Diglycosyl diselenides alter redox homeostasis and glucose consumption of infective African trypanosomes

Abstract

With the aim to develop compounds able to target multiple metabolic pathways and, thus, to lower the chances of drug resistance, we investigated the anti-trypanosomal activity and selectivity of a series of symmetric diglycosyl diselenides and disulfides. Of 18 compounds tested the fully acetylated forms of di-β-D-glucopyranosyl and di-β-D-galactopyranosyl diselenides (13 and 15, respectively) displayed strong growth inhibition against the bloodstream stage of African trypanosomes (EC50 0.54 μM for 13 and 1.49 μM for 15) although with rather low selectivity (SI < 10 assayed with murine macrophages). Nonacetylated versions of the same sugar diselenides proved to be, however, much less efficient or completely inactive to suppress trypanosome growth. Significantly, the galactosyl (15), and to a minor extent the glucosyl (13), derivative inhibited glucose catabolism but not its uptake. Both compounds induced redox unbalance in the pathogen. In vitro NMR analysis indicated that diglycosyl diselenides react with glutathione, under physiological conditions, via formation of selenenylsulfide bonds. Our results suggest that non-specific cellular targets as well as actors of the glucose and the redox metabolism of the parasite may be affected. These molecules are therefore promising leads for the development of novel multitarget antitrypanosomal agents.

Glucose 6-Phosphate Dehydrogenase from Trypanosomes: Selectivity for Steroids and Chemical Validation in Bloodstream Trypanosoma brucei

“…On the other hand, inhibition of G6PDH has consequences on the downstream output of the PPP pathway, i.e., it causes a shortage of ribose 5-phosphate that impairs cell proliferation (the cytostatic effect). Thus, in order to verify the on-target effect of the steroid derivatives and their metabolic effect in the pathogen, we used a redox-reporter cell line of bloodstream T. brucei [ 46 ]. Based on the higher potency of the derivatives against bloodstream T. brucei and the fact that this parasite stage represents a clinically more relevant biological model to investigate compounds mode of action, the studies were performed on this parasite and not on T. cruzi epimastigotes.…”

“…The redox biosensor (hGrx1-roGFP2) [ 47 ] consists of a redox sensitive green fluorescent protein (roGFP2) fused to human glutaredoxin 1 (hGrx1) that enables a rapid redox equilibration between roGFP2 and the pool of reduced and oxidized glutathione and trypanothione [ 46 , 48 , 49 ].…”

“…Bloodstream T. b. brucei expressing the redox biosensor hGrxroGFP2 was grown as described elsewhere [ 46 ]. Briefly, 2 × 10 6 cells/ mL were seeded per well in a 96-well plate and incubated with different steroids derivatives each at 2× their EC 50 , 250 µM menadione, 250 µM diamide or 1% v / v DMSO for 4 h (5% CO 2 and 37 °C).…”

“…Next, 50 µL from each well were transferred to a 96-well plate, containing 100 µL of sterile PBS with glucose 1% ( w / v ). For each condition tested, a second sample was incubated with 1 mM DTT 20 min prior to analysis by flow cytometry, while propidium iodide (exclusion dye; PI) was added at a final concentration of 2 µg/mL All samples were analyzed with a C6Accuri flow cytometer (BD) as described in [ 46 ]. GFP fluorescence (filter λem = 530/40 nm) was only analyzed for viable cells (PI negative).…”

5-Vinylquinoline-substituted nitrofurans as inhibitors of trypanothione reductase and antitrypanosomal agents

“…The antitrypanosomal activity of the compounds was evaluated against the bloodstream form of T. b. brucei strain 427, line 449 hGrx-roGFP2, sensitive to nifurtimox with EC 50 = 15 ± 2.5 µM (compound concentration causing 50% parasite growth inhibition) [20]. The parasites were grown in the HMI-9 medium complemented with 10% (v/v) fetal bovine serum (FBS) and antibiotics in a humidified incubator with 5% CO 2 and at 37°C [16].…”