Abstract
New antifungal drugs are urgently needed due to the currently limited selection, the emergence of drug resistance, and the toxicity of several commonly used drugs. To identify drug leads, we screened small molecules using a Saccharomyces cerevisiae reporter bioassay in which S. cerevisiae heterologously expresses Hik1, a group III hybrid histidine kinase (HHK) from Magnaporthe grisea. Group III HHKs are integral in fungal cell physiology, and highly conserved throughout this kingdom; they are absent in mammals, making them an attractive drug target. Our screen identified compounds 13 and 33, which showed robust activity against numerous fungal genera including Candida spp., Cryptococcus spp. and molds such as Aspergillus fumigatus and Rhizopus oryzae. Drug-resistant Candida albicans from patients were also highly susceptible to compounds 13 and 33. While the compounds do not act directly on HHKs, microarray analysis showed that compound 13 induced transcripts associated with oxidative stress, and compound 33, transcripts linked with heavy metal stress. Both compounds were highly active against C. albicans biofilm, in vitro and in vivo, and exerted synergy with fluconazole, which was inactive alone. Thus, we identified potent, broad-spectrum antifungal drug leads from a small molecule screen using a high-throughput, S. cerevisiae reporter bioassay.
Highlights
Over the past twenty years, the incidence of fungal infections has risen sharply as advances in medicine have increased the number of immunocompromised patients [1]
Fludioxonil and 314 (1.6%) of the small molecules inhibited S. cerevisiae growth by more than 50% (Table 1)
In the secondary screen to assess whether the small molecules acted in a Hik1dependent manner, 57 of the small molecules inhibited the growth of the reporter strain by more than 50%, and the parental S. cerevisiae strain by less than 10% (Table 1)
Summary
Over the past twenty years, the incidence of fungal infections has risen sharply as advances in medicine have increased the number of immunocompromised patients [1]. The antifungal drug armamentarium has not kept pace. Current antifungal therapeutics are plagued with problems including limited spectrum of activity, the emergence of resistant strains, and patient toxicity [2]. New drugs are required to meet the growing need for antifungal therapy. The identification of novel antifungals is hindered by the limited number of drug targets that are unique to fungi due to the close evolutionary relationship between fungi and mammals. Hybrid histidine kinases (HHKs) are an appealing antifungal drug target due to their central role in fungal physiology, conservation throughout the fungal kingdom, and absence in mammals. HHKs regulate two-component signaling pathways in response to a variety of environmental stimuli, including osmotic, nitrosative, and oxidative and stress in bacteria and fungi [3]
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