To test whether prime editing could repair a disease-causing GATA2 mutation in patient-derived blood stem and progenitor cells.
Approach:
Editing strategy: Researchers used prime editing to correct a seven-base-pair deletion in patient CD34-positive hematopoietic stem and progenitor cells.
Preclinical testing: They assessed editing, cell function, engraftment in immunodeficient mice, and unintended genetic changes.
Complementary method: They also tested PASSIGE to insert a larger corrective GATA2 sequence.
Key Findings:
Prime editing reached efficiencies above 70% in optimized CD34-positive cells, with limited effects on cell fitness.
The proportion of functional GATA2 alleles increased from 50% to as much as 77%, alongside increased GATA2 protein expression.
Edited cells retained colony-forming capacity and achieved up to 2.73-fold greater engraftment than untreated cells in immunodeficient mice; editing persisted in repopulating stem cells 16 weeks after transplantation.
Researchers reported limited unintended changes at the target site and no editing at the 20 highest-ranked predicted off-target sites.
Reducing stem-cell stimulation to less than 24 hours before editing reduced the DNA damage response and improved multilineage engraftment.
PASSIGE inserted the larger corrective sequence into 4.6% of alleles.
Interpretation:
In this preclinical study, prime editing corrected the tested GATA2 mutation in patient-derived cells and improved their engraftment in mice. The authors described the work as a step toward addressing the genetic cause of the disease.
Limitations:
The findings are preclinical; the described transplantation experiments were conducted in immunodeficient mice.
PASSIGE integration efficiency was 4.6% of alleles, and the researchers stated that further optimization is needed.
