Combining fluconazole with benzo[a]phenoxazine derivatives as a promising strategy against fluconazole-resistant candida species

The rise in non-albicans Candida species, exhibiting unpredictable antifungal resistance, complicates treatment and contributes to the growing threat of invasive, life-threatening infections. This study evaluates the antifungal activity of four benzo[a]phenoxazine derivatives (C34, C35, A42, and A44) against 14 Candida strains following EUCAST standards. Fluconazole interactions are analysed through fractional inhibitory concentration index (FICI) calculation and response surface analysis based on the Bliss model. Macrophage-like J774A.1 cells are used to assess Candida killing in the presence of synergistic compounds. The MIC values against the different strains vary, with C34 showing the strongest activity, followed by C35, while A42 has the highest MIC values, indicating lower efficacy. However, A42 demonstrates the best synergy with fluconazole against fluconazole-resistant Candida strains. Cytotoxicity assays reveal that the chloropropyl group present in C35 and A42 enhances cytocompatibility. Co-culture with macrophages shows significant yeast killing for C. albicans and C. auris when fluconazole and A42 are combined, requiring concentrations 4 and 16 times lower than their MIC values, enhancing antifungal activity. Given fluconazole’s fungistatic nature and the emergence of drug-resistant strains, benzo[a]phenoxazine derivatives’ ability to enhance fluconazole’s efficacy present a promising strategy to address antifungal resistance in critical pathogens. These findings align with global research priorities, offering new potential avenues for developing more effective antifungal therapies.

This work was supported by FEDER (European Fund for Regional Development)-COMPETEQREN-EU in the framework of the Strategic Funding UIDB/04050/2020 (https://doi.org/10.54499/UIDB/04050/2020) and UID/QUI/00686/2020. FCT is also acknowledged for the Ph.D. scholarships UI/BD/150872/2021 (https://doi.org/10.54499/UI/BD/150872/2021) (M.I.P.), 2020.08235.BD (https://doi.org/10.54499/2020.08235.BD) (P.P.-S.), SFRH/BD/133513/2017 and COVID/BD/152169/2021 (A.C.-B.).