Malaria, particularly in its severe forms, remains a global health and economic burden. Researchers at EMBL Barcelona, the University of Texas, the University of Copenhagen, and the Scripps Research Institute have identified two broadly reactive human monoclonal antibodies (mAbs) that can recognize and target some of the proteins that cause severe malaria. In vitro tests using organ-on-a-chip technology showed that the antibodies prevented parasite-infected blood cells from sticking to blood vessels. The investigators suggest that their breakthrough could pave the way for future vaccines or anti-malaria treatments.
“This study opens the door to targeting new ways of protecting people from severe malaria, like a vaccine or other treatments,” said Maria Bernabeu, PhD, group leader at EMBL Barcelona. “This is thanks to the international and interdisciplinary collaboration that is key to understanding diseases like malaria. Our collaborators are from across the globe, studying malaria from different angles. We have to continue working together to tackle big challenges like this one.”
Bernabeu is co-senior author of the team’s published report in Nature, titled “Broadly inhibitory antibodies to severe malaria virulence proteins,” in which they concluded, “These broadly reactive antibodies are likely to represent a common mechanism of acquired immunity to severe malaria and offer novel insights for the design of a vaccine or treatment targeting severe malaria.”
Severe malaria caused by the parasite Plasmodium falciparum results in about 600,000 deaths every year, mostly among young children living in sub-Saharan Africa, the authors wrote. “Ten times as many suffer from severe disease, often with long-lasting health and socioeconomic consequences.”
The parasite infects and modifies red blood cells, which stick to the walls of the tiny blood vessels in the brain. This accumulation results in impaired blood flow and blood vessel blockage, which causes brain swelling and can develop into cerebral malaria. “In severe cases this may lead to organ failure and death,” the team continued.
The blockage of blood flow is primarily driven by a family of about 60 virulent proteins, called PfEMP1s, present on the surface of infected red blood cells. “Parasite-infected erythrocytes bind to endothelial cell receptors on the microvasculature via the polymorphic multi-domain P. falciparum erythrocyte membrane proteins, PfEMP1s, expressed on their cell surface,” the team further explained. Some types of PfEMP1 proteins can attach to another human protein called EPCR on the surface of cells lining blood vessels. This interaction damages blood vessels and is closely linked to the development of life-threatening complications. “Severe malaria is caused by parasites binding to human EPCR through the subset of PfEMP1s that has cysteine-rich interdomain region α1 (CIDRα1) domains,” the team continued.
When children in Africa grow older, they progressively develop immunity, and teenagers and adults rarely suffer from lethal disease complications. This protection was thought to be mediated by antibodies that target PfEMP1. “PfEMP1 proteins are major targets of the humoral immune response to P. falciparum infection, and antibody reactivity to the CIDRα1 domain family correlates with protection from severe malaria,” the investigators explained.
PfEMP1 is a highly variable protein and has long been considered a technically difficult vaccine target. A long-standing question therefore has been whether the immune system can generate antibodies that can target the wide variety of PfEMP1 types in circulation. The team noted, “The identification of the interaction of PfEMP1 with EPCR as the common trait of parasites causing severe malaria has reinforced the notion that individuals exposed to P. falciparum may develop broadly reactive antibodies that are capable of inhibiting this central driver of severe malaria pathogenesis.”
Through their newly reported work the investigators isolated, from two individuals, two broadly reactive and inhibitory monoclonal antibodies to CIDRα1, designated C7 and C74. “We were hesitant whether we could identify a single antibody that could recognize them all,” acknowledged Bernabeu. “And it turned out that our improved immunological screening methods developed at the University of Texas quickly identified two examples of human antibodies broadly effective against different versions of the PfEMP1 protein.” The authors also reported, “The antibodies isolated from two different individuals exhibited similar and consistent EPCR-binding inhibition of diverse CIDRα1 domains, representing five of the six subclasses of CIDRα1.” Bernabeu added, “They both targeted a part of the protein known as CIDRα1 which interacts with the EPCR receptor.”
The team needed to test if the antibodies could successfully block EPCR binding in living blood vessels. While animal models for in vivo testing are available for many diseases, for malaria this is not possible because the virulent proteins of the parasites that infect mice are very different from those of their human counterparts.
The researchers came up with an innovative approach to overcome this challenge. They developed a way to grow a network of human blood vessels in the laboratory and to pass human blood infected with live parasites through the vessels, thereby reconstructing the disease in a dish. These experiments demonstrated that the antibodies were able to prevent the infected cells from accumulating, suggesting that they might help stop the blockage that leads to severe malaria symptoms.
“We used our organ-on-a-chip technology to recreate brain microvessels in 3D, which we then infected with malaria parasites,” said co-first author Viola Introini, PhD, the Marie Skłodowska-Curie postdoctoral fellow in Bernabeu’s Group at EMBL Barcelona. “We introduced the two antibodies into the vasculature and were impressed at how well they prevented infected blood cells from sticking to the vessels. It was striking to see inhibition readily apparent by eye.” The authors added, “Both antibodies inhibited EPCR binding of both recombinant full-length and native PfEMP1 proteins, as well as parasite sequestration in bioengineered 3D human brain microvessels under physiologically relevant flow conditions.”
Structural and immunology analysis by collaborators at the University of Copenhagen and the Scripps Research Institute revealed that these antibodies prevent parasite binding by a similar mechanism—recognizing three highly conserved amino acids on CIDRα1. The authors say their results provide evidence that humans frequently exposed to P. falciparum develop broadly reactive and inhibitory mAbs to severe malaria-associated CIDRα1 PfEMP1 proteins.
“The functional and molecular characterization of the mAbs C7 and C74 presented in this study not only provides conclusive evidence for the presence of broadly inhibitory antibodies to EPCR-binding PfEMP1 but also unveils that such antibodies are likely to share a uniform mode of binding,” they wrote. Bernabeu also noted, “At EMBL Barcelona, we believe that tissue engineering and growing organs-on-a-chip allow us to study diseases with much more complexity and detail, as well as provide useful platforms for screening vaccine candidates.”
