| RRC ID |
62313
|
| 著者 |
Li C, Psatha N, Gil S, Wang H, Papayannopoulou T, Lieber A.
|
| タイトル |
HDAd5/35++ Adenovirus Vector Expressing Anti-CRISPR Peptides Decreases CRISPR/Cas9 Toxicity in Human Hematopoietic Stem Cells.
|
| ジャーナル |
Mol Ther Methods Clin Dev
|
| Abstract |
We generated helper-dependent HDAd5/35++ adenovirus vectors expressing CRISPR/Cas9 for potential hematopoietic stem cells (HSCs) gene therapy of β-thalassemia and sickle cell disease through re-activation of fetal γ-globin expression (HDAd-globin-CRISPR). The process of CRISPR/Cas9 gene transfer using these vectors was not associated with death of human CD34+ cells and did not affect their in vitro expansion and erythroid differentiation. However, functional assays for primitive HSCs, e.g., multi-lineage progenitor colony formation and engraftment in irradiated NOD/Shi-scid/interleukin-2 receptor γ (IL-2Rγ) null (NSG) mice, revealed toxicity of HDAd-globin-CRISPR vectors related to the prolonged expression and activity of CRISPR/Cas9. To control the duration of CRISPR/Cas9 activity, we generated an HDAd5/35++ vector that expressed two anti-CRISPR (Acr) peptides (AcrII4 and AcrII2) capable of binding to the CRISPR/Cas9 complex (HDAd-Acr). CD34+ cells that were sequentially infected with HDAd-CRISPR and HDAd-Acr engrafted at a significantly higher rate. Target site disruption frequencies in engrafted human cells were similar to those in pre-transplantation CD34+ cells, indicating that genome-edited primitive HSCs survived. In vitro differentiated HSCs isolated from transplanted mice demonstrated increased γ-globin expression as a result of genome editing. Our data indicate that the HDAd-Acr vector can be used as a tool to reduce HSC cytotoxicity of the CRISPR/Cas9 complex.
|
| 巻・号 |
9
|
| ページ |
390-401
|
| 公開日 |
2018-6-15
|
| DOI |
10.1016/j.omtm.2018.04.008
|
| PII |
S2329-0501(18)30046-9
|
| PMID |
30038942
|
| PMC |
PMC6054697
|
| IF |
4.533
|
| リソース情報 |
| ヒト・動物細胞 |
HUDEP-2(RCB4557) |
