Prime editing in hematopoietic stem cells—From ex vivo to in vivo CRISPR-based treatment of blood disorders

Abstract

Prime editing of human hematopoietic stem cells has the potential to become a safe and efficient way of treating diseases of the blood directly in patients. By allowing site-targeted gene intervention without homology-directed repair donor templates and DNA double-stranded breaks, the invention of prime editing fuels the exploration of alternatives to conventional recombination-based ex vivo genome editing of hematopoietic stem cells. Prime editing is as close as we get today to a true genome editing drug that does not require a separate DNA donor. However, to adapt the technology to perform in vivo gene correction, key challenges remain to be solved, such as identifying effective prime editing guide RNAs for clinical targets as well as developing efficient vehicles to deliver prime editors to stem cells in vivo. In this review, we summarize the current progress in delivery of prime editors both in vitro and in vivo and discuss future challenges that need to be adressed to allow in vivo prime editing as a cure for blood disorders.

Post-Transplant Administration of G-CSF Impedes Engraftment of Gene Edited Human Hematopoietic Stem Cells by Exacerbating the p53-Mediated DNA Damage Response

“…The synergistic activation of p53 observed in HSPCs exposed to G-CSF and CRISPR-Cas9 RNPs is analogous to the enhanced p53 stimulation previously reported when HSPC gene editing was performed in conjunction with transduction of AAV6 DNA donor template delivery vectors [10][11][12] . In the future, it will be important to ascertain the impact of post-transplant G-CSF on human HSPCs subjected to emerging gene correction technologies that do not rely on DSB formation (i.e., base editing [38][39][40] and prime editing [40][41][42][43] ) or are independent of the cellular DDR (i.e., recombinases 40,[44][45][46][47] and RNA-guided transposon systems 40,47 ). Globally, our findings further expand the body of evidence delineating p53 as a central hub in a molecular network orchestrating a variety of DDR pathways, including cell cycle checkpoints, apoptosis and immune responses.…”