Over the last decade, CRISPR/Cas gene-editing technology has been a transformational tool to allow efficient and accurate DNA manipulations for rapid analysis of gene function, mutation and variations in model systems. This allows for human disease models, which can then be used to test new therapeutic and diagnostic strategies. The technology itself is also being used for clinical gene therapy, involving the manipulation and correction of genetic mutations that contribute to disease. There has already been an FDA-approved therapy based on this technology and there are many clinical trials in progress.
Despite this rapid progress and the huge potential for gene editors as a new class of therapeutics, a major scientific bottleneck exists, namely the ability to target a wide variety of cell types in vivo. Due to the avid uptake of the gene editing vector, whether as lipid nanoparticles or a virus, by the liver, much of the recent work has focused on correcting genes in hepatocytes which cause rare diseases. In order to deploy this technology more widely for a variety of diseases, it is important to understand how to target cells beyond the liver, such as in the heart or central nervous system. In parallel, there needs to be a greater understanding of the genetic stability of cells which have been edited in vivo. This project will develop next generation gene editors which are honed to specific cell types and explore the safety profiles such as on and off target effects.
The student will work with genetically modified mouse models and pluripotent stem cells to develop components of base and prime editors that can be packaged efficiently for delivery to target tissues. The ultimate goal is to create a modular delivery system for somatic editing, enabling the rapid exchange of different CRISPR/Cas tools and tissue-targeting modalities to address specific mutations in defined target cells. In the first year, the student would establish and characterise a genetically modified mouse line that expresses an inactivated CRISPR/Cas9, which serves as an RNA-guided DNA-binding protein. They would begin to optimise methodologies for linking this deactivated Cas9 to base- and prime-editing machinery using in vitro systems such as induced pluripotent stem cells. Years 2–3 would see the student focus on testing delivery methods in vivo, once the mouse model and modular system have been established. In the final year, they would address a proof-of-concept disease model, applying the developed systems in vivo to correct underlying mutations in the target organ.
The rotation project will focus on validating gene editing technologies in vitro. This includes selecting a candidate Prime and Base editor, producing the necessary components for in vitro delivery, delivering them into various cell lines, and analysing the outcomes and efficiency of gene editing.