Antimalarial Activity of Artefenomel Against Asexual Parasites and Transmissible Gametocytes During Experimental Blood-Stage Plasmodium vivax Infection.

BACKGROUND: Interventions that effectively target Plasmodium vivax are critical for the future control and elimination of malaria. We conducted a P. vivax volunteer infection study to characterize the antimalarial activity of artefenomel, a new drug candidate. METHODS: Eight healthy, malaria-naive participants were intravenously inoculated with blood-stage P. vivax and subsequently received a single oral 200-mg dose of artefenomel. Blood samples were collected to monitor the development and clearance of parasitemia, and plasma artefenomel concentration. Mosquito feeding assays were conducted before artefenomel dosing to investigate parasite transmissibility. RESULTS: Initial parasite clearance occurred in all participants after artefenomel administration (log10 parasite reduction ratio over 48 hours, 1.67; parasite clearance half-life, 8.67 hours). Recrudescence occurred in 7 participants 11-14 days after dosing. A minimum inhibitory concentration of 0.62 ng/mL and minimum parasiticidal concentration that achieves 90% of maximum effect of 0.83 ng/mL were estimated, and a single 300-mg dose was predicted to clear 109 parasites per milliliter with 95% certainty. Gametocytemia developed in all participants and was cleared 4-8 days after dosing. At peak gametocytemia, 75% of participants were infectious to mosquitoes. CONCLUSIONS: The in vivo antimalarial activity of artefenomel supports its further clinical development as a treatment for P. vivax malaria. CLINICAL TRIALS REGISTRATION: NCT02573857.

Lipid Compositions in Infant Formulas Affect the Solubilization of Antimalarial Drugs Artefenomel (OZ439) and Ferroquine during Digestion.

Recent studies have shown that the solubilization of two antimalarial drug candidates, artefenomel (OZ439) and ferroquine (FQ), designed to provide a single-dose combination therapy for uncomplicated malaria can be enhanced using milk as a lipid-based formulation. However, milk as an excipient faces significant quality and regulatory hurdles. We therefore have investigated infant formula as a potential alternative formulation approach. The significance of the lipid species present in a formula with different lipid compositions upon the solubilization of OZ439 and FQ during digestion has been investigated. Synchrotron small-angle X-ray scattering was used to measure the diffraction from a dispersed drug during digestion and thereby determine the extent of drug solubilization. High-performance liquid chromatography was used to quantify the amount of drug partitioned into the digested lipid phases. Our results show that both the lipid species and the amount of lipids administered were key determinants for the solubilization of OZ439, while the solubilization of FQ was independent of the lipid composition. Infant formulas could therefore be designed and used as milk substitutes to tailor the desired level of drug solubilization while circumventing the variability of components in naturally derived milk. The enhanced solubilization of OZ439 was achieved during the digestion of medium-chain triacylglycerols (MCT), indicating the potential applicability of MCT-fortified infant formula powder as a lipid-based formulation for the oral delivery of OZ439 and FQ.

Spray drying OZ439 nanoparticles to form stable, water-dispersible powders for oral malaria therapy.

BACKGROUND: OZ439 is a new chemical entity which is active against drug-resistant malaria and shows potential as a single-dose cure. However, development of an oral formulation with desired exposure has proved problematic, as OZ439 is poorly soluble (BCS Class II drug). In order to be feasible for low and middle income countries (LMICs), any process to create or formulate such a therapeutic must be inexpensive at scale, and the resulting formulation must survive without refrigeration even in hot, humid climates. We here demonstrate the scalability and stability of a nanoparticle (NP) formulation of OZ439. Previously, we applied a combination of hydrophobic ion pairing and Flash NanoPrecipitation (FNP) to formulate OZ439 NPs 150 nm in diameter using the inexpensive stabilizer hydroxypropyl methylcellulose acetate succinate (HPMCAS). Lyophilization was used to process the NPs into a dry form, and the powder's in vitro solubilization was over tenfold higher than unprocessed OZ439. METHODS: In this study, we optimize our previous formulation using a large-scale multi-inlet vortex mixer (MIVM). Spray drying is a more scalable and less expensive operation than lyophilization and is, therefore, optimized to produce dry powders. The spray dried powders are then subjected to a series of accelerated aging stability trials at high temperature and humidity conditions. RESULTS: The spray dried OZ439 powder's dissolution kinetics are superior to those of lyophilized NPs. The powder's OZ439 solubilization profile remains constant after 1 month in uncapped vials in an oven at 50 °C and 75% RH, and for 6 months in capped vials at 40 °C and 75% RH. In fasted-state intestinal fluid, spray dried NPs achieved 80-85% OZ439 dissolution, to a concentration of 430 µg/mL, within 3 h. In fed-state intestinal fluid, 95-100% OZ439 dissolution is achieved within 1 h, to a concentration of 535 µg/mL. X-ray powder diffraction and differential scanning calorimetry profiles similarly remain constant over these periods. CONCLUSIONS: The combined nanofabrication and drying process described herein, which utilizes two continuous unit operations that can be operated at scale, is an important step toward an industrially-relevant method of formulating the antimalarial OZ439 into a single-dose oral form with good stability against humidity and temperature.

A randomised, double-blind clinical phase II trial of the efficacy, safety, tolerability and pharmacokinetics of a single dose combination treatment with artefenomel and piperaquine in adults and children with uncomplicated Plasmodium falciparum malaria.

BACKGROUND: The clinical development of a single encounter treatment for uncomplicated malaria has the potential to significantly improve the effectiveness of antimalarials. Exploratory data suggested that the combination of artefenomel and piperaquine phosphate (PQP) has the potential to achieve satisfactory cure rates as a single dose therapy. The primary objective of the study was to determine whether a single dose of artefenomel (800 mg) plus PQP in ascending doses is an efficacious treatment for uncomplicated Plasmodium falciparum malaria in the 'target' population of children ≤ 5 years of age in Africa as well as Asian patients of all ages. METHODS: Patients in six African countries and in Vietnam were randomised to treatment with follow-up for 42-63 days. Efficacy, tolerability, safety and pharmacokinetics were assessed. Additional key objectives were to characterise the exposure-response relationship for polymerase chain reaction (PCR)-adjusted adequate clinical and parasitological response at day 28 post-dose (ACPR28) and to further investigate Kelch13 mutations. Patients in Africa (n = 355) and Vietnam (n = 82) were included, with 85% of the total population being children < 5 years of age. RESULTS: ACPR28 in the per protocol population (95% confidence interval) was 70.8% (61.13-79.19), 68.4% (59.13-76.66) and 78.6% (70.09-85.67) for doses of 800 mg artefenomel with 640 mg, 960 mg and 1440 mg of PQP respectively. ACPR28 was lower in Vietnamese than in African patients (66.2%; 54.55-76.62 and 74.5%; 68.81-79.68) respectively. Within the African population, efficacy was lowest in the youngest age group of ≥ 0.5 to ≤ 2 years, 52.7% (38.80-66.35). Initial parasite clearance was twice as long in Vietnam than in Africa. Within Vietnam, the frequency of the Kelch13 mutation was 70.1% and was clearly associated with parasite clearance half-life (PCt1/2). The most significant tolerability finding was vomiting (28.8%). CONCLUSIONS: In this first clinical trial evaluating a single encounter antimalarial therapy, none of the treatment arms reached the target efficacy of > 95% PCR-adjusted ACPR at day 28. Achieving very high efficacy following single dose treatment is challenging, since > 95% of the population must have sufficient concentrations to achieve cure across a range of parasite sensitivities and baseline parasitaemia levels. While challenging, the development of tools suitable for deployment as single encounter curative treatments for adults and children in Africa and to support elimination strategies remains a key development goal. TRIAL REGISTRATION: ClinicalTrials.gov, NCT02083380 . Registered on 7 March 2014.

Evaluation of the QT effect of a combination of piperaquine and a novel anti-malarial drug candidate OZ439, for the treatment of uncomplicated malaria.

AIMS: The aim was to investigate the QT effect of a single dose combination regimen of piperaquine phosphate (PQP) and a novel aromatic trioxolane, OZ439, for malaria treatment. METHODS: Exposure-response (ER) analysis was performed on data from a placebo-controlled, single dose, study with OZ439 and PQP. Fifty-nine healthy subjects aged 18 to 55 years received OZ439 alone or placebo in a first period, followed by OZ439 plus PQP or matching placebos in period 2. OZ439 and PQP doses ranged from 100-800 mg and 160-1440 mg, respectively. Twelve-lead ECG tracings and PK samples were collected serially pre- and post-dosing. RESULTS: A significant relation between plasma concentrations and placebo-corrected change from baseline QTc F (ΔΔQTc F) was demonstrated for piperaquine, but not for OZ439, with a mean slope of 0.047 ms per ng ml(-1) (90% CI 0.038, 0.057). Using an ER model that accounts for plasma concentrations of both piperaquine and OZ439, a largest mean QTc F effect of 14 ms (90% CI 10, 18 ms) and 18 ms (90% CI 14, 22 ms) was predicted at expected plasma concentrations of a single dose 800 mg OZ439 combined with PQP 960 mg (188 ng ml(-1) ) and 1440 mg (281 ng ml(-1) ), respectively, administered in the fasted state. CONCLUSIONS: Piperaquine prolongs the QTc interval in a concentration-dependent way. A single dose regimen combining 800 mg OZ439 with 960 mg or 1440 mg PQP is expected to result in lower peak piperaquine plasma concentrations compared with available 3 day PQP-artemisinin combinations and can therefore be predicted to cause less QTc prolongation.

A Proteasome Mutation Sensitizes P. falciparum Cam3.II K13C580Y Parasites to DHA and OZ439.

Artemisinin-based combination therapies (ACTs), the World Health Organization-recommended first-line therapy for uncomplicated falciparum malaria, has led to significant decreases in malaria-associated morbidity and mortality in the past two decades. Decreased therapeutic efficacy of artemisinins, the cornerstone of ACTs, is threatening the gains made against this disease. As such, novel therapeutics with uncompromised mechanisms of action are needed to combat parasite-mediated antimalarial resistance. We have previously reported the antimalarial activity of Plasmodium falciparum-specific proteasome inhibitors in conjunction with a variety of antimalarials in clinical use or in preclinical investigations and of proteasome mutants generated in response to these inhibitors. Here, we discover that despite harboring K13C580Y, which has conventionally mediated artemisinin resistance in vitro as measured by increased survival in ring-stage survival assays (RSA), the Cam3.II strain parasites of Cambodian origin that have acquired an additional mutation in the proteasome display increased susceptibility to DHA and OZ439. This discovery implicates the proteasome in peroxide susceptibilities and has favorable implications on the use of peroxide and proteasome inhibitor combination therapy for the treatment of artemisinin-resistant malaria.

Cytochrome P450-Mediated Metabolism and CYP Inhibition for the Synthetic Peroxide Antimalarial OZ439.

OZ439 is a potent synthetic ozonide evaluated for the treatment of uncomplicated malaria. The metabolite profile of OZ439 was characterized in vitro using human liver microsomes combined with LC/MS-MS, chemical derivatization, and metabolite synthesis. The primary biotransformations were monohydroxylation at the three distal carbon atoms of the spiroadamantane substructure, with minor contributions from N-oxidation of the morpholine nitrogen and deethylation cleavage of the morpholine ring. Secondary transformations resulted in the formation of dihydroxylation metabolites and metabolites containing both monohydroxylation and morpholine N-oxidation. With the exception of two minor metabolites, none of the other metabolites had appreciable antimalarial activity. Reaction phenotyping indicated that CYP3A4 is the enzyme responsible for the metabolism of OZ439, and it was found to inhibit CYP3A via both direct and mechanism-based inhibition. Elucidation of the metabolic pathways and kinetics will assist with efforts to predict potential metabolic drug-drug interactions and support physiologically based pharmacokinetic (PBPK) modeling.

Single dose treatment of malaria - current status and perspectives.

INTRODUCTION: Despite increased international efforts for control and ultimate elimination, malaria remains a major health problem. Currently, artemisinin-based combination therapies are the treatment of choice for uncomplicated malaria exhibiting high efficacy in clinical trial settings in sub-Saharan Africa. However, their administration over a three-day period is associated with important problems of treatment adherence resulting in markedly reduced effectiveness of currently recommended antimalarials under real world settings. AREAS COVERED: Antimalarial drug candidates and antimalarial drug combinations currently under advanced clinical development for the indication as single dose antimalarial therapy. Expert commentary: Several new drug candidates and combinations are currently undergoing pivotal proof-of-concept studies or clinical development programmes. The development of a single dose combination therapy would constitute a breakthrough in the control of malaria. Such an innovative treatment approach would simultaneously close the effectiveness gap of current three-day therapies and revolutionize population based interventions in the context of malaria elimination campaigns.

Antimalarial compounds in Phase II clinical development.

INTRODUCTION: Malaria is a major health problem in endemic countries and chemotherapy remains the most important tool in combating it. Treatment options are limited and essentially rely on a single drug class - the artemisinins. Efforts are ongoing to restrict the evolving threat of artemisinin resistance but declining sensitivity has been reported. Fueled by the ambitious aim of malaria eradication, novel antimalarial compounds, with improved properties, are now in the progressive phase of drug development. AREAS COVERED: Herein, the authors describe antimalarial compounds currently in Phase II clinical development and present the results of these investigations. EXPERT OPINION: Thanks to recent efforts, a number of promising antimalarial compounds are now in the pipeline. First safety data have been generated for all of these candidates, although their efficacy as antimalarials is still unclear for most of them. Of particular note are KAE609, KAF156 and DSM265, which are of chemical scaffolds new to malaria chemotherapy and would truly diversify antimalarial options. Apart from SAR97276, which also has a novel chemical scaffold that has had its development stopped, all other compounds in the pipeline belong to already known substance classes, which have been chemically modified. At this moment in time, there is not one standout compound that will revolutionize malaria treatment but several compounds that will add to its control in the future.