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Dissertation
Towards a multivalent, multistage malaria vaccine: evaluation of a Plasmodium-specific carrier protein to enhance production and immunogenicity of a transmission-blocking vaccine candidate
Doctor of Philosophy (Ph.D.), Drexel University
Apr 2018
DOI:
https://doi.org/10.17918/D87Q2K
Abstract
Progress towards the eradication of Plasmodium falciparum would be greatly enhanced by the development of a multivalent, multistage vaccine containing a transmission-blocking component. Unfortunately, several challenges have impeded the development of such a formulation, including 1) the recombinant production of complex targets in their proper native conformations, 2) the inherent, suboptimal immunogenicity of many protective candidates and 3) the induction and maintenance of protective responses against individual components in a multivalent context. Previous studies with PfMSP1, a protective blood-stage target that embodies these challenges, demonstrated that many of these issues could be addressed through the use of merozoite surface protein 8 (PfMSP8) as a carrier protein. We therefore applied this strategy to transmission-blocking vaccine candidate Pfs25. Antibodies directed against conformational epitopes within the four EGF-like domains of Pfs25 are capable of blocking sexual stage development in the mosquito vector. Using an E. coli expression system, we produced an rPfs25-PfMSP8 fusion protein (rPfs25/8(C[delta]S)) and an unfused, single rPfs25 antigen (rPfs25). rPfs25 was purified with a modest yield and required refolding procedures to obtain proper conformation. Alternatively, rPfs25/8(C[delta]S) was purified in higher yield without the need for refolding, via denaturation and renaturation, of the Pfs25 domain. Both antigens were immunogenic in rabbits, inducing IgG that bound native, parasite-associated Pfs25 and exhibited potent transmission-reducing activity. Comparative mouse studies were conducted to assess the effects of adjuvant selection (Alum vs. GLA-SE) and antigen dose (2.5 vs. 0.5 microgram) on the immunogenicity of these antigens. When formulated with Alum, a Th2-biasing adjuvant, both antigens elicited strong and comparable anti-Pfs25 titers irrespective of antigen dose. However, formulations with GLA-SE, a synthetic TLR4 agonist that supports a Th1-biased response, elicited divergent responses based on antigen. Immunization with rPfs25/8(C[delta]S) induced significantly higher anti-Pfs25 titers than immunization with rPfs25 at both doses, highlighting the requirement for genetic linkage of Pfs25 to the PfMSP8(C[delta]S) carrier in order to benefit from the adjuvant effect of GLA-SE. Of note, antigenic competition was observed when rPfs25 and rPfMSP8(C[delta]S) were co-administered as individual antigens, which was avoided through immunization with chimeric rPfs25/8(C[delta]S). Importantly, IgG induced by immunization with rPfs25/8(C[delta]S) demonstrated potent transmission-reducing activity, as measured by the in vivo standard membrane feeding assay (SMFA), irrespective of adjuvant. Based on these data, the chimeric rPfs25/8(C[delta]S) antigen was selected for further testing in a bivalent formulation with blood-stage vaccine candidate rPfMSP1/8(C[delta]S). Outbred mice that were co-immunized with rPfs25/8(C[delta]S) and rPfMSP1/8(C[delta]S) antigens produced strong antibody titers against both Pfs25 and PfMSP119 that were comparable to corresponding monovalent formulations, indicating the successful avoidance of antigenic competition. Consistent with previous results, immunization with rPfs25/8(C[delta]S) elicited comparable anti-Pfs25 antibody titers irrespective of adjuvant. However, immunization with rPfMSP1/8(C[delta]S) induced anti-PfMSP119 responses that were significantly higher when adjuvanted with GLA-SE relative to Alum-based formulations. These data are promising as we continue to assess and optimize additional blood-stage antigens (PfMSP2 and PfRh5), moving towards the ultimate goal of producing a highly efficacious, multivalent vaccine to reduce both blood stage-mediated clinical disease and transmission of P. falciparum parasites.
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Details
- Title
- Towards a multivalent, multistage malaria vaccine
- Creators
- Elizabeth M. Parzych - DU
- Contributors
- James M. Burns Jr. (Advisor) - Drexel University (1970-)
- Awarding Institution
- Drexel University
- Degree Awarded
- Doctor of Philosophy (Ph.D.)
- Publisher
- Drexel University; Philadelphia, Pennsylvania
- Number of pages
- xv, 209 pages
- Resource Type
- Dissertation
- Language
- English
- Academic Unit
- Microbiology and Immunology; College of Medicine; Drexel University
- Other Identifier
- 7959; 991014632082804721