Fig 2.

Identification of antifungal compounds from a purinome-focused chemical library. (A) Determination of the Z′ factor for microtiter plate-based assays of growth inhibition by measurements of culture turbidity (absorbance) and RFP fluorescence. OSU76 yeast cells were exposed to AmB at concentrations from 0.003 μM to 1.7 μM, and growth was measured at day 4 and normalized to growth of yeast in the absence of AmB. Values represent the means ± standard deviations of replicate cultures (n = 3). Data are representative of ≥3 independent experiments. (B) Overview of hit compound identification. A total of 3,600 compounds were screened for their ability to inhibit Histoplasma yeast growth by measurement of RFP fluorescence and absorbance at 595 nm. Compounds were initially screened at 10 μM. Dose-response tests further narrowed the 59 hits to 28, which were effective at concentrations of <5 μM. Macrophage (Mϕ) toxicity was determined by inhibition of P388D1-lacZ macrophage cells and identified 16 compounds that were nontoxic to host cells at 5 μM. Overall selectivity (yeast IC₅₀ determined by the absorbance at 595 nm relative to the P388D1-lacZ IC₅₀) was used to identify the top 7 hits with selectivity indices of >5. (C) Summary of IC₅₀ and selectivity data for hit compounds. All concentrations are in μM.