Abstract
The Plasmodium falciparum Pfs230 and Pfs48/45 proteins are leading candidates for a malaria transmission-blocking vaccine (TBV). Previously, we showed that a Pfs230–Pfs48/45 fusion protein elicits higher levels of functional antibodies than the individual antigens, but low yields hampered progression to clinical evaluation. Here we identified a modified construct (ProC6C) with a circumsporozoite protein (CSP) repeat-linker sequence that enhances expression. A scalable and reproducible process in the Lactococcus lactis expression system was developed and ProC6C was successfully transferred for manufacturing under current Good Manufacturing Practices (cGMP). In addition, a panel of analytical assays for release and stability were developed. Intact mass spectrometry analysis and multiangle light scattering showed that the protein contained correct disulfide bonds and was monomeric. Immunogenicity studies in mice showed that the ProC6C adsorbed to Alhydrogel®, with or without Matrix-MTM, elicited functional antibodies that reduced transmission to mosquitoes and sporozoite invasion of human hepatocytes. Altogether, our data support manufacture and clinical evaluation of ProC6C as a multistage malaria-vaccine candidate.
Highlights
Malaria is a vector-borne disease caused by Plasmodium parasites, of which Plasmodium falciparum causes the highest morbidity and mortality worldwide
Malaria transmissionblocking interventions have largely focused on vector control, but the increase in insecticide resistance calls for new tools
While others have focused on Pfs[230] Domain 15, here we focused on the N-terminal Pro-domain, absent of cysteines to prevent scrambled disulfide bonds, while maintaining some transmission-reducing activity[6,7]
Summary
Malaria is a vector-borne disease caused by Plasmodium parasites, of which Plasmodium falciparum causes the highest morbidity and mortality worldwide. Malaria transmission-blocking vaccines (TBV) hold the potential to block malaria transmission in the population, thereby contributing to malaria elimination Such vaccines aim to produce specific antibodies against functionally important proteins expressed during parasite development in the mosquito. Pfs[230] and Pfs48/ 45 are expressed during gametocyte development in humans as well as in mosquitoes where they form a protein complex on the surface of the P. falciparum gamete[3]. To explore whether there is an additive or synergistic activity between Pfs[230] and Pfs48/45 antigens, we have screened a series of chimeric vaccine antigens composed of different Pfs48/ 45 and Pfs[230] domains in preclinical models[13] In these studies, dual-antigen vaccines elicited higher levels of functional antibodies than the corresponding single-antigen vaccines. Assuming that complexity of disulfide-bond formation might reduce the overall secreted-protein yield of the chimera, we decided to focus on the N-terminal Pro domain of
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