Artemisinin—the next generation: Efficacies of artemisone against the malaria parasite are substantially greater than those of the current artemisinin “gold standard”, artesunate. Also, in contrast to most current artemisinins it displays low lipophilicity and negligible neuro‐ and cytotoxicity in in vitro and in vivo assays. Thus, the drug offers promise for use in artemisinin‐based combination therapy.

Malarial dihydrofolate reductase (DHFR) is the target of antifolate antimalarial drugs such as pyrimethamine and cycloguanil, the clinical efficacy of which have been compromised by resistance arising through mutations at various sites on the enzyme. Here, we describe the use of cocrystal structures with inhibitors and substrates, along with efficacy and pharmacokinetic profiling for the design, characterization, and preclinical development of a selective, highly efficacious, and orally available antimalarial drug candidate that potently inhibits both wild-type and clinically relevant mutated forms of Plasmodium falciparum (Pf) DHFR. Important structural characteristics of P218 include pyrimidine side-chain flexibility and a carboxylate group that makes charge-mediated hydrogen bonds with conserved Arg122 (PfDHFR-TS amino acid numbering). An analogous interaction of P218 with human DHFR is disfavored because of three species-dependent amino acid substitutions in the vicinity of the conserved Arg. Thus, P218 binds to the active site of PfDHFR in a substantially different fashion from the human enzyme, which is the basis for its high selectivity. Unlike pyrimethamine, P218 binds both wild-type and mutant PfDHFR in a slow-on/slow-off tight-binding mode, which prolongs the target residence time. P218, when bound to PfDHFR-TS, resides almost entirely within the envelope mapped out by the dihydrofolate substrate, which may make it less susceptible to resistance mutations. The high in vivo efficacy in a SCID mouse model of P. falciparum malaria, good oral bioavailability, favorable enzyme selectivity, and good safety characteristics of P218 make it a potential candidate for further development.

Plasmodium falciparum, the causative agent of malaria, relies extensively on glycolysis coupled with homolactic fermentation during its blood-borne stages for energy production. Selective inhibitors of the parasite lactate dehydrogenase (LDH), central to NAD ؉ regeneration, therefore potentially provide a route to new antimalarial drugs directed against a novel molecular target. A series of heterocyclic, azole-based compounds are described that preferentially inhibit P. falciparum LDH at sub-micromolar concentrations, typically at concentrations about 100-fold lower than required for human lactate dehydrogenase inhibition. Crystal structures show these competitive inhibitors form a network of interactions with amino acids within the active site of the enzyme, stacking alongside the nicotinamide ring of the NAD ؉ cofactor. These compounds display modest activity against parasitized erythrocytes, including parasite strains with known resistance to existing anti-malarials and against Plasmodium berghei in BALB/c mice. Initial toxicity data suggest the azole derivatives have generally low cytotoxicity, and preliminary pharmocokinetic data show favorable bioavailability and circulation times. These encouraging results suggest that further enhancement of these structures may yield candidates suitable for consideration as new therapeutics for the treatment of malaria. In combination these studies also provide strong support for the validity of targeting the Plasmodium glycolytic pathway and, in particular, LDH in the search for novel anti-malarials.Plasmodium parasites are believed to lack a functional Krebs (citric acid) cycle for at least part of their life cycle and hence rely extensively on ATP generation via the anaerobic fermentation of glucose (see Ref. 1 for review). The energy requirement of the parasitized erythrocyte is such that utilization of glucose is up to 100 times greater than in nonparasitized erythrocytes (2, 3), and virtually all glucose can be accounted for by production of lactate (2). Lactate dehydrogenase (LDH), 1 the last enzyme in the glycolytic pathway in Plasmodium falciparum, is a 2-hydroxy acid oxidoreductase that converts pyruvate to lactate and simultaneously the conversion of NADH to NAD ϩ . As a constant supply of NADH is a prerequisite for glycolysis, and LDH acts as the primary source in Plasmodium for the regeneration of NADH from NAD ϩ , inhibition of LDH is expected to stop production of ATP, with subsequent P. falciparum cell death. Any compound that blocks the LDH enzyme is a potentially potent antimalarial with a different mode of action to existing drugs. As such, P. falciparum lactate dehydrogenase (pfLDH) has been suggested as a drug target by several authors (4 -6). One well recognized difficulty is that the drug must potently inhibit pfLDH yet show much less activity against the three human LDH (hsLDH) isoforms.A comparison of the crystal structures of both P. falciparum and human LDH (7,8) shows the following two key differences: namely positioning of the NADH factor, re...