Francis M. Ndungu

Plasmodium falciparum malaria causes an estimated 500 million cases per year and hundreds of thousands of deaths annually. Substantial morbidity and mortality occurs in Africa. An effective vaccine is needed, and vaccine development is essential to provide an effective sustainable and cost-effective control of disease. Rational vaccine development would be based on clearly described mechanisms that lead to clearance of parasites and the correlates of immunity. The current lead vaccine, based on the first falciparum gene ever cloned, delivers ~30% protection over 4 years. An understanding and identification of the underlying mechanisms that lead to naturally acquired immunity has the potential to inform rational vaccine design. We need to identify the antigenic targets for protective immune responses

and the threshold required and functional properties. The study of naturally acquired immunity has been conducted using observational field studies to date, based on surveillance of naturally exposed children. However, adults living in endemic areas acquire resistance to infection and/or disease with repeated exposure to malaria causing parasites. To understand how this immunity impacts infection, we recently conducted a deliberate infection study using parasite stages that are infective from the mosquito, sporozoites. Out of a total of 142 volunteers, 33 were observed to be malaria parasite free and remained uninfected throughout monitoring for infection in the study. These studies in non-endemic volunteers, with 100% developing infection, have recently been used to investigate pathophysiological mechanisms of disease utilising the stage of the parasite responsible for disease, blood stages i.e. deliberate infection with parasite infected red blood cells. We now propose to deliberately infect volunteers who we found to be previously malaria-free with parasite-infected red blood cells that directly results in the signs and symptoms associated with disease. We intend to evaluate pathophysiological mechanisms of disease and determine the contribution and mechanisms of immunity resulting from the disease stage of infection. We will adopt a biomarker and systems immunology approach to identify mechanisms associated with disease outcome including detailed characterisation of biomarkers of immune activation including endothelial activation and microvascular function

typing of antibody specificities and function, analysis of cellular immunity

and analysis of the metabolome. This study will define mechanisms associated with disease in the context of pre-existing immunity and help inform targets for the next generation blood-stage vaccines. Technical Summary Plasmodium falciparum malaria causes substantial morbidity and mortality in Africa. A high-efficacy vaccine is essential. The current lead malaria vaccine confers ~30% protection over 4 years and is based on the major antigen on the surface of pre-erythrocytic sporozoite parasite stages. An attractive strategy to build on this partial efficacy is a multi-stage vaccine including immunity against blood-stages of the parasites. However, there is no analogous single major antigen for blood stage parasites, and vaccines based on several different blood stage antigens have not been successful. Adults in endemic areas acquire immunity to blood-stage parasites after repeated malaria exposure. Identifying the underlying mechanisms of naturally acquired immunity has the potential to inform rational vaccine design e.g. antigen selection. We recently completed controlled human malaria infection (CHMI) studies in Kenyan adults using cryopreserved sporozoites (PfSPZ) and identified 33 out of 142 volunteers who remained uninfected by sensitive blood PCR for malaria parasites. The PCR negative volunteers are likely to have high levels of acquired immunity, and imply an excellent model for vaccine development. However the protection may have resulted either from immunity to pre-erythrocytic or to blood-stage parasites. The distinction is critical for further study and vaccine design. We now propose a blood-stage challenge experiment for these previously protected volunteers since this will bypass pre-erythrocytic stages to determine the contribution of blood-stage immunity and mechanisms. We aim to: (a) test whether a key component of malaria immunity is through an immune response to blood-stage infection

and (b) test whether individuals who are especially immune are able to control higher inoculums of parasites

and (c) define immunological mechanisms associated protection and/or disease pathogenesis to inform the development of the next generation of blood-stage malaria vaccines.