TY - JOUR
T1 - Evaluating CRISPR-based prime editing for cancer modeling and CFTR repair in organoids
AU - Geurts, Maarten H.
AU - de Poel, Eyleen
AU - Pleguezuelos-Manzano, Cayetano
AU - Oka, Rurika
AU - Carrillo, Léo
AU - Andersson-Rolf, Amanda
AU - Boretto, Matteo
AU - Brunsveld, Jesse E.
AU - van Boxtel, Ruben
AU - Beekman, Jeffrey M.
AU - Clevers, Hans
N1 - Funding Information:
This work was supported by Cancer Research UK C6307/A29058 and The Mark Foundation for Cancer Research (H Clevers and MH Geurts). This work was supported by the NWO building blocks of life project: Cell dynamics within lung and intestinal organoids (737.016.009) (MH Geurts) and grants of the Dutch Cystic Fibrosis Foundation (NCFS), The Netherlands, as part of the HIT-CF Program; the Dutch Health Organization ZonMw, The Netherlands. This work was supported by the Netherlands Organ-on-Chip Initiative, an NWO Gravitation project (024.003.001) funded by the Ministry of Education, Culture and Science of the government of The Netherlands (H Clevers, C Pleguezuelos-Manzano). This work is part of the Oncode Institute which is partly financed by the Dutch Cancer Society and was funded by the gravitation program Can-cerGenomiCs.nl from The Netherlands Organisation for Scientific Research (NWO). M Boretto and A Andersson-Rolf are postdoctoral researchers supported by a long-term fellowship of the European Organization for Molecular Biology (EMBO/ALTF 765-2019, and EMBO ALTF 332-2018, resp.).
Publisher Copyright:
© 2021 Geurts et al.
PY - 2021/10
Y1 - 2021/10
N2 - Prime editing is a recently reported genome editing tool using a nickase-cas9 fused to a reverse transcriptase that directly synthesizes the desired edit at the target site. Here, we explore the use of prime editing in human organoids. Common TP53 mutations can be correctly modeled in human adult stem cell-derived colonic organoids with efficiencies up to 25% and up to 97% in hepatocyte organoids. Next, we functionally repaired the cystic fibrosis CFTR-F508del mutation and compared prime editing to CRISPR/Cas9-mediated homology-directed repair and adenine base editing on the CFTR-R785* mutation. Whole-genome sequencing of prime editing-repaired organoids revealed no detectable off-target effects. Despite encountering varying editing efficiencies and undesired mutations at the target site, these results underline the broad applicability of prime editing for modeling oncogenic mutations and showcase the potential clinical application of this technique, pending further optimization.
AB - Prime editing is a recently reported genome editing tool using a nickase-cas9 fused to a reverse transcriptase that directly synthesizes the desired edit at the target site. Here, we explore the use of prime editing in human organoids. Common TP53 mutations can be correctly modeled in human adult stem cell-derived colonic organoids with efficiencies up to 25% and up to 97% in hepatocyte organoids. Next, we functionally repaired the cystic fibrosis CFTR-F508del mutation and compared prime editing to CRISPR/Cas9-mediated homology-directed repair and adenine base editing on the CFTR-R785* mutation. Whole-genome sequencing of prime editing-repaired organoids revealed no detectable off-target effects. Despite encountering varying editing efficiencies and undesired mutations at the target site, these results underline the broad applicability of prime editing for modeling oncogenic mutations and showcase the potential clinical application of this technique, pending further optimization.
UR - http://www.scopus.com/inward/record.url?scp=85113285818&partnerID=8YFLogxK
U2 - 10.26508/LSA.202000940
DO - 10.26508/LSA.202000940
M3 - Article
C2 - 34373320
AN - SCOPUS:85113285818
VL - 4
JO - Life Science Alliance
JF - Life Science Alliance
IS - 10
M1 - e202000940
ER -