Evaluation of a Plasmodium-Specific Carrier Protein To Enhance Production of Recombinant s25, a Leading Transmission-Blocking Vaccine Candidate.

Challenges with the production and suboptimal immunogenicity of malaria vaccine candidates have slowed the development of a multiantigen vaccine. Attempting to resolve these issues, we focused on the use of highly immunogenic merozoite surface protein 8 (MSP8) as a vaccine carrier protein. Previously, we showed that a genetic fusion of the C-terminal 19-kDa fragment of merozoite surface protein 1 (MSP1) to MSP8 (MSP8) facilitated antigen production and folding and the induction of neutralizing antibodies to conformational B cell epitopes of MSP1 Here, using the MSP1/8 construct, we further optimized the recombinant MSP8 (rMSP8) carrier by the introduction of two cysteine-to-serine substitutions (CΔS) to improve the yield of the monomeric product. We then sought to test the broad applicability of this approach using the transmission-blocking vaccine candidate s25. The production of rs25-based vaccines has presented challenges. Antibodies directed against the four highly constrained epidermal growth factor (EGF)-like domains of s25 block sexual-stage development in mosquitoes. The sequence encoding mature s25 was codon harmonized for expression in We produced a rs25-MSP8 fusion protein [rs25/8(CΔS)] as well as unfused, mature rs25. rs25 was purified with a modest yield but required the incorporation of refolding protocols to obtain a proper conformation. In comparison, chimeric rs25/8(CΔS) was expressed and easily purified, with the s25 domain bearing the proper conformation without renaturation. Both antigens were immunogenic in rabbits, inducing IgG that bound native s25 and exhibited potent transmission-reducing activity. These data further demonstrate the utility of MSP8 as a parasite-specific carrier protein to enhance the production of complex malaria vaccine targets.