CRISPR/Cas9 and p53: An Odd Couple Requiring Relationship Management
Authors: Seppo Ylä-Herttuala
Journal: Molecular Therapy (2018)

CRISPR/Cas9 is undoubtedly a very useful tool for genome editing and numerous other applications in genomics, gene therapy, and regenerative medicine. CRIPSR/Cas9 greatly facilitates genomic modification, and these reagents are widely available to the research community. It is thus not surprising when new aspects of CRISPR/Cas9 technology are discovered by the increasing number of studies using this technology. One example is a set of recent publications that highlighted a connection between the efficacy of CRISPR/Cas9-mediated genome editing and cellular DNA repair processes and, in particular, the well-known tumor suppressor gene p53.1Ihry R.J. Worringer K.A. Salick M.R. Frias E. Ho D. Theriault K. Kommineni S. Chen J. Sondey M. Ye C. et al.p53 inhibits CRISPR-Cas9 engineering in human pluripotent stem cells.Nat. Med. 2018; 24: 939-946Crossref PubMed Scopus (512) Google Scholar, 2Haapaniemi E. Botla S. Persson J. Schmierer B. Taipale J. CRISPR-Cas9 genome editing induces a p53-mediated DNA damage response.Nat. Med. 2018; 24: 927-930Crossref PubMed Scopus (602) Google Scholar, 3Janic A. Valente L.J. Wakefield M.J. Di Stefano L. Milla L. Wilcox S. Yang H. Tai L. Vandenberg C.J. Kueh A.J. et al.DNA repair processes are critical mediators of p53-dependent tumor suppression.Nat. Med. 2018; 24: 947-953Crossref PubMed Scopus (83) Google Scholar CRISPR/Cas9 genome editing involves the generation of DNA double-strand breaks (DSBs) induced by Cas9 that are then repaired by either non-homologous end joining or homology-directed repair. In targeted genome editing, a repair template homologous to the targeted locus is introduced into the cells to drive homology-directed repair of the DSBs to insert new genetic material into the target site. Since the generation of DSBs is a major threat to cells, it is not surprising that they protect their genome using several DNA repair mechanisms. Indeed, it has recently been shown that CRISPR/Cas9 editing induces a p53-mediated DNA damage response and that this process can significantly affect the efficiency of precise genomic editing, which could lead to potential problems, especially for direct in vivo gene editing applications. Since wild-type p53 can counteract DNA editing, it is possible that the CRISPR/Cas9 system might be more effective in subsets of cells in which the p53 tumor suppressor has been inactivated by mutations.1Ihry R.J. Worringer K.A. Salick M.R. Frias E. Ho D. Theriault K. Kommineni S. Chen J. Sondey M. Ye C. et al.p53 inhibits CRISPR-Cas9 engineering in human pluripotent stem cells.Nat. Med. 2018; 24: 939-946Crossref PubMed Scopus (512) Google Scholar This could obviously lead to malignant transformation and a selective growth advantage of unwanted cell populations. On the other hand, it has been suggested that transient inhibition of p53 might improve homology-directed repair and the efficacy of genome editing in cell types that are difficult to modify with CRISPR/Cas9 technology.2Haapaniemi E. Botla S. Persson J. Schmierer B. Taipale J. CRISPR-Cas9 genome editing induces a p53-mediated DNA damage response.Nat. Med. 2018; 24: 927-930Crossref PubMed Scopus (602) Google Scholar It is well known that several cancer cell lines are easy to engineer with CRISPR/Cas9 technology, whereas many normal cell lines, especially human pluripotent stem cells (hPSCs), are very difficult targets for genome editing. It has been shown that, in hPSCs, CRISPR/Cas9-induced DSBs are toxic and often kill the cells.1Ihry R.J. Worringer K.A. Salick M.R. Frias E. Ho D. Theriault K. Kommineni S. Chen J. Sondey M. Ye C. et al.p53 inhibits CRISPR-Cas9 engineering in human pluripotent stem cells.Nat. Med. 2018; 24: 939-946Crossref PubMed Scopus (512) Google Scholar Since the toxic response and efficiency of precise genomic editing are significantly reduced by p53 activity, it is important to realize that genome editing can occur more easily in p53 mutated cells and that cell replacement therapies using CRISPR/Cas9-engineered hPSCs should be carefully monitored for p53 and DNA repair functions. It is now evident that DNA repair processes may also affect other techniques used for precise genome editing. Generation of chromatin rearrangements and insertions or deletions (Indels) at the target site needs to be carefully monitored. Currently, off-target effects of CRISPR/Cas9 technology after in vivo applications remain unclear, although it appears that careful design of guide RNAs can substantially reduce off-target effects.4Akcakaya P. Bobbin M.L. Guo J.A. Malagon-Lopez J. Clement K. Garcia S.P. Fellows M.D. Porritt M.J. Firth M.A. Carreras A. et al.In vivo CRISPR editing with no detectable genome-wide off-target mutations.Nature. 2018; 561: 416-419Crossref PubMed Scopus (212) Google Scholar On the other hand, on-target alterations are equally important since targeted gene editing can generate Indels at the target site with the possibility of even larger genomic rearrangements. While we are seeing an exponential growth in applications using efficient CRISPR/Cas9 genome editing, the potential to modify DNA repair mechanisms and their consequences must be kept in mind. With a greater understanding of the multiple DNA repair mechanisms and their activities in different cell types and during different phases of the cell cycle, it is expected that the efficacy and safety of precise genome editing will be significantly improved both in experimental settings and, ultimately, in ex vivo and in vivo clinical applications.