Lead Hopping for PfDHODH Inhibitors as Antimalarials Based on Pharmacophore Mapping, Molecular Docking and Comparative Binding Energy Analysis (COMBINE): A Three‐Layered Virtual Screening Approach

Malaria is a major worldwide public health threat securing the fifth position among the top ten causes of worldwide death with worrying social and economic burdens due to the rapid emergence of resistance to the currently available antimalarial drugs like chloroquine, sulfadoxine-pyrimethamine including Artemisinin. The causative agent for the disease is a parasite belonging to the Plasmodium species transmitted to human by the aid of female Anopheles mosquito. Although several antimalarial drugs have been reported till date, the efficacy of these drugs has been severely limited by widespread drug resistance necessitating new target based therapy. The enzymes governing the pyrimidine biosynthesis within the malarial parasite constitute essential targets for a number of clinically effective therapies since the pyrimidine metabolism pathway proves to be a vulnerable component of the parasite’s biology. Amongst the chemically validated targets, P. falciparum dihydroorotate dehydrogenase (PfDHODH), present in the mitochondria, is one of the essential druggable targets identified against P. falciparum that catalyses fourth reaction (formation of dihydroorotate to orotate which represents the rate limiting step in de novo pyrimidine biosynthesis) of pyrimidine de novo biosynthesis. Inhibition of the enzyme results in the shutdown of the mitochondrial electron transport chain thereby arresting crucial metabolic pathways within the microorganism leading to the inhibition of pyrimidine biosynthesis and consequent parasite death, rendering this enzyme a valid and attractive drug target against P. falciparum. Subsequently, the PfDHODH enzyme has been reviewed by several authors 7] as a promising drug target for novel antimalarial chemotherapy. The human and parasitic DHODH enzymes differ extensively in their amino acid sequence at the inhibitor binding-pocket thereby providing the structural basis for the identification of species-specific inhibitors. Several attempts have been made by different authors’ groups [8–10] to identify potent PfDHODH inhibitors based on in silico methodologies. Quantitative structure-activity relationship (QSAR) techniques constitute an essential in silico tool aiming to develop statistically valid models that may be utilized for database screening and activity prediction of untested molecules. The in silico virtual screening and computer-aided drug design methodologies enable an initial screening of large databases based on molecular properties, thereby saving both time and money involved in synthesizing and analyzing each of the molecules available in the database. The in silico screening techniques thus reduce the number of molecules to be synthesized and analyzed by identifying the hit compounds only. In the present work, 3D pharmacophore models have been developed based on a series of triazolopyrimidine derivatives exhibiting PfDHODH inhibitory activity. 13] Statistical validation of the model was performed based on a test set with already reported PfDHODH inhibitory activity and the essential pharmacophoric features were identified. Thus, the best pharmacophore model was used as 3D search queries for screening the NCI database (http://cactus.nci.nih.gov/download/nci) to identify new hits having potent PfDHODH inhibitory activity. The Lipinski filter and the ADMET filter were employed for the selection of the drug like molecules with the requisite pharmacokinetic properties. Finally, the lead compounds, selected based on the best fit values of the molecules, were subjected to molecular docking studies to refine the retrieved hits. The virtual screening approach in combination with pharmacophore modeling, molecular docking and COMBINE (comparative binding energy) based QSAR has been utilized in this work for identifying the requisite pharmacophoric features and screening the lead compounds with potential PfDHODH inhibitory activity. Ten pharmacophore hypotheses (Table 1) thus developed yielded acceptable results in terms of cost functions and