695. An AAV8 Mutant with Better Transduction in Murine Muscle and Nasal Airway Than AAV8

Abstract

Adeno-associated virus (AAV)-based gene therapy is showing increasing promise, stimulated by encouraging results from clinical trials in recent years. To advance the platform, we have been exploring AAV capsid manipulation. By combining three mutations at hyper-variable regions (HVRs) I, IV, and VIII of the AAV8 capsid - mutations isolated from saturation mutagenesis followed by several rounds of in vivo selection - we created an AAV8 mutant, referred to as AAV8.Triple. In vitro, the AAV8.Triple mutant showed resistance to various antisera of monkey and human, as well as human IVIG (titers that are at levels 2 to 4 fold that of AAV8, with respect to human IVIG). All three mutations contributed to the observed resistance. In mice, the liver transduction efficiency of AAV8.Triple was reduced compared with AAV8, however its muscle transduction was higher than that of AAV8 by approximately 10 fold. In addition, AAV8.Triple demonstrated a higher heparin affinity than AAV8. Interestingly, reducing the positive charges of the HVR.IV mutation decreased the vector's heparin affinity while liver transduction was partially restored. Similar to the trend observed in muscle, intranasal administration of AAV8.Triple resulted in a transduction efficiency 2 to 3 fold greater than that of AAV8, which was further improved to levels approximately 10 fold greater than AAV8 by swapping the VP1 unique region of AAV8.Triple with that of another AAV serotype. Overall, our findings indicate that AAV capsid manipulation was able to generate an AAV8 mutant with improved transduction in both murine muscle and nasal airway compared with wild-type AAV8. These findings may be relevant to disease models where high-efficiency intramuscular or intranasal gene delivery and resistance to pre-existing neutralizing antibodies are desired.