The Problem

Severe malaria causes a million deaths each year, out of the 500 million total malaria infections that occur each year. While this percentage is small, deaths from severe malaria nonetheless represent a serious health challenge. There are two difficulties:

1. We currently have no effective way to identify individuals who will develop life-threatening malaria. This prevents effective triage and management of severe malaria and results in the misallocation of scarce health resources.
2. There is a paucity of effective therapies for those who develop life-threatening illness. Like other serious infectious diseases, severe malaria is associated with high fatality rates despite the use of potent antimicrobial agents and optimal supportive care. This is because the body’s own immune response to the infection may cause organ injury and provoke a severe and sometimes fatal outcome – and anti-malarial treatment does not affect this response.

How are we tackling the problem?

The capacity to rapidly identify individuals at risk of critical illness and the ability to deliver novel interventions to improve survival would represent a transformative advance in the management of life-threatening infectious diseases. Therefore, our work is focused on finding biomarkers of severe malaria that are effective in identifying individuals at risk of this life-threatening illness and on devising practical therapies that modify the host response to improve health outcomes in conjunction with anti-malarial drugs. Our goal is to translate our findings into clinically relevant solutions that can be delivered in resource poor settings.

Our approach to exploring these areas involves: (i) the identification of candidate proteins and pathways that may play a role in progression to severe malaria; (ii) confirmation of their role in development of critical illness; and (iii) intervention with novel therapies to prevent severe disease. This approach embraces several models of severe disease development:

1. The endothelium plays a central role in severe malaria. The endothelium is the largest interconnected organ in the human body, linking all vital organs. A common pathway of injury induced by multiple life-threatening conditions, endothelial activation and loss of integrity directly contributes to critical illness, multi-organ failure and death, and therefore represents a highly attractive target for the identification of new markers of disease severity and novel interventions to improve survival.
2. The innate immune response dictates malarial disease severity. The innate immune system is the first-line of defense against pathogens. It consists of reactions that require no prior exposure to the pathogen, and thus represents the main response of those who have limited immunity to malaria and are at highest risk of progressing to severe disease (children, pregnant women, and travelers, for instance). Promoting beneficial responses and blocking harmful ones are novel ways to therapeutically intervene in life-threatening malarial infections.
3. Genetic adaptations insights lead to novel intervention strategies. Genetic variations (polymorphisms) that provide protection against severe malarial disease have evolved in humans as a result of selective pressure by malaria. Teasing apart how these polymorphisms prevent life-threatening illness provides important insight into severe disease development and suggests innovative new strategies to intervene and improve outcome.

I arrived in the lab with a goal to apply my immunology and genetics background to an issue specific to women’s health. An estimated 125 million women are pregnant in areas of malaria transmission each year. These women are among the most susceptible to detrimental outcomes with malaria infections including maternal death and anemia, spontaneous abortions, preterm delivery and poor fetal growth. Due to ethical and technical issues surrounding studies of pregnant women, little is known about this manifestation of malarial infection. Building off the finding of a labmate that excessive systemic complement activation is detected in women with placental malaria, I set out to determine if this contributes to the poor maternal and fetal effects associated with placental malaria. Harnessing a newly identified placental malaria mouse model, genetically modified mouse strains and reagents that block complement activation, I have been able to show that the C5a-C5aR pathway does contribute to malaria-induced fetal growth restriction and spontaneous miscarriage. Identification of such pathways is a critical step to providing a rational intervention strategy. ~Karlee Silver, Postdoctoral Fellow 

What have we accomplished?

The endothelium plays a central role in development of severe malaria

We have shown that angiopoietins – proteins that directly determine the activation state of the endothelium – are excellent biomarkers of severe malaria. In a healthy individual, angiopoietin (Ang)-1 is always binding the shared receptor, Tie-2, to maintain endothelial integrity and prevent inflammation. Ang-2 can displace Ang-1 and sensitize the endothelium to become responsive to low levels of pro-inflammatory cytokines such as TNF, resulting in the up-regulation of adhesion molecules in the brain endothelium to which parasitized erythrocytes bind. We have shown a clinical association between an increased Ang-2/Ang-1 ratio and increased malarial disease severity in children, adults and pregnant women. Findings from these studies were covered by 14 news agencies in 6 languages. Of particular importance for translation into a clinical diagnostic tool, we have shown that these biomarkers can be measured in whole blood. More recent studies with samples of malaria infected children in Malawi and Uganda are confirming the diagnostic efficacy of Ang-1, Ang-2 and Tie-2.

In order to prevent endothelial activation via Ang-2 release in malarial infections, we have been conducting studies with inhaled nitric oxide. Nitric oxide inhibits Ang-2 release and, as a result, is a potent endothelial stabilizer. Our studies in the experimental cerebral malaria model have shown that inhaled nitric oxide alone prolongs survival. Since nitric oxide is safe for use (even in premature neonates), cheaply manufactured, easily stored and almost off patent, if it is proven effective in controlling severe malaria in conjunction with antimalarials, then this would be a major advance in malaria treatment.

The innate immune response dictates malarial disease severity

Key innate responses in malaria are clearance of infected red blood cells by immune cells and activation of the complement system, which in turn leads to inflammation and initiation of the acquired immune response. We recently discovered a role for innate immunity in both disease and protection.

We provided the first evidence showing that blocking excessive activation of complement component C5 increases survival to experimental cerebral malaria. The identification of C5a-C5aR as a potential target for intervention in severe malaria and the availability of inhibitors of C5a-C5aR, including an FDA-approved humanized monoclonal antibody against C5, now present an exciting opportunity to translate these findings into the clinical setting.

Our work also showed that promoting the protective effect of increased clearance of malaria-infected red blood cells with the diabetes drug rosiglitazone also improves outcomes in an experimental cerebral malaria model. These findings have been taken all the way to human clinical trials.

Genetic adaptations insights lead to novel intervention strategies

We have shown that a common human genetic red cell disorder (pyruvate kinase deficiency, PKD) protects against human Plasmodium falciparum infection and replication. These exciting findings indicate that mutant PK alleles may confer a protective advantage against malaria in humans similar to sickle cell and thalassemia. This work was published in the New England Journal of Medicine, accompanied by an editorial and received considerable media and scientific attention.

We have also provided a new explanation for the clinically observed protection of individuals with blood group O, also involving increased clearance of infected red blood cells. We can now exploit this knowledge to design novel therapies based on evolutionary strategies that provide natural protection and that, as a result, would not be subject to drug resistance, a problem that plagues standard antimicrobial drugs.

What are the next steps?

In the coming years we will be focused on the following priorities:

1. A blinded, controlled clinical trial of inhaled nitric oxide as an adjunctive therapy for severe malaria in Ugandan children.
2. An NIH-funded collaborative field study of pregnant women in Tanzania to validate biomarkers for placental malaria.
3. Follow-up on exciting endothelial and innate immune response candidates, including CD13 and complement component C3. Therapeutic interventions for both of these candidates are already in the pipeline based on their roles in cancer and sepsis, respectively.

Key Publications

Pyruvate kinase deficiency (PKD) is the second most common erythrocyte enzyme abnormality. In this paper, we show for the first time that PKD protects against infection and replication of P. falciparum malaria in human erythrocytes. The mechanism of protection was found to be associated with reduced invasion of erythrocytes and enhanced clearance of parasitized erythrocytes. These findings suggest that mutant pyruvate kinase alleles may confer a protective advantage against malaria similar to sickle cell and thalassemia:

Ayi K, Min-Oo G, Serghides L, Crockett M, Kirby-Allen M, Quirt I, Gros P, Kain KC. Pyruvate Kinase deficiency protects against malaria. New England Journal of Medicine 2008; 358:1805-10.

In this study, we provide the first evidence implicating excessive activation of complement in the development of cerebral malaria (CM) in the PbA model of experimental severe malaria and suggest a potential role for complement activation and C5a generation in particular, in the pathophysiology of human CM. The identification of C5a-C5aR as a potential target for intervention in severe and cerebral malaria and the availability of inhibitors of C5a-C5aR, including an FDA-approved humanized monoclonal antibody against C5, now permit the exciting opportunity to directly test this hypothesis:

Patel S, Berghout J, Lovegrove FL, Ayi K, Conroy A, Serghides L, Min-oo G, Gowda C, Sarma JV, Rittirsch D, Ward PA, Liles WC, Gros P, Kain KC. C5 deficiency and C5a or C5aR blockade protects against cerebral malaria. J Exp Med 2008; 205 1133-1143.  

In this study, we show that malarial disease severity is associated with dysregulated angiopoietin-1 and angiopoietin-2 expression in two different populations. These findings suggest that these biomarkers may provide important prognostic information regarding who will progress to severe malaria (and thus requires more intensive medical intervention). Furthermore, they suggest that therapies that prevent dysregulation of the angiopoietins could ameliorate the outcome of severe malaria:

Lovegrove FE, Tangpukdee N, Opoka R, Lafferty EI, Rajwans N, Hawkes M, Krudsood S, Looareesuwan S, John CC, Liles WC, Kain KC. Serum angiopoietin-1 and -2 levels discriminate cerebral malaria from uncomplicated malaria and predict clinical outcome in African children. /PLoS ONE /2009; 4:e491.