Significance: Genome editing technologies enable the permanent repair of disease-causing genetic mutations. However, the application of this technology has been limited by the technical challenge of achieving safe, effective, and specific in vivo delivery of the CRISPR-Cas9 genome editing components. Here, we report the development of a newly identified lipid nanoparticle (LNP) for specific delivery of CRISPR-Cas9 mRNA to the liver. While LNPs have been FDA approved for delivery of siRNA to the liver, here we examine their application for genome editing. When compared head-to-head, our delivery platform significantly outperforms the FDA-approved LNP in the efficient delivery of Cas9 mRNA for knockdown of theAngptl3gene and subsequent regulation of hypercholesterolemia, while matching the safety and specificity of the approved platform.

Leukemia stem cells (LSCs) sustain the disease and contribute to relapse in acute myeloid leukemia (AML). Therapies that ablate LSCs may increase the chance of eliminating this cancer in patients. To this end, we used a bioreducible lipidoid-encapsulated Cas9/single guide RNA (sgRNA) ribonucleoprotein [lipidoid nanoparticle (LNP)–Cas9 RNP] to target the critical gene interleukin-1 receptor accessory protein (IL1RAP) in human LSCs. To enhance LSC targeting, we loaded LNP-Cas9 RNP and the chemokine CXCL12α onto mesenchymal stem cell membrane–coated nanofibril (MSCM-NF) scaffolds mimicking the bone marrow microenvironment. In vitro, CXCL12α release induced migration of LSCs to the scaffolds, and LNP-Cas9 RNP induced efficient gene editing. IL1RAP knockout reduced LSC colony-forming capacity and leukemic burden. Scaffold-based delivery increased the retention time of LNP-Cas9 in the bone marrow cavity. Overall, sustained local delivery of Cas9/IL1RAP sgRNA via CXCL12α-loaded LNP/MSCM-NF scaffolds provides an effective strategy for attenuating LSC growth to improve AML therapy.

In situ cancer vaccination by lipidoid nanoparticles enhances antigen cross-presentation and STING activation.