Image-Based In Vitro Screening Reveals the Trypanostatic Activity of Hydroxymethylnitrofurazone against Trypanosoma cruzi

Abstract

Hydroxymethylnitrofurazone (NFOH) is a therapeutic candidate for Chagas disease (CD). It has negligible hepatotoxicity in a murine model compared to the front-line drug benznidazole (BZN). Here, using Trypanosoma cruzi strains that express bioluminescent and/or fluorescent reporter proteins, we further investigated the in vitro and in vivo activity of NFOH to define whether the compound is trypanocidal or trypanostatic. The in vitro activity was assessed by exploiting the fluorescent reporter strain using wash-out assays and real-time microscopy. For animal experimentation, BALB/c mice were inoculated with the bioluminescent reporter strain and assessed by highly sensitive in vivo and ex vivo imaging. Cyclophosphamide treatment was used to promote parasite relapse in the chronic stage of infection. Our data show that NFOH acts by a trypanostatic mechanism, and that it is more active than BZN in vitro against the infectious trypomastigote form of Trypanosoma cruzi. We also found that it is more effective at curing experimental infections in the chronic stage, compared with the acute stage, a feature that it shares with BZN. Therefore, given its reduced toxicity, enhanced anti-trypomastigote activity, and curative properties, NFOH can be considered as a potential therapeutic option for Chagas disease, perhaps in combination with other trypanocidal agents.

Synthesis, Characterization, and Activity of Hydroxymethylnitrofurazone Nanocrystals against Trypanosoma cruzi and Leishmania spp.

“…The NFOH nanocrystals described in the present study (NFOH-F1 and NFOH-F2) exhibited similar activities compared with free NFOH [14][15][16][17]. Likewise, considering the fact that the NFOH nanocrystals can be tested against other strains of T. cruzi or Leishmania in animal assays, the present results are important to future studies on NFOH (free, nanocrystals, or polymers).…”

“…Hydroxymethylnitrofurazone (NFOH) is a nitrofurazone prodrug (Figure 1), which has been considered a crucial and promising anti-T. cruzi candidate in vitro [14,15]. Its activity was evaluated and confirmed in murine models with acute and chronic stages of Chagas disease [15][16][17].…”

The Use of AlphaFold for In Silico Exploration of Drug Targets in the Parasite Trypanosoma cruzi

“…Regarding BNZ, NFX, hydroxymethylnitrofurazone (NFOH) and fexinidazole, they are all described to be prodrugs, which upon being metabolized generate highly reactive intermediary compounds that can target many cellular components ( Maya et al., 2007 ; Hall et al., 2011 ; Scarim et al., 2021 ). In particular, these drugs would be mainly metabolized by T. cruzi nitroreductase (TcCLB.510611.60, UniProtKB Q4D8D9) ( Maya et al., 2007 ; Hall et al., 2011 ).…”

Drug/Lead Compound Hydroxymethylation as a Simple Approach to Enhance Pharmacodynamic and Pharmacokinetic Properties

“…Also, NFOH (91) hepatotoxicity was assessed in in vitro experiments with HepG2 cells, and it caused an enhancement in ROS generation and in the oxidative damage to DNA biomarker 8-oxo-29-deoxyguanosine (8oxo-dG). Despite this, cell viability was not significantly (Davis et al, 2010;Ekins et al, 2015;Scarim et al, 2018;Scarim et al, 2021a changed by the administration of NFOH (91). On the other hand, the cells treated with BNZ showed no increase in this biomarker, but they exhibited a reduction in cell viability, displaying 33% cell death at 100 μM (Davies et al, 2014).…”