Plasmodium falciparum is unable to synthesize purine bases and relies upon purine salvage and purine recycling to meet its purine needs. We report that purines formed as products of polyamine synthesis are recycled in a novel pathway in which 5 -methylthioinosine is generated by adenosine deaminase. The action of P. falciparum purine nucleoside phosphorylase is a convergent step of purine salvage, converting both 5 -methylthioinosine and inosine to hypoxanthine. We used accelerator mass spectrometry to verify that 5 -methylthioinosine is an active nucleic acid precursor in P. falciparum. Prior studies have shown that inhibitors of purine salvage enzymes kill malaria, but potent malaria-specific inhibitors of these enzymes have not been described previously. 5 -Methylthio-immucillin-H, a transition state analogue inhibitor that is selective for malarial relative to human purine nucleoside phosphorylase, kills P. falciparum in culture. Immucillins are currently in clinical trials for other indications and may also have application as anti-malarials.

Background. Vaccines that block human-to-mosquito Plasmodium transmission are needed for malaria eradication and clinical trials have targeted zygote antigen Pfs25 for decades. We reported that a Pfs25 protein-protein conjugate vaccine formulated in alum adjuvant induced significant serum functional activity in both US and Malian adults. However, antibody levels declined rapidly, and transmission-reducing activity required four vaccine doses. Functional immunogenicity and durability must be improved before advancing TBV further in clinical development. We hypothesized that the pre-fertilization protein Pfs230 alone or in combination with Pfs25 would improve functional activity. Methods. Transmission-blocking vaccine candidates based on gamete antigen Pfs230 or Pfs25 were conjugated with Exoprotein A, formulated in Alhydrogel ®, and administered to mice, rhesus macaques, and humans. Antibody levels were measured by ELISA and transmission-reducing activity was assess by the Standard Membrane Feeding Assay. Results. Pfs25-EPA/Alhydrogel ® and Pfs230D1-EPA/Alhydrogel ® induced similar serum functional activity in mice, but Pfs230D1-EPA induced significantly greater activity in rhesus monkeys that was enhanced by complement. In U.S. adults, two vaccine doses induced complement-dependent activity in 4 of 5 Pfs230D1-EPA/Alhydrogel® recipients but no significant activity in five Pfs25-EPA recipients, and combination with Pfs25-EPA did not increase activity over Pfs230D1-EPA alone. Conclusion. The complement-dependent functional immunogenicity of Pfs230D1-EPA represents a significant improvement over Pfs25-EPA in this comparative study. The rhesus model is more predictive of the functional human immune response to Pfs230D1 than is the mouse model.

BLI is a noninvasive technique that is useful for screening potential drugs and candidate vaccines with which to combat malaria. The prospect of cross-stage protective immunity increases the number of avenues to be explored in the development of an effective vaccine against malaria.

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BackgroundVaccines that block human-to-mosquito Plasmodium transmission are needed for malaria eradication and clinical trials have targeted zygote antigen Pfs25 for decades. We reported that a Pfs25 protein-protein conjugate vaccine formulated in alum adjuvant induced significant serum functional activity in both US and Malian adults. However, antibody titers declined rapidly, and transmission-reducing activity required four vaccine doses. Functional immunogenicity and durability must be improved before advancing TBV further in clinical development. We hypothesized that the pre-fertilization protein Pfs230 alone or in combination with Pfs25 would improve functional activity.MethodsTransmission-blocking vaccine candidates based on gamete antigen Pfs230 or Pfs25 were conjugated with Exoprotein A, formulated in Alhydrogel®, and administered to mice, rhesus macaques, and humans. Antibody titers were measured by ELISA and transmission-reducing activity was assess by the Standard Membrane Feeding Assay.ResultsPfs25-EPA/Alhydrogel® and Pfs230D1-EPA/Alhydrogel® induced similar serum functional activity in mice, but Pfs230D1-EPA induced significantly greater activity in rhesus monkeys that was enhanced by complement. In U.S. adults, two vaccine doses induced complement-dependent activity in 4 of 5 Pfs230D1-EPA/Alhydrogel® recipients but no significant activity in five Pfs25-EPA recipients, and combination with Pfs25-EPA did not increase activity over Pfs230D1-EPA alone.ConclusionThe complement-dependent functional immunogenicity of Pfs230D1-EPA represents a significant improvement over Pfs25-EPA. The rhesus model is more predictive of the functional human immune response to Pfs230D1 than is the mouse model.Trial RegistrationClinicalTrials.govNCT02334462FundingThis work was supported by the Intramural Research Program of the National Institute of Allergy and Infectious Diseases, National Institutes of Health.