*2.4.1 Arginase (L-arginine amidinohydrolase, ARG, E.C. 3.5.3.1)*

Arg is an enzyme that catalyzes the conversion of L-arginine amino acid to L-ornithine and urea.

The expression of the *Leishmania amazonensis* ARG in a bacterial host was done [44]. da Silva et al. expressed the recombinant enzyme in *E. coli* and performed biochemical and biophysical characterization studies [45].

**Figure 4.** *Polyamine metabolism and enzymes in the pathway.*

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

Reguera et al. suggest that broad inhibition of ARG activity alone will be insufficient to achieve therapeutically useful control of leishmaniasis, but combined inhibition of ARG with downstream enzymes leading to polyamine synthesis could result in improved therapeutic responses [46]. 3′-methoxy-cinnamoyl-1,3,4 thiadiazolium-2-phenylamine, an ARG inhibitory compound, exhibited moderate antileishmanial activity upon amastigotes of *L. amazonensis* [47].

[1,2,4]triazolo[1,5-a]pyrimidine derivatives [48], pyrazolo[3,4-d]pyrimidine derivatives [49], α,α-difluorohydrazide derivatives [50], chalcone derivatives [51], cinnamide derivatives [52], and 7,8-dihydroxyflavone—gold nanoparticles [53] were also studied as antileishmanial compounds with the mechanism of ARG inhibition.

On the other hand, antileishmanial natural products exhibiting ARG inhibitor activity with antileishmanial properties were reported—flavonoid and quercetin derivative [54], orientin and isovitexin [55], verbascoside [56], fisetin [57], rosmarinic acid, and caffeic acid [58].

#### *2.4.2 Ornithine decarboxylase (ODC, EC 4.1.1.17)*

ODC metabolizes ornithine to the diamine putrescine by its catalytic action [59]. Although alpha-difluoromethylornithine (DFMO) is an irreversible inhibitor of ODC, DFMO has not shown any antileishmanial activity [60]. Therefore, inhibition of ODC serves as a promising therapeutic paradigm for the treatment of leishmaniasis [61].

3-aminooxy-1-aminopropane was reported as a selective ODC inhibitor with potent antileishmanial activity against *Labrus donovani* (*L. donovani* promastigotes IC50 of 42 μM and *L. donovani* amastigotes IC50 of 5 μM) [62].

Gama-guanidinooxypropylamine [63], diospyrin [64], oxochromen, xanthone, and azaspirodecene derivatives [65] are reported in the literature with their ability to inhibit ODC enzyme and antileishmanial activity.

#### *2.4.3 Spermidine synthase (SpdSyn, SpdS, EC 2.5.1.16)*

SpdS catalyzes the conversion of putrescine to spermidine, a crucial polyamine for parasite proliferation. Genetic studies proved that SpdS is an essential gene in *L.donovani* [66]. Additionally, it was demonstrated that *L. donovani* amastigotes require SpdS activity to sustain a robust infection in mice; which is required for virulence [67].

Up to our knowledge, the only reported SpdS inhibitor with antileishmanial properties is natural compound hypericin [68].

#### *2.4.4 S-Adenosylmethionine decarboxylase (AdoMetDC, EC 4.1.1.50)*

AdoMetDC is involved in the synthesis of spermidine and spermine, an essential polyamine for *Leishmania*. Therefore, AdoMetDC may be a potential therapeutic target for leishmaniasis [69].

CGP40215A, a specific AdoMetDC inhibitor, was also reported with the antileishmanial effect that verified the potential of AdoMetDC enzyme inhibition strategy [70].

#### *2.4.5 Trypanothione synthetase (Trypanothione synthase, TryS; EC 6.3.1.9)*

TryS bifunctionally catalyzes both biosynthesis and hydrolysis of the glutathione-spermidine adduct trypanothione, which is the main regulator in intracellular

**Figure 5.** *Examples of TryS inhibitor structures with antileishmanial activity.*

thiol-redox metabolite for parasitic trypanosomatids. As TryS is absent in humans, targeting this enzyme provides selectivity. Inhibition of TryS results in controlling relative levels of the critical metabolites, trypanothione, glutathionylspermidine, and spermidine in *Leishmania* [71]. Genetic and chemical analyses reveal that TryS is essential for *Leishmania infantum* [72].

In a computational screening campaign, oxabicyclo[3.3.1]nonanone skeleton was identified not only as a TryS inhibitor but also with TR inhibitory properties. A modest antileishmanial activity was reported for compound PS203 upon *L. donovani* promastigotes (**Figure 5**) [73]. In another study, TryS from *L. donovani* was characterized and inhibition studies with the natural compounds selected from an earlier Micro Source discovery natural product data set were performed [74]. Among the tested natural compounds, conessine and uvaol showed good TryS inhibition (Ki of 3.12 μM and 3.55 μM, respectively) with significant antileishmanial activity on *L. donovani* promastigotes (IC50 of 13,42 μM and 11,23 μM, respectively) (**Figure 5**) [74].

About 144 compounds belonging to seven different scaffolds were tested for TyrS inhibitory properties in a study by Benitez et al. One of the most promising inhibitors (IC50 of 0.15 μM) namely MOL2008, an *N*<sup>5</sup> -substituted paullone derivative was evaluated upon *L. infantum* promastigotes (EC50 of 12.6 μM) (**Figure 5**) [75]. Following these results, 36 different derivatives of MOL2008 were developed by the same group [76]. Based on intriguing TyrS inhibition of compound 20 (IC50 0.3 μM), it was tested on both *L. infantum* promastigotes and *L. infantum* amastigotes. The metabolic changes exerted by 20 in both promastigote form and amastigote form of *L. infantum* are compatible with TryS inhibition (**Figure 5**) [76].
