A programmable, cell-agnostic DNA nano-technology platform for CRISPR gene editing

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CRISPR-Cas9 gene editing has warranted its developers the 2020 Nobel Prize for Chemistry, in view of the massive impact that this technology is having on biological, biotechnological and medical research. In particular CRISPR gene editing is central to next generation cell-based therapies to treat cancer and other diseases, in which some of the patient's cells are extracted, genetically modified to fulfil specific functions, and then re-injected into the patient. This process is however time consuming and very costly, limiting the diffusion of these potentially life-saving therapies. One of the reasons behind the prohibitive costs is the low efficiency with which one can deliver to the cells the biological machinery required to perform CRISPR gene editing, both at scale and without excessive toxicity (leading to cell death).In this project, we will develop a novel and alternative approach to delivering CRISPR machinery to mammalian cells in vitro. Our approach will rely on specifically designed vectors, which we dub Editosomes. These are microscopic enclosures constructed from lipid membranes, similar to cell membranes, and containing large quantities of the CRISPR machinery.For Editosomes to deliver the machinery to the target cells the two would have to fuse. We will induce fusion by decorating both Editosomes and the target cells with artificial "fusogenic" nanomachines, that by binding to each other bring the cell and Editosome membranes to within a very short distance, ultimately making them merge. The fusogenic nanostructures will be constructed from synthetic DNA molecules, which are particularly suitable for engineering nanodevices in view of the very high selectivity and programmability of the base-pairing interactions.We envisage that Editosome technology will have a direct and profound impact on our ability to perform high-throughput, efficient, CRISPR gene editing in vivo, and thus on the accessibility and economic sustainability of the therapeutic technologies that rely on it.Additionally, we will clarify fundamental aspects of the (bio)physics underling lipid membrane stability, fusion, and the ability of DNA nanostructures to modulate them.