دانلود رایگان مقاله سیستم CRISPR / Cas9 و کاربرد آن در سلول های خونساز انسان

Abstract

Since 2012, the CRISPR-Cas9 system has been quickly and successfully tested in a broad range of organisms and cells including hematopoietic cells. The application of CRISPR-Cas9 in human hematopoietic cells mainly involves the genes responsible for HIV infection, β-thalassemia and sickle cell disease (SCD). The successful disruption of CCR5 and CXCR4 genes in T cells by CRISPR-Cas9 promotes the prospect of the technology in the functional cure of HIV. More recently, eliminating CCR5 and CXCR4 in induced pluripotent stem cells (iPSCs) derived from patients and targeting the HIV genome have been successfully carried out in several laboratories. The outcome from these approaches bring us closer to the goal of eradicating HIV infection. For hemoglobinopathies the ability to produce iPSC-derived from patients with the correction of hemoglobin (HBB) mutations by CRISPR-Cas9 has been tested in a number of laboratories. These corrected iPSCs also show the potential to differentiate into mature erythrocytes expressing high-level and normal HBB. In light of the initial success of CRESPR-Cas9 in target mutated gene(s) in the iPSCs, a combination of genomic editing and autogenetic stem cell transplantation would be the best strategy for root treatment of the diseases, which could replace traditional allogeneic stem cell transplantation.

6. Summary and outlook

Over the last few years, the development of several gene editing technologies, ZFN, TALEN, and CRESPR-Cas9, has greatly promoted biology research and clinical trials for gene therapy. Among these three biotechnologies CRESPR-Cas9 is the latest player in the field but received great attention and applications. The main advantage of CRISPR-Cas9 is its simplicity, efficiency, and versatility for gene manipulations. Targeting efficiency is one of the most important parameters for assessing a genome-editing tool especially the one used for gene therapy. Although a number of reports have confirmed that CRISPR-Cas9 has better targeting efficiency than ZFNs and TALENs, the efficiency still varies with cell types and research labs. For example, in zebrafish the targeting efficiency by CRISPR-Cas9 system has reached to 100% [46] and in one-cell mouse embryo 78% [108]. In contrast, in human CD4+ T cells the Cas9 system obtains efficiencies ranging from 5% to 20% [71,86] and in induced pluripotent stem cells only 2–5% [70]. Therefore, further improvement of the target efficiency and stability in hematopoietic cells is necessary. Moreover, it is unclear whether and how CRISPRCas9 is able to target CCR5 or CXCR4 effectively in all CD4+ T cells in vivo.