**1. Introduction**

Leishmaniasis is a disease caused by a single-cell eukaryotic parasite of the *Leishmania* species. This protozoan parasite causes a substantial level of morbidity and mortality. *Leishmania* has a digenetic life cycle [1]. In mammals, the parasite colonizes macrophages, transforming into intracellular amastigotes. The parasite has an adaptive way to life conditions. The amastigotes tolerate low pH and are hydrolase resistant [2].

Trypanothione is a major product of the trypanothione biosynthesis pathway in trypanosomes which is crucial in maintaining cellular redox potential and is essential for the parasite's survival. This molecule is catalyzed by so many enzymes for which *Leishmania major* trypanothione reductase (*Lm*TR, E.C. 1.6.4.8) plays a critical role in the biosynthetic pathway. TR reduces trypanothione (T[S]2) to dithiol (T[SH]2). They catalyze the transfer of electrons from NADPH to their specific substrate via an FAD prosthetic group [3]. The reduced form is critical in regulating oxidative stress by reacting with reactive oxygen species (ROS) that are produced by the macrophage. T[SH]2 is not only needed for detoxification of peroxides but also required for the synthesis of DNA precursors, homeostasis of ascorbate, sequestration and export of thiol conjugate [4].

Trypanothione reductase is a member of the disulphide oxidoreductase family of enzymes. It has an analogue in the human body, glutathione reductase (GR) which also carries out oxidoreductive reactions. But *Lm*TR does not process GSSG and host GR does not reduce T[S]2 [5, 6]. The ascribed reasons for targeting *Lm*TR include the following: (i) trypanothione reductase is a key enzyme in regulating a reducing environment aiding in disease pathogenesis, (ii) this parasite does not depend on the host for reduced trypanothione, (iii) it has a less close known homologous protein in humans; (iv) the availability of template homologs for modeling purpose; and (v) moreover, *in vitro* trials have proven *Lm*TR to be a good therapeutic target [7].

Several inhibitors have been screened against this enzyme causing a reduction of infectivity and decreased capacity of the parasite to survive within intracellular macrophages. Potent compounds, such as 7-chloro-4-nitro-5-quinazolin-4 ylsulfanyl-2,1,3-benzothiadia-zole (CNQB) and 4-phenyl-5-(4-nitro-cinnamoyl)- 1,3,4-thiadiazolium-2-phenylamine-chloride (PNTPC) with IC50 values 0.58 and 1.63 μM, respectively have already been tested in an *in vitro* assay against trypanothione reductase of trypanosomatids [8, 9].

Computer-aided drug designing is an *in-silico* approach for drug discovery that combines computational and pharmaceutical research [10]. This application helps in spanning the drug discovery pipeline and helps to speed up and rationalize the drug design process while reducing costs [11]. Ehrlich in 1909 first defined the term pharmacophore as 'a molecular framework that carries (*phoros*) the essential features responsible for a drug's (*pharmacon*) biological activity [12]. These features are essential functional groups of atoms in a three-dimensional position that interact with a receptor. Ligand-based drug design can be performed in association with molecular docking. These methods can be combined to identify a number of new hit compounds with potent inhibitory activity and to understand the main interactions at the binding sites. Appropriate use of these methods can improve the ability to identify and optimize hits and confirm their potential to serve as scaffolds for producing new therapeutic agents [13].

Drugs currently used for the treatment of human leishmaniasis are toxic, having severe adverse reactions which limit their use. Aside this includes, increase in resistance by the parasite, high cost of available drugs, lack of efficacy against VL \HIV co-infections with standard chemotherapy, and the development of a single drug or formulation for all forms of leishmaniasis [14–17]. Therefore, the

development of novel, effective drugs with reduced side effects, is still a major priority for health researchers, in spite of many compelling research reports published on antileishmanial agents in the last 10 years [18]. In this study, *in silico* method of identifying leads was used incorporating the knowledge of pharmacophore and virtual screening to arrive at lead molecules. The overall goal of the study was to predict with high degree potent selective inhibitors of *Lm*TR from the African Natural Product Database (AfroDb) and North African Natural Product Database (NANPDB).
