Engineering Human ARMMs as Therapeutic Non-Viral Vehicles forin vivoDelivery of Genome Editing Payloads

• DOI: 10.1101/2024.09.24.614725
• Publication Date: 2024-09-26T16:45:28Z
• Authors: Qiyu Wang; Shu-Lin Liu; Wen-Ning Zhao; Ryan von Kleeck; Michael Thomas; Kristin Luther; Zahra Vahramian; Yumei Sun; Komal Vyas; Pearl Moharil; Chris Chatzinakos; Carson O. Semple; Jennifer A. Badji; Leah Gens; Alysia Bryant; Steve Greenway; Chou-Wei Chang; Adam Francoeur; Nedyalka Valkov; Killian O’Brien; Joseph F. Nabhan

The ability to edit the human genome treat previously incurable diseases or modify undruggable targets has potential transform lives of patients. However, delivery editing machineries in vivo, and a transient manner, continues be major challenge. We engineered cell-derived ARRDC1-mediated microvesicles (ARMMs) as non-viral vehicles that mediate intracellular proteins, including DNA-modifying enzymes such Cre recombinase Cas9/guide RNA ribonucleoproteins (RNPs). ARMMs packaged multiple payloads per vesicle our production system, yielding dose-dependent functional Cas9/gRNA RNP variety cell types, primary mouse cells. Oropharyngeal aspiration loaded with led biodistribution alveolar macrophages (AMs), whereas intravenous administration predominantly Kupffer cells (KCs), liver sinusoidal endothelial (LSECs), splenocytes. Suprisingly, one Cas9 RNPs by oropharyngeal intravenously resulted efficient knockout genes AMs KCs, suggesting therapeutic utility. Through Cas9/NLRP3 gRNA we showed amelioration drug-induced injury model, supporting use for editor vivo.