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J. Biol. Chem., Vol. 279, Issue 2, 1526-1532, January 9, 2004

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Mannose 6-Phosphate Receptor-mediated Uptake Is Defective in Acid Sphingomyelinase-deficient Macrophages

IMPLICATIONS FOR NIEMANN-PICK DISEASE ENZYME REPLACEMENT THERAPY^*

Rajwinder Dhami and Edward H. Schuchman¶

From the Department of Human Genetics and Carl C. Icahn Institute for Gene Therapy and Molecular Medicine, Mount Sinai School of Medicine, New York, New York 10029

Progressive accumulation of lipid-laden macrophages is a hallmark of the acid sphingomyelinase (ASM)-deficient forms of Niemann-Pick disease (i.e. Types A and B NPD). To investigate the mechanisms underlying enzyme replacement therapy for this disorder, we studied the uptake of recombinant, human ASM (rhASM) by alveolar macrophages from ASM knock-out (ASMKO) mice. The recombinant enzyme used for these studies was produced in Chinese hamster ovary cells and contained complex type, N-linked oligosaccharides. Binding of radiolabeled, rhASM to the ASMKO macrophages was enhanced as compared with normal macrophages, consistent with their larger size and increased surface area. However, internalization of the enzyme by the ASMKO cells was markedly reduced when compared with normal cells. Studies using receptor-specific ligands to inhibit enzyme uptake revealed that in normal cells rhASM was taken up by a combination of mannose and mannose 6-phosphate receptors (MR and M6PR, respectively), whereas in the ASMKO cells the M6PR had a minimal role in rhASM uptake. Expression of M6PR mRNA was normal in the ASMKO cells, although Western blotting revealed more receptors in these cells when compared with normal. We therefore hypothesized that lipid accumulation in ASMKO macrophages led to abnormalities in M6PR trafficking and/or degradation, resulting in reduced enzyme uptake. Consistent with this hypothesis, we also found that, when rhASM was modified to expose terminal mannose residues and target mannose receptors, the uptake of this modified enzyme form by ASMKO cells was 10-fold greater when compared with the "complex" type rhASM. These findings have important implications for NPD enzyme replacement therapy, particularly in the lung.

Received for publication, August 26, 2003 , and in revised form, October 13, 2003.

^* This work was supported by National Institutes of Health Grant R01-HD-28607. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

^¶ To whom correspondence should be addressed: Dept. of Human Genetics, Box 14-98, Mount Sinai School of Medicine, 1425 Madison Ave., Rm. 14-20A, New York, NY 10029. Tel.: 212-659-6711; Fax: 212-849-2447; E-mail: Edward.Schuchman{at}mssm.edu.

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				C. Garnacho, R. Dhami, E. Simone, T. Dziubla, J. Leferovich, E. H. Schuchman, V. Muzykantov, and S. Muro Delivery of Acid Sphingomyelinase in Normal and Niemann-Pick Disease Mice Using Intercellular Adhesion Molecule-1-Targeted Polymer Nanocarriers J. Pharmacol. Exp. Ther., May 1, 2008; 325(2): 400 - 408. [Abstract] [Full Text] [PDF]