Background Ebselen, an organoselenium compound and a clinically safe molecule has been reported to possess potent antifungal activity, but its antifungal mechanism of action and in vivo antifungal activity remains unclear. Methods The antifungal effect of ebselen was tested against Candida albicans, C. glabrata, C. tropicalis, C. parapsilosis, Cryptococcus neoformans, and C. gattii clinical isolates. Chemogenomic profiling and biochemical assays were employed to identify the antifungal target of ebselen. Ebselen’s antifungal activity in vivo was investigated in a Caenorhabditis elegans animal model. Results Ebselen exhibits potent antifungal activity against both Candida spp. and Cryptococcus spp, at concentrations ranging from 0.5 – 2 μg/ml. Ebselen rapidly eradicates a high fungal inoculum within two hours of treatment. Investigation of the drug’s antifungal mechanism of action indicates that ebselen depletes intracellular glutathione (GSH) levels, leading to increased production of reactive oxygen species (ROS), and thereby disturbs the redox homeostasis in fungal cells. Examination of ebselen’s in vivo antifungal activity in two Caenorhabditis elegans models of infection demonstrate that ebselen is superior to conventional antifungal drugs (fluconazole, flucytosine and amphotericin) in reducing Candida and Cryptococcus fungal load. Conclusion Ebselen possesses potent antifungal activity against clinically relevant isolates of both Candida and Cryptococcus by regulating GSH and ROS production. The potent in vivo antifungal activity of ebselen supports further investigation for repurposing it for use as an antifungal agent. General significance The present study shows that ebselen targets glutathione and also support that glutathione as a potential target for antifungal drug development.

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The limited therapeutic options and the recent emergence of multidrug-resistant Candida species present a significant challenge to human medicine and underscore the need for novel therapeutic approaches. Drug repurposing appears as a promising tool to augment the activity of current azole antifungals, especially against multidrug-resistant C. auris. In this study, we evaluated the fluconazole chemosensitization activity of 1547 FDA-approved drugs and clinical molecules against azole-resistant C. auris. This led to the discovery that lopinavir, an HIV protease inhibitor, is a potent agent capable of sensitizing C. auris to the effect of azole antifungals. At a therapeutically achievable concentration, lopinavir exhibited potent synergistic interactions with azole drugs, particularly with itraconazole against C. auris (ΣFICI ranged from 0.04 to 0.09). Additionally, the lopinavir/itraconazole combination enhanced the survival rate of C. auris-infected Caenorhabditis elegans by 90% and reduced the fungal burden in infected nematodes by 88.5% (P < 0.05), relative to the untreated control. Furthermore, lopinavir enhanced the antifungal activity of itraconazole against other medically important Candida species including C. albicans, C. tropicalis, C. krusei, and C. parapsilosis. Comparative transcriptomic profiling and mechanistic studies revealed lopinavir was able to significantly interfere with the glucose permeation and ATP synthesis. This compromised the efflux ability of C. auris and consequently enhanced the susceptibility to azole drugs, as demonstrated by Nile red efflux assays. Altogether, these findings present lopinavir as a novel, potent, and broad-spectrum azole chemosensitizing agent that warrants further investigation against recalcitrant Candida infections.