Membrane chromatography for AAV full capsid enrichment: Process development to scale up.

Area of Science:

• Biotechnology and Bioprocessing
• Gene Therapy Manufacturing
• Chromatographic Separations

Background:

• Increasing demand for adeno-associated virus (AAV)-based gene therapies necessitates efficient manufacturing processes.
• A critical challenge in AAV production is the removal of empty capsids, which lack therapeutic genetic material.
• Anion exchange chromatography is a key technology for enriching full AAV capsids.

Purpose of the Study:

• To develop and optimize a scalable process for separating empty and full AAV capsids.
• To establish baseline separation using anion exchange membrane chromatography.
• To achieve high purity and recovery of full AAV capsids for gene therapy applications.

Main Methods:

• Utilized anion exchange membrane chromatography for AAV capsid separation.
• Employed response surface Design of Experiments (DoE) to optimize elution conditions (conductivity, elution step duration).
• Developed statistical models to define the optimal operating space for the purification process.
• Validated model predictions at small scale and scaled up the purification process.

Main Results:

• Achieved baseline separation of empty and full AAV capsids (serotypes 8 and 9).
• Developed statistical models predicting optimal conditions for purity and recovery.
• Demonstrated recovery and purity exceeding 70% under optimized conditions.
• Validated model predictions and showed consistent results at manufacturing scale.

Conclusions:

• Anion exchange membrane chromatography is an effective method for purifying full AAV capsids.
• The developed process, guided by statistical modeling, enables scalable and efficient AAV manufacturing.
• Optimized conditions ensure high purity and yield, critical for advancing gene therapy production.