Maintaining immunogenicity of blood stage and sexual stage subunit malaria vaccines when formulated in combination

BACKGROUND: Eradication of Plasmodium falciparum malaria will likely require a multivalent vaccine, but the development of a highly efficacious subunit-based formulation has been challenging. We previously showed that production and immunogenicity of two leading vaccine targets, PfMSP1(19) (blood-stage) and Pfs25 (sexual stage), could be enhanced upon genetic fusion to merozoite surface protein 8 (PfMSP8). Here, we sought to optimize a Pfs25-based formulation for use in combination with rPfMSP1/8 with the goal of maintaining the immunogenicity of each subunit. METHODS: Comparative mouse studies were conducted to assess the effects of adjuvant selection (Alhydrogel vs. glucopyranosyl lipid adjuvant-stable emulsion (GLA-SE)) and antigen dose (2.5 vs. 0.5 μg) on the induction of anti-Pfs25 immune responses. The antibody response (magnitude, IgG subclass profile, and transmission-reducing activity (TRA)) and cellular responses (proliferation, cytokine production) generated in response to each formulation were assessed. Similarly, immunogenicity of a bivalent vaccine containing rPfMSP1/8 and rPfs25/8 was evaluated. RESULTS: Alum-based formulations elicited strong and comparable humoral and cellular responses regardless of antigen form (unfused rPfs25 or chimeric rPfs25/8) or dose. In contrast, GLA-SE based formulations elicited differential responses as a function of both parameters, with 2.5 μg of rPfs25/8 inducing the highest titers of functional anti-Pfs25 antibodies. Based on these data, chimeric rPfs25/8 was selected and tested in a bivalent formulation with rPfMSP1/8. Strong antibody titers against Pfs25 and PfMSP1(19) domains were induced with GLA-SE based formulations, with no indication of antigenic competition. CONCLUSIONS: We were able to generate an immunogenic bivalent vaccine designed to target multiple parasite stages that could reduce both clinical disease and parasite transmission. The use of the same PfMSP8 carrier for two different vaccine components was effective in this bivalent formulation. As such, the incorporation of additional protective targets fused to the PfMSP8 carrier into the formulation should be feasible, further broadening the protective response.