Integrated computational and experimental identification of <i>p53, KRAS</i> and <i>VHL</i> mutant selection associated with CRISPR-Cas9 editing

Sinha, Sanju; Barbosa, Karina; Cheng, Kuoyuan; Leiserson, Mark D.M.; Wilson, David M.; Ryan, Bríd M.; Ronai, Ze’ev; Lee, Joo Sang; Deshpande, Aniruddha J.; Ruppin, Eytan

doi:10.1101/407767

Cited by 4 publications

(4 citation statements)

References 41 publications

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“…5c and Supplementary Data 6 ). This suggests that Cas9-induced DNA damage in TP53 -WT cells increased the dependency on functional DNA repair machinery, consistent with a similar recent analysis 19 . Finally, we compared the dependency on TP53 itself between TP53 -WT lines in which p53 pathway activation was identified and TP53 -WT lines in which such activation was not observed.…”

supporting

confidence: 91%

Cas9 activates the p53 pathway and selects for p53-inactivating mutations

et al. 2020

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Cas9 is commonly introduced into cell lines to enable CRISPR/Cas9-mediated genome editing. Here we studied the genetic and transcriptional consequences of Cas9 expression per se . Gene expression profiling of 165 pairs of human cancer cell lines and their Cas9-expressing derivatives revealed upregulation of the p53 pathway upon Cas9 introduction, specifically in TP53 -WT cell lines. This was confirmed at the mRNA and protein levels. Moreover, elevated levels of DNA repair were observed in Cas9-expressing cell lines. Genetic characterization of 42 cell line pairs showed that Cas9 introduction can lead to the emergence and expansion of p53-inactivating mutations. This was confirmed by competition experiments in isogenic TP53 -WT/ TP53 -null cell lines. Lastly, Cas9 was less active in TP53 -WT than in TP53 -mutant cell lines, and Cas9-induced p53 pathway activation affected cellular sensitivity to both genetic and chemical perturbations. These findings may have broad implications for the proper use of CRISPR/Cas9-mediated genome editing.

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supporting

confidence: 91%

Cas9 activates the p53 pathway and selects for p53-inactivating mutations

et al. 2020

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“…Examples of such potential consequences may include unintentional mutations in the cancer driver genes VHL or KRAS , as a result of GET activity yielding neoplastic events. 53 More intense research is needed to understand, and perhaps in the future control, which repair pathway is activated following DNA cutting by GETs. In this context, as molecules that increase the frequency or extent of mutations, GETs fit the definition of “targeted mutagens.” Therefore, it is unrealistic to expect that GETs alone will guarantee the desired outcomes of their use.…”

Section: Main Textmentioning

confidence: 99%

Variability in Genome Editing Outcomes: Challenges for Research Reproducibility and Clinical Safety

et al. 2020

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Genome editing tools have already revolutionized biomedical research and are also expected to have an important impact in the clinic. However, their extensive use in research has revealed much unpredictability, both off and on target, in the outcome of their application. We discuss the challenges associated with this unpredictability, both for research and in the clinic. For the former, an extensive validation of the model is essential. For the latter, potential unpredicted activity does not preclude the use of these tools but requires that molecular evidence to underpin the relevant risk:benefit evaluation is available. Safe and successful clinical application will also depend on the mode of delivery and the cellular context.

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“…A recent report by Sinha et al [ 54 ] addressed the question of whether CRISPR/Cas9 editing could lead to oncogenic mutations in other genes. Authors analysed genome-wide CRISPR and RNAi screens to reveal the mutation selection potential of CRISPR knockouts across the whole exome.…”

Section: Crispr/cas9 System and Tp53 Pathway Gementioning

confidence: 99%

Therapeutic Editing of the TP53 Gene: Is CRISPR/Cas9 an Option?

Mirgayazova

Khadiullina

Chasov

et al. 2020

Genes

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The TP53 gene encodes the transcription factor and oncosuppressor p53 protein that regulates a multitude of intracellular metabolic pathways involved in DNA damage repair, cell cycle arrest, apoptosis, and senescence. In many cases, alterations (e.g., mutations of the TP53 gene) negatively affect these pathways resulting in tumor development. Recent advances in genome manipulation technologies, CRISPR/Cas9, in particular, brought us closer to therapeutic gene editing for the treatment of cancer and hereditary diseases. Genome-editing therapies for blood disorders, blindness, and cancer are currently being evaluated in clinical trials. Eventually CRISPR/Cas9 technology is expected to target TP53 as the most mutated gene in all types of cancers. A majority of TP53 mutations are missense which brings immense opportunities for the CRISPR/Cas9 system that has been successfully used for correcting single nucleotides in various models, both in vitro and in vivo. In this review, we highlight the recent clinical applications of CRISPR/Cas9 technology for therapeutic genome editing and discuss its perspectives for editing TP53 and regulating transcription of p53 pathway genes.

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Integrated computational and experimental identification of p53, KRAS and VHL mutant selection associated with CRISPR-Cas9 editing

Cited by 4 publications

References 41 publications

Cas9 activates the p53 pathway and selects for p53-inactivating mutations

Cas9 activates the p53 pathway and selects for p53-inactivating mutations

Variability in Genome Editing Outcomes: Challenges for Research Reproducibility and Clinical Safety

Therapeutic Editing of the TP53 Gene: Is CRISPR/Cas9 an Option?

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