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ABSTRACT Organoids bridge the gap between 2D cell lines and in vivo studies. With their 3D organization and cellular heterogeneity, adult stem cell-derived organoids closely resemble their
tissue of origin. The development of CRISPR-mediated genome engineering and the recent additions of base and prime editing to the CRISPR toolbox have greatly simplified the generation of
exact, isogenic models for Mendelian diseases. Here, we review recent developments in CRISPR-mediated genome engineering and its application in human adult-stem-cell-derived organoids in the
construction of isogenic disease models. These models allow accurate qualification of the impact of allelic disease variants observed in patients. Furthermore, we discuss the use of
organoids as models for safety and efficacy of CRISPR for gene repair. Although transplantation of repaired tissue remains challenging, benchmarking CRISPR tools in organoids can bring
genome engineering one step closer to patients. KEY POINTS * CRISPR–Cas9-mediated genome engineering acts by introducing double-stranded DNA breaks into the genome. The damage repair process
can be used for gene knockout or precise targeted introduction of exogenous DNA. * Next-generation CRISPR tools, including base and prime editing, allow for induction of precise base
changes and small insertions and deletions, bypassing potentially deleterious double-stranded DNA breaks. * Owing to their 3D organization, adult-stem-cell-derived organoids closely resemble
the tissue of origin and are therefore a good model system to study human health and disease. * CRISPR–Cas9-mediated genome engineering can be used to create isogenic models to investigate
the onset, cause and treatment of human diseases. * CRISPR tools can be benchmarked for efficiency and safety by studying gene repair ex vivo in adult-stem-cell-derived organoids,
facilitating CRISPR–Cas9 clinical translation. * Ex vivo repaired adult-stem-cell-derived organoids can potentially be transplanted into patients to relieve disease phenotypes. Access
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support SIMILAR CONTENT BEING VIEWED BY OTHERS CRISPR SOMATIC GENOME ENGINEERING AND CANCER MODELING IN THE MOUSE PANCREAS AND LIVER Article 14 March 2022 PRIME EDITING FOR FUNCTIONAL
REPAIR IN PATIENT-DERIVED DISEASE MODELS Article Open access 23 October 2020 THE NIH SOMATIC CELL GENOME EDITING PROGRAM Article Open access 07 April 2021 REFERENCES * Visscher, P. M. et al.
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_Nature_ 592, 99–104 (2021). Google Scholar Download references ACKNOWLEDGEMENTS The authors thank J. Beumer for providing confocal images of human intestinal organoids, S. Gandhi for
providing confocal images of human fetal hepatocyte organoids and J. van der Vaart for providing confocal images of murine thyroid organoids. AUTHOR INFORMATION Author notes * Maarten H.
Geurts Present address: Xilis BV, Utrecht, The Netherlands * Hans Clevers Present address: Pharma Research Early Development, Roche, Basel, Switzerland AUTHORS AND AFFILIATIONS * Hubrecht
Institute, Royal Netherlands Academy of Arts and Sciences (KNAW) and University Medical Center Utrecht, Utrecht, The Netherlands Maarten H. Geurts & Hans Clevers * Oncode Institute,
Hubrecht Institute, Utrecht, The Netherlands Maarten H. Geurts & Hans Clevers Authors * Maarten H. Geurts View author publications You can also search for this author inPubMed Google
Scholar * Hans Clevers View author publications You can also search for this author inPubMed Google Scholar CORRESPONDING AUTHOR Correspondence to Hans Clevers. ETHICS DECLARATIONS COMPETING
INTERESTS H.C. is inventor on several patents related to organoid technology; his full disclosure is given at https://www.uu.nl/staff/JCClevers/. H.C. is currently head of pharma Research
Early Development (pRED) at Roche. H.C. holds several patents on organoid technology. Their application numbers, followed by their publication numbers (if applicable), are as follows:
PCT/NL2008/050543, WO2009/022907; PCT/NL2010/000017, WO2010/090513; PCT/IB2011/002167, WO2012/014076; PCT/IB2012/052950, WO2012/168930; PCT/EP2015/060815, WO2015/173425; PCT/EP2015/077990,
WO2016/083613; PCT/EP2015/077988, WO2016/083612; PCT/EP2017/054797, WO2017/149025; PCT/EP2017/065101, WO2017/220586; PCT/EP2018/086716, n/a; and GB1819224.5, n/a. M.H.G. is currently a
scientist at Xilis BV. PEER REVIEW PEER REVIEW INFORMATION _Nature Reviews Bioengineering_ thanks Nicholas Zachos and the other, anonymous, reviewer(s) for their contribution to the peer
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permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Geurts, M.H., Clevers, H. CRISPR engineering in organoids for gene repair and disease modelling. _Nat Rev Bioeng_ 1, 32–45 (2023).
https://doi.org/10.1038/s44222-022-00013-5 Download citation * Accepted: 17 November 2022 * Published: 19 January 2023 * Issue Date: January 2023 * DOI:
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