Generation of quinolone antimalarials targeting the Plasmodium falciparum mitochondrial respiratory chain for the treatment and prophylaxis of malaria

Abstract

There is an urgent need for new antimalarial drugs with novel mechanisms of action to deliver effective control and eradication programs. Parasite resistance to all existing antimalarial classes, including the artemisinins, has been reported during their clinical use. A failure to generate new antimalarials with novel mechanisms of action that circumvent the current resistance challenges will contribute to a resurgence in the disease which would represent a global health emergency. Here we present a unique generation of quinolone lead antimalarials with a dual mechanism of action against two respiratory enzymes, NADH:ubiquinone oxidoreductase (Plasmodium falciparum NDH2) and cytochrome bc 1 . Inhibitor specificity for the two enzymes can be controlled subtly by manipulation of the privileged quinolone core at the 2 or 3 position. Inhibitors display potent (nanomolar) activity against both parasite enzymes and against multidrug-resistant P. falciparum parasites as evidenced by rapid and selective depolarization of the parasite mitochondrial membrane potential, leading to a disruption of pyrimidine metabolism and parasite death. Several analogs also display activity against liver-stage parasites (Plasmodium cynomolgi) as well as transmission-blocking properties. Lead optimized molecules also display potent oral antimalarial activity in the Plasmodium berghei mouse malaria model associated with favorable pharmacokinetic features that are aligned with a single-dose treatment. The ease and low cost of synthesis of these inhibitors fulfill the target product profile for the generation of a potent, safe, and inexpensive drug with the potential for eventual clinical deployment in the control and eradication of falciparum malaria.T he discovery of atovaquone 20 years ago validated the malaria parasite's mitochondrial electron transport chain (ETC) as an exploitable drug target. Atovaquone targets the ETC at the level of the bc 1 complex (1), with inhibition preventing proton pumping, resulting in a loss of mitochondrial membrane potential (2) and eventual organelle dysfunction, an important function of which is to provide intermediates for pyrimidine synthesis (3, 4). The bc 1 complex requires reducing equivalents provided by ubiquinol, which in turn is generated by membrane-bound dehydrogenases upstream in the ETC that catalyze redox reactions by reducing ubiquinone. The parasite lacks the canonical protonmotive NADH dehydrogenase (Complex I) but instead harbors a bacterial-like type II NADH:ubiquinone oxidoreductase, Plasmodium falciparum NDH2 (PfNDH2) (5). Based on these key observations, we undertook a drug-discovery initiative to develop costeffective inhibitors capable of inhibiting PfNDH2 with the goal of providing antimalarials that overcome the limitations of the expensive atovaquone. Although our initial drug-discovery efforts were focused on optimization of activity versus PfNDH2, we found, during hit-to-lead development, that optimized structures with single-digit nanomolar activity versus the primary target ...

Metabolomics: A Tool Ahead for Understanding Molecular Mechanisms of Drugs and Diseases

“…For the antimalarial, atovaquone, its inhibition of cytochrome bc (1), results in loss of mitochondrial membrane potential, which inhibits dihydroorotate dehydrogenase, thus inhibiting pyrimidine synthesis. Metabolomics confirmed this by showing accumulation of dihydroorotate and carbamoyl-L-aspartate, its precursor [21].…”

Metabolomic-Based Strategies for Anti-Parasite Drug Discovery

“…81 Atovaquone and its quinolone analogue CK-2-68 were shown to inhibit two enzymes in the bc 1 complex: NADH:ubiquinone oxidoreductase and cytochrome bc 1 . Using an MRM-targeted metabolomics approach looking for 35 metabolites, it was shown that following atovaquone-induced membrane depolarization, proton pumping was disrupted but dihydroorotate dehydrogenase activity is also diminished, leading to a depletion in orotate levels 37 (Fig. 4).…”

“…The rise of drug resistance in every class of antimalarial 37 is compromising antimalaria therapy, and combination therapies are increasingly preferred. 80 More drugs with novel MOAs are required that will circumvent the current resistance mechanisms.…”

“…The lack of orotate then affects pyrimidine synthesis. 37 Despite the fact that atovaquone acts against more than one target, resistance has been relatively simple to select. 82,83 Both mutations in the catalytic domain of the bc1 complex 84 and dihydroorotate dehydrogenase overexpression 85 have been identified as mechanisms to reduce sensitivity to atovaquone.…”

4(1H)-Quinolones with Liver Stage Activity against Plasmodium berghei

“…Salzer et al (11) found that endochin had liver and blood stage activity in avian malaria models; however, due to the lack of appropriate preclinical models, additional studies were not conducted (11). The compounds related to endochin also have activity against Plasmodium cynomolgi liver stages (12), and more recently, studies have shown these compounds to be potent in vitro and in vivo against Plasmodium falciparum and rodent malaria blood stage parasites (13)(14)(15)(16)(17)(18) …”