Mechanism of azole resistance in Candida glabrata in the presence of immunosuppressant mycophenolic acidTome, Miha (Avtor) Raspor, Peter (Mentor) Kuchkler, Karl (Komentor) Candida glabratadrug resistancemechanism of drug resistancedrug combinationsimmunosuppressive drugsantifungalsazolesfunctional genomicsfluconazolemycophenolic acidSaccharomyces cerevisiaeCandida glabrata is the second most common cause of Candidemia and other forms of invasive candidiasis in the western world. It has an intrinsic high tolerance and often develops resistance to various antifungals. Understanding the resistance mechanisms is of utmost importance to tackle this problem. The most studied among the yeast pathogens is Candida albicans, however C. glabrata and its pathogenic traits differ from it, especially the proficiency of C. glabrata to adapt to harsh environment and develop resistance. C. glabrata is a close relative to the ale yeast Saccharomyces cerevisiae, they share a high degree of homology, although the regulation and function of some genes can be different. Drug combinations are a valid strategy to combat the resistance, and many combinations are already present in the clinic. Unfortunately, the drug-drug interactions are still mostly only considered for their potentially toxic effect on the host, and their effects on pathogens are usually ignored. On this basis, we tested different combinations of immunosuppressive and antifungal drugs against C. glabrata and S. cerevisiae clinical isolates. We successfully confirmed the synergistic interaction between calcineurin inhibitors (cyclosporine A, Fk506) and antifungals (amphotericin B, itraconazole, and fluconazole). We also discovered clinically relevant antagonism between purine biosynthesis inhibitor mycophenolic acid (MPA) and azole antifungals and explored the mechanism behind it. MPA alleviates the effect of azoles through enhanced activity of efflux pumps, which lowers the bioavailability of azoles thus reducing their effect. The MPA induction of the efflux pumps comes as a cell response to weak lipophilic acid, and via dysfunctional mitochondria regulating the Hog1 osmotic/oxidative stress response and Pkc1 cell wall integrity pathways involving genes HSP12, SSA3, RCK2, ROX1, and YPK1. Ypk1 serine/threonine protein kinase seems to be an integral regulator for drug response and a potential connection between the signaling from the dysfunctional mitochondria, by sensing the sphingolipid homeostasis and triggering the cell wall integrity pathway and drug response. Ypk1 is a promising target for the drug development, since its deletion greatly reduces the tolerance to fluconazole and MPA and diminishes the suppressive antagonistic interaction between the drugs. We also discovered/confirmed 28 gene deletions that significantly change the susceptibility of C. glabrata to fluconazole, 26 for MPA and 17 for the combination of both drugs. All of these genes present potential for further drug development.[M. Tome]20182018-07-13 07:45:28Doktorsko delo/naloga101911sl