In Vivo Selection of Transduced Hematopoietic Stem Cells

镰状细胞病和其他血液疾病的基因治疗策略必须解决两个关键挑战：1)实现治疗所需的足够数量的校正血细胞；2)建立可接受的风险/效益比，特别是关于插入性突变的风险。我们已经开发了体内选择转导造血干细胞（hsc）的系统，以增加治疗水平的数量，基于耐药基因对细胞毒性药物治疗的保护能力。我们现在建议在大型动物模型中进一步测试这种方法，与常规模型不同，它更准确

Strategies for gene therapy of sickle cell disease and other blood disorders must address 2 key challenges: 1) achievement of a therapeutically adequate number of corrected blood cells 2) establishment of an acceptable risk/benefit ratio, partieularly regarding the risk of insertional mutagenesis. We have developed systems for in vivo selection of transduced hematopoietic stem eells (HSCs) to increase the number to therapeutic levels, based on the ability of drug-resistance genes to protect against treatment with cytotoxic drugs. We now propose to further test this approach in a large animal model, which unlike routine models, accurately simulates clinical applications. We will also explore new ways of using the HOXB4 transcription factor to augment HSC selection. Given the ability of HOXB4 to augment HSC self-renewal, we have developed strategies to regulate HOXB4 function, and to co-express this gene with a drug resistance gene. In terms of the safety, the field currently lacks a proven in vivo assay system for studying vector-induced transformation. Experiments in this proposal will develop a model using mice bearing pre-oncogenie lesions, and will test a number of specific hypotheses regarding the role of the vector transgene, the viral regulatory sequences within the vector, and the role of immunodefieicney in vector-induced T cell malignancies. Overall, these studies should lead to a safe and effective HSC selection system for use in hemoglobin disorders, and address important questions broadly relevant to the field ofhematopoietic gene therapy.