PROJECT SUMMARY OVERALL While the development of high-throughput sequencing technology and its application to clinical diagnostics has yielded the genetic basis for many rare genetic diseases, the development of effective treatments has not kept pace. Although gene replacement and modulation therapies can be powerful, sometimes even lifesaving treatment options, they come with many risks, such as immunogenicity and oncogenicity. Programmable nucleases such as CRISPR/Cas9 have revolutionized our ability to manipulate the genome, and provide the potential to achieve lasting, precise genome modification for therapeutic benefit. The proposed U19 program seeks to address these challenges through the development, validation and translation of gene editing– based therapeutic solutions for rare neurological genetic diseases. We propose to focus on four neurological conditions that each represent a significant unmet clinical need: Spinal Muscular Atrophy, Friedrich's Ataxia, Huntington's Disease, and Rett Syndrome. Members of our team have developed a suite of base and prime genome editing tools that can install precise alterations without creating a DSB or requiring a donor template. We also have developed validated in vivo mouse models for each of these diseases and bring deep expertise in the IND-enabling preclinical evaluation of gene-editing therapeutics. We propose to merge these considerable assets with disease-specific expertise in each of the four neurological conditions, supported by expertise and resources for scaled production of AAV-based delivery vectors for delivery of precision gene- editing therapies to tissues, and for navigating the regulatory path to IND submission. The proposed U19 team has a track record of individual and collaborative success at every step of the preclinical pipeline pathway and is thus well positioned to achieve our milestones, which include an IND package submitted to FDA for at least one therapy and neurological condition. Our Overall Aims are to: 1) Assemble a multi-disciplinary team with unique strengths and expertise to develop and implement innovative genome editing strategies to address important disease of the CNS, including Spinal Muscular Atrophy, Friedreich's Ataxia, Huntington's Disease, and Rett Syndrome; 2) Optimize lead base editor and prime editor candidates for each disease area, utilizing in vitro platforms and validated animal models; 3) Execute definitive preclinical in vivo pharmacology studies on optimized leads to develop reproducible efficacy data, while monitoring biodistribution, PK/PD, tolerability, and toxicology; and 4) Advance one lead candidate to an allowable investigational new drug (IND) application through coordinated communication with the FDA INTERACT program, the research project team, and the project Cores.

PROJECT NARRATIVE/RELEVANCE TO PUBLIC HEALTH Science and medicine have revolutionized our ability to diagnose rare genetic diseases, but patients and their families still have vanishingly limited treatment options. This is because of the challenges in precisely correcting genetic diseases using gene therapies that are both safe and effective. This U19 program seeks to address this vexing gap for four neurological conditions, with the goal of bringing at least one gene-editing therapeutic to the FDA within 5 years. Success in our program could revolutionize treatment for at least one of the conditions we will study and pave the path for the others in the near future.