Real-Time Imaging of the Azole Class of Antifungal Drugs in Live Candida Cells
Open accessAbstract
Azoles are the most commonly used class of antifungal drugs, yet where they localize within fungal cells and how they are imported remain poorly understood. Azole antifungals target lanosterol 14α-demethylase, a cytochrome P450, encoded by ERG11 in Candida albicans, the most prevalent fungal pathogen. We report the synthesis of fluorescent probes that permit visualization of antifungal azoles within live cells. Probe 1 is a dansyl dye-conjugated azole, and probe 2 is a Cy5-conjugated azole. Docking computations indicated that each of the probes can occupy the active site of the target cytochrome P450. Like the azole drug fluconazole, probe 1 is not effective against a mutant that lacks the target cytochrome P450. In contrast, the azole drug ketoconazole and probe 2 retained some antifungal activity against mutants lacking the target cytochrome P450, implying that both act against more than one target. Both fluorescent azole probes colocalized with the mitochondria, as determined by fluorescence microscopy with MitoTracker dye. Thus, these fluorescent probes are useful molecular tools that can lead to detailed information about the activity and localization of the important azole class of antifungal drugs.
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Drug uptake and efflux appear to contribute at least partially to tolerance, based on studies with fluorescent azole probes and strains with similar MIC levels but different tolerance levels.
“In addition to affecting resistance, drug uptake and efflux appears to contribute, at least partially, to tolerance9, based on studies with fluorescent azole probes and strains with similar MIC levels but different tolerance levels (measured as FoG and SMG, see Box ##)89,90”
From Drug resistance and tolerance in fungi. (2020) · cites this paper as [89]
The Introduction describes fluorescent azole probes for studying drug uptake, and Results and Discussion reports that increased efflux is associated with fluconazole resistance in one Candida strain. The paper does not report that uptake or efflux contributes to tolerance, or compare strains with similar MICs but different FoG or SMG tolerance levels. No page numbers are available in our copy. (LLM, conf=0.86)
Experimental — accuracy not yet measuredAzole drugs accumulate in mitochondria, at least upon contact with Candida albicans cells.
“Recent studies have shown that azole drugs accumulate in mitochondria, at least upon contact with C. albicans cells [43].”
From In Candida glabrata, ERMES Component GEM1 Controls Mitochondrial Morphology, mtROS, and Drug Efflux Pump Expression, Resulting in Azole Susceptibility (2023) · cites this paper as [43]
The Conclusion reports that two fluorescent azole probes accumulated in mitochondria in C. albicans. The Results and Discussion proposes that many or all azole antifungals may do so, but does not establish mitochondrial accumulation as a finding for azole drugs generally. (LLM, conf=0.87)
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- [45]Friesner RA Banks JL Murphy RB Halgren TA Klicic JJ Mainz DT Glide: a new approach for rapid, accurate docking and scoring. 1. Method and assessment of docking accuracy J Med Chem 2004 47 1750 1759 15027865
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- Authors
- Benhamou, R. I., Bibi, M., Steinbuch, K. B., Engel, H., Levin, M., Roichman, Y., Berman, J., & Fridman, M.
- Affiliations
- Tel Aviv University (IL)
- Corresponding author
- Berman, Judith
- Fridman, Micha
- ORCID
- Venue
- ACS Chemical Biology
- Publisher
- American Chemical Society (ACS)
- Published
- 2017-05-12
- Volume
- 12
- Issue
- 7
- Pages
- 1769-1777
- ISSN
- 1554-8929 (ISSN-L)
- 1554-8937
- PMC
- PMC7030953
- OpenAlex
- W2610314569
- Cited by
- 71
- References
- 54
- Citations by year
- 2026: 3 · 2025: 8 · 2024: 7 · 2023: 10 · 2022: 8 · 2021: 10 · 2020: 7 · 2019: 8 · 2018: 9 · 2017: 1
- FWCI
- 5.26
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- ACS Chemical Biology (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- Funders
- United States-Israel Binational Science Foundation (2014314)
- Israel Science Foundation (6/14)
- FP7 Ideas: European Research Council (340087)
- Topics
- Health Sciences › Medicine › Infectious Diseases › Antifungal resistance and susceptibility
- Physical Sciences › Chemistry › Spectroscopy › Molecular Sensors and Ion Detection
- Physical Sciences › Chemistry › Organic Chemistry › Click Chemistry and Applications
- Keywords
- azole antifungals, Candida albicans, ERG11, lanosterol 14α-demethylase, cytochrome P450, fluorescent probes, Cy5, drug localization, drug importation, mitochondria, fluconazole, ketoconazole, drug target specificity, live-cell imaging
- MeSH terms
- Animals, Antifungal Agents, Azoles, Candida, Cells, Cultured, Cytochrome P-450 Enzyme System, Fluorescent Dyes, Microscopy, Fluorescence, Mutation, Catalytic Domain, Molecular Docking Simulation
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