Synergy and Mechanism of Leflunomide Plus Fluconazole Against Resistant Candida albicans: An in vitro Study

Abstract

Objective

The global rise in the resistance of Candida albicans to conventional antifungals makes Candida albicans infections harder to treat. The main objective of this study was to investigate the antifungal effects and underlying mechanisms of leflunomide in combination with triazoles against resistant Candida albicans.

Methods

In this study, the microdilution method was used to determine the antifungal effects of leflunomide in combination with three triazoles on planktonic cells in vitro. The morphological transition from yeast to hyphae was observed under a microscope. The effects on ROS, metacaspase, efflux pumps, and intracellular calcium concentration were investigated, respectively.

Results

Our findings suggested that leflunomide + triazoles showed a synergistic effect against resistant Candida albicans in vitro. Further study concluded that the synergistic mechanisms were resulted from multiple factors, including the inhibited efflux of triazoles, the inhibition of yeast-to-hyphae transition, ROS increasing, metacaspase activation, and [Ca²⁺]_i disturbance.

Discussion

Leflunomide appears to be a potential enhancer of current antifungal agents for treating candidiasis caused by resistant Candida albicans. This study can also serve as an example to inspire the exploration of new approaches to treating resistant Candida albicans.

Keywords:

• Candida albicans
• triazoles
• leflunomide
• synergy
• synergistic mechanism

Abbreviations

LEF, Leflunomide; FLC, Fluconazole; ITR, Itraconazole; VRC, Voriconazole; MIC, Minimum inhibitory concentration; FICI, Fractional inhibitory concentration index; SMIC, Sessile minimum inhibitory concentration; CA, Candida albicans; CG, Candida glabrata; CK, Candida krusei.

Acknowledgments

We are very grateful to Dr. Yueling Wang (Department of Clinical Laboratory, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, 250014, Shandong Province, P.R. China) for his assistance in the identification of strains and the determination of drug susceptibility.

Author Contributions

All authors made a significant contribution to the work reported, whether that is in the conception, study design, execution, acquisition of data, analysis and interpretation, or in all these areas; took part in drafting, revising or critically reviewing the article; gave final approval of the version to be published; have agreed on the journal to which the article has been submitted; and agree to be accountable for all aspects of the work.

Disclosure

The authors report no conflicts of interest in this work.

Additional information

Funding

This work was supported by the [National Natural Science Foundation of China] under Grant [82173703], [Key Research and Development Plan of Shandong Province] under Grant [GG201809270073] and [Shandong Provincial Natural Science Foundation] under Grant [ZR2020QH365]. These funding organizations had no role in the design of the study and the collection, analysis and interpretation of the data.