The Mechanism Of Using GGTase I As A Drug Target To Reverse Echinocandin-resistance In Candidiasis

Candida is the most common opportunistic pathogenic fungus in humans.In healthy people,it usually does not cause disease or only causes slight superficial infection.When the human immune system is damaged,however,it causes severe invasive infection.Although Candida albicans accounts for most of candidiasis,the infections caused by non-albicans Candida species increased significantly.The newest class of antifungal to reach the clinic is the echinocandins,which includes the clinically approved drugs micafungin,caspofungin,and anidulafungin.The echinocandins noncompetitively inhibit the ?-1,3-D-glucan synthase enzyme encoded by FKS1.Targeting Fks1 disrupts the synthesis of the major cell wall biopolymer ?-1,3-glucan,resulting in loss of cell wall integrity and imparting a severe cell wall stress on the fungus.Echinocandin resistance is increasing in prevalence in the clinic as their increased use for prophylaxis and prolonged treatment regimens,which poses grave concern.Echinocandin resistance resulting in clinical failures is conferred by amino acid substitutions that occur in two limited but highly conserved Fks hot-spot regions in C.albicans and most other Candida species.This highlights the urgent need for novel treatments that can tackle echinocandin-resistant candidasis.Here,we report that deletion of Cdc43,the ? subunit of GGTase I,an important enzyme that catalyses the prenylation of proteins in C.albicans,confers hypersensitivity to echinocandins,which renders GGTase I a tractable target in combatting echinocandin resistance.We found that geranylgeranylation of Rho1 by GGTase I is required for its membrane localization.Disrupting GGTase I blocks the membrane localization of Rho1,the regulatory subunit of Fks1,leading to the decreased amounts of glucan in cell wall,and thereby exacerbating the cell wall stress induced by echinocandins.We identified a fungal selective GGTase I inhibitor L-269289 by screening library of small-molecule compoundes.Inhibiting GGTase I by L-269289 mislocalizes Rho1 by preventing it from geranylgeranylation modification,which resembles the cdc43 mutant,resulting in disruption of ?-1,3-D-glucan synthase activity.L-269289 synergizes with echinocandins against C.albicans and enhances the efficacy of echinomycin in vitro,eliminating echinocandin-resistant C.albicans at lower concentrations.In animal models for disseminated infection,L-269289 improves the efficacy of caspofungin in the treatment of echinomycin-resistant C.albicans infection.Furthermore,L-269289 and echinocandins also act in a synergistic manner for the treatment of Candida tropicalis and Candida parapsilosis.Importantly,deletion of CDC43 is lethal in Candida glabrata.L-269289 is active on its own to kill Candida glabrata,and its fungicidal activity is enhanced when combined with caspofungin.Thus,targeting GGTase I has therapeutic potential to address the clinical challenge of echinocandin-resistant candidiasis.