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A peptide-linked recombinant glucocerebrosidase for targeted neuronal delivery: Design, production, and assessment

Paper ID Volume ID Publish Year Pages File Format Full-Text
22716 43373 2016 12 PDF Available
Title
A peptide-linked recombinant glucocerebrosidase for targeted neuronal delivery: Design, production, and assessment
Abstract

•Gaucher's disease is caused by mutations in lysosomal enzyme glucocerebrosidase.•There is currently no treatment for the neuronopathic forms of Gaucher's disease.•Commercially available enzymes do not bind neurons well or cross the BBB.•A rabies-derived peptide and specific linker targeted glucocerebrosidase to neurons.•RDP-IgAh-linked glucocerebrosidase degraded >90% of pathogenic glucosylsphingosine

Although recombinant glucocerebrosidase (GCase) is the standard therapy for the inherited lysosomal storage disease Gaucher's disease (GD), enzyme replacement is not effective when the central nervous system is affected. We created a series of recombinant genes/proteins where GCase was linked to different membrane binding peptides including the Tat peptide, the rabies glycoprotein derived peptide (RDP), the binding domain from tetanus toxin (TTC), and a tetanus like peptide (Tet1). The majority of these proteins were well-expressed in a mammalian producer cell line (HEK 293F). Purified recombinant Tat-GCase and RDP-GCase showed similar GCase protein delivery to a neuronal cell line that genetically lacks the functional enzyme, and greater delivery than control GCase, Cerezyme (Genzyme). This initial result was unexpected based on observations of superior protein delivery to neurons with RDP as a vector. A recombinant protein where a fragment of the flexible hinge region from IgA (IgAh) was introduced between RDP and GCase showed substantially enhanced GCase neuronal delivery (2.5 times over Tat-GCase), suggesting that the original construct resulted in interference with the capacity of RDP to bind neuronal membranes. Extended treatment of these knockout neuronal cells with either Tat-GCase or RDP-IgAh-GCase resulted in an >90% reduction in the lipid substrate glucosylsphingosine, approaching normal levels. Further in vivo studies of RDP-IgAh-GCase as well as Tat-GCase are warranted to assess their potential as treatments for neuronopathic forms of GD. These peptide vectors are especially attractive as they have the potential to carry a protein across the blood–brain barrier, avoiding invasive direct brain delivery.

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Keywords
GCase, glucocerebrosidase; GD, Gaucher’s disease; RDP, rabies-derived peptide; TTC, tetanus toxin fragment C; IgAh, peptide from flexible hinge region of IgA; ERT, enzyme replacement therapy; GlcCer, glucosylceramide; GlcSph, glucosylsphingosine; 4-MUG, 4
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A peptide-linked recombinant glucocerebrosidase for targeted neuronal delivery: Design, production, and assessment
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 221, 10 March 2016, Pages 1–12
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us