Rationally designed nanoparticle delivery of Cas9 ribonucleoprotein for effective gene editing

Area of Science:

• Biotechnology
• Molecular Biology
• Gene Therapy

Background:

• CRISPR/Cas9 technology offers therapeutic potential for genetic diseases.
• Efficient and safe intracellular delivery of CRISPR/Cas9 components remains a significant challenge for clinical applications.

Purpose of the Study:

• To develop and evaluate a novel porous nanoparticle system, termed 'BALL', for the intracellular delivery of Cas9 ribonucleoprotein (RNP).
• To assess the efficacy and safety of BALL-mediated Cas9 RNP delivery for gene editing in vitro and in vivo.

Main Methods:

• Porous nanoparticles (BALL) were engineered for Cas9 RNP encapsulation and delivery.
• In vitro gene knockout efficiency was assessed using a green fluorescent protein (GFP) model.
• In vivo studies involved delivering Cas9 RNP targeting the myostatin (MSTN) gene via intramuscular injection in a mouse model.

Main Results:

• BALL nanoparticles demonstrated improved bioavailability and serum stability for Cas9 RNP delivery.
• Efficient gene knockout was achieved with approximately 40% indel efficiency in vitro and 20% in vivo.
• MSTN gene knockout led to increased muscle mass and improved motor function in vivo.

Conclusions:

• The BALL nanoparticle system provides a safe and effective platform for intracellular delivery of CRISPR/Cas9 RNP.
• This delivery system holds significant promise for advancing CRISPR-based genome editing therapies for various genetic disorders.