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The ontogeny of hepatic CYP3A, CYP2C and CYP2E enzyme activity in conventional pigs

Joske Millecam (UGent) , Mathias Devreese (UGent) , Lies De Bock, Jan Van Bocxlaer (UGent) , Elke Gasthuys (UGent) , Patrick De Backer (UGent) and Siska Croubels (UGent)
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  • IWT-SBO-141427
Abstract
Introduction: The anatomy and physiology of pigs is closely related to human characteristics. Therefore, the use of pigs as an animal model to study the pharmacokinetic (PK) behavior of drugs in therapeutic subpopulations, including pediatrics, could be of interest. One of the key PK processes, biotransformation, is primarily mediated by the cytochrome P450 (CYP) enzyme system. Literature reports have demonstrated a high homology between human and porcine CYP3A, 2C and 2E in adults, namely at least 75, 62 and 79% amino acid sequence identity, respectively. However, data regarding the ontogeny of porcine CYP enzymes are lacking. Therefore, in order to assess whether piglets might serve as a model for pediatric PK studies, knowledge regarding the ontogeny of the CYP enzymes in pigs is mandatory. Materials and methods: Liver samples were collected immediately after euthanasia from 16 pigs (8 males and 8 females, Hybrid sow x Piétrain boar) of different ages (2 days, 4 and 8 weeks, 6 months-old). Samples were snap-frozen and stored at <-80°C until analysis. Microsomes were prepared by a differential centrifugation method. Midazolam, tolbutamide and chlorzoxazone probe drugs were used to determine the in vitro CYP3A, 2C and 2E catalytic activity, respectively. The corresponding metabolites, namely 1-hydroxy-midazolam, 4-hydroxy-tolbutamide and 6-hydroxy-chlorzoxazone, were quantified using a validated UHPLC-MS/MS method (1). Furthermore, the microsomal protein per gram of liver was determined as it is an important scaling factor in the extrapolation of the obtained in vitro enzyme activities to in vivo (2). Results and conclusions: The biotransformation of midazolam, tolbutamide and chlorzoxazone increased with age. The mean (±SD) CYP3A activity was 60.5 (±45.7) and 83.3 (±20.7) pmol/min/mg protein at the age of 2 days, 971.1 (±367.8) and 1072.7 (±371.7) pmol/min/mg protein at the age of 4 weeks and 723.4 (±146.3) and 1134.7 (±282.6) pmol/min/mg protein at 8 weeks of age for the barrows and sows, respectively. CYP2C activity at the same ages increased from 20.1 (±12.3) and 29.1 (±18.5) to 78.3 (±25.6) and 106.7 (±69.1) and 103.5 (±39.6) and 170.2 (±71.9) pmol/min/mg protein, while the activity of CYP2E was 539.3 (±251.0) and 643.3 (±220.3), 747.7 (±134.8) and 948.9 (±246.2) and 957.9 (±221.7) and 1549.8 (±345.0) pmol/min/mg protein, respectively for the barrows and sows. Significant sex differences (P<0.05) were only observed at 8 weeks of age. These data show similar trends with human CYP ontogeny.
Keywords
cytochrome P450, ontogony, pigs

Citation

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MLA
Millecam, Joske, et al. “The Ontogeny of Hepatic CYP3A, CYP2C and CYP2E Enzyme Activity in Conventional Pigs.” JOURNAL OF VETERINARY PHARMACOLOGY AND THERAPEUTICS, edited by Jim Riviere and Johanna Fink-Gremmels, vol. 38, no. suppl. 1, 2015, pp. 48–49.
APA
Millecam, J., Devreese, M., De Bock, L., Van Bocxlaer, J., Gasthuys, E., De Backer, P., & Croubels, S. (2015). The ontogeny of hepatic CYP3A, CYP2C and CYP2E enzyme activity in conventional pigs. In J. Riviere & J. Fink-Gremmels (Eds.), JOURNAL OF VETERINARY PHARMACOLOGY AND THERAPEUTICS (Vol. 38, pp. 48–49). Nantes, France.
Chicago author-date
Millecam, Joske, Mathias Devreese, Lies De Bock, Jan Van Bocxlaer, Elke Gasthuys, Patrick De Backer, and Siska Croubels. 2015. “The Ontogeny of Hepatic CYP3A, CYP2C and CYP2E Enzyme Activity in Conventional Pigs.” In JOURNAL OF VETERINARY PHARMACOLOGY AND THERAPEUTICS, edited by Jim Riviere and Johanna Fink-Gremmels, 38:48–49.
Chicago author-date (all authors)
Millecam, Joske, Mathias Devreese, Lies De Bock, Jan Van Bocxlaer, Elke Gasthuys, Patrick De Backer, and Siska Croubels. 2015. “The Ontogeny of Hepatic CYP3A, CYP2C and CYP2E Enzyme Activity in Conventional Pigs.” In JOURNAL OF VETERINARY PHARMACOLOGY AND THERAPEUTICS, ed by. Jim Riviere and Johanna Fink-Gremmels, 38:48–49.
Vancouver
1.
Millecam J, Devreese M, De Bock L, Van Bocxlaer J, Gasthuys E, De Backer P, et al. The ontogeny of hepatic CYP3A, CYP2C and CYP2E enzyme activity in conventional pigs. In: Riviere J, Fink-Gremmels J, editors. JOURNAL OF VETERINARY PHARMACOLOGY AND THERAPEUTICS. 2015. p. 48–9.
IEEE
[1]
J. Millecam et al., “The ontogeny of hepatic CYP3A, CYP2C and CYP2E enzyme activity in conventional pigs,” in JOURNAL OF VETERINARY PHARMACOLOGY AND THERAPEUTICS, Nantes, France, 2015, vol. 38, no. suppl. 1, pp. 48–49.
@inproceedings{6890416,
  abstract     = {Introduction: The anatomy and physiology of pigs is closely related to human characteristics. Therefore, the use of pigs as an animal model to study the pharmacokinetic (PK) behavior of drugs in therapeutic subpopulations, including pediatrics, could be of interest. One of the key PK processes, biotransformation, is primarily mediated by the cytochrome P450 (CYP) enzyme system. Literature reports have demonstrated a high homology between human and porcine CYP3A, 2C and 2E in adults, namely at least 75, 62 and 79% amino acid sequence identity, respectively. However, data regarding the ontogeny of porcine CYP enzymes are lacking. Therefore, in order to assess whether piglets might serve as a model for pediatric PK studies, knowledge regarding the ontogeny of the CYP enzymes in pigs is mandatory.
Materials and methods: Liver samples were collected immediately after euthanasia from 16 pigs (8 males and 8 females, Hybrid sow x Piétrain boar) of different ages (2 days, 4 and 8 weeks, 6 months-old). Samples were snap-frozen and stored at <-80°C until analysis. Microsomes were prepared by a differential centrifugation method. Midazolam, tolbutamide and chlorzoxazone probe drugs were used to determine the in vitro CYP3A, 2C and 2E catalytic activity, respectively. The corresponding metabolites, namely 1-hydroxy-midazolam, 4-hydroxy-tolbutamide and 6-hydroxy-chlorzoxazone, were quantified using a validated UHPLC-MS/MS method (1). Furthermore, the microsomal protein per gram of liver was determined as it is an important scaling factor in the extrapolation of the obtained in vitro enzyme activities to in vivo (2).
Results and conclusions: The biotransformation of midazolam, tolbutamide and chlorzoxazone increased with age. The mean (±SD) CYP3A activity was 60.5 (±45.7) and 83.3 (±20.7) pmol/min/mg protein at the age of 2 days, 971.1 (±367.8) and 1072.7 (±371.7) pmol/min/mg protein at the age of 4 weeks and 723.4 (±146.3) and 1134.7 (±282.6) pmol/min/mg protein at 8 weeks of age for the barrows and sows, respectively. CYP2C activity at the same ages increased from 20.1 (±12.3) and 29.1 (±18.5) to 78.3 (±25.6) and 106.7 (±69.1) and 103.5 (±39.6) and 170.2 (±71.9) pmol/min/mg protein, while the activity of CYP2E was 539.3 (±251.0) and 643.3 (±220.3), 747.7 (±134.8) and 948.9 (±246.2) and 957.9 (±221.7) and 1549.8 (±345.0) pmol/min/mg protein, respectively for the barrows and sows. Significant sex differences (P<0.05) were only observed at 8 weeks of age. These data show similar trends with human CYP ontogeny.},
  articleno    = {abstract 16.4},
  author       = {Millecam, Joske and Devreese, Mathias and De Bock, Lies and Van Bocxlaer, Jan and Gasthuys, Elke and De Backer, Patrick and Croubels, Siska},
  booktitle    = {JOURNAL OF VETERINARY PHARMACOLOGY AND THERAPEUTICS},
  editor       = {Riviere, Jim and Fink-Gremmels, Johanna},
  issn         = {0140-7783},
  keywords     = {cytochrome P450,ontogony,pigs},
  language     = {eng},
  location     = {Nantes, France},
  number       = {suppl. 1},
  pages        = {abstract 16.4:48--abstract 16.4:49},
  title        = {The ontogeny of hepatic CYP3A, CYP2C and CYP2E enzyme activity in conventional pigs},
  url          = {http://dx.doi.org/10.1111/jvp.12246},
  volume       = {38},
  year         = {2015},
}

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