High-level production of recombinant human lysosomal acid alpha-glucosidase in Chinese hamster ovary cells which targets to heart muscle and corrects glycogen accumulation in fibroblasts from patients with Pompe disease

Proc Natl Acad Sci U S A. 1996 Jan 9;93(1):65-70. doi: 10.1073/pnas.93.1.65.


Infantile Pompe disease is a fatal genetic muscle disorder caused by a deficiency of acid alpha-glucosidase, a glycogen-degrading lysosomal enzyme. We constructed a plasmid containing a 5'-shortened human acid alpha-glucosidase cDNA driven by the cytomegalovirus promoter, as well as the aminoglycoside phosphotransferase and dihydrofolate reductase genes. Following transfection in dihydrofolate reductase-deficient Chinese hamster ovary cells, selection with Geneticin, and amplification with methotrexate, a cell line producing high levels of the alpha-glucosidase was established. In 48 hr, the cells cultured in Iscove's medium with 5 mM butyrate secreted 110-kDa precursor enzyme that accumulated to 91 micrograms.ml-1 in the medium (activity, > 22.6 mumol.hr-1.ml-1). This enzyme has a pH optimum similar to that of the mature form, but a lower Vmax and Km for 4-methylumbelliferyl-alpha-D-glucoside. It is efficiently taken up by fibroblasts from Pompe patients, restoring normal levels of acid alpha-glucosidase and glycogen. The uptake is blocked by mannose 6-phosphate. Following intravenous injection, high enzyme levels are seen in heart and liver. An efficient production system now exists for recombinant human acid alpha-glucosidase targeted to heart and capable of correcting fibroblasts from patients with Pompe disease.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Biological Transport
  • CHO Cells
  • Cells, Cultured
  • Cricetinae
  • DNA Primers / chemistry
  • Fibroblasts / metabolism
  • Glycogen / metabolism*
  • Glycogen Storage Disease Type II / enzymology*
  • Glycogen Storage Disease Type II / therapy
  • Humans
  • Liver / metabolism
  • Lysosomes / enzymology
  • Molecular Sequence Data
  • Muscles / metabolism
  • Myocardium / metabolism
  • Recombinant Proteins / metabolism
  • Transfection
  • alpha-Glucosidases / metabolism*


  • DNA Primers
  • Recombinant Proteins
  • Glycogen
  • alpha-Glucosidases