*2.4.6 Trypanothione reductase (TR, TryR, Trypanothione-disulfide reductase 1, EC 1.8.1.12)*

One of the main strategies of the host organism to overcome the infection is oxidative stress. TR has been purified from *T.cruzi* [77], first, and then from *Labrus donovani* [78]. TR enzyme is responsible for keeping trypanothione in the reduced state that is a variant of glutathione in *Leishmania* parasites. These enzyme

#### *Toward New Antileishmanial Compounds: Molecular Targets for Leishmaniasis Treatment DOI: http://dx.doi.org/10.5772/intechopen.101132*

inhibitors have been investigated in antileishmanial drug discovery as the enzyme is essential for the parasite survival and its absence in the host, in which glutathione reductase (GR) is found, provides selectivity [79]. Although both TR and GR are inhibited by trivalent antimonials, TR is considerably more sensitive [80]. TR enzyme is also a target for anti-Chagas compounds and antimalarials. The main limitation of TR becoming a target in antileishmanial drug discovery is that in order to obtain a considerable effect in parasites' redox state, a minimum of *85%* inhibition is required [81]. Additionally, GR should be considered as an off-target for TR inhibitors and the selectivity over TR enzyme of the compounds may be presented. Apart from being an interesting target for antileishmanial drug design, it is also a popular target for antimalarial compounds.

The early discovery of tricyclic inhibitors that are specific for TR over GR led to the design and synthesis of a group of phenothiazine derivatives and their openedring analogs.

The first rational drugs with TR inhibitor activity over GR inhibition are tricyclic structures like phenothiazine and imipramine. Based on this, among several of quaternary phenothiazines, [3-(2-chloro-4a,10a-dihydrophenothiazin-10-yl) propyl] - (3,4-dichlorobenzyl) dimethylammonium derivative (Ki 0.12 μM) was reported possessing improved activity up to 2-fold compared to chlorpromazine on *L. donovani* species [82]. Compound **10**, an opened ring analog of phenothiazine, showed antileishmanial activity upon *L. donovani* (IC50 of 3.9 μg/mL). Expectedly, it was one of the most active compounds for TR enzyme with the Ki value of 6.5 μM [83].

A series of bis (2-amino diphenyl sulfides) were designed and synthesized to inhibit TR [84]. Among them, compound **15** was found to be the most active with the IC50 value of 200 nM. Although there was no correlation between TR inhibition and antileishmanial activity, the compounds showed activity upon *L. infantum* amastigotes (**Figure 6**) [84]. Sulfonamide and urea derivatives of quinacrine with varying methylene spacer lengths were designed as TR inhibitors and their antiprotozoal activities were evaluated [85]. Compound **2b** (TR IC50 of 3.3 μM and GR IC50 of 27.2 μM) was also one of the most active compounds upon *L. donovani* among with *Trypanosoma cruzi and Trypanosoma brucei* [85] (**Figure 6**).

In the pursuit of discovering novel lead heteroaromatic frameworks, harmaline, pyrimidobenzothiazine, and aspidospermine scaffolds were tested against TR inhibition (Ki of 35.1 μM, Ki of 26.9 and Ki of 64.6 μM, respectively) and *L. amazonensis* promastigote toxicity. Moreover, compounds have not exhibited any GR inhibitory activity [86]. Interestingly, Blackie et al. has introduced ferrocenic 4-aminoquinoline urea compounds with TR inhibitory and antileishmanial properties to the literature [87]. Although compounds inhibited TR in a low μM range with good selectivity over GR and showed antileishmanial activity on *L. donovani* amastigotes, unfortunately, these compounds were found to be toxic to macrophages (**Figure 6**) [87].

In an HTS campaign, 100,000 lead-like compounds were evaluated for their TR inhibition. As our focus on antileishmanial compounds, 2 series of compounds namely, nitrogenous heterocycles (triazine and pyrimidine derivatives) and conjugated indole derivatives took our interest in their potential on *L. donovani* amastigotes (**Figure 6**) [88].

Various chemical structures were reported with TR inhibitor activity and leishmaniacidal activity to the literature: Ag(0) nanoparticles encapsulated by ferritin molecules [89], Cu(II) diketonates [90], oxabicyclo[3.3.1]nonanones [73], azole-based compounds – e. pyrrole [91], β-carboline–quinazolinone hybrid [92], phenothiazine and phenoxazine derived chloroacetamides [93], selenocyanates and diselenide compounds [94, 95], iminodibenzyl derivatives with ethylenediamine,

**Figure 6.** *Examples of TR inhibitor structures with antileishmanial activity.*

ethanolamine and diethylenetriamine and their copper(II) complexes [96], diaryl sulfide derivatives [97], ammonium trichloro [1,2-ethanediolato-*O,O*′]-tellurat [98], all-hydrocarbon stapled peptides [99] chalcone derivatives [100], thiophene derivatives [101], imidazole-phenyl-thiazole compounds [102], isothiocyanate derivatives [103], (phenylthio)pyrimidin-4-amine derivatives [104], ferrocenylquinoline derivatives [105], triazole-phenyl-thiazoles derivatives [106], fluorene derivatives [107], adamantan derivatives, and their gold complexes [108] and natural products [109, 110] (**Figure 6**).
