In Vivo Zinc Finger Nuclease-mediated Targeted Integration of a Glucose-6-phosphatase Transgene Promotes Survival in Mice With Glycogen Storage Disease Type IA

Abstract

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). Furthermore, transgene integration has been confirmed by sequencing in the majority of the mice treated with both vectors. Targeted alleles were 4.6-fold more common in livers of mice with GSD Ia, as compared with normal littermates, at 8 months following vector administration (P < 0.02). This suggests a selective advantage for vector-transduced hepatocytes following ZFN-mediated integration of the G6Pase vector. A short-term experiment also showed that 3-month-old mice receiving the ZFN had significantly-improved biochemical correction, in comparison with mice that received the donor vector alone. These data suggest that the use of ZFNs to drive integration of G6Pase at a safe harbor locus might improve vector persistence and efficacy, and lower mortality in GSD Ia.

Long‐term complications of glycogen storage disease type Ia in the canine model treated with gene replacement therapy

“…It is likely that genome editing would increase the viability of current gene replacement techniques by allowing more cells to be corrected over time (Landau et al 2016). Currently transgene expression from AAV-G6Pase is gradually lost (Supplementary Fig.…”

Exploiting epigenetics for the treatment of inborn errors of metabolism

“…To date, CRISPR‐Cas9 has already been successfully used in various in vitro studies and in vivo animal models. In preclinical models for IEMs including OTC deficiency, HT1, mucopolysaccharidosis type II (MPS II; OMIM #309900), and GSD Ia, ZFNs and CRISPR‐Cas9 techniques have been used to correct the mutated gene or insert the correct cDNA. These techniques have progressed to clinical trials, for instance to treat lung cancer patients (OMIM #211980) by administration of their own T cells after these were isolated and genetically modified ex vivo .…”

Genome editing for inborn errors of metabolism: advancing towards the clinic

“…In addition to albumin, there is the potential for integration into other safe harbor loci. Targeting of the glucose-6-phosphatase ( G6PC ) cDNA into the Rosa26 locus using ZFNs led to increased survival in a mouse model of glycogen storage disease type Ia (GSD1a) [ 45 ]. A dual AAV system was employed, with one vector delivering ZFNs and a second the G6PC transgene.…”