Knock-out mouse for Canavan disease: a model for gene transfer to the central nervous system

• Published in: The journal of gene medicine 2000-05, Vol.2 (3), p.165-175
• Main Authors: Matalon, Reuben, Rady, Peter L., Platt, Kenneth A., Skinner, Henry B., Quast, Michael J., Campbell, Gerald A., Matalon, Kimberlee, Ceci, Jeffrey D., Tyring, Stephen K., Nehls, Michael, Surendran, Sankar, Wei, Jingna, Ezell, Ed L., Szucs, Sylvia
• Format: Article
• Language: English
• Subjects: Amidohydrolases - genetics; Animals; aspartoacylase deficiency; Brain - abnormalities; Brain - pathology; Canavan disease; Canavan Disease - genetics; Canavan Disease - therapy; Central Nervous System - metabolism; central nervous system model; Cloning, Molecular; Disease Models, Animal; Gene therapy; Gene Transfer Techniques; Genetic Therapy; knock-out mouse; Magnetic Resonance Imaging; Mice; Mice, Knockout; N-acetylaspartic acid (NAA); Phenotype; spongy degeneration of the brain
• ISSN: 1099-498X; 1521-2254
• DOI: 10.1002/(SICI)1521-2254(200005/06)2:3<165::AID-JGM107>3.0.CO;2-R

Background Canavan disease (CD) is an autosomal recessive leukodystrophy characterized by deficiency of aspartoacylase (ASPA) and increased levels of N‐acetylaspartic acid (NAA) in brain and body fluids, severe mental retardation and early death. Gene therapy has been attempted in a number of children with CD. The lack of an animal model has been a limiting factor in developing vectors for the treatment of CD. This paper reports the successful creation of a knock‐out mouse for Canavan disease that can be used for gene transfer. Methods Genomic library λ knock‐out shuttle (λKOS) was screened and a specific pKOS/Aspa clone was isolated and used to create a plasmid with 10 base pair (bp) deletion of exon four of the murine aspa. Following linearization, the plasmid was electroporated to ES cells. Correctly targeted ES clones were identified following positive and negative selection and confirmed by Southern analysis. Chimeras were generated by injection of ES cells to blastocysts. Germ line transmission was achieved by the birth of heterozygous mice as confirmed by Southern analysis. Results Heterozygous mice born following these experiments have no overt phenotype. The homozygous mice display neurological impairment, macrocephaly, generalized white matter disease, deficient ASPA activity and high levels of NAA in urine. Magnetic resonance imaging (MRI) and spectroscopy (MRS) of the brain of the homozygous mice show white matter changes characteristic of Canavan disease and elevated NAA levels. Conclusion The newly created ASPA deficient mouse establishes an important animal model of Canavan disease. This model should be useful for developing gene transfer vectors to treat Canavan disease. Vectors for the central nervous system (CNS) and modulation of NAA levels in the brain should further add to the understanding of the pathophysiology of Canavan disease. Data generated from this animal model will be useful for developing strategies for gene therapy in other neurodegenerative diseases. Copyright © 2000 John Wiley & Sons, Ltd.