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Recent advances have made modeling human small intestines in vitro possible, but it remains a challenge to recapitulate fully their structural and functional characteristics. We suspected interstitial flow within the intestine, powered by circulating blood plasma during embryonic organogenesis, to be a vital factor. We aimed to construct an in vivo-like multilayered small intestinal tissue by incorporating interstitial flow into the system and, in turn, developed the micro-small intestine system by differentiating definitive endoderm and mesoderm cells from human pluripotent stem cells simultaneously on a microfluidic device capable of replicating interstitial flow. This approach enhanced cell maturation and led to the development of a three-dimensional small intestine-like tissue with villi-like epithelium and an aligned mesenchymal layer. Our micro-small intestine system not only overcomes the limitations of conventional intestine models but also offers a unique opportunity to gain insights into the detailed mechanisms underlying intestinal tissue development., Competing Interests: Declaration of interests K.T. and S.D. are inventors on JP patent application no. 2023-132372 “Construction of micro-small intestine systems”. K.T. receives research funds for developing micro-small intestine systems under the contract with Kanto Chemical Co., Inc. M.I. is a scientific adviser for xFOREST Therapeutics without salary., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Renal transporters play critical roles in predicting potential drug-drug interactions. However, current in vitro models often fail to adequately express these transporters, particularly solute carrier proteins, including organic anion transporters (OAT1/3), and organic cation transporter 2 (OCT2). Here, we developed a hiPSC-derived kidney organoids-based proximal tubule-on-chip (OPTC) model that emulates in vivo renal physiology to assess transporter function. Compared to chips based on immortalized cells, OPTC derived from the two most commonly used differentiation protocols exhibited significant improvement in expression level and polarity of OAT1/3 and OCT2. Hence, the OPTC demonstrates enhanced functionality in efflux and uptake assessments, and nephrotoxicity. Furthermore, these functionalities are diminished upon adding inhibitors during substrate-inhibitor interactions, which were closer to in vivo observations. Overall, these results support that OPTC can reliably assess the role of renal transporters in drug transport and nephrotoxicity, paving the way for personalized models to assess renal transport and disease modeling., Competing Interests: R.Y., C.M., R.B.S., K.F., T.A., M.T., and R.N. are inventors on JP patent application no. 2024–73489 “Construction of proximal tubules micro-physiological system”., (© 2024 The Author(s).)

In the evolving field of drug discovery and development, multiorgans-on-a-chip and microphysiological systems are gaining popularity owing to their ability to emulate in vivo biological environments. Among the various gut-liver-on-a-chip systems for studying oral drug absorption, the chip developed in this study stands out with two distinct features: incorporation of perfluoropolyether (PFPE) to effectively mitigate drug sorption and a unique enterohepatic single-passage system, which simplifies the analysis of first-pass metabolism and oral bioavailability. By introducing a bolus drug injection into the liver compartment, hepatic extraction alone could be evaluated, further enhancing our estimation of intestinal availability. In a study on midazolam (MDZ), PFPE-based chips showed more than 20-times the appearance of intact MDZ in the liver compartment effluent compared to PDMS-based counterparts. Notably, saturation of hepatic metabolism at higher concentrations was confirmed by observations when the dose was reduced from 200 μM to 10 μM. This result was further emphasized when the metabolism was significantly inhibited by the coadministration of ketoconazole. Our chip, which is designed to minimize the dead volume between the gut and liver compartments, is adept at sensitively observing the saturation of metabolism and the effect of inhibitors. Using genome-edited CYP3A4/UGT1A1-expressing Caco-2 cells, the estimates for intestinal and hepatic availabilities were 0.96 and 0.82, respectively; these values are higher than the known human in vivo values. Although the metabolic activity in each compartment can be further improved, this gut-liver-on-a-chip can not only be used to evaluate oral bioavailability but also to carry out individual assessment of both intestinal and hepatic availability.

CYP2D6 variants contain various single nucleotide polymorphisms as well as differing levels of metabolic activity. Among these, one of the less active variants CYP2D6*10 (100C > T) is the most prevalent mutation in East Asians, including Japanese. This mutation leads to an amino acid substitution from proline to serine, which reduces the stability of CYP2D6 and consequently decreases its metabolic activity. In this study, we used a genome editing technology called the Precise Integration into Target Chromosome (PITCh) system to stably express six drug-metabolizing enzymes (CYP3A4, POR, uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1), CYP1A2, CYP2C19, CYP2C9, and CYP2D6*10) in HepG2 (CYP2D6*10 KI-HepG2) cells to examine the effect of CYP2D6*10 on drug metabolism prediction. The protein expression levels of CYP2D6 in CYP2D6*10 KI-HepG2 cells were reduced relative to those in the CYP3A4-POR-UGT1A1-CYP1A2-CYP2C19-CYP2C9-CYP2D6 knock-in-HepG2 (CYPs-UGT1A1 KI-HepG2) cells. Consistent with the CYP2D6 protein expression results, CYP2D6 metabolic activity in CYP2D6*10 KI-HepG2 cells was reduced relative to CYPs-UGT1A1 KI-HepG2 cells. We successfully generated CYP2D6*10 KI-HepG2 cells with highly expressed, functional CYP2D6*10, as well as CYP1A2, 2C9, 2C19 and 3A4. CYP2D6*10 KI-HepG2 cells could be an invaluable model for hepatic metabolism and hepatotoxicity studies in East Asians, including Japanese.