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CRISPR engineering in organoids for gene repair and disease modelling

Nature Reviews Bioengineering volume 1, pages 32–45 (2023)Cite this article

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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.

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Fig. 1: CRISPR engineering in adult-stem-cell-derived organoids.
Fig. 2: Principles of CRISPR–Cas9-mediated genome engineering.
Fig. 3: Principles of cytosine and adenine base editing.
Fig. 4: Principles of prime editing.
Fig. 5: The process of creating isogenic disease models in adult-stem-cell-derived organoids.

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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.

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Author notes

  1. Maarten H. Geurts

    Present address: Xilis BV, Utrecht, The Netherlands

  2. Hans Clevers

    Present address: Pharma Research Early Development, Roche, Basel, Switzerland

Authors and Affiliations

  1. Hubrecht Institute, Royal Netherlands Academy of Arts and Sciences (KNAW) and University Medical Center Utrecht, Utrecht, The Netherlands

    Maarten H. Geurts & Hans Clevers

  2. Oncode Institute, Hubrecht Institute, Utrecht, The Netherlands

    Maarten H. Geurts & Hans Clevers

Authors
  1. Maarten H. Geurts
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  2. Hans Clevers
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Correspondence to Hans Clevers.

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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.

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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

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