Robert E. Sinden

ChAd63-MVA-vectored blood-stage malaria vaccines targeting MSP1 and AMA1: assessment of efficacy against mosquito bite challenge in humans

The induction of cellular immunity, in conjunction with antibodies, may be essential for vaccines to protect against blood-stage infection with the human malaria parasite Plasmodium falciparum. We have shown that prime-boost delivery of P. falciparum blood-stage antigens by chimpanzee adenovirus 63 (ChAd63) followed by the attenuated orthopoxvirus MVA is safe and immunogenic in healthy adults. Here, we report on vaccine efficacy against controlled human malaria infection delivered by mosquito bites. The blood-stage malaria vaccines were administered alone, or together (MSP1+AMA1), or with a pre-erythrocytic malaria vaccine candidate (MSP1+ME-TRAP). In this first human use of coadministered ChAd63-MVA regimes, we demonstrate immune interference whereby responses against merozoite surface protein 1 (MSP1) are dominant over apical membrane antigen 1 (AMA1) and ME-TRAP. We also show that induction of strong cellular immunity against MSP1 and AMA1 is safe, but does not impact on parasite growth rates in the blood. In a subset of vaccinated volunteers, a delay in time to diagnosis was observed and sterilizing protection was observed in one volunteer coimmunized with MSP1+AMA1-results consistent with vaccine-induced pre-erythrocytic, rather than blood-stage, immunity. These data call into question the utility of T cell-inducing blood-stage malaria vaccines and suggest that the focus should remain on high-titer antibody induction against susceptible antigen targets.

A novel multiple-stage antimalarial agent that inhibits protein synthesis

There is an urgent need for new drugs to treat malaria, with broad therapeutic potential and novel modes of action, to widen the scope of treatment and to overcome emerging drug resistance. Here we describe the discovery of DDD107498, a compound with a potent and novel spectrum of antimalarial activity against multiple life-cycle stages of the Plasmodium parasite, with good pharmacokinetic properties and an acceptable safety profile. DDD107498 demonstrates potential to address a variety of clinical needs, including single-dose treatment, transmission blocking and chemoprotection. DDD107498 was developed from a screening programme against blood-stage malaria parasites; its molecular target has been identified as translation elongation factor 2 (eEF2), which is responsible for the GTP-dependent translocation of the ribosome along messenger RNA, and is essential for protein synthesis. This discovery of eEF2 as a viable antimalarial drug target opens up new possibilities for drug discovery.

A long-duration dihydroorotate dehydrogenase inhibitor (DSM265) for prevention and treatment of malaria

The antimalarial drug DSM265 displays activity against blood and liver stages of Plasmodium falciparum and has a long predicted half-life in humans. Long-acting new treatment for drug-resistant malaria Malaria kills 0.6 million people annually, yet current malaria drugs are no longer fully effective because the parasite that causes malaria is becoming resistant to these agents. Phillips et al. have identified a new drug that kills both drug-sensitive and drug-resistant malaria parasites by targeting the ability of the parasite to synthesize the nucleotide precursors required for synthesis of DNA and RNA. This drug kills parasites in both the blood and liver and is sufficiently long-acting that it is expected to cure malaria after a single dose or to be effective if dosed weekly for chemoprevention. Malaria is one of the most significant causes of childhood mortality, but disease control efforts are threatened by resistance of the Plasmodium parasite to current therapies. Continued progress in combating malaria requires development of new, easy to administer drug combinations with broad-ranging activity against all manifestations of the disease. DSM265, a triazolopyrimidine-based inhibitor of the pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase (DHODH), is the first DHODH inhibitor to reach clinical development for treatment of malaria. We describe studies profiling the biological activity, pharmacological and pharmacokinetic properties, and safety of DSM265, which supported its advancement to human trials. DSM265 is highly selective toward DHODH of the malaria parasite Plasmodium, efficacious against both blood and liver stages of P. falciparum, and active against drug-resistant parasite isolates. Favorable pharmacokinetic properties of DSM265 are predicted to provide therapeutic concentrations for more than 8 days after a single oral dose in the range of 200 to 400 mg. DSM265 was well tolerated in repeat-dose and cardiovascular safety studies in mice and dogs, was not mutagenic, and was inactive against panels of human enzymes/receptors. The excellent safety profile, blood- and liver-stage activity, and predicted long half-life in humans position DSM265 as a new potential drug combination partner for either single-dose treatment or once-weekly chemoprevention. DSM265 has advantages over current treatment options that are dosed daily or are inactive against the parasite liver stage.

Evaluation of the Efficacy of ChAd63-MVA Vectored Vaccines Expressing Circumsporozoite Protein and ME-TRAP Against Controlled Human Malaria Infection in Malaria-Naive Individuals.

Background. Circumsporozoite protein (CS) is the antigenic target for RTS,S, the most advanced malaria vaccine to date. Heterologous prime-boost with the viral vectors simian adenovirus 63 (ChAd63)-modified vaccinia virus Ankara (MVA) is the most potent inducer of T-cells in humans, demonstrating significant efficacy when expressing the preerythrocytic antigen insert multiple epitope–thrombospondin-related adhesion protein (ME-TRAP). We hypothesized that ChAd63-MVA containing CS may result in a significant clinical protective efficacy. Methods. We conducted an open-label, 2-site, partially randomized Plasmodium falciparum sporozoite controlled human malaria infection (CHMI) study to compare the clinical efficacy of ChAd63-MVA CS with ChAd63-MVA ME-TRAP. Results. One of 15 vaccinees (7%) receiving ChAd63-MVA CS and 2 of 15 (13%) receiving ChAd63-MVA ME-TRAP achieved sterile protection after CHMI. Three of 15 vaccinees (20%) receiving ChAd63-MVA CS and 5 of 15 (33%) receiving ChAd63-MVA ME-TRAP demonstrated a delay in time to treatment, compared with unvaccinated controls. In quantitative polymerase chain reaction analyses, ChAd63-MVA CS was estimated to reduce the liver parasite burden by 69%–79%, compared with 79%–84% for ChAd63-MVA ME-TRAP. Conclusions. ChAd63-MVA CS does reduce the liver parasite burden, but ChAd63-MVA ME-TRAP remains the most promising antigenic insert for a vectored liver-stage vaccine. Detailed analyses of parasite kinetics may allow detection of smaller but biologically important differences in vaccine efficacy that can influence future vaccine development. Clinical Trials Registration. NCT01623557.

Transmission-blocking interventions eliminate malaria from laboratory populations

Transmission-blocking interventions aim to reduce the prevalence of infection in endemic communities by targeting Plasmodium within the insect host. Although many studies have reported the successful reduction of infection in the mosquito vector, direct evidence that there is an onward reduction in infection in the vertebrate host is lacking. Here we report the first experiments using a population, transmission-based study of Plasmodium berghei in Anopheles stephensi to assess the impact of a transmission-blocking drug upon both insect and host populations over multiple transmission cycles. We demonstrate that the selected transmission-blocking intervention, which inhibits transmission from vertebrate to insect by only 32%, reduces the basic reproduction number of the parasite by 20%, and in our model system can eliminate Plasmodium from mosquito and mouse populations at low transmission intensities. These findings clearly demonstrate that use of transmission-blocking interventions alone can eliminate Plasmodium from a vertebrate population, and have significant implications for the future design and implementation of transmission-blocking interventions within the field.

Comparative Assessment of Transmission-Blocking Vaccine Candidates against Plasmodium falciparum

Malaria transmission-blocking vaccines (TBVs) target the development of Plasmodium parasites within the mosquito, with the aim of preventing malaria transmission from one infected individual to another. Different vaccine platforms, mainly protein-in-adjuvant formulations delivering the leading candidate antigens, have been developed independently and have reported varied transmission-blocking activities (TBA). Here, recombinant chimpanzee adenovirus 63, ChAd63, and modified vaccinia virus Ankara, MVA, expressing AgAPN1, Pfs230-C, Pfs25, and Pfs48/45 were generated. Antibody responses primed individually against all antigens by ChAd63 immunization in BALB/c mice were boosted by the administration of MVA expressing the same antigen. These antibodies exhibited a hierarchy of inhibitory activity against the NF54 laboratory strain of P. falciparum in Anopheles stephensi mosquitoes using the standard membrane feeding assay (SMFA), with anti-Pfs230-C and anti-Pfs25 antibodies giving complete blockade. The observed rank order of inhibition was replicated against P. falciparum African field isolates in A. gambiae in direct membrane feeding assays (DMFA). TBA achieved was IgG concentration dependent. This study provides the first head-to-head comparative analysis of leading antigens using two different parasite sources in two different vector species, and can be used to guide selection of TBVs for future clinical development using the viral-vectored delivery platform.

The biology of sexual development of Plasmodium: the design and implementation of transmission-blocking strategies

A meeting to discuss the latest developments in the biology of sexual development of Plasmodium and transmission-control was held April 5-6, 2011, in Bethesda, MD. The meeting was sponsored by the Bill & Melinda Gates Foundation and the National Institutes of Health, National Institute of Allergy and Infectious Diseases (NIH/NIAID) in response to the challenge issued at the Malaria Forum in October 2007 that the malaria community should re-engage with the objective of global eradication. The consequent rebalancing of research priorities has brought to the forefront of the research agenda the essential need to reduce parasite transmission. A key component of any transmission reduction strategy must be methods to attack the parasite as it passes from man to the mosquito (and vice versa). Such methods must be rationally based on a secure understanding of transmission from the molecular-, cellular-, population- to the evolutionary-levels. The meeting represented a first attempt to draw together scientists with expertise in these multiple layers of understanding to discuss the scientific foundations and resources that will be required to provide secure progress toward the design and successful implementation of effective interventions.

Imaging-Based High-Throughput Screening Assay To Identify New Molecules with Transmission-Blocking Potential against Plasmodium falciparum Female Gamete Formation

ABSTRACT In response to a call for the global eradication of malaria, drug discovery has recently been extended to identify compounds that prevent the onward transmission of the parasite, which is mediated by Plasmodium falciparum stage V gametocytes. Lately, metabolic activity has been used in vitro as a surrogate for gametocyte viability; however, as gametocytes remain relatively quiescent at this stage, their ability to undergo onward development (gamete formation) may be a better measure of their functional viability. During gamete formation, female gametocytes undergo profound morphological changes and express translationally repressed mRNA. By assessing female gamete cell surface expression of one such repressed protein, Pfs25, as the readout for female gametocyte functional viability, we developed an imaging-based high-throughput screening (HTS) assay to identify transmission-blocking compounds. This assay, designated the P. falciparum female gametocyte activation assay (FGAA), was scaled up to a high-throughput format (Z′ factor, 0.7 ± 0.1) and subsequently validated using a selection of 50 known antimalarials from diverse chemical families. Only a few of these agents showed submicromolar 50% inhibitory concentrations in the assay: thiostrepton, methylene blue, and some endoperoxides. To determine the best conditions for HTS, a robustness test was performed with a selection of the GlaxoSmithKline Tres Cantos Antimalarial Set (TCAMS) and the final screening conditions for this library were determined to be a 2 μM concentration and 48 h of incubation with gametocytes. The P. falciparum FGAA has been proven to be a robust HTS assay faithful to Plasmodium transmission-stage cell biology, and it is an innovative useful tool for antimalarial drug discovery which aims to identify new molecules with transmission-blocking potential.

The cell biology of malaria infection of mosquito: advances and opportunities

Recent reviews (Feachem et al.; Alonso et al.) have concluded that in order to have a sustainable impact on the global burden of malaria, it is essential that we knowingly reduce the global incidence of infected persons. To achieve this we must reduce the basic reproductive rate of the parasites to < 1 in diverse epidemiological settings. This can be achieved by impacting combinations of the following parameters: the number of mosquitoes relative to the number of persons, the mosquito/human biting rate, the proportion of mosquitoes carrying infectious sporozoites, the daily survival rate of the infectious mosquito and the ability of malaria‐infected persons to infect mosquito vectors.

The design and interpretation of laboratory assays measuring mosquito transmission of Plasmodium

Since 2010 two global reviews of malaria research have recognized that local elimination and eradication of Plasmodium parasites are key drivers for further experimentation. To achieve these ambitious objectives it is universally recognized we must reduce malaria transmission through the mosquito vectors. A plethora of new laboratory assays are being developed to interrogate malaria transmission from the gametocyte to the sporozoite stage: assays that augment well-established field protocols to determine the entomological inoculation rate. However, the diverse readouts of these assays are not directly comparable. Here we attempt to identify the utility of each assay and provide rational frameworks by which to compare the impacts recorded by the diverse methodologies.