Francisco Javier Gamo

Quinolone-3-Diarylethers: A New Class of Antimalarial Drug

ELQ-300, an investigational drug for treating and preventing malaria, shows potent transmission-blocking activity in rodent models of malaria. Taking the Bite Out of Malaria Malaria is spread from person to person by mosquitoes that inject 8 to 10 sporozoite forms of the parasite in a single bite. The sporozoites reproduce in the liver to produce 10,000 to 30,000 merozoites before the liver schizont ruptures and parasites flood into the bloodstream where the absolute parasite burden may increase to a thousand billion (1012) circulating parasites. Some of these parasites develop into gametocytes that may be ingested by another mosquito where they progress through ookinete, oocyst, and sporozoite stages to complete the cycle. Like quinine, most antimalarial drugs in use today target only the symptomatic blood stage. The efficacy of these drugs has been compromised by resistance, and so there is a pressing need for new drugs that target multiple stages of the parasite life cycle for use in malaria treatment and prevention. Clearly, it is advantageous to strike at the liver stage where parasite numbers are low, to diminish the likelihood of selecting for a resistant mutant and before the infection has a chance to weaken the defenses of the human host. In a new study, Nilsen and colleagues describe ELQ-300, a 4(1H)-quinolone-3-diarylether, which targets the liver and blood stages, including the forms that are crucial to disease transmission (gametocytes, zygotes, and ookinetes). In mouse models of malaria, a single oral dose of 0.03 mg/kg prevented sporozoite-induced infections, whereas four daily doses of 1 mg/kg achieved complete cures of patent infections. ELQ-300 is a preclinical candidate that may be coformulated with other antimalarials to prevent and treat malaria, with the potential to aid in eradication of the disease. The goal for developing new antimalarial drugs is to find a molecule that can target multiple stages of the parasite’s life cycle, thus impacting prevention, treatment, and transmission of the disease. The 4(1H)-quinolone-3-diarylethers are selective potent inhibitors of the parasite’s mitochondrial cytochrome bc1 complex. These compounds are highly active against the human malaria parasites Plasmodium falciparum and Plasmodium vivax. They target both the liver and blood stages of the parasite as well as the forms that are crucial for disease transmission, that is, the gametocytes, the zygote, the ookinete, and the oocyst. Selected as a preclinical candidate, ELQ-300 has good oral bioavailability at efficacious doses in mice, is metabolically stable, and is highly active in blocking transmission in rodent models of malaria. Given its predicted low dose in patients and its predicted long half-life, ELQ-300 has potential as a new drug for the treatment, prevention, and, ultimately, eradication of human malaria.

Structure-guided lead optimization of triazolopyrimidine-ring substituents identifies potent Plasmodium falciparum dihydroorotate dehydrogenase inhibitors with clinical candidate potential

Drug therapy is the mainstay of antimalarial therapy, yet current drugs are threatened by the development of resistance. In an effort to identify new potential antimalarials, we have undertaken a lead optimization program around our previously identified triazolopyrimidine-based series of Plasmodium falciparum dihydroorotate dehydrogenase (PfDHODH) inhibitors. The X-ray structure of PfDHODH was used to inform the medicinal chemistry program allowing the identification of a potent and selective inhibitor (DSM265) that acts through DHODH inhibition to kill both sensitive and drug resistant strains of the parasite. This compound has similar potency to chloroquine in the humanized SCID mouse P. falciparum model, can be synthesized by a simple route, and rodent pharmacokinetic studies demonstrated it has excellent oral bioavailability, a long half-life and low clearance. These studies have identified the first candidate in the triazolopyrimidine series to meet previously established progression criteria for efficacy and ADME properties, justifying further development of this compound toward clinical candidate status.

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.

Falcipain inhibitors: optimization studies of the 2-pyrimidinecarbonitrile lead series.

Falcipain-2 and falcipain-3 are papain-family cysteine proteases of the malaria parasite Plasmodium falciparum that are responsible for host hemoglobin hydrolysis to provide amino acids for parasite protein synthesis. Different heteroarylnitrile derivatives were studied as potential falcipain inhibitors and therefore potential antiparasitic lead compounds, with the 5-substituted-2-cyanopyrimidine chemical class emerging as the most potent and promising lead series. Through a sequential lead optimization process considering the different positions present in the initial scaffold, nanomolar and subnanomolar inhibitors at falcipains 2 and 3 were identified, with activity against cultured parasites in the micromolar range. Introduction of protonable amines within lead molecules led to marked improvements of up to 1000 times in activity against cultured parasites without noteworthy alterations in other SAR tendencies. Optimized compounds presented enzymatic activities in the picomolar to low nanomolar range and antiparasitic activities in the low nanomolar range.

An Invitation to Open Innovation in Malaria Drug Discovery: 47 Quality Starting Points from the TCAMS

In 2010, GlaxoSmithKline published the structures of 13533 chemical starting points for antimalarial lead identification. By using an agglomerative structural clustering technique followed by computational filters such as antimalarial activity, physicochemical properties, and dissimilarity to known antimalarial structures, we have identified 47 starting points for lead optimization. Their structures are provided. We invite potential collaborators to work with us to discover new clinical candidates.

Pyrazoleamide compounds are potent antimalarials that target Na+ homeostasis in intraerythrocytic Plasmodium falciparum

The quest for new antimalarial drugs, especially those with novel modes of action, is essential in the face of emerging drug-resistant parasites. Here we describe a new chemical class of molecules, pyrazoleamides, with potent activity against human malaria parasites and showing remarkably rapid parasite clearance in an in vivo model. Investigations involving pyrazoleamide-resistant parasites, whole-genome sequencing and gene transfers reveal that mutations in two proteins, a calcium-dependent protein kinase (PfCDPK5) and a P-type cation-ATPase (PfATP4), are necessary to impart full resistance to these compounds. A pyrazoleamide compound causes a rapid disruption of Na+ regulation in blood-stage Plasmodium falciparum parasites. Similar effect on Na+ homeostasis was recently reported for spiroindolones, which are antimalarials of a chemical class quite distinct from pyrazoleamides. Our results reveal that disruption of Na+ homeostasis in malaria parasites is a promising mode of antimalarial action mediated by at least two distinct chemical classes.

Exploration of 4(1H)-pyridones as a novel family of potent antimalarial inhibitors of the plasmodial cytochrome bc1

A novel family of antimalarials based on the 4(1H)-pyridone scaffold is described. The compounds display potent antimalarial activity against Plasmodium falciparum in vitro and in vivo. Like atovaquone, 4(1H)-pyridones exert their antimalarial action by inhibiting selectively the electron-transport chain in P. falciparum at the cytochrome bc1 level (complex III). However, despite the similar mechanism of action, no cross-resistance with atovaquone has been found, suggesting that the binding mode of 4(1H)-pyridones might be different from that of atovaquone. The medicinal chemistry program, focused on improving potency and physicochemical properties, ultimately led to the discovery of GSK932121, which was progressed efficiently into first time in human studies. However, progression of GSK932121 was terminated when new toxicology results were obtained in the rat with a soluble phosphate prodrug of the candidate, indicating a potentially narrow therapeutic index.

A new in vivo screening paradigm to accelerate antimalarial drug discovery.

The emergence of resistance to available antimalarials requires the urgent development of new medicines. The recent disclosure of several thousand compounds active in vitro against the erythrocyte stage of Plasmodium falciparum has been a major breakthrough, though converting these hits into new medicines challenges current strategies. A new in vivo screening concept was evaluated as a strategy to increase the speed and efficiency of drug discovery projects in malaria. The new in vivo screening concept was developed based on human disease parameters, i.e. parasitemia in the peripheral blood of patients on hospital admission and parasite reduction ratio (PRR), which were allometrically down-scaled into P. berghei-infected mice. Mice with an initial parasitemia (P0) of 1.5% were treated orally for two consecutive days and parasitemia measured 24 h after the second dose. The assay was optimized for detection of compounds able to stop parasite replication (PRR = 1) or induce parasite clearance (PRR >1) with statistical power >99% using only two mice per experimental group. In the P. berghei in vivo screening assay, the PRR of a set of eleven antimalarials with different mechanisms of action correlated with human-equivalent data. Subsequently, 590 compounds from the Tres Cantos Antimalarial Set with activity in vitro against P. falciparum were tested at 50 mg/kg (orally) in an assay format that allowed the evaluation of hundreds of compounds per month. The rate of compounds with detectable efficacy was 11.2% and about one third of active compounds showed in vivo efficacy comparable with the most potent antimalarials used clinically. High-throughput, high-content in vivo screening could rapidly select new compounds, dramatically speeding up the discovery of new antimalarial medicines. A global multilateral collaborative project aimed at screening the significant chemical diversity within the antimalarial in vitro hits described in the literature is a feasible task.

Antimalarial efficacy of MMV390048, an inhibitor of Plasmodium phosphatidylinositol 4-kinase

MMV390048, a member of a new class of inhibitors of the Plasmodium phosphatidylinositol 4-kinase, shows potential for both treatment and prophylaxis. A new antimalarial in the armamentarium Paquet et al. screened a small-molecule library against the human malaria parasite, Plasmodium falciparum, and identified the 2-aminopyridine chemical class with potent activity. The optimized compound from this class, MMV390048, was active against multiple parasite life cycle stages, in both the mammalian host and the mosquito vector, and also killed drug-resistant parasites. MMV390048 killed the malaria parasite by blocking the parasite’s phosphatidylinositol 4-kinase (PI4K) and was able to protect monkeys from malaria infection. MMV390048 has potential as a new antimalarial drug that may contribute to global malaria eradication efforts. As part of the global effort toward malaria eradication, phenotypic whole-cell screening revealed the 2-aminopyridine class of small molecules as a good starting point to develop new antimalarial drugs. Stemming from this series, we found that the derivative, MMV390048, lacked cross-resistance with current drugs used to treat malaria. This compound was efficacious against all Plasmodium life cycle stages, apart from late hypnozoites in the liver. Efficacy was shown in the humanized Plasmodium falciparum mouse model, and modest reductions in mouse-to-mouse transmission were achieved in the Plasmodium berghei mouse model. Experiments in monkeys revealed the ability of MMV390048 to be used for full chemoprotection. Although MMV390048 was not able to eliminate liver hypnozoites, it delayed relapse in a Plasmodium cynomolgi monkey model. Both genomic and chemoproteomic studies identified a kinase of the Plasmodium parasite, phosphatidylinositol 4-kinase, as the molecular target of MMV390048. The ability of MMV390048 to block all life cycle stages of the malaria parasite suggests that this compound should be further developed and may contribute to malaria control and eradication as part of a single-dose combination treatment.

In Vitro Resistance Selections for Plasmodium Falciparum Dihydroorotate Dehydrogenase Inhibitors Give Mutants with Multiple Point Mutations in the Drug-Binding Site and Altered Growth.

Background: Inhibiting PfDHODH kills malaria parasites, but the potential for drug resistance is unknown. Results: Selections gave several categories of resistance mutations. Several mutants were hypersensitive to other drugs. Conclusion: Resistance to PfDHODH inhibitors is largely though mutations in or amplification of the target gene, PfDHODH. Significance: Resistance to PfDHODH inhibitors is possible but often increases sensitivity to other compounds. Malaria is a preventable and treatable disease; yet half of the worlds population lives at risk of infection, and an estimated 660,000 people die of malaria-related causes every year. Rising drug resistance threatens to make malaria untreatable, necessitating both the discovery of new antimalarial agents and the development of strategies to identify and suppress the emergence and spread of drug resistance. We focused on in-development dihydroorotate dehydrogenase (DHODH) inhibitors. Characterizing resistance pathways for antimalarial agents not yet in clinical use will increase our understanding of the potential for resistance. We identified resistance mechanisms of Plasmodium falciparum (Pf) DHODH inhibitors via in vitro resistance selections. We found 11 point mutations in the PfDHODH target. Target gene amplification and unknown mechanisms also contributed to resistance, albeit to a lesser extent. These mutant parasites were often hypersensitive to other PfDHODH inhibitors, which immediately suggested a novel combination therapy approach to preventing resistance. Indeed, a combination of wild-type and mutant-type selective inhibitors led to resistance far less often than either drug alone. The effects of point mutations in PfDHODH were corroborated with purified recombinant wild-type and mutant-type PfDHODH proteins, which showed the same trends in drug response as the cognate cell lines. Comparative growth assays demonstrated that two mutant parasites grew less robustly than their wild-type parent, and the purified protein of those mutants showed a decrease in catalytic efficiency, thereby suggesting a reason for the diminished growth rate. Co-crystallography of PfDHODH with three inhibitors suggested that hydrophobic interactions are important for drug binding and selectivity.