Ultrastructural alterations induced by ∆24(25)-sterol methyltransferase inhibitors on Trichomonas vaginalis

7 p. : il.Trichomonas vaginalis is an important human parasite that causes trichomoniasis, a cosmopolitan sexually transmitted disease. Currently, the treatment of choice for T. vaginalis infections is metronidazole. The increase in metronidazole-resistant parasites and undesirable side effects of this drug make the search for alternative chemotherapeutic approaches a priority for the management of trichomoniasis. Here, the antiproliferative and ultrastructural effects of sterol biosynthesis inhibitors against T. vaginalis were investigated. It was found that 22,26-azasterol (5 mM) and 24(R,S),25-epiminolanosterol (10 mM), known inhibitors of D24(25)- sterol methyltransferase, exhibited antiproliferative effects on T. vaginalis trophozoites cultured in vitro. Morphological analyses showed that azasterols induced changes in the ultrastructure of T. vaginalis. The most significant alterations were (1) membrane blebbing and disruption, (2) wrinkled cells and (3) the formation of cell clusters. In addition, autophagic vacuoles, Golgi duplication arrest, an abnormal Golgi enlargement and damaged hydrogenosomes were also observed. Nonspecific cytotoxicity assays using the cultured mammalian cell lines Madin–Darby canine kidney cells showed no effect of the azasterols on the viability and proliferation of these cells at a concentration that significantly inhibited the proliferation of T. vaginalis, indicating a selective antiparasitic action. Taken together, these results suggest that azasterols could be important compounds in the development of novel chemotherapeutic approaches against T. vaginalis

Synthetic arylquinuclidine derivatives exhibit antifungal activity against Candida albicans, Candida tropicalis and Candida parapsilopsis

10 p. : il.Background: Sterol biosynthesis is an essential pathway for fungal survival, and is the biochemical target of many antifungal agents. The antifungal drugs most widely used to treated fungal infections are compounds that inhibit cytochrome P450-dependent C14a-demethylase (CYP51), but other enzymes of this pathway, such as squalene synthase (SQS) which catalyses the first committed step in sterol biosynthesis, could be viable targets. The aim of this study was to evaluate the antifungal activity of SQS inhibitors on Candida albicans, Candida tropicalis and Candida parapsilopsis strains. Methods: Ten arylquinuclidines that act as SQS inhibitors were tested as antiproliferative agents against three ATCC strains and 54 clinical isolates of Candida albicans, Candida tropicalis and Candida parapsilopsis. Also, the morphological alterations induced in the yeasts by the experimental compounds were evaluated by fluorescence and transmission electron microscopy. Results: The most potent arylquinuclidine derivative (3-[1’-{4’-(benzyloxy)-phenyl}]-quinuclidine-2-ene) (WSP1267) had a MIC50 of 2 μg/ml for all species tested and MIC90 varying from 4 μg/ml to 8 μg/ml. Ultrathin sections of C. albicans treated with 1 μg/ml of WSP1267 showed several ultrastructural alterations, including (a) loss of cell wall integrity, (b) detachment of the plasma membrane from the fungal cell wall, (c) accumulation of small vesicles in the periplasmic region, (d) presence of large electron-dense vacuoles and (e) significantly increased cell size and cell wall thickness. In addition, fluorescence microscopy of cells labelled with Nile Red showed an accumulation of lipid droplets in the cytoplasm of treated yeasts. Nuclear staining with DAPI revealed the appearance of uncommon yeast buds without a nucleus or with two nuclei. Conclusion: Taken together, our data demonstrate that arylquinuclidine derivatives could be useful as lead compounds for the rational synthesis of new antifungal drugs