The Anti-helminthic Compound Mebendazole Has Multiple Antifungal Effects against Cryptococcus neoformans

Abstract

Cryptococcus neoformans is the most lethal pathogen of the central nervous system. The gold standard treatment of cryptococcosis, a combination of amphotericin B with 5-fluorocytosine, involves broad toxicity, high costs, low efficacy, and limited worldwide availability. Although the need for new antifungals is clear, drug research and development (R&D) is costly and time-consuming. Thus, drug repurposing is an alternative to R&D and to the currently available tools for treating fungal diseases. Here we screened a collection of compounds approved for use in humans seeking for those with anti-cryptococcal activity. We found that benzimidazoles consist of a broad class of chemicals inhibiting C. neoformans growth. Mebendazole and fenbendazole were the most efficient antifungals showing in vitro fungicidal activity. Since previous studies showed that mebendazole reaches the brain in biologically active concentrations, this compound was selected for further studies. Mebendazole showed antifungal activity against phagocytized C. neoformans, affected cryptococcal biofilms profoundly and caused marked morphological alterations in C. neoformans, including reduction of capsular dimensions. Amphotericin B and mebendazole had additive anti-cryptococcal effects. Mebendazole was also active against the C. neoformans sibling species, C. gattii. To further characterize the effects of the drug a random C. gattii mutant library was screened and indicated that the antifungal activity of mebendazole requires previously unknown cryptococcal targets. Our results indicate that mebendazole is as a promising prototype for the future development of anti-cryptococcal drugs.

Fenbendazole controlsin vitrogrowth, virulence potential and animal infection in theCryptococcusmodel

“…We asked whether the mechanism of anti-cryptococcal activity of fenbendazole was similar to what was previously described for C. gattii (16) or if it was related to its well characterized anthelmintic effect (19–21). Mutant strains of C. gattii lacking expression of the Aim25 scramblase or the nucleolar protein Nop16 were resistant to mebendazole (16), suggesting that these proteins are potential targets for the antifungal activity of benzimidazoles. To investigate the mechanism of action of fenbendazole, we first evaluated its antifungal activity against mutant strains of C. gattii lacking the AIM25 or NOP16 genes and observed that both strains and wild type cells were similarly sensitive to this benzimidazole (Figures 2A and B).…”

“…The in vitro anti-cryptococcal activity of benzimidazoles has been consistently reported (13–16). A general analysis of these reports on the anti-cryptococcal activity of benzimidazoles indicated that, among all members of this drug family, fenbendazole required the lowest MIC against C. neoformans and C. gattii , in comparison to other benzimidazoles (13–16). In addition, a detailed study on the toxicity of fenbendazole suggested high levels of safety in humans (17, 18).…”

“…In mice, orally administered flubendazole resulted in an expressive reduction in fungal burden in comparison with controls, but in the rabbit model, there were no quantifiable drug concentrations or antifungal activity in the cerebrospinal fluid or brain (15). Mebendazole, another member of the family of benzimidazoles, also showed antifungal activity against C. neoformans , including phagocytized yeast cells and cryptococcal biofilms (16).…”

“…The anti-cryptococcal effects of other benzimidazoles have been superficially examined. In comparison to other benzimidazoles, fenbendazole was the most efficient compound showing in vitro fungicidal activity (16), but mechanistic approaches and in vivo activity of this compound were not evaluated. Fenbendazole has been licensed worldwide for the treatment and control of helminth infections in food and non-food producing animals for more than 30 years and its safety is well established.…”

“…Based on the anti-cryptococcal effects of fenbendazole (16) and negligible toxicity to humans and animals (17, 18), we evaluated the therapeutic potential of this benzimidazole against pathogenic cryptococci. Our results demonstrated that fenbendazole was inhibitory against several strains of C. neoformans and C. gattii .…”