Illuminating gene therapy

项目摘要 在本计画中，我们将建立一种新的基因治疗定量小鼠模型。为了提高严谨性， 重复性，我们从单基因报告系统和过渡到多路报告 这些配置适合于宽范围的基因编辑实施例。具体来说，我们有 设计了由单体近红外荧光蛋白组成的表达盒， 深层组织测量，一种LacZ转基因，可在整个器官和组织学中提供快速可见染色 切片和实验室进化的红移荧光素酶，其被设计用于高度敏感的体内 测量.这种灵活的配置将有利于研究界，提供多个读出 调查员的选择。 所描述的小鼠被工程化以检测宽范围的编辑实施方案，包括胞嘧啶碱基突变。 编辑器，腺嘌呤碱基编辑器，Prime-Editors，CasFx，也许还有其他。这种“一体化”设计将允许 研究人员可以最大限度地减少所需动物的数量，从而节省时间和成本。它还促进严谨性， 由于使用相同动物的不同编辑技术可以更容易地定量地 比较不同群体进行的研究，使用不同的输送剂， CRISPR实施方案。在撰写本文时，还没有这样的鼠标模型可用，因此该项目将 为基因治疗领域提供了一种新颖独特的试剂。 我们已经组建了一个强大的团队来解决这个项目，由基础编辑器和主要编辑器的专家指导 （大卫刘和詹妮弗杜德纳）和小鼠遗传学（迈克尔麦克马纳斯）。该项目还带来了一个 具有小鼠模型基因治疗专业知识的博士后合作研究员（Wilson Poon），他开发了一种 设计用于wtCas 9基因编辑的小鼠传感器。不断增长的势头和不断增加的 有证据表明，基础编辑和主要编辑将成为临床竞赛的领跑者， 治疗学小鼠模型的开发将大大加快我们将基因编辑带入人类的目标。 帮助确保更安全和更有效的治疗。

PROJECT SUMMARY In this project, we will develop a new mouse model for quantitating gene therapy. To promote rigor and reproducibility, we move away from single gene reporter systems and transition to multiplexed reporter configurations that are amenable for a broad range of gene editing embodiments. Specifically, we have designed an expression cassette composed of a monomeric near-infrared fluorescence protein suitable for deep tissue measurements, a LacZ transgene that provides a rapid visible stain in whole organs and histology sections, and laboratory evolved red-shifted luciferase that is designed for highly sensitive in vivo measurements. This flexible configuration will benefit the research community, offering multiple readout options for investigators. The described mouse is engineered to detect a broad range of editing embodiments including Cytosine Base Editors, Adenine Base Editors, Prime-Editors, CasFx, and perhaps others. This ‘all-in-one’ design will allow researchers to minimize the number of animals needed, thus saving time and cost. It also promotes rigor and reproducibility since different editing technologies that use the same animal can be more readily quantitatively compared between research studies performed by different groups, using different delivery agents and CRISPR embodiments. As of this writing, there are no such mouse models available, hence this project will provide a novel and unique reagent for the gene therapy field. We have assembled a strong team to tackle this project, guided by experts in base editors and prime editors (David Liu and Jennifer Doudna) and mouse genetics (Michael McManus). The project also brings on board a postdoc co-investigator with expertise in gene therapy in mouse models (Wilson Poon) who has developed a mouse sensor designed for wtCas9 gene editing. Growing momentum and an ever-increasing body of evidence indicates that base editors and prime editors will become front-runners in the race to clinical therapeutics. The development of mouse models will greatly accelerate our goal to bring gene editing to the clinic, helping to ensure a safer and more efficacious treatment.