A functional CFTR assay using primary cystic fibrosis intestinal organoids

• Published in: Nature medicine Vol. 19; no. 7; pp. 939 - 945
• Main Authors: Dekkers, Johanna F, Wiegerinck, Caroline L, de Jonge, Hugo R, Bronsveld, Inez, Janssens, Hettie M, de Winter-de Groot, Karin M, Brandsma, Arianne M, de Jong, Nienke W M, Bijvelds, Marcel J C, Scholte, Bob J, Nieuwenhuis, Edward E S, van den Brink, Stieneke, Clevers, Hans, van der Ent, Cornelis K, Middendorp, Sabine, Beekman, Jeffrey M
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01-07-2013; Nature Publishing Group
• Subjects: 1678/631/45/269; 1877/631/532/2437; 347/631/1647/1407/652; 600/631/1647/767/2199; Animals; Biological Assay - methods; Biomedicine; Biotechnology; Cancer Research; Cells, Cultured; Cystic fibrosis; Cystic Fibrosis - diagnosis; Cystic Fibrosis - genetics; Cystic Fibrosis - pathology; Cystic Fibrosis Transmembrane Conductance Regulator - analysis; Cystic Fibrosis Transmembrane Conductance Regulator - genetics; Cystic Fibrosis Transmembrane Conductance Regulator - metabolism; Drug therapy; Female; HEK293 Cells; Humans; Incubation; Infectious Diseases; Intestinal Mucosa - metabolism; Intestinal Mucosa - pathology; Low temperature; Male; Metabolic Diseases; Mice; Mice, Inbred C57BL; Mice, Inbred CFTR; Molecular Medicine; Mutagenesis; Mutant Proteins - analysis; Mutant Proteins - genetics; Mutant Proteins - metabolism; Mutant Proteins - physiology; Mutation; Neurosciences; Organoids - metabolism; Organoids - pathology; Primary Cell Culture - methods; technical-report; Technology application; Tissues; Water - metabolism; Netherlands
• ISSN: 1078-8956; 1546-170X
• DOI: 10.1038/nm.3201

Building on earlier work, Dekkers
et al
. describe the first application of their intestinal organoid culture technology to the study of human disease, in this case cystic fibrosis. These so called 'mini-guts', which recapitulate the essential
in vivo
intestinal tissue architecture
in vitro
, are used to develop a rapid and quantitative assay to measure mutant cystic fibrosis transmembrane conductance regulator (CFTR) function, as well as test the efficacy of correctors and potentiators of mutant CFTR.
We recently established conditions allowing for long-term expansion of epithelial organoids from intestine, recapitulating essential features of the
in vivo
tissue architecture. Here we apply this technology to study primary intestinal organoids of people suffering from cystic fibrosis, a disease caused by mutations in
CFTR
, encoding cystic fibrosis transmembrane conductance regulator. Forskolin induces rapid swelling of organoids derived from healthy controls or wild-type mice, but this effect is strongly reduced in organoids of subjects with cystic fibrosis or in mice carrying the Cftr F508del mutation and is absent in
Cftr
-deficient organoids. This pattern is phenocopied by CFTR-specific inhibitors. Forskolin-induced swelling of
in vitro
–expanded human control and cystic fibrosis organoids corresponds quantitatively with forskolin-induced anion currents in freshly excised
ex vivo
rectal biopsies. Function of the CFTR F508del mutant protein is restored by incubation at low temperature, as well as by CFTR-restoring compounds. This relatively simple and robust assay will facilitate diagnosis, functional studies, drug development and personalized medicine approaches in cystic fibrosis.