽

Blood Stage ◽

Malaria Vaccine Candidate

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Bacterially Expressed Full-Length Recombinant Plasmodium falciparum RH5 Protein Binds Erythrocytes and Elicits Potent Strain-Transcending Parasite-Neutralizing Antibodies

Infection and Immunity ◽

10.1128/iai.00970-13 ◽

2013 ◽

Vol 82 (1) ◽

pp. 152-164 ◽

Cited By ~ 54

Author(s):

K. Sony Reddy ◽

Alok K. Pandey ◽

Hina Singh ◽

Tajali Sahar ◽

Amlabu Emmanuel ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Neutralizing Antibodies ◽

Malaria Vaccine ◽

Expression System ◽

Full Length ◽

Blood Stage ◽

Parasite Protein ◽

Content Type ◽

Erythrocyte Binding ◽

Blood Stage Malaria

ABSTRACTPlasmodium falciparumreticulocyte binding-like homologous protein 5 (PfRH5) is an essential merozoite ligand that binds with its erythrocyte receptor, basigin. PfRH5 is an attractive malaria vaccine candidate, as it is expressed by a wide number ofP. falciparumstrains, cannot be genetically disrupted, and exhibits limited sequence polymorphisms. Viral vector-induced PfRH5 antibodies potently inhibited erythrocyte invasion. However, it has been a challenge to generate full-length recombinant PfRH5 in a bacterial-cell-based expression system. In this study, we have produced full-length recombinant PfRH5 inEscherichia colithat exhibits specific erythrocyte binding similar to that of the native PfRH5 parasite protein and also, importantly, elicits potent invasion-inhibitory antibodies against a number ofP. falciparumstrains. Antibasigin antibodies blocked the erythrocyte binding of both native and recombinant PfRH5, further confirming that they bind with basigin. We have thus successfully produced full-length PfRH5 as a functionally active erythrocyte binding recombinant protein with a conformational integrity that mimics that of the native parasite protein and elicits potent strain-transcending parasite-neutralizing antibodies.P. falciparumhas the capability to develop immune escape mechanisms, and thus, blood-stage malaria vaccines that target multiple antigens or pathways may prove to be highly efficacious. In this regard, antibody combinations targeting PfRH5 and other key merozoite antigens produced potent additive inhibition against multiple worldwideP. falciparumstrains. PfRH5 was immunogenic when immunized with other antigens, eliciting potent invasion-inhibitory antibody responses with no immune interference. Our results strongly support the development of PfRH5 as a component of a combination blood-stage malaria vaccine.

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The Plasmodium falciparum circumsporozoite protein produced in Lactococcus lactis is pure and stable

Journal of Biological Chemistry ◽

10.1074/jbc.ra119.011268 ◽

2019 ◽

Vol 295 (2) ◽

pp. 403-414 ◽

Cited By ~ 1

Author(s):

Susheel K. Singh ◽

Jordan Plieskatt ◽

Bishwanath Kumar Chourasia ◽

Vandana Singh ◽

Judith M. Bolscher ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Lactococcus Lactis ◽

Malaria Vaccine ◽

Vaccine Development ◽

Expression System ◽

Surface Protein ◽

Circumsporozoite Protein ◽

Full Length

The Plasmodium falciparum circumsporozoite protein (PfCSP) is a sporozoite surface protein whose role in sporozoite motility and cell invasion has made it the leading candidate for a pre-erythrocytic malaria vaccine. However, production of high yields of soluble recombinant PfCSP, including its extensive NANP and NVDP repeats, has proven problematic. Here, we report on the development and characterization of a secreted, soluble, and stable full-length PfCSP (containing 4 NVDP and 38 NANP repeats) produced in the Lactococcus lactis expression system. The recombinant full-length PfCSP, denoted PfCSP4/38, was produced initially with a histidine tag and purified by a simple two-step procedure. Importantly, the recombinant PfCSP4/38 retained a conformational epitope for antibodies as confirmed by both in vivo and in vitro characterizations. We characterized this complex protein by HPLC, light scattering, MS analysis, differential scanning fluorimetry, CD, SDS-PAGE, and immunoblotting with conformation-dependent and -independent mAbs, which confirmed it to be both pure and soluble. Moreover, we found that the recombinant protein is stable at both frozen and elevated-temperature storage conditions. When we used L. lactis–derived PfCSP4/38 to immunize mice, it elicited high levels of functional antibodies that had the capacity to modify sporozoite motility in vitro. We concluded that the reported yield, purity, results of biophysical analyses, and stability of PfCSP4/38 warrant further consideration of using the L. lactis system for the production of circumsporozoite proteins for preclinical and clinical applications in malaria vaccine development.

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Tailoring a Combination Preerythrocytic Malaria Vaccine

Infection and Immunity ◽

10.1128/iai.01063-15 ◽

2015 ◽

Vol 84 (3) ◽

pp. 622-634 ◽

Cited By ~ 13

Author(s):

Karolis Bauza ◽

Erwan Atcheson ◽

Tomas Malinauskas ◽

Andrew M. Blagborough ◽

Arturo Reyes-Sandoval

Keyword(s):

Viral Vectors ◽

Malaria Vaccine ◽

Subunit Vaccines ◽

Content Type ◽

Adhesive Protein ◽

Order Of Magnitude ◽

Antibody Titers ◽

Malaria Vaccine Candidate ◽

Low Levels ◽

Combined Trap

The leading malaria vaccine candidate, RTS,S, based on thePlasmodium falciparumcircumsporozoite protein (CSP), will likely be the first publicly adopted malaria vaccine. However, this and other subunit vaccines, such as virus-vectored thrombospondin-related adhesive protein (TRAP), provide only intermediate to low levels of protection. In this study, thePlasmodium bergheihomologues of antigens CSP and TRAP are combined. TRAP is delivered using adenovirus- and vaccinia virus-based vectors in a prime-boost regime. Initially, CSP is also delivered using these viral vectors; however, a reduction of anti-CSP antibodies is seen when combined with virus-vectored TRAP, and the combination is no more protective than either subunit vaccine alone. Using an adenovirus-CSP prime, protein-CSP boost regime, however, increases anti-CSP antibody titers by an order of magnitude, which is maintained when combined with virus-vectored TRAP. This combination regime using protein CSP provided 100% protection in C57BL/6 mice compared to no protection using virus-vectored TRAP alone and 40% protection using adenovirus-CSP prime and protein-CSP boost alone. This suggests that a combination of CSP and TRAP subunit vaccines could enhance protection against malaria.

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Antibody Targets and Properties for Complement-Fixation Against the Circumsporozoite Protein in Malaria Immunity

Frontiers in Immunology ◽

10.3389/fimmu.2021.775659 ◽

2021 ◽

Vol 12 ◽

Author(s):

Liriye Kurtovic ◽

Damien R. Drew ◽

Arlene E. Dent ◽

James W. Kazura ◽

James G. Beeson

Keyword(s):

Plasmodium Falciparum ◽

Malaria Vaccine ◽

Vaccine Development ◽

Complement Fixation ◽

Terminal Region ◽

Antibody Responses ◽

Circumsporozoite Protein ◽

Full Length ◽

Acquired Immunity ◽

Vaccine Candidates

The Plasmodium falciparum circumsporozoite protein (CSP) forms the basis of leading subunit malaria vaccine candidates. However, the mechanisms and specific targets of immunity are poorly defined. Recent findings suggest that antibody-mediated complement-fixation and activation play an important role in immunity. Here, we investigated the regions of CSP targeted by functional complement-fixing antibodies and the antibody properties associated with this activity. We quantified IgG, IgM, and functional complement-fixing antibody responses to different regions of CSP among Kenyan adults naturally exposed to malaria (n=102) and using a series of rabbit vaccination studies. Individuals who acquired functional complement-fixing antibodies had higher IgG, IgM and IgG1 and IgG3 to CSP. Acquired complement-fixing antibodies targeted the N-terminal, central-repeat, and C-terminal regions of CSP, and positive responders had greater antibody breadth compared to those who were negative for complement-fixing antibodies (p<0.05). Using rabbit vaccinations as a model, we confirmed that IgG specific to the central-repeat and non-repeat regions of CSP could effectively fix complement. However, vaccination with near full length CSP in rabbits poorly induced antibodies to the N-terminal region compared to naturally-acquired immunity in humans. Poor induction of N-terminal antibodies was also observed in a vaccination study performed in mice. IgG and IgM to all three regions of CSP play a role in mediating complement-fixation, which has important implications for malaria vaccine development.

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Assessment of Genetic Diversity of Plasmodium Falciparum Circumsporozoite Protein in Sudan; The RTS,S Leading Malaria Vaccine Candidate

10.21203/rs.3.rs-619335/v1 ◽

2021 ◽

Author(s):

Nouh Saad Mohamed ◽

Hanadi AbdElbagi ◽

Ahad R. Elsadig ◽

Abdalla Elssir Ahmed ◽

Yassir Osman Mohammed ◽

...

Keyword(s):

Genetic Diversity ◽

Plasmodium Falciparum ◽

Malaria Vaccine ◽

Geographical Origin ◽

Vaccine Candidate ◽

Haplotype Diversity ◽

Terminal Region ◽

Circumsporozoite Protein ◽

High Conservation ◽

Malaria Vaccine Candidate

Abstract The currently used malaria vaccine; the RTS,S, is designed based on the Plasmodium falciparum circumsporozoite protein (PfCSP). The PfCSP gene, besides having different polymorphic patterns, can vary between P. falciparum isolates due to geographical origin and host immune response. Such aspects are essential when considering the deployment of the RTS,S vaccine in a certain region. Therefore, in this study we assessed the genetic diversity of P. falciparum in Sudan based on the PfCSP gene by investigating the diversity at the N-terminal, central repeat, and the C-terminal regions. The results of the N-terminal region showed the presence of 2 different haplotypes with a haplotype diversity (Hapd) of 0.425 ± 0.00727. The presence of the unique insertion of NNNGDNGREGKDEDKRDGNN was reported. The KLKQP motif was conserved in all the studied isolates. At the central repeat region, 11 haplotypes were seen with a Hapd of 0.779 ± 0.00097. The analysis of the genetic diversity in the C-terminal region showed the presence of 10 haplotypes with a Hapd of 0.457 ± 0.073. In this study, the results indicated a high conservation at the PfCSP gene. This may further contribute in understanding the genetic polymorphisms of P. falciparum prior to the deployment of the RTS,S vaccine in Sudan.

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