Overlooked aspects of scaling enzyme activity through abundance across tissues and individuals: Preliminary Insights from kcat measurements in matched liver and intestinal samples

Prediction of human intestinal metabolism within physiologically-based pharmacokinetic models is now well established within drug development. Extrapolation of in vitro kinetic parameters accounts for differences in abundance between different in vitro systems and tissues. The existing data assume that the activity of cytochrome P450 (CYP)3A4 is consistent between intestine and liver once adjusted for its tissue-specific expression level. However, the validity of this assumption for other enzymes and other tissues remains uncertain. In the current study, indicators of ‘activity per unit of enzyme’, namely turnover number (kcat) or specificity constant (ksp), were measured for 7 enzymes (CYP2C9, CYP2C19, CYP2D6, CYP3A4, UGT1A6, UGT2B7, and UGT2B17) in microsomes prepared from 4 paired (same donor) intestine and liver tissue samples. After excluding one donor with low intestinal activity, the intestinal kcat and ksp for the studied CYPs were within 2-fold of the liver values, with the exception of one donor with 4-fold lower CYP2D6 kcat in the intestine compared with the liver. Conversely, the UGT1A1 ksp and UGT2B7 kcat were 5-fold and 7-fold higher in intestinal microsomes compared with liver microsomes, respectively. Trends in inter-donor variability in kcat were noted and require further evaluation in a larger set of donors. The current paradigm of extrapolation of hepatic metabolism data to predict in vivo first-pass metabolism in the intestine using tissue abundances appears to be valid for CYPs but should be approached with caution when predicting intestinal glucuronidation.