ABSTRACT Recombinant adeno-associated viruses (AAV) have emerged as safe and effective vectors for clinical gene therapy applications including systemic treatment of neuromuscular diseases such as Spinal Muscular Atrophy (SMA), Duchenne Muscular Dystrophy (DMD), and Giant Axonal Neuropathy (GAN) amongst others. However, enabling gene editing therapeutics to treat human disease requires improved systems that achieve effective genome editing at low systemic AAV vector doses in multiple animal models. Additionally, the ideal target cell types for gene editing, including progenitor cells, are distinct from the target cells that conventional gene delivery vectors have been optimized for. Genome editing in neuromuscular tissue, in particular, is challenging. To date, no effective non-viral delivery vehicles have been identified for widespread genome editing in musculoskeletal and neurological tissue following systemic administration. Further, in case of AAV vectors, several challenges pertinent to delivery of genome editors remain. The rationale for our current proposal stems from barriers posed by (i) species-related differences observed in AAV tropism, (ii) presence of pre-existing neutralizing antibodies to natural AAV, (iii) the need for high systemic AAV doses to achieve neuromuscular gene transfer, (iv) vector dose-related toxicity as indicated by detection of liver transaminases in patients, and (v) the unique opportunity to correct progenitor cells in situ. To address these aspects, we have assembled a collaborative team with cross- cutting expertise and developed a comprehensive and innovative approach to evolve high potency AAV variants for systemic neuromuscular genome editing. Specifically, we will focus our efforts on evolving high potency AAV vectors for neuromuscular genome editing using a three-tiered approach applied across different species, tissues and cell types.