The broad objective of the proposed research is to investigate the pathogenesis of selected animal models of human mucopolysaccharidoses (MPS) and use them to develop and evaluate various therapeutic strategies. Inherent in these studies is the characterization of the molecular pathology of each animal model to determine the degree of homology to the respective human disease at the clinical, pathologic, biochemical, and genetic levels. Since the studies necessary cannot always be done in human patients, these models are an important source of knowledge. The analogues to be studied include: MPS I (Hurler syndrome, alpha-L- iduronidase deficiency), MPS VI (Maroteaux-Lamy syndrome, arylsulfatase B deficiency), and MPS VII (Sly disease, beta- glucuronidase deficiency). In addition, a new model will be created, a double homozygote MPS I/VI (alpha-L- iduronidase/arylusulfase B deficiency), that will allow new investigations into pathogenesis.
The specific aims i nclude: (1) The characterization of the natural history and pathologic manifestations of the animal analogues, in particular the disease of the brain and spinal cord, skeleton, heart, white blood cells, pigment epithelium, and liver; (2) Characterizing the biochemical defects in the MPS homologues will be done by purifying and determining the physicokinetic properties of the normal and abnormal enzymes, developing monoclonal antibodies, and measuring the glycosaminoglycan substrates; (3) Isolating cDNA and/or genomic clones encoding the normal enzymes using synthesized oligonucleotide probes and/or antibodies, collinear sequencing, and cDNA to probe genomic libraries; (4) Characterizing the molecular lesions in the MPS homologues by northern blotting, RNase analysis, and isolating, sequencing, and identifying the MPS mutations; (5) Developing and evaluating therapeutic modalities including: (a) enzyme manipulation therapy using compounds that increase residual enzyme activity in vitro and in vivo; (b) cellular manipulation therapy using bone marrow transplantation; (c) gene therapy using retroviral vectors developed to transfer the appropriate normal cloned gene into cultured fibroblasts, retinal pigment epithelium, and bone marrow cells prior to in vivo transplantation of transfection autologous bone marrow stem cells; (d) evaluate the therapy clinically, biochemically, and pathologically to determine the safety and efficacy of the procedures.
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