Single-cell analysis reveals effective siRNA delivery in brain tumors with microbubble-enhanced ultrasound and cationic nanoparticles.

RNA-based therapies offer unique advantages for treating brain tumors. However, tumor penetrance and uptake are hampered by RNA therapeutic size, charge, and need to be "packaged" in large carriers to improve bioavailability. Here, we have examined delivery of siRNA, packaged in 50-nm cationic lipid-polymer hybrid nanoparticles (LPHs:siRNA), combined with microbubble-enhanced focused ultrasound (MB-FUS) in pediatric and adult preclinical brain tumor models. Using single-cell image analysis, we show that MB-FUS in combination with LPHs:siRNA leads to more than 10-fold improvement in siRNA delivery into brain tumor microenvironments of the two models. MB-FUS delivery of Smoothened (SMO) targeting siRNAs reduces SMO protein production and markedly increases tumor cell death in the SMO-activated medulloblastoma model. Moreover, our analysis reveals that MB-FUS and nanoparticle properties can be optimized to maximize delivery in the brain tumor microenvironment, thereby serving as a platform for developing next-generation tunable delivery systems for RNA-based therapy in brain tumors.

Introduction

Purpose Drug delivery with BBB opening

Study Objective To evaluate whether microbubble-enhanced focused ultrasound (MB-FUS) delivery of siRNA-loaded 50-nm lipid–polymer hybrid nanoparticles enhances tumor uptake and induces oncogene-targeted cell death in preclinical brain tumor models.

Disease model Medulloblastoma (SMO-activated brain tumor models)

Cargo name and characteristics siRNA (oncogene-targeting, Smoothened/SMO-targeting) packaged in ~50 nm cationic lipid–polymer hybrid nanoparticles (LPHs)

Outcomes and Safety

Summary of Outcomes Microbubble-enhanced focused ultrasound (MB-FUS) combined with 40–50 nm cationic LPH:siRNA increased nanoparticle accumulation ~10–13-fold and siRNA uptake ~10-fold in brain tumors, produced ~5-fold reduction in SMO protein and induced a ~16-fold increase in tumor cell apoptosis in an SHH‑activated medulloblastoma model. The effective FUS approach used microbubble‑enhanced, MR‑guided or ultrasound‑guided FUS (sonication of tumor regions) with strong harmonic emissions (~22 dB) and <1% broadband emissions (safety window); no other FUS parameter variants were reported as tested.

Duration of biological effect 30 hours

Safety-related matter The authors report that their MB-FUS exposure settings produced strong harmonic emissions with <1% broadband emissions (within safety levels) and state delivery was achieved in immunocompetent mice "without significant adverse effects." They also show apoptosis was specific to SMO‑siRNA (non‑targeting LPH:Cy5‑siRNA did not increase apoptosis), note prior studies of similar lipid nanoparticles reported negligible toxicity, and recommend extended toxicologic analyses in future work.

Brain Region

Visualization unavailable

Ultrasound Parameters

Focal Characteristics Focal depth: None; Focal length: None; Aperture size: None

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