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Correction of glycogen storage disease type II by enzyme replacement with a recombinant human acid maltase produced by over-expression in a CHO-DHFR(neg) cell line.
Biochem Biophys Res Commun 2000; 276(3):917-23BB

Abstract

Inherited genetic deficiency of lysosomal acid alpha glucosidase or acid maltase (GAA) results in the autosomal recessive glycogen storage disease type II (GSD II). To investigate whether we could generate a functional recombinant human GAA (rhGAA) for enzyme replacement therapy, we subcloned the cDNAs for human GAA and mouse dihydrofolate reductase (DHFR) into DHFR(neg) Chinese hamster ovary cells and established a stable cotransformant that expressed rhGAA. We cultured the recombinant cells in media with progressively increasing concentrations of methotrexate and found that human GAA enzyme activity increased to over 2,000 IU per gram protein. Importantly, the human GAA enzyme activity correlated to equivalent amounts of human GAA protein by rocketimmunoelectrophoresis. We confirmed that the human GAA enzyme activity corresponded to an amplification in human GAA mRNA by Northern analysis and human GAA cDNA copy number by Southern analysis. Exposing the rhGAA to human GSDII fibroblast cells or patient's lymphocytes or monocytes resulted in uptake of the rhGAA and reversal of the enzymatic defect. Mannose-6-phosphate in the media blocked uptake. GAA -/- mice were treated with the rhGAA at 1 mg/kg, which resulted in heterozygous levels of GAA in tissues, most notably skeletal muscle, heart and diaphragm after two infusions. More importantly, after multiple infusions, hind, and fore-limb muscle weakness was reversed. This rhGAA would be ideal for enzyme replacement therapy in GSD II.

Authors+Show Affiliations

Department of Medicine, Pulmonary Division and Critical Care Medicine, New York University Medical Center, New Bellevue 6N5, 550 First Avenue, New York, New York, 10016, USA. martif02@popmail.med.nyu.eduNo affiliation info availableNo affiliation info availableNo 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

11027569

Citation

Martiniuk, F, et al. "Correction of Glycogen Storage Disease Type II By Enzyme Replacement With a Recombinant Human Acid Maltase Produced By Over-expression in a CHO-DHFR(neg) Cell Line." Biochemical and Biophysical Research Communications, vol. 276, no. 3, 2000, pp. 917-23.
Martiniuk F, Chen A, Donnabella V, et al. Correction of glycogen storage disease type II by enzyme replacement with a recombinant human acid maltase produced by over-expression in a CHO-DHFR(neg) cell line. Biochem Biophys Res Commun. 2000;276(3):917-23.
Martiniuk, F., Chen, A., Donnabella, V., Arvanitopoulos, E., Slonim, A. E., Raben, N., ... Rom, W. N. (2000). Correction of glycogen storage disease type II by enzyme replacement with a recombinant human acid maltase produced by over-expression in a CHO-DHFR(neg) cell line. Biochemical and Biophysical Research Communications, 276(3), pp. 917-23.
Martiniuk F, et al. Correction of Glycogen Storage Disease Type II By Enzyme Replacement With a Recombinant Human Acid Maltase Produced By Over-expression in a CHO-DHFR(neg) Cell Line. Biochem Biophys Res Commun. 2000 Oct 5;276(3):917-23. PubMed PMID: 11027569.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Correction of glycogen storage disease type II by enzyme replacement with a recombinant human acid maltase produced by over-expression in a CHO-DHFR(neg) cell line. AU - Martiniuk,F, AU - Chen,A, AU - Donnabella,V, AU - Arvanitopoulos,E, AU - Slonim,A E, AU - Raben,N, AU - Plotz,P, AU - Rom,W N, PY - 2000/10/12/pubmed PY - 2001/2/28/medline PY - 2000/10/12/entrez SP - 917 EP - 23 JF - Biochemical and biophysical research communications JO - Biochem. Biophys. Res. Commun. VL - 276 IS - 3 N2 - Inherited genetic deficiency of lysosomal acid alpha glucosidase or acid maltase (GAA) results in the autosomal recessive glycogen storage disease type II (GSD II). To investigate whether we could generate a functional recombinant human GAA (rhGAA) for enzyme replacement therapy, we subcloned the cDNAs for human GAA and mouse dihydrofolate reductase (DHFR) into DHFR(neg) Chinese hamster ovary cells and established a stable cotransformant that expressed rhGAA. We cultured the recombinant cells in media with progressively increasing concentrations of methotrexate and found that human GAA enzyme activity increased to over 2,000 IU per gram protein. Importantly, the human GAA enzyme activity correlated to equivalent amounts of human GAA protein by rocketimmunoelectrophoresis. We confirmed that the human GAA enzyme activity corresponded to an amplification in human GAA mRNA by Northern analysis and human GAA cDNA copy number by Southern analysis. Exposing the rhGAA to human GSDII fibroblast cells or patient's lymphocytes or monocytes resulted in uptake of the rhGAA and reversal of the enzymatic defect. Mannose-6-phosphate in the media blocked uptake. GAA -/- mice were treated with the rhGAA at 1 mg/kg, which resulted in heterozygous levels of GAA in tissues, most notably skeletal muscle, heart and diaphragm after two infusions. More importantly, after multiple infusions, hind, and fore-limb muscle weakness was reversed. This rhGAA would be ideal for enzyme replacement therapy in GSD II. SN - 0006-291X UR - https://www.unboundmedicine.com/medline/citation/11027569/Correction_of_glycogen_storage_disease_type_II_by_enzyme_replacement_with_a_recombinant_human_acid_maltase_produced_by_over_expression_in_a_CHO_DHFR_neg__cell_line_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0006-291X(00)93555-1 DB - PRIME DP - Unbound Medicine ER -