Exosomen-vermittelte Nanomedizin für die Behandlung von Leukodystrophien
Molekulare Biologie und Physiologie von Nerven- und Gliazellen
Zusammenfassung der Projektergebnisse
Leukodystrophies are fatal inherited myelin diseases and to date there is no efficient treatment available. The fact that they are caused by single gene mutations makes them the ideal model to explore targeted CNS gene therapy strategies. In this study we established mouse models for the two leukodystrophies, Canavan disease (CD) and Hypomyelination with Brain stem and Spinal cord involvement and Leg spasticity (HBSL), in order to test novel gene delivery approaches. Aspa-null mice serve as an accurate model for CD with pathology similar to CD patients. Homozygous Dars-null mice die in utero but the robust pre-pulse inhibition phenotype of heterozygous Darsnull mice, although not resembling the full clinical spectrum, could be utilized as an endpoint for future preclinical therapeutic efficacy studies. For the targeted gene delivery to the CNS we genetically engineered and tested recombinant adeno associated viruses (rAAVs) and exosomes. While exosomes failed to yield sufficient transduction efficiency in vitro, rAAVs equipped with cell-type specific promoters were highly effective in targeting cells of the CNS in vitro as well as in vivo. Transduction with reporter rAAVs carrying the enhanced green fluorescent protein (EGFP) under control of the ubiquitous chicken β-actin (CBA) promoter resulted almost exclusively in neuronal EGFP expression. Swapping the CBA promoter either with the oligodendrocyte specific myelin basic protein (MBP) or myelin-associated glycoprotein (MAG) promoter or the astrocyte specific glial fibrillary acidic protein (GFAP) promoter resulted in targeted transgene expression in oligodendrocytes and astrocytes, respectively. To test the efficacy of rAAV for CD therapy we replaced EGFP with Aspa cDNA under control of the oligodendrocyte specific MBP promoter. Postsymptomatic AAV-MBP-Aspa injection into the brain of Aspa-null mice resulted in widespread ASPA expression throughout the brain and reversed CD pathology in Aspa-null mice significantly by restoring myelination and reducing vacuolization and ventricle dilation. Behavioral analysis of AAV- MBP-Aspa treated Aspa-null mice revealed partial but significant restoration of their locomotor skills. Taken together these results show that rAAVs with cell-type specific promoters are the vector of choice for targeted gene delivery to the brain and that they will be promising tools for the treatment of CD and other leukodystrophies.
Projektbezogene Publikationen (Auswahl)
-
Modeling and treatment of the novel white-matter disorder HBSL. ISN Meeting, Cairns, 2015
Dominik Fröhlich, Ziggy Harrison-Tikisci, Georg von Jonquieres, Andre Bongers, Matthias Klugmann
-
Modeling and treatment of the novel white-matter disorder HBSL. ISN Myelin Satellite Meeting, Fitzroy Island, 2015
Dominik Fröhlich, Ziggy Harrison-Tikisci, Georg von Jonquieres, Andre Bongers, Matthias Klugmann
-
(2016). "In vivo characterization of the aspartyl-tRNA synthetase DARS: Homing in on the leukodystrophy HBSL." Neurobiol Dis. 97:24-35
Fröhlich, D., Suchowerska, A. K., Spencer, Z. H., von Jonquieres, G., Klugmann, C. B., Bongers, A., Delerue, F., Stefen, H., Ittner, L. M., Fath, T., Housley, G. D. and Klugmann, M.
-
(2016). "Recombinant Human Myelin-Associated Glycoprotein Promoter Drives Selective AAV-Mediated Transgene Expression in Oligodendrocytes." Front Mol Neurosci 9: 13
von Jonquieres, G., Fröhlich, D., Klugmann, C. B., Wen, X., Harasta, A. E., Ramkumar, R., Spencer, Z. H., Housley, G. D. and Klugmann, M.
