Liver-targeted AAV gene therapy vectors produced by a clinical scale manufacturing process result in high, continuous therapeutic levels of enzyme activity and effective substrate reduction in mouse model of Fabry disease

Fabry disease (FD), an X-linked lysosomal storage disease, is caused by mutations in the GLA gene encoding α-galactosidase A (α-GalA). FD is characterized by progressive systemic accumulation of the enzyme’s substrates, globotriaosylceramide (Gb3) and lyso-Gb3, leading to renal, cardiac and/or cerebrovascular disease and culminating in premature demise. FD is treated by enzyme replacement therapy (ERT), however the short enzyme half-life necessitates life-long biweekly infusions. A more effective and long-lasting treatment would benefit FD patients. An AAV-mediated liver-targeted gene therapy was evaluated in a mouse model (GLAKO) that lacks α-GalA activity and accumulates high levels of Gb3/lyso-Gb3 in plasma and tissues. This strategy employs an episomal AAV (serotype 2/6) vector encoding human GLA cDNA (hGLA) driven by a liver-specific promoter. One-time administration of increasing amounts of AAV-hGLA cDNA generated using a clinical scale manufacturing process resulted in supraphysiological expression of plasma α-GalA (over 300-fold of WT) by study day 15, was well tolerated, and was stable for 3 months post-injection. Dose-dependent increases in α-GalA activities were achieved in liver, heart and kidney with a corresponding reduction of Gb3/lyso-Gb3. This initial vector was compared to an improved cDNA vector in a 1-month study using two different AAV doses in wild type C57BL/6 mice. The improved cDNA vector produced on average 7-fold higher levels of plasma α-GalA activity at study day 28 than mice administered the same dose of the initial cDNA. The high levels of α-GalA activity seen in these studies, along with the concomitant marked reduction in the accumulated Gb3/lyso-Gb3 in key tissues of the GLAKO mouse model, provide “proof-of-concept” for AAV-mediated targeting of hepatocytes to express therapeutic levels of human α-GalA. The clinical scale manufacturing process developed for these studies will enable rapid development and production in 2019 of clinical-grade material featuring the improved cDNA construct.