AAVone: A Cost-Effective, Single-Plasmid Solution for Efficient AAV Production with Reduced DNA Impurities

Area of Science:

• Biotechnology
• Gene Therapy Manufacturing

Background:

• Current GMP-grade adeno-associated virus (AAV) vector production relies on triple-plasmid transfection.
• This method faces limitations in efficiency, batch consistency, and cost due to complex transfection requirements.

Purpose of the Study:

• To develop an improved AAV production system.
• To enhance efficiency, consistency, and cost-effectiveness for GMP-grade AAV vector manufacturing.

Main Methods:

• Developed AAVone, an all-in-one single-plasmid system consolidating adenovirus helper genes, AAV packaging genes (rep and cap), and the vector transgene.
• Compared AAVone to traditional triple-plasmid systems in mammalian cell transfection.

Main Results:

• AAVone achieved a 2- to 4-fold increase in AAV vector yields.
• Demonstrated reduced batch-to-batch variation and simplified production by eliminating plasmid ratio adjustments.
• Generated AAV vectors with comparable in vitro and in vivo transduction efficiency, improved purity (reduced backbone sequences), and fewer non-functional genomes.

Conclusions:

• The AAVone platform offers a straightforward, cost-effective, and highly consistent method for GMP-grade AAV vector production.
• This single-plasmid system addresses key limitations of traditional multi-plasmid approaches.