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Strategies for determining correct cytochrome P450 contributions in hepatic clearance predictions : in vitro-in vivo extrapolation as modelling approach and tramadol as proof-of concept compound

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Abstract
Background and Objective: Although the measurement of cytochrome P450 (CYP) contributions in metabolism assays is straightforward, determination of actual in vivo contributions might be challenging. How representative are in vitro for in vivo CYP contributions? This article proposes an improved strategy for the determination of in vivo CYP enzyme-specific metabolic contributions, based on in vitro data, using an in vitro-in vivo extrapolation (IVIVE) approach. Approaches are exemplified using tramadol as model compound, and CYP2D6 and CYP3A4 as involved enzymes. Methods: Metabolism data for tramadol and for the probe substrates midazolam (CYP3A4) and dextromethorphan (CYP2D6) were gathered in human liver microsomes (HLM) and recombinant human enzyme systems (rhCYP). From these probe substrates, an activity-adjustment factor (AAF) was calculated per CYP enzyme, for the determination of correct hepatic clearance contributions. As a reference, tramadol CYP contributions were scaled-back from in vivo data (retrograde approach) and were compared with the ones derived in vitro. In this view, the AAF is an enzyme-specific factor, calculated from reference probe activity measurements in vitro and in vivo, that allows appropriate scaling of a test drug's in vitro activity to the 'healthy volunteer' population level. Calculation of an AAF, thus accounts for any 'experimental' or 'batch-specific' activity difference between in vitro HLM and in vivo derived activity. Results: In this specific HLM batch, for CYP3A4 and CYP2D6, an AAF of 0.91 and 1.97 was calculated, respectively. This implies that, in this batch, the in vitro CYP3A4 activity is 1.10-fold higher and the CYP2D6 activity 1.97-fold lower, compared to in vivo derived CYP activities. Conclusion: This study shows that, in cases where the HLM pool does not represent the typical mean population CYP activities, AAF correction of in vitro metabolism data, optimizes CYP contributions in the prediction of hepatic clearance. Therefore, in vitro parameters for any test compound, obtained in a particular batch, should be corrected with the AAF for the respective enzymes. In the current study, especially the CYP2D6 contribution was found, to better reflect the average in vivo situation. It is recommended that this novel approach is further evaluated using a broader range of compounds.
Keywords
ISOLATED HEPATOCYTES, LIVER-MICROSOMES, METABOLISM, IMPACT, DRUGS

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Chicago
T’jollyn, Huybrecht, Jan Snoeys, Jan Van Bocxlaer, Lies De Bock, Pieter Annaert, Achiel Van Peer, Karel Allegaert, Geert Mannens, An Vermeulen, and Koen Boussery. 2017. “Strategies for Determining Correct Cytochrome P450 Contributions in Hepatic Clearance Predictions : in Vitro-in Vivo Extrapolation as Modelling Approach and Tramadol as Proof-of Concept Compound.” European Journal of Drug Metabolism and Pharmacokinetics 42 (3): 537–543.
APA
T’jollyn, H., Snoeys, J., Van Bocxlaer, J., De Bock, L., Annaert, P., Van Peer, A., Allegaert, K., et al. (2017). Strategies for determining correct cytochrome P450 contributions in hepatic clearance predictions : in vitro-in vivo extrapolation as modelling approach and tramadol as proof-of concept compound. EUROPEAN JOURNAL OF DRUG METABOLISM AND PHARMACOKINETICS, 42(3), 537–543.
Vancouver
1.
T’jollyn H, Snoeys J, Van Bocxlaer J, De Bock L, Annaert P, Van Peer A, et al. Strategies for determining correct cytochrome P450 contributions in hepatic clearance predictions : in vitro-in vivo extrapolation as modelling approach and tramadol as proof-of concept compound. EUROPEAN JOURNAL OF DRUG METABOLISM AND PHARMACOKINETICS. 2017;42(3):537–43.
MLA
T’jollyn, Huybrecht, Jan Snoeys, Jan Van Bocxlaer, et al. “Strategies for Determining Correct Cytochrome P450 Contributions in Hepatic Clearance Predictions : in Vitro-in Vivo Extrapolation as Modelling Approach and Tramadol as Proof-of Concept Compound.” EUROPEAN JOURNAL OF DRUG METABOLISM AND PHARMACOKINETICS 42.3 (2017): 537–543. Print.
@article{8548352,
  abstract     = {Background and Objective: Although the measurement of cytochrome P450 (CYP) contributions in metabolism assays is straightforward, determination of actual in vivo contributions might be challenging. How representative are in vitro for in vivo CYP contributions? This article proposes an improved strategy for the determination of in vivo CYP enzyme-specific metabolic contributions, based on in vitro data, using an in vitro-in vivo extrapolation (IVIVE) approach. Approaches are exemplified using tramadol as model compound, and CYP2D6 and CYP3A4 as involved enzymes. 
Methods: Metabolism data for tramadol and for the probe substrates midazolam (CYP3A4) and dextromethorphan (CYP2D6) were gathered in human liver microsomes (HLM) and recombinant human enzyme systems (rhCYP). From these probe substrates, an activity-adjustment factor (AAF) was calculated per CYP enzyme, for the determination of correct hepatic clearance contributions. As a reference, tramadol CYP contributions were scaled-back from in vivo data (retrograde approach) and were compared with the ones derived in vitro. In this view, the AAF is an enzyme-specific factor, calculated from reference probe activity measurements in vitro and in vivo, that allows appropriate scaling of a test drug's in vitro activity to the 'healthy volunteer' population level. Calculation of an AAF, thus accounts for any 'experimental' or 'batch-specific' activity difference between in vitro HLM and in vivo derived activity. 
Results: In this specific HLM batch, for CYP3A4 and CYP2D6, an AAF of 0.91 and 1.97 was calculated, respectively. This implies that, in this batch, the in vitro CYP3A4 activity is 1.10-fold higher and the CYP2D6 activity 1.97-fold lower, compared to in vivo derived CYP activities. 
Conclusion: This study shows that, in cases where the HLM pool does not represent the typical mean population CYP activities, AAF correction of in vitro metabolism data, optimizes CYP contributions in the prediction of hepatic clearance. Therefore, in vitro parameters for any test compound, obtained in a particular batch, should be corrected with the AAF for the respective enzymes. In the current study, especially the CYP2D6 contribution was found, to better reflect the average in vivo situation. It is recommended that this novel approach is further evaluated using a broader range of compounds.},
  author       = {T'jollyn, Huybrecht and Snoeys, Jan and Van Bocxlaer, Jan and De Bock, Lies and Annaert, Pieter and Van Peer, Achiel and Allegaert, Karel and Mannens, Geert and Vermeulen, An and Boussery, Koen},
  issn         = {0378-7966},
  journal      = {EUROPEAN JOURNAL OF DRUG METABOLISM AND PHARMACOKINETICS},
  language     = {eng},
  number       = {3},
  pages        = {537--543},
  title        = {Strategies for determining correct cytochrome P450 contributions in hepatic clearance predictions : in vitro-in vivo extrapolation as modelling approach and tramadol as proof-of concept compound},
  url          = {http://dx.doi.org/10.1007/s13318-016-0355-0},
  volume       = {42},
  year         = {2017},
}

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