Efficacy of Helper-dependent Adenovirus Vector-mediated Gene Therapy in Murine Glycogen Storage Disease Type Ia  Dwight D Koeberl, B Sun, A Bird, YT Chen, K Oka, L Chan  Molecular Therapy  Volume 15, Issue 7, Pages 1253-1258 (July 2007) DOI: 10.1038/sj.mt.6300188 Copyright © 2007 The American Society of Gene Therapy Terms and Conditions

Figure 1 Reversal of glycogen storage disease type Ia complications in affected genetic deficiency of glucose-6-phosphatase knockout (G6Pase-KO) mice following helper-dependent adenovirus (HDAd) vector administration. (a) Proportion of affected G6Pase-KO mice surviving following administration of the indicated number of HDAd vector particles: 5 × 1012/kg (5E+12; n = 13), 2 × 1012/kg (2E+12; n = 27), or no vector (n = 11). (b) Weights of affected mice, following administration of 2 × 1012/kg (2E+12; n = 10) or 5 × 1012/kg (5E+12; n = 5) vector particles/kg, and weights of wild-type mice (n = 5) are shown. w.o., week old. Molecular Therapy 2007 15, 1253-1258DOI: (10.1038/sj.mt.6300188) Copyright © 2007 The American Society of Gene Therapy Terms and Conditions

Figure 2 Restoration of normalcy of body habitus following helper-dependent adenovirus (HDAd) vector administration. Affected mice indicated (*), shown in comparison to unaffected age-matched mice. (a) Glucose-6-phosphatase knockout (G6Pase-KO) mice at 2 week old (w.o.). (b) G6Pase-KO mice at 2 w.o. with coin (U.S. quarter) for comparison of sizes. (c) One-year-old G6Pase-KO mice following HDAd vector administration (5 × 1012vector particles/kg). Molecular Therapy 2007 15, 1253-1258DOI: (10.1038/sj.mt.6300188) Copyright © 2007 The American Society of Gene Therapy Terms and Conditions

Figure 3 Biochemical correction during fasting in glucose-6-phosphatase (G6Pase) mice. Affected mice did not survive until 3 weeks old (w.o.), if the helper-dependent adenovirus (HDAd) vector was not administered. Affected mice (n = 4); affected mice following administration of the indicated number of vector particles: 2 × 1012/kg (2E+12; n = 6) or 5 × 1012/kg (5E+12; n = 4); and wild-type mice (n = 6) were evaluated. (a) Plasma glucose. (b) Plasma cholesterol. One mouse each died the low dose affected (2E+12) group and in the wild-type group at 6 months-old (n = 5 for each). P values were calculated with Kruskal–Wallis and Dunn's multiple comparison tests. (c) Plasma glucose at the latest available time point, shown for mice euthanized at 28 w.o. and for those that died early. Molecular Therapy 2007 15, 1253-1258DOI: (10.1038/sj.mt.6300188) Copyright © 2007 The American Society of Gene Therapy Terms and Conditions

Figure 4 Correction of glucose-6-phosphatase (G6Pase) deficiency in the livers of affected G6Pase knocknot (G6Pase-KO) mice following helper-dependent adenovirus (HDAd) vector administration. (a) G6Pase activity following administration of 2 × 1012/kg (2E+12) or 5 × 1012/kg (5E+12) vector particles at day 3 after injection (n = 3 in each group) and 26 weeks after injection (n = 4 in each group). G6Pase was quantitated as the degradation of glucose-6-phosphate. Mean ± SD values are shown. Wild-type (n = 4) and affected (n = 3) littermates were analyzed for comparison. (b) Glycogen content of mouse liver in (a). Glycogen was quantitated as the release of glucose. (c) Urinary lactate in affected G6Pase-KO mice, in comparison to wild-type mice, following administration of 2 × 1012/kg (2E+12; n = 4) vector particles, analyzed at 26 weeks after injection (P= 0.0002); and following administration of 5 × 1012/kg (5E+12) vector particles, analyzed at 3 days after injection (P= 0.003; n = 3), and at 26 weeks after injection (P = 0.04; n = 3). Molecular Therapy 2007 15, 1253-1258DOI: (10.1038/sj.mt.6300188) Copyright © 2007 The American Society of Gene Therapy Terms and Conditions

Figure 5 Histomorphological correction of glycogen accumulation in the livers of glucose-6-phosphatase knockout (G6Pase-KO) mice following helper-dependent adenovirus (HDAd) vector administration. Periodic acid-Schiff (PAS) staining for glycogen (a–c) and hematoxylin and eosin (H&E) staining (d–f) in an affected mouse following HDAd vector administration (5 × 1012 particles/kg) at 3 w.o (a and d) and 7 month-old (m.o.) (b and e), and in a 7 m.o. wild-type mouse (c and f). Representative images are shown (three mice were evaluated for each image shown). Original magnification ×200. Molecular Therapy 2007 15, 1253-1258DOI: (10.1038/sj.mt.6300188) Copyright © 2007 The American Society of Gene Therapy Terms and Conditions