Targeting the IL-33-inflammasome axis in therapy for cerebral malaria

Malaria continues to be a major cause of morbidity and mortality in many regions of the world. Cerebral malaria (HCM) is the most severe manifestation of malaria infection, being responsible for the death of approximately 300,000 people, mainly young children in Africa, each year. Currently anti-malarial drugs are the only available treatment for HCM but by itself such treatment is unsuccessful in 15-25% of cases. Indeed, the majority of individuals that die from HCM are given anti-malarial drugs but the treatment fails to prevent death. Moreover, in the individuals that survive HCM, many exhibit long-lasting neurological deficits that impact on their quality of life creating a significant socio-economic burden in some of the poorest countries in the world. Thus, there is an urgent need to develop better treatments for HCM.

There is some evidence that the level of inflammation within the brain (neuroinflammation), may contribute to the sub-optimal recovery from HCM. However, as we are only able to obtain brains from individuals that died of HCM, we are unable to study solely in human brains the processes that are responsible for development of HCM nor can we identify the ways through which treatments improve recovery from HCM. Thus, animal models of HCM are required to identify and test new treatments for HCM. The murine experimental cerebral malaria (ECM) model is currently the most frequently utilised model to study HCM and, relevantly, we have recently shown that the nature of brain pathology in HCM and ECM are very similar, validating the model for development of new treatments for HCM. Using this model, we have found that administration of the anti-inflammatory molecule interleukin-IL-33 (IL-33), which is garnering interest as a critical controller of brain health, significantly improves the effectiveness of anti-malarial drug treatment of mice with established ECM. Importantly, we have found that IL-33 appears to be protective through ameliorating a specific type of neuroinflammation that is promoted by an immune complex called the inflammasome. Consequently, the overall aim of this project is to define the way in which rIL-33 and inhibitors of the inflammasome improve the recovery from HCM.

To address this aim, in objective 1 we will utilise the ECM model to perform a series of analyses of how IL-33 and inhibitors of the inflammasome improve the success of anti-malarial drug treatment of ECM. Specifically, we will examine how the treatments reduce the level of cerebral pathology and cerebral inflammation within the brain post-anti-malarial drug chemotherapy of the syndrome, improving survival and reducing the severity of neurological deficits. Because parasite-adhesion to cerebral endothelial cells is an important event in HCM, in objective 2 we will complement the analyses in the ECM model with studies in a parasite-human endothelial cell co-culture system. Using this system we will examine how IL-33 and inflammasome inhibitors improve the health of human brain endothelial cells exposed to malaria parasites and anti-malarial drugs. Finally, in objective 3 of the study we will obtain and perform studies in plasma from individuals with HCM, individuals with malaria but who did not have HCM, and from individuals that were not infected with malaria. We will also obtain brain samples from individuals that died of HCM and from individuals with other causes of death. By analysing these samples we will assess the relationship between systemic and brain-specific IL-33 levels, inflammasome activation, and the development of HCM and the outcome of anti-malarial drug chemotherapy of HCM. Overall, the successful completion of this project will demonstrate how therapy with IL-33 and inflammasome inhibitors can improve the treatment and recovery from CM. This will prioritise the development of clinical trials to assess the effectiveness of IL-33 and inflammasome inhibitors as treatments for CM.

Cerebral malaria (HCM) is the most severe complication of malaria infection, being responsible for approximately 300,000 deaths each year. The majority of individuals that die from HCM are treated with anti-malarial drugs, which is the only current treatment for the condition, but which is ineffective in 15-25% of cases. Thus, there is an urgent need to develop better treatments for CM to reduce the burden of this devastating syndrome. Our preliminary data strongly suggest that rIL-33 is a novel and highly effective potential adjunct immunotherapy for treatment of CM through its capacity to inhibit NLRP3 inflammasome-mediated neuroinflammation. Indeed, adjunct treatment with the inflammasome inhibitor MCC950 phenocopies the protective effects of rIL-33 in improving the recovery of mice with CM. These data, therefore, underline the central importance of the IL-33-inflammasome axis in determining the treatment success of CM. In this application we propose to definitively define, using a validated murine model of CM and an established in vitro human brain tripartite culture system mimicking parasite sequestration and monocyte-mediated parasite killing within the brain microvasculature, the mechanism through which adjunctive administration of rIL-33 and inflammasome inhibitors improve the recovery from established CM. Furthermore, we will perform highly resolved and quantitative analyses of IL-33 expression and inflammasome activity within plasma and post-mortem brain samples from individuals with CM and in case controls samples from individuals with asymptomatic malaria or non-CM causes of death. These investigations will reveal the relationship between impaired IL-33 function, dysregulated inflammasome activation, and neuroinflammation in determining the development and resultant treatment failure of CM. We anticipate that our results will support the design of clinical trials to study the adjunct activity of rIL-33 and inflammasome inhibitors in treatment of CM.

Malaria still kills 438,000 people every year and causes severe morbidity in up to 214 million people per year across the tropical world. In endemic countries, malaria places a major constraint on education, economic growth and social development. The majority of malarial deaths, and a large proportion of malaria-associated long term morbidity, is due to cerebral malaria, which is one of the major severe disease complications of Plasmodium falciparum infection. Even with anti-malarial drug chemotherapy, CM is fatal in 15-25% of cases.

There is no specific treatment for CM because we do not fully understand the pathogenesis of the syndrome and we have very limited insight into why anti-malarial drug chemotherapy works in some individuals but is ineffective in other individuals. The inability to examine the brains of CM sufferers prior to and at defined time points post-drug treatment, or to contrast the pathology in brains of individuals that survive or die from CM post-drug treatment, means it is impossible to obtain this information solely through the study of human CM. Thus, first and foremost, we need to develop better treatments for CM. Secondly, to develop and test new treatments, we need to establish more effective translational programmes of work using all valuable models and tools available for study of HCM.

In this project we propose to utilise the ECM model, the pathology of which we have extensively characterised, in a direct, integrated, programme of work alongside investigations in HCM to interrogate the utility of IL-33 and inflammasome inhibitors as therapies for malaria-induced cerebral pathology. The successful completion of this project will therefore: 1. Establish a translatable programme of work for the study of HCM; 2. significantly enhance our understanding of the determinants that specify the treatment success and failure of CM and; 3. Develop IL-33 and inflammasome inhibitors as new potential immunotherapeutic adjunctive treatments for HCM. Collectively, these studies will benefit researchers in academia and in the private sector.