@article{brooks_landau_everitt_brown_grady_waskowicz_bass_d'angelo_asfaw_williams_et al._2018, title={Long-term complications of glycogen storage disease type Ia in the canine model treated with gene replacement therapy}, volume={41}, ISSN={["1573-2665"]}, DOI={10.1007/s10545-018-0223-y}, abstractNote={Background Glycogen storage disease type Ia (GSD Ia) in dogs closely resembles human GSD Ia. Untreated patients with GSD Ia develop complications associated with glucose-6-phosphatase (G6Pase) deficiency. Survival of human patients on intensive nutritional management has improved; however, long-term complications persist including renal failure, nephrolithiasis, hepatocellular adenomas (HCA), and a high risk for hepatocellular carcinoma (HCC). Affected dogs fail to thrive with dietary therapy alone. Treatment with gene replacement therapy using adeno-associated viral vectors (AAV) expressing G6Pase has greatly prolonged life and prevented hypoglycemia in affected dogs. However, long-term complications have not been described to date. Methods Five GSD Ia-affected dogs treated with AAV-G6Pase were evaluated. Dogs were euthanized due to reaching humane endpoints related to liver and/or kidney involvement, at 4 to 8 years of life. Necropsies were performed and tissues were analyzed. Results Four dogs had liver tumors consistent with HCA and HCC. Three dogs developed renal failure, but all dogs exhibited progressive kidney disease histologically. Urolithiasis was detected in two dogs; uroliths were composed of calcium oxalate and calcium phosphate. One affected and one carrier dog had polycystic ovarian disease. Bone mineral density was not significantly affected. Conclusions Here, we show that the canine GSD Ia model demonstrates similar long-term complications as GSD Ia patients in spite of gene replacement therapy. Further development of gene therapy is needed to develop a more effective treatment to prevent long-term complications of GSD Ia.}, number={6}, journal={JOURNAL OF INHERITED METABOLIC DISEASE}, author={Brooks, Elizabeth D. and Landau, Dustin J. and Everitt, Jeffrey I. and Brown, Talmage T. and Grady, Kylie M. and Waskowicz, Lauren and Bass, Cameron R. and D'Angelo, John and Asfaw, Yohannes G. and Williams, Kyha and et al.}, year={2018}, month={Dec}, pages={965–976} } @article{demaster_luo_curtis_williams_landau_drake_kozink_bird_crane_sun_et al._2012, title={Long-Term Efficacy Following Readministration of an Adeno-Associated Virus Vector in Dogs with Glycogen Storage Disease Type Ia}, volume={23}, ISSN={["1557-7422"]}, DOI={10.1089/hum.2011.106}, abstractNote={Glycogen storage disease type Ia (GSD-Ia) is the inherited deficiency of glucose-6-phosphatase (G6Pase), primarily found in liver and kidney, which causes life-threatening hypoglycemia. Dogs with GSD-Ia were treated with double-stranded adeno-associated virus (AAV) vectors encoding human G6Pase. Administration of an AAV9 pseudotyped (AAV2/9) vector to seven consecutive GSD-Ia neonates prevented hypoglycemia during fasting for up to 8 hr; however, efficacy eventually waned between 2 and 30 months of age, and readministration of a new pseudotype was eventually required to maintain control of hypoglycemia. Three of these dogs succumbed to acute hypoglycemia between 7 and 9 weeks of age; however, this demise could have been prevented by earlier readministration an AAV vector, as demonstrated by successful prevention of mortality of three dogs treated earlier in life. Over the course of this study, six out of nine dogs survived after readministration of an AAV vector. Of these, each dog required readministration on average every 9 months. However, two were not retreated until >34 months of age, while one with preexisting antibodies was re-treated three times in 10 months. Glycogen content was normalized in the liver following vector administration, and G6Pase activity was increased in the liver of vector-treated dogs in comparison with GSD-Ia dogs that received only with dietary treatment. G6Pase activity reached approximately 40% of normal in two female dogs following AAV2/9 vector administration. Elevated aspartate transaminase in absence of inflammation indicated that hepatocellular turnover in the liver might drive the loss of vector genomes. Survival was prolonged for up to 60 months in dogs treated by readministration, and all dogs treated by readministration continue to thrive despite the demonstrated risk for recurrent hypoglycemia and mortality from waning efficacy of the AAV2/9 vector. These preclinical data support the further translation of AAV vector–mediated gene therapy in GSD-Ia. Demaster and colleagues report preclinical results of treating dogs with glycogen storage disease type Ia (GSD-Ia) with double-stranded adeno-associated viral vector type 9 (AAV2/9) encoding human glucose-6-phosphatase (G6Pase). Vector treatment was able to prevent hypoglycemia and led to normalized liver glycogen levels and increased G6Pase activity, but carefully timed readministration with a different pseudotype was required to prevent the therapeutic effect from waning. Survival was prolonged for up to 60 months in dogs treated by readministration, with all dogs continuing to thrive despite the demonstrated risk for recurrent hypoglycemia and mortality from waning vector efficacy.}, number={4}, journal={HUMAN GENE THERAPY}, author={Demaster, Amanda and Luo, Xiaoyan and Curtis, Sarah and Williams, Kyha D. and Landau, Dustin J. and Drake, Elizabeth J. and Kozink, Daniel M. and Bird, Andrew and Crane, Bayley and Sun, Francis and et al.}, year={2012}, month={Apr}, pages={407–418} } @article{crane_luo_demaster_williams_kozink_zhang_brown_pinto_oka_sun_et al._2012, title={Rescue administration of a helper-dependent adenovirus vector with long-term efficacy in dogs with glycogen storage disease type Ia}, volume={19}, ISSN={["0969-7128"]}, DOI={10.1038/gt.2011.86}, abstractNote={Glycogen storage disease type Ia (GSD-Ia) stems from glucose-6-phosphatase (G6Pase) deficiency and causes hypoglycemia, hepatomegaly, hypercholesterolemia and lactic acidemia. Three dogs with GSD-Ia were initially treated with a helper-dependent adenovirus encoding a human G6Pase transgene (HDAd-cG6Pase serotype 5) on postnatal day 3. Unlike untreated dogs with GSD-Ia, all three dogs initially maintained normal blood glucose levels. After 6–22 months, vector-treated dogs developed hypoglycemia, anorexia and lethargy, suggesting that the HDAd-cG6Pase serotype 5 vector had lost efficacy. Liver biopsies collected at this time revealed significantly elevated hepatic G6Pase activity and reduced glycogen content, when compared with affected dogs treated only by frequent feeding. Subsequently, the HDAd-cG6Pase serotype 2 vector was administered to two dogs, and hypoglycemia was reversed; however, renal dysfunction and recurrent hypoglycemia complicated their management. Administration of a serotype 2 HDAd vector prolonged survival in one GSD-Ia dog to 12 months of age and 36 months of age in the other, but the persistence of long-term complications limited HDAd vectors in the canine model for GSD-Ia.}, number={4}, journal={GENE THERAPY}, author={Crane, B. and Luo, X. and Demaster, A. and Williams, K. D. and Kozink, D. M. and Zhang, P. and Brown, T. T. and Pinto, C. R. and Oka, K. and Sun, F. and et al.}, year={2012}, month={Apr}, pages={443–452} } @article{fry_brown_lloyd_hansen_legleiter_robarge_spears_2011, title={Effect of dietary boron on physiological responses in growing steers inoculated with bovine herpesvirus type-1}, volume={90}, ISSN={["1532-2661"]}, DOI={10.1016/j.rvsc.2010.04.016}, abstractNote={Thirty-six Angus and Angus × Simmental steers were fed one of three dietary treatments; (1) control (no supplemental B), (2) 5 mg supplemental B/kg, and (3) 15 mg supplemental B/kg for 47 days to determine the effects of dietary boron (B) on disease resistance following an inoculation with bovine herpesvirus type-1 (BHV-1). On day 34 of the study steers were inoculated intranasally with BHV-1. Rectal temperatures began to elevate at day 2, and plasma tumor necrosis factor-α concentrations increased (P < 0.05) by day 2 following BHV-1 inoculation. Plasma acute phase proteins were increased (P < 0.01) while plasma interferon-γ was decreased (P < 0.05) by day 4 post-inoculation. Supplementation of B increased (P < 0.001) plasma B concentrations in a dose-responsive manner. However, dietary B did not affect the duration and severity of clinical signs of BHV-1 and had minimal effects on plasma acute phase proteins and cytokines.}, number={1}, journal={RESEARCH IN VETERINARY SCIENCE}, author={Fry, R. S. and Brown, T. T., Jr. and Lloyd, K. E. and Hansen, S. L. and Legleiter, L. R. and Robarge, W. P. and Spears, J. W.}, year={2011}, month={Feb}, pages={78–83} } @article{luo_hall_li_bird_lavin_winn_kemper_brown_koeberl_2011, title={Hepatorenal Correction in Murine Glycogen Storage Disease Type I With a Double-stranded Adeno-associated Virus Vector}, volume={19}, ISSN={["1525-0016"]}, DOI={10.1038/mt.2011.126}, abstractNote={Glycogen storage disease type Ia (GSD-Ia) is caused by the deficiency of glucose-6-phosphatase (G6Pase). Long-term complications of GSD-Ia include life-threatening hypoglycemia and proteinuria progressing to renal failure. A double-stranded (ds) adeno-associated virus serotype 2 (AAV2) vector encoding human G6Pase was pseudotyped with four serotypes, AAV2, AAV7, AAV8, and AAV9, and we evaluated efficacy in 12-day-old G6pase (−/−) mice. Hypoglycemia during fasting (plasma glucose <100 mg/dl) was prevented for >6 months by the dsAAV2/7, dsAAV2/8, and dsAAV2/9 vectors. Prolonged fasting for 8 hours revealed normalization of blood glucose following dsAAV2/9 vector administration at the higher dose. The glycogen content of kidney was reduced by >65% with both the dsAAV2/7 and dsAAV2/9 vectors, and renal glycogen content was stably reduced between 7 and 12 months of age for the dsAAV2/9 vector-treated mice. Every vector-treated group had significantly reduced glycogen content in the liver, in comparison with untreated G6pase (−/−) mice. G6Pase was expressed in many renal epithelial cells of with the dsAAV2/9 vector for up to 12 months. Albuminuria and renal fibrosis were reduced by the dsAAV2/9 vector. Hepatorenal correction in G6pase (−/−) mice demonstrates the potential of AAV vectors for the correction of inherited diseases of metabolism. Glycogen storage disease type Ia (GSD-Ia) is caused by the deficiency of glucose-6-phosphatase (G6Pase). Long-term complications of GSD-Ia include life-threatening hypoglycemia and proteinuria progressing to renal failure. A double-stranded (ds) adeno-associated virus serotype 2 (AAV2) vector encoding human G6Pase was pseudotyped with four serotypes, AAV2, AAV7, AAV8, and AAV9, and we evaluated efficacy in 12-day-old G6pase (−/−) mice. Hypoglycemia during fasting (plasma glucose <100 mg/dl) was prevented for >6 months by the dsAAV2/7, dsAAV2/8, and dsAAV2/9 vectors. Prolonged fasting for 8 hours revealed normalization of blood glucose following dsAAV2/9 vector administration at the higher dose. The glycogen content of kidney was reduced by >65% with both the dsAAV2/7 and dsAAV2/9 vectors, and renal glycogen content was stably reduced between 7 and 12 months of age for the dsAAV2/9 vector-treated mice. Every vector-treated group had significantly reduced glycogen content in the liver, in comparison with untreated G6pase (−/−) mice. G6Pase was expressed in many renal epithelial cells of with the dsAAV2/9 vector for up to 12 months. Albuminuria and renal fibrosis were reduced by the dsAAV2/9 vector. Hepatorenal correction in G6pase (−/−) mice demonstrates the potential of AAV vectors for the correction of inherited diseases of metabolism.}, number={11}, journal={MOLECULAR THERAPY}, author={Luo, Xiaoyan and Hall, Gentzon and Li, Songtao and Bird, Andrew and Lavin, Peter J. and Winn, Michelle P. and Kemper, Alex R. and Brown, Talmage T. and Koeberl, Dwight D.}, year={2011}, month={Nov}, pages={1961–1970} } @article{koeberl_pinto_brown_chen_2009, title={Gene Therapy for Inherited Metabolic Disorders in Companion Animals}, volume={50}, ISSN={["1084-2020"]}, DOI={10.1093/ilar.50.2.122}, abstractNote={Scientists first described inborn errors of metabolism, also termed inherited disorders of metabolism, early in the 20th century and since then have determined the biochemical and genetic bases of a great number of these disorders both in humans and in an increasing number of companion animals. The availability of metabolic screening tests has advanced the biochemical and genetic characterization in affected breeds of companion animals of inherited metabolic disorders involving amino acid, carbohydrate, fatty acid, and metal metabolism. Advances in gene therapy have led to the development of new treatments for inherited disorders of metabolism, and animal models have played a critical role in this research. For example, glycogen storage disease type Ia in dogs was highly responsive to adeno-associated viral vector–mediated gene therapy, which prolonged survival and for more than a year prevented hypoglycemia during fasting. Gene therapy for other glycogen storage diseases and metabolic disorders will also be feasible. The establishment of a breeding colony and the ability to sustain affected animals are critical steps toward evaluating the safety and efficacy of gene therapy with clinically relevant endpoints. The further development of gene therapy for inherited disorders of metabolism could lead to curative therapy for affected humans and animals alike.}, number={2}, journal={ILAR JOURNAL}, author={Koeberl, Dwight D. and Pinto, Carlos and Brown, Talmage and Chen, Y. T.}, year={2009}, pages={122–127} } @article{hill_hopkins_davidson_bolt_diaz_brownie_brown_huntington_whitlow_2009, title={The addition of cottonseed hulls to the starter and supplementation of live yeast or mannanoligosaccharide in the milk for young calves}, volume={92}, ISSN={["1525-3198"]}, DOI={10.3168/jds.2008-1320}, abstractNote={The objectives of this study were to investigate the effects of the addition of cottonseed hulls (CSH) to the starter and the supplementation of live yeast product (YST) or mannanoligosaccharide product (MOS) to milk, on growth, intake, rumen development, and health parameters in young calves. Holstein (n = 116) and Jersey (n = 46) bull (n = 74) and heifer (n = 88) calves were assigned randomly within sex at birth to treatments. All calves were fed 3.8 L of colostrum daily for the first 2 d. Holstein calves were fed 3.8 L of whole milk, and Jersey calves were fed 2.8 L of whole milk through weaning at 42 d. Calves continued on trial through 63 d. Six treatments were arranged as a 2 × 3 factorial. Calves received either a corn-soybean meal-based starter (21% crude protein and 6% acid detergent fiber; −CSH) or a blend of 85% corn-soybean meal-based starter and 15% CSH (18% crude protein and 14% acid detergent fiber; +CSH) ad libitum. In addition, calves received whole milk with either no supplement (NONE) or supplemented with 3 g/d of mannanoligosaccharide product (MOS) or 4 g/d of live yeast product (YST) through weaning at 42 d. Twelve Holstein steers [n = 6 (per starter type); n = 4 (per supplement type)] were euthanized for collection and examination of rumen tissue samples. Dry matter intake (DMI) was greater for Holstein calves fed +CSH (0.90 kg/d) than −CSH (0.76 kg/d). Final body weight at 63 d of Holstein calves fed +CSH (75.8 kg) was greater than that of those fed −CSH (71.0 kg). Average daily gain (ADG) was greater for Holstein calves fed +CSH (0.58 kg/d) than −CSH (0.52 kg/d). However, Holstein calves fed −CSH had a greater feed efficiency (FE; 0.71 kg of ADG/kg of DMI) than those fed +CSH (0.65 kg of ADG/kg of DMI). Also, Holstein calves fed +CSH had narrower rumen papillae (0.32 mm) compared with those fed −CSH (0.41 mm). There were no significant effects of CSH on DMI, ADG, or FE in Jersey calves. There were no significant effects of YST or MOS on DMI, ADG, FE, or rumen papillae measures in Holstein calves. Jersey calves fed YST or MOS had greater final body weight at 63 d (51.2 kg and 51.0 kg, respectively) than calves fed NONE (47.5 kg). However, there were no significant effects of YST or MOS on DMI, ADG, or FE in Jersey calves.}, number={2}, journal={JOURNAL OF DAIRY SCIENCE}, author={Hill, S. R. and Hopkins, B. A. and Davidson, S. and Bolt, S. M. and Diaz, D. E. and Brownie, C. and Brown, T. and Huntington, G. B. and Whitlow, L. W.}, year={2009}, month={Feb}, pages={790–798} } @article{koeberl_pinto_sun_li_kozink_benjamin_demaster_kruse_vaughn_hillman_et al._2008, title={AAV vector-mediated reversal of hypoglycemia in canine and murine glycogen storage disease type Ia}, volume={16}, ISSN={["1525-0016"]}, DOI={10.1038/mt.2008.15}, abstractNote={Glycogen storage disease type Ia (GSD-Ia) profoundly impairs glucose release by the liver due to glucose-6-phosphatase (G6Pase) deficiency. An adeno-associated virus (AAV) containing a small human G6Pase transgene was pseudotyped with AAV8 (AAV2/8) to optimize liver tropism. Survival was prolonged in 2-week-old G6Pase (-/-) mice by 600-fold fewer AAV2/8 vector particles (vp), in comparison to previous experiments involving this model (2 x 10(9) vp; 3 x 10(11) vp/kg). When the vector was pseudotyped with AAV1, survival was prolonged only at a higher dose (3 x 10(13) vp/kg). The AAV2/8 vector uniquely prevented hypoglycemia during fasting and fully corrected liver G6Pase deficiency in GSD-Ia mice and dogs. The AAV2/8 vector has prolonged survival in three GSD-Ia dogs to >11 months, which validated this strategy in the large animal model for GSD-Ia. Urinary biomarkers, including lactate and 3-hydroxybutyrate, were corrected by G6Pase expression solely in the liver. Glycogen accumulation in the liver was reduced almost to the normal level in vector-treated GSD-Ia mice and dogs, as was the hepatocyte growth factor (HGF) in GSD-Ia mice. These preclinical data demonstrated the efficacy of correcting hepatic G6Pase deficiency, and support the further preclinical development of AAV vector-mediated gene therapy for GSD-Ia.}, number={4}, journal={MOLECULAR THERAPY}, author={Koeberl, Dwight D. and Pinto, Carlos and Sun, Baodong and Li, Songtao and Kozink, Daniel M. and Benjamin, Daniel K., Jr. and Demaster, Amanda K. and Kruse, Meghan A. and Vaughn, Valerie and Hillman, Steven and et al.}, year={2008}, month={Apr}, pages={665–672} } @article{sun_young_li_di_brown_salva_li_bird_yan_auten_et al._2008, title={Correction of multiple striated muscles in murine Pompe diseasethrough adeno-associated virus-mediated gene therapy}, volume={16}, ISSN={["1525-0016"]}, DOI={10.1038/mt.2008.133}, abstractNote={Glycogen storage disease type II (Pompe disease; MIM 232300) stems from the deficiency of acid α-glucosidase (GAA; acid maltase; EC 3.2.1.20), which primarily involves cardiac and skeletal muscles. An adeno-associated virus 2/8 (AAV2/8) vector containing the muscle creatine kinase (MCK) (CK1) reduced glycogen content by ∼50% in the heart and quadriceps in GAA-knockout (GAA-KO) mice; furthermore, an AAV2/8 vector containing the hybrid α-myosin heavy chain enhancer-/MCK enhancer-promoter (MHCK7) cassette reduced glycogen content by >95% in heart and >75% in the diaphragm and quadriceps. Transduction with an AAV2/8 vector was higher in the quadriceps than in the gastrocnemius. An AAV2/9 vector containing the MHCK7 cassette corrected GAA deficiency in the distal hindlimb, and glycogen accumulations were substantially cleared by human GAA (hGAA) expression therein; however, the analogous AAV2/7 vector achieved much lower efficacy. Administration of the MHCK7-containing vectors significantly increased striated muscle function as assessed by increased Rotarod times at 18 weeks after injection, whereas the CK1-containing vector did not increase Rotarod performance. Importantly, type IIb myofibers in the extensor digitalis longus (EDL) were transduced, thereby correcting a myofiber type that is unresponsive to enzyme replacement therapy. In summary, AAV8 and AAV9-pseudotyped vectors containing the MHCK7 regulatory cassette achieved enhanced efficacy in Pompe disease mice. Glycogen storage disease type II (Pompe disease; MIM 232300) stems from the deficiency of acid α-glucosidase (GAA; acid maltase; EC 3.2.1.20), which primarily involves cardiac and skeletal muscles. An adeno-associated virus 2/8 (AAV2/8) vector containing the muscle creatine kinase (MCK) (CK1) reduced glycogen content by ∼50% in the heart and quadriceps in GAA-knockout (GAA-KO) mice; furthermore, an AAV2/8 vector containing the hybrid α-myosin heavy chain enhancer-/MCK enhancer-promoter (MHCK7) cassette reduced glycogen content by >95% in heart and >75% in the diaphragm and quadriceps. Transduction with an AAV2/8 vector was higher in the quadriceps than in the gastrocnemius. An AAV2/9 vector containing the MHCK7 cassette corrected GAA deficiency in the distal hindlimb, and glycogen accumulations were substantially cleared by human GAA (hGAA) expression therein; however, the analogous AAV2/7 vector achieved much lower efficacy. Administration of the MHCK7-containing vectors significantly increased striated muscle function as assessed by increased Rotarod times at 18 weeks after injection, whereas the CK1-containing vector did not increase Rotarod performance. Importantly, type IIb myofibers in the extensor digitalis longus (EDL) were transduced, thereby correcting a myofiber type that is unresponsive to enzyme replacement therapy. In summary, AAV8 and AAV9-pseudotyped vectors containing the MHCK7 regulatory cassette achieved enhanced efficacy in Pompe disease mice. Glycogen storage disease type II (Pompe disease; MIM 232300) is a classical lysosomal storage disease that causes death in infancy as a result of cardiomyopathy and cardiorespiratory failure. The single gene deficiency in acid α-glucosidase (GAA; acid maltase; EC 3.2.1.20) results in lysosomal accumulation of glycogen in various tissues, primarily in heart and skeletal muscle. Pompe disease could be effectively treated by the correction of GAA deficiency in striated muscle. GAA expression with pseudotyped adeno-associated virus (AAV) vectors1Sun B Zhang H Franco LM Young SP Schneider A Bird A et al.Efficacy of an adeno-associated virus 8-pseudotyped vector in glycogen storage disease type II.Mol Ther. 2005; 11: 57-65Abstract Full Text Full Text PDF PubMed Scopus (121) Google Scholar,2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar,3Franco LM Sun B Yang X Bird A Zhang H Schneider A et al.Evasion of immune responses to introduced human acid alpha-glucosidase by liver-restricted expression in glycogen storage disease type II.Mol Ther. 2005; 12: 876-884Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar,4Mah C Cresawn KO Fraites TJ Jr Pacak CA Lewis MA Zolotukhin I et al.Sustained correction of glycogen storage disease type II using adeno-associated virus serotype 1 vectors.Gene Ther. 2005; 12: 1405-1409Crossref PubMed Scopus (60) Google Scholar,5Cresawn KO Fraites TJ Wasserfall C Atkinson M Lewis M Porvasnik S et al.Impact of humoral immune response on distribution and efficacy of recombinant adeno-associated virus-derived acid alpha-glucosidase in a model of glycogen storage disease type II.Hum Gene Ther. 2005; 16: 68-80Crossref PubMed Scopus (56) Google Scholar or with a helper-dependent adenovirus vector has achieved prolonged efficacy6Kiang A Hartman ZC Liao SX Xu F Serra D Palmer DJ et al.Fully deleted adenovirus persistently expressing GAA accomplishes long-term skeletal muscle glycogen correction in tolerant and nontolerant GSD-II mice.Mol Ther. 2006; 13: 127-134Abstract Full Text Full Text PDF PubMed Scopus (37) Google Scholar in Pompe disease mice. None of the aforementioned vectors completely corrected the glycogen content of all skeletal muscles, and none of these studies evaluated the distal hindlimb muscles. The clinical presentation of Pompe disease resembles that of the muscular dystrophies, featuring weakness of the proximal leg muscles that generalizes to all skeletal muscles.7Hirschhorn R Reuser AJJ Scriver CR Beaudet AL Sly WS Valle D Glycogen storage disease type II: acid α-glucosidase (acid maltase) deficiency.The Metabolic and Molecular Basis for Inherited Disease. McGraw-Hill, New York2001: 3389-3420Google Scholar Gene therapy in the muscular dystrophies represents a unique challenge, because humoral and cytotoxic immune responses occur frequently in response to introduced proteins.8Ferrer A Wells KE Wells DJ Immune responses to dystrophin: implications for gene therapy of Duchenne muscular dystrophy.Gene Ther. 2000; 7: 1439-1446Crossref PubMed Scopus (84) Google Scholar,9Cordier L Gao GP Hack AA McNally EM Wilson JM Chirmule N et al.Muscle-specific promoters may be necessary for adeno-associated virus-mediated gene transfer in the treatment of muscular dystrophies.Hum Gene Ther. 2001; 12: 205-215Crossref PubMed Scopus (129) Google Scholar Cytotoxic T lymphocyte responses against gene therapy vectors have been reduced by substituting muscle-specific regulatory cassettes for ubiquitously active viral promoter/enhancers. An AAV2 vector containing a muscle-specific creatine kinase (MCK) regulatory cassette evoked an attenuated immune response in mdx mice, in comparison with an analogous AAV vector containing the cytomegalovirus (CMV) promoter.10Yuasa K Sakamoto M Miyagoe-Suzuki Y Tanouchi A Yamamoto H Li J et al.Adeno-associated virus vector-mediated gene transfer into dystrophin-deficient skeletal muscles evokes enhanced immune response against the transgene product.Gene Ther. 2002; 9: 1576-1588Crossref PubMed Scopus (113) Google Scholar An AAV2/6 vector containing the MCK CK6 cassette11Hauser MA Robinson A Hartigan-O'Connor D Williams-Gregory D Buskin JN Apone S et al.Analysis of muscle creatine kinase regulatory elements in recombinant adenoviral vectors.Mol Ther. 2000; 2: 16-25Abstract Full Text Full Text PDF PubMed Scopus (80) Google Scholar transduced skeletal muscle with lower efficiency, in comparison with an analogous vector containing a CMV promoter/enhancer; however, the MCK-driven β-galactosidase expression persisted longer than CMV-driven expression.12Gregorevic P Blankinship MJ Allen JM Crawford RW Meuse L Miller DG et al.Systemic delivery of genes to striated muscles using adeno-associated viral vectors.Nat Med. 2004; 10: 828-834Crossref PubMed Scopus (544) Google Scholar An AAV2/6 vector containing another MCK regulatory cassette [CK1 (refs. 13Shield MA Haugen HS Clegg CH Hauschka SD E-box sites and a proximal regulatory region of the muscle creatine kinase gene differentially regulate expression in diverse skeletal muscles and cardiac muscle of transgenic mice.Mol Cell Biol. 1996; 16: 5058-5068Crossref PubMed Scopus (99) Google Scholar,14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar)] produced high-level human GAA (hGAA) expression and glycogen clearance in the injected gastrocnemius muscle, and the analogous AAV2/7 vector partially cleared glycogen storage in multiple muscle groups of GAA-knockout (GAA-KO) mice following intravenous administration.2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar AAV2/8 vectors have efficiently transduced striated muscle following systemic delivery in mice.15Nakai H Fuess S Storm TA Muramatsu S Nara Y Kay MA Unrestricted hepatocyte transduction with adeno-associated virus serotype 8 vectors in mice.J Virol. 2005; 79: 214-224Crossref PubMed Scopus (255) Google Scholar As few as 3 × 1011 vector particles transduced the majority of cardiomyocytes.15Nakai H Fuess S Storm TA Muramatsu S Nara Y Kay MA Unrestricted hepatocyte transduction with adeno-associated virus serotype 8 vectors in mice.J Virol. 2005; 79: 214-224Crossref PubMed Scopus (255) Google Scholar,16Wang Z Zhu T Qiao CP Zhou LQ Wang B Zhang J et al.Adeno-associated virus serotype 8 efficiently delivers genes to muscle and heart.Nat Biotechnol. 2005; 23: 321-328Crossref PubMed Scopus (503) Google Scholar More relevant to Pompe disease, an AAV2/1 vector encoding GAA reduced glycogen storage following intravenous administration to neonatal GAA-KO mice.4Mah C Cresawn KO Fraites TJ Jr Pacak CA Lewis MA Zolotukhin I et al.Sustained correction of glycogen storage disease type II using adeno-associated virus serotype 1 vectors.Gene Ther. 2005; 12: 1405-1409Crossref PubMed Scopus (60) Google Scholar Thus, current data from Pompe and muscular dystrophy mice endorse the further investigation of AAV pseudotypes with enhanced muscle tropism in these models.2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar,4Mah C Cresawn KO Fraites TJ Jr Pacak CA Lewis MA Zolotukhin I et al.Sustained correction of glycogen storage disease type II using adeno-associated virus serotype 1 vectors.Gene Ther. 2005; 12: 1405-1409Crossref PubMed Scopus (60) Google Scholar,17Rutledge EA Halbert CL Russell DW Infectious clones and vectors derived from adeno-associated virus (AAV) serotypes other than AAV type 2.J Virol. 1998; 72: 309-319Crossref PubMed Google Scholar,18Bostick B Ghosh A Yue Y Long C Duan D Systemic AAV-9 transduction in mice is influenced by animal age but not by the route of administration.Gene Ther. 2007; 14: 1605-1609Crossref PubMed Scopus (137) Google Scholar Salva et al. designed a series of highly active muscle-specific regulatory cassettes that were evaluated in striated muscle following systemic delivery via tail vein injections of AAV6 vectors.14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar The most active cassette in a variety of anatomical muscles combined a 190-bp enhancer from the murine α-myosin heavy chain gene with a 570 bp abbreviated MCK regulatory cassette, termed the MHCK7. The latter expressed human placental alkaline phosphatase at very high levels in murine heart and skeletal muscle, exceeding levels achieved with the CMV promoter/enhancer in the heart, and expressed high levels of microdystrophin in skeletal and cardiac muscles of mdx mice.14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar We hypothesized that systemic administration of an AAV vector containing a muscle-specific regulatory cassette could achieve long-term correction of multiple muscles in GAA-KO mice. An AAV vector containing the MHCK7 cassette was pseudotyped with AAV serotypes 7, 8, and 9, and the efficacy of each serotype was evaluated in GAA-KO mice. The transduction of striated muscle with AAV2/8 vectors containing either an abbreviated MCK regulatory cassette (AAV2/8-CK1hGAApA2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar) or a hybrid α-myosin heavy chain enhancer/MCK cassette (AAV2/8-MHCK7hGAApA) were evaluated, following intravenous administration in adult GAA-KO mice (1 × 1011 or 1 × 1012 vector particles/mouse). The former vector contains the CK1 promoter13Shield MA Haugen HS Clegg CH Hauschka SD E-box sites and a proximal regulatory region of the muscle creatine kinase gene differentially regulate expression in diverse skeletal muscles and cardiac muscle of transgenic mice.Mol Cell Biol. 1996; 16: 5058-5068Crossref PubMed Scopus (99) Google Scholar and the latter contains MHCK7.14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar GAA activity and glycogen content in the heart, liver, and skeletal muscles were analyzed 18 weeks following vector administration (Figure 1). GAA activity and glycogen content were significantly affected by both dosage and vector type for all tissues examined (P < 0.05 with a two-way ANOVA). Bonferroni post-test comparisons of the two vectors showed that GAA levels were significantly greater in all tissues treated with the higher dose of AAV2/8-MHCK7hGAApA compared with the equivalent dose of AAV2/8-CK1hGAApA(Figure 1). At the low dose the difference between the two vectors was not significant for any of the tissues, although the mean GAA levels were greater in each tissue for the AAV2/8-MHCK7hGAApA group. The lower GAA activity in the liver following AAV2/8-CK1hGAApA administration suggested a more stringent muscle-restricted expression with the CK1 cassette relative to the MHCK7 cassette (Figure 1a).14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar Real-time reverse transcriptase–PCR revealed that liver expression of GAA was significantly increased with AAV2/8-MHCK7hGAApA, in comparison with AAV2/8-CK1hGAApA, when each was administered at the higher dose (2.2 ± 1.2% of β-actin RNA versus 0.20 ± 0.05%, respectively; P = 0.006). However, secretion of hGAA from transduced muscles and uptake by the liver also could have contributed to the higher GAA activity in the liver following AAV2/8-MHCK7hGAApA administration. Glycogen was significantly lower in all muscles treated with AAV2/8-MHCK7hGAApA at both low and high doses, in comparison with the equivalent dose of AAV2/8-CK1hGAApA (Figure 1b). The presence of residual glycogen accumulation in hindlimb muscles, particularly the gastrocnemius, following AAV2/8 vector administration prompted a comparison with the AAV2/7 and AAV2/9 pseudotypes of AAV-MCHK7hGAApA. The GAA activities of the heart and skeletal muscles were significantly increased following AAV2/8-MHCK7hGAApA or AAV2/9- MHCK7hGAApA administration (P < 0.05), in comparison with AAV2/7-MHCK7hGAApA (Figure 1c). The glycogen contents of the heart and skeletal muscles were significantly decreased following AAV2/8-MHCK7hGAApA or AAV2/9-MHCK7hGAApA administration (P < 0.05), in comparison with AAV2/7-MHCK7hGAApA (Figure 1d). Vector genomes were quantified to allow a comparison of the transduction efficiency with each pseudotype of AAV-MHCK7hGAApA (Figure 1e). Long-term hGAA expression in heart and skeletal muscles was associated with the persistence of up to ∼1 vector genome per nuclear genome in the heart, gastrocnemius, and quadriceps, whereas more than tenfold higher amounts of vector DNA were present in the liver (Figure 1e). The enhanced correction with the AAV2/8 and AAV2/9 vectors, in comparison with AAV2/7, correlated with slightly increased numbers of vector genomes in the heart, quadriceps, and gastrocnemius (Figure 1e); moreover, the AAV2/8 and AAV2/9 vectors demonstrated significantly increased normalized GAA expression in striated muscle and heart (Figure 1f) (P < 0.05). Taken together, these data indicated that the AAV2/8 and AAV2/9 vectors transduced striated muscle more efficiently and produced higher GAA activity in transduced myofibers, in comparison with the AAV2/7 vector. Western blot analysis revealed the ∼76 kd processed form of GAA in the heart, quadriceps, gastrocnemius, and liver of GAA-KO mice following administration of the higher number of particles of AAV2/8-CK1hGAApA and AAV2/8-MHCK6hGAApA, whereas a very low signal was detected in the diaphragm following transduction with AAV2/8-CK1hGAApA (Figure 2, liver GAA not shown). GAA activity and glycogen content were not significantly changed in diaphragm following administration of AAV2/8-CK1hGAApA (Figure 1), consistent with the low activity for the CK1 cassette in the diaphragm.12Gregorevic P Blankinship MJ Allen JM Crawford RW Meuse L Miller DG et al.Systemic delivery of genes to striated muscles using adeno-associated viral vectors.Nat Med. 2004; 10: 828-834Crossref PubMed Scopus (544) Google Scholar,13Shield MA Haugen HS Clegg CH Hauschka SD E-box sites and a proximal regulatory region of the muscle creatine kinase gene differentially regulate expression in diverse skeletal muscles and cardiac muscle of transgenic mice.Mol Cell Biol. 1996; 16: 5058-5068Crossref PubMed Scopus (99) Google Scholar,14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar Glycogen staining revealed the basis for incomplete correction of skeletal gastrocnemius with AAV2/8-MHCK7hGAApA (Figure 1b). Multiple glycogen laden myofibers were detected in the gastrocnemius, consistent with the lack of correction of GAA deficiency within individual myofibers (Figure 3a). The quadriceps contained a higher number of normal-appearing myofibers (Figure 3a, arrows), consistent with the lower glycogen content in the quadriceps (Figure 1b). Thus, the relatively higher residual glycogen in the gastrocnemius could be attributed to the greater prevalence of untransduced myofibers. Lymphocytic infiltrates were absent in skeletal muscle, indicating a lack of cytotoxic T-cell responses in transduced muscle (data not shown). To evaluate the distribution of functionally transduced myofibers, mice were injected with 1 × 1012 vector particles of an AAV2/8 vector carrying a human placental alkaline phosphatase reporter cDNA driven by the MHCK7 regulatory cassette. The highest level of functional transduction was detected in the heart, followed by the quadriceps, whereas fewer transduced myofibers were present in the gastrocnemius (Figure 3b). The soleus and extensor digitalis longus (EDL) were transduced at even lower frequency than the gastrocnemius (less than three positive myofibers/field; data not shown). The low transduction of the soleus and EDL demonstrated a clear limitation for the AAV2/8 pseudotype. The GAA activities of the EDL and soleus were significantly increased following AAV2/9-MHCK7hGAApA administration (P < 0.05), in comparison with AAV2/7-MHCK7hGAApA (Figure 4a). As expected, the glycogen contents of the EDL and soleus were significantly decreased following AAV2/9- MHCK7hGAApA administration (Figure 4b). Thus, the transduction of the small hindlimb muscles was superior for the AAV9-pseudotyped vector. Antibodies against hGAA have prevented efficient cross-correction of GAA deficiency through receptor-mediated uptake, despite the presence of secreted hGAA in the blood.2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar,3Franco LM Sun B Yang X Bird A Zhang H Schneider A et al.Evasion of immune responses to introduced human acid alpha-glucosidase by liver-restricted expression in glycogen storage disease type II.Mol Ther. 2005; 12: 876-884Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar,19Ding EY Hodges BL Hu H McVie-Wylie AJ Serra D Migone FK et al.Long-term efficacy after [E1-,polymerase-] adenovirus-mediated transfer of human acid-alpha-glucosidase gene into glycogen storage disease type II knockout mice.Hum Gene Ther. 2001; 12: 955-965Crossref PubMed Scopus (63) Google Scholar Anti-hGAA antibodies were significantly elevated by 6 weeks following administration of both low and high doses of AAV2/8 vector (P < 0.05), regardless of the regulatory cassette type (Figure 5a). Despite the presence of anti-GAA antibodies, Rotarod times were significantly increased (P < 0.05) at 12 and 18 weeks following administration of AAV-MHCK7hGAApA-injected mice for the AAV2/7, AAV2/8, and AAV2/9 pseudotypes (Figure 5b). However, Rotarod times were not significantly increased at 12 or 18 weeks for AAV-CK1hGAApA-treated mice. A glucotetrasaccharide biomarker for increased glycogen storage, Glcα1-6Glcα1-4Glcα1-4Glc, (Glc4), was significantly reduced (P < 0.05) at 18 weeks for the high-dose group of all vector-injected GAA-KO mice, regardless of the regulatory cassette or pseudotype in comparison with phosphate-buffered saline–injected mice (Figure 5c). There was no difference in mean Glc4 levels between the different vector-treated groups. Hence, a significant increase in Rotarod time and reduction of Glc4 biomarker were achieved, despite the presence of an antibody response against hGAA. Wide-spread clearance of glycogen in individual myofibers was demonstrated following AAV2/7 and AAV2/9 vector administration (Figure 6). Fiber typing confirmed the high prevalence of type I, slow-twitch fibers in the soleus of GAA-KO mice, whereas the EDL was comprised mainly of type IIb myofibers (data not shown). The clearance of glycogen vacuolation within multiple myofibers in the EDL and soleus indicated that both type I and IIb myofibers were transduced by the AAV2/9 vector (Figure 6). Gene therapy in the muscular dystrophies will likely require therapeutic gene expression in striated muscle generally, particularly for lethal muscular dystrophies such as Duchenne muscular dystrophy. Pompe disease uniquely responds to infused or secreted therapeutic protein, because unlike other muscular dystrophies Pompe disease is a lysosomal storage disorder amenable to enzyme replacement therapy. Owing to the high enzyme level requirements in enzyme replacement therapy and complicating antibody responses to GAA, muscle-targeted gene therapy is under development in Pompe disease.2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar,20Raben N Plotz P Byrne BJ Acid alpha-glucosidase deficiency (glycogenosis type II, Pompe disease).Curr Mol Med. 2002; 2: 145-166Crossref PubMed Scopus (192) Google Scholar,21Fraites TJ Jr Schleissing MR Shanely RA Walter GA Cloutier DA Zolotukhin I et al.Correction of the enzymatic and functional deficits in a model of Pompe disease using adeno-associated virus vectors.Mol Ther. 2002; 5: 571-578Abstract Full Text Full Text PDF PubMed Scopus (106) Google Scholar Systemic delivery of an AAV2/8 vector encoding muscle-restricted hGAA achieved significant efficacy; however, the transduction of myofibers was partial, preventing the complete correction of glycogen storage in the gastrocnemius. The analogous AAV2/9 vector demonstrated significantly increased transduction of the small hindlimb muscles, in direct comparison with the AAV2/7 vector. Transduction of the EDL and soleus were inefficient with an AAV2/8 vector encoding human placental alkaline phosphatase, implying that the AAV2/9 pseudotype might have significant advantages with regard to transduction of the distal hindlimb muscles. Type I fibers were more easily cleared of glycogen during enzyme replacement therapy, indicating the need to focus on the transduction of type II fibers in Pompe disease.22Raben N Danon M Gilbert AL Dwivedi S Collins B Thurberg BL et al.Enzyme replacement therapy in the mouse model of Pompe disease.Mol Genet Metab. 2003; 80: 159-169Abstract Full Text Full Text PDF PubMed Scopus (178) Google Scholar,23Fukuda T Ewan L Bauer M Mattaliano RJ Zaal K Ralston E et al.Dysfunction of endocytic and autophagic pathways in a lysosomal storage disease.Ann Neurol. 2006; 59: 700-708Crossref PubMed Scopus (248) Google Scholar Type I myofibers were transduced less efficiently with an AAV2 vector than with an AAV2/6 vector, reflecting the potential advantage of newer serotypes over AAV2.24Blankinship MJ Gregorevic P Allen JM Harper SQ Harper H Halbert CL et al.Efficient transduction of skeletal muscle using vectors based on adeno-associated virus serotype 6.Mol Ther. 2004; 10: 671-678Abstract Full Text Full Text PDF PubMed Scopus (197) Google Scholar,25Pruchnic R Cao B Peterson ZQ Xiao X Li J Samulski RJ et al.The use of adeno-associated virus to circumvent the maturation-dependent viral transduction of muscle fibers.Hum Gene Ther. 2000; 11: 521-536Crossref PubMed Scopus (74) Google Scholar Currently, the AAV2/9 serotype efficiently transduced both type I and IIb myofibers, as well as cardiomyocytes, following intravenous administration. This study further endorses the role of AAV2/9 vectors for gene therapy in Pompe disease, demonstrating significant correction of not only the heart but all skeletal muscles examined. Previously AAV2/9 vectors have transduced striated muscle more efficiently than either an AAV2/1 vector in neonatal mice26Pacak CA Mah CS Thattaliyath BD Conlon TJ Lewis MA Cloutier DE et al.Recombinant adeno-associated virus serotype 9 leads to preferential cardiac transduction in vivo.Circ Res. 2006; 99: e3-e9Crossref PubMed Scopus (316) Google Scholar or an AAV2/8 vector in adult mice.27Inagaki K Fuess S Storm TA Gibson GA McTiernan CF Kay MA et al.Robust systemic transduction with AAV9 vectors in mice: efficient global cardiac gene transfer superior to that of AAV8.Mol Ther. 2006; 14: 45-53Abstract Full Text Full Text PDF PubMed Scopus (467) Google Scholar Neither of these earlier studies demonstrated significant correction of multiple skeletal muscles in adult mice.26Pacak CA Mah CS Thattaliyath BD Conlon TJ Lewis MA Cloutier DE et al.Recombinant adeno-associated virus serotype 9 leads to preferential cardiac transduction in vivo.Circ Res. 2006; 99: e3-e9Crossref PubMed Scopus (316) Google Scholar,27Inagaki K Fuess S Storm TA Gibson GA McTiernan CF Kay MA et al.Robust systemic transduction with AAV9 vectors in mice: efficient global cardiac gene transfer superior to that of AAV8.Mol Ther. 2006; 14: 45-53Abstract Full Text Full Text PDF PubMed Scopus (467) Google Scholar The improved efficacy of this new AAV2/9 vector further strengthens preclinical data in favor of clinical trials of gene therapy in Pompe disease. The successful development of gene therapy in GAA-KO mice indicates that curative therapy for Pompe disease may become available in the foreseeable future; however, the efficacy of gene therapy in these experiments was inversely related to the presence of immune responses. Immunocompetent GAA-KO mice produced high titer anti-hGAA immunoglobulin G in response to an AAV vector containing the ubiquitously active CMV promoter/chicken β-actin (CB) promoter, packaged as either AAV2/6 or AAV2/8.2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar,3Franco LM Sun B Yang X Bird A Zhang H Schneider A et al.Evasion of immune responses to introduced human acid alpha-glucosidase by liver-restricted expression in glycogen storage disease type II.Mol Ther. 2005; 12: 876-884Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar The CB-containing vector failed to secrete detectable hGAA in the plasma for >2 weeks or to reduce glycogen storage in the muscle of GAA-KO mice.3Franco LM Sun B Yang X Bird A Zhang H Schneider A et al.Evasion of immune responses to introduced human acid alpha-glucosidase by liver-restricted expression in glycogen storage disease type II.Mol Ther. 2005; 12: 876-884Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar Immune responses to ubiquitously expressed hGAA included lymphocytic infiltrates and activation of CD4+ and CD8+ lymphocytes in injected skeletal muscle, and in the liver following intravenous injection.2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar,3Franco LM Sun B Yang X Bird A Zhang H Schneider A et al.Evasion of immune responses to introduced human acid alpha-glucosidase by liver-restricted expression in glycogen storage disease type II.Mol Ther. 2005; 12: 876-884Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar Substitution of the CK1 regulatory cassette in place of the CB promoter prevented CD8+ lymphocytic responses in the injected muscle. Therefore, both antibody production and cytotoxic T lymphocytes were elicited in response to hGAA production from a ubiquitously active CB promoter; however, vectors containing muscle-specific cassettes did not provoke lymphocytic infiltrates in transduced muscles and expressed hGAA for >18 weeks.2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar The latter experiment also demonstrated the imperviousness of muscle-specific hGAA expression to circulating anti-GAA antibodies. The presence of anti-GAA antibodies has prevented cross-correction of untransduced muscle cells,1Sun B Zhang H Franco LM Young SP Schneider A Bird A et al.Efficacy of an adeno-associated virus 8-pseudotyped vector in glycogen storage disease type II.Mol Ther. 2005; 11: 57-65Abstract Full Text Full Text PDF PubMed Scopus (121) Google Scholar,2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar,3Franco LM Sun B Yang X Bird A Zhang H Schneider A et al.Evasion of immune responses to introduced human acid alpha-glucosidase by liver-restricted expression in glycogen storage disease type II.Mol Ther. 2005; 12: 876-884Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar,19Ding EY Hodges BL Hu H McVie-Wylie AJ Serra D Migone FK et al.Long-term efficacy after [E1-,polymerase-] adenovirus-mediated transfer of human acid-alpha-glucosidase gene into glycogen storage disease type II knockout mice.Hum Gene Ther. 2001; 12: 955-965Crossref PubMed Scopus (63) Google Scholar implicating the transduction of individual muscle cells as the source of glycogen clearance in the current study. The MHCK7 regulatory cassette has driven highly efficacious hGAA expression in the Pompe disease mouse model in this study. A comparison of AAV2/6 vectors containing either the MHCK7 or the CK1 cassette demonstrated high transgene expression within cardiac muscle and skeletal muscles,14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar which has now been replicated in Pompe disease mice for AAV2/7, AAV2/8, and AAV2/9 vectors. Virtually all myofibers were transduced in the muscles examined with the aforementioned AAV2/6 vector, although transgene expression was reduced in the diaphragm; moreover, transduction was enhanced by the co-administration of the vascular permeabilizing agent vascular endothelial growth factor at lower vector doses.12Gregorevic P Blankinship MJ Allen JM Crawford RW Meuse L Miller DG et al.Systemic delivery of genes to striated muscles using adeno-associated viral vectors.Nat Med. 2004; 10: 828-834Crossref PubMed Scopus (544) Google Scholar The current AAV2/8 vector achieved partial clearance of glycogen from the diaphragm in Pompe mice through the administration of reasonably low particle numbers, increasing the likelihood of translation to clinical applications in Pompe disease. The activity of the MCHK7 regulatory cassette was higher than that for the CK1 cassette in heart, quadriceps, gastrocnemius, and diaphragm, all critical targets for gene therapy in Pompe disease and other forms of muscular dystrophy. The combination of highly active muscle-specific regulatory cassettes and novel AAV serotypes promises to advance gene therapy for muscular dystrophy by providing curative therapy for these devastating disorders. Preparation of pseudotyped AAV vectors. AAV-MHCK7hGAApA contains the MHCK7 regulatory cassette,14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar the hGAA cDNA, and a human growth hormone polyadenylation sequence. The vector plasmid, pAAV-MHCK7hGAApA, was derived from pAAV-CBhGAApA.28Sun B Chen YT Bird A Xu F Hou YX Amalfitano A et al.Packaging of an AAV vector encoding human acid alpha-glucosidase for gene therapy in glycogen storage disease type II with a modified hybrid adenovirus-AAV vector.Mol Ther. 2003; 7: 467-477Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar The pAAV-MHCK7hGAApA was digested with KpnI, blunt-ended with the Klenow fragment of DNA polymerase, then digested with XbaI; subsequently, the 5.7-kb blunt/XbaI fragment from pAAV-MHCK7hGAApA was ligated with a 0.8-kb XbaI/blunt SalI fragment containing MHCK7 cassette from αMHCKChAP14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar (provided by Dr. Stephen Hauschka, University of Washington, Seattle, WA). AAV-CK1hGAApA has been described (formerly AAV-MCKhGAApA).2Sun B Zhang H Franco LM Brown T Bird A Schneider A et al.Correction of glycogen storage disease type II by an adeno-associated virus vector containing a muscle-specific promoter.Mol Ther. 2005; 11: 889-898Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar Briefly, 293 cells were transfected with an AAV vector, the AAV packaging plasmid29Gao GP Alvira MR Wang L Calcedo R Johnston J Wilson JM Novel adeno-associated viruses from rhesus monkeys as vectors for human gene therapy.Proc Natl Acad Sci USA. 2002; 99: 11854-11859Crossref PubMed Scopus (1237) Google Scholar (courtesy of Dr. James M. Wilson, University of Pennsylvania, Philadelphia, PA), and pAdHelper (Stratagene, La Jolla, CA). Cell lysate was harvested 48 hours following infection and freeze-thawed three times, and isolated by sucrose cushion pelleting followed by two cesium chloride gradient centrifugation steps. AAV stocks were dialyzed against three changes of Hanks buffer, and aliquots were stored at −80 °C. The number of vector DNA–containing particles in viral stocks was determined by DNase I digestion, DNA extraction, and Southern blot analysis. The Southern blot signal for vector genomes was quantified by comparison with the signals from standards consisting of AhdI-digested vector plasmid. All viral vector stocks were handled according to Biohazard Safety Level 2 guidelines published by the National Institutes of Health. In vivo analysis of AAV vector. The AAV vector stocks were administered intravenously (via the retroorbital sinus) in 3-month-old GAA-KO mice.30Raben N Nagaraju K Lee E Kessler P Byrne B Lee L et al.Targeted disruption of the acid alpha-glucosidase gene in mice causes an illness with critical features of both infantile and adult human glycogen storage disease type II.J Biol Chem. 1998; 273: 19086-19092Crossref PubMed Scopus (223) Google Scholar At the indicated time points after injection, plasma or tissue samples were obtained and processed as described below. All animal procedures were performed in accordance with Duke University Institutional Animal Care and Use Committee–approved guidelines. Rotarod testing was performed as previously described.1Sun B Zhang H Franco LM Young SP Schneider A Bird A et al.Efficacy of an adeno-associated virus 8-pseudotyped vector in glycogen storage disease type II.Mol Ther. 2005; 11: 57-65Abstract Full Text Full Text PDF PubMed Scopus (121) Google Scholar GAA activity and glycogen content were analyzed as described, and real-time RT-PCR was performed with standard methods using primers previously described.31Amalfitano A McVie-Wylie AJ Hu H Dawson TL Raben N Plotz P et al.Systemic correction of the muscle disorder glycogen storage disease type II after hepatic targeting of a modified adenovirus vector encoding human acid-alpha-glucosidase.Proc Natl Acad Sci USA. 1999; 96: 8861-8866Crossref PubMed Scopus (133) Google Scholar Western blotting of hGAA was performed as previously described1Sun B Zhang H Franco LM Young SP Schneider A Bird A et al.Efficacy of an adeno-associated virus 8-pseudotyped vector in glycogen storage disease type II.Mol Ther. 2005; 11: 57-65Abstract Full Text Full Text PDF PubMed Scopus (121) Google Scholar using the hGAA monoclonal antibody (courtesy of Genzyme, Framingham, MA). Alkaline phosphatase staining was performed as previously described.14Salva MZ Himeda CL Tai PW Nishiuchi E Gregorevic P Allen JM et al.Design of tissue-specific regulatory cassettes for high-level rAAV-mediated expression in skeletal and cardiac muscle.Mol Ther. 2007; 15: 320-329Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar The enzyme-linked immunosorbent assay was performed as previously described.3Franco LM Sun B Yang X Bird A Zhang H Schneider A et al.Evasion of immune responses to introduced human acid alpha-glucosidase by liver-restricted expression in glycogen storage disease type II.Mol Ther. 2005; 12: 876-884Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar All samples yielded absorbance values that were within the linear range of the assay at this dilution. Urinary Glc4 concentrations were determined relative to creatinine by stable isotope-dilution electrospray tandem mass spectrometry as previously described.32Young SP Stevens RD An Y Chen YT Millington DS Analysis of a glucose tetrasaccharide elevated in Pompe disease by stable isotope dilution-electrospray ionization tandem mass spectrometry.Anal Biochem. 2003; 316: 175-180Crossref PubMed Scopus (59) Google Scholar Statistical analyses. Two-way ANOVAs were performed using the independent variables of vector type and dose of vector, with Bonferroni post-tests for specific comparison between groups (Prism 3.0, GraphPad, San Diego, CA). Comparison of two groups was assessed by a homoscedastic Student's t-test. A P value of <0.05 indicated a significant difference between the observed values for each group. This work was supported by National Institutes of Health grant R01 HL081122-01A1 from the National Heart, Lung, and Blood Institute. D.D.K. was supported by the Muscular Dystrophy Association and Genzyme Corporation. B.S. was supported by a Development Grant from the Muscular Dystrophy Association. Development and evaluation of the MCK regulatory cassettes were supported by National Institutes of Health grants RO1 AR18860, 1 U54 HD047175, and R24 HL64387, and the Muscular Dystrophy Association to S.D.H. GAA-KO mice were provided courtesy of Nina Raben at the National Institutes of Health (Bethesda, MD). The AAV8 packaging plasmid, p5E18-VD 2/8, was provided courtesy of James M. Wilson at the University of Pennsylvania (Philadelphia, PA).}, number={8}, journal={MOLECULAR THERAPY}, author={Sun, Baodong and Young, Sarah P. and Li, Ping and Di, Chunhui and Brown, Talmage and Salva, Maja Z. and Li, Songtao and Bird, Andrew and Yan, Zhen and Auten, Richard and et al.}, year={2008}, month={Aug}, pages={1366–1371} } @article{mackillop_olby_linder_brown_2007, title={Intramedullary cavernous malformation of the spinal cord in two dogs}, volume={44}, ISSN={["1544-2217"]}, DOI={10.1354/vp.44-4-528}, abstractNote={Intramedullary cavernous malformations (CVMs) of the spinal cord were diagnosed in 2 adult dogs that presented for paraparesis. An intramedullary spinal cord lesion was identified on a myelogram in the first dog, and expansion of the vertebral canal was evident on radiographs in the second. Extensive intraparenchymal hemorrhage was found on gross postmortem examination in both dogs, and a distinct lobulated intramedullary mass was evident in the second dog. Microscopically, both lesions were composed of dilated, thin-walled vascular channels with little-to-no intervening neural parenchyma. Both dogs had evidence of channel thrombosis along with perilesional hemorrhage and hemosiderin accumulation. The second dog had additional degenerative changes, including thickened fibrous channel walls with hyalinization, foci of mineralization, and occasional tongues of entrapped gliotic neuropil. CVMs appear to be an uncommon cause of both acute and chronic spinal cord disease in the dog.}, number={4}, journal={VETERINARY PATHOLOGY}, author={Mackillop, E. and Olby, N. J. and Linder, K. E. and Brown, T. T.}, year={2007}, month={Jul}, pages={528–532} } @article{koeberl_sun_damodaran_brown_millington_benjamin_bird_schneider_hillman_jackson_et al._2006, title={Early, sustained efficacy of adeno-associated virus vector-mediated gene therapy in glycogen storage disease type Ia}, volume={13}, ISSN={["1476-5462"]}, DOI={10.1038/sj.gt.3302774}, abstractNote={The deficiency of glucose-6-phosphatase (G6Pase) underlies life-threatening hypoglycemia and growth retardation in glycogen storage disease type Ia (GSD-Ia). An adeno-associated virus (AAV) vector encoding G6Pase was pseudotyped as AAV8 and administered to 2-week-old GSD-Ia mice (n = 9). Median survival was prolonged to 7 months following vector administration, in contrast to untreated GSD-Ia mice that survived for only 2 weeks. Although GSD-Ia mice were initially growth-retarded, treated mice increased fourfold in weight to normal size. Blood glucose was partially corrected by 2 weeks following treatment, whereas blood cholesterol normalized. Glucose-6-phosphatase activity was partially corrected to 25% of the normal level at 7 months of age in treated mice, and blood glucose during fasting remained lower in treated, affected mice than in normal mice. Glycogen storage was partially corrected in the liver by 2 weeks following treatment, but reaccumulated to pre-treatment levels by 7 months old (m.o.). Vector genome DNA decreased between 3 days and 3 weeks in the liver following vector administration, mainly through the loss of single-stranded genomes; however, double-stranded vector genomes were more stable. Although CD8+ lymphocytic infiltrates were present in the liver, partial biochemical correction was sustained at 7 m.o. The development of efficacious AAV vector-mediated gene therapy could significantly reduce the impact of long-term complications in GSD-Ia, including hypoglycemia, hyperlipidemia and growth failure.}, number={17}, journal={GENE THERAPY}, author={Koeberl, D. D. and Sun, B. D. and Damodaran, T. V. and Brown, T. and Millington, D. S. and Benjamin, D. J., Jr. and Bird, A. and Schneider, A. and Hillman, S. and Jackson, M. and et al.}, year={2006}, month={Sep}, pages={1281–1289} } @article{sun_zhang_benjamin_brown_bird_young_mcvie-wylie_chen_koeberl_2006, title={Enhanced efficacy of an AAV vector encoding chimeric, highly secreted acid alpha-glucosidase in glycogen storage disease type II}, volume={14}, ISSN={["1525-0024"]}, DOI={10.1016/j.ymthe.2006.08.001}, abstractNote={