High-efficiency purification of divergent AAV serotypes using AAVX affinity chromatography

Area of Science:

• Biotechnology
• Gene Therapy
• Viral Vector Production

Background:

• Adeno-associated viral vectors (AAV) are crucial for gene therapy, but their production faces scalability and cost challenges.
• Current purification methods often lack efficiency and require serotype-specific optimization.
• Developing robust purification techniques is essential for advancing AAV-based therapeutics.

Purpose of the Study:

• To evaluate the efficacy of the POROS CaptureSelect AAVX affinity resin for broad AAV serotype purification.
• To assess the scalability, cost-effectiveness, and reusability of the AAVX resin.
• To compare the potency of AAV vectors purified by AAVX resin versus traditional methods.

Main Methods:

• Affinity chromatography using POROS CaptureSelect AAVX resin.
• Purification of 15 divergent AAV serotypes.
• Regeneration and reusability testing of the AAVX resin.
• Comparison of AAVX purification with iodixanol ultracentrifugation.
• In vitro and in vivo potency assays.

Main Results:

• AAVX resin efficiently captured a panel of 15 diverse AAV serotypes.
• The resin demonstrated excellent reusability without loss of efficiency or contamination.
• AAVX-purified vectors showed comparable potency to those purified by ultracentrifugation, despite a higher fraction of empty capsids.
• Optimized protocol achieved 65%-80% overall efficiency across scales and serotypes.

Conclusions:

• POROS CaptureSelect AAVX affinity chromatography is a versatile and efficient method for purifying a wide range of AAV serotypes.
• This approach addresses key bottlenecks in AAV production, enabling scalable and cost-effective gene therapy manufacturing.
• AAVX resin represents a significant advancement for the commercialization of AAV-based gene therapies.