key: cord-0261432-mxw2olzw authors: Larsen, Mads Delbo; Lopez-Perez, Mary; Dickson, Emmanuel Kakra; Ampomah, Paulina; Ndam, Nicaise Tuikue; Nouta, Jan; Koeleman, Carolien A M; Ederveen, Agnes L Hipgrave; Mordmüller, Benjamin; Salanti, Ali; Nielsen, Morten Agertoug; Massougbodji, Achille; van der Schoot, C. Ellen; Ofori, Michael F.; Wuhrer, Manfred; Hviid, Lars; Vidarsson, Gestur title: Afucosylated Plasmodium falciparum-specific IgG is induced by infection but not by subunit vaccination date: 2021-04-23 journal: bioRxiv DOI: 10.1101/2021.04.23.441082 sha: aa2afb48eef3a2670eb4598bc2d798aac5e7b74c doc_id: 261432 cord_uid: mxw2olzw IgG specific for members of the Plasmodium falciparum erythrocyte membrane protein 1(PfEMP1) family, which mediates receptor- and tissue-specific sequestration of infected erythrocytes (IEs), is a central component of naturally acquired malaria immunity. PfEMP1-specific IgG is thought to protect via inhibition of IE sequestration, and through IgG-Fc Receptor (FcγR) mediated phagocytosis and killing of antibody-opsonized IEs. The affinity of afucosylated IgG to FcγRIIIa is elevated up to 40-fold compared to fucosylated IgG, resulting in enhanced antibody-dependent cellular cytotoxicity. Most IgG in plasma is fully fucosylated, but afucosylated IgG is elicited in response to enveloped viruses and to paternal alloantigens during pregnancy. Here we show that naturally acquired PfEMP1-specific IgG is likewise markedly afucosylated in a stable and exposure-dependent manner, and efficiently induces FcγRIIIa-dependent natural killer (NK) cell degranulation. In contrast, immunization with a soluble subunit vaccine based on VAR2CSA-type PfEMP1 resulted in fully fucosylated specific IgG. These results have implications for understanding natural and vaccine-induced antibody-mediated protective immunity to malaria. Summary Afucosylated IgG has enhanced Fc-receptor affinity and functionality, and is formed specifically against membrane proteins of enveloped viruses. We show that this also applies to Plasmodium falciparum erythrocyte membrane-specific IgG induced by natural infection, but not by soluble PfEMP1 vaccination. The most severe form of malaria is caused by the protozoan parasite Plasmodium falciparum. 57 The disease is currently estimated to cost around 400,000 lives a year, mostly of young children and 58 pregnant women in sub-Saharan Africa. In addition, nearly 900,000 babies are born with a low birth 59 weight as a consequence of placental malaria (PM) (World Health Organization, 2020). The Acquired immunity mediated by PfEMP1-specific IgG is generally thought to rely on their 92 ability to interfere directly with IE sequestration (i.e., neutralizing, adhesion-inhibitory antibodies). 93 However, antibody-mediated opsonization of IEs is a likely additional effector function of these 94 antibodies, since the antibody response to most P. falciparum asexual blood-stage antigens 95 (including PfEMP1) is completely dominated by the cytophilic subclasses IgG1 and (to a lesser In this study, we tested the hypothesis that antibody responses to P. falciparum antigens expressed 133 on the IE surface are also a subject to afucosylation. To this end, we examined naturally acquired 134 IgG responses to the PfEMP1 antigens VAR6 and VAR2CSA and to the merozoite antigen 135 GLURP, and VAR2CSA-specific IgG induced by subunit vaccination. 137 Naturally acquired PfEMP1-specific IgG is highly afucosylated 138 We first used a set of plasma samples collected from 127 pregnant Ghanaian women at the 139 time of their first visit to antenatal clinics (Ofori et al., 2009), to assess N297 glycosylation of IgG 140 with specificity for three P. falciparum recombinant antigens. We used the full ectodomains of 141 VAR2CSA, and the non-pregnancy-restricted Group A-type VAR6, which are both naturally 142 expressed on the IE surface. We also included the merozoite antigen GLURP, which is not Fc fucosylation was generally high, also in line with our hypothesis (Fig. 2A) . A few women 155 showed marked afucosylation of GLURP-specific IgG1 ( Fig. 2A) Fig. 1A-B) . Levels of 160 bisecting GlcNAc were lower for VAR2CSA-and VAR6-specific IgG1, and higher for GLURP- 161 specific IgG1 compared to total IgG1 ( Supplementary Fig. 1C ). These results indicate that antigen-162 specific IgG levels are modulated in complex ways according to exposure and antigen context. 163 Afucosylation of VAR2CSA-specific IgG1 was generally less pronounced than that of VAR6-164 specific IgG1 ( Fig. 2A) . Exposure to VAR2CSA-type PfEMP1 occurs later in life, as it is restricted 177 The above findings support the hypothesis that afucosylated IgG specific for host membrane-178 associated immunogens is attained following repeated exposure and that the phenotype is stable 179 once acquired. To examine this hypothesis further, and to consolidate the findings described above, 180 we proceeded to determine the Fc fucosylation of IgG with specificity for the same three antigens, Acknowledgments 386 We are grateful to all the individuals donating blood samples for this study, and to the 387 scientists and health workers participating in the studies for which they were originally collected. 388 We thank Michael Theisen (University of Copenhagen and Statens Seruminstitut) for GLURP C q qq q q q q q q q qq q q qq q q q q q q qqq q q q q q q q q q q q qq q q q q q q q q q q q q q q q q q q q q q q q q q q **** Plasma antibodies from malaria-exposed pregnant women recognize variant surface 602 antigens on Plasmodium falciparum-infected erythrocytes in a parity-dependent manner and 603 block parasite adhesion to chondroitin sulphate A Malaria in pregnancy: 605 pathogenesis and immunity Evidence for the 608 involvement of VAR2CSA in pregnancy-associated malaria A human immunoglobulin G receptor exists in both polypeptide-anchored and 611 phosphatidylinositol-glycan-anchored forms Plasmodium 614 falciparum infection early in pregnancy has profound consequences for fetal growth Changes in antigen-specific IgG1 Fc N-618 glycosylation upon influenza and tetanus vaccination Lack of fucose on human IgG1 N-linked oligosaccharide improves binding to human FcgRIII and antibody-dependent cellular toxicity PRIMVAC vaccine adjuvanted with Alhydrogel or GLA-SE to 628 prevent placental malaria: a first-in-human, randomised, double-blind, placebo-controlled 629 study Expression of a recombinant high affinity IgG Fc receptor by engineered 632 NK cells as a docking platform for therapeutic mAbs to target cancer cells Hipgrave 635 Antigen 636 specificity determines anti-red blood cell IgG-Fc alloantibody glycosylation and thereby 637 severity of haemolytic disease of the fetus and newborn Glycosylation pattern of 640 anti-platelet IgG is stable during pregnancy and predicts clinical outcome in alloimmune 641 thrombocytopenia Var2CSA minimal CSA binding region is located within the N-terminal region Full-length extracellular region of the var2CSA variant of 648 PfEMP1 is required for specific, high-affinity binding to CSA Acquisition and decay of antibodies to pregnancy-associated variant 652 antigens on the surface of Plasmodium falciparum infected erythrocytes that are associated 653 with protection against placental parasitemia Variant 655 surface antigen-specific IgG and protection against the clinical consequences of pregnancy-656 associated Plasmodium falciparum malaria Investigating the function of F c -specific binding of IgM to Plasmodium 659 falciparum erythrocyte membrane protein 1 mediating erythrocyte rosetting Functional attributes of antibodies, effector cells, and target cells 664 affecting NK cell-mediated antibody-dependent cellular cytotoxicity Antigenicity and 667 immunogenicity of recombinant glutamate-rich protein of Plasmodium falciparum expressed 668 in Escherichia coli High level of var2csa transcription by Plasmodium 671 falciparum isolated from the placenta Dynamics of anti-VAR2CSA immunoglobulin G 674 response in a cohort of Senegalese pregnant women Severe malaria is associated with parasite binding to endothelial protein C 678 receptor A pilot study showing differences in glycosylation 681 patterns of IgG subclasses induced by pneumococcal, meningococcal, and two types of 682 influenza vaccines IgG subclasses and allotypes: from structure to 684 effector functions A single member of the Plasmodium falciparum var multigene family 687 determines cytoadhesion to the placental receptor chondroitin sulphate A IgG antibodies to dengue enhanced for FcgRIIIA binding determine 692 disease severity Regulated glycosylation patterns of IgG during alloimmune responses against human 696 platelet antigens ADCC: Antibody-dependent cellular cytotoxicity; CM: Cerebral malaria; Fc: fragment 699 crystallizable; FcγR: Fcγ receptor IgG: Immunoglobulin G; IE = Infected erythrocyte; MBD: Minimal-binding domain 701 PBS: Phosphate-buffered saline; PBS-T = PBS supplemented with TWEEN20 PfEMP1: Plasmodium falciparum erythrocyte membrane protein-1; PM: placental malaria