Parasite Biology and Host Interactions
The Kappe Lab studies the biology of the mosquito stages of the malaria parasite called sporozoites, which are transmitted to humans when a mosquito bites. Within humans, sporozoites infect the liver and develop as liver stages within liver cells called hepatocytes, which is another focus of the lab. During this stage, each parasite replicates many times within the confines of a single hepatocyte and ultimately leads to the formation and release of tens of thousands of new parasite forms, which infect red blood cells. During initial infection in the liver, parasite numbers are small and infection is asymptomatic, making it a vulnerable target for prevention of malaria infection. The Kappe Lab pioneered functional genomics studies of both sporozoites and liver stages and thus laid the groundwork for a systems approach to their analysis. The Lab uses rodent malaria models (Plasmodium yoelii, Plasmodium berghei) and the most lethal human malaria parasite Plasmodium falciparum. The Lab made major contributions to the field by elucidating the molecular underpinnings that regulate sporozoite infectivity for the mammalian host, factors that are critical for parasite liver infection and key parasite and host factors that are critical for liver stage growth. The Lab has also more recently contributed major findings in the area of host responses to liver stage infection. The Kappe Lab is at the cutting edge of establishing small animal models that allow the direct study of human malaria parasite liver infection.
Models for Human Malaria
Human malaria parasite liver stages are exquisitely hard to study due to the requirement for human liver cells for liver stage development. The Kappe Lab has overcome this obstacle by using a human-liver chimeric mouse known as the FRG huHep mouse. We have successfully infected these mice with human malaria parasites and have been able to follow liver stage development over time. Additionally, FRG huHep mice infused with human red blood cells support the parasite transition from liver infection to blood stage infection in vivo and thus for the first time, we are able to complete the Plasmodium falciparum life cycle in the lab without the need for primates. This breakthrough has allowed us to complete experimental genetic crosses between distinct Plasmodium falciparum strains and should dramatically aid in our understanding of Plasmodium genetics, including drug resistance phenotypes. The Lab has also utilized this system as a way to study the ability of antibodies that are elicited by whole sporozoite immunizations of humans, to prevent Plasmodium falciparum infection of the liver.
The FRG huHep mouse has also allowed us to study Plasmodium vivax liver stage development alongside the formation, persistence and relapse of the elusive hypnozoite – a dormant liver stage form that can activate many weeks, months or even years after the infectious mosquito bite, leading to a relapse of blood stage infection and disease. Malaria elimination programs will have to find better ways of killing the Plasmodium vivax hypnozoite and our work in this arena will greatly increase our understanding of hypnozoite biology and will aid in the discovery of new drugs to kill hypnozoites.
Immunology and Vaccine Development
All vaccines rely on one simple principle: give the immune system a head start advantage over the pathogen by teaching it to recognize the pathogen before an actual infection occurs. The immune system then forms a “memory” of the pathogen and can build defenses capable of eliminating it and its associated disease during a real infection. However, vaccines capable of completely protecting against a pathogen as complex as the malaria parasite have never been successfully created. The failures in developing an effective malaria vaccine largely stem from both our limited understanding of what parts of the parasite are recognized by the immune system, what elements are required of the immune system to eliminate the parasite, and from a limited set of tools to associate immune responses with protection.
The Kappe Lab pioneered the creation of genetically attenuated Plasmodium parasites (or GAPs) that arrest and die in the liver before they can progress to the blood stages infection, which cause disease and death. The arrest of these GAPs in the liver exposes them to the immune system and engenders a powerful and effective immune response that can block a new malaria infection from the moment of an infectious mosquito bite. The Lab has found that this recalled immune response can attack the parasite through sporozoite infection, the liver stages and into the blood stages. This not only prevents disease in the infected individual, but it also stops the mosquito-human-mosquito transmission cycle and could greatly accelerate the elimination of malaria from the planet.
Previous malaria vaccine candidates have targeted on only one stage of the parasite life cycle, utilizing only one arm of the immune system. In contrast, the Kappe Lab has found that GAPs which halt late in the liver generate an immune response as complex as the malaria parasite itself—targeting all host stages of the parasite life cycle with both T cells and antibodies. Researchers in the lab have also shed light on how the parasite is sensed early in infection by the “innate” immune response and are investigating how this can program the robust “adaptive” or memory immune response during GAP vaccination. The Lab has also expanded the tools to study the immune responses to malaria by pioneering methods to evaluate prevention of liver stage development in both rodent malaria models and human malaria utilizing human liver-chimeric mice. The Lab is also using information gleaned from studying the basic biology of the parasite to identify new immune targets for use in conventional subunit (single protein) vaccines. These combined efforts are guiding the lab in developing the next generation of malaria vaccines capable of generating a multifaceted and potent immune response that can match and defeat the most complex disease ever targeted for eradication.
Genetically Attenuated Parasite (GAP) Strains for Vaccination
The rodent malaria work has enabled us to identify genes that, when deleted, cause a complete block in parasite development in the liver. The fact that mutant parasites infect the liver but cannot develop to a disease-causing blood stage infection allowed us to test their potential as live-attenuated vaccines. For example, mice immunized with sporozoites from rodent malaria parasites that lack the genes UIS3, UIS4, or P52/P36 are completely protected against subsequent challenge with infectious sporozoites. This protection is long lasting. Thus, attenuated parasites are powerful vaccines and completely protect against malaria. The Lab has identified the orthologous genes in P. falciparum and created the first dual gene deletion strain, Pf p52(-)/p36(-). This strain has been assessed for growth defects in hepatocyte culture and a first generation humanized mouse model that harbors human hepatocytes. A phase I clinical trial showed that Pf p52(-)/p36(-) GAP was severely but not completely attenuated in human infection.
To achieve full attenuation we thus recently created a triple gene deletion Pf GAP by introduction of an additional gene deletion in the SAP1 locus (Pf p52(-)/p36(-)/sap1(-) GAP), which was previously shown by us to be essential for successful liver stage infection in rodent malaria parasites. This next generation GAP 3KO strain has completed a phase I clinical study in early 2015. Using the rodent malaria model, the Kappe Lab is continuously refining live-attenuated vaccine design to achieve complete attenuation and optimal immunogenicity.
Our work and the work of colleagues critically depend on malaria parasite infection in mosquitoes and production of sporozoites for lab experiments. We maintain state-of-the-art insectaries that breed and house Anopheles mosquitoes. Mosquitoes are infected with rodent malaria parasites and with Plasmodium falciparum, which are maintained under safe containment conditions. Malaria parasite mosquito stages are then isolated and used for experimentation. Infected mosquitoes are also used for malaria challenge studies of human volunteers in our Malaria Clinical Trials Center.