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

doi:10.1016/j.ymthe.2018.10.008

Editing 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:	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 mostra contributor esterni nascondi contributor esterni
Digital Object Identifier (DOI):	10.1016/j.ymthe.2018.10.008
Rivista:	MOLECULAR THERAPY
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

File in questo prodotto:

Non ci sono file associati a questo prodotto.

Utilizza questo identificativo per citare o creare un link a questo documento:
http://hdl.handle.net/11380/1172498

Citazioni

I documenti presenti in Iris Unimore sono rilasciati con licenza Creative Commons Attribuzione - Non commerciale - Non opere derivate 3.0 Italia, salvo diversa indicazione.
In caso di violazione di copyright, contattare Supporto Iris