The lysosomal storage disease fucosidosis: towards enzyme replacement therapy

Wolf H (2016)
Bielefeld: Universität Bielefeld.

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Bielefeld Dissertation | English
Abstract
Fucosidosis is a rare lysosomal storage disease caused by the deficiency of the lysosomal glycosidase a-L-fucosidase resulting in the accumulation of fucosylated glycocompounds. Human fucosidosis patients are mainly characterized by progressive mental retardation and neurological deterioration leading to premature death. In this study, the generation of a constitutive fucosidosis mouse model is reported. The gene for the lysosomal a-L-fucosidase (Fuca1) was disrupted by gene targeting resulting in the complete absence of a-L-fucosidase activity in Fuca1(-/-) mice. The phenotype of the mouse model closely resembled that of a milder form of the human disease and due to animal welfare, the Fuca1(-/-) mice had to be euthanized at 9 - 11 months of age. A lysosomal storage pathology was detected in nearly all investigated organs of the mouse model, e. g. in liver, kidney, spleen as well as in the CNS, and was characterized by foam-like storage vacuoles to variable degree. An immense accumulation of water-soluble fucosylated glycocompounds was demonstrated and the glycoasparagine Asn-GlcNAc-Fuc was identified as main storage material in kidney and brain and was also excreted with the urine of the Fuca1(-/-) mice. The neuropathological alterations of the mouse model were analyzed in more detail, as these are the leading symptoms in human fucosidosis. The Fuca1(-/-) mice exhibited neuroinflammatory signs including prominent micro- and astrogliosis and suffered from progressive loss of Purkinje cells. Particularly the latter might contribute to behavioral abnormalities of the animals including progressive coordinatory and motor deficits as well as a reduced overall activity and cognitive impairment.

In order to enable the development of an enzyme replacement therapy (ERT) for the fucosidosis disease, an expression system for the production of the human a-L-fucosidase was generated in CHO-K1 cells. While a purification strategy for the His6-tagged enzyme was established using a Ni2+-affinity chromatography followed by a strong cation exchange chromatography, the purification of the untagged a-L-fucosidase was more challenging and needs further efforts. Finally, a Fuca1(-/-) mouse was intravenously treated with purified His6-tagged enzyme resulting in an efficient uptake of the recombinant a-L-fucosidase into visceral organs and in a complete clearance of storage material in the spleen. These preliminary results provide promising data with regard to develop an ERT also for human fucosidosis patients.
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Wolf H. The lysosomal storage disease fucosidosis: towards enzyme replacement therapy. Bielefeld: Universität Bielefeld; 2016.
Wolf, H. (2016). The lysosomal storage disease fucosidosis: towards enzyme replacement therapy. Bielefeld: Universität Bielefeld.
Wolf, H. (2016). The lysosomal storage disease fucosidosis: towards enzyme replacement therapy. Bielefeld: Universität Bielefeld.
Wolf, H., 2016. The lysosomal storage disease fucosidosis: towards enzyme replacement therapy, Bielefeld: Universität Bielefeld.
H. Wolf, The lysosomal storage disease fucosidosis: towards enzyme replacement therapy, Bielefeld: Universität Bielefeld, 2016.
Wolf, H.: The lysosomal storage disease fucosidosis: towards enzyme replacement therapy. Universität Bielefeld, Bielefeld (2016).
Wolf, Heike. The lysosomal storage disease fucosidosis: towards enzyme replacement therapy. Bielefeld: Universität Bielefeld, 2016.
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