Generation of Human-Induced Pluripotent Stem Cell-Derived Functional Enterocyte-Like Cells for Pharmacokinetic Studies

Shinpei Yoshida,Teruhiko Watanabe,Masaya Ishikawa,Takayuki Honjo,Nobuaki Shiraki,Sylvia Leo,Shoen Kume,Keita Iino,Ryunosuke Ishibe,Hiroyuki Kusuhara,Kazuya Maeda,Tomoka Shimada

doi:10.1016/j.stemcr.2020.12.017

Abstract

SummaryWe aimed to establish an in vitro differentiation procedure to generate matured small intestinal cells mimicking human small intestine from human-induced pluripotent stem cells (iPSCs). We previously reported the efficient generation of CDX2-expressing intestinal progenitor cells from embryonic stem cells (ESCs) using 6-bromoindirubin-3′-oxime (BIO) and (3,5-difluorophenylacetyl)-L-alanyl-L-2-phenylglycine tert-butyl ester (DAPT) to treat definitive endodermal cells. Here, we demonstrate the generation of enterocyte-like cells by culturing human iPSC-derived intestinal progenitor cells on a collagen vitrigel membrane (CVM) and treating cells with a simple maturation medium containing BIO, DMSO, dexamethasone, and activated vitamin D3. Functional tests further confirmed that these iPSC-derived enterocyte-like cells exhibit P-gp- and BCRP-mediated efflux and cytochrome P450 3A4 (CYP3A4)-mediated metabolism. We concluded that hiPS cell-derived enterocyte-like cells can be used as a model for the evaluation of drug transport and metabolism studies in the human small intestine.

Highlights

The small intestine is tasked with absorbing drugs as well as nutrients, ions, and water through its enterocytes
Collagen vitrigel supports the differentiation of human induced pluripotent stem cells (iPSCs) into intestinal enterocyte-like cells characterized by intestinal marker expression In this study, we used the collagen vitrigel membrane (CVM) in an attempt to induce the differentiation of human iPS cells into mature intestinal enterocytes
The results indicate that hiPSC-derived intestinal cells expressed intestinal markers, transporters, and cytochrome P450 (CYP) enzymes that resemble those of the intact human enterocytes

Summary

Introduction

The small intestine is tasked with absorbing drugs as well as nutrients, ions, and water through its enterocytes. Caco-2 cells form a tight monolayer and show drug uptake/efflux mediated by some transporter isoforms such as P-glycoprotein (P-gp), but the absolute expression levels of metabolic enzymes and transporters are often different from those in the intact intestinal tissue (Sun et al, 2002). Cytochrome P450 3A4 (CYP3A4) is recognized as a critical element for the drug metabolism in the intestine since CYP3A4 is most abundantly expressed in the small intestine among CYP isoforms and about half of the approved drugs are metabolized by CYP3A4. Previous reports indicated the clinical significance of CYP3A4 in the suppression of intestinal absorption of various drugs; Caco-2 cells lack CYP3A4 expression and cannot be used for the evaluation of the intestinal availability of drugs. A more appropriate in vitro model system for evaluating intestinal absorption of compounds in humans is needed

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