Alternative oxidase inhibitors: Mitochondrion‐targeting as a strategy for new drugs against pathogenic parasites and fungi

Abstract

The alternative oxidase (AOX) is a ubiquitous terminal oxidase of plants and many fungi, catalyzing the four‐electron reduction of oxygen to water alongside the cytochrome‐based electron transfer chain. Unlike the classical electron transfer chain, however, the activity of AOX does not generate adenosine triphosphate but has functions such as thermogenesis and stress response. As it lacks a mammalian counterpart, it has been investigated intensely in pathogenic fungi. However, it is in African trypanosomes, which lack cytochrome‐based respiration in their infective stages, that trypanosome alternative oxidase (TAO) plays the central and essential role in their energy metabolism. TAO was validated as a drug target decades ago and among the first inhibitors to be identified was salicylhydroxamic acid (SHAM), which produced the expected trypanocidal effects, especially when potentiated by coadministration with glycerol to inhibit anaerobic energy metabolism as well. However, the efficacy of this combination was too low to be of practical clinical use. The antibiotic ascofuranone (AF) proved a much stronger TAO inhibitor and was able to cure Trypanosoma vivax infections in mice without glycerol and at much lower doses, providing an important proof of concept milestone. Systematic efforts to improve the SHAM and AF scaffolds, aided with the elucidation of the TAO crystal structure, provided detailed structure‐activity relationship information and reinvigorated the drug discovery effort. Recently, the coupling of mitochondrion‐targeting lipophilic cations to TAO inhibitors has dramatically improved drug targeting and trypanocidal activity while retaining target protein potency. These developments appear to have finally signposted the way to preclinical development of TAO inhibitors.

Discovery of trypanocidal coumarins with dual inhibition of both the glycerol kinase and alternative oxidase of Trypanosoma brucei brucei

“…This difference may explain the difference in the kinetic inhibitory mechanism for AF, which has previously been reported to be a mixed-type inhibitor of TAO (27). Importantly, this report is an additional contribution to our recent efforts to find more potent inhibitors targeting TAO with the overall aim of contributing toward the design of new trypanocides (18,(51)(52)(53).…”

The cytochrome bc1 complex as an antipathogenic target

“…It must also be noted that the native AOX activity is important for maintaining redox (and metabolite) homeostasis in plants, and inhibition of this enzyme may be deleterious under osmotic and light stress conditions [83,84]. Nevertheless, the intriguing possibility exists of the development of dual fungal cyt bc 1 /AOX inhibitors, as compounds such as the quinoline aurachins, produced by the myxobacterium Stigmatella aurantiaca , are inhibitors of both enzymes [85–88]. We note also the potential for complex I/cyt bc 1 inhibition for fungicidal activity as Δp would be expected to be severely diminished under such circumstances, regardless of the activity of AOX.…”

Discovery of Sustainable Drugs for Neglected Tropical Diseases: Cashew Nut Shell Liquid (CNSL)‐Based Hybrids Target Mitochondrial Function and ATP Production in Trypanosoma brucei

“…All this does not exclude at arget insidet he mitochondrion, but it is not (principally) TAOo rg lycerol-3-phosphate dehydrogenase( G3PDH), which together enable the re-oxidationo fg lycolysis-produced NAD + via the cyanide-resistant, non-protonmotive oxidation of ubiquinol by TAO. [41] This is because the activity of the hybrids was not enhanced in the presence of glycerol. The target linked to rapid ATPd epletion could also be in glycosomes but we saw no evidence of glycosomal damagei nt he TEM images, nor consistenti nhibition of GAPDH.…”

“…However, ATP depletion precedes mitochondrial abnormalities and the depolarization of Ψ m can be observed within 30 min, and we propose that the hybrids, as intended, exert more than one trypanocidal effect, consistent with the complex return of the metabolomics experiment, (although it was not possible to derive any clear conclusion from the metabolomics data, at this point). All this does not exclude a target inside the mitochondrion, but it is not (principally) TAO or glycerol‐3‐phosphate dehydrogenase (G3PDH), which together enable the re‐oxidation of glycolysis‐produced NAD + via the cyanide‐resistant, non‐protonmotive oxidation of ubiquinol by TAO . This is because the activity of the hybrids was not enhanced in the presence of glycerol.…”