The objective of this project is to accomplish the requisite preclinical studies for--and to initiate a clinical trial of--hematopoietic stem cell (HSC) gene therapy for Hurler syndrome (mucopolysaccharidosis type I._ Retroviral vectors will be constructed containing the coding sequence (i.e., cDNA) for human alpha-L-iduronidase (IDUA). Potentially- therapeutic vectors will be evaluated in vitro on the basis of optimal transduction frequency and IDUA gene expression (i.e., quantity of IDUA activity) when targeted to lymphoblastoid cell lines (LCL) and to HSC isolated from bone marrow, peripheral blood stem cells, and umbilical cord blood. With the selected vector, allogeneic donor HSC will be transduced ex vivo and then infused into patients with Hurler syndrome concomitant to conventional bone marrow transplantation. Subsequently, patients will be studied for the presence of IDUA cDNA sequence and normal IDUA enzyme activity. In long-term follow-up, this project will study the response to genetically-modified HSC, specifically, determining the level of recombinant IDUA enzyme activity achieved, the duration of expression, and any augmentation of clinical response. The clinical trial will study the response in the central nervous system by serial MRI and psychometric examinations. Unlike any gene therapy study currently in progress, the proposed human trial has the potential to provide clinical efficacy by virtue of substantially increasing the level of IDUA enzyme available to the patient undergoing bone marrow transplantation. Furthermore, it will provide information of more broad significance to disorders causing mental retardation by assessing the potential to provide recombinant protein across the blood-brain barrier. It is anticipated that by year 3 of this project, accomplishments by Project 1 will have demonstrated the capability of transducing non-mitotic HSC which are not efficiently transduced by retroviral vectors and thus less accessible. At that time, this project will exploit novel adeno- associated virus (AAV) vectors and evaluate alternative strategies for more complete HSC gene transfer using this vector system. It is anticipated that such advancements will lead to AAV-based vectors to deliver IDUA cDNA to HSC without requiring prior marrow ablation. Such vectors will be constructed and tested in vitro and, based on these results, will be considered for clinical trials of HSC gene therapy without myeloablative chemotherapy.
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