In Vivo Prime Editing by Lipid Nanoparticle Co-delivery of Chemically Modified pegRNA and Prime Editor mRNA

Area of Science:

• Gene editing technologies
• Molecular and cellular biology
• Biotechnology

Background:

• Prime editing is a powerful gene editing tool with significant therapeutic potential.
• In vivo delivery of prime editors faces challenges, particularly with adeno-associated virus (AAV) vectors.
• Lipid nanoparticles (LNPs) offer a promising non-viral alternative for in vivo gene editing delivery.

Purpose of the Study:

• To demonstrate successful co-delivery of prime editor mRNA and chemically modified pegRNA using LNPs for in vivo prime editing.
• To investigate the impact of pegRNA chemical modifications and dosing regimens on editing efficiency and toxicity.
• To evaluate the influence of LNP composition on in vivo prime editing performance.

Main Methods:

• Co-delivery of chemically modified pegRNA and prime editor mRNA encapsulated in LNPs.
• Evaluation of different dosing regimens (daily vs. weekly) in mouse models.
• Assessment of editing efficiency and liver toxicity.
• Testing LNP formulations with varying ionizable lipids and phospholipids.
• Prime editing efficacy assessment in NSG immunodeficient mice.

Main Results:

• Weekly LNP dosing was well-tolerated and increased editing efficacy 1.8-fold compared to daily dosing, which caused liver toxicity.
• LNP-delivered prime editing showed a 2.8-fold enhancement in NSG immunodeficient mice.
• The chemical modifications of pegRNA and the specific ionizable lipids and phospholipids in LNPs significantly impacted in vivo editing efficiency.

Conclusions:

• LNPs are a viable and advantageous platform for in vivo prime editor delivery, overcoming limitations of viral vectors.
• Optimized dosing strategies and LNP formulations are crucial for maximizing prime editing efficacy and safety.
• These findings support the advancement of LNP-based prime editing for potential therapeutic applications.