Pregnancy associated malaria (PAM) causing maternal anaemia, low birth weight and stillbirth, is a severe manifestation of
Plasmodium falciparum infection [
1]. PAM is caused by infected erythrocytes (IE) that sequester in the intervillous space of the placenta [
2]. The ability to sequester in the vascular bed whereby the parasite avoids immune mechanisms in the spleen is a hallmark of the particular virulence of
P. falciparum. The IE bind host receptors on various endothelia through antigens called
P. falciparum erythrocyte membrane protein 1 (PfEMP1). The PfEMP1 protein family, encoded by the
var genes, is constituted of large proteins of 150-350 kDa with several Duffy-binding-like (DBL) domains. Different PfEMP1 molecules have different receptor specificities, and clonal switching between expression of the various
var gene products, in a mutually exclusive manner, allows the parasite to modify its adhesion properties accordingly (reviewed in [
3]). There are about 60 copies of highly different
var genes in each parasite genome and a high sequence variation among genomes [
4‐
6]. Expression of different PfEMP1 variants allow the parasites to escape previously acquired antibody responses, and clinical protection occurs when a large repertoire of variant specific antibodies allows the host to control the infection [
7]. After repeated infections during childhood in endemic areas the acquired repertoire of antibodies against the variant surface antigens will progressively protect against variants expressed during severe, mild and asymptomatic infections [
8,
9]. During pregnancy the parasite again escapes acquired immunity by expressing variants not encountered during childhood disease. This is mediated by parasites occupying a new niche, the developing placenta. The interaction between parasite antigens on the surface of the IE and chondroitin sulphate A (CSA) in the placenta is one of the most direct associations between binding phenotype and disease outcome in
falciparum malaria. Placental parasites and parasite lines selected for CSA binding
in vitro express a unique
var gene named
var2csa [
10,
11]. VAR2CSA is expressed on the surface of IE panned on CSA and on IE isolated from infected placentas [
12,
13] and parasite clones where the
var2csa gene is disrupted lose the ability to bind CSA [
14]. Several domains and regions of VAR2CSA have been shown to bind CSA
in vitro, however, the specificity of single VAR2CSA domain binding to CSA does not seem to be exclusive for CSA type glycans [
15‐
18].
Women in malaria endemic areas acquire antibodies that protect against PAM as a function of parity [
1]. The mechanism of protection is suggested to be based on antibodies that block binding of IE to CSA [
19]. Likewise, high anti-VAR2CSA IgG levels are correlated with protection against the clinical consequences of PAM [
13]. These findings suggest that it is feasible to develop a VAR2CSA-based vaccine to protect women in malaria endemic areas against PAM. A challenge for vaccine development is to define VAR2CSA constructs of a size compatible with protein-vaccine production, which elicit pan-reactive antibodies that abrogate binding of parasites in the placenta. It has previously been shown that antibodies induced against DBL6-FCR3 partially inhibited parasite binding and that this inhibitory activity was only present in serum collected during the immunization but absent in the final bleed [
20]. Recently, it was shown that DBL4-FCR3 induced a broadly IgG based parasite adhesion-blocking response, which increased during the immunizations and was highly reproducible in subsequent immunizations with the same antigen [
21]. These DBL4 antibodies are currently being tested for the capacity to inhibit binding to CSA in a large panel of placental parasites. It is possible that the DBL4-FCR3 antibodies do not inhibit all isolates due to sequence variation within VAR2CSA. In that case an effective PAM vaccine would need to be based on multiple VAR2CSA serotypes. The aim of this study was to define additional DBL domain, which can induce parasite adhesion-blocking antibodies in order to make a multidomain vaccine, which presumable could protect women against multiple
P. falciparum genotypes.
Two domains, DBL1-3D7 and DBL3-HB3 T1 were shown to induce a cross-reactive inhibitory response, however, in line with the published data on DBL6-FCR3 the inhibitory response was either not sustained during the entire immunization protocol or was not reproducible.