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Simultaneously Assessing Concentration Changes in 17 Biochemical Pathways as a Result of Drug Dosing and Cytochrome P450 and Non-cytochrome P450-Mediated Metabolism: A Quasi-Untargeted Metabolomics LC/MS Assay

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Cytochrome P450

Part of the book series: Methods in Pharmacology and Toxicology ((MIPT))

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Abstract

From a number of cellular metabolism enzymes, including cytochrome P450 (CYP) and non-CYP enzymes, dozens of intracellular metabolites that are structurally related to a dosed drug molecule are produced. The drug or its metabolites potentially may cause adverse effects on the normal physiological function of one or more cell components, and thus possibly inducing toxicity. A targeted liquid chromatography, mass spectrometry (LC/MS) metabolomics analysis, which allows interrogation of 17 common biochemical pathways of human plasma, is described that allows for the investigation of possible toxicity in hepatic cell culture, animal, or human fluids. The assay utilizes three liquid chromatographic modes of separation, including ion pairing, reversed-phase, and hydrophobic interaction chromatography, to achieve quantitation of 93 metabolites. Plasma samples are prepared using a simple protein precipitation step followed by drying of supernatant and reconstitution in method-specific diluents. Quantitation is performed using both positive and negative ionization electrospray mass spectrometry in the range of endogenous concentrations. Results for six lots of plasma are reported and generally agree with published data. This method can provide rapid evaluation for off-target effects and adverse toxicity events that might be observed in preclinical or clinical studies supporting various therapeutic areas.

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Acknowledgments

The authors thank Drs. Wensheng Lang, Gregory Leo, and Allan Xu for helpful discussions. The views expressed here are solely those of the authors and do not reflect the opinions of Janssen Research & Development, LLC.

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Authors and Affiliations

  1. Janssen Research and Development LLC, Spring House, PA, USA

    John A. Masucci & Kerem Bingol

  2. Keystone Bioanalytical, West Point, PA, USA

    Feng Liang & Vince Windisch

  3. Blue Bell, PA, USA

    Gary W. Caldwell

Authors
  1. John A. Masucci
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  2. Feng Liang
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  3. Kerem Bingol
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  4. Vince Windisch
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  5. Gary W. Caldwell
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Corresponding author

Correspondence to John A. Masucci .

Editor information

Editors and Affiliations

  1. Genentech Inc., South San Francisco, CA, USA

    Zhengyin Yan

  2. Blue Bell, PA, USA

    Gary W. Caldwell

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Masucci, J.A., Liang, F., Bingol, K., Windisch, V., Caldwell, G.W. (2021). Simultaneously Assessing Concentration Changes in 17 Biochemical Pathways as a Result of Drug Dosing and Cytochrome P450 and Non-cytochrome P450-Mediated Metabolism: A Quasi-Untargeted Metabolomics LC/MS Assay. In: Yan, Z., Caldwell, G.W. (eds) Cytochrome P450. Methods in Pharmacology and Toxicology. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1542-3_21

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  • DOI: https://doi.org/10.1007/978-1-0716-1542-3_21

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1541-6

  • Online ISBN: 978-1-0716-1542-3

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