Recent advances in the use of ZFN-mediated gene editing for human gene therapy

Abstract

Targeted genome editing with programmable nucleases has revolutionized biomedical research. The ability to make site-specific modifications to the human genome, has invoked a paradigm shift in gene therapy. Using gene editing technologies, the sequence in the human genome can now be precisely engineered to achieve a therapeutic effect. Zinc finger nucleases (ZFNs) were the first programmable nucleases designed to target and cleave custom sites. This article summarizes the advances in the use of ZFN-mediated gene editing for human gene therapy and discusses the challenges associated with translating this gene editing technology into clinical use.

Gene editing technology for improving life quality: A dream coming true?

“…The main characteristics of the designer nucleases [11][12][13][14] Guanine-rich region The ability to recognize any certain sequence of DNA triplets; Small protein size High-priced, labor-intensive, and timetaking; Unintended change in DNA; restricted target length to multiples of three; laborious zinc finger engineering and sequence selection TALEN Recently, Anzalone et al 22 have reported construction of a searchand-replace genome editing CRISPR based method, named prime editing, which consist of a catalytically impaired Cas9 fused to an engineered reverse transcriptase, employing a prime editing guide RNA (pegRNA) with the ability to specify the target sequence and edit the favored position. Nickase Cas9 (nCas9) proteins are another form of Cas9 modifications involving a single mutated nucleolytic domain while the other domain preserves its cleavage activity and therefore produce single strand breaks (SSBs).…”

“…ZFN-mediated CCR5 disruption, was the first application of genome editing tools in human in order to prevent the entry of HIV into the cells. 12 Table 1 summarizes the main properties of this nuclease along with the other genome editing tools. The key for opening the third gate to the gene editing world was in the hands of a plant pathogen called Xanthomonas which in 2009 was discovered to have DNA sequence specific transcription activator-like effector (TALE) proteins 19 with each repeat domain having the ability to distinguish a single base.…”

Creating cell and animal models of human disease by genome editing using CRISPR/Cas9

“…Amino acid replacement in DNA-binding proteins from ZFNs has also been a topic of interest for research. 18 Nevertheless, increasing the efficiency of ZFNs, as a tool that facilitates genetic manipulation for purposes such as the development of novel human disease models and gene therapy strategies, is still a matter of debate (Table 1).…”

“…In a study recently carried out aiming to improve specificity and reduce toxicity, an increase in the number of ZF motifs, as well as the inactivation of the catalytic component of ZFNs, was shown that lead to the creation of an enzyme called ZF nickase. Amino acid replacement in DNA‐binding proteins from ZFNs has also been a topic of interest for research . Nevertheless, increasing the efficiency of ZFNs, as a tool that facilitates genetic manipulation for purposes such as the development of novel human disease models and gene therapy strategies, is still a matter of debate (Table ).…”

Designer Nucleases: Gene-Editing Therapies using CCR5 as an Emerging Target in HIV

“…In fact, the higher the nuclease specificity, the lower the nuclease off-target cleavage and henceforth associated-toxicity. 104 Several studies highlighted the fact that the existent analogous architecture between ZFNs and TALENs seemed preponderant to achieve a fruitful clinical setting through the development of the following mutual nuclease-criteria: 105 A. High specificity of the DNA-binding domain…”