Intestinal tissue organoids to study drug transport andmetabolism from infants to adults,22,31
1
41,5Eva J. Streekstra¹ , M. Keuper-Navis , R. Greupink , F.G.M. Russel , Kazuhiro Ariga ,E. van de Steeg², S.N. de Wildt
1 Division of Pharmacology and Toxicology, Department of Pharmacy, Radboud University Medical Center, Nijmegen, The Netherlands#1P-154

2 Human Cell Biology, Department of Metabolic Health Research, TNO, Leiden, The Netherlands
3 Experimental Pharmacology, Department of Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands
4 TNO, Pharma, Japan
5 Intensive Care and Department of Pediatric Surgery, Erasmus MC Sophia Children Hospital, Rotterdam, The Netherlands

BackgroundThe small intestine plays a very important role in the oral bioavailability ofdrugs. Therefore, it is important to understand intestinal absorption,
potential regional differences, and age-related effects during drug
development. Existing in vitro models lack the ability to recapitulate the invivo intestinal physiology, neither mimic population variation. Moreover, GI
toxicity studies are profoundly dependent on animal studies1. Patientderived small intestinal organoids may overcome these limitations.
Differentiated organoids show most types of intestinal cells present inepithelial barrier, and express drug transporters and metabolizing enzymes
thereby being a promising tool for pharma- and toxicokinetic research in
special populations.

Why an intestinal model?

Organoid monolayer methodology

Objective
To study the applicability of patient derived intestinalorganoids for predicting region and population differencesin drug transport and metabolism

Results in fresh tissue vs patient derived organoids
Adult regional specificity fresh tissue vs organoid monolayers
Regional specific efflux transport

Fresh tissue

Organoids

Papp (x10-6 cm/s)

Papp (x10-6 cm/s)

Ileum

10500

A>B B>A A>B B>A

A>B B>A A>B B>AFresh tissue

20Organoids

Fresh tissue

A>B B>A A>B B>A

15105A>B B>A A>B B>A

Adult ileum

10500
A>B B>A A>B B>A A>B B>A A>B B>A

Adult monolayer metabolism by CYP3A4

More CYP3A4 metabolism in organoid monolayers compared to Caco-2

A>B B>A A>B B>A A>B B>A A>B B>A

2000
100050100150time (min)Adults50on
ths

01-3 Months
8-13 Months
Adult ileum

10000

500000m

Caco-2

pmol/mgprotein

3000

100813Ileum

Caco-2onths

Jejunum

Ileum organoids

4000

0150

Jejunum organoids

m0.2
0.0

1OH-midazolam formation
15000

1310Higher CYP3A4 metabolism in children ±1 year old

Metabolic rate
(pmol/mg protein/min)

pmol/mgprotein

Metabolic rate
(pmol/mg protein/min)20

Pediatric vs adult CYP3A4 metabolism in organoid monolayers
Metabolic rate CYP3A45000

30A>B = apical to basolateral transport; B>A = basolateral to apical transport

1OH-Midazolam formation40

Paedaitric ileum

15A>B = apical to basolateral transport; B>A = basolateral to apical transport

Metabolic rate CYP3A4

Organoids

Jejunum

155

20

Papp (x10-6 cm/s)

20

Organoids

Rosuvastatin-BCRP

Talinolol-P-gp

Papp (x10-6 cm/s)

Fresh tissue

Similar efflux (B>A) transport in pediatric organoids

Rosuvastatin-BCRP

Talinolol-P-gp
10Pediatric ≤1 year & adult in fresh tissue vs organoid monolayers

050

100150time (min)Take home messages:• Patient derived adult and pediatric organoids can be cultured and applied for PK research• Intestinal organoid monolayers from adults show region specific functionality trends of MDR1 and BCRP 2,3 and more CYP3A4 functionality
compared to Caco-2 monolayers• Pediatric intestinal organoid monolayers 8-13 months old show a possible higher CYP3A4 metabolism compared to adults• Pediatric intestinal organoid monolayers show similar drug transport compared to adults• More (young) donors will be included to further study population variation in intestinal drug transport and metabolism• Organoid metabolism will be compared with fresh tissue functionality data and gene expression

Questions? Eva Streekstra, MSc; eva.streekstra@radboudumc.nl;

1 Matsui, T., & Shinozawa, T. (2021). Human organoids for predictive toxicology research and drug development. Frontiers in Genetics, 2119.doi: 10.3389/fgene.2021.767621 2 Müller J, et al. Expression, regulation and function of intestinal drug transporters: an update. Biol Chem.

2017 Feb 1;398(2):175-192. doi: 10.1515/hsz-2016-0259. 3 Grangeon A, et al. Determination of CYP450 Expression Levels in the Human Small Intestine by Mass Spectrometry-Based Targeted Proteomics. Int J Mol Sci. 2021 Nov 26;22(23):12791. doi: 10.3390/ijms222312791.