Sphingolipid Inhibitors as an Alternative to Treat Candidiasis Caused by Fluconazole-Resistant Strains

Candida species are fungal pathogens known to cause a wide spectrum of diseases, and Candida albicans and Candida glabrata are the most common associated with invasive infections. A concerning aspect of invasive candidiasis is the emergence of resistant isolates, especially those highly resistant to fluconazole, the first choice of treatment for these infections. Fungal sphingolipids have been considered a potential target for new therapeutic approaches and some inhibitors have already been tested against pathogenic fungi. The present study therefore aimed to evaluate the action of two sphingolipid synthesis inhibitors, aureobasidin A and myriocin, against different C. albicans and C. glabrata strains, including clinical isolates resistant to fluconazole. Susceptibility tests of aureobasidin A and myriocin were performed using CLSI protocols, and their interaction with fluconazole was evaluated by a checkerboard protocol. All Candida strains tested were sensitive to both inhibitors. Regarding the evaluation of drug interaction, both aureobasidin A and myriocin were synergic with fluconazole, demonstrating that sphingolipid synthesis inhibition could enhance the effect of fluconazole. Thus, these results suggest that sphingolipid inhibitors in conjunction with fluconazole could be useful for treating candidiasis cases, especially those caused by fluconazole resistant isolates.

Chromosome 1 trisomy overcomes the block of sphingolipid biosynthesis caused by aureobasidin A and confers pleiotropic effects on susceptibility to antifungals in Candida albicans

Sphingolipids are important membrane lipid components of eukaryotic cells. In Candida albicans, chromosome 1 trisomy not only overcame the block of sphingolipid biosynthesis caused by aureobasidin A, but also altered tolerance to three of the four major classes of antifungal drugs. Two haploinsufficient genes on chromosome 1, PDR16 and IPT1, were associated with tolerance to aureobasidin A. This study illustrates an example of multi-drug tolerance caused by aneuploidy in the human fungal pathogen C. albicans.

Hope on the Horizon: Novel Fungal Treatments in Development

The treatment of invasive fungal infections remains challenging due to limitations in currently available antifungal therapies including toxicity, interactions, restricted routes of administration, and drug resistance. This review focuses on novel therapies in clinical development, including drugs and a device. These drugs have novel mechanisms of action to overcome resistance, and some offer new formulations providing distinct advantages over current therapies to improve safety profiles and reduce interactions. Among agents that target the cell wall, 2 glucan synthesis inhibitors are discussed (rezafungin and ibrexafungerp), as well as fosmanogepix and nikkomycin Z. Agents that target the cell membrane include 3 fourth-generation azoles, oral encochleated amphotericin B, and aureobasidin A. Among agents with intracellular targets, we will review olorofim, VL-2397, T-2307, AR-12, and MGCD290. In addition, we will describe neurapheresis, a device used as adjunctive therapy for cryptococcosis. With a field full of novel treatments for fungal infections, the future looks promising.

A Simple Method to Detect the Inhibition of Transcription Factor-DNA Binding Due to Protein–Protein Interactions In Vivo

Binding of transcription factors (TFs) to cis-regulatory elements (DNA) could modulate the expression of downstream genes, while interactions between TFs and other proteins might inhibit them binding to DNA. Nowadays, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) approaches are usually employed to detect the inhibitory effect. However, EMSA might not reflect the inhibitory effect in vivo. ChIP requires preparation of specific antibody or stable genetic transformation and complicated experimental steps, making it laborious and time-consuming. Here, based on the yeast one-hybrid (Y1H) system, we present a simple method to detect the inhibition of TF–DNA binding due to protein–protein interactions in vivo. When interactions between TFs and other proteins inhibit TFs binding to DNA, the reporter (Aureobasidin A resistance) gene is not activated, thereby inhibiting yeast growth on media containing the AbA antibiotic. Two examples were tested with the newly developed method to demonstrate its feasibility. In conclusion, this method provides an alternative strategy for detecting the inhibition of DNA-binding of TFs due to their interactions with other proteins in vivo.

Sphingolipids activate the endoplasmic reticulum stress surveillance pathway

Proper inheritance of functional organelles is vital to cell survival. In the budding yeast, Saccharomyces cerevisiae, the endoplasmic reticulum (ER) stress surveillance (ERSU) pathway ensures that daughter cells inherit a functional ER. Here, we show that the ERSU pathway is activated by phytosphingosine (PHS), an early biosynthetic sphingolipid. Multiple lines of evidence support this: (1) Reducing PHS levels with myriocin diminishes the ability of cells to induce ERSU phenotypes. (2) Aureobasidin A treatment, which blocks conversion of early intermediates to downstream complex sphingolipids, induces ERSU. (3) orm1Δorm2Δ cells, which up-regulate PHS, show an ERSU response even in the absence of ER stress. (4) Lipid analyses confirm that PHS levels are indeed elevated in ER-stressed cells. (5) Lastly, the addition of exogenous PHS is sufficient to induce all ERSU phenotypes. We propose that ER stress elevates PHS, which in turn activates the ERSU pathway to ensure future daughter-cell viability.