Abstract
Naturally occurring adeno-associated virus (AAV) serotypes that bind to ligands such as AVB sepharose or heparin can be purified by affinity chromatography, which is a more efficient and scalable method than gradient ultracentrifugation. Wild-type AAV8 does not bind effectively to either of these molecules, which constitutes a barrier to using this vector when a high throughput design is required. Previously, AAV8 was engineered to contain a SPAKFA amino acid sequence to facilitate purification using AVB sepharose resin; however, in vivo studies were not conducted to examine whether these capsid mutations altered the transduction profile. To address this gap in knowledge, a mutant AAV8 capsid was engineered to bind to AVB sepharose and heparan sulfate (AAV8-AVB-HS), which efficiently bound to both affinity columns, resulting in elution yields of >80% of the total vector loaded compared to <5% for wild-type AAV8. However, in vivo comparison by intramuscular, intravenous, and intraperitoneal vector administration demonstrated a significant decrease in AAV8-AVB-HS transduction efficiency without alteration of the transduction profile. Therefore, although it is possible to engineer AAV capsids to bind various affinity ligands, the consequences associated with mutating surface exposed residues have the potential to negatively impact other vector characteristics including in vivo potency and production yield. This study demonstrates the importance of evaluating all aspects of vector performance when engineering AAV capsids.