EnglishEditing the β-globin locus in hematopoietic stem cells is an alternative therapeutic approach for gene therapy of β-thalassemia and sickle cell disease. Using the CRISPR/Cas9 system, we genetically modified human hematopoietic stem and progenitor cells (HSPCs) to mimic the large rearrangements in the β-globin locus associated with hereditary persistence of fetal hemoglobin (HPFH), a condition that mitigates the clinical phenotype of patients with β-hemoglobinopathies. We optimized and compared the efficiency of plasmid-, lentiviral vector (LV)-, RNA-, and ribonucleoprotein complex (RNP)-based methods to deliver the CRISPR/Cas9 system into HSPCs. Plasmid delivery of Cas9 and gRNA pairs targeting two HPFH-like regions led to high frequency of genomic rearrangements and HbF reactivation in erythroblasts derived from sorted, Cas9+ HSPCs but was associated with significant cell toxicity. RNA-mediated delivery of CRISPR/Cas9 was similarly toxic but much less efficient in editing the β-globin locus. Transduction of HSPCs by LVs expressing Cas9 and gRNA pairs was robust and minimally toxic but resulted in poor genome-editing efficiency. Ribonucleoprotein (RNP)-based delivery of CRISPR/Cas9 exhibited a good balance between cytotoxicity and efficiency of genomic rearrangements as compared to the other delivery systems and resulted in HbF upregulation in erythroblasts derived from unselected edited HSPCs.
| Data di pubblicazione: | 2019 |
| Titolo: | Optimization of CRISPR/Cas9 Delivery to Human Hematopoietic Stem and Progenitor Cells for Therapeutic Genomic Rearrangements |
| Autore/i: | Lattanzi, Annalisa; Meneghini, Vasco; Pavani, Giulia; Amor, Fatima; Ramadier, Sophie; Felix, Tristan; Antoniani, Chiara; Masson, Cecile; Alibeu, Olivier; Lee, Ciaran; Porteus, Matthew H.; Bao, Gang; Amendola, Mario; Mavilio, Fulvio; Miccio, Annarita |
| Autore/i UNIMORE: | |
| Digital Object Identifier (DOI): | 10.1016/j.ymthe.2018.10.008 |
| Rivista: | |
| Volume: | 27 |
| Fascicolo: | 1 |
| Pagina iniziale: | 137 |
| Pagina finale: | 150 |
| Codice identificativo ISI: | WOS:000454708300015 |
| Codice identificativo Scopus: | 2-s2.0-85056268645 |
| Codice identificativo Pubmed: | 30424953 |
| Citazione: | Optimization of CRISPR/Cas9 Delivery to Human Hematopoietic Stem and Progenitor Cells for Therapeutic Genomic Rearrangements / Lattanzi, Annalisa; Meneghini, Vasco; Pavani, Giulia; Amor, Fatima; Ramadier, Sophie; Felix, Tristan; Antoniani, Chiara; Masson, Cecile; Alibeu, Olivier; Lee, Ciaran; Porteus, Matthew H.; Bao, Gang; Amendola, Mario; Mavilio, Fulvio; Miccio, Annarita. - In: MOLECULAR THERAPY. - ISSN 1525-0016. - 27:1(2019), pp. 137-150. |
| Tipologia | Articolo su rivista |
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