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The development of a vaccine against Plasmodium falciparum

The development of a vaccine against Plasmodium falciparumStudent number: 06901380

Malaria due to infection with Plasmodium falciparum is a leading cause of death in children under the age of 5 years. Discuss why there is not a useful vaccine against malaria when other diseases that used to be common in this age group (such as measles and polio) have been successfully controlled by vaccination.It is estimated that malaria is the ninth leading cause of death in low income countries, causing between one and three million deaths per annum worldwide (Guerra et al., 2008, Snow et al., 2005). The majority of these deaths are caused by Plasmodium falciparum, the most deadly of the four species of malaria affecting humans (Guerra et al., 2008). Current research efforts are targeted at vaccine prevention of Plasmodium falciparum malaria. It is not possible to create a vaccine that mimics natural immunity, as with vaccines for polio and measles, since the inherent immune response in an immunocompetent host does not provide sterile immunity. This is a consequence of the complex lifecycle of the malaria parasite and a number of immune evasion techniques the parasite employs. This essay will briefly discuss the lifecycle of Plasmodium falciparum, the normal immune response to infection and the immune evasion techniques the parasite employs. The potential vaccine targets will then be discussed with reference to the efficacy of vaccines currently in development and trials.The life-cycle of Plasmodium falciparum

The lifecycle of the protozoan, Plasmodium falciparum, involves transmission from human to human via the Anopheles mosquito vector, as illustrated in figure 1. The salivary glands of an infected Anopheles mosquito contain a large number of sporozoites, the infective form of the Plasmodium parasite. As the Anopheles initiates feeding it injects a small amount of saliva containing sporozoites into the wound. These are rapidly taken up by the liver where they multiply inside hepatocytes as merozoites (Kumar and Clark, 2002). These two phases are collectively known as the pre-erythrocytic stage. After several days the hepatocytes rupture releasing the merozoites into the blood. The merozoites then infect erythrocytes and replicate in a phase termed the asexual blood-stage. In this stage the merozoites differentiate first into a trophozoite and then a schizont. At the schizont stage the Plasmodium undergoes DNA replication followed by cellular segmentation to form between 8 and 24 new merozoites (Kumar and Clark, 2002). Eventually the erythrocyte ruptures and the merozoites are released to infect further erythrocytes. This cycle is known as erythrocytic schizogony and is responsible for many of the clinical features of malaria. A minority of merozoites differentiate into gametocytes, the sexual form of the parasite, which are not released from the erythrocytes but remain intracellular until they are taken up by a feeding mosquito (Kumar and Clark, 2002). If the gametocytes are successful in infecting a mosquito they differentiate into gametes which then form zygotes. These then multiply again, changing from zygote to ookinete, oocyst and then sporozoites ready for infection of the next human host (Hall et al., 2005).

The immune response and immune evasion techniques

The immune response to malaria has a complexity which complements the parasites complex lifecycle. Different stages of the parasite present different antigens, requiring multiple different immune responses to eradicate the infection. However, the malaria protozoan has a number of highly effective methods of evading the bodies immune response (Schofield and Tachado, 1996). Even in areas where malaria is endemic, immunity is slowly acquired, unstable and is usually incomplete (Reeder and Brown, 1996). Identified immune responses against the pre-erythrocytic stage include the development of antibodies against the circumsporozoite protein which block hepatocyte invasion of sporozoites (Holder, 1994). If the sporozoites successfully invade hepatocytes a cytotoxic T cell response is triggered by parasitic protein derived peptides presented on the surface of the infected hepatocytes (Hill et al., 1992).

The most significant immune response is targeted at the asexual blood-stage parasites. Antibodies against surface proteins of the merozoites which can prevent erythrocyte invasion have been demonstrated (Holder, 1996). These surface antigens include apical membrane antigen 1 (AMA-1) and merozoites surface antigens 1 and 2 (MSA-1 and MSA-2). Once inside the erythrocyte the merozoite is initially shielded from immune surveillance mechanisms. However, as the parasite develops inside the erythrocyte changes occur in the erythrocyte membrane triggering its sequestration and destruction within the spleen (Foley and Tilley, 1995). The developing parasite is able to prevent this destruction by causing adherence of the erythrocyte to the postcapillary venular endothelium (Brown and Rogerson, 1996, Gruarin et al., 2001). The occurrence of this sequestration within the brain is the likely pathological mechanism underlying the development of cerebral malaria (Aikawa et al., 1990). Cytoadhesion is achieved by the insertion of parasitic adhesion molecules into the erythrocyte cell membrane such as Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1). These adhesion molecules also act as foreign antigens with antibodies developed against them causing opsonisation of the infected erythrocyte and prevention of cytoadhesion (Reeder and Brown, 1996).

The genome of the parasite has evolved to maximise the array of immunogenic proteins through two distinct mechanisms; antigenic variation and antigenic diversity. Antigenic variation is the ability of a single clonal parasite population to alter the antigens presented to the immune system. The parasite exhibits polymorphisms in many immunogenic proteins. For example a monoclonal parasite population contains around 40 alleles for the surface antigen PfEMP1 (Saul, 1999). Switching of antigenic phenotype occurs via a stochastic process at rates comparable with the time taken to generate a high-titre antibody response (Saul, 1999). High rates of antigenic phenotype switching may allow immune mediated negative selection pressures to permit replication of parasites which subsequently express unrecognised antigens and thereby determine this rate of switching (Reeder and Brown, 1996). More explicitly; the rate at which novel antigens are displayed by the parasite may be determined by the rate at which the immune system responds to the previously displayed antigens (Saul, 1999). Antigen variation permits chronic infection with successive generations of parasites unimpeded by the antibodies generated to combat their predecessors.By contrast with antigenic variation, antigenic diversity is defined as the expression of different alleles in different Plasmodium populations. A number of different forms of the MSA-1 antigen have been identified in Plasmodium falciparum and their corresponding genes sequenced (Kemp, 1992). These genes (shown in figure 2) show a level of amino acid sequence homology as low as 10% in some regions (Tanabe et al., 1987). Mechanisms by which intragenic recombination occurs at the meiosis stage of parasite reproduction have been suggested (Kemp, 1992) and may provide a method to facilitate the transformation of the antigenic repertoire over time. The MSA-2 gene has also been reported to have a number of different alleles (Kemp, 1992).Currently it is unclear how natural immunity to malaria develops. The primary controversy is whether immunity is achieved by generating response to a finite number of antigenic variations or by generating a response to a single poorly immunogenic epitope such as a conserved domain of PfEMP (Reeder and Brown, 1996). If immunity is developed through recognition of the diverse repertoire of antigens, a polyclonal reservoir of responding B cells would be required. If immunity is conferred by response to a single epitope it may be possible to induce this response through vaccination with similar epitope of increased immungenicity. Alternatively if immunity is achieved by developing a polyclonal reservoir of responding B cells then a vaccine may need to present multiple immune targets, thus replicating this polyclonal response.Vaccine targets

As the antigens presented to the immune system are different at each stage of the malaria life cycle, current vaccines are designed only to target a single parasite stage. Pre-erythrocytic stage vaccines target surface antigens on the merozoites in order to prevent hepatocyte invasion or trigger the destruction of parasites within infected hepatocytes via T-cell recognition of surface antigens. However these vaccines offer no protection against parasites which emerge from the pre-erythrocytic stage and are subsequently able to undergo asexual replication unimpeded (Cheng et al., 1997). Asexual stage vaccines either target merozoite surface antigens and prevent erythrocyte invasion or infect erythrocyte surface molecules to enhance destruction of infected cells. A final type of vaccine is aimed at preventing sporogenic development within the mosquito vector via the production of antibodies against the mosquito stages of the parasite which would be transmitted to the mosquito with the gametocytes during feeding. These are termed transmission blocking vaccines. Pre-erythrocytic stage vaccines

Sterile immunity to Plasmodium falciparum has been achieved by exposure to around 1000-3000 mosquitoes containing irradiated sporozoites (Hoffman et al., 2002). However this method of vaccination is highly impractical and the current focus is directed towards recombinant and peptide vaccines (Targett, 2005). There a currently a wide range of vaccine technologies under investigation including the use of plasmid DNA, virosomes and synthetic polypeptides but only one construct has shown significant promise in clinical trials, the RTS,S vaccine (Targett, 2005). This is perhaps the most promising malaria vaccine to date. The RTS,S vaccine is a recombinant protein comprised of the Plasmodium falciparum circumsporozoite protein and the hepatitis B surface antigen molecule. It has shown considerable promise in clinical trials including significant protection against infection and clinical disease in children (Bojang et al., 2001). A phase II randomised control trial in Mozambique demonstrated a 58% decrease in severe malaria episodes in children six months after vaccination with RTS,S (Alonso et al., 2004).Asexual stage vaccines

Most potential vaccines against the blood stage of malaria target molecules which are involved with the invasion of erythrocytes. Unfortunately these invasion pathways are redundant in Plasmodium falciparum requiring blockade of multiple targets simultaneously. Some promising results have been demonstrated in vaccines targeting MSA-1, MSA-2 and MSA-3 (Druilhe et al., 2005, Genton et al., 2002). These results are, however, strain specific as a result of the antigenic diversity of these targets. Other vaccine targets include the parasite molecules on the erythrocyte surface. As these molecules express high levels of antigenic variation, vaccine development will be challenging. However one surface antigen VAR2CSA is responsible for erythrocyte sequestration in the placenta and the cause of severe malaria sequelae in pregnancy (Duffy, 2007). This provides the possibility of developing a vaccine beneficial in pregnancy (Duffy, 2007). Transmission blocking vaccines

These vaccines would not provide protection for malaria to the recipient but if successfully developed could form part of a malaria eradication program. Mosquito stage antigens are not exposed to the human immune system and therefore have not developed a high level of antigenic diversity. However some identified gametocyte antigens have been found to have variants and some are strain specific (Kaslow, 1993). Nevertheless animal models of transmission blocking vaccines have shown some success (Saul, 2007). Combination vaccines

It is likely that combination vaccines will offer the highest level of protection against malaria. The combination of a pre-erythrocytic vaccine with an asexual stage vaccine will prevent the problem of parasitic break through that occurs when a pre-erythrocytic vaccine is used alone. Incorporation of a transmission blocking vaccine may reduce the probability of emergence of vaccine resistant clones by limiting malaria spread from vaccinated individuals.

Combination of vaccines which target the same parasite stage through different antigens is also likely to have beneficial outcomes: If the individual component vaccines fail to provide sterile immunity such a vaccine may have increased potency. The existence of a variety of HLA (human leukocyte antigen) types within the human population result in varying responsiveness to current vaccines (Nardin et al., 2000), combination of different target vaccines will increase the chance of generating immunity. A combination vaccine of this type is would also reduce the probability of vaccine resistance emerging.Although the benefits of combination vaccines appear evident the cost of vaccine production rises rapidly with increased complexity and may prove to render such vaccines impractical. For this reason optimisation of individual vaccines should be the principal consideration of current research.

Conclusion

Although an effective and practical malaria vaccine appears to be close many obstacles remain. It is only through an understanding of the molecular and genetic mechanism that underpin the immune evasion of the malaria parasite and the human host interaction with these mechanisms that we can hope to develop such a vaccine. It is likely that a combination vaccine, targeting different parasite stages, will provide the greatest level of protection against clinical malaria and development of vaccine resistant malaria.Word count: 1993References

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Figure 2. A schematic of MSA-1 genes from Plasmodium falciparum. Regions 1 and 17 are involved in membrane transport and attachment and are highly conserved. The remaining variable and semiconserved regions are combined in various ways providing antigenic diversity. Figure adapted from Kemp (1992)

Figure 1. The lifecycle of malaria. Sporozoites are injected via the Anopheles mosquito and travel to the liver (A). Replication and release of merozoites then occurs in hepatocytes (B). Released merozoites repeatedly infect erythrocytes (C) by replication and haemolysis. Some merozoites differentiate into gametocytes which are taken up the feeding of a second Anopheles (D). Finally fertilization and production of infective sporozoites occurs within the mosquito (E). Figure adapted from Richie and Saul (2002).

ADDIN EN.CITE Richie2002282817TL RichieA SaulProgress and challenges for malaria vaccinesNatureNature694-7014152002(Richie and Saul, 2002)

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