Analytical characterization of full, intermediate, and empty AAV capsids

Area of Science:

• Biotechnology
• Gene Therapy
• Viral Vector Manufacturing

Background:

• Recombinant adeno-associated virus (AAV) vector production yields full, intermediate, and empty capsids.
• Intermediate and empty capsids are impurities that may affect AAV product safety and efficacy.
• These impurities can reduce efficacy by competing for cellular transduction and increase safety concerns due to higher capsid load.

Purpose of the Study:

• To investigate the impact of intermediate capsids on the potency of AAV products.
• To determine if intermediate capsids contribute to the biological activity and efficacy of AAV vectors.

Main Methods:

• Separation of an AAV preparation into fractions enriched for full, intermediate, or empty capsids.
• Utilized a comprehensive panel of in vitro assays (infectivity, gene expression, biological activity).
• Employed in vivo potency assays to assess AAV product performance based on capsid content.

Main Results:

• Intermediate capsids were found to contribute to the vector genome titer.
• Intermediate capsids demonstrated infectivity comparable to full capsids.
• Crucially, intermediate capsids did not contribute to the overall potency of the AAV product.

Conclusions:

• Intermediate capsids, despite being infectious, do not enhance AAV product potency.
• Reducing and controlling intermediate capsid levels is essential for manufacturing efficacious AAV gene therapy products.
• This study highlights the importance of capsid purity for optimal AAV vector performance.