Identification of Inhibitors of Fungal Fatty Acid Biosynthesis

[Image: see text] Fungal fatty acid (FA) synthase and desaturase enzymes are essential for the growth and virulence of human fungal pathogens. These enzymes are structurally distinct from their mammalian counterparts, making them attractive targets for antifungal development. However, there has been little progress in identifying chemotypes that target fungal FA biosynthesis. To accomplish this, we applied a whole-cell-based method known as Target Abundance-based FItness Screening using Candida albicans. Strains with varying levels of FA synthase or desaturase expression were grown in competition to screen a custom small-molecule library. Hit compounds were defined as preferentially inhibiting the growth of the low target-expressing strains. Dose–response experiments confirmed that 16 hits (11 with an acyl hydrazide core) differentially inhibited the growth of strains with an altered desaturase expression, indicating a specific chemical–target interaction. Exogenous unsaturated FAs restored C. albicans growth in the presence of inhibitory concentrations of the most potent acyl hydrazides, further supporting the primary mechanism being inhibition of FA desaturase. A systematic analysis of the structure–activity relationship confirmed the acyl hydrazide core as essential for inhibitory activity. This collection demonstrated broad-spectrum activity against Candida auris and mucormycetes and retained the activity against azole-resistant candida isolates. Finally, a preliminary analysis of toxicity to mammalian cells identified potential lead compounds with desirable selectivities. Collectively, these results establish a scaffold that targets fungal FA biosynthesis with a potential for development into novel therapeutics.