Abstract
We describe here a high-throughput assay to support rapid evaluation of drug discovery compounds for possible drug–drug interaction (DDI). Each compound is evaluated for its DDI potential by incubating over a range of eight concentrations and against a panel of six cytochrome P450 (CYP) enzymes: 1A2, 2C8, 2C9, 2C19, 2D6, and 3A4. The method utilizes automated liquid handling for sample preparation, and online solid-phase extraction/tandem mass spectrometry (SPE/MS/MS) for sample analyses. The system is capable of generating two 96-well assay plates in 30 min, and completes the data acquisition and analysis of both plates in about 30 min. Many laboratories that perform the CYP inhibition screening automate only part of the processes leaving a throughput bottleneck within the workflow. The protocols described in this chapter are aimed to streamline the entire process from assay to data acquisition and processing by incorporating automation and utilizing high-precision instrument to maximize throughput and minimize bottleneck.
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Acknowledgements
We would like to thank Ms. Melinda Manuel, Mr. Joshua Cramlett, and Dr. Beverly Knight from Takeda San Diego for their support in the development and testing of the high-throughput P450 inhibition assay.
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Lim, K.B., Ozbal, C.C., Kassel, D.B. (2013). High-Throughput Mass Spectrometric Cytochrome P450 Inhibition Screening. In: Phillips, I., Shephard, E., Ortiz de Montellano, P. (eds) Cytochrome P450 Protocols. Methods in Molecular Biology, vol 987. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-321-3_3
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DOI: https://doi.org/10.1007/978-1-62703-321-3_3
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Publisher Name: Humana Press, Totowa, NJ
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Online ISBN: 978-1-62703-321-3
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