Generation of Enterocyte-Like Cells with Pharmacokinetic Functions from Human Induced Pluripotent Stem Cells Using Small-Molecule Compounds

Abstract

The small intestine plays an important role in all aspects of pharmacokinetics, but there is no system for the comprehensive evaluation of small-intestinal pharmacokinetics, including drug metabolism and absorption. In this study, we aimed to construct an intestinal pharmacokinetics evaluation system and to generate pharmacokinetically functional enterocytes from human induced pluripotent stem cells. Using activin A and fibroblast growth factor 2, we differentiated these stem cells into intestinal stem cell-like cells, and the resulting cells were differentiated into enterocytes in a medium containing epidermal growth factor and small-molecule compounds. The differentiated cells expressed intestinal marker genes and drug transporters. The expression of sucrase-isomaltase, an intestine-specific marker, was markedly increased by small-molecule compounds. The cells exhibited activities of drug-metabolizing enzymes expressed in enterocytes, including CYP1A1/2, CYP2C9, CYP2C19, CYP2D6, CYP3A4/5, UGT, and sulfotransferase. Fluorescence-labeled dipeptide uptake into the cells was observed and was inhibited by ibuprofen, an inhibitor of the intestinal oligopeptide transporter solute carrier 15A1/PEPT1. CYP3A4 mRNA expression level was increased by these compounds and induced by the addition of 1a,25-dihydroxyvitamin D 3 . CYP3A4/5 activity was also induced by 1a,25-dihydroxyvitamin D 3 in cells differentiated in the presence of the compounds. All these results show that we have generated enterocyte-like cells that have pharmacokinetic functions, and we have identified small-molecule compounds that are effective for promoting intestinal differentiation and the gain of pharmacokinetic functions. Our enterocyte-like cells would be useful material for developing a novel evaluation system to predict human intestinal pharmacokinetics.

Characteristic Analysis of Intestinal Transport in Enterocyte-Like Cells Differentiated from Human Induced Pluripotent Stem Cells

“…We previously reported that human iPS cell-derived enterocyte-like cells had metabolic functions, such as drug-metabolizing enzyme activities and CYP3A4 inducibility (Iwao et al, 2015). Moreover, in this study, we demonstrated the drug transport characteristics in the enterocyte-like cells.…”

“…Moreover, it was indicated that the F a of drugs in humans can be estimated from permeability data of the enterocyte-like cell membrane. In our previous study, we found that the differentiated cells performed drug-metabolizing enzyme activities and CYP3A4 inducibility (Iwao et al, 2015). Taken together, the enterocyte-like cells may be useful as an appropriate model to comprehensively predict drug transport and metabolism in the intestine.…”

“…The human iPS cells were differentiated into enterocytes on the basis of our previous report (Iwao et al, 2015). Briefly, human iPS cells were differentiated into endodermal cells by incubating the cells in the presence of 100 ng/ml activin A for 72 hours, and these endodermal-like cells were then differentiated into intestinal stem cells via 250 ng/ml FGF2 for 96 hours.…”

“…We have previously established a method for differentiating human iPS cells into enterocytes (Iwao et al, , 2015 and found several small-molecule compounds that were effective in promoting the differentiation of human iPS cells. In addition, these enterocytes were associated with a gain of pharmacokinetic function.…”

Generation of intestinal organoids derived from human pluripotent stem cells for drug testing

“…The hiPSCs can serve as an attractive tool for drug development and studies of disease mechanisms 14 . Recently, differentiation procedures into IECs from hESCs or hiPSCs have been reported [8][9][10]15 . Human iPSCs possess high proliferative activity and can be differentiated into preferred cell types.…”