Malaria Vaccines

BACKGROUND

Human malaria is caused by five species of the Plasmodium protozoan parasites: P. falciparum , P. vivax , P. ovale , P. malariae , and P. knowlesi . Malaria caused by these protozoan parasites occupies a remarkable place in the history of infectious diseases. In ancient Chinese, Egyptian, Greek, and Roman writings, intermittent fevers and malaria symptoms of headaches and chills were mentioned. These symptoms were associated with three demons carrying a hammer, a pail of water, or a stove in Chinese beliefs. In the 1st century, malaria spread northward from Greece and Italy to England and Denmark, becoming established throughout Europe for the next 2000 years. In the Americas, P. vivax malaria was probably imported by European explorers and colonists, and P. falciparum , with African slaves. Both parasites were common in the United States (U.S.) between the 18th and the early 20th century, draining the physical and economic health of entire regions of the country and leading to the foundation of the U.S. public health agency (Centers for Disease Control and Prevention). Until the early 20th century, malaria was endemic across every continent except Antarctica.

EPIDEMIOLOGY AND BURDEN OF DISEASE

Malaria occurs where the environment supports competent Anopheles mosquito vectors with access to malaria-infected hosts. By the 1950s, malaria was controlled in the U.S. Europe, and Australia through the use of insecticides and environmental strategies. In the U.S. the number of malaria cases diagnosed has been stable at approximately 2000 per year, with most cases occurring in travelers and immigrants returning from countries where malaria transmission occurs. In the past century, more than 100 countries eliminated malaria (no indigenous malaria cases for 3 consecutive years), encouraging national malaria control programs to consider elimination to be an attainable goal and resulting in the addition of malaria eradication in the global health agenda. , The dramatic fall in malaria incidence, resulting in its elimination in some countries, was driven by a multifaceted strategy including environmental control, insecticides, bed nets, improved rapid diagnostic tests, chemotherapy, artemisinin-combination therapy, and better case management. ^,

P. falciparum is responsible for the vast majority of malaria cases and most malaria deaths globally, and is the most prevalent on the African continent where it causes approximately 99% of the 241 million cases annually ( Online Supplement 1 ). ^{, ,} P. vivax is the second most common species, causing about 3% of malaria cases globally in 2019, and accounting for most malaria cases occurring in temperate regions. ^, P. vivax is responsible for less than 1% of cases in Africa (mainly in Ethiopia), approximately 25% of cases in the Middle East, 33% of cases in the Western Pacific, 50% of cases in South-East Asia, and almost 75% of cases in Central and South America ( Online Supplement 1 ). ^{, ,} P. ovale is endemic in Africa, South-East Asia, and the Western Pacific region. P. malariae is distributed worldwide, with cases reported in Africa, South-East Asia, South America, and Western Pacific. P. knowlesi was recognized in 2008 as the fifth Plasmodium species potentially leading to fatal outcome in humans. P. knowlesi transmission is zoonotic, as the parasite’s natural reservoir are macaques living in South-East Asia. P. ovale , P. malariae , and P. knowlesi represent only a small percentage of infections globally, although higher proportions of cases may occasionally be reported (up to 15% of malaria cases were caused by P. ovale in Papua New Guinea, 24.3% of cases were caused by P. malariae at the Thai–Myanmar border, and 41% of cases were caused by P. knowlesi in Sarawak, Malaysia ). In addition, these Plasmodia are often detected as mixed infections with P. falciparum . While they are usually causing less severe disease, severe cases and long-term consequences have been reported for each of them.

The world has achieved tremendous progress in the fight against malaria between 2000 and 2015, reducing malaria cases incidence by 41% and malaria deaths by 62%. However, more than three billion people still live in malaria-endemic areas, and a slowing in the rate of decline was observed for both uncomplicated clinical malaria and malaria deaths between 2015 and 2019. In the most recent World Malaria Report, there were an estimated 241 million malaria cases in 85 endemic countries in 2020, showing an increase compared with the 238 million cases reported in 2000 and the 227 million cases reported in 2019. The estimated number of annual malaria deaths declined from 896,000 in 2000 to 558,000 in 2019, and 627,000 in 2020. During the COVID-19 pandemic, disruptions in the delivery of malaria services contributed to the considerable increases seen in malaria cases (14 million) and deaths (69,000) between 2019 and 2020. Of note, the quantification of the clinical burden of malaria is imprecise and is impacted by variable access to diagnosis and treatment, differences in case definitions, and low specificity of diagnosis. ^,

Nowhere has malaria exacted a greater toll than in sub-Saharan Africa. Testimony to malaria’s ancient hold on Africa includes the survival and fitness of the heterozygous hemoglobin S mutated allele, which is the cause of an inherited hemoglobin disorder (sickle cell anemia), owing to a balanced polymorphism in which strong selective pressure in favor of the sickle cell allele confers protection against P. falciparum malaria. Another evidence is the near absence in West Africa of the Duffy blood group Fy antigen on the surface of red blood cells (RBCs), which is an essential receptor for the invasion of P. vivax asexual stage parasites into erythrocytes. ^, In the 20th and 21st centuries, 80–90% of malaria cases and deaths were reported in sub-Saharan Africa, primarily in children younger than 5 years of age. In 2019, six countries (Nigeria, Democratic Republic of the Congo, the United Republic of Tanzania, Mozambique, Niger, and Burkina Faso) accounted for 51% of all cases globally.

According to the World Health Organization (WHO), the milestones established by Global Technical Strategy for 2016–2030 to reduce malaria morbidity will not be achieved without accelerated implementation of existing strategies and rapid expansion of new tools, including malaria vaccines. Despite advances in effective treatments and vector control, a safe and effective vaccine remains an essential missing tool in the fight against malaria.

PARASITE LIFE CYCLE

An understanding of the parasite’s life cycle is important to exploit vulnerable targets on their surface for vaccine development ( Fig. 37.1 ). All malaria species have a complex life cycle that begins when female malaria-infected Anopheles mosquitoes bite the skin of humans seeking a blood meal. Approximately 15–100 infectious stage sporozoites are injected into the skin and migrate to the liver either through the lymphatics or direct blood circulation. They traverse cell membranes of endothelial cells, Kupffer cells, and several liver hepatocytes before invading a single hepatocyte wherein they develop into exoerythrocytic stage parasites. This stage of the life cycle is known as the preerythrocytic stage , which is asymptomatic in the host and is the primary target of vaccines against sporozoites and developing liver stage parasites. During the 9–14 days following invasion of a hepatocyte, parasites develop, multiply thousands of times, enter blood circulation, and invade human erythrocytes. The majority of blood stage parasites replicate asexually in RBCs, with repeated cycles of amplification (every 24 hours for P. knowlesi , 48 hours for P. falciparum , P. vivax , and P. ovale, and 72 hours for P. malariae ). Once merozoites erupt and invade circulating RBCs, symptoms of clinical disease become apparent depending upon parasite density in the circulation and level of blood-stage immunity. All malaria symptoms, morbidity, and mortality are attributed to the asexual blood stage of infection. Following RBC invasion, merozoites develop through ring, trophozoite, and schizont stages. They replicate to produce 16–32 daughter merozoites over 24–48 hours depending on the species. The newly formed intraerythrocytic merozoites are released into the circulation and the replication cycle is repeated, allowing for exponential expansion of the parasites in the host. A small proportion of blood stage parasites develop into male and female gametocytes during the sexual stage of the parasite’s life cycle. At least one male and one female gametocyte need to be taken up by the bite of an Anopheles mosquito when it obtains a blood meal to allow the formation of a zygote. The zygote is transformed into a mobile ookinete, which traverses the mosquito’s midgut and develops into the oocyst stage from which infective sporozoites are formed. Sporozoites find their way to the mosquito’s salivary glands where they can initiate a new infection upon feeding on a susceptible host.

Human malaria parasites share common life cycle stages with notable exceptions. Following invasion of hepatocytes, some P. vivax and P. ovale sporozoites can enter a dormant period of quiescence (termed hypnozoites ). ^{, ,} Following a time period ranging from a few weeks to a year or more, hypnozoites may resume development within the hepatocyte, multiply, and emerge into the blood stream to initiate repeated episodes of symptomatic blood stage infection. ^, Additionally, P. vivax asexual blood stage parasites preferentially invade reticulocytes (rather than mature RBCs invaded by P. falciparum merozoites), and can convert to gametocytes and be transmitted to mosquitoes earlier in the course of infection.

CLINICAL COURSE OF DISEASE

Characteristic signs and symptoms of malaria include fever, malaise, headache, muscle aches, anorexia, nausea, and vomiting. ^{, ,} Symptoms appear to coincide with the synchronized lysis of RBCs by malarial toxins released along with merozoites. Severe disease occurs most frequently with P. falciparum , leading to severe malarial anemia, cerebral malaria, and multiorgan failure. Severe malarial anemia is primarily observed in very young children and results from both erythrocyte destruction from parasite multiplication and dyserythropoiesis. Sequestration of mature schizont-infected RBCs, which cytoadhere to postcapillary venules, may result in serious complications in the brain (leading to cerebral malaria), heart, gastrointestinal tract, and skin in children and nonimmune adults. ^{, , ,} Severe clinical sequelae (e.g., acute renal failure, severe anemia, hypoglycemia, and pulmonary edema) occur most commonly in malaria-naïve adult populations. ^{, , ,} Pregnant women and the developing fetus are at increased risk for severe disease, indicating unique mechanisms of pathogenesis due to sequestration of blood schizonts in the placenta. Coinfection with human immunodeficiency virus (HIV) predisposes to more severe disease. ^,

RATIONALE AND STRATEGIES FOR VACCINE DEVELOPMENT

Is a malaria vaccine feasible when compared to other well-described vaccines against bacterial and viral pathogens? Classical vaccine development models are based on the fundamental principle that natural infection elicits immune responses that clear the pathogen and prevent re-infection. Naturally acquired immunity against clinical and severe malaria takes years to develop, and is only maintained through constant exposure to repeated infectious mosquito bites in high transmission settings. ^, Immunity is species-specific (immunity to P. falciparum does not prevent P. vivax infection), stage-specific (preerythrocytic stage protection does not protect against blood stage infection), and transient (protection against clinical malaria wanes rapidly in the absence of constant exposure).

Malaria is different from classical acute viral infections in that individuals infected with Plasmodium parasites infrequently clear blood stage parasites. Chronic asymptomatic infections with low level submicroscopic asexual stage parasites persist and are thought to be important to maintain naturally acquired immunity. These asymptomatic infections may be accompanied by long-term persistence of sexual stage gametocytes and sustained ongoing transmission.

The complexity of the Plasmodium parasites, with a genome that contains more than 5000 open reading frames from antigenically diverse and variant parasites, creates a complex setting for the identification of suitable antigenic targets for vaccine development. Unlike most bacteria and viruses, in which the full repertoire of antigenic targets are present during the entire incubation period prior to onset of illness, many potential targets of protective immunity against the malaria parasite are stage-specific and exposed to the immune system for short periods of time, hampering anamnestic and memory recall responses upon re-exposure. Most candidate vaccines currently in development focus on a single stage of the parasite’s life cycle. Although some target antigens are expressed in multiple stages, a highly efficacious vaccine with durable protection might require distinct antigens from different stages as proposed over 20 years ago. ^,

Throughout its life cycle, the malaria parasite passes through two “biological bottlenecks,” where the limited number of parasites creates a compelling rationale for stage-specific vaccines ( Fig. 37.2 ). The first chokepoint, which is the target of most candidate malaria vaccines, is when a low number of sporozoites are delivered into the skin following a mosquito bite. The aim of these vaccines is to prevent invasion and development of parasites within the host hepatocytes. Vaccine-induced antibodies, which kill or inactivate sporozoites, and cellular immune effector cells against the remaining liver stage schizonts form the basis for most vaccines under development, including the RTS,S vaccine, other circumsporozoite (CS)-based vaccines (e.g., R21), and attenuated sporozoites. The responses to vaccines targeting this life cycle bottleneck are an “all-or-none” phenomenon, meaning that any sporozoite or liver stage schizont that is not killed or inactivated will proceed to blood stage, multiply exponentially as asexual stage parasites in erythrocytes in the circulation, precipitate clinical illness, and potentially lead to severe and lethal consequences.

Since the primary objective of vaccine developers is to reduce malaria-attributed morbidity and mortality, the development of blood stage vaccines could appear as a priority at first glance. However, to be effective, blood stage vaccines must kill millions of parasites and rapidly reduce parasite multiplication rates. This proved to be challenging because the exponential multiplication of merozoites during the blood stage is an ideal biological environment for selection of variant or mutant parasite strains that permit the parasite to escape from naturally acquired or vaccine-induced immune responses. Another challenge is that blood stage parasites use many redundant mechanisms to invade erythrocytes, providing them with alternative routes of entry if one is blocked by a vaccine targeting a single invasion pathway.

The second chokepoint, which is target of so called transmission-blocking vaccines, occurs when a limited number of gametocytes are responsible for the transmission from the human host to the mosquito vector. The aim of these transmission-blocking vaccines is to prevent sporozoite development in the midgut and salivary glands of mosquitoes by targeting sexual stage gametocytes, gametes, or ookinetes within the host or the mosquito. Limiting the force of infection at the sporozoite stage (preerythrocytic stage vaccines) or the transmission stage (transmission-blocking vaccines) by vaccine-induced antibodies greatly diminishes the probability that parasites continue development within the host or mosquito.

A large number of malaria vaccine candidates targeting each of the distinct stages of the parasite’s life cycle in humans have been tested preclinically or in Phase I/II/III clinical trials ( Online Supplement 2 ), ^, but only one (the RTS,S vaccine) has completed late-stage development ( Online Supplement 3 ).

CONTROLLED HUMAN MALARIA INFECTION MODEL

Human challenge models for malaria are major accelerators for vaccine development and have informed our knowledge of immunologic correlates of protection. ^, Two types of human challenge models exist for malaria: the sporozoite challenge model (particularly suited for preerythrocytic vaccines but also useful for vaccines targeting other stages of the parasite’s life cycle) and the induced blood stage malaria model (useful for asexual and sexual blood stage vaccines).

One of the key applications of the human challenge models for malaria is to assess the potential for vaccine efficacy in small numbers of volunteers early during vaccine development. This has been particularly relevant for preerythrocytic malaria vaccines, for which the primary endpoint in Controlled Human Malaria Infection (CHMI) vaccine trials is the presence or absence of blood stage parasites in volunteers immunized with an experimental vaccine candidate. ^, The biologic basis for the most prevalent challenge models is that the bite of an infectious Anopheles mosquito initiates infection by injecting small numbers of sporozoites into the skin, which culminates with the appearance of ring stage asexual parasites in the circulation approximately 7–10 days later. The establishment of a continuous culture system of P. falciparum blood stage parasites improved the challenge model, allowing for reproducibility, consistency, and predictability of infection in volunteers. ^, Since the 1980s, dozens of P. falciparum malaria vaccine trials using the mosquito bite challenge models have been conducted around the world. Compared to P. falciparum parasites, P. vivax parasites cannot be continuously cultured in vitro to produce gametocytes that infect Anopheles mosquitoes. Therefore, P. vivax challenge models are limited to using naturally acquired parasites obtained from patients who present to health treatment facilities with clinical P. vivax malaria infection.

In more recent years, an alternative P. falciparum challenge model has been developed, in which volunteers are infected with live unattenuated cryopreserved sporozoites by direct venous inoculation. ^, Because mosquito bite challenge is laborious and requires extensive insectary facilities that are not amenable to transfer across clinical trial sites, a developmental program was initiated more than a decade ago to assess whether sporozoites harvested from a mosquito under Good Manufacturing Practice standards could be cryopreserved, thawed, and injected into human volunteers to establish a liver-stage infection with subsequent emergence of blood stage parasites into the peripheral circulation. While a human infection requires as few as 15–100 sporozoites delivered through the bite of a single infected mosquito, ^, many more cryopreserved sporozoites need to be injected to establish infection, probably because the processes required to harvest, freeze, thaw, and administer sporozoites with a needle and syringe take a toll on parasite viability and infectiousness. Nevertheless, many studies have been conducted to assess whether such cryopreserved sporozoites could be administered in a similar manner to delivery by an infectious mosquito bite. This includes several studies that assessed intradermal, ^, subcutaneous, intramuscular, ^{, ,} and intravenous administration by a procedure called direct venous inoculation. ^,

VACCINES TARGETING THE PREERYTHROCYTIC STAGE

Vaccine development strategies targeting the infectious sporozoites or liver stage schizonts aim to elicit protective responses that neutralize sporozoites’ ability to invade or develop within hepatocytes ( Fig. 37.2 ).