Gene correction of HBB mutations in CD34+ hematopoietic stem cells using Cas9 mRNA and ssODN donors.

ABSTRACT: BACKGROUND:?-Thalassemia is an inherited hematological disorder caused by mutations in the human hemoglobin beta (HBB) gene that reduce or abrogate ?-globin expression. Although lentiviral-mediated expression of ?-globin and autologous transplantation is a promising therapeutic approach, the risk of insertional mutagenesis or low transgene expression is apparent. However, targeted gene correction of HBB mutations with programmable nucleases such as CRISPR/Cas9, TALENs, and ZFNs with non-viral repair templates ensures a higher safety profile and endogenous expression control. METHODS:We have compared three different gene-editing tools (CRISPR/Cas9, TALENs, and ZFNs) for their targeting efficiency of the HBB gene locus. As a proof of concept, we studied the personalized gene-correction therapy for a common ?-thalassemia splicing variant HBBIVS1-110 using Cas9 mRNA and several optimally designed single-stranded oligonucleotide (ssODN) donors in K562 and CD34+ hematopoietic stem cells (HSCs). RESULTS:Our results exhibited that indel frequency of CRISPR/Cas9 was superior to TALENs and ZFNs (P?

SUBMITTER: Antony JS 

PROVIDER: S-EPMC6236008 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature