Evaluation of the efflux inhibitory potential of aminopyrazole derivative to restore azole sensitivity in Candida albicans.

Candida albicans is an opportunistic fungus that causes infections in people with weakened immune system. Candida has the propensity to gain drug resistance by mutation, and its ability to extrude antifungals through drug transporters. Efflux pump inhibitors increase the intracellular drug concentration and restore drug sensitivity in resistant strains. We synthesized and screened 10 azole heterocyclic derivatives (pyrazole-5-amine and 2H-indazole) against Mdr1p/Cdr1p overexpressing and knock-out strains of Candida albicans. Among the derivatives screened, sulfenylated pyrazole-5-amine (designated as 4a) displayed strong efflux inhibitory potential and caused a 64-fold reduction in the fluconazole MIC. Compound 4a inhibited the Mdr1p pump, as MIC reversal was not observed in the Mdr1p knock-out strain. 4a exhibited synergy with fluconazole only against Mdr1p harboring strains, as discerned by the Checkerboard assay. Combining 4a with fluconazole restricted microbial growth and reduced cell counts by 4 log fold relative to treatment with fluconazole in a time-kill assay. Infection of RAW macrophages followed by treatment with a combination of sulfenylated aminopyrazole & fluconazole resulted in a significant 2 log reduction in intracellular cell counts relative to treatment with fluconazole. 4a also inhibited biofilm formation and yeast to hyphal shift in Candida, thereby reducing virulence. Combining EPIs with antifungal drugs has the potential to improve the treatment of C. albicans infections and reduce the risk of drug resistance. However, further research is needed to determine the safety and efficacy of these compounds in humans.