Novel, synthetic gene editing technology for gene knock-ins without DNA cleavage

基因组编辑领域的最新进展使我们更接近修复人类患者缺陷基因的梦想。在过去的20年里，可编程核酸酶技术的连续王朝已被应用于这个问题。从锌指核酸酶，通过TALEN到CRISPR-Cas-核酸酶，存在增加此类系统灵活性的趋势。几个系统已经被改造为使用替代的效应部分嵌入在这些系统的第一个版本的核酸酶，允许靶细胞修饰通过更好的耐受性和更可控的变化，染色体DNA.What是失踪的，是一个灵活和有效的方法，引入新的遗传物质在一个基因组的一个确定的基因座，而不会产生双链（ds）断裂。这种ds断裂如果不迅速修复是致命的，它们也被认为是对基因组完整性的威胁，因此可以很容易地触发靶细胞中的程序性细胞死亡。生物科学公司已经创建并正在开发一种完全合成的基因组调控系统（ApGet），并寻求资金来调整该系统，以在关键靶位点提供高效的供体DNA序列敲入。ApGet系统体积小（约0.7 kb），模块化，组成非CRISPR。它能够通过RNA引导将不同的效应蛋白募集到宿主基因组中的特定位点。ApGet计划将ApGet的一些组件替换为其他组件，从而通过一种根本不同的机制进行敲入，这种机制不会诱导dsDNA断裂。其目的是将供体DNA募集到目标位点，以便为基因修复和工程细胞疗法的产生提供安全、耐受性良好和高效的敲入编辑。从长远来看，基因编辑可能会为减轻遗传疾病的负担做出重大贡献，仅在英国就有超过300万人受到遗传疾病的影响。这些疾病中的许多最好用“一次性”治疗，包括基因敲入介导的等位基因缺陷部分的替换。

Recent advances in the field of genome editing have moved us closer to the dream of repairing defective genes in human patients. Over the last 20 years, successive dynasties of programmable nuclease technologies have been applied to this problem. From zinc-finger nucleases, through TALENs to CRISPR--Cas- nucleases there has been a trend towards increasing flexibility of such systems. Several of the systems have been adapted to use alternative effector moieties to the nucleases embedded in the first versions of these systems, allowing target cell modification via better tolerated and more controllable changes to chromosomal DNA.What is missing however, is a flexible and effective method for introducing new genetic material at a defined locus of a genome without creating double-strand (ds) breaks. Such ds breaks are lethal if not rapidly repaired, and they are also recognized as a threat to genome integrity and hence can easily trigger programmed cell death in target cells.Pencil Biosciences has created, and is developing, a fully-synthetic genome modulation system (ApGet) and seeks funding to adapt the system to deliver highly-efficient knock-in of donor DNA sequences at key target sites. The ApGet system is small in size (~0.7 kb), modular and non-CRISPR in composition. It is capable of recruiting diverse effector proteins to specific sites in the host genome by means of RNA-guides.Pencil plans to replace some of ApGet's components with others allowing knock-ins via a fundamentally different mechanism that does not induce a dsDNA break. The aim is to recruit donor DNA to target sites of interest in order to deliver safe, well-tolerated and highly efficient knock-in edits for gene repair and the generation of engineered cell therapies.In the long term gene editing may make a significant contribution to reducing the burden of genetic diseases, which in the UK alone affect more than 3 million people. Many of these diseases would be best treated with a 'one and done' treatment involving a knock-in-mediated replacement of the defective section of an allele.