Dihydrofolate Reductase Is a Valid Target for Antifungal Development in the Human Pathogen Candida albicans

Abstract

While the folate biosynthetic pathway has provided a rich source of antibacterial, antiprotozoal, and anticancer therapies, it has not yet been exploited to develop uniquely antifungal agents. Although there have been attempts to develop fungal-specific inhibitors of dihydrofolate reductase (DHFR), the protein itself has not been unequivocally validated as essential for fungal growth or virulence. The purpose of this study was to establish dihydrofolate reductase as a valid antifungal target. Using a strain with doxycycline-repressible transcription of DFR1 (PTETO-DFR1 strain), we were able to demonstrate that Dfr1p is essential for growth in vitro. Furthermore, nutritional supplements of most forms of folate are not sufficient to restore growth when Dfr1p expression is suppressed or when its activity is directly inhibited by methotrexate, indicating that Candida albicans has a limited capacity to acquire or utilize exogenous sources of folate. Finally, the PTETO-DFR1 strain was rendered avirulent in a mouse model of disseminated candidiasis upon doxycycline treatment. Collectively, these results confirm the validity of targeting dihydrofolate reductase and, by inference, other enzymes in the folate biosynthetic pathway as a strategy to devise new and efficacious therapies to combat life-threatening invasive fungal infections. IMPORTANCE The folate biosynthetic pathway is a promising and understudied source for novel antifungals. Even dihydrofolate reductase (DHFR), a well-characterized and historically important drug target, has not been conclusively validated as an antifungal target. Here, we demonstrate that repression of DHFR inhibits growth of Candida albicans, a major human fungal pathogen. Methotrexate, an antifolate, also inhibits growth but through pH-dependent activity. In addition, we show that C. albicans has a limited ability to take up or utilize exogenous folates as only the addition of high concentrations of folinic acid restored growth in the presence of methotrexate. Finally, we show that repression of DHFR in a mouse model of infection was sufficient to eliminate host mortality. Our work conclusively establishes DHFR as a valid antifungal target in C. albicans.

Antibacterial, Antifungal, and Antioxidant Activities of Silver Nanoparticles Biosynthesized from Bauhinia tomentosa Linn

“…Moreover, AgNPs can interact with pivotal microbial proteins to inhibit their activities and cause cell death [ 78 , 79 , 80 , 81 ]. Accordingly, we selected representative proteins for each species to study the possible three-dimensional (3D) interaction of AgNPs with bacterial DNA gyrase [ 82 , 83 ], fungal CYP51 (cytochrome P450 monooxygenase (CYP) superfamily) [ 51 , 84 ] and fungal dihydrofolate reductase [ 85 ]. To predict the biological interactions of the biosynthesized B. tomentosa Linn flower-powder extract-derived AgNPs with these possible microbial target proteins, we performed molecular docking analysis using a Patch dock server for the 3D structures of PDB proteins 3G7B (DNA gyrase, S. aureus ), 4WUB (DNA gyrase, E. coli ), 5TZI (cytochrome P450, C. albicans ), and 6DRS (dihydrofolate reductase, A. flavus ).…”

“…The AA residues interacted with silver through hydrophobic contact (Figure 11). (cytochrome P450 monooxygenase (CYP) superfamily) [51,84] and fungal dihydrofolate reductase [85]. To predict the biological interactions of the biosynthesized B. tomentosa Linn flower-powder extract-derived AgNPs with these possible microbial target proteins, we performed molecular docking analysis using a Patch dock server for the 3D structures of PDB proteins 3G7B (DNA gyrase, S. aureus), 4WUB (DNA gyrase, E. coli), 5TZI (cytochrome P450, C. albicans), and 6DRS (dihydrofolate reductase, A. flavus).…”

Repurposing Benzbromarone as Antifolate to Develop Novel Antifungal Therapy for Candida Albicans

“…[6] Echinocandins is a recently approved antifungal agent effective only against the Candida species by targeting their cell wall biosynthesis via intravenous administrations. [7,8] It is a matter of high concern that the mediocre e cacy and limited clinical utility of the available three major classes of antifungal agents is responsible for increasing mortality rates in patients having IFI with a high rate of transmission of the infection among humans. More than one-third of the patients with Candida infections resisted the therapy with azoles like uconazole, voriconazole, as well as isavuconazole.…”

“…Targeting the folate biosynthetic pathway of pathogenic microbes by developing a suitable antifolate is a highly successful approach in developing different types of antimicrobial therapies for humans. [7,10] Drug repurposing is a trending approach to identi cation of a newer pharmacological role for an existing approved drug, whereas establishing the newer mechanism of an existing approved drug by identifying its potent binding a nity towards an alternate drug target is called drug repositioning. The high cost as well as the slow pace involved in the traditional drug development process are becoming a barrier in the combat against the new diseases affecting mankind.…”

Antifungal docking studies of Anacyclus pyrethrum(L.) and Senna Obtusifolia(L.) against candida albicans .

“…The folate biosynthetic pathway is an important and effective mechanism that serves as source for novel Antifungal. Historically Dihydrofolate reductase (DHFR), a well-characterized and important drug target for many antineoplastic, antibacterial drugs but has not been mostly validated as an antifungal target [1]. Here, wecarried out some in-silico experiments with help of some software (Autodock Vina) of DHFR inhibits growth of Candida albicans which is a major human fungal pathogen.…”