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Identification of Plasmodium falciparum proteoforms from liver stage models.

ABSTRACT: BACKGROUND:Immunization with attenuated malaria sporozoites protects humans from experimental malaria challenge by mosquito bite. Protection in humans is strongly correlated with the production of T cells targeting a heterogeneous population of pre-erythrocyte antigen proteoforms, including liver stage antigens. Currently, few T cell epitopes derived from Plasmodium falciparum, the major aetiologic agent of malaria in humans are known. METHODS:In this study both in vitro and in vivo malaria liver stage models were used to sequence host and pathogen proteoforms. Proteoforms from these diverse models were subjected to mild acid elution (of soluble forms), multi-dimensional fractionation, tandem mass spectrometry, and top-down bioinformatics analysis to identify proteoforms in their intact state. RESULTS:These results identify a group of host and malaria liver stage proteoforms that meet a 5% false discovery rate threshold. CONCLUSIONS:This work provides proof-of-concept for the validity of this mass spectrometry/bioinformatic approach for future studies seeking to reveal malaria liver stage antigens towards vaccine development.

SUBMITTER: Winer B

PROVIDER: S-EPMC6947969 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Identification of Plasmodium falciparum proteoforms from liver stage models.

Winer Benjamin B Edgel Kimberly A KA Zou Xiaoyan X Sellau Julie J Hadiwidjojo Sri S Garver Lindsey S LS McDonough Christin E CE Kelleher Neil L NL Thomas Paul M PM Villasante Eileen E Ploss Alexander A Gerbasi Vincent R VR

Malaria journal 20200107 1

<h4>Background</h4>Immunization with attenuated malaria sporozoites protects humans from experimental malaria challenge by mosquito bite. Protection in humans is strongly correlated with the production of T cells targeting a heterogeneous population of pre-erythrocyte antigen proteoforms, including liver stage antigens. Currently, few T cell epitopes derived from Plasmodium falciparum, the major aetiologic agent of malaria in humans are known.<h4>Methods</h4>In this study both in vitro and in vi ...[more]

PMID: 31910830

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Project description:Immunization with radiation-attenuated sporozoites (RAS) has been shown to protect against malaria infection, primarily through CD8 T cell responses, but protection is limited based on parasite strain. Therefore, while CD8 T cells are an ideal effector population target for liver stage malaria vaccine development strategies, such strategies must incorporate conserved epitopes that cover a large range of class I human leukocyte antigen (HLA) supertypes to elicit cross-strain immunity across the target population. This approach requires identifying and characterizing a wide range of CD8 T cell epitopes for incorporation into a vaccine such that coverage across a large range of class I HLA alleles is attained. Accordingly, we devised an experimental framework to identify CD8 T cell epitopes from novel and minimally characterized antigens found at the pre-erythrocytic stage of parasite development. Through in silico analysis we selected conserved P. falciparum proteins, using P. vivax orthologues to establish stringent conservation parameters, predicted to have a high number of T cell epitopes across a set of six class I HLA alleles representative of major supertypes. Using the decision framework, five proteins were selected based on the density and number of predicted epitopes. Selected epitopes were synthesized as peptides and evaluated for binding to the class I HLA alleles in vitro to verify in silico binding predictions, and subsequently for stimulation of human T cells using the Modular IMmune In-vitro Construct (MIMIC®) technology to verify immunogenicity. By combining the in silico tools with the ex vivo high throughput MIMIC platform, we identified 15 novel CD8 T cell epitopes capable of stimulating an immune response in alleles across the class I HLA panel. We recommend these epitopes should be evaluated in appropriate in vivo humanized immune system models to determine their protective efficacy for potential inclusion in future vaccines.

Project description:Background: Primaquine (PQ) is the only currently licensed antimalarial that prevents Plasmodium vivax (Pv) relapses. It also clears mature P. falciparum (Pf) gametocytes, thereby reducing post-treatment transmission. Randomized PQ treatment in a treatment-to-reinfection cohort in Papua New Guinean children permitted the study of Pv and Pf gametocyte carriage after radical cure and to investigate the contribution of Pv relapses.Methods: Children received radical cure with Chloroquine, Artemether-Lumefantrine plus either PQ or placebo. Blood samples were subsequently collected in 2-to 4-weekly intervals over 8 months. Gametocytes were detected by quantitative reverse transcription-PCR targeting pvs25 and pfs25.Results: PQ treatment reduced the incidence of Pv gametocytes by 73%, which was comparable to the effect of PQ on incidence of blood-stage infections. 92% of Pv and 79% of Pf gametocyte-positive infections were asymptomatic. Pv and to a lesser extent Pf gametocyte positivity and density were associated with high blood-stage parasite densities. Multivariate analysis revealed that the odds of gametocytes were significantly reduced in mixed-species infections compared to single-species infections for both species (ORPv = 0.39 [95% CI 0.25-0.62], ORPf = 0.33 [95% CI 0.18-0.60], p<0.001). No difference between the PQ and placebo treatment arms was observed in density of Pv gametocytes or in the proportion of Pv infections that carried gametocytes. First infections after blood-stage and placebo treatment, likely caused by a relapsing hypnozoite, were equally likely to carry gametocytes than first infections after PQ treatment, likely caused by an infective mosquito bite.Conclusion: Pv relapses and new infections are associated with similar levels of gametocytaemia. Relapses thus contribute considerably to the Pv reservoir highlighting the importance of effective anti-hypnozoite treatment for efficient control of Pv.Trial registration: ClinicalTrials.gov NCT02143934.

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Identification of Plasmodium falciparum proteoforms from liver stage models.

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Identification of Plasmodium falciparum proteoforms from liver stage models.

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