(M0930-05-33) A Gut-Hepatobiliary Organ Perfusion Model: Demonstrator Study Showing the Absorption, Metabolism, and Excretion of Midazolam

Purpose: In order to determine the oral bioavailability of a drug, characterization of the absorption, metabolism, distribution and excretion (ADME) is of great importance. To date, complete ADME profiles can only be studied in in vivo models as it remains difficult to recapitulate all organ functions within a single model. The current developments in organ perfusion techniques, opens the possibility to study physiologic processes in an ex vivo environment. Aim of this study was to explore the feasibility of multi-organ perfusion model the first-pass effect and subsequently oral bioavailability of midazolam.

Methods: Porcine en-bloc organs were procured from a slaughterhouse (n=4). After obtaining the organs, the lungs and heart were dissected and the aorta was cannulated, ligated and a cold flush (HTK) was initiated. In the meantime the colon was dissected. Thereafter, the organs were flushed with HTK and an additional portal flush was applied. Upon arrival in the laboratory, the stomach was dissected and the small intestine was shortened to ~2 meter. The organs were perfused via the aorta (60-80 mmHg) and portal vein (8-11 mmHg) with a red blood cell based perfusate. After 120 min of normothermic perfusion, 20 mg of midazolam was dosed via the duodenum. Perfusate, portal vein, bile and tissue samples were taken in time. Hourly blood gas analysis was performed to monitor organ viability.

Results: After intraduodenal administration, midazolam was quickly detected in the portal vein along with 1-OH midazolam (E_G-pv of 0.16±0.1), indicating gut wall metabolism through oxidation. In the intestinal fecal effluent, 1-OH midazolam and 1-OH midazolam glucuronide (E_G-intestine 0.051±0.03) were observed, resulting from local gut glucuronidation. Biliary elimination of midazolam (0.04±0.01%) and its glucuronide (0.01±0.01%) contributed minimally to the enterohepatic circulation. Hepatic metabolism (F_H 0.35±0.07) was more extensive than intestinal metabolism (F_G 0.78±0.11), leading to an oral bioavailability of 0.27±0.05.

Conclusion: We have successfully applied ex vivo multi-organ perfusion as a novel method for characterizing the pre-systemic extraction of midazolam by measuring both intestinal and hepatic extraction. This model can provide valuable insights into the absorption and metabolism of new drugs. Additional innovations could include a predigestion protocol to more accurately stimulate gastric digestion and including additional intestinal regions with varying expression of transporters and metabolic enzymes can better simulate the in vivo situation. Ex vivo multi-organ perfusion, complemented with physiologically based pharmacokinetic modelling is a valuable approach to investigate the first-pass effect and oral bioavailability of novel pharmaceutical compounds in human subjects.

References: Stevens, L J et al. “Ex vivo gut-hepato-biliary organ perfusion model to characterize oral absorption, gut-wall metabolism, pre-systemic hepatic metabolism and biliary excretion; application to midazolam.” European journal of pharmaceutical sciences: official journal of the European Federation for Pharmaceutical Sciences vol. 196 (2024): 106760. doi:10.1016/j.ejps.2024.106760