Moderate to high throughput in vitro binding kinetics for drug discovery.
This review provides a concise summary for state of the art, moderate to high throughput in vitro technologies being employed to study drug-target binding kinetics. These technologies cover a wide kinetic timescale spanning up to nine orders of magnitude from milliseconds to days. Automated stopped flow measures transient and (pre)steady state kinetics from milliseconds to seconds. For seconds to hours timescale kinetics we discuss surface plasmon resonance-based biosensor, global progress curve analysis for high throughput kinetic profiling of enzyme inhibitors and activators, and filtration plate-based radioligand or fluorescent binding assays for receptor binding kinetics. Jump dilution after pre-incubation is the preferred method for very slow kinetics lasting for days. The basic principles, best practices and simulated data for these technologies are described. Finally, the application of a universal label-free technology, liquid chromatography coupled tandem mass spectrometry (LC/MS/MS), is briefly reviewed. Select literature references are highlighted for in-depth understanding. A new reality is dawning wherein binding kinetics is an integral and routine part of mechanism of action elucidation and translational, quantitative pharmacology for drug discovery.
In vitro Pharmacology, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, New Jersey, rumin.zhang@merck.com.
Rumin
R
Zhang
In vitro Pharmacology, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, New Jersey, USA.
Christopher M
CM
Barbieri
In vitro Pharmacology, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, New Jersey, USA.
Margarita
M
Garcia-Calvo
In vitro Pharmacology, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, New Jersey, USA.
Robert W
RW
Myers
In vitro Pharmacology, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, New Jersey, USA.
David
D
McLaren
In vitro Pharmacology, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, New Jersey, USA.
Michael
M
Kavana
D015374:Biosensing Techniques / Q000379:methods; D055808:Drug Discovery / Q000379:methods*; D020536:Enzyme Activators / Q000737:chemistry / Q000493:pharmacokinetics* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000493:pharmacokinetics* / Q000494:pharmacology; D057166:High-Throughput Screening Assays / Q000379:methods*; D010599:Pharmacokinetics; D020349:Surface Plasmon Resonance / Q000379:methods; D053719:Tandem Mass Spectrometry
D020536:Enzyme Activators; D004791:Enzyme Inhibitors