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Complete correction of acid alpha-glucosidase deficiency in Pompe disease fibroblasts in vitro, and lysosomally targeted expression in neonatal rat cardiac and skeletal muscle.
Gene Ther 1998; 5(4):473-80GT

Abstract

The enzyme acid alpha-glucosidase catalyzes the breakdown of lysosomal glycogen. Absence of this enzyme results in infantile Pompe disease, characterized by hypertrophic cardiomyopathy, skeletal muscle weakness and fatal heart failure by 2 years of age. We have examined the possibility of gene replacement therapy for this disease, by constructing an E1-deleted recombinant adenovirus encoding human acid alpha-glucosidase (Ad-GAA). The dose-response in fibroblasts from patients with Pompe disease transduced with this vector is linear over the range tested (one to 2000 plaque forming units (p.f.u.) of Ad-GAA per cell), and acid alpha-glucosidase activity comparable to that of normal fibroblasts is achieved at 100 p.f.u. per cell. Targeting of the recombinant protein to the lysosomal compartment was confirmed by immunocytochemistry. In vivo expression was examined by injecting Ad-GAA into newborn rats; intracardiac administration produced 10 times the normal level of acid alpha-glucosidase activity in whole heart lysates, while a hind-limb i.m. injection increased activity in that muscle to six times the normal level. Western blotting of these tissues defected species at 76 kDa consistent with the size of processed lysosomal enzyme, and levels of expression as high as 1.0 mg recombinant protein per gram of tissue wet weight were produced. These data demonstrate high-level, lysosomal expression of recombinant acid alpha-glucosidase in treated target tissues and support the feasibility of gene replacement strategies for Pompe disease.

Authors+Show Affiliations

Peter Belfer Cardiac Laboratory, Johns Hopkins University School of Medicine, Baltimore, MD, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

9614571

Citation

Pauly, D F., et al. "Complete Correction of Acid Alpha-glucosidase Deficiency in Pompe Disease Fibroblasts in Vitro, and Lysosomally Targeted Expression in Neonatal Rat Cardiac and Skeletal Muscle." Gene Therapy, vol. 5, no. 4, 1998, pp. 473-80.
Pauly DF, Johns DC, Matelis LA, et al. Complete correction of acid alpha-glucosidase deficiency in Pompe disease fibroblasts in vitro, and lysosomally targeted expression in neonatal rat cardiac and skeletal muscle. Gene Ther. 1998;5(4):473-80.
Pauly, D. F., Johns, D. C., Matelis, L. A., Lawrence, J. H., Byrne, B. J., & Kessler, P. D. (1998). Complete correction of acid alpha-glucosidase deficiency in Pompe disease fibroblasts in vitro, and lysosomally targeted expression in neonatal rat cardiac and skeletal muscle. Gene Therapy, 5(4), pp. 473-80.
Pauly DF, et al. Complete Correction of Acid Alpha-glucosidase Deficiency in Pompe Disease Fibroblasts in Vitro, and Lysosomally Targeted Expression in Neonatal Rat Cardiac and Skeletal Muscle. Gene Ther. 1998;5(4):473-80. PubMed PMID: 9614571.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Complete correction of acid alpha-glucosidase deficiency in Pompe disease fibroblasts in vitro, and lysosomally targeted expression in neonatal rat cardiac and skeletal muscle. AU - Pauly,D F, AU - Johns,D C, AU - Matelis,L A, AU - Lawrence,J H, AU - Byrne,B J, AU - Kessler,P D, PY - 1998/6/6/pubmed PY - 1998/6/6/medline PY - 1998/6/6/entrez SP - 473 EP - 80 JF - Gene therapy JO - Gene Ther. VL - 5 IS - 4 N2 - The enzyme acid alpha-glucosidase catalyzes the breakdown of lysosomal glycogen. Absence of this enzyme results in infantile Pompe disease, characterized by hypertrophic cardiomyopathy, skeletal muscle weakness and fatal heart failure by 2 years of age. We have examined the possibility of gene replacement therapy for this disease, by constructing an E1-deleted recombinant adenovirus encoding human acid alpha-glucosidase (Ad-GAA). The dose-response in fibroblasts from patients with Pompe disease transduced with this vector is linear over the range tested (one to 2000 plaque forming units (p.f.u.) of Ad-GAA per cell), and acid alpha-glucosidase activity comparable to that of normal fibroblasts is achieved at 100 p.f.u. per cell. Targeting of the recombinant protein to the lysosomal compartment was confirmed by immunocytochemistry. In vivo expression was examined by injecting Ad-GAA into newborn rats; intracardiac administration produced 10 times the normal level of acid alpha-glucosidase activity in whole heart lysates, while a hind-limb i.m. injection increased activity in that muscle to six times the normal level. Western blotting of these tissues defected species at 76 kDa consistent with the size of processed lysosomal enzyme, and levels of expression as high as 1.0 mg recombinant protein per gram of tissue wet weight were produced. These data demonstrate high-level, lysosomal expression of recombinant acid alpha-glucosidase in treated target tissues and support the feasibility of gene replacement strategies for Pompe disease. SN - 0969-7128 UR - https://www.unboundmedicine.com/medline/citation/9614571/Complete_correction_of_acid_alpha_glucosidase_deficiency_in_Pompe_disease_fibroblasts_in_vitro_and_lysosomally_targeted_expression_in_neonatal_rat_cardiac_and_skeletal_muscle_ L2 - http://dx.doi.org/10.1038/sj.gt.3300609 DB - PRIME DP - Unbound Medicine ER -