Multiplex enzyme assays and inhibitor screening by mass spectrometry.
Current methods for high-throughput screening (HTS) use a serial process to evaluate compounds as inhibitors toward a single therapeutic target, but as the demand to reduce screening time and cost continues to grow, one solution is the development of multiplex technology. In this communication, the multiplex assay capability of a mass spectrometry (MS)-based readout system is verified using a kinase and esterase reaction simultaneously. Furthermore, the MS-based readout is shown to be compatible with a typical HTS workflow by identifying and validating several new inhibitors for each enzyme from a small library of compounds. These data confirm that it is possible to monitor inhibition of multiple therapeutic targets with one pass through the compound repository, thus demonstrating the potential for MS-based methods to become a method of choice for HTS of isolated enzymes.
Rathore
Rakesh
R
Department of Cancer and Cell Biology, University of Cincinnati College of Medicine, Cincinnati, OH 45237, USA.
Pribil
Patrick
P
Corr
Jay J
JJ
Seibel
William L
WL
Evdokimov
Artem
A
Greis
Kenneth D
KD
D000110:Acetylcholinesterase / Q000378:metabolism; D002138:Calibration; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D057075:Enzyme Assays / Q000379:methods* / Q000592:standards; D004791:Enzyme Inhibitors / Q000032:analysis* / Q000302:isolation & purification* / Q000494:pharmacology; D058851:GPI-Linked Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000379:methods* / Q000592:standards; D006801:Humans; D020128:Inhibitory Concentration 50; D013058:Mass Spectrometry / Q000379:methods* / Q000592:standards; D008954:Models, Biological; D054852:Small Molecule Libraries / Q000032:analysis
D004791:Enzyme Inhibitors; D058851:GPI-Linked Proteins; D054852:Small Molecule Libraries; D017868:Cyclic AMP-Dependent Protein Kinases; C548244:ACHE protein, human; D000110:Acetylcholinesterase