Mass Spectrometry-Based Quantification of CYP Enzymes to Establish In Vitro/In Vivo Scaling Factors for Intestinal and Hepatic Metabolism in Beagle Dog

• Published in: Pharmaceutical research Vol. 29; no. 7; pp. 1832 - 1842
• Main Authors: Heikkinen, Aki T., Friedlein, Arno, Lamerz, Jens, Jakob, Peter, Cutler, Paul, Fowler, Stephen, Williamson, Tara, Tolando, Roberto, Lave, Thierry, Parrott, Neil
• Format: Journal Article
• Language: English
• Published: Boston Springer US 01.07.2012; Springer; Springer Nature B.V
• Subjects: Amino Acid Sequence; Animals; Aryl Hydrocarbon Hydroxylases - analysis; Aryl Hydrocarbon Hydroxylases - metabolism; Biochemistry; Biological and medical sciences; Biomedical and Life Sciences; Biomedical Engineering and Bioengineering; Biomedicine; Cytochrome P-450 Enzyme System - analysis; Cytochrome P-450 Enzyme System - metabolism; Cytochrome P450 Family 2; Dogs; Enzymes; General pharmacology; Intestines - chemistry; Intestines - enzymology; Liver - chemistry; Liver - enzymology; Mass Spectrometry; Medical Law; Medical sciences; Metabolism; Microsomes - chemistry; Microsomes - enzymology; Molecular Sequence Data; Pharmaceutical technology. Pharmaceutical industry; Pharmacology. Drug treatments; Pharmacology/Toxicology; Pharmacy; Research Paper; Steroid Hydroxylases - analysis; Steroid Hydroxylases - metabolism; MRM; scaling; CYP; beagle dog; intestinal metabolism; /; Fissipedia; Carnivora; Pharmaceutical technology; Enzyme; Liver; Gut; Metabolism; In vitro; In vivo; Vertebrata; Mammalia; Animal; in vitro/in vivo scaling; Pharmacokinetics; Dog; Mass spectrometry
• ISSN: 0724-8741; 1573-904X
• DOI: 10.1007/s11095-012-0707-7

ABSTRACT Purpose Physiologically based models, when verified in pre-clinical species, optimally predict human pharmacokinetics. However, modeling of intestinal metabolism has been a gap. To establish in vitro/in vivo scaling factors for metabolism, the expression and activity of CYP enzymes were characterized in the intestine and liver of beagle dog. Methods Microsomal protein abundance in dog tissues was determined using testosterone-6β-hydroxylation and 7-hydroxycoumarin-glucuronidation as markers for microsomal protein recovery. Expressions of 7 CYP enzymes were estimated based on quantification of proteotypic tryptic peptides using multiple reaction monitoring mass spectrometry. CYP3A12 and CYP2B11 activity was evaluated using selective marker reactions. Results The geometric mean of total microsomal protein was 51 mg/g in liver and 13 mg/cm in intestine, without significant differences between intestinal segments. CYP3A12, followed by CYP2B11, were the most abundant CYP enzymes in intestine. Abundance and activity were higher in liver than intestine and declined from small intestine to colon. Conclusions CYP expression in dog liver and intestine was characterized, providing a basis for in vitro/in vivo scaling of intestinal and hepatic metabolism.