RESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMO
Gene correction in human long-term hematopoietic stem cells (LT-HSCs) could be an effective therapy for monogenic diseases of the blood and immune system. Here we describe an approach for X-linked sSevere cCombined iImmunodeficiency (SCID-X1) using targeted integration of a cDNA into the endogenous start codon to functionally correct disease-causing mutations throughout the gene. Using a CRISPR-Cas9/AAV6 based strategy, we achieve up to 20% targeted integration frequencies in LT-HSCs. As measures of the lack of toxicity we observe no evidence of abnormal hematopoiesis following transplantation and no evidence of off-target mutations using a high-fidelity Cas9 as a ribonucleoprotein complex. We achieve high levels of targeting frequencies (median 45%) in CD34+ HSPCs from six SCID-X1 patients and demonstrate rescue of lymphopoietic defect in a patient derived HSPC population in vitro and in vivo. In sum, our study provides specificity, toxicity and efficacy data supportive of clinical development of genome editing to treat SCID-Xl.
Assuntos
DNA Complementar/genética , Edição de Genes/métodos , Transplante de Células-Tronco Hematopoéticas , Subunidade gama Comum de Receptores de Interleucina/genética , Doenças por Imunodeficiência Combinada Ligada ao Cromossomo X/terapia , Animais , Antígenos CD34/metabolismo , Sistemas CRISPR-Cas/genética , Linhagem Celular , Códon de Iniciação/genética , Dependovirus , Éxons/genética , Sangue Fetal/citologia , Vetores Genéticos/genética , Voluntários Saudáveis , Células-Tronco Hematopoéticas/metabolismo , Humanos , Masculino , Camundongos , Mutação , Parvovirinae/genética , Cultura Primária de Células , Fatores de Tempo , Transdução Genética/métodos , Quimeras de Transplante/genética , Transplante Heterólogo/métodos , Doenças por Imunodeficiência Combinada Ligada ao Cromossomo X/genéticaRESUMO
The original version of this Article omitted the following from the Acknowledgements: "G.B. acknowledges the support from the Cancer Prevention and Research Institute of Texas (RR140081 and RR170721)."This has now been corrected in both the PDF and HTML versions of the Article.