Induction of metabolism and transport in human intestine:Validation of precision-cut slices as a tool to study induction of drug metabolism in human intestine in vitro

Induction of drug enzyme activity in the intestine can strongly determine plasma levels of drugs. It is therefore important to pre-dict drug-drug interactions in human intestine in vitro. We evalu-ated the applicability of human intestinal precision-cut slices for induction studies in vitro. Morphological examination and intracel-lular ATP levels indicated tissue integrity up to 24 h of incubation, whereas in proximal jejunum slices, the metabolic rate toward most substrates remained at 40 to 50 % of initial values. In colon slices, the cytochrome P450 conversions were below the detection limit, but conjugation rates remained relatively constant during incubation. The inducibility of drug-metabolizing enzymes and P-glycoprotein was evaluated using prototypical inducers for five induction pathways. -Naphthoflavone (aryl hydrocarbon receptor ligand) induced CYP1A1 (132-fold in colon and 362-fold in proximal jejunum) and UDP glucuronosyltransferase (UGT) 1A

Innovative methods to study human intestinal drug metabolism in vitro:Precision-cut slices compared with Ussing chamber preparations

Predictive in vitro methods to investigate drug metabolism in the human intestine using intact tissue are of high importance. Therefore, we studied the metabolic activity of human small intestinal and colon slices and compared it with the metabolic activity of the same human intestinal segments using the Ussing chamber technique. The metabolic activity was evaluated using substrates to both phase I and phase II reactions: testosterone, 7-hydroxy-coumarin ( 7HC), and a mixture of cytochrome P450 ( P450) substrates ( midazolam, diclofenac, coumarin, and bufuralol). In slices of human proximal jejunum, the metabolic activity of several P450-mediated and conjugation reactions remained constant up to 4 h of incubation. In the colon slices, conjugation rates were virtually equal to those in small intestine, whereas P450-mediated conversions occurred much slower. In both organs, morphological evaluation and ATP content implied tissue integrity within this period. P450 conversions using the Ussing chamber technique showed that the metabolic rate ( sum of metabolites measured in apical, basolateral, and tissue compartments) was constant up to 3 h. For 7HC conjugations, the metabolic rate remained constant up to 4 h. The distribution of the metabolites in the compartments differed between the substrates. Overall, metabolic rates were surprisingly similar in both techniques and appear similar to or even higher than in liver. In conclusion, this study shows that both human intestinal precision-cut slices and Ussing chamber preparations provide useful tools for in vitro biotransformation studies

A proof of concept using the Ussing chamber methodology to study pediatric intestinal drug transport and age-dependent differences in absorption

Little is known about the impact of age on the processes governing human intestinal drug absorption. The Ussing chamber is a system to study drug transport across tissue barriers, but it has not been used to study drug absorption processes in children. This study aimed to explore the feasibility of the Ussing chamber methodology to assess pediatric intestinal drug absorption. Furthermore, differences between intestinal drug transport processes of children and adults were explored as well as the possible impact of age. Fresh terminal ileal leftover tissues from both children and adults were collected during surgery and prepared for Ussing chamber experiments. Paracellular (enalaprilat), transcellular (propranolol), and carrier-mediated drug transport by MDR1 (talinolol) and BCRP (rosuvastatin) were determined with the Ussing chamber methodology. We calculated apparent permeability coefficients and efflux ratios and explored their relationship with postnatal age. The success rate for the Ussing chamber experiments, as determined by electrophysiological measurements, was similar between children (58%, N = 15, median age: 44 weeks; range 8 weeks to 17 years) and adults (67%, N = 13). Mean serosal to mucosal transport of talinolol by MDR1 and rosuvastatin by BCRP was higher in adult than in pediatric tissues (p = 0.0005 and p = 0.0091). In contrast, within our pediatric cohort, there was no clear correlation for efflux transport across different ages. In conclusion, the Ussing chamber is a suitable model to explore pediatric intestinal drug absorption and can be used to further elucidate ontogeny of individual intestinal pharmacokinetic processes like drug metabolism and transport

No evidence for the involvement of the multidrug resistance-associated protein and/or the monocarboxylic acid transporter in the intestinal transport of fluvastatin in rats

Fluvastatin, an amphiphilic anion, shows a nonlinear increase in effective intestinal permeability (Peff) with increasing lumenal concentrations in rats. The main objective of this study was to investigate whether or not this observation could be attributed to an efflux-mediated transport by the multidrug resistance-associated protein (MRP). In parallel, we investigated the possible involvement of the monocarboxylic acid transporter (MCT) in the rapid intestinal absorption of fluvastatin. Single-pass perfusions were performed in the ileum and colon of the rat, with and without the presence of well-established inhibitors/substrates for the MRP (probenecid) and the MCT (nicotinic acid). The results suggest that neither the MRP nor the MCT are involved to any significant extent in the absorption process of fluvastatin in the rat intestine. Thus, the previously reported concentration-dependent Peff of fluvastatin in these intestinal regions of the rat is probably not attributable to saturation of any efflux mediated by MRP