%0 Journal Article
%T Engineered human Diamond-Blackfan anemia disease model confirms therapeutic effects of clinically applicable lentiviral vector at single-cell resolution.
%A Liu Y
%A Schmiderer L
%A Hjort M
%A Lang S
%A Bremborg T
%A Rydström A
%A Schambach A
%A Larsson J
%A Karlsson S
%J Haematologica
%V 108
%N 11
%D 2023 11 1
%M 37199130
%F 11.047
%R 10.3324/haematol.2022.282068
%X Diamond-Blackfan anemia is a rare genetic bone marrow failure disorder which is usually caused by mutations in ribosomal protein genes. In the present study, we generated a traceable RPS19-deficient cell model using CRISPR-Cas9 and homology-directed repair to investigate the therapeutic effects of a clinically applicable lentiviral vector at single-cell resolution. We developed a gentle nanostraw delivery platform to edit the RPS19 gene in primary human cord bloodderived CD34+ hematopoietic stem and progenitor cells. The edited cells showed expected impaired erythroid differentiation phenotype, and a specific erythroid progenitor with abnormal cell cycle status accompanied by enrichment of TNFα/NF-κB and p53 signaling pathways was identified by single-cell RNA sequencing analysis. The therapeutic vector could rescue the abnormal erythropoiesis by activating cell cycle-related signaling pathways and promoted red blood cell production. Overall, these results establish nanostraws as a gentle option for CRISPR-Cas9- based gene editing in sensitive primary hematopoietic stem and progenitor cells, and provide support for future clinical investigations of the lentiviral gene therapy strategy.