Parity-Dependent Recognition of DBL1X-3X Suggests an Important Role of the VAR2CSA High-Affinity CSA-Binding Region in the Development of the Humoral Response against Placental Malaria

Abstract

f Plasmodium falciparum multidomain protein VAR2CSA stands today as the leading vaccine candidate against pregnancy-associated malaria (PAM). Most of the studies aiming to decrypt how naturally acquired immunity develops have assessed the immune recognition of individual VAR2CSA Duffy-binding-like (DBL) domains, thus overlooking the presence of conformational epitopes resulting from the overall folding of the full-length protein. In order to characterize the development of humoral immunity toward VAR2CSA, we made use of a large cohort of 293 Senegalese pregnant women to assess the level of recognition by plasma IgG of the full-length VAR2CSA protein of the 3D7 parasite strain (3D7-VAR2CSA), the CSA-binding multidomains 3D7-DBL1X to -DBL3X (3D7-DBL1X-3X), and the CSA nonbinding multidomains 3D7-DBL4 to -DBL6 (3D7-DBL4-6), as well as individual 3D7-DBL domains. Our results revealed a parity-dependent recognition of the full-length 3D7-VAR2CSA and of the CSA-binding region, 3D7-DBL1X-3X. Indeed, multigravid women possess significantly higher levels of antibodies directed against these constructs than primigravidae. Our results suggest an important role of antibodies targeting the CSA-binding region in the development of immunity against PAM, therefore providing new insights on how natural protection might be acquired and further information for the design of VAR2CSA-based vaccines. E ach year, pregnancy-associated malaria (PAM) is responsible for the deaths of as many as 363,000 neonates and for at least 10,000 maternal deaths worldwide (1). The massive accumulation of Plasmodium falciparum-infected erythrocytes (IEs) to the syncytiotrophoblast layer composing the maternal face of the placenta limits maternal-fetal exchanges, leading to clinical complications for both mother and child (2). This phenomenon is mediated by the VAR2CSA protein, a member of the P. falciparum erythrocyte membrane protein 1 (PfEMP1) adhesins, that is preferentially expressed by placental parasites (3-5) and binds to the placental chondroitin-4-sulfate A (CSA) (6). After one or two pregnancies, women acquire antibodies targeting VAR2CSA that inhibit IE sequestration and decrease the negative outcomes of PAM (7,8). In vitro assays have shown that antibodies from multigravid women are able to block the interactions between CSA and IEs originating from different parts of the world (9), showing the existence of a cross-reactive antibody response and suggesting that a relative conservation of important inhibitory epitopes within VAR2CSA proteins may exit.VAR2CSA is a 350-kDa transmembrane protein composed of six Duffy-binding-like (DBL) domains. Previous works have described the CSA-binding properties of single DBL domains (3, 10, 11). Insight derived from structural studies of DBL domains revealed the importance of the DBL3X domain in VAR2CSA CSA binding but also suggested a higher-order structure organization of VAR2CSA where the positioning of different surfaces involved in chondroitin sulfate proteoglycan (CSPG) binding forms a specific bin...

Beninese children with cerebral malaria do not develop humoral immunity against the IT4-VAR19-DC8 PfEMP1 variant linked to EPCR and brain endothelial binding

“…Optical density obtained by ELISA was converted into arbitrary units (AU) using the following formula: AU = 100 × (LnOD test − LnOD negative sample )/(LnOD positive sample − LnOD negative sample ) [ 32 ]. Negative and positive plasma samples were chosen based on their reactivity to AMA1 already tested [ 33 ].…”

Antibody responses to the full-length VAR2CSA and its DBL domains in Cameroonian children and teenagers

“…These differences in responses are important in the light of correlates of protection from PM. Correlates of protection from PM include breadth of responses to multiple VAR2CSA domains, high avidity Ab to FV2 and high levels of Ab to DBL1-2, DBL3, DBL1-3x [37, 47–50]. In addition, it is not clear how long Ab to DBL2 and DBL4 persist.…”

Clinical development of placental malaria vaccines and immunoassays harmonization: a workshop report

“…The advantages of the ELISA are its robustness and its amenability to automation, standardization and harmonization and easy transfer of protocols/standard operating procedures (SOPs) between different laboratories. ELISA has been extensively used by both teams in pre-clinical studies and was proven as adequate to detect naturally acquired human antibodies directed towards VAR2CSA-based proteins, such as the ones comprising the current vaccine candidates [ 27 , 28 ].…”