b-thalassemias (b-thal) are a group of blood disorders caused by mutations in the b-globin gene (HBB) cluster. b-globin associates with a-globin to form adult hemoglobin (HbA, a2b2), the main oxygen-carrier in erythrocytes. When b-globin chains are absent or limiting, free a-globins precipitate and damage cell membranes, causing hemolysis and ineffective erythropoiesis. Clinical data show that severity of b-thal correlates with the number of inherited a-globin genes (HBA1 and HBA2), with a-globin gene deletions having a beneficial effect for patients. Here, we describe a novel strategy to treat b-thal based on genome editing of the a-globin locus in human hematopoietic stem/progenitor cells (HSPCs). Using CRISPR/Cas9, we combined 2 therapeutic approaches: (1) a-globin downregulation, by deleting the HBA2 gene to recreate an a-thalassemia trait, and (2) b-globin expression, by targeted integration of a b-globin transgene downstream the HBA2 promoter. First, we optimized the CRISPR/Cas9 strategy and corrected the pathological phenotype in a cellular model of b-thalassemia (human erythroid progenitor cell [HUDEP-2] b0). Then, we edited healthy donor HSPCs and demonstrated that they maintained long-term repopulation capacity and multipotency in xenotransplanted mice. To assess the clinical potential of this approach, we next edited b-thal HSPCs and achieved correction of a/b globin imbalance in HSPC-derived erythroblasts. As a safer option for clinical translation, we performed editing in HSPCs using Cas9 nickase showing precise editing with no InDels. Overall, we described an innovative CRISPR/Cas9 approach to improve a/b globin imbalance in thalassemic HSPCs, paving the way for novel therapeutic strategies for b-thal.

Correction of b-thalassemia by CRISPR/Cas9 editing of the a-globin locus in human hematopoietic stem cells / Pavani, G.; Fabiano, A.; Laurent, M.; Amor, F.; Cantelli, E.; Chalumeau, A.; Maule, G.; Tachtsidi, A.; Concordet, J. -P.; Cereseto, A.; Mavilio, F.; Ferrari, G.; Miccio, A.; Amendola, M.. - In: BLOOD ADVANCES. - ISSN 2473-9529. - 5:5(2021), pp. 1137-1153. [10.1182/bloodadvances.2020001996]

Correction of b-thalassemia by CRISPR/Cas9 editing of the a-globin locus in human hematopoietic stem cells

Mavilio F.;
2021

Abstract

b-thalassemias (b-thal) are a group of blood disorders caused by mutations in the b-globin gene (HBB) cluster. b-globin associates with a-globin to form adult hemoglobin (HbA, a2b2), the main oxygen-carrier in erythrocytes. When b-globin chains are absent or limiting, free a-globins precipitate and damage cell membranes, causing hemolysis and ineffective erythropoiesis. Clinical data show that severity of b-thal correlates with the number of inherited a-globin genes (HBA1 and HBA2), with a-globin gene deletions having a beneficial effect for patients. Here, we describe a novel strategy to treat b-thal based on genome editing of the a-globin locus in human hematopoietic stem/progenitor cells (HSPCs). Using CRISPR/Cas9, we combined 2 therapeutic approaches: (1) a-globin downregulation, by deleting the HBA2 gene to recreate an a-thalassemia trait, and (2) b-globin expression, by targeted integration of a b-globin transgene downstream the HBA2 promoter. First, we optimized the CRISPR/Cas9 strategy and corrected the pathological phenotype in a cellular model of b-thalassemia (human erythroid progenitor cell [HUDEP-2] b0). Then, we edited healthy donor HSPCs and demonstrated that they maintained long-term repopulation capacity and multipotency in xenotransplanted mice. To assess the clinical potential of this approach, we next edited b-thal HSPCs and achieved correction of a/b globin imbalance in HSPC-derived erythroblasts. As a safer option for clinical translation, we performed editing in HSPCs using Cas9 nickase showing precise editing with no InDels. Overall, we described an innovative CRISPR/Cas9 approach to improve a/b globin imbalance in thalassemic HSPCs, paving the way for novel therapeutic strategies for b-thal.
2021
5
5
1137
1153
Correction of b-thalassemia by CRISPR/Cas9 editing of the a-globin locus in human hematopoietic stem cells / Pavani, G.; Fabiano, A.; Laurent, M.; Amor, F.; Cantelli, E.; Chalumeau, A.; Maule, G.; Tachtsidi, A.; Concordet, J. -P.; Cereseto, A.; Mavilio, F.; Ferrari, G.; Miccio, A.; Amendola, M.. - In: BLOOD ADVANCES. - ISSN 2473-9529. - 5:5(2021), pp. 1137-1153. [10.1182/bloodadvances.2020001996]
Pavani, G.; Fabiano, A.; Laurent, M.; Amor, F.; Cantelli, E.; Chalumeau, A.; Maule, G.; Tachtsidi, A.; Concordet, J. -P.; Cereseto, A.; Mavilio, F.; F...espandi
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11380/1248012
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