Development and Optimization of a Hydrophobic Interaction Chromatography-Based Method of AAV Harvest, Capture, and Recovery.

ABSTRACT: With many ongoing clinical trials utilizing adeno-associated virus (AAV) gene therapy, it is necessary to find scalable and serotype-independent primary capture and recovery methods to allow for efficient and robust manufacturing processes. Here, we demonstrate the ability of a hydrophobic interaction chromatography membrane to capture and recover AAV1, AAV5, AAV8, and AAV "Mutant C" (a novel serotype incorporating elements of AAV3B and AAV8) particles from cell culture media and cell lysate with recoveries of 76%-100% of loaded material, depending on serotype. A simple, novel technique that integrates release and recovery of cell-associated AAV capsids is demonstrated. We show that by the addition of lyotropic salts to AAV-containing cell suspensions, AAV is released at an equivalent efficiency to mechanical lysis. The addition of the lyotropic salt also promotes a phase separation, which allows physical removal of large amounts of DNA and insoluble cellular debris from the AAV-containing aqueous fraction. The AAV is then captured and eluted from a hydrophobic interaction chromatography membrane. This integrated lysis and primary capture and recovery technique facilitates substantial removal of host-cell DNA and host-cell protein impurities.

SUBMITTER: McNally DJ 

PROVIDER: S-EPMC7569186 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature