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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)
 
 
 

A quantitative high-throughput trapping assay as a measurement of potential for bioactivation.

Idiosyncratic adverse drug reactions (ADRs) are one of the most common causes of pharmaceutical withdrawals and labeling changes. Most ADRs are caused by drugs that form reactive species that can bind covalently to macromolecules such as proteins. The current methodology for the measurement of covalent binding relies on the use of radiolabeled material that requires an investment in time and resources not typically expended until later in the discovery process. Efforts are also made to identify reactive intermediates by the use of chemical trapping agents, such as reduced glutathione and cyanide, to form stable adducts that are characterized by liquid chromatography-tandem mass spectrometry and/or nuclear magnetic resonance spectroscopy. Here, we describe a high-throughput assay for the measurement of reactive intermediate formation. The method involves incubation of cold compound with liver microsomes in the presence of [14C]potassium cyanide. Hard electrophilic species would react with the trapping agent, resulting in the formation of a radiolabeled conjugate. Unreacted trapping agent is removed using solid-phase extraction, and the amount of radiolabeled conjugate present is determined by liquid scintillation counting. This newly developed screen has proved to be specific, sensitive, robust, and a powerful tool for assessing bioactivation potential.[1]

References

  1. A quantitative high-throughput trapping assay as a measurement of potential for bioactivation. Meneses-Lorente, G., Sakatis, M.Z., Schulz-Utermoehl, T., De Nardi, C., Watt, A.P. Anal. Biochem. (2006) [Pubmed]
 
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