Characterization of a novel inhibitory human monoclonal antibody directed against Plasmodium falciparum Apical Membrane Antigen 1.

Dominika J Maskus,Otchere Addai-Mensah,Rolf Fendel,Rainer Fischer,Michał Królik,Susanne Bethke,Stefan Barth,Andreas Reimann,Melanie Seidel,Torsten Klockenbring,Holger Spiegel,Stephanie Kapelski

doi:10.1038/srep39462

Abstract

Malaria remains a major challenge to global health causing extensive morbidity and mortality. Yet, there is no efficient vaccine and the immune response remains incompletely understood. Apical Membrane Antigen 1 (AMA1), a leading vaccine candidate, plays a key role during merozoite invasion into erythrocytes by interacting with Rhoptry Neck Protein 2 (RON2). We generated a human anti-AMA1-antibody (humAbAMA1) by EBV-transformation of sorted B-lymphocytes from a Ghanaian donor and subsequent rescue of antibody variable regions. The antibody was expressed in Nicotiana benthamiana and in HEK239-6E, characterized for binding specificity and epitope, and analyzed for its inhibitory effect on Plasmodium falciparum. The generated humAbAMA1 shows an affinity of 106–135 pM. It inhibits the parasite strain 3D7A growth in vitro with an expression system-independent IC50-value of 35 μg/ml (95% confidence interval: 33 μg/ml–37 μg/ml), which is three to eight times lower than the IC50-values of inhibitory antibodies 4G2 and 1F9. The epitope was mapped to the close proximity of the RON2-peptide binding groove. Competition for binding between the RON2-peptide and humAbAMA1 was confirmed by surface plasmon resonance spectroscopy measurements. The particularly advantageous inhibitory activity of this fully human antibody might provide a basis for future therapeutic applications.

Highlights

Of erythrocyte invasion by merozoites, (2) the neutralization of merozoites by agglutination, (3) the initiation of the complement cascade resulting in further opsonization and lysis, and (4) the recruitment of neutrophilic granulocytes and monocytes/macrophages[12,13,14,15,16,17,18,19,20]
In order to choose a promising candidate for the generation of an Apical Membrane Antigen 1 (AMA1)-specific human monoclonal antibody, plasma of 31 adult Ghanaian blood donors were screened by indirect ELISA for IgG reactivity against different recombinant variants of AMA1, including the allelic variant of Plasmodium falciparum strain 3D7, as well as a mixture of three artificial diversity covering variants of AMA1
We found that P. falciparum strain 3D7A was reproducibly inhibited by plant-produced humAbAMA1 with an IC50 value of 35 μg/ml (95% confidence interval (CI): 33–37 μg/ml)

Summary

Introduction

Of erythrocyte invasion by merozoites, (2) the neutralization of merozoites by agglutination, (3) the initiation of the complement cascade resulting in further opsonization and lysis, and (4) the recruitment of neutrophilic granulocytes and monocytes/macrophages[12,13,14,15,16,17,18,19,20]. Many anti-plasmodial monoclonal antibodies (mAbs) have been generated in mice or other rodents which helped to gain valuable insights into the functions of a plethora of plasmodial proteins[40,41,42,43,44]. Such mAbs do not necessarily reflect the naturally acquired anti-plasmodial immunoglobulin repertoire in humans which takes years – or even decades – to develop. Only few human anti-plasmodial monoclonal antibodies (humAbs) have been generated Among these are humAbs directed at Merozoite Surface Protein 1 (MSP1), MSP2, MSP3, MSP10, NPNA1, Pfs48/45, and VAR2CSA45–52. We report the isolation, expression and characterization of the first human monoclonal antibody recognizing P. falciparum AMA1, called humAbAMA1

Methods

Results

Conclusion