Ultrasound-Mediated Blood-Brain Barrier Opening Improves Whole Brain Gene Delivery in Mice

Abstract

Gene therapy represents a powerful therapeutic tool to treat diseased tissues and provide a durable and effective correction. The central nervous system (CNS) is the target of many gene therapy protocols, but its high complexity makes it one of the most difficult organs to reach, in part due to the blood-brain barrier that protects it from external threats. Focused ultrasound (FUS) coupled with microbubbles appears as a technological breakthrough to deliver therapeutic agents into the CNS. While most studies focus on a specific targeted area of the brain, the present work proposes to permeabilize the entire brain for gene therapy in several pathologies. Our results show that, after i.v. administration and FUS sonication in a raster scan manner, a self-complementary AAV9-CMV-GFP vector strongly and safely infected the whole brain of mice. An increase in vector DNA (19.8 times), GFP mRNA (16.4 times), and GFP protein levels (17.4 times) was measured in whole brain extracts of FUS-treated GFP injected mice compared to non-FUS GFP injected mice. In addition to this increase in GFP levels, on average, a 7.3-fold increase of infected cells in the cortex, hippocampus, and striatum was observed. No side effects were detected in the brain of treated mice. The combining of FUS and AAV-based gene delivery represents a significant improvement in the treatment of neurological genetic diseases.

The engineered AAV2-HBKO promotes non-invasive gene delivery to large brain regions beyond ultrasound targeted sites

“…However, a previous study delivering AAV9 encoding GFP under a cytomegalovirus promoter to the entire brain using FUS reported no immune response 1 month after viral delivery. 42 Differences in mouse strains, AAVs, promoters, transgenes, cells, and brain regions transduced may influence the immune responses observed across studies. 23 , 42 With AAV2-HBKO, when the area of FUS delivery was increased to two MRIgFUS spots unilaterally in the striatum or bilaterally in the thalamus, GFAP was upregulated and microglia morphed toward an amoeboid phenotype.…”

“… 42 Differences in mouse strains, AAVs, promoters, transgenes, cells, and brain regions transduced may influence the immune responses observed across studies. 23 , 42 With AAV2-HBKO, when the area of FUS delivery was increased to two MRIgFUS spots unilaterally in the striatum or bilaterally in the thalamus, GFAP was upregulated and microglia morphed toward an amoeboid phenotype. In the thalamus, a minimal increase in MHCII expression and T cell infiltration 3 weeks after viral delivery was also observed.…”

Perfluorocarbon Nanodroplets as Potential Nanocarriers for Brain Delivery Assisted by Focused Ultrasound-Mediated Blood–Brain Barrier Disruption

“…To administer SonoVue ® MB by retro-orbital injections in mice, a 1 mL tuberculin syringe (PentaFerte, Ferrara, Italy) with a 26-gauge, 12.5 mm needle (SOL-M TM , Sol-Millenium, Shanghai, China) was used. Then, ultrasound transcranial sonication (FUS) was started immediately after MB injection and was applied above the right hemisphere with an estimated in situ peak negative acoustic pressure of 0.33 MPa (0.45 MPa in free water) considering a mean transmission loss through the mouse skull of 25.7% [ 34 ]. Concretely, the FUS sequence was composed of ultrasonic waves transmitted for 5.6 s at 1.5 MHz with DC of 72%.…”

Current Strategies to Enhance Delivery of Drugs across the Blood–Brain Barrier

“…In the brains of rats treated, no harmful consequences were seen. The use of FUS and AAV-based gene transfer to treat neurological genetic diseases is a significant step forward [ 93 ].…”