Prototype Mouse Models for SEND-Based mRNA Delivery and Technology Development - Many human diseases are caused by faulty genes. The best way to treat them is by gene therapy, a concept that has been around for over four decades. One of the major challenges of gene therapy is the lack of reliable technologies to deliver healthy gene copies to target cells. Some commonly used delivery approaches include viral vectors or nucleic acids coated with lipid-based nanoparticles, but their success in clinical applications, particularly for gene therapy is limited. The revolutionary CRISPR-Cas gene editing system, developed about a decade ago, offers a solution, but it too suffers from the delivery problem. A recent report uncovered a surprising inter cellular mRNA transfer process, termed SEND (for selective endogenous encapsidation for cellular delivery; Segel et al., Science: V373; 6557, p882), which provides an elegant solution to the delivery problem. The SEND system utilizes a cellular protein, PEG10, which has the ability to form a capsid-like protein coat on its own mRNA. PEG10 viral-like particles (VLPs) containing cargo (either Peg10 mRNA itself or an engineered heterologous mRNA), coated with another protein called fusogen, exit cells and attach to other cells, delivering their cargo. Our preliminary results show that the SEND-VLP system works in vivo in mice. While this initial work demonstrates the power of the SEND system for gene-therapy applications, the real potential of SEND—as a much-needed delivery tool—could be exploited once we understand more about the biogenesis of VLPs in mammalian tissues and their role in development, physiology, and pathology. The task of systematic in vivo investigation of this newly discovered phenomenon requires the development of a variety of carefully designed genetically engineered mouse models (GEMMs). Specifically, we have conceived a suite of GEMMs having inducible and conditional potential to produce VLPs in a tissue of choice or at a specified time in the mouse lifespan, including aging, as well as in disease conditions including fetal diseases (applicable for Intra Uterine Gene Therapy; IUGT). As part of this award, we have selected three prototypes of GEM models suitable for SEND-VLP research—one each for understanding the physiology of intercellular mRNA transfer, for delivering CRISPR-Cas9 tools to a target tissue (e.g., liver) to treat genetic diseases in adult mouse and for IUGT to treat fetal diseases. One of the major advantages of the SEND system is that it uses endogenous proteins and therefore it is thought to be non-immunogenic and non-toxic. Once we learn more about the SEND-VLP system, it could be re-purposed as a gene therapy tool to apply for numerous human diseases for which there are on-going clinical trials that rely on viral vectors or nano-formulations; the drawbacks of which are immunogenicity and toxicity. The ultimate goal of this proposal is to repurpose the SEND system as a delivery tool for gene therapy.
