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Dissertation
The 19-kDa carboxyl-terminal region of merozoite surface protein-1 as a vaccine candidate against malaria
Doctor of Philosophy (Ph.D.), Allegheny University of the Health Sciences
May 1998
DOI:
https://doi.org/10.17918/00002807
Abstract
Recent results with primate plasmodia and rodent models of infection have focused attention on the carboxyl-terminal (C-terminal) region of the merozoite surface protein-1 (MSP-1₁₉) as one of the leading candidates for vaccination against the erythrocytic stages of malaria. This thesis contains two parts, which reveal different aspects of MSP-1₁₉ as a vaccine candidate. While the C-terminal region of MSP-1 from the two prototypic alleles of P. falciparum has been shown to be relatively conserved in laboratory-maintained strains, little data exist on sequence heterogeneity of this region in field isolates from diverse geographic areas. To address this question, DNA encoding P. falciparum MSP-1₁₉ from field samples was analyzed by a PCR-direct sequencing method. In fifteen isolates from Africa, Asia, and Latin America, only a few nucleotide changes were found leading to amino acid alterations at four positions out of 102 residues. All the cysteine residues were conserved. The four changes were E->Q at position 1644 and TSR->KNG, or KNG->TSR at positions 1691, 1700 and 1701. Importantly, we first identified a novel variant (Q-TSR) of PfMSP-1₁₉, thus this and previous data established that PfMSP-1₁₉ were also very conserved in field samples from different areas with four major variants, E-TSR, E-KNG, Q-TSR, and Q-KNG. The sequence conservation of PfMSP-1₁₉ supports its potential utility as a vaccine candidate. Immunization with plasmid DNA containing sequences encoding relevant antigens (Ags) can induce both cell-mediated and humoral immune responses against pathogenic microorganisms in several animal models. This new technology is well suited for malarial vaccine development, since ideally an effective malarial vaccine should be multivalent and induce different immune responses against multi-stages of its life cycle. Our previous results demonstrated that when the C-terminus of Plasmodium yoelii MSP-1 (PyC2) was expressed as a fusion protein (GST-PyC2) with glutathione S-transferase (GST), it elicited antibody (Ab)-mediated protective immune responses in BALB/c mice. In the second part of my thesis, I extended this study to DNA vaccination in the same rodent model and characterized the humoral immune responses induced by a DNA vaccine encoding GST-PyC2 (V3). The GST-PyC2 expressed in V3-transfected Cos 7 cells was recognized by a protective monoclonal Ab to PyC2 (mAb302), although the secreted product had undergone N-linked glycosylation. When BALB/c mice were immunized with V3 plasmid, anti-PyC2 Abs were successfully induced. These Abs immunoprecipitated native PyMSP-1 protein and competed with mAb302 for binding to its epitope at a level similar to those elicited by GST-PyC2 protein immunization. However, DNA-induced Abs had significantly lower titers and avidities, and different isotype profiles and protective capacities against a lethal erythrocytic stage challenge than those resulting from immunization with GST-PyC2 protein. Most surprising was the finding that, in contrast to protein immunization, there was no significant increase in the avidity of either GST-specific or PyC2-specific IgG Abs during the course of DNA immunization. This suggests that there may be no or little affinity maturation of specific Abs during DNA immunization in this system. To our knowledge, this is the first demonstration of lack of affinity maturation of Abs in DNA vaccinations. This may be very important in further understanding the mechanism involved in DNA vaccination, especially in cases that humoral immune responses are desired. Furthermore, when mice were immunized with V3 DNA vaccine in the Ribi adjuvant system (RAS), specific antibodies with slightly higher titers and similar avidities were induced. Importantly after lethal blood stage challenge, all V3+RAS immunized mice delayed their peak of parasitemias by two days and half of them cleared the parasitemias by day 18. These data indicate it is possible that M SP-1₁₉-based DNA vaccines will achieve complete protection against blood stages of malaria.
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Details
- Title
- The 19-kDa carboxyl-terminal region of merozoite surface protein-1 as a vaccine candidate against malaria
- Creators
- Yang Kang
- Contributors
- Carole A. Long (Advisor) - Drexel University, Allegheny University of the Health Sciences (1996-1998)
- Awarding Institution
- Allegheny University of the Health Sciences
- Degree Awarded
- Doctor of Philosophy (Ph.D.)
- Publisher
- Allegheny University of the Health Sciences; Philadelphia, Pennsylvania
- Number of pages
- xiii, 147 pages
- Resource Type
- Dissertation
- Language
- English
- Academic Unit
- Microbiology and Immunology (1996-1998); Allegheny University of the Health Sciences (1996-1998); School of Medicine (1996-1998)
- Other Identifier
- 991021888171804721