Life science
Medical & heathcare, Drug discovery

Monolayer platform using human biopsy-derived small
intestinal organoids for pharmaceutical researchLaboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences
Professor Hiroyuki Mizuguchi
Researchmap https://researchmap.jp/read0165079

The human small intestine is the key organ for absorption, metabolism, and excretion of orally administered drugs. To preclinically predict these reactions in drug discovery research, a cell modelthat can precisely recapitulate the in vivo human intestinal monolayer is desired. Here, we developed a monolayer platform using
human biopsy-derived small intestinal organoids for application topharmacokinetic studies. The human small intestinal organoid-derived monolayer was prepared by a simple method in 3–8 days. It
consisted of polarized absorptive cells and had tight junctions. It
showed much higher drug-metabolizing enzyme activities than theexisting models (Caco-2 cells). It also showed transporter activity.
Based on these findings, this monolayer assay system using biopsy-derived human small intestinal organoids is likely to be widelyadopted.

▍ Significance of the research and Future perspective
The human small intestinal organoid-derived monolayer canrecapitulate the characteristics of the in vivo small intestine, andtherefore this monolayer could be a powerful tool to predict thepharmacokinetics of orally administered drugs.

▍ Background & Results
The human small intestine plays a central role in the pharmacokinetics of orally administrated drugs, which account for around
70% of FDA-approved drugs (U. S. Food and Drug Administration,2019). The process of absorbing, metabolizing, and excreting oforally administered drugs begins in the small intestine. These series of reactions are called the first-pass effect and are an essentialconsideration in drug discovery research. An in vitro system torecapitulate the human in vivo drug absorption, metabolism, andexcretion is crucially important for the structural design of compounds in the early stages of drug discovery research.
It has been difficult to culture primary human intestinal epithelial cells for a long period without diminishing their functions andviability. Therefore, small intestinal tissues obtained from rodents
or a human colon cancer-derived cell line, i.e., Caco-2 cells, havebeen broadly used and have made a major contribution to pharmacokinetic studies. Nonetheless, the former has the problem ofspecies differences, and the latter has the problem that some ofthe drug-metabolizing enzymes and drug transporters are absent
or poorly expressed in Caco-2 cells. To solve these problems, analternative system is urgently needed.
We successfully produced a monolayer from biopsy-derived
human small intestinal organoids by means of a very simple andstraightforward method. Organoids are self-organized three-dimensional tissue cultures. They can proliferate, be passaged, andmaintained for a long period in in vitro culture. The human small
intestinal organoid-derived monolayer can be generated from theorganoids in 3–8 days by using a single medium without complicated procedures (Fig.1). The monolayer showed higher expressions and functions of major pharmacokinetic-related enzymes andtransporters than Caco-2 cells, and these profiles in the monolayer
were close to those of the human intestine in general. In addition,
the human small intestinal organoid-derived monolayer exhibitedgene expression profiles similar to those of the adult small intestine(Fig.2).

Fig 1. Graphical abstract of this studyAB

500 μm
C100 umDrelative gene expression(GAPDH = 1.0)Life science

Abstract

600 nm

100 1-1100.1-2100.01-30.001
10-40.0001
100.00001
10-5

CYP3A4CES2organoid monolayer
オルガノイド単層膜
Caco-2 cells
Caco-2細胞
成⼈⼗⼆指腸adult intestineFig 2. Characteristics of human small intestinal organoids(A) Established human small intestinal organoids(B) Human small intestinal organoids-derived monolayer(C)	Transmission electron micrographs of the monolayer. A brush border with microvilli (white arrow) and tight junctions (black arrow) were observed.
(D)	The gene expression levels of the major drug-metabolizing enzymes (CYP3A4and CES2) in the human small intestinal organoids-derived monolayer werecompared with those in Caco-2 cells and human adult intestine.

P a t e n t Japanese Patent Application No. 2020-016592
Yamashita, Tomoki; Mizuguchi, Hiroyuki et al. Monolayer platform using human biopsy-derived duodenal organoids for pharmaceutical research. MolecuTr e a t i s e
lar Therapy - Methods & Clinical Development. 2021; 22: 263-278. doi: 10.1016/j.omtm.2021.05.005.
URL https://resou.osaka-u.ac.jp/ja/research/2021/20210520_2
K e y w o r d small intestine, pharmacokinetics, organoid