Background Plasmodium falciparum merozoite surface protein 3 is a malaria vaccine candidate that was identified, characterised, and developed based on a unique immuno-clinical approach. The vaccine construct was derived from regions fully conserved among various strains and containing B cell epitopes targeted by human antibodies (from malaria-immune adults) that are able to mediate a monocyte-dependent parasite killing effect. The corresponding long synthetic peptide was administered to 36 volunteers, with either alum or Montanide ISA720 as adjuvant.Methods and FindingsBoth formulations induced cellular and humoral immune responses. With alum, the responses lasted up to 12 mo. The vaccine-induced antibodies were predominantly of cytophilic classes, i.e., able to cooperate with effector cells. In vitro, the antibodies induced an inhibition of the P. falciparum erythrocytic growth in a monocyte-dependent manner, which was in most instances as high as or greater than that induced by natural antibodies from immune African adults. In vivo transfer of the volunteers' sera into P. falciparum–infected humanized SCID mice profoundly reduced or abrogated parasitaemia. These inhibitory effects were related to the antibody reactivity with the parasite native protein, which was seen in 60% of the volunteers, and remained in samples taken 12 mo postimmunisation.ConclusionThis is the first malaria vaccine clinical trial to clearly demonstrate antiparasitic activity by vaccine-induced antibodies by both in vitro and in vivo methods. The results, showing the induction of long-lasting antibodies directed to a fully conserved polypeptide, also challenge current concepts about malaria vaccines, such as unavoidable polymorphism, low antigenicity, and poor induction of immune memory.

Immunoglobulins from individuals with immunity to malaria have a strong antiparasitic effect when transferred to Plasmodium falciparum malaria infected patients. One prominent target of antiparasitic antibodies is the merozoite surface antigen 3 (MSP-3). We have investigated the antibody response against MSP-3 residues 194 to 257 (MSP-3 194-257 ) on the molecular level. mRNA from peripheral blood leukocytes from clinically immune individuals was used as a source of Fab (fragment antibody) genes. A Fab-phage display library was made, and three distinct antibodies designated RAM1, RAM2, and RAM3 were isolated by panning. Immunoglobulin G1 (IgG1) and IgG3 full-length antibodies have been produced in CHO cells. Reactivity with the native parasite protein was demonstrated by immunofluorescence microscopy, flow cytometry, and immunoblotting. Furthermore, the antiparasitic effect of RAM1 has been tested in vitro in an antibody-dependent cellular inhibition (ADCI) assay. Both the IgG1 and the IgG3 versions of the antibody show an inhibitory effect on parasite growth.Clinical immunity to Plasmodium falciparum malaria is gradually acquired over a dozen years of intense exposure to the parasite (12). Acquired immunity to malaria has been termed premunition and is characterized as being nonsterile and incomplete (43). The exact mechanism responsible for premunition is not known with certainty. However, a number of clinical studies carried out in the early sixties (9, 16, 23)-and subsequently confirmed and extended in the nineties (2, 33)-showed an unambiguous antiparasitic effect of antibodies transferred from adults with immunity to malaria to malariainfected infants. Clinical effects observed in one of these studies correlated with the effect measured in the in vitro assay termed antibody-dependent cellular inhibition (ADCI) (2, 4). In the ADCI assay, immune antibody cooperates with monocytes in an in vitro malaria culture, and the antiparasitic effect is demonstrated by parasite growth inhibition. It has been shown that the antibody-merozoite complex by a contact-dependent mechanism stimulates the monocyte to secrete substances toxic to the asexual blood stages. The specific substances responsible for the subsequent, non-contact-dependent parasite growth inhibition include tumor necrosis factor alpha together with other molecules that are yet to be identified (4). The ADCI assay has been used for identification and characterization of the merozoite surface protein 3 (MSP-3) (27).An invariable structural feature of all reported MSP-3 sequences is the presence of three regions each of which contains three, four, or five conserved heptad repeat units. Previously published structural analyses suggest that the heptad repeat regions have an amphipathic alpha-helical secondary structure. A coiled-coil bundle conformation including these regions is a theoretical possibility supported by experimental data (24). The C-terminal part of MSP-3 contains a leucine zipper-like domain possibly implicated in dimerization and the formation...