Abstract
Recombinant adeno-associated viruses (AAVs) with precise genome editing and cell-virus interaction have become a promising delivery tool for gene therapy. A robust AAV purification process is crucial for ensuring therapeutic efficacy. The challenges of AAV purification process development encompass limited material availability during early-stage development, high cost-of-goods compared to traditional biologics, and short development timelines for the critical first-in-human stages. The key to overcoming these challenges is to leverage high throughput (HTP) methods. In this article, an integrated end-to-end HTP workflow is proposed, utilizing a resin tip as the purification module and incorporating an HTP analytical toolkit on one platform. Purification parameters, including binding capacity, resin selection, and buffer composition screening for AAV full/partial/empty capsids separation, are efficiently determined using a 25 μL resin tip and HTP analytical tools with only micro-volume sample requirements. The process parameters determined from the HTP workflow predict the trends of full capsid enrichment and partial capsid removal for the bench-scale purification. This HTP workflow is also applied for the assessment of the AAV quality attributes to accelerate early-stage cell line and cell culture development. Comparable AAV quality attributes are demonstrated to Robocolumn as the benchmark HTP purification method. By leveraging HTP analytical tools to instantly interpret the purification data, this integrated HTP workflow effectively accelerates AAV purification process development, with a 2% material volume requirement compared to the benchmark method, 96-well format screening, short turnaround time for analytical assays, and significant cost-of-goods savings for downstream process development.