摘要：SummaryBackgroundA point mutation in sickle cell disease (SCD) alters one amino acid in the β-globin subunit of hemoglobin, with resultant anemia and multiorgan damage that typically shortens lifespan by decades. Because SCD is caused by a single mutation, and hematopoietic stem cells (HSCs) can be harvested, manipulated, and returned to an individual, it is an attractive target for gene correction.ResultsAn optimized Cas9 ribonucleoprotein (RNP) with an ssDNA oligonucleotide donor together generated correction of at least one β-globin allele in more than 30% of long-term engrafting human HSCs. After adopting a high-fidelity Cas9 variant, efficient correction with minimal off-target events also was observed.In vivoerythroid differentiation markedly enriches for corrected β-globin alleles, indicating that erythroblasts carrying one or more corrected alleles have a survival advantage.SignificanceThese findings indicate that the sickle mutation can be corrected in autologous HSCs with an optimized protocol suitable for clinical translation.Graphical abstractDisplay OmittedHighlights•The gene editing protocol corrects the sickle mutation in ∼30% of engrafting cells•Random assortment of engrafting stem cell clones without clonal dominance was shown•Corrected erythroid cells are preferentially enriched compared with unedited cellsGenetics; Molecular genetics; Genetic engineering
