CHIMERISM IN CORD BLOOD TRANSPLANTATION IN MICE

描述：（研究者摘要）镰状细胞病（SCD）可以治愈 通过骨髓移植（BMT）;然而，由于发病率和 与手术相关的死亡率，BMT仅用作最后一种 为最严重的病人提供治疗。BMT和/或基因 治疗成为大多数镰状细胞的现实治愈性治疗选择 患者，必须制定毒性较小的准备方案，增加 现有捐赠者的数量，制定基因治疗/移植方案 其中可以使用遗传操作的自体细胞，并确定 预防SCD后遗症所需的混合嵌合体比例。 脐带血（CB）已被用作干细胞的替代来源 为BMT提供了一个独特的干细胞来源， 它的容易和安全的可用性从个人产前诊断为 遗传性疾病，如SCD。目标是提供有效、安全、 用于SCD患者的最低毒性BMT和/或基因疗法。假设是 测试的是：CB移植可以提供稳定的混合嵌合体在非消融 或最小程度消融的受体。具体目标是利用近期 胎儿/新生小鼠血液（FNPB）CB移植模型，我们和 其他人已经发展到：1）确定FNPB细胞的最小数量， 可以在非消融和非移植的患者中提供长期的完全植入或混合嵌合体， 最小程度消融的受体; 2）移植转基因小鼠， 人SCD与正常FNPB干/祖细胞（S/PC），并确定 改善或"治愈"镰状病所需的供体：受体嵌合体比例 细胞后遗症; 3）使用含有人 γ-和β-珠蛋白基因能够长期抑制FNPB S/PC 植入并将这些转导的细胞移植到未消融的和/或 最低程度消融的正常和镰状细胞小鼠。这些研究旨在 定义FNPB S/PC在频率、干细胞 循环动力学、"干性"和基因转导。BMT，包括 竞争性再增殖试验，短期和长期液体培养，以及 将使用甲基纤维素培养物。这些"翻译前"研究 为开发有效的人CB移植和自体移植奠定了基础。 CB基因治疗方案。

DESCRIPTION: (Investigator's abstract) Sickle cell disease (SCD) can be cured by bone marrow transplantation (BMT); however because of the morbidity and mortality associated with the procedure, BMT has been used only as a last resort for the most severely affected patients. In order for BMT and/or gene therapy to become a realistic curative therapeutic option for most sickle cell patients, it is imperative to develop less toxic preparative regimens, increase the number of available donors, develop gene therapy/transplantation protocols in which genetically manipulated autologous cells can be used, and determine the proportion of mixed chimerism required to prevent the sequelae of SCD. Umbilical cord blood (CB) has been used as an alternative source of stem cells for BMT and may provide a unique source of stem cells for gene therapy because of its easy and safe availability from individuals prenatally diagnosed with genetic disorders such as SCD. The goal is to provide effective, safe, and minimally toxic BMT and/or gene therapy for SCD patients. The hypothesis to be tested is: CB transplantation can provide stable mixed chimerism in non-ablated or minimally ablated recipients. The specific aims are to utilize a near-term fetal/newborn mouse blood (FNPB) model of CB transplantation which we and others have developed to: 1) determine the minimum number of FNPB cells which can provide long-term full engraftment or mixed chimerism in non-ablated and minimally ablated recipients; 2) transplant transgenic mice that are models of human SCD with normal FNPB stem/progenitor cells (S/PC) and to determine the proportion of donor:recipient chimerism required to ameliorate or "cure" sickle cell sequelae in these recipients; 3) use retroviral vectors containing human gamma- and beta-globin genes to transduce FNPB S/PC capable of long term engraftment and to transplant these transduced cells into non-ablated and/or minimally ablated normal and sickle cell mice. These studies are designed to define the characteristics of FNPB S/PC with regard to frequency, stem cell cycle kinetics, "stemness," and gene transducibility. BMT, including competitive repopulation assay, short-term and long-term liquid cultures, and methylcellulose cultures will be employed. These "pre-translational" studies form the basis for developing effective human CB transplantation and autologous CB gene therapy protocols.