Δ24-Sterol Methyltransferase Plays an Important Role in the Growth and Development of Sporothrix schenckii and Sporothrix brasiliensis

Abstract

Inhibition of Δ24-sterol methyltransferase (24-SMT) in Sporothrix schenckii sensu stricto and Sporothrix brasiliensis was investigated in vitro. The effects on fungal growth and sterol composition of the 24-SMT inhibitor 22-hydrazone-imidazolin-2-yl-chol-5-ene-3β-ol (H3) were compared to those of itraconazole. MIC and MFC analysis showed that H3 was more effective than itraconazole against both species in both their filamentous and yeast forms. H3 showed fungistatic activity in a time-kill assay, with inhibitory activity stronger than that of itraconazole. GC analysis of cell sterol composition showed that sterols present in control cells (ergosterol and precursors) were completely replaced by 14α-methylated sterols after H3 exposure. Itraconazole only partially inhibited ergosterol synthesis but completely arrested synthesis of other sterols found in control cells, promoting accumulation of nine 14α-methyl sterols. Based on these results, we propose a schematic model of sterol biosynthesis pathways in S. schenckii and S. brasiliensis. Effects on cell morphology due to 24-SMT inhibition by H3 as analyzed by SEM and TEM included irregular cell shape, reduced cytoplasmic electron-density, and reduced thickness of the microfibrillar cell wall layer. Moreover, 24-SMT inhibition by H3 promoted mitochondrial disturbance, as demonstrated by alterations in MitoTracker® Red CMXRos fluorescence intensity evaluated by flow cytometry. When used in conjunction with itraconazole, H3 enhanced the effectiveness of itraconazole against all tested strains, reducing at least half (or more) the MIC values of itraconazole. In addition, cytotoxicity assays revealed that H3 was more selective toward these fungi than was itraconazole. Thus, 24-SMT inhibition by H3 was an effective antifungal strategy against S. schenckii and S. brasiliensis. Inhibition of the methylation reaction catalyzed by 24-SMT has a strong antiproliferative effect via disruption of ergosterol homeostasis, suggesting that this enzyme is a promising target for novel antifungal therapies against sporotrichosis, either as sole treatments or in combination with itraconazole.

Chemical Composition and Antifungal Properties of Essential Oil of <i>Origanum vulgare</i> Linnaeus (Lamiaceae) against <i>Sporothrix schenckii</i> and <i>Sporothrix brasiliensis</i>

“…In the presence of ketoconazole, the cells showed up twisted and with kinks. In support of our results, Santos [21] observed changes in the length and width of S. schenckii and S. brasiliensis cells in the yeast form, when exposed to drugs azole and amphotericin B.…”

Zoonotic Epidemic of Sporotrichosis: Cat to Human Transmission

“…Identifying these epidemiological trends associated with the emergence of drug resistance is important to adjust antifungal therapy and to encourage the development of new drugs to treat sporotrichosis. Currently, potential alternative therapies to impair Sporothrix development and tackle sporotrichosis include terpinen-4-ol and farnesol [23], miltefosine [24], TCAN26 (a structural analogue of miltefosine) [25], and H3 (a 24-sterol methyltransferase inhibitor) [26]. …”

Metal–azasterol complexes: Synthesis, characterization, interaction studies with DNA and TrxR and Biological Evaluation

“…One of them had high field shifts from 3.63 ppm to 3.49 ppm, overlapping with H-3, while, the other proton remains in the same position at 3.32 ppm. In the 13 C-NMR spectrum of V, C-22 appears at 43.7 ppm indicating that bromine substitution effect shifts this carbon resonance to high-field.…”

“…It is important to highlight that AH3 was synthesized as analogue of azasterol H3 (Sterol hydrazone class), because it has shown a significant anti-proliferative activity against S. schenckii and S. brasiliensis, even better than itraconazole [13]. Thus, it can be seen from Table 4 that, this new azasterol called AH3, also inhibited fungal growth at low micromolar concentration.…”

“…A new 24-SMT inhibitor belonging to the sterol hydrazone class, named 22-hydrazoneimidazolin-2-yl-chol-5-ene-3β-ol (H3) showed a significant anti-proliferative activity against the dimorphic fungus Paracoccidioides brasiliensis [9,12], and Cryptococcus neoformans [11]. More recently, its effects on the growth of Sporothrix schenckii and Sporothrix brasiliensis was described, and compared to that of itraconazole [13]. GC-MS analysis of cell sterol composition showed that sterols present in control cells (ergosterol and precursors) were replaced mainly by lanosterol after H3 exposure.…”