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Currently, CRISPR\/Cas9 delivery into cells relies primarily on viral vectors, which suffer from limitations in packaging capacity and safety concerns. These issues with a nonviral delivery strategy are addressed, where Cas9\u2022sgRNA ribonucleoprotein (RNP) complexes can be encapsulated into supramolecular nanoparticles (SMNP) to form RNP\u2282SMNPs, which can then be delivered into targeted cells via supramolecular nanosubstrate\u2010mediated delivery. Utilizing the U87 glioblastoma cell line as a model system, a variety of parameters for cellular\u2010uptake of the RNP\u2010laden nanoparticles are examined. Dose\u2010 and time\u2010dependent CRISPR\/Cas9\u2010mediated gene disruption is further examined in a green fluorescent protein (GFP)\u2010expressing U87 cell line (GFP\u2010U87). The utility of an optimized SMNP formulation in co\u2010delivering Cas9 protein and two sgRNAs that target deletion of exons 45\u201355 (708 kb) of the dystrophin gene is demonstrated. Mutations in this region lead to Duchenne muscular dystrophy, a severe genetic muscle wasting disease. Efficient delivery of these gene deletion cargoes is observed in a human cardiomyocyte cell line (AC16), induced pluripotent stem cells, and mesenchymal stem cells.<\/jats:p>","DOI":"10.1002\/smll.202100546","type":"journal-article","created":{"date-parts":[[2021,6,9]],"date-time":"2021-06-09T12:47:22Z","timestamp":1623242842000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":16,"title":["Supramolecular Nanosubstrate\u2010Mediated Delivery for CRISPR\/Cas9 Gene Disruption and Deletion"],"prefix":"10.1002","volume":"17","author":[{"given":"Qian","family":"Ban","sequence":"first","affiliation":[{"name":"School of Life Sciences Center for Stem Cell and Translational Medicine Anhui University  Hefei 230601 China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peng","family":"Yang","sequence":"additional","affiliation":[{"name":"Department of Molecular and Medical Pharmacology David Geffen School of Medicine 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