Introduction and Background of the Project

Introduction

Malaria caused by Plasmodium vivax is a leading tropical disease in Southeast Asia, South Asia, Mesoamerica, South America, and Oceania. The disease poses a major challenge for the global malaria eradication because P. vivax can cause recurring blood infections. These ‘relapses’ are caused by hypnozoites, the latent form of the parasite which can persists for a long time in the liver even after clearance of blood infection by anti-malarial drugs. Interventions that directly or indirectly reduce the hypnozoite reservoir in the affected communities, including new vaccines that block parasite transmission, will be highly required. Although vaccines are the most cost-effective tools to fight many infectious diseases, malaria vaccine targeting P. vivax is still not available.

Project objective

This project aims to develop a novel mRNA-based malaria transmission-blocking vaccine of P. vivax. It targets Pvs230, a sexual stage protein of the parasite, that induces potent and long-lasting transmission blocking immunity and is able to interrupt transmission of P. vivax from human to mosquito.

Project design

We will combine the expertise in malaria vaccine development from Mahidol and Ehime Universities to generate new mRNA vaccines that block transmission of P. vivax. The vaccine target is the parasite protein called Pvs230, a well-known vaccine candidate expressed during the sexual-stage development of the parasite. Since Pvs230 is a large protein, we will first screen several Pvs230 fragments to identify the subdomain that induces strongest functional immunity. Then we will employ both the classical linear nucleoside-modified mRNA and our newly developed circular mRNA to devise the best performing vaccine construct. Vaccine efficacy will be determined by the ability of the vaccine to induce antibodies that block transmission of the parasite from humans to mosquitoes. 

How can your partnership (project) address global health challenges?

Malaria continues to place a heavy toll on human health in low- and middle-income countries. The WHO’s Global Technical Strategy for Malaria (2016–2030) aims to eliminate malaria in a further 35 countries by 2030 compared to 2015. Through a single mosquito bite, P. vivax can cause recurring malaria episodes, making it difficult to control and eliminate. A vaccine that interrupts parasite transmission is expected to be an essential tool that helps eliminate the disease.

Short-term Impact: We will develop an mRNA vaccine that elicits potent transmission-blocking immunity in mice. The vaccine will be the basis for further development and evaluation in nonhuman primates and humans.

Long-term Impact: We aim to eventually combine potent transmission-blocking vaccine with a pre-erythrocytic vaccine in a multi-valent vaccine formulation to block transmission both from humans to mosquitoes and from mosquitos to humans. Such a combination vaccine is expected to quickly bring malaria transmission to cessation and thereby accelerate malaria eradication.

What sort of innovation are you bringing in your project?

To date, two transmission-blocking vaccines for P. vivax utilized recombinant Pvs25 protein were tested in human volunteers, but the results were disappointing due to the low immunogenicity and adverse events. Thus, there is a need to develop better vaccines. Nucleoside-modified mRNA formulated with lipid nanoparticle (mRNA-LNP) recently emerged as an effective vaccine platform for COVID-19. In this project, we will apply this well-established mRNA technology to help develop a new malaria transmission-blocking vaccine using Pvs230 as a target. In addition to this, we will also explore the use of circular mRNA which can potentially provide better vaccine stability, enhanced immunogenicity, and lower costs of production. 

Role and Responsibility of Each Partner

In this project, Ehime University will lead the target and antigen selection, experiment planning, recombinant protein production for immunogenicity testing, and data analysis. Mahidol University will be responsible for project administration, mRNA vaccine design and production, immune response characterization, and functional assays to evaluate the vaccine potency using field isolates.

Others (including references if necessary)

WHO Global Technical Strategy for Malaria 2016-2030.
https://www.who.int/malaria/publications/atoz/9789241564991/en/

Final Report

1. Project objective

Plasmodium vivax, the major cause of malaria morbidity outside Africa, is transmitted by mosquito bites. The parasite is widely distributed, with a high burden in the remote populations that have limited access to healthcare services. This project aims to develop a novel mRNA-based transmission-blocking vaccine targeting P. vivax. The vaccine focuses on Pvs230, a sexual-stage protein of the parasite, and is designed to induce potent and long-lasting transmission-blocking immunity, thereby interrupting the transmission of P. vivax from humans to mosquitoes. Implementation of this vaccine in endemic communities could substantially cut the spread of the parasite, contributing to malaria elimination efforts.

2. Project design

Because Pvs230 is a large protein, we first screened several Pvs230 fragments to identify a subdomain capable of inducing transmission-blocking immunity in mice. Linear nucleoside-modified mRNA vaccines were produced and formulated with lipid nanoparticles (LNP) for mouse immunization. Antibody responses were measured by ELISA using wheat germ cell-free system (WGCFS)-produced protein fragments. Vaccine efficacy was evaluated against P. vivax from Thai malaria patients by direct membrane-feeding assay (DMFA), in which mouse antibodies were added to infected blood before mosquito feeding. We also compared our newly developed circular mRNA vaccine platform to the conventional linear mRNA format.

3. Results, lessons learned

Six individual linear mRNAs encoding Pvs230 fragments spanning the full-length Pvs230 were successfully generated and formulated with LNP to produce test vaccines. Following a two-dose immunization regimen in mice, all six linear mRNA vaccines elicited high-titer antibodies that recognize the native Pvs230 protein expressed on the surface of P. vivax gametocytes - the parasite stage that develops within mosquitoes. In addition to robust humoral responses, these vaccines also induced strong cellular immune responses. Notably, antibodies induced by two of the six Pvs230 constructs completely block parasite transmission, as demonstrated by DMFA. These two promising constructs were subsequently selected for evaluation using our circular mRNA vaccine platform.

With two-dose administration, both the circular mRNA vaccines elicited strong antibody responses in mice. These antibodies recognize Pvs230 on the surface of gametocytes; however, neither circular mRNA vaccine-induced antibodies displayed detectable transmission-blocking activities by DMFA, thus excluding them for further development.

In summary, we systematically screened Pvs230-based mRNA-LNP vaccine candidates and successfully identified novel transmission-blocking vaccine constructs for P. vivax. The findings reinforce observations from our previous GHIT-funded project, confirming that the mRNA vaccine platform can be effectively employed at the discovery stage to explore and evaluate new vaccine candidates with strong translation potential for human use.