Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system

Abstract Caco-2 cells are widely used as an in vitro intestinal epithelial cell model because they can form a monolayer and predict drug absorption with high accuracy. However, Caco-2 cells hardly express cytochrome P450 (CYP), a drug-metabolizing enzyme. It is known that CYP3A4 is the dominant drug...

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Autores principales:	Moe Ichikawa, Hiroki Akamine, Michika Murata, Sumito Ito, Kazuo Takayama, Hiroyuki Mizuguchi
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/c0b45f99dce54b83a6c5de5e16934232
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spelling	oai:doaj.org-article:c0b45f99dce54b83a6c5de5e169342322021-12-02T15:56:49ZGeneration of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system10.1038/s41598-021-91160-z2045-2322https://doaj.org/article/c0b45f99dce54b83a6c5de5e169342322021-06-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-91160-zhttps://doaj.org/toc/2045-2322Abstract Caco-2 cells are widely used as an in vitro intestinal epithelial cell model because they can form a monolayer and predict drug absorption with high accuracy. However, Caco-2 cells hardly express cytochrome P450 (CYP), a drug-metabolizing enzyme. It is known that CYP3A4 is the dominant drug-metabolizing enzyme in human small intestine. In this study, we generated CYP3A4-expressing Caco-2 (CYP3A4-Caco-2) cells and attempted to establish a model that can simultaneously evaluate drug absorption and metabolism. CYP3A4-Caco-2 cells were generated by piggyBac transposon vectors. A tetracycline-controllable CYP3A4 expression cassette (tet-on system) was stably transduced into Caco-2 cells, thus regulating the levels of CYP3A4 expression depending on the doxycycline concentration. The CYP3A4 expression levels in CYP3A4-Caco-2 cells cultured in the presence of doxycycline were similar to or higher than those of adult small intestine. The CYP3A4-Caco-2 cells had enough ability to metabolize midazolam, a substrate of CYP3A4. CYP3A4 overexpression had no negative effects on cell proliferation, barrier function, and P-glycoprotein activity in Caco-2 cells. Thus, we succeeded in establishing Caco-2 cells with CYP3A4 metabolizing activity comparable to in vivo human intestinal tissue. This cell line would be useful in pharmaceutical studies as a model that can simultaneously evaluate drug absorption and metabolism.Moe IchikawaHiroki AkamineMichika MurataSumito ItoKazuo TakayamaHiroyuki MizuguchiNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-9 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Moe Ichikawa Hiroki Akamine Michika Murata Sumito Ito Kazuo Takayama Hiroyuki Mizuguchi Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system
description	Abstract Caco-2 cells are widely used as an in vitro intestinal epithelial cell model because they can form a monolayer and predict drug absorption with high accuracy. However, Caco-2 cells hardly express cytochrome P450 (CYP), a drug-metabolizing enzyme. It is known that CYP3A4 is the dominant drug-metabolizing enzyme in human small intestine. In this study, we generated CYP3A4-expressing Caco-2 (CYP3A4-Caco-2) cells and attempted to establish a model that can simultaneously evaluate drug absorption and metabolism. CYP3A4-Caco-2 cells were generated by piggyBac transposon vectors. A tetracycline-controllable CYP3A4 expression cassette (tet-on system) was stably transduced into Caco-2 cells, thus regulating the levels of CYP3A4 expression depending on the doxycycline concentration. The CYP3A4 expression levels in CYP3A4-Caco-2 cells cultured in the presence of doxycycline were similar to or higher than those of adult small intestine. The CYP3A4-Caco-2 cells had enough ability to metabolize midazolam, a substrate of CYP3A4. CYP3A4 overexpression had no negative effects on cell proliferation, barrier function, and P-glycoprotein activity in Caco-2 cells. Thus, we succeeded in establishing Caco-2 cells with CYP3A4 metabolizing activity comparable to in vivo human intestinal tissue. This cell line would be useful in pharmaceutical studies as a model that can simultaneously evaluate drug absorption and metabolism.
format	article
author	Moe Ichikawa Hiroki Akamine Michika Murata Sumito Ito Kazuo Takayama Hiroyuki Mizuguchi
author_facet	Moe Ichikawa Hiroki Akamine Michika Murata Sumito Ito Kazuo Takayama Hiroyuki Mizuguchi
author_sort	Moe Ichikawa
title	Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system
title_short	Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system
title_full	Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system
title_fullStr	Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system
title_full_unstemmed	Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system
title_sort	generation of tetracycline-controllable cyp3a4-expressing caco-2 cells by the piggybac transposon system
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/c0b45f99dce54b83a6c5de5e16934232
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Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system

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