The iminosugar isofagomine increases the activity of N370S mutant acid β-glucosidase in Gaucher fibroblasts by several mechanisms

doi:10.1073/pnas.0605928103

The iminosugar isofagomine increases the activity of N370S mutant acid β-glucosidase in Gaucher fibroblasts by several mechanisms

*Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110; and
^†Amicus Therapeutics, 6 Cedar Brook Drive, Cranbury, NJ 08512

Contributed by Stuart A. Kornfeld, July 14, 2006

Abstract

Gaucher disease is a lysosomal storage disorder caused by deficiency in lysosomal acid β-glucosidase (GlcCerase), the enzyme responsible for the catabolism of glucosylceramide. One of the most prevalent disease-causing mutations, N370S, results in an enzyme with lower catalytic activity and impaired exit from the endoplasmic reticulum. Here, we report that the iminosugar isofagomine (IFG), an active-site inhibitor, increases GlcCerase activity 3.0 ± 0.6-fold in N370S fibroblasts by several mechanisms. A major effect of IFG is to facilitate the folding and transport of newly synthesized GlcCerase in the endoplasmic reticulum, thereby increasing the lysosomal pool of the enzyme. In addition, N370S GlcCerase synthesized in the presence of IFG exhibits a shift in pH optimum from 6.4 to 5.2 and altered sensitivity to SDS. Although IFG fully inhibits GlcCerase in the lysosome in an in situ assay, washout of the drug leads to partial recovery of GlcCerase activity within 4 h and full recovery by 24 h. These findings provide support for the possible use of active-site inhibitors in the treatment of some forms of Gaucher disease.

Footnotes

^‡To whom correspondence should be addressed. E-mail: skornfel{at}im.wustl.edu
Author contributions: R.A.S., S.C., B.W., A.P., H.D., and S.A.K. designed research; R.A.S., S.C., and A.P. performed research; R.A.S., S.C., B.W., A.P., H.D., and S.A.K. analyzed data; and R.A.S. and S.A.K. wrote the paper.
Conflict of interest statement: S.A.K. is a member of the Scientific Advisory Board of Amicus Therapeutics.
Abbreviations:

CBE,

conduritol β-epoxide;

ER,

endoplasmic reticulum;

ERT,

enzyme-replacement therapy;

GlcCerase,

acid β-glucosidase;

IFG,

isofagomine;

PFB-FDGlu,

5-(pentafluorobenzoylamino)fluorescein di-β-d-glucopyranoside;

PNGase F,

peptide:N-glycosidase F.