title: Transduction of fibroblasts and CD34+ progenitors using a selectable retroviral vector containing cDNAs encoding arylsulfatase A and CD24.
abstract: Metachromatic leukodystrophy (MLD) is an autosomal recessive, inherited, lysosomal storage disease caused by a deficiency in arylsulfatase A (ASA). This disease is characterized by progressive demyelination leading to severe neurological symptoms. Allogenic bone marrow transplantation at an early stage of clinical course is only effective treatment currently available. Accordingly the corrective transfer of the ASA gene into hematopoietic stem cells is thought to be an important option for curative treatment for MLD. We have recently developed a selectable vector system based on ex vivo sorting of transduced cells (Migita et al. 1995). In this study, we applied this selectable system for development of MLD gene therapy. A bicistronic retroviral vector containing ASA cDNA and CD24 cDNA as a selectable marker gene was constructed. This vector was successfully transduced on fibroblasts from MLD patients, ASA activity was increased 7-fold compared to normal untransduced cells. PCR Southern analysis of hematopoietic colonies showed that transduction efficiency of CD34+ cells was 11-22%. However, after fluorescence-activated cell sorting using anti-CD24 antibody, 75-100% of colonies became vector positive. The sorting raised the ASA activity several fold compared to untransduced CD34+ progenitors. These results suggest that a bicistronic ASA vector containing a CD24 selectable marker could be a useful component of gene therapy for MLD.
authors.affiliation: Department of Biochemistry and Molecular Bilogy, Nippon Medical School, Tokyo, Japan.
authors.forename: A
authors.initials: A
authors.lastname: Tsutsudaasano
authors.forename: M
authors.initials: M
authors.lastname: Migita
authors.forename: K
authors.initials: K
authors.lastname: Takahashi
authors.forename: T
authors.initials: T
authors.lastname: Shimada
mesh_terms: D015703:Antigens, CD / Q000235:genetics*; D018952:Antigens, CD34 / Q000032:analysis*; D015415:Biomarkers; D051927:CD24 Antigen; D002553:Cerebroside-Sulfatase / Q000172:deficiency / Q000235:genetics*; D018076:DNA, Complementary / Q000235:genetics; D005347:Fibroblasts / Q000378:metabolism*; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000627:therapeutic use*; D006412:Hematopoietic Stem Cells / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D007966:Leukodystrophy, Metachromatic / Q000201:enzymology / Q000235:genetics / Q000628:therapy; D008562:Membrane Glycoproteins; D012190:Retroviridae / Q000235:genetics; D014161:Transduction, Genetic / Q000235:genetics
chemical_list: D015703:Antigens, CD; D018952:Antigens, CD34; D015415:Biomarkers; D051927:CD24 Antigen; C497889:CD24 protein, human; D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D002553:Cerebroside-Sulfatase