Immunoprofiling Identifies Functional B and T Cell Subsets Induced by an Attenuated Whole Parasite Malaria Vaccine as Correlates of Sterile Immunity

Immune correlates of protection remain elusive for most vaccines. An identified immune correlate would accelerate the down-selection of vaccine formulations by reducing the need for human pathogen challenge studies that are currently required to determine vaccine efficacy. Immunization via mosquito-delivered, radiation-attenuated P. falciparum sporozoites (IMRAS) is a well-established model for efficacious malaria vaccines, inducing greater than 90% sterile immunity. The current immunoprofiling study utilized samples from a clinical trial in which vaccine dosing was adjusted to achieve only 50% protection, thus enabling a comparison between protective and non-protective immune signatures. In-depth immunoprofiling was conducted by assessing a wide range of antigen-specific serological and cellular parameters and applying our newly developed computational tools, including machine learning. The computational component of the study pinpointed previously un-identified cellular T cell subsets (namely, TNFα-secreting CD8+CXCR3−CCR6− T cells, IFNγ-secreting CD8+CCR6+ T cells and TNFα/FNγ-secreting CD4+CXCR3−CCR6− T cells) and B cell subsets (i.e., CD19+CD24hiCD38hiCD69+ transitional B cells) as important factors predictive of protection (92% accuracy). Our study emphasizes the need for in-depth immunoprofiling and subsequent data integration with computational tools to identify immune correlates of protection. The described process of computational data analysis is applicable to other disease and vaccine models.

Merozoite Proteins Discovered by qRT-PCR-Based Transcriptome Screening of Plasmodium falciparum

The development of malaria vaccines and medicines depends on the discovery of novel malaria protein targets, but the functions of more than 40% of P. falciparum genes remain unknown. Asexual parasites are the critical stage that leads to serious clinical symptoms and that can be modulated by malaria treatments and vaccines. To identify critical genes involved in the development of Plasmodium parasites within erythrocytes, the expression profile of more than 5,000 genes distributed across the 14 chromosomes of the PF3D7 strain during its six critical developmental stages (merozoite, early-ring, late-ring, early trophozoite, late-trophozoite, and middle-schizont) was evaluated. Hence, a qRT-PCR-based transcriptome of the erythrocytic developmental process of P. falciparum was revealed. Weighted gene coexpression network analyses revealed that a large number of genes are upregulated during the merozoite release process. Further gene ontology analysis revealed that a cluster of genes is associated with merozoite and may be apical complex components. Among these genes, 135 were comprised within chromosome 14, and 80% of them were previously unknown in functions. Western blot and immunofluorescence assays using newly developed corresponding antibodies showed that some of these newly discovered proteins are highly expressed in merozoites. Further invasion inhibition assays revealed that specific antibodies against several novel merozoite proteins can interfere with parasite invasion. Taken together, our study provides a developmental transcriptome of the asexual parasites of P. falciparum and identifies a group of previously unknown merozoite proteins that may play important roles in the process of merozoite invasion.

Reproducibility of malaria sporozoite challenge model in humans for evaluating efficacy of vaccines and drugs: a systematic review

Background The development of novel malaria vaccines and antimalarial drugs is limited partly by emerging challenges to conduct field trials in malaria endemic areas, including unknown effects of existing immunity and a reported fall in malaria incidence. As a result, Controlled Human Malaria Infection (CHMI) has become an important approach for accelerated development of malarial vaccines and drugs. We conducted a systematic review of the literature to establish aggregate evidence on the reproducibility of a malaria sporozoite challenge model. Methods A systematic review of research articles published between 1990 and 2018 on efficacy testing of malaria vaccines and drugs using sporozoite challenge and sporozoite infectivity studies was conducted using Pubmed, Scopus, Embase and Cochrane Library, ClinicalTrials.gov and Trialtrove. The inclusion criteria were randomized and non-randomized, controlled or open-label trials using P. falciparum or P. vivax sporozoite challenges. The data were extracted from articles using standardized data extraction forms and descriptive analysis was performed for evidence synthesis. The endpoints considered were infectivity, prepatent period, parasitemia and safety of sporozoite challenge. Results Seventy CHMI trials conducted with a total of 2329 adult healthy volunteers were used for analysis. CHMI was induced by bites of mosquitoes infected with P. falciparum or P. vivax in 52 trials and by direct venous inoculation of P. falciparum sporozoites (PfSPZ challenge) in 18 trials. Inoculation with P. falciparum-infected mosquitoes produced 100% infectivity in 40 studies and the mean/median prepatent period assessed by thick blood smear (TBS) microscopy was ≤ 12 days in 24 studies. On the other hand, out of 12 infectivity studies conducted using PfSPZ challenge, 100% infection rate was reproduced in 9 studies with a mean or median prepatent period of 11 to 15.3 days as assessed by TBS and 6.8 to 12.6 days by PCR. The safety profile of P. falciparum and P.vivax CHMI was characterized by consistent features of malaria infection. Conclusion There is ample evidence on consistency of P. falciparum CHMI models in terms of infectivity and safety endpoints, which supports applicability of CHMI in vaccine and drug development. PfSPZ challenge appears more feasible for African trials based on current evidence of safety and efficacy.

Identification of Novel Malaria Transmission-Blocking Vaccine Candidates

Control measures have significantly reduced malaria morbidity and mortality in the last two decades; however, the downward trends have stalled and have become complicated by the emergence of COVID-19. Significant efforts have been made to develop malaria vaccines, but currently only the RTS,S/AS01 vaccine against Plasmodium falciparum has been recommended by the WHO, for widespread use among children in sub-Saharan Africa. The efficacy of RTS,S/AS01 is modest, and therefore the development of more efficacious vaccines is still needed. In addition, the development of transmission-blocking vaccines (TBVs) to reduce the parasite transmission from humans to mosquitoes is required toward the goal of malaria elimination. Few TBVs have reached clinical development, and challenges include low immunogenicity or high reactogenicity in humans. Therefore, novel approaches to accelerate TBV research and development are urgently needed, especially novel TBV candidate discovery. In this mini review we summarize the progress in TBV research and development, novel TBV candidate discovery, and discuss how to accelerate novel TBV candidate discovery.

Epitope-coated polymer particles elicit neutralising antibodies against Plasmodium falciparum sporozoites

The current Malaria RTS,S vaccine is based on virus-like particles (VLPs) comprising the NANP repetitive epitopes from the cicumsporozoite protein (CSP) of Plasmodium falciparum. This vaccine has limited efficacy, only preventing severe disease in about 30% of vaccinated individuals. A more efficacious vaccine is urgently needed to combat malaria. Here we developed a particulate malaria vaccine based on the same CSP epitopes but using biopolymer particles (BPs) as an antigen carrier system. Specific B- and T-cell epitope-coated BPs were assembled in vivo inside an engineered endotoxin-free mutant of Escherichia coli. A high-yield production process leading to ~27% BP vaccine weight over biomass was established. The epitope-coated BPs were purified and their composition, i.e., the polymer core and epitope identity, was confirmed. Epitope-coated BPs were used alongside soluble peptide epitopes and empty BPs to vaccinate sheep. Epitope-coated BPs showed enhanced immunogenicity by inducing anti-NANP antibody titre of EC50 > 150,000 that were at least 20 times higher than induced by the soluble peptides. We concluded that the additional T-cell epitope was not required as it did not enhance immunogenicity when compared with the B-cell epitope-coated BPs. Antibodies specifically bound to the surface of Plasmodium falciparum sporozoites and efficiently inhibited sporozoite motility and traversal of human hepatocytes. This study demonstrated the utility of biologically self-assembled epitope-coated BPs as an epitope carrier for inclusion in next-generation malaria vaccines.

Molecular interactions between parasite and mosquito during midgut invasion as targets to block malaria transmission

Despite considerable effort, malaria remains a major public health burden. Malaria is caused by five Plasmodium species and is transmitted to humans via the female Anopheles mosquito. The development of malaria vaccines against the liver and blood stages has been challenging. Therefore, malaria elimination strategies advocate integrated measures, including transmission-blocking approaches. Designing an effective transmission-blocking strategy relies on a sophisticated understanding of the molecular mechanisms governing the interactions between the mosquito midgut molecules and the malaria parasite. Here we review recent advances in the biology of malaria transmission, focusing on molecular interactions between Plasmodium and Anopheles mosquito midgut proteins. We provide an overview of parasite and mosquito proteins that are either targets for drugs currently in clinical trials or candidates of promising transmission-blocking vaccines.

Insights from modelling malaria vaccines for policy decisions: the focus on RTS,S

Mathematical models are increasingly used to inform decisions throughout product development pathways from pre-clinical studies to country implementation of novel health interventions. This review illustrates the utility of simulation approaches by reviewing the literature on malaria vaccine modelling, with a focus on its link to the development of policy guidance for the first licensed product, RTS,S/AS01. The main contributions of modelling studies have been in inferring the mechanism of action and efficacy profile of RTS,S; to predicting the public health impact; and economic modelling mainly comprising cost-effectiveness analysis. The value of both product-specific and generic modelling of vaccines is highlighted.

A vaccine targeting the L9 epitope of the malaria circumsporozoite protein confers protection from blood-stage infection in a mouse challenge model

Pre-erythrocytic malaria vaccines that induce high-titer, durable antibody responses can potentially provide protection from infection. Here, we engineered a virus-like particle (VLP)-based vaccine targeting a recently described vulnerable epitope at the N-terminus of the central repeat region of the Plasmodium falciparum circumsporozoite protein (CSP) that is recognized by the potently inhibitory monoclonal antibody L9 and show that immunization with L9 VLPs induces strong antibody responses that provide protection from blood-stage malaria in a mouse infection model.

A mosquito AgTRIO monoclonal antibody reduces early Plasmodium infection of mice

Malaria begins when an infected mosquito injects saliva containing Plasmodium sporozoites into the skin of a vertebrate host. Passive immunization of mice with mosquito AgTRIO antisera offers significant protection against Plasmodium infection of mice. Furthermore, passive transfer of both AgTRIO antisera and an anti-circumsporozoite protein monoclonal antibody provides synergistic protection. In this study, we generated monoclonal antibodies against AgTRIO to delineate the regions of AgTRIO associated with protective immunity. Monoclonal antibody 13F-1 markedly reduced Plasmodium infection in mice and recognized a region, VDDLMAKFN, in the carboxyl terminus of AgTRIO. 13F-1 is an IgG2a isotype monoclonal antibody and the Fc region is required for protection. These data will aid in the generation of future malaria vaccines that may include both pathogen and vector antigens.