CRISPR-Based Adenine Editors Correct Nonsense Mutations in a Cystic Fibrosis Organoid Biobank

Geurts, Maarten H., de Poel, Eyleen, Amatngalim, Gimano D., Oka, Rurika, Meijers, Fleur M., Kruisselbrink, Evelien, van Mourik, Peter, Berkers, Gitte, de Winter-de Groot, Karin M., Michel, Sabine, Muilwijk, Danya, Aalbers, Bente L., Mullenders, Jasper, Boj, Sylvia F., Suen, Sylvia W.F., Brunsveld, Jesse E., Janssens, Hettie M., Mall, Marcus A., Graeber, Simon Y., van Boxtel, Ruben, van der Ent, Cornelis K., Beekman, Jeffrey M. & Clevers, Hans

Published: 02/04/2020


Adenine base editing (ABE) enables enzymatic conversion from A-T into G-C base pairs. ABE holds promise for clinical application, as it does not depend on the introduction of double-strand breaks, contrary to conventional CRISPR/Cas9-mediated genome engineering. Here, we describe a cystic fibrosis (CF) intestinal organoid biobank, representing 664 patients, of which ~20% can theoretically be repaired by ABE. We apply SpCas9-ABE (PAM recognition sequence: NGG) and xCas9-ABE (PAM recognition sequence: NGN) on four selected CF organoid samples. Genetic and functional repair was obtained in all four cases, while whole-genome sequencing (WGS) of corrected lines of two patients did not detect off-target mutations. These observations exemplify the value of large, patient-derived organoid biobanks representing hereditary disease and indicate that ABE may be safely applied in human cells.

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