Restricted valency (NPNA)n repeats and junctional epitope-based circumsporozoite protein vaccines against Plasmodium falciparum

Robin Miller,Adrian H Batchelor,Sofya Savransky,Xiaoyan Zou,Arasu Balasubramaniyam,Mark D Langowski,Dallas R Brown,Lorraine A Soisson,William B Harrison,Farhat A Khan,Sheetij Dutta,Zoltan Beck,Jason A Regules,Gary R Matyas

doi:10.1038/s41541-022-00430-y

Abstract

The Circumsporozoite Protein (CSP) of Plasmodium falciparum contains an N-terminal region, a conserved Region I (RI), a junctional region, 25–42 copies of major (NPNA) and minor repeats followed by a C-terminal domain. The recently approved malaria vaccine, RTS,S/AS01 contains NPNAx19 and the C-terminal region of CSP. The efficacy of RTS,S against natural infection is low and short-lived, and mapping epitopes of inhibitory monoclonal antibodies may allow for rational improvement of CSP vaccines. Tobacco Mosaic Virus (TMV) was used here to display the junctional epitope (mAb CIS43), Region I (mAb 5D5), NPNAx5, and NPNAx20 epitope of CSP (mAbs 317 and 580). Protection studies in mice revealed that Region I did not elicit protective antibodies, and polyclonal antibodies against the junctional epitope showed equivalent protection to NPNAx5. Combining the junctional and NPNAx5 epitopes reduced immunogenicity and efficacy, and increasing the repeat valency to NPNAx20 did not improve upon NPNAx5. TMV was confirmed as a versatile vaccine platform for displaying small epitopes defined by neutralizing mAbs. We show that polyclonal antibodies against engineered VLPs can recapitulate the binding specificity of the mAbs and immune-focusing by reducing the structural complexity of an epitope may be superior to immune-broadening as a vaccine design approach. Most importantly the junctional and restricted valency NPNA epitopes can be the basis for developing highly effective second-generation malaria vaccine candidates.

Highlights

Malaria is a disease of significant public health importance and according to the WHO 2020 World Malaria Report, 409,000 deaths globally were attributed to malaria in 2019
Sporozoites are covered with the Circumsporozoite protein (CSP) which has a N-terminal ‘domain’ encompassing Region I (RI), that includes an inter-species conserved 5 amino-acid motif (KLKQP), followed by a junctional sequence NP-DPNA-NPNV-DPNA, 25-42 copies of the major NPNA repeats, interspersed minor NPNV-DPNA repeat[3,4,5], and a C-terminal region consisting of an alpha thrombospondin type-I repeat domain (Supplementary Fig. 1)
Our data support the continued development of CSP vaccines based on Tobacco Mosaic Virus (TMV) VLPs and we show that epitope focusing by restricting (NPNA)n valency or by displaying the junctional epitope can elicit potent protective antibodies

Summary

Introduction

Malaria is a disease of significant public health importance and according to the WHO 2020 World Malaria Report, 409,000 deaths globally were attributed to malaria in 2019. Of the five species of Plasmodium that can infect humans, P. falciparum (Pf) is the primary cause of malaria-associated mortality, with most deaths occurring in children under 5 years of age. Sporozoites are covered with the Circumsporozoite protein (CSP) which has a N-terminal ‘domain’ encompassing Region I (RI), that includes an inter-species conserved 5 amino-acid motif (KLKQP), followed by a junctional sequence NP-DPNA-NPNV-DPNA, 25-42 copies of the major NPNA repeats, interspersed minor NPNV-DPNA repeat[3,4,5], and a C-terminal region consisting of an alpha thrombospondin type-I repeat domain (Supplementary Fig. 1). P. falciparum strains show a high degree of conservation in the N-terminal and the repeat regions but the C-terminal region of CSP contains polymorphic residues surrounding a hydrophobic pocket[6]. CSP is the target of the most advanced malaria vaccine candidate, RTS,S/AS01

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Conclusion