311. In Vivo Zinc Finger Nuclease-Mediated Targeted Integration of a Glucose-6-Phosphatase Transgene Enhances Biochemical Correction in Mice with Glycogen Storage Disease Type IA

Glycogen storage disease type Ia (GSD Ia) is caused by glucose-6-phosphatase (G6Pase) deficiency in association with severe, life-threatening hypoglycemia that necessitates lifelong dietary therapy. Here we show that use of a zinc-finger nuclease (ZFN) targeted to the ROSA26 safe harbor locus and a ROSA26-targeting vector containing a G6PC donor transgene, both delivered with adeno-associated virus (AAV) vectors, markedly improved survival of G6Pase knockout (G6Pase-KO) mice compared with mice receiving the donor vector alone (p<0.04) out to 8 months of age. Transgene integration has been confirmed by sequencing in the majority of the mice treated with both vectors. Surviving G6Pase-KO mice at 8 months of age had decreased glycogen content compared with young untreated G6Pase-KO controls, which correlated with the long-term survival of these mice that otherwise perish before weaning. A short-term experiment resolved difficulty observing biochemical differences between the dual-vector and single-vector groups, caused by a selective advantage for strongly-responding mice: 3-month-old mice receiving the ZFN had significantly reduced hepatic glycogen accumulation and improved G6Pase activity, compared with mice that received the donor vector alone. These data demonstrate that the use of ZFNs to drive integration of G6Pase at a safe harbor locus might improve transgene persistence and efficacy, and lower mortality in GSD Ia.