The mucopolysaccharidoses (MPS), a major class of lysosomal storage diseases, are a promising target for somatic cell gene therapy. That normal enzyme is synthesized and exported by normal cells and can be taken up by, and can correct, the metabolic defect in affected cells (defined as cross-correction) has been demonstrated in cocultures of affected and normal cells, as well as by bone marrow transplantation in a limited number of human and animal patients. These studies indicate that if a sufficient number of cells producing normal enzyme can be introduced into the patient, significant improvement can be expected. The number of cells need not be large. If the transplanted cells engraft and continue to multiply in sufficient numbers, improvement should be long-term. However, problems associated with histocompatibility limit heterologous transplantation as an approach to therapy. Somatic cell gene therapy is potentially a way of circumventing the problems associated with the immune response, as well as providing a more adequate source of normal enzyme for diverse tissues. In gene therapy studies of a number of different cultured cell types from human and canine patients with MPS VII, we have previously shown that the metabolic defect, including storage of glycosaminoglycans (GAG), is corrected by retroviral transfer of the normal gene for beta- glucuronidase (GUSB). In this proposal, we extend those in vitro studies to therapeutic strategies conducted in vivo in the mouse and canine homologues of MPS VII. These studies will serve as models for correction of MPS diseases by somatic cell gene therapy, and more generally, for the large class of lysosomal storage diseases. The following approaches will be tested: 1) An organoid system using autologous vector-corrected fibroblasts first in MPS VII mice and then dogs to choose long-term, high expression candidate vectors; 2) Autologous bone marrow transplantation in canine MPS VII using vector-mediated transfer to hematopoietic cells; and 3) Gene transfer to hepatocytes in vivo. The effects of the therapeutic strategies will be determined by experiments utilizing littermate controls and will include extensive clinical evaluations, comprehensive light and electron microscopic studies of tissues, enzyme assays, and quantitation of stored substrate in various tissues.
