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Tackling fungal resistance with new ketoconazole derivatives targeting plasma membrane sphingolipids

Publication . Bento-Oliveira, Andreia; Almeida, Rodrigo Freire Martins de

The search for new antifungal drugs is a relevant and timely subject due to fungal resistance to antimicrobial agents which is an emerging public health threat. Recently developed diphenylphosphane derivatives of ketoconazole exhibited potential as antifungal therapeutic agents. The mode of action of these compounds is not yet fully uncovered, although evidence suggests that they might interact with sphingolipids in the fungal membrane, which are major component of sphingolipid-enriched gel domains (SLEDs). Moreover, it was shown that the antifungal polyene nystatin has pore-forming activity in highly ordered gel membranes that do not contain sterols, contradicting the classical ergosterol-dependent mode of action. Hence, SLEDs which are not found in animal membranes are promising targets for antifungal drugs and are possibly involved in certain antifungal drug action and resistance mechanisms. The objectives of this thesis were 1) to explore the biophysical properties of the plasma membrane of the budding yeast Saccharomyces cerevisiae focusing on SLEDs, and 2) to uncover the preferential interaction of ketoconazole and diphenylphosphane derivatives, and nystatin with membranes containing specific fungal lipids and distinct biophysical properties as compared to mammalian ones, correlating this interaction with the drugs activity. Fluorescence spectroscopy was used together with an array of membrane systems, from intact cells in suspension to simpler systems such as isolated plasma membrane and liposomes prepared from S. cerevisiae lipid extracts or synthetic lipids. For ketoconazole derivatives, this work also allowed to unravel how structural differences influence their interaction (partition and relative distribution) with membranes and how that could correlate with their biological activity through direct interference on the permeability of the plasma membrane. Overall, the results presented herein point to SLEDs as promising targets for antifungal agents, with potential to uncover new approaches to fight antifungal drug resistance.

2025-04-14Doctoral thesis

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