In vitro assay for quantitative prediction of human intestinal drug absorption with cryopreserved human intestinal mucosal epithelium-derived intestinal stem cells and 96-well Vitrigel inserts.

This study aimed to establish a novel in vitro model by combining jejunal stem cells from commercially available cryopreserved human intestinal mucosal epithelium (CHIM), which can be used without ethical concerns, with 96-well Vitrigel inserts for high-throughput screening. Jejunal stem cells were successfully isolated from CHIM, and stable long-term expansion in spheroid culture was achieved. The differentiated cells exhibited mRNA expression and function of major pharmacokinetic-related genes. Comparable metabolic and transport activities were maintained even with 96-well Vitrigel inserts instead of the conventional 24-well culture inserts. In a 96-well transcellular transport assay, a significant correlation was observed between the in vitro Papp values and the reported FaFg values (the fraction of orally administered drugs reaching the portal vein) in humans for 10 compounds known to be primarily absorbed via passive diffusion, allowing us to construct a fitting curve. Applying this curve, the FaFg values of transporter substrates could also be estimated with reasonably high accuracy. Furthermore, to estimate human Fg values (the fraction of orally administered drugs escaping from intestinal metabolism), we assessed 5 compounds known to be metabolized by intestinal enzymes. Their predicted Fg values closely matched the reported Fg ones, indicating that our cell model enables their accurate prediction. These findings suggest that a combination of CHIM-derived jejunal stem cells and 96-well Vitrigel inserts provides a practical in vitro model for the quantitative prediction of human intestinal absorption of orally administered drugs. SIGNIFICANCE STATEMENT: In this study, we demonstrated that a combination of commercially available cryopreserved human intestinal mucosal epithelium-derived intestinal stem cells, whose use is free from ethical concerns and restricted tissue availability, and 96-well Vitrigel inserts enables a high-throughput assay that quantitatively predicts human intestinal absorption with high accuracy.