Intracranial Delivery of Lipid Nanoparticles in the Murine Model

Abstract

INTRODUCTION: Lipid nanoparticles (LNPs) offer several benefits over viral vectors for delivery of gene editing technology, including lower antigenicity and higher biocompatibility, scalability, modifiability, and the ability to package more variable cargo. In addition, their lipophilic nature allows them to target difficult-to-transduce cell populations, making them ideal for brain delivery. In this study, we demonstrate that intracranial (IC) delivery of LNPs is an effective method for brain targeting. METHODS:ROSA26mtmg mice at postnatal day 0 were injected with LNPs containing Cre-recombinase mRNA (LNP-Cre) or saline directly into the intraventricular space; these mice express tdTomato (red) fluorescence at baseline, which is excised in the presence of Cre-recombinase to express eGFP (green) fluorescence. Mice received 100 ng of LNP-Cre (n = 4) or saline (n = 1) in 4 μL total volume bilaterally. The pups were monitored for 10 days and then euthanized for analysis. RESULTS: At harvest, all pups were viable with no gross neurologic abnormalities. Brains injected with LNP-Cre showed strong eGFP positivity on stereomicroscopy (Fig. 1A) compared with control (Fig. 1B) and eGFP+ cells lining the ventricular spaces on histology (Fig. 1C). Quantitative polymerase chain reaction showed an average 20-fold increase in eGFP mRNA expression in the cortex, 16-fold increase in the midbrain, and 3-fold increase in the cerebellum compared with control (Fig. 1D).Figure 1CONCLUSION: This study shows that IC LNP injection is a safe and effective technique for brain editing with 100% survival over 10 days. Further optimization is critical for use in preclinical gene editing studies for neurocognitive disorders where brain targeting is the primary focus.