Establishment of a system to evaluate the therapeutic effect and the dynamics of an investigational drug on ulcerative colitis using human colonic organoids

Abstract

Background

Ulcerative colitis (UC) is a chronic inflammatory disease of the colon with an intractable, recurrent course. The goal of UC therapy is to target mucosal healing because immune-suppressive therapy for UC frequently results in relapse. However, few drugs directly target mucosal healing. We, therefore, aim to evaluate the therapeutic effect of an investigational drug on intestinal epithelial cells in an in vitro UC model using human colonic organoids.

Methods

Colonic organoids were isolated from human colon and cultured. A mixture of cytokines and bacterial components were used to mimic UC in humans. The effect of the investigational drug on colonic organoid was evaluated by microarray analysis and 3D immunofluorescence. The enrichment of stem cells was assessed with a colony formation assay.

Results

Inflammatory stimulation resulted in a significant induction of inflammatory-related genes in colonic organoids whereas cell differentiation was suppressed. Treatment with the investigational drug KAG-308 showed reciprocal dynamics of gene expression to inflammatory stimulation, which resulted in not only the suppression of immune response but also the promotion of cellular differentiation towards secretory lineages. Moreover, SPDEF and Reg4 were identified as novel targets for the enrichment of intestinal epithelial stem cells and mucosal healing.

Conclusions

The establishment of in vitro UC model using human colonic organoid could reveal the effects and targets of investigational drugs in intestinal epithelial cells under inflammation conditions. Further maturation of this system might be more efficient to predict the effect on UC, as compared with the use of animal model, for the development of new drugs.

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Abbreviations

IBD:

Inflammatory bowel disease

UC:

Ulcerative colitis

DSS:

Dextran sulphate sodium

AOM:

Azoxymethane

IECs:

Intestinal epithelial cells

NF-κB:

Nuclear factor-κB

PGE2:

Prostaglandin E2

EP4:

Prostaglandin E2 receptor 4

GSEA:

Gene set enrichment analysis

TLRs:

Toll-like receptors

IL:

Interleukin

ROS:

Reactive oxygen species

LPS:

Lipopolysaccharide

TNF:

Tumour necrosis factor

CLDN18:

Claudin-18

CIITA:

Class II major histocompatibility complex transactivator

DAPI:

4′,6-diamidino-2-phenylindole

Reg4:

Regenerating family member 4

SPDEF:

SAM-pointed domain containing ETS transcription factor

Duoxa2:

Dual oxidase maturation factor 2

ALDH1:

Aldehyde dehydrogenase 1

Lgr5:

Leucine-rich repeat-containing G-protein coupled receptor 5

Hes1:

Hairy and enhancer of split 1

Atoh1:

Atonal bHLH transcription factor 1

TFF3:

Trefoil factor 3

MUC2:

Mucin 2

CgA:

Chromogranin A

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Acknowledgements

This work was supported by scientific Research, from the Japanese Ministry of Education, Culture, Sports, Science and Technology as follows (KAKENHI grant numbers 24590935, 25114703, 25130704, 26221307, 15H04808, 16H06770, 17H06654, 17K15930, 17K19513); Japan Foundation for Applied Enzymology; the Health and Labor Sciences Research Grant from the Japanese Ministry of Health, Labor and Welfare (MHLW) as follows (Grant number 14526073); The Practical Research for Innovative Cancer Control, Rare/Intractable Diseases from Japan Agency for Medical Research and Development (AMED) as follows (Grant numbers 15Ack0106017h0002, 16ck0106017h0003, 16ek0109456h0003); Research Grant of the Princess Takamatsu Cancer Research Fund; Naoki Tsuchida Research Grant.

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Correspondence to Kiichiro Tsuchiya.

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Conflict of interest

YM and YW are employed by Kaken Pharmaceutical Co., Ltd. KT has received research funding (collaborative research) provided by Kaken Pharmaceutical Co., Ltd.

Author contributions

MW, TN and KT designed the experiments and acquired data. TS and SH established inflammatory stimulation of the organoids. RN and SW assessed the effect of KAG-308 on the organoids. YM and YW assessed the effect of KAG-308 on DSS colitis model in mice. KT and MW coordinated the projects and drafted the manuscript.

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Nishimura, R., Shirasaki, T., Tsuchiya, K. et al. Establishment of a system to evaluate the therapeutic effect and the dynamics of an investigational drug on ulcerative colitis using human colonic organoids. J Gastroenterol 54, 608–620 (2019). https://doi.org/10.1007/s00535-018-01540-y

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Keywords

  • Inflammatory bowel disease
  • In vitro human model
  • Evaluation system
  • Investigational drug