New Approaches for the Identification of Drug Targets in Protozoan Parasites - International Review of Cell and Molecular Biology - page 382

metronidazole (as a prodrug) is reduced to a nitro radical ( Sisson et al., 2002 )
by electrons coming from PFOR ( Upcroft and Upcroft, 2001 ). This radi-
cal causes irreversible damage on intracellular structures. More recent find-
ings in the microaerophilic parasite T. vaginalis indicate, however, another
mode of action. By comparing 2-D-protein electrophoresis patterns from
treated and untreated cells, the authors suggest that metronidazole binds
to the sulfhydryl group in the active center of various enzymes includ-
ing thioredoxin reductase thereby impairing essential cellular functions, and
that resistant cells overcome this blocking by re-regulating other enzymes
involved in oxidoreductive processes, such as PFORs ( Leitsch et al., 2009a ).
In this model, downregulation of PFOR would be a consequence rather
than a prerequisite of resistance formation. Also in this model, an activated
form of metronidazole would act on multiple targets, the number being
limited to enzymes with sulfhydryl groups in their active center. Moreover,
metronidazole is thought to form adducts with free cysteine since addition
of free cysteine to the culture medium renders E. histolytica more resis-
tant to metronidazole ( Leitsch et al., 2009b ). In a study on G. lamblia , even
a positive correlation between metronidazole-resistance formation and
PFOR mRNA quantity has been observed ( Argüello-García et al., 2009 ).
A more likely direct target of nitro drugs in anaerobic parasites are nitro-
reductases (NRs), as identified in G. lamblia ( Müller et al., 2007b ; Nillius
et al., 2011 ). Furthermore, possible giardial metronidazole-resistance mech-
anisms involving complex changes in the gene expression pattern have been
described, including alterations of those genes encoding variant surface pro-
teins (VSPs), which mediate antigenic variation ( Müller et al., 2007a ).
3.5.4. Nitro Compounds Effective against Aerobic Parasites
Interestingly, trypanosomal mitochondria contain a bacterial-like NR
activating the antitrypanosomal prodrugs benznidazole and nifurtimox,
loss of a single copy of NR in T. cruzi being sufficient to cause significant
cross-resistance to these drugs and to other nitroheterocyclic compounds
( Wilkinson et al., 2008 ). The recombinant NR from T. cruzi and T. brucei are
used in target-based screenings in order to identify novel antitrypanosomal
agents ( Hall et al., 2010 ). Trypanosomal NR catalyzes an oxygen-insensitive
reaction with the interaction of enzyme, reductant, and prodrug occurring
through a ping-pong mechanism. In the case of benznidazole as a pro-
drug, the major product is 4,5-dihydro-4,5-dihydroxyimidazole releasing
the reactive dialdehyde glyoxal during his further breakdown. In the pres-
ence of guanosine, cytotoxic guanosine-glyoxal adducts are generated ( Hall