Expanding the Malaria Antibody Toolkit: Development and Characterisation of Plasmodium falciparum RH5, CyRPA, and CSP Recombinant Human Monoclonal Antibodies

Nacer, Adéla; Kivi, Gaily; Pert, Raini; Juronen, Erkki; Holenya, Pavlo; Aliprandini, Eduardo; Amino, Rogério; Silvie, Olivier; Quinkert, Doris; Duff, Yann Le; Hurley, Margaret M.; Reimer, Ulf; Tover, Andres; Draper, Simon J.; Gilbert, Sarah C.; Ho, Mei Mei; Bowyer, Paul W.

doi:10.3389/fcimb.2022.901253

Cited by 9 publications

(7 citation statements)

References 61 publications

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“…Cy.003 14, 35 was transiently expressed in Expi293F™ cells. Culture supernatants were harvested and 0.45 µm filtered before loading on to a 1 ml HiTrap™ Protein G HP column (Cytiva) pre-equilibrated in PBS (79382, Sigma-Aldrich).…”

Section: Methodsmentioning

confidence: 99%

Structure of the PfRCR complex which bridges the malaria parasite and erythrocyte during invasion

Farrell

Alam

Hart

et al. 2023

Preprint

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The symptoms of malaria occur during the blood stage of infection, when parasites invade and replicate within human erythrocytes. The five-component PfPCRCR complex, containing PfRH5, PfCyRPA, PfRIPR, PfCSS and PfPTRAMP, is essential for erythrocyte invasion by the deadliest human malaria parasite, Plasmodium falciparum. Invasion can be prevented by antibodies or nanobodies against each of these five conserved proteins, making them the leading blood stage malaria vaccine candidates. However, little is known about the molecular mechanism by which PfPCRCR functions during invasion. Here we present the structure of the PfRCR complex, containing PfRH5, PfCyRPA and PfRIPR, determined by cryogenic-electron microscopy. This reveals that PfRIPR consists of an ordered multi-domain core flexibly linked to an elongated tail. We test the hypothesis that PfRH5 opens to insert into the membrane, but instead show that a rigid, disulphide-locked PfRH5 can mediate efficient erythrocyte invasion. Finally, we show that the elongated tail of PfRIPR, which is the target of growth-neutralising antibodies, binds to the PfCSS-PfPTRAMP complex on the parasite membrane. Therefore, a modular PfRIPR is linked to the merozoite membrane through an elongated tail, while its structured core presents PfCyRPA and PfRH5 to interact with erythrocyte receptors. This provides novel insight into the mechanism of erythrocyte invasion and opens the way to new approaches in rational vaccine design.

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Section: Methodsmentioning

confidence: 99%

Structure of the PfRCR complex which bridges the malaria parasite and erythrocyte during invasion

Farrell

Alam

Hart

et al. 2023

Preprint

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“…Generation of anti-RH5 and anti-CyRPA recombinant and chimeric mAbs ( Table S1 ) has been previously described 16,36,37,43,57 . These mAbs were transiently expressed in Expi293F HEK cells.…”

Section: Methodsmentioning

confidence: 99%

Development of an improved blood-stage malaria vaccine targeting the essential RH5-CyRPA-RIPR invasion complex

Williams,

King,

Pulido

et al. 2024

Preprint

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In recent years, reticulocyte-binding protein homologue 5 (RH5) has emerged as a leading blood-stage Plasmodium falciparum malaria vaccine antigen. The most advanced blood-stage vaccine candidate in a Phase 2b clinical trial, RH5.1/Matrix-M[TM], is based on a full-length soluble protein-with-adjuvant formulation. RH5 interacts with cysteine-rich protective antigen (CyRPA) and RH5-interacting protein (RIPR) to form an essential heterotrimeric RCR-complex. Here, we investigated whether a vaccine candidate based on the ternary RCR-complex could substantially improve upon the leading clinical candidate RH5.1/Matrix-M[TM] in preclinical studies. Using a panel of monoclonal antibodies (mAbs) we confirm that parasite growth-inhibitory epitopes on each antigen are exposed on the surface of the RCR-complex and that mAb pairs binding to different antigens can function additively or synergistically to mediate parasite growth inhibition activity (GIA) in vitro. However, immunisation of rats with the RCR-complex consistently fails to outperform RH5.1 alone. We show this is due to immuno-dominance of RIPR coupled with the inferior potency of anti-full length RIPR polyclonal IgG antibodies as compared to the anti-RH5 and anti-CyRPA response. To address this, we identified the growth-inhibitory antibody epitopes of RIPR are clustered within C-terminal EGF-like domains of RIPR. A fusion of these EGF domains to CyRPA, called R78C, combined with RH5.1, provided a new vaccination strategy that improves upon the levels of in vitro GIA seen with RH5.1 alone. Superiority of the combination antigen vaccine candidate was achieved by the induction of a quantitatively higher, but qualitatively similar, polyclonal antibody response that demonstrated additive GIA across the three antigen targets. These preclinical data justified the advancement of the RH5.1+R78C/Matrix-M[TM] combination vaccine to a Phase 1 clinical trial.

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“…Cy.009, Cy.007 and Cy.003 were provided by Icosagen AS through the Centre for AIDS Reagents repository at the National Institute for Biological Standards and Control, UK. These mAbs were produced through the European Commission FP7 EURIPRED project [39]. Cy.009, Cy.007 and Cy.003 were produced by Icosagen using HybriFree Technology.…”

Section: Methodsmentioning

confidence: 99%

The Dual Action of Human Antibodies Specific toPlasmodium falciparumPfRH5 and PfCyRPA: Blocking Invasion and Inactivating Extracellular Merozoites

Weiss

Ragotte

Quinkert

et al. 2023

Preprint

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The Plasmodium falciparum reticulocyte-binding protein homolog 5 (PfRH5) is the current leading blood-stage malaria vaccine candidate. PfRH5 functions as part of the pentameric PCRCR complex containing PTRAMP, CSS, PfCyRPA and PfRIPR, all of which are essential for infection of human red blood cells (RBCs). To trigger RBC invasion, PfRH5 engages with RBC protein basigin in a step termed the RH5-basigin binding stage. Although we know increasingly more about how antibodies specific for PfRH5 can block invasion, much less is known about how antibodies recognizing other members of the PCRCR complex can inhibit invasion. To address this, we performed live cell imaging using monoclonal antibodies (mAbs) which bind PfRH5 and PfCyRPA. We measured the degree and timing of the invasion inhibition, the stage at which it occurred, as well as subsequent events. We show that parasite invasion is blocked by individual mAbs, and the degree of inhibition is enhanced when combining a mAb specific for PfRH5 with one binding PfCyRPA. In addition to directly establishing the invasion-blocking capacity of the mAbs, we identified a secondary action of certain mAbs on extracellular parasites that had not yet invaded where the mAbs appeared to inactivate the parasites by triggering a developmental pathway normally only seen after successful invasion. These findings suggest that epitopes within the PfCyRPA-PfRH5 sub-complex that elicit these dual responses may be more effective immunogens than neighboring epitopes by both blocking parasites from invading and rapidly inactivating extracellular parasites. These two protective mechanisms, prevention of invasion and inactivation of uninvaded parasites, resulting from antibody to a single epitope indicate a possible route to the development of more effective vaccines.

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Expanding the Malaria Antibody Toolkit: Development and Characterisation of Plasmodium falciparum RH5, CyRPA, and CSP Recombinant Human Monoclonal Antibodies

Cited by 9 publications

References 61 publications

Structure of the PfRCR complex which bridges the malaria parasite and erythrocyte during invasion

Structure of the PfRCR complex which bridges the malaria parasite and erythrocyte during invasion

Development of an improved blood-stage malaria vaccine targeting the essential RH5-CyRPA-RIPR invasion complex

The Dual Action of Human Antibodies Specific toPlasmodium falciparumPfRH5 and PfCyRPA: Blocking Invasion and Inactivating Extracellular Merozoites

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