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Efficient CRISPR-Cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients

Gene editing by the CRISPR-Cas9 nuclease system technology can be considered among the most promising strategies to correct hereditary mutations in a variety of monogenic diseases. In this paper, we present for the first time the correction, by CRISPR-Cas9 gene editing, of the β(0)39-thalassemia mut...

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Autores principales: Cosenza, Lucia Carmela, Gasparello, Jessica, Romanini, Nicola, Zurlo, Matteo, Zuccato, Cristina, Gambari, Roberto, Finotti, Alessia
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society of Gene & Cell Therapy 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8091488/
https://www.ncbi.nlm.nih.gov/pubmed/33997100
http://dx.doi.org/10.1016/j.omtm.2021.03.025
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author Cosenza, Lucia Carmela
Gasparello, Jessica
Romanini, Nicola
Zurlo, Matteo
Zuccato, Cristina
Gambari, Roberto
Finotti, Alessia
author_facet Cosenza, Lucia Carmela
Gasparello, Jessica
Romanini, Nicola
Zurlo, Matteo
Zuccato, Cristina
Gambari, Roberto
Finotti, Alessia
author_sort Cosenza, Lucia Carmela
collection PubMed
description Gene editing by the CRISPR-Cas9 nuclease system technology can be considered among the most promising strategies to correct hereditary mutations in a variety of monogenic diseases. In this paper, we present for the first time the correction, by CRISPR-Cas9 gene editing, of the β(0)39-thalassemia mutation, one of the most frequent in the Mediterranean area. The results obtained demonstrated the presence of normal β-globin genes after CRISPR-Cas9 correction of the β(0)39-thalassemia mutation performed on erythroid precursor cells from homozygous β(0)39-thalassemia patients. This was demonstrated by allele-specific PCR and sequencing. Accumulation of corrected β-globin mRNA and relevant “de novo” production of β-globin and adult hemoglobin (HbA) were found with high efficiency. The CRISPR-Cas9-forced HbA production levels were associated with a significant reduction of the excess of free α-globin chains. Genomic toxicity of the editing procedure (low indels and no off-targeting) was analyzed. The protocol might be the starting point for the development of an efficient editing of CD34(+) cells derived from β(0)39 patients and for the design of combined treatments using, together with the CRISPR-Cas9 editing of the β-globin gene, other therapeutic approaches, such as, for instance, induction of HbA and/or fetal hemoglobin (HbF) using chemical inducers.
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spelling pubmed-80914882021-05-14 Efficient CRISPR-Cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients Cosenza, Lucia Carmela Gasparello, Jessica Romanini, Nicola Zurlo, Matteo Zuccato, Cristina Gambari, Roberto Finotti, Alessia Mol Ther Methods Clin Dev Original Article Gene editing by the CRISPR-Cas9 nuclease system technology can be considered among the most promising strategies to correct hereditary mutations in a variety of monogenic diseases. In this paper, we present for the first time the correction, by CRISPR-Cas9 gene editing, of the β(0)39-thalassemia mutation, one of the most frequent in the Mediterranean area. The results obtained demonstrated the presence of normal β-globin genes after CRISPR-Cas9 correction of the β(0)39-thalassemia mutation performed on erythroid precursor cells from homozygous β(0)39-thalassemia patients. This was demonstrated by allele-specific PCR and sequencing. Accumulation of corrected β-globin mRNA and relevant “de novo” production of β-globin and adult hemoglobin (HbA) were found with high efficiency. The CRISPR-Cas9-forced HbA production levels were associated with a significant reduction of the excess of free α-globin chains. Genomic toxicity of the editing procedure (low indels and no off-targeting) was analyzed. The protocol might be the starting point for the development of an efficient editing of CD34(+) cells derived from β(0)39 patients and for the design of combined treatments using, together with the CRISPR-Cas9 editing of the β-globin gene, other therapeutic approaches, such as, for instance, induction of HbA and/or fetal hemoglobin (HbF) using chemical inducers. American Society of Gene & Cell Therapy 2021-04-03 /pmc/articles/PMC8091488/ /pubmed/33997100 http://dx.doi.org/10.1016/j.omtm.2021.03.025 Text en © 2021 The Authors https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Original Article
Cosenza, Lucia Carmela
Gasparello, Jessica
Romanini, Nicola
Zurlo, Matteo
Zuccato, Cristina
Gambari, Roberto
Finotti, Alessia
Efficient CRISPR-Cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients
title Efficient CRISPR-Cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients
title_full Efficient CRISPR-Cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients
title_fullStr Efficient CRISPR-Cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients
title_full_unstemmed Efficient CRISPR-Cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients
title_short Efficient CRISPR-Cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients
title_sort efficient crispr-cas9-based genome editing of β-globin gene on erythroid cells from homozygous β(0)39-thalassemia patients
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8091488/
https://www.ncbi.nlm.nih.gov/pubmed/33997100
http://dx.doi.org/10.1016/j.omtm.2021.03.025
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