One-step generation of tumor models by base editor multiplexing in adult stem cell-derived organoids

Maarten H Geurts, Shashank Gandhi, Matteo G Boretto, Ninouk Akkerman, Lucca L M Derks, Gijs van Son, Martina Celotti, Sarina Harshuk-Shabso, Flavia Peci, Harry Begthel, Delilah Hendriks, Paul Schürmann, Amanda Andersson-Rolf, Susana M Chuva de Sousa Lopes, Johan H van Es, Ruben van Boxtel, Hans Clevers

Research output: Contribution to journal/periodicalArticleScientificpeer-review

4 Citations (Scopus)

Abstract

Optimization of CRISPR/Cas9-mediated genome engineering has resulted in base editors that hold promise for mutation repair and disease modeling. Here, we demonstrate the application of base editors for the generation of complex tumor models in human ASC-derived organoids. First we show efficacy of cytosine and adenine base editors in modeling CTNNB1 hot-spot mutations in hepatocyte organoids. Next, we use C > T base editors to insert nonsense mutations in PTEN in endometrial organoids and demonstrate tumorigenicity even in the heterozygous state. Moreover, drug sensitivity assays on organoids harboring either PTEN or PTEN and PIK3CA mutations reveal the mechanism underlying the initial stages of endometrial tumorigenesis. To further increase the scope of base editing we combine SpCas9 and SaCas9 for simultaneous C > T and A > G editing at individual target sites. Finally, we show that base editor multiplexing allow modeling of colorectal tumorigenesis in a single step by simultaneously transfecting sgRNAs targeting five cancer genes.

Original languageEnglish
Pages (from-to)4998
JournalNature Communications
Volume14
Issue number1
DOIs
Publication statusPublished - 17 Aug 2023

Keywords

  • Adult
  • Humans
  • RNA, Guide, CRISPR-Cas Systems
  • Oncogenes
  • Carcinogenesis/genetics
  • Cell Transformation, Neoplastic
  • Adult Stem Cells
  • Organoids

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