In these first-in-human studies, MSP142(FVO)/AS01 was evaluated, first in 26 malaria-naïve adults in a dose-escalation fashion, followed by administration of only the 50 μg dose to 20 malaria-exposed adults with a rabies comparator vaccine given to 10 concomitantly enrolled subjects. The MSP142/AS01 vaccine in both studies was shown to be well tolerated, with the solicited AEs profile similar to what has been seen with past studies of AS01 in combination with other recombinant proteins [21–24].
In subjects in the US study, this vaccine formulation proved to be equally immunogenic at both 10 and 50 μg doses, with relatively similar antibody titres between the two dosage groups at all time points. While not directly comparable due to population differences and malaria exposure, there were generally higher titres induced by the 50 μg dose vaccine in the US study compared to the same dose in the Kenya study. Given the relatively high levels of antibodies to MSP142 at all time points measured in both the MSP142/AS01 and rabies vaccine groups, it is possible that immune responses elicited in Kenyan adults with significant malaria exposure may interfere with the induction of antibody responses relative to the responses observed in US subjects. This issue may be less relevant for young children who have had less sustained malaria exposure and/or lower antibody titres, and thus, in this context, the vaccine may not have to overwrite clonally imprinted responses, i e, pre-existing immunity . For both studies, there was a significant increase in MSP1-specific antibody titres after the second vaccination but no further boosting of responses after the third vaccination. This phenomenon has been seen with other recombinant protein antigen/adjuvant system vaccines in malaria-naïve adults [8, 22], although the quality of the antibody responses with respect to avidity or subclass would require further evaluation. Antibody titres also appeared to be relatively stable through the follow-up period after the third vaccination.
The role that MSP119 plays in the formation of the parasitic food vacuole  suggests that antibodies that are directed against MSP119 may be able to interfere with the intra-erythrocytic parasite development . In order to evaluate functional antibody responses induced by the vaccine, the in vitro GIA was employed to measure the effect of antibody on parasite growth and development. In both the US and Kenya studies, it appears that MSP142 FVO/AS01 is capable of inducing some functional GIA activity in a few subjects when tested at 20% (v/v), primarily directed against the homologous FVO parasites and not the heterologous 3D7 P. falciparum parasites. In the previous clinical trial using the alternative allele, MSP142 3D7/AS02 , albeit with a different Adjuvant System, three out of five subjects from the 50 μg dose achieved similar levels of inhibitory responses against the homologous allele parasites; however, when this same vaccine was tested in 200 children aged 12 to 48 months residing in Western Kenya in a Phase 2 trial, there was no apparent efficacy . Evaluation of their immune serum tested at 5% (v/v) showed no vaccine-related GIA activity; however significant inhibition was detected predominantly toward FVO allele parasites, which were not vaccine related and most likely due to natural exposure [manuscript in preparation, E Angov]. Other trials with MSP1 vaccine candidates have reported similar discordance between the magnitude of antibody titres and functional GIA activity [15, 27, 28].
Although the functional antibodies detected by GIA to blood-stage vaccine antigens such as MSP142 have been proposed as putative surrogates of antigen quality and protective potential, there is little direct evidence supporting this concept outside of findings from epidemiologic studies evaluating the relationship of MSP1 antibody titres and functional inhibition by GIA with reduced parasite densities or clinical disease [29–33]. Studies from endemic areas suggest that the acquisition of inhibitory antibodies may be dependent upon the target antigen (MSP1, AMA1, etc.) , and the age of the individual whereby growth inhibitory activity had an inverse association with increasing age [29, 31, 32]. In a study in Kenyan children and adults, increased levels of growth inhibitory antibodies correlated with a modest delay to time to infection but only in younger children . Similar findings were recently also reported for West Africa where reduced malaria risk in children and the presence of inhibitory antibodies contributed to, but were not solely implicated in, the acquisition of protective immunity . Conversely, in a longitudinal study of children and adults in Kenya , support for a link between levels of MSP1-specific antibodies, growth inhibition and risk of symptomatic malaria was not observed. Further complicating this picture is the existence of “blocking” antibodies, which “block” the ability of “inhibitory” antibodies to act against the parasite as well as neutral antibodies, those that do not inhibit nor block . Several studies report that children naturally exposed to malaria may develop both blocking and/or neutral antibodies to MSP1 in addition to inhibitory antibodies [35, 36]. Thus the humoral immune response induced in malaria-naïve vaccinated subjects may differ qualitatively from those living in endemic areas who may already have established blocking and/or neutral antibody specificities.
It should be cautioned that one should not rely solely on an unvalidated in vitro functional assay such as the GIA when making clinical development decisions on progression of blood stage vaccine candidates. Since no surrogate markers of protection have been identified using active immunization, it is not known whether the absence of significant GIA responses in this study is meaningful. Induction of antibodies that function to inhibit erythrocyte invasion in malaria-experienced populations is influenced by many factors, including age and transmission level, as well as the heterogeneity of circulating parasite strains, and since malaria parasites appear to use redundant invasion pathways, the current GIA readout methods may not fully capture the complete effect of anti-parasite activity. The malaria vaccine researchers at WRAIR are currently re-examining the allele-specific effects from the Phase 2 study that used the 3D7 MSP142/AS02 formulation in order to assess whether specific parasite genotypes in breakthrough malaria infections in the vaccinated group signified selection against the vaccine strain . In addition, the evaluation of new surrogate functional assays is urgently required to assist in down selection of promising blood stage vaccine candidates. Towards this end, the WRAIR has recently improved the development of a promising new functional in vivo assay which measures the ability of passively transferred human immunoglobulin to protected mice challenged with transgenic Plasmodium berghei parasites containing the p19 of P. falciparum D10 strain . The advantage of such a transgenic model is that immune effectors cells are present, thus enabling participation of Fc-receptor positive cells in the clearance of the parasite as proposed previously [38, 39]. In addition to the current formulations, alternative vaccine platforms such as particle-based delivery and/or viral vectors could enhance the immunogenicity of MSP1 and increase its utility as a component of second-generation malaria vaccines.