scholarly journals Optimization of a Higher Throughput Microsomal Stability Screening Assay for Profiling Drug Discovery Candidates

2003 ◽  
Vol 8 (4) ◽  
pp. 453-462 ◽  
Author(s):  
Li Di ◽  
Edward H. Kerns ◽  
Yan Hong ◽  
Teresa A. Kleintop ◽  
Oliver J. Mc Connell ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Drug Discovery ◽  
High Throughput ◽  
High Throughput Screening ◽  
Parent Compound ◽  
Metabolic Stability ◽  
Screening Assay ◽  
Drug Candidates ◽  
Increasing Demand ◽  
The Stability

Metabolic stability plays an important role in the success of drug candidates. First-pass metabolism is one of the major causes of poor oral bioavailability and short half-life. Traditionally, metabolic stability was evaluated at a later stage of drug discovery and required laborious manual manipulations. With the advance of high-throughput screening, combinatorial chemistry, and early profiling of drug-like properties, automated and rapid stability assays are needed to meet the increasing demand of throughput, speed, and reproducibility at earlier stages of drug discovery. The authors describe optimization of a simple, robust, high-throughput microsomal stability assay developed in a 96-well format. The assay consists of 2 automated components: robotic sample preparation for incubation and cleanup and rapid liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS) analysis to determine percent remaining of the parent compound. The reagent solutions and procedural steps were optimized for automation. Variables affecting assay results were investigated. The variability introduced by microsome preparations from different sources (various vendors and batches) was studied and indicates the need for careful control. Quality control and normalization of the stability results are critical when applying the screening data, generated at different times or research sites, to discovery projects.

High-Throughput Affinity Selection Mass Spectrometry Using SAMDI-MS to Identify Small-Molecule Binders of the Human Rhinovirus 3C Protease

2021 ◽  
pp. 247255522110232
Author(s):  
Michael D. Scholle ◽  
Doug McLaughlin ◽  
Zachary A. Gurard-Levin
Keyword(s):  
Mass Spectrometry ◽  
Drug Discovery ◽  
High Throughput ◽  
High Throughput Screening ◽  
Human Rhinovirus ◽  
Binding Potential ◽  
Affinity Selection ◽  
Response Curves ◽  
Desorption Ionization ◽  
Self Assembled

Affinity selection mass spectrometry (ASMS) has emerged as a powerful high-throughput screening tool used in drug discovery to identify novel ligands against therapeutic targets. This report describes the first high-throughput screen using a novel self-assembled monolayer desorption ionization (SAMDI)–ASMS methodology to reveal ligands for the human rhinovirus 3C (HRV3C) protease. The approach combines self-assembled monolayers of alkanethiolates on gold with matrix-assisted laser desorption ionization time-of-flight (MALDI TOF) mass spectrometry (MS), a technique termed SAMDI-ASMS. The primary screen of more than 100,000 compounds in pools of 8 compounds per well was completed in less than 8 h, and informs on the binding potential and selectivity of each compound. Initial hits were confirmed in follow-up SAMDI-ASMS experiments in single-concentration and dose–response curves. The ligands identified by SAMDI-ASMS were further validated using differential scanning fluorimetry (DSF) and in functional protease assays against HRV3C and the related SARS-CoV-2 3CLpro enzyme. SAMDI-ASMS offers key benefits for drug discovery over traditional ASMS approaches, including the high-throughput workflow and readout, minimizing compound misbehavior by using smaller compound pools, and up to a 50-fold reduction in reagent consumption. The flexibility of this novel technology opens avenues for high-throughput ASMS assays of any target, thereby accelerating drug discovery for diverse diseases.

scholarly journals A Novel High-Throughput Screening Assay for Sickle Cell Disease Drug Discovery

2009 ◽  
Vol 14 (4) ◽  
pp. 330-336 ◽  
Author(s):  
Eszter Pais ◽  
John S. Cambridge ◽  
Cage S. Johnson ◽  
Herbert J. Meiselman ◽  
Timothy C. Fisher ◽  
...  
Keyword(s):  
Sickle Cell Disease ◽  
Sickle Cell ◽  
High Throughput ◽  
High Throughput Screening ◽  
Rapid Screening ◽  
Cell Disease ◽  
Screening Assay ◽  
Drug Candidates ◽  
Test Molecule ◽  
High Throughput Screening Assay

Although the pathophysiology and molecular basis of sickle cell disease (SCD) were described more than half a century ago, an effective and safe therapy is not yet available. This may be explained by the lack of a suitable high-throughput technique that allows rapid screening of thousands of compounds for their antisickling effect. The authors have thus developed a novel high-throughput screening (HTS) assay based on detecting the ability of red blood cells (RBC) to traverse a column of tightly packed Sephacryl chromatography beads. When deoxygenated, sickle RBC are rigid and remain on the top of the column. However, when deoxygenated and treated with an effective antisickling agent, erythrocytes move through the Sephacryl media and produce a red dot on the bottom of the assay tubes. This approach has been adapted to wells in a 384-well microplate. Results can be obtained by optical scanning: The size of the red dot is proportional to the antisickling effect of the test molecule. The new assay is simple, inexpensive, reproducible, requires no special reagents, and should be readily adaptable to robotic HTS systems. It has the potential to identify novel drug candidates, allowing the development of new therapeutic options for individuals affected with SCD. ( Journal of Biomolecular Screening. 2009:330-336)

Development of a high-throughput screening assay for drug discovery in SOD1-mediated ALS

2009 ◽  
Vol 65 ◽  
pp. S120
Author(s):  
Gaku Murakami ◽  
Haruhisa Inoue ◽  
Kayoko Tsukita ◽  
Yasuyuki Asai ◽  
Kazuhiro Aiba ◽  
...  
Keyword(s):  
Drug Discovery ◽  
High Throughput ◽  
High Throughput Screening ◽  
Screening Assay ◽  
High Throughput Screening Assay

scholarly journals High-Throughput Docking and Molecular Dynamics Simulations towards the Identification of Potential Inhibitors against Human Coagulation Factor XIIa

2020 ◽  
Vol 2020 ◽  
pp. 1-13
Author(s):  
Dongfang Xu ◽  
Guangpu Xue ◽  
Bangya Peng ◽  
Zanjie Feng ◽  
Hongling Lu ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Md Simulation ◽  
Coagulation Factor ◽  
Inhibition Mechanism ◽  
Drug Candidates ◽  
Factor Xiia ◽  
The Stability ◽  
Dynamics Simulations ◽  
Potential Inhibitors

Human coagulation factor XIIa (FXIIa) is a trypsin-like serine protease that is involved in pathologic thrombosis. As a potential target for designing safe anticoagulants, FXIIa has received a great deal of interest in recent years. In the present study, we employed virtual high-throughput screening of 500,064 compounds within Enamine database to acquire the most potential inhibitors of FXIIa. Subsequently, 18 compounds with significant binding energy (from -65.195 to -15.726 kcal/mol) were selected, and their ADMET properties were predicted to select representative inhibitors. Three compounds (Z1225120358, Z432246974, and Z146790068) exhibited excellent binding affinity and druggability. MD simulation for FXIIa-ligand complexes was carried out to reveal the stability and inhibition mechanism of these three compounds. Through the inhibition of activated factor XIIa assay, we tested the activity of five compounds Z1225120358, Z432246974, Z45287215, Z30974175, and Z146790068, with pIC50 values of 9.3∗10−7, 3.0∗10−5, 7.8∗10−7, 8.7∗10−7, and 1.3∗10−6 M, respectively; the AMDET properties of Z45287215 and Z30974175 show not well but have better inhibition activity. We also found that compounds Z1225120358, Z45287215, Z30974175, and Z146790068 could be more inhibition of FXIIa than Z432246974. Collectively, compounds Z1225120358, Z45287215, Z30974175, and Z146790068 were anticipated to be promising drug candidates for inhibition of FXIIa.

A high-throughput screening assay for drug discovery in SOD1-mediated ALS targeting the transcription of SOD1

2010 ◽  
Vol 68 ◽  
pp. e311
Author(s):  
Gaku Murakami ◽  
Haruhisa Inoue ◽  
Kayoko Tsukita ◽  
Yasuyuki Asai ◽  
Kazuhiro Aiba ◽  
...  
Keyword(s):  
Drug Discovery ◽  
High Throughput ◽  
High Throughput Screening ◽  
Screening Assay ◽  
High Throughput Screening Assay
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