CRISPR-Cas9 genome editing induces megabase-scale chromosomal truncations

Nat Commun. 2019 Mar 8;10(1):1136. doi: 10.1038/s41467-019-09006-2.


CRISPR-Cas9 is a promising technology for genome editing. Here we use Cas9 nuclease-induced double-strand break DNA (DSB) at the UROS locus to model and correct congenital erythropoietic porphyria. We demonstrate that homology-directed repair is rare compared with NHEJ pathway leading to on-target indels and causing unwanted dysfunctional protein. Moreover, we describe unexpected chromosomal truncations resulting from only one Cas9 nuclease-induced DSB in cell lines and primary cells by a p53-dependent mechanism. Altogether, these side effects may limit the promising perspectives of the CRISPR-Cas9 nuclease system for disease modeling and gene therapy. We show that the single nickase approach could be safer since it prevents on- and off-target indels and chromosomal truncations. These results demonstrate that the single nickase and not the nuclease approach is preferable, not only for modeling disease but also and more importantly for the safe management of future CRISPR-Cas9-mediated gene therapies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CRISPR-Associated Protein 9 / genetics
  • CRISPR-Associated Protein 9 / metabolism
  • CRISPR-Cas Systems*
  • Chromosome Deletion
  • Chromosomes, Human, Pair 10*
  • Clustered Regularly Interspaced Short Palindromic Repeats
  • DNA / genetics
  • DNA / metabolism
  • DNA Breaks, Double-Stranded*
  • Deoxyribonuclease I / genetics*
  • Deoxyribonuclease I / metabolism
  • Fibroblasts / cytology
  • Fibroblasts / metabolism
  • Gene Editing / methods*
  • Genetic Therapy / methods*
  • Genome, Human
  • HEK293 Cells
  • High-Throughput Nucleotide Sequencing
  • Humans
  • K562 Cells
  • Models, Biological
  • Porphyria, Erythropoietic / genetics
  • Porphyria, Erythropoietic / metabolism
  • Porphyria, Erythropoietic / pathology
  • Porphyria, Erythropoietic / therapy
  • Primary Cell Culture
  • RNA, Guide, CRISPR-Cas Systems / genetics
  • RNA, Guide, CRISPR-Cas Systems / metabolism
  • Recombinational DNA Repair
  • Tumor Suppressor Protein p53 / genetics
  • Tumor Suppressor Protein p53 / metabolism
  • Uroporphyrinogen III Synthetase / genetics*
  • Uroporphyrinogen III Synthetase / metabolism


  • RNA, Guide, CRISPR-Cas Systems
  • Tumor Suppressor Protein p53
  • DNA
  • CRISPR-Associated Protein 9
  • Deoxyribonuclease I
  • Uroporphyrinogen III Synthetase