Antifungal effects and potential mechanism of bortezomib in combination with fluconazole against Candida albicans

Abstract

The clinical management of Candida albicans (C. albicans) infections presents considerable problems, intensified by the few therapeutic choices available for this disease. The increasing resistance to fluconazole (FLC) following its widespread clinical use underscores the need for novel therapeutic strategies. Drug repurposing represents a promising approach to expand the antifungal armamentarium, and bortezomib (BTZ), a proteasome inhibitor originally approved for multiple myeloma, has emerged as a candidate for this purpose. This work systematically assessed the synergistic effects of fluconazole (FLC) with bortezomib (BTZ) on the susceptibility of C. albicans, focusing specifically on the planktonic and biofilm development phases. The results exhibited a synergistic antifungal effect of BTZ and FLC against FLC-resistant C. albicans in planktonic cells. The minimal inhibitory concentration (MIC) of BTZ decreased from 64-128 µg/ml to 8 µg/ml, whereas the MIC of FLC reduced from >128 µg/ml to 0.25 µg/ml, resulting in a fractional inhibitory concentration index (FICI) of less than 0.5. For FLC-susceptible C. albicans strains, the medication combination exhibited an indifferent interaction, with FICI above 0.5. The combination exhibited synergistic effects against C. albicans during the early stages of biofilm formation (4 and 8 h). The sessile minimum inhibitory concentration (sMIC) of FLC significantly lowered from ≥128 µg/mL to 0.25 µg/mL, whereas the sMIC of BTZ diminished from ≥128 µg/mL to 8-16 µg/mL, with a FICI of less than 0.5, signifying significant synergy. Time-kill assays revealed a delayed but sustained inhibitory effect against planktonic cells (24–48 h), while CFU enumeration confirmed a concentration-dependent reduction in biofilm viability, with the higher concentration (0.25 + 16) maintaining efficacy through 24 h. In vivo, using the Galleria mellonella (G. mellonella) infection model, the FLC+BTZ combination significantly improved larval survival, reduced fungal burden, and alleviated tissue damage compared with FLC monotherapy. Mechanistic studies demonstrated that this synergistic effect was mediated by multiple mechanisms, including the suppression of hyphal growth, inhibition of efflux pump activity, and intracellular formation of reactive oxygen species (ROS). These findings highlight the potential of drug repurposing in combating antifungal resistance and provide mechanistic insights into overcoming C. albicans drug resistance.