GENE TRANSFER IN LEUKOCYTE ADHERENCE DEFICIENCY

这项提议的目的是利用逆转录病毒载体来指导 外周血白细胞整合素CD 18亚单位的体外表达 血液祖细胞和骨髓细胞来自正常个体和来自 患有白细胞粘附缺陷或LAD的儿童，以及体内骨 来自小鼠的骨髓细胞，其中CD 18已被基因破坏， 面向.患有LAD的儿童会出现复发性、危及生命的 细菌感染，由于白细胞不能从这些 儿童粘附在血管壁上并迁移到 感染这种疾病的临床表现源于 LAD患儿的白细胞不能表达 细胞表面的白细胞整合素CD 11/CD 18分子， CD 18亚基的缺陷。 白细胞粘附缺陷症是人类基因治疗的候选疾病 基于基因转移到造血祖细胞中的治疗， 干细胞：l）疾病在严重的情况下危及生命 缺乏型，2）除骨髓移植外， 支持疗法以外的治疗，3）正常CD 18的转导 基因导入LAD EBV B细胞已被证明可以纠正生物化学和 功能缺陷，和4）基于基因表达的白细胞在 LAD中度缺乏表型，CD 11/CD 18正常水平的5% 表达似乎足以纠正严重的临床 疾病的表现。 目前的研究旨在确定最佳条件， 使用逆转录病毒载体恢复外周血中的CD 18表达 最终目的是从LAD儿童中获得祖细胞和骨髓细胞， 通过将CD 18基因转移到造血干细胞， 干细胞在这些研究中，我们的目标是：1）确定条件 所需的逆转录病毒载体LgCD 18 SN和LgCD 18，编码人 CD 18亚基，以产生高效率的基因转导和基因 外周血祖细胞中CD 18亚单位的表达， 正常人和LAD患儿的骨髓细胞; 2） 为了确定转导的功效、效率和安全性， 人CD 18亚单位的小鼠骨髓细胞中，其中CD 18亚单位 亚基已被基因靶向破坏。 LAD是一种有吸引力人基因治疗模型， 外周血祖细胞和骨髓细胞与CD 18编码 离体载体然后再输注转导的细胞可以 对LAD儿童的主要治疗获益。在急性发作期间， 感染的粒细胞和单核细胞分化， CD 18转导的祖细胞可以运输到炎症部位， 参与宿主对感染因子的反应。因此，在本发明中， 造血干细胞的转导可能不是造血干细胞治疗所必需的。 LAD的临床反应。

The aim of this proposal is to use retroviral vectors to direct the expression of the leukocyte integrin CD18 subunit in vitro in peripheral blood progenitor and bone marrow cells from normal individuals and from children with leukocyte adherence deficiency or LAD, and in vivo in bone marrow cells from mice in which the CD18 has been disrupted by gene targeting. Children with LAD experience recurrent, life-threatening bacterial infections due to the inability of the leukocytes from these children to adhere to the vessel wall and migrate to the site of infection. The clinical manifestations of the disease stem from the inability of leukocytes from children with LAD to express the members of the leukocyte integrin CD11/CD18 molecules on the cell surface due to defects in the CD18 subunit. Leukocyte adherence deficiency is a candidate disease for human gene therapy based upon gene transfer into hematopoietic progenitor cells and stem cells in that: l) the disease is life-threatening in the severe deficiency form, 2) except for bone marrow transplantation there is no treatment other than supportive therapy, 3) transduction of a normal CD18 gene into LAD EBV B-cells has been shown to correct the biochemical and functional defect, and 4) based upon gene expression by leukocytes in the moderate deficiency phenotype of LAD, 5% of normal levels of CD11/CD18 expression appears to be sufficient to correct the severe clinical manifestations of the disease. The current studies are designed to determine the optimal conditions for using retroviral vectors to restore CD18 expression in peripheral blood progenitors and bone marrow cells from LAD children with the ultimate goal of long-term disease correction by CD18 gene transfer into hematopoietic stem cells. In these studies we aim to: 1) determine the conditions required for retroviral vectors LgCD18SN and LgCD18, encoding the human CD18 subunit, to produce high efficiency gene transduction and gene expression of the CD18 subunit in peripheral blood progenitor cells and bone marrow cells from normal individuals and from children with LAD; 2) to determine the efficacy, efficiency, and safety of transduction of the human CD18 subunit into bone marrow cells from mice in whom the CD18 subunit has been disrupted by gene targeting. LAD is an attractive model for human gene therapy in that transduction of peripheral blood progenitor cells and bone marrow cells with CD18-encoding vectors ex vivo followed by re-infusion of the transduced cells may be major therapeutic benefit to children with LAD. During acute episodes of infection the granulocytes and monocytes which differentiate from the CD18-transduced progenitor cells may traffic to the inflammatory site and participate in the host response to the infectious agent. Thus, transduction of a hematopoietic stem cell may not be required for a clinical response in LAD.