Abstract
Mucopolysaccharidosis (MPS) type II (Hunter's syndrome) is caused by mutations in the iduronate 2-sulfatase (IDS) fusion protein. MPS-II affects the brain, and enzyme replacement therapy is not effective in the brain, because the enzyme does not cross the blood-brain barrier. To treat mouse models of MPS-II with brain-penetrating IDS, the lysosomal enzyme was reengineered as an IgG-IDS fusion protein. The mature human IDS was fused to the carboxyl terminus of both heavy chains of the chimeric monoclonal antibody (MAb) against the mouse transferrin receptor (TfR), and the fusion protein is designated cTfRMAb-IDS. The purity and identity of the fusion protein was confirmed by electrophoresis and Western blotting with antibodies to mouse IgG and human IDS. The EC50 of binding of the cTfRMAb-IDS fusion protein to the mouse TfR (0.85 ± 0.15 nM) was comparable to the EC50 of binding of the cTfRMAb (0.78 ± 0.05 nM). The IDS enzyme activity of the cTfRMAb-IDS fusion protein was 126 ± 1 nmol · h−1 · μg−1 protein. After intravenous injection in the mouse, the cTfRMAb-IDS fusion protein was rapidly removed from plasma and distributed to tissues, including brain and spinal cord. The uptake of the fusion protein by brain or spinal cord was 1.3 ± 0.1 and 2.2 ± 0.2% injected dose/g, respectively, which is 100-fold greater than the brain uptake of IDS alone. This work shows that a lysosomal sulfatase can be reengineered as an IgG-enzyme fusion protein that rapidly penetrates the brain after intravenous administration.
Footnotes
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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ABBREVIATIONS:
- MPS
- mucopolysaccharidosis
- IDS
- iduronate 2-sulfatase
- BBB
- blood-brain barrier
- MAb
- monoclonal antibody
- HIR
- human insulin receptor
- HIRMAb
- engineered MAb against the HIR
- TfR
- transferrin receptor
- cTfRMAb
- chimeric MAb against the mouse TfR
- IDUA
- iduronidase
- SUMF1
- sulfatase modifying factor 1
- RT
- reverse transcription
- ODN
- oligodeoxynucleotide
- TV
- tandem vector
- LC
- light chain
- orf
- open reading frame
- PCR
- polymerase chain reaction
- CM
- conditioned medium
- ELISA
- enzyme-linked immunosorbent assay
- PAGE
- polyacrylamide gel electrophoresis
- CHO
- Chinese hamster ovary
- ECD
- extracellular domain
- DMEM
- Dulbecco's modified Eagle's medium
- TCA
- trichloroacetic acid
- VD
- volume of distribution
- ID
- injected dose
- AUC
- area under the curve
- PS
- permeability-surface area
- AA
- amino acid
- HC
- heavy chain.
- Received September 19, 2011.
- Accepted November 7, 2011.
- Copyright © 2012 by The American Society for Pharmacology and Experimental Therapeutics
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