Combining Fluconazole with Benzo[a]phenoxazine Derivatives as a Promising Strategy Against Fluconazole-Resistant Candida Species

The rise in non-<i>albicans Candida</i> 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[<i>a</i>]phenoxazine derivatives (<b>C34</b>, <b>C35</b>, <b>A42</b>, and <b>A44</b>) against 14 <i>Candida</i> 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 <i>Candida</i> killing in the presence of synergistic compounds. The MIC values against the different strains vary, with <b>C34</b> showing the strongest activity, followed by <b>C35</b>, while <b>A42</b> has the highest MIC values, indicating lower efficacy. However, <b>A42</b> demonstrates the best synergy with fluconazole against fluconazole-resistant <i>Candida</i> strains. Cytotoxicity assays reveal that the chloropropyl group present in <b>C35</b> and <b>A42</b> enhances cytocompatibility. Co-culture with macrophages shows significant yeast killing for <i>C. albicans</i> and <i>C. auris</i> when fluconazole and <b>A42</b> 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[<i>a</i>]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.