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Gene Editing Therapy 1

source:未知 author:admin Release time:2020-02-15 12:04 Browse volume:
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Recently, researchers at the Tongji Medical College of Huazhong University of Science and Technology have used gene editing technology-TALENs-to prepare syngeneic leukemia cell clones and destroy the FLT3 gene, thus proving that this genome editing method is the molecular basis for exploring gene mutation Powerful platform. The relevant research results were published in the most recent Nature journal, Scientific Reports.

Acute leukemia (AL) is the most common hematological malignancy, showing significant heterogeneity in genetic makeup and clinical outcomes. Thanks to a deeper understanding of cytogenetics / gene mutations and the introduction of corresponding targeted therapies, amazing success has been achieved in the treatment of certain leukemia subtypes. However, many genetically high-risk subtypes currently use standard protocols It remains difficult to treat and shows a poor prognosis. The World Health Organization and the National Comprehensive Cancer Network increasingly classify AL based on genetic predisposition, highlighting the profound impact of cytogenetics / gene mutations on clinical management decision making. Therefore, it is urgent to analyze the molecular pathological basis of these AL patients in order to develop new treatment strategies.

 

Recently, researchers at the Tongji Medical College of Huazhong University of Science and Technology have used gene editing technology-TALENs-to prepare syngeneic leukemia cell clones and destroy the FLT3 gene, thus proving that this genome editing method is the molecular basis for exploring gene mutation Powerful platform. The relevant research results were published in the most recent Nature journal, Scientific Reports. Professor Zhou Jianfeng and Zhen Shang of Tongji Medical College, Huazhong University of Science and Technology are the co-corresponding authors of this article.

 

Compared with candidate gene analysis, the progress made by high-throughput sequencing technology allows us to characterize AL in a fair and comprehensive manner from a genome-wide perspective. The identification of high-recurrence gene mutations in AL provides valuable basic principles for improving diagnostic methods, patient stratification, and targeted therapy. However, although detailed AL genome sketches have emerged, it is difficult to clarify how these frequently occurring genetic mutations correlate with clinical phenotypes and treatment outcomes. It is necessary to determine the clinical relevance of these individuals' genetic abnormalities. More importantly, unveiling a key molecular event in leukemia biology—a major challenge in the era of the human genome—is fundamentally important for future genetic medicine.

 

Several methods have been widely used to explore the molecular mechanism of a given gene mutation. Using genetic engineering strategies, researchers have established many animal models of leukemia and provided persuasive insights into the drivers of leukemia. In the next few years, genetically engineered animal models will remain the most reliable tool for “studying the association between genetic defects and clinical phenotypes” and will help design and develop new molecular targeting strategies.

 

However, the animal model approach has some inherent disadvantages. For example, animal models may not always accurately simulate clinical background-related leukemia phenotypes, and the process of generating and maintaining transgenic models is also time-consuming and expensive. In addition, cell line and master sample-based models are also widely used to explore key molecular events that lead to tumor cell phenotypes. The huge genetic differences between the main samples, the non-physiological level of transgene overexpression / knockout, and the interference introduced by random integration of viral vectors all limit the actual molecular mechanisms. Therefore, in the post-genomic era, new analytical tools are urgently needed to elucidate the molecular mechanisms of leukemia-related genes.

 

Transcription activator-like effector (TALE) nucleases (TALENs)-an efficient genome editing tool, is an artificial fusion protein, contains the catalytic domain of the endonuclease FokI and a designed TALE DNA binding domain — -Identify specific DNA sequences. Two separate TALENs bind to adjacent DNA sequences, enabling FokI to dimerize and the target DNA to be cleaved, thereby introducing site-specific double-strand breaks (DSBs).
 

Cellular DNA repair is then activated through homology-directed repair or non-homologous end-junction (NHEJ) pathways. To date, genome editing technology has been successfully used to induce bone marrow malignancies in normal hematopoietic stem cells in mice. However, few studies have used this technique to investigate molecular events caused by individual genetic abnormalities in leukemia.

 

In this study, the researchers prepared syngeneic leukemia cell clones and used TALENs to disrupt the FLT3 gene. The allogeneic clone of FLT3 with single allele disruption was compared with the wild-type control clone of the allele and the transcriptional expression, downstream FLT3 signal and proliferation ability of the parental leukemia cells. Using RNA-seq, researchers compared the entire gene expression profiles of the mutant K562 clone and its corresponding wild-type control group.

 

It was found that the transcription level and ligand-dependent autophosphorylation of FLT3 were decreased in mutant clones. TALENs-mediated deficiency of the FLT3 gene can damage cell proliferation and colony formation in vitro. This inhibitory effect is maintained in vivo, which improves the survival rate of NOD / SCID mice transplanted with the mutant K562 clone. Cluster analysis showed that the gene expression pattern of syngeneic clones was determined by the state of the FLT3 mutant-not the deviation between individual syngeneic clones. Genes that are differentially expressed between the mutant and wild-type, reveal the non-sense-mediated activation of the decay pathway in mutant K562 clones and suppressed FLT3 signaling.

 

All in all, this research supports that this method of genome editing is a powerful and universally applicable platform for exploring the molecular basis of gene mutations.
 TALENs-mediated gene disruption of FLT3 in leukemia cells: Using genome-editing approach for exploring the molecular basis of gene abnormality
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TALENs-mediated gene disruption of FLT3 in leukemia cells: Using genome-editing approach for exploring the molecular basis of gene abnormality
TALENs-mediated gene disruption of FLT3 in leukemia cells: Using genome-editing approach for exploring the molecular basis of gene abnormality
Literature search: doi: 10.1038 / srep18454 Link source: http://www.biodiscover.com/news/resear

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