The focus of TRICONT is to put together a concerted effort to discover high affinity ligands that can be developed into more efficacious and less toxic drugs against trypanosomatidae. The precise goals of this project will be structure elucidation and determination of inhibitor binding site(s), designing of high-affinity scaffolds capable of binding tightly to the targets, synthesis of small-ligand libraries based on the scaffold. Finally, these ligands can be tested for their efficacy by in vitro biochemical assay and cell based assay (against L. infantum). An international team of interdisciplinary experts with exclusive working experience in the area of computational biology, protozoal biology and chemistry are put together for achieving the goals of TRICONT. Briefly, the main objectives of the project are as follows.

1. Derivation of small ligands as effective inhibitors of the target proteins using innovative insilico strategies, synthesis and validation of the identified inhibitors for the enzymes of L. infantum followed by T. brucei, T. cruzi and L. major.

An important prerequisite for rational drug design is the knowledge of the three dimensional structure of the drug target. Though some knowledge about the active site of these parasite targets is available, the structures of these targets are not known. However six structures of rpiB (from four different pathogens two complexes with substrate) and two structures of aspA (complexes with substrate and cofactor) are available in PDB. Indian group will be dedicated to the determination of the structure of these two enzymes by using predictive modeling (homology modeling). Structural characterization of protein-ligand complexes will specifically help to identification of the high affinity and low affinity ligands and find out the best affinity inhibitor. Based on further in silico analysis of these putative inhibitors by virtual screening methods such as docking, binding affinity prediction the most promising compounds will be identified. Other protocol will also be used such as fragment-based drug design and ligand–based design when appropriate. These compounds will be collectively analyzed and the scaffolds will be identified. These results will be used by Italy group to carry out the synthesis of the compounds that belong to the identified scaffolds. The synthesized compounds will be used for rapid screening assay using purified enzymes.

2. Development of potential drug candidate(s) further as safer therapeutic agent(s) for the treatment of Leishmaniasis, African trypanosomiasis and Chagas’ disease.

This objective represents the most complex objective for iteration of synthesis/testing cycle by sequential application of in vitro enzyme based assay, in silico screening, QSAR and in vivo cell based assay. After initial round of screening, physico chemical properties of active molecules will be used for in silico screening. Shape based screening algorithm and various surface topology & charge-based prediction algorithms will be used screen available compound database (ACD) for searching similar compounds. Structure based screening using pharmacophore mapping and docking approaches will be used to filter ACD results for target specific ligands. Validated QSAR model will be used to predict the biological activity of screened compound library. Generated library compounds will also be subjected for ADME/T properties and drug likeliness properties for evaluating drug likeliness. Finally enzyme based screening assay will be carried out for pool of compounds showing good predicted activity followed by cell based screening of compounds showing good activity in enzyme assay. Final tests for compounds evaluation will be performed in vivo against mice models of leishmaniasis. To further improve the biopharmaceutical properties of the new synthesized small molecules drug delivery systems, based on liposomal and nanoparticles formulations, specific to Trypanosomatidae will be developed.