Combinatorial Chemistry & High Throughput Screening - Volume 18, Issue 5, 2015

A high throughput, semi-automated clearance screening assay in hepatocytes was developed allowing a scientist to generate data for 96 compounds in one week. The 384-well format assay utilizes a Thermo Multidrop Combi and an optimized LC-MS/MS method. The previously reported LCMS/ MS method reduced the analytical run time by 3-fold, down to 1.2 min injection-to-injection. The Multidrop was able to deliver hepatocytes to 384-well plates with minimal viability loss. Comparison of results from the new 384-well and historical 24-well assays yielded a correlation of 0.95. In addition, results obtained for 25 marketed drugs with various metabolism pathways had a correlation of 0.75 when compared with literature values. Precision was maintained in the new format as 8 compounds tested in ≥39 independent experiments had coefficients of variation ≤21%. The ability to predict in vivo clearances using the new stability assay format was also investigated using 22 marketed drugs and 26 AbbVie compounds. Correction of intrinsic clearance values with binding to hepatocytes (in vitro data) and plasma (in vivo data) resulted in a higher in vitro to in vivo correlation when comparing 22 marketed compounds in human (0.80 vs 0.35) and 26 AbbVie Discovery compounds in rat (0.56 vs 0.17), demonstrating the importance of correcting for binding in clearance studies. This newly developed high throughput, semi-automated clearance assay allows for rapid screening of Discovery compounds to enable Structure Activity Relationship (SAR) analysis based on high quality hepatocyte stability data in sufficient quantity and quality to drive the next round of compound synthesis

The aim of this study was to examine the anticancer activities and phytochemical profiles of Dicranum scoparium against HeLa cell lines. The bio-guided fractionation studies of dichloromethane extract have high antiproliferative activities. Fractions 7, 9, 19, 20 are rich source of unsaturated fatty acids, and- in the case of Fr-19 may improve the antiproliferative activities as well as increase the unsaturated fatty acid content. The effect of proliferative activities in hexane extract can be attributed to the saturated fatty acid composition of D. scoparium. The Fr-9 exhibited strong antiproliferative activity at concentrations of 100 and 50 μg mL-1 compared to 5-FU. The fractions of 7, 9, 19 and 20 from dichloromethane extracts exhibited antiproliferative activities at a concentration of 100 μg mL-1. The HPLC-TOF/MS studies gave nine compounds from the most active fraction of dichloromethane at concentrations of 250 and 100 μg mL-1. The lower activities were obtained from the fractions including steroid derivatives.

HIV-1 protease is a key enzyme for viral maturation because it cleaves precursor polypeptides into mature structural and functional proteins. The introduction of protease inhibitors into therapy in the mid-1990s has dramatically changed the AIDS panorama worldwide. However, resistance to currently available protease inhibitors remains a serious challenge that must be overcome. Herein, we report a fragment-based QSAR study of a series of highly potent HIV-1 protease inhibitors, as well as the structural basis of their binding affinity. Hologram QSAR (HQSAR) analyses were performed, resulting in robust statistical models that consistently correlated the bioactivity profile with the twodimensional molecular descriptors. The robustness of the best model was assessed based on the correlation coefficients (q2 = 0.70 and r2 = 0.90), as well as the prediction of the activity of an external test set (r2 pred = 0.75). Structure-based molecular modeling studies were performed to investigate the binding mode of the best inhibitor in the active site of the enzyme. The HQSAR model and the structural findings provide valuable insights for the rational design of structurally related HIV-1 protease inhibitors.

The breast cancer resistant protein (BCRP) is an important transporter and its inhibitors play an important role in cancer treatment by improving the oral bioavailability as well as blood brain barrier (BBB) permeability of anticancer drugs. In this work, a computational model was developed to predict the compounds as BCRP inhibitors or non-inhibitors. Various machine learning approaches like, support vector machine (SVM), k-nearest neighbor (k-NN) and artificial neural network (ANN) were used to develop the models. The Matthews correlation coefficients (MCC) of developed models using ANN, k-NN and SVM are 0.67, 0.71 and 0.77, and prediction accuracies are 85.2%, 88.3% and 90.8% respectively. The developed models were tested with a test set of 99 compounds and further validated with external set of 98 compounds. Distribution plot analysis and various machine learning models were also developed based on druglikeness descriptors. Applicability domain is used to check the prediction reliability of the new molecules.

An efficient and selective synthesis of substituted chromene derivatives via threecomponent reaction of 4-hydroxycoumarin or 1,3-dicarbonyl compounds, activated acetylenic compounds and N-nucleophiles is described. The reaction was conducted under solvent-free conditions at 70°C using potassium fluoride impregnated on natural zeolite as a cheap and available solid base. The procedure has several advantages involving selectivity, excellent yields and a convenient work-up method.

Malaria is the world’s most fatal disease - causing up to 2.7 million deaths annually all over the world. The ability of organisms to develop resistance against existing antimalarial drugs exacerbates the problem. There is a clear cut need for more effective, affordable and accessible drugs that act by novel modes of action. Glutathione synthetase (GS) from Plasmodium falciparum represents an important potential drug target due to its defensive role; hence ceasing the respective metabolic step will destroy the parasite. A three dimensional model of Plasmodium GS was constructed by de novo modelling method and potential GS inhibitors were identified from a library of glutathione (GSH) analogues retrieved from Ligand-info database and filtered using Lipinski and ADME rules. Two common feature pharmacophore models were generated from the individual inhibitor clusters to provide insight into the key pharmacophore features that are crucial for the GS inhibition. Molecular docking of selective compounds into the predicted GS binding site revealed that the compound CMBMB was the best GS inhibitor when compared to the standard reference Chloroquine (CQ). This was taken as indicating that CMBMB was the best effective and safest drug against P. falciparum.

This study was focused on evaluation of the cytotoxicity and apoptotic affects of benzofuran derivative on MCF-7 breast cancer cell line. This effective compound was isolated from the root of Petasites hybridus plant. For experiments, the MCF-7 cells were treated with several concentrations (0-500μM) of 1-(6-hydroxy-2- isopropenyl-1-benzofuran-5-yl)-1-ethanone 1 at different times. In this study, test of neutral red was also employed for cytotoxicity assay and quantity of P53, P21. Bax genes expression was analyzed using Real-Time PCR and ELISA techniques. Results show that compound 1 has cytotoxicity and apoptotic effects on Human breast cancer (Michigan Cancer Foundation-7) MCF-7 cells.

Rat renal tubular epithelial cell (RTEC) cultured with high glucose has been used to observe the protective effect of Ginkgo biloba extract (GBE) against diabetic nephropathy (DN). The compounds in GBE binding with cell membrane or entering into cell are still unknown, which may be potential bioactive components. In this paper, a powerful method for screening and analyzing the potential bioactive components from GBE was developed using cell extraction coupled with high performance liquid chromatography tandem mass spectrometry (LC-MS/MS). 8 prototype compounds and 5 metabolites were obtained, among which 6 prototype compounds and 1 metabolite were identified or tentatively characterized as rutin, bilobalide, ginkgolide B, ginkgolide C, genkwanin, apigenin and diosmetin by comparing their retention times and MS spectra with those of authentic standards or literature data. The 6 prototype compounds were further quantitatively analyzed using electrospray ionization in negative mode multiple reaction monitoring (MRM). The results showed that high glucose changed the Tmax, MRT(0-t), Cmax and AUC(0-t) of all observed compounds and decreased the t1/2 of genkwanin and apigenin, significantly. The overall findings indicate that 8 prototype compounds may be the potential bioactive components of GBE with preventive effect against DN and the method of RTEC extraction coupled with LC-MS/MS technology screening method we developed is a feasible, rapid, and useful tool for screening and analyzing potential bioactive components.