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Expression, immunogenicity, histopathology, and potency of a mosquito-based malaria transmission-blocking recombinant vaccine.

ABSTRACT: Vaccines have been at the forefront of global research efforts to combat malaria, yet despite several vaccine candidates, this goal has yet to be realized. A potentially effective approach to disrupting the spread of malaria is the use of transmission-blocking vaccines (TBV), which prevent the development of malarial parasites within their mosquito vector, thereby abrogating the cascade of secondary infections in humans. Since malaria is transmitted to human hosts by the bite of an obligate insect vector, mosquito species in the genus Anopheles, targeting mosquito midgut antigens that serve as ligands for Plasmodium parasites represents a promising approach to breaking the transmission cycle. The midgut-specific anopheline alanyl aminopeptidase N (AnAPN1) is highly conserved across Anopheles vectors and is a putative ligand for Plasmodium ookinete invasion. We have developed a scalable, high-yield Escherichia coli expression and purification platform for the recombinant AnAPN1 TBV antigen and report on its marked vaccine potency and immunogenicity, its capacity for eliciting transmission-blocking antibodies, and its apparent lack of immunization-associated histopathologies in a small-animal model.

SUBMITTER: Mathias DK

PROVIDER: S-EPMC3318422 | biostudies-literature | 2012 Apr

REPOSITORIES: biostudies-literature

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Expression, immunogenicity, histopathology, and potency of a mosquito-based malaria transmission-blocking recombinant vaccine.

Mathias D K DK Plieskatt J L JL Armistead J S JS Bethony J M JM Abdul-Majid K B KB McMillan A A Angov E E Aryee M J MJ Zhan B B Gillespie P P Keegan B B Jariwala A R AR Rezende W W Bottazzi M E ME Scorpio D G DG Hotez P J PJ Dinglasan R R RR Dinglasan R R RR

Infection and immunity 20120206 4

Vaccines have been at the forefront of global research efforts to combat malaria, yet despite several vaccine candidates, this goal has yet to be realized. A potentially effective approach to disrupting the spread of malaria is the use of transmission-blocking vaccines (TBV), which prevent the development of malarial parasites within their mosquito vector, thereby abrogating the cascade of secondary infections in humans. Since malaria is transmitted to human hosts by the bite of an obligate inse ...[more]

PMID: 22311924

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Project description:BACKGROUND:Control and elimination of malaria can be accelerated by transmission-blocking interventions such as vaccines. A surface antigen of Plasmodium falciparum gametocytes, Pfs230, is a leading vaccine target antigen, and has recently progressed to experimental clinical trials. To support vaccine product development, an N-terminal Pfs230 antigen was designed to increase yield, as well as to improve antigen quality, integrity, and homogeneity. METHODS:A scalable baculovirus expression system was used to express the Pfs230D1+ construct (aa 552-731), which was subsequently purified and analysed. Pfs230D1+ was designed to avoid glycosylation and protease digestion, thereby potentially increasing homogeneity and stability. The resulting Pfs230D1+ protein was compared to a previous iteration of the Pfs230 N-terminal domain, Pfs230C1 (aa 443-731), through physiochemical characterization and in vivo analysis. The induction of functional antibody responses was confirmed via the standard membrane feeding assay (SMFA). RESULTS:Pfs230D1+ was produced and purified to an overall yield of 23 mg/L culture supernatant, a twofold yield increase over Pfs230C1. The Pfs230D1+ protein migrated as a single band via SDS-PAGE and was detected by anti-Pfs230C1 monoclonal antibodies. Evaluation by SDS-PAGE, chromatography (size-exclusion and reversed phase) and capillary isoelectric focusing demonstrated the molecule had improved homogeneity in terms of size, conformation, and charge. Intact mass spectrometry confirmed its molecular weight and that it was free of glycosylation, a key difference to the prior Pfs230C1 protein. The correct formation of the two intramolecular disulfide bonds was initially inferred by binding of a conformation specific monoclonal antibody and directly confirmed by LC/MS and peptide mapping. When injected into mice the Pfs230D1+ protein elicited antibodies that demonstrated transmission-reducing activity, via SMFA, comparable to Pfs230C1. CONCLUSION:By elimination of an O-glycosylation site, a potential N-glycosylation site, and two proteolytic cleavage sites, an improved N-terminal Pfs230 fragment was produced, termed D1+, which is non-glycosylated, homogeneous, and biologically active. An intact protein at higher yield than that previously observed for the Pfs230C1 fragment was achieved. The results indicate that Pfs230D1+ protein produced in the baculovirus expression system is an attractive antigen for transmission-blocking vaccine development.

Dataset Information

Expression, immunogenicity, histopathology, and potency of a mosquito-based malaria transmission-blocking recombinant vaccine.

Publications

Expression, immunogenicity, histopathology, and potency of a mosquito-based malaria transmission-blocking recombinant vaccine.

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