ACUVRA: Anion-Exchange Chromatography UV-Ratio Analysis-A QC-Friendly Method for Monitoring Adeno-Associated Virus...

Area of Science:

• Biotechnology
• Gene Therapy
• Analytical Chemistry

Background:

• Adeno-associated virus (AAV) vectors are crucial for gene therapy, with capsid genome content impacting safety and efficacy.
• Empty capsids are critical quality attributes (CQAs) requiring strict control during AAV-based product manufacturing.
• Traditional methods using anion-exchange chromatography for capsid separation need extensive optimization for different AAV types.

Purpose of the Study:

• To develop and validate a novel analytical method for quantifying empty and full AAV capsids.
• To provide a QC-friendly alternative that bypasses the need for physical separation of empty and full capsids.
• To establish a robust method for AAV vector characterization throughout production.

Main Methods:

• Anion-exchange chromatography combined with UV absorbance ratio (A260/A280) analysis.
• The ACUVRA (Anion-exchange Chromatography UV-Ratio Analysis) method leverages the DNA/protein absorbance ratio.
• Application demonstrated using recombinant AAV2 (rAAV2) process intermediates and drug substance.

Main Results:

• ACUVRA accurately determines the relative proportion of empty and full capsids from a single chromatographic peak.
• Method qualification and GMP validation confirmed its suitability for quality control laboratories.
• Demonstrated applicability across process development, characterization, and GMP release testing.

Conclusions:

• ACUVRA offers a streamlined and efficient approach for AAV capsid quantitation.
• The method is a fit-for-purpose tool for process development and a QC-friendly option for GMP release testing.
• ACUVRA supports the development and manufacturing of safe and potent AAV-based gene therapies.