301. AAV Capsid Engineering to Improve Transduction in Retina and Brain

Gene therapy vectors based on adeno-associated virus (AAV) are currently in clinical studies for numerous disease indications including Leber's congenital amaurosis, age-related macular degeneration, hemophilia, muscular dystrophy and Parkinson's disease. AAV vectors hold considerable promise as therapeutic agents; however there is potential to further improve the efficiency of AAV gene delivery and efficacy by making modifications to the AAV capsid. The AAV capsid can be engineered to incorporate mutations that alter its transduction activity, tropism, biodistribution and immunogenicity. We have constructed variant AAV vectors harboring a variety of capsid modifications including those that negate receptor binding and have tested these vectors in several tissues including the eye and brain. One variant, AAV2HBKO, is an AAV2 based vector containing mutations of critical amino acids known to be required for binding to its receptor, heparin sulfate proteoglycan. Interestingly, an AAV2HBKO vector delivering a secreted transgene, sFLT02, unexpectedly resulted in a 2-log increase in transduction compared to parental AAV2 when delivered subretinally to the mouse eye. Subretinal delivery of an AAV2HBKO vector expressing EGFP demonstrated that these capsid modifications resulted in an increase in photoreceptor transduction compared to the unmodified AAV2 vector. In contrast, the AAV2HBKO vector demonstrated a lack of transduction activity following intravitreal delivery to the mouse eye. In addition, we evaluated the transduction and tropism of AAV2HBKO in the mouse brain. In a head to head comparison with AAV2, the AAV2HBKO vector facilitated widespread striatal and cortical expression following an intrastriatal injection while AAV2-mediated expression was restricted to the site of injection. Similar to AAV2, the tropism of AAV2HBKO was primarily neuronal with little to no transduction of astrocytes or microglia. Biodistribution data suggests that this vector, when delivered systemically in the mouse, has significantly reduced liver transduction but a higher propensity to be delivered to skeletal muscle and heart compared to the wild-type AAV2 vector. We will present data evaluating the transduction activity, tropism and biodistribution of the AAV2HBKO variant. These studies illustrate the potential for improving the efficiency of AAV gene transfer via targeted capsid engineering.