Potential Hydrophobic Pocket of Squalene Synthase: An In Silico Analysis

Cardiovascular disease cases increase due to consumption cholesterol dietary habit. It is well-known that squalence synthase (SQS) is the first committed enzyme for cholesterol synthesis. Therefore, SQS become target of anti-cholesterol. This paper aims to determine the potential binding pocket of SQS (PDB ID: 1EZF). Dogsitescorer, siteFinder, and DEPTH were used for binding pocket prediction and MOE 2009.10 was performed for molecular docking. We found that there are five out of 37 pockets which have druggability score above 0.8. Pocket_5 is the highest drugability and favorable for hydrophobic interaction, yet lower number of hydrogen bond with the ligand. However, Pocket_2, and Pocket_3 are suitable for hydrogen bond formation of ligand-protein. Molecular docking study showed that TAK-475, D99, and Cynarin inhibitors were embedded on the P_2 and P_3 of SQS, showing that P2_and P3 are promising binding pocket for ligand interactions. These results show a promising alternative to design anti-cholesterol using these potential pocket in silico

Parasite, Compartments, and Molecules: Trick versus Treatment on Chagas Disease

Chagas disease, caused by the protozoan Trypanosoma cruzi, is endemic to Latin America, standing out as a socio-economic problem for low-income tropical populations. Such disease affects millions of people worldwide and emerges in nonendemic areas due to migration and climate changes. The current chemotherapy is restricted to two nitroderivatives (benznidazole and nifurtimox), which is unsatisfactory due to limited efficacy (particularly in chronic phase) and adverse side effects. T. cruzi life cycle is complex, including invertebrate and vertebrate hosts and three developmental forms (epimastigotes, trypomastigotes, and amastigotes). In this chapter, we will discuss promising cellular and molecular targets present in the vertebrate-dwelling forms of the parasite (trypomastigotes and amastigotes). Among the cellular targets, the mitochondrion is the most frequently studied; while among the molecular ones, we highlight squalene synthase, C14α-sterol demethylase, and cysteine proteases. In this scenario, proteomics becomes a valuable tool for the identification of other molecular targets, and some previously identified candidates will be also discussed. Multidisciplinary studies are needed to identify novel key molecules in T. cruzi in order to increase trypanocidal activity and reduce mammalian toxicity, ensuring the development of novel drugs for Chagas disease

Non-bisphosphonate inhibitors of isoprenoid biosynthesis identified via computer-aided drug design.

The relaxed complex scheme, a virtual-screening methodology that accounts for protein receptor flexibility, was used to identify a low-micromolar, non-bisphosphonate inhibitor of farnesyl diphosphate synthase. Serendipitously, we also found that several predicted farnesyl diphosphate synthase inhibitors were low-micromolar inhibitors of undecaprenyl diphosphate synthase. These results are of interest because farnesyl diphosphate synthase inhibitors are being pursued as both anti-infective and anticancer agents, and undecaprenyl diphosphate synthase inhibitors are antibacterial drug leads

Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins

Certain species of pathogenic bacteria damage tissues by secreting cholesterol‐dependent cytolysins, which form pores in the plasma membranes of animal cells. However, reducing cholesterol protects cells against these cytolysins. As the first committed step of cholesterol biosynthesis is catalyzed by squalene synthase, we explored whether inhibiting this enzyme protected cells against cholesterol‐dependent cytolysins. We first synthesized 22 different nitrogen‐containing bisphosphonate molecules that were designed to inhibit squalene synthase. Squalene synthase inhibition was quantified using a cell‐free enzyme assay, and validated by computer modeling of bisphosphonate molecules binding to squalene synthase. The bisphosphonates were then screened for their ability to protect HeLa cells against the damage caused by the cholesterol‐dependent cytolysin, pyolysin. The most effective bisphosphonate reduced pyolysin‐induced leakage of lactate dehydrogenase into cell supernatants by >80%, and reduced pyolysin‐induced cytolysis from >75% to <25%. In addition, this bisphosphonate reduced pyolysin‐induced leakage of potassium from cells, limited changes in the cytoskeleton, prevented mitogen‐activated protein kinases cell stress responses, and reduced cellular cholesterol. The bisphosphonate also protected cells against another cholesterol‐dependent cytolysin, streptolysin O, and protected lung epithelial cells and primary dermal fibroblasts against cytolysis. Our findings imply that treatment with bisphosphonates that inhibit squalene synthase might help protect tissues against pathogenic bacteria that secrete cholesterol‐dependent cytolysins

Current and future chemotherapy for Chagas disease

Luís Gaspar is thankful to FCT for funding (scholarship reference: SFRH/BD/81604/2011). The research leading to these results has received funding from the European Community’s Seventh Framework Programme under grant agreement No.602773 (Project KINDRED) and No. 603240 (Project NMTrypI).American trypanosomiasis, commonly called Chagas disease, is one of the most neglected illnesses in the world and remains one of the most prevalent chronic infectious diseases of Latin America with thousands of new cases every year. The only treatments available have been introduced five decades ago. They have serious, undesirable side effects and disputed benefits in the chronic stage of the disease – a characteristic and debilitating cardiomyopathy and/or megavisceras. Several laboratories have therefore focused their efforts in finding better drugs. Although recent years have brought new clinical trials, these are few and lack diversity in terms of drug mechanism of action, thus resulting in a weak drug discovery pipeline. This fragility has been recently exposed by the failure of two candidates, posaconazole and E1224, to sterilely cure patients in phase 2 clinical trials. Such setbacks highlight the need for continuous, novel and high quality drug discovery and development efforts to discover better and safer treatments. In this article we will review past and current findings on drug discovery for Trypanosoma cruzi made by academic research groups, industry and other research organizations over the last half century. We will also analyze the current research landscape that is now better placed than ever to deliver alternative treatments for Chagas disease in the near futurePostprintPeer reviewe

In silico analysis of Mentha pipertia (phyto-constituents) as HMG coa reductase and squalene synthase inhibitors

Mentha piperita has been well known for its hypolipidemic activity. This prompted the present study to be carried out on a selected 12 phyto-constituents of Mentha piperita which are naringin, eriodictyol, eriodictyol 7-glucuronide, eriocitrin, hesperidin, isorohifolin, luteolin 7-glucoside, diosmin, rosmarinic acid, piperitoside, menthoside and caffeic acid. These phyto-constituents were evaluated on the docking behaviour of HMG CoA reductase (HMGR) and Squalene synthase (SQS) using Discovery Studio Version 3.1. In addition, molecular physicochemical, drug-likeness, ADMET (Absorption, Distribution, Metabolism, Excretion and Toxicity) and TOPKAT (Toxicity Prediction by Komputer Assisted Technology) analyses were done. The molecular physicochemical analysis revealed that eriodictyol, rosmarinic acid and caffeic acid (3 ligands) complied with Lipinski’s rule of five. ADMET analysis showed that eriodictyol and caffeic acid exhibited good intestinal absorption property. Docking studies and binding free energy calculations revealed that menthoside (-70.0 kcal/mol) and piperitoside (-65.32 kcal/mol) exhibited the maximum interaction energy with HMGR and SQS respectively. Caffeic acid exhibited very least binding energy irrespective of its target protein. Caffeic acid showed interaction with Leu546 and Gln212 amino acid residue of HMGR and SQS. Hence, the results of this present study exhibited the potential of these twelve ligands as hypolipidemic agents

A Mapping of Drug Space from the Viewpoint of Small Molecule Metabolism

Small molecule drugs target many core metabolic enzymes in humans and pathogens, often mimicking endogenous ligands. The effects may be therapeutic or toxic, but are frequently unexpected. A large-scale mapping of the intersection between drugs and metabolism is needed to better guide drug discovery. To map the intersection between drugs and metabolism, we have grouped drugs and metabolites by their associated targets and enzymes using ligand-based set signatures created to quantify their degree of similarity in chemical space. The results reveal the chemical space that has been explored for metabolic targets, where successful drugs have been found, and what novel territory remains. To aid other researchers in their drug discovery efforts, we have created an online resource of interactive maps linking drugs to metabolism. These maps predict the “effect space” comprising likely target enzymes for each of the 246 MDDR drug classes in humans. The online resource also provides species-specific interactive drug-metabolism maps for each of the 385 model organisms and pathogens in the BioCyc database collection. Chemical similarity links between drugs and metabolites predict potential toxicity, suggest routes of metabolism, and reveal drug polypharmacology. The metabolic maps enable interactive navigation of the vast biological data on potential metabolic drug targets and the drug chemistry currently available to prosecute those targets. Thus, this work provides a large-scale approach to ligand-based prediction of drug action in small molecule metabolism

The Identification of Small Molecule Inhibitors to Candida albicans Phosphatidylserine Synthase

Candida albicans phosphatidylserine (PS) synthase, encoded by the CHO1 gene, has been identified as a potential drug target for new antifungals against systemic candidiasis due to its importance in virulence, absence in the host and conservation among fungal pathogens. This dissertation is focused on the identification of inhibitors for this membrane enzyme. Cho1 has two substrates: cytidyldiphosphate-diacylglycerol (CDP-DAG) and serine. Previous studies identified a conserved CDP-alcohol phosphotransferase (CAPT) binding motif present within Cho1, and here we revealed that mutations in all but one conserved amino acid within the CAPT motif resulted in decreased Cho1. For serine, we have predicted a serine-binding site based on sequence alignment and found that some of the residues in this putative serine-binding site are required for Cho1 function. One residue, R189, is particularly interesting because it was suggested to be involved in serine binding. Then, we attempted to perform a small molecule screening on C .albicans Cho1, which will be facilitated by purified Cho1 protein. Due to the transmembrane nature, several solubilizing reagents were used to solubilize Cho1 protein. Digitonin was determined to be the best detergent as it retained the most PS synthase activity. Pull-downs of HA-tagged Cho1 in the digitonin-solubilized fraction reveal an apparent MW of Cho1 consistent with a hexamer. Biochemical and electron microscopy analysis suggest that the hexamer is composed of a trimer of dimers. Cho1 protein was then purified to near-homogeneity as a hexamer and was optimized for high activity to be used in the small drug screening. For the screening, we developed a nucleotidase-coupled malachite green-based screen against purified Cho1. Over 7,300 molecules curated from repurposing chemical libraries were interrogated in primary and dose-responsivity assays using this platform, and seven compounds were identified to inhibit purified Cho1. Among all, compound CBR-5884 disrupted in vivo Cho1 function by inducing phenotypes consistent with the cho1∆∆ mutant, including a reduction of cellular PS levels. Kinetic curves and computational docking suggest that CBR-5884 competes with serine for binding of Cho1 with a Ki of 1,550 ± 245.6 nM, thus this compound has the potential for further drug design

Toward New Antileishmanial Compounds: Molecular Targets for Leishmaniasis Treatment

The leishmaniases are a group of diseases caused by protozoan parasites—Leishmania sp. Leishmaniasis is classified among the 20 neglected diseases by WHO. Although the disease has been known for more than 120 years, the number of drugs used for the treatment is still limited to 5–6. The first-line drugs against leishmaniasis are pentavalent antimonials, which were introduced to the treatment 70 years ago—despite all their side effects. Molecular targets are becoming increasingly important for efficacy and selectivity in postgenomic drug research studies. In this chapter, we have discussed potential therapeutic targets of antileishmanial drug discovery such as pteridine reductase (PTR1), trypanothione reductase (TR), N-myristoyltransferase (NMT), trypanothione synthetase (TryS), IU-nucleoside hydrolase, and topoisomerases, enzymes and their inhibitors reported in the literature