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Mechanobiology

Nonviral Delivery of CRISPR-Cas9 Using Protein-Agnostic, High-Loading Porous Silicon and Polymer Nanoparticles

AUTHORS

Fletcher RBR Brock , Stokes LDLarry D , Kelly IBIsom B , Henderson KMKatelyn M , Vallecillo-Viejo ICIsabel C , Colazo JMJuan M , Wong BVBenjamin V , Yu FFang , d'Arcy RRichard , Struthers MNMorgan N , Evans BCBrian C , Ayers JJacob , Castanon MMatthew , Weirich MJMichael J , Reilly SKSarah K , Patel SSShrusti S , Ivanova YIYoanna I , Silvera Batista CACarlos A , Weiss SMSharon M , Gersbach CACharles A , Brunger JMJonathan M , Duvall CLCraig L . ACS nano. 2023 08 15; ().

ABSTRACT

The complexity of CRISPR machinery is a challenge to its application for nonviral therapeutic gene editing. Here, we demonstrate that proteins, regardless of size or charge, efficiently load into porous silicon nanoparticles (PSiNPs). Optimizing the loading strategy yields formulations that are ultrahigh loading─>40% cargo by volume─and highly active. Further tuning of a polymeric coating on the loaded PSiNPs yields nanocomposites that achieve colloidal stability under cryopreservation, endosome escape, and gene editing efficiencies twice that of the commercial standard Lipofectamine CRISPRMAX. In a mouse model of arthritis, PSiNPs edit cells in both the cartilage and synovium of knee joints, and achieve 60% reduction in expression of the therapeutically relevant MMP13 gene. Administered intramuscularly, they are active over a broad dose range, with the highest tested dose yielding nearly 100% muscle fiber editing at the injection site. The nanocomposite PSiNPs are also amenable to systemic delivery. Administered intravenously in a model that mimics muscular dystrophy, they edit sites of inflamed muscle. Collectively, the results demonstrate that the PSiNP nanocomposites are a versatile system that can achieve high loading of diverse cargoes and can be applied for gene editing in both local and systemic delivery applications.