Fetal Stem Cell Gene Therapy

描述（改编自应用程序） 越来越多的血液病可以在出生前诊断出来。在 在某些情况下，早期治疗可能有利于胎儿的健康和生存。 子宫内干细胞移植（IUT）或胎儿基因治疗可以治疗 血红蛋白病等疾病。本申请旨在确定 将基因导入胎儿造血干细胞的最佳方法 （HSC）。胎儿HSC比其成人对应物更具增殖性， 因此，假设更容易被转导， 基于鼠白血病病毒或人免疫缺陷病毒的逆转录病毒载体 病毒IUT提供了另一种治疗多种血液病的方法， 产生造血嵌合状态。然而，在没有任何 对于供体HSC的优势，可以通过以下方式实现的嵌合体水平 IUT很低。这限制了这种治疗方法在极少数疾病中的应用。我们的目标 是将IUT的应用扩展到地中海贫血等疾病的治疗 和镰状细胞性贫血，通过改造HSC使其具有增殖优势， 超过正常的HSC。此应用程序将测试是否引入 促红细胞生成素受体（EpoR）进入HSC将使这些改变的细胞 促红细胞生成素（EPO）。这将反过来导致改变 HSC及其后代比正常HSC具有增殖优势 祖先还将检测EpoR的截短形式（tEpoR）。这些tepoR， 在其细胞质的负调控区有缺失 结构域，提供比EpoR更强的增殖信号。的影响 导入EpoR基因对HSC增殖和分化的影响， 它们的祖先后代将使用各种体外培养物 系统.假设EpoR或tEpoR的异位表达 将赋予造血干细胞和早期祖细胞增殖的能力， 对EPO的反应，对这些细胞的分化程序的影响最小， 细胞为了测试HSC上的异位EpoR或tEpoR表达是否可以使这些细胞 比正常的HSC更具竞争力， 在人造血的小鼠模型中针对对照HSC。的能力 表达异位EpoR的HSC在没有或仅在最小程度上不表达EpoR后移植骨髓。 还将测试细胞消融。这些体内实验将进一步 确定使HSC对EPO产生反应是否会对 长期重建和多谱系潜力 HSC。拟议研究的积极成果将有助于发展 基于产生造血细胞的血红蛋白病的治疗 异源嵌合

DESCRIPTION (adapted from the application) A growing number of hematological diseases can be diagnosed before birth. In some cases, early treatment may benefit the health and survival of the fetus. Either in utero stem cell transplantation (IUT) or fetal gene therapy may treat diseases such as the hemaglobinopathies. This application aims to determine the best method for the introduction of genes into fetal hematopoietic stem cells (HSCs). Fetal HSCs are more proliferative than their adult counterparts and are, therefore, hypothesized to be more susceptible to transduction by retroviral vectors based on murine leukemia virus or human immunodeficiency virus. IUT offers another means of curing a number of hematological diseases by generating a state of hematopoietic chimerism. However, in the absence of any advantage for the donor HSCs, the levels of chimerism that can be achieved by IUT are low. This limits the use of this therapy to very few diseases. Our aim is to extend the use of IUT to the treatment of diseases, such as thalassemia and sickle cell anemia, by engineering HSCs to have a proliferative advantage over normal HSCs. This application will test if introduction of the erythropoietin receptor (EpoR) into HSCs will render these altered cells responsive to erythropoietin (EPO). This will in turn result in the altered HSCs and their progeny having a proliferative advantage over normal progenitors. Truncated forms of EpoR (tEpoR) will also be tested. These tEpoR, having deletions in the negative regulatory region of their cytoplasmic domains, deliver stronger proliferative signals than EpoR. The effects of introducing the EpoR genes on the proliferation and differentiation of HSCs and their progenitor progeny will be determined using various in vitro culture systems. It is hypothesized that ectopic expression of either EpoR or tEpoR will confer the ability of HSCs and early progenitors to proliferate in response to EPO with minimal effect on the differentiation program of these cells. To test if ectopic EpoR or tEpoR expression on HSCs can make these cells more competitive than their normal counterparts, modified HSCs will be tested against control HSCs in a mouse model of human hematopoiesis. The ability of HSCs expressing ectopic EpoR to engraft bone marrow after no or only minimal cytoablation will also be tested. These in vivo experiments will further determine if making HSCs responsive to EPO will have any detrimental effect on the long‑term reconstituting‑ and multilineage‑ potential of HSCs. A positive outcome from the proposed studies would aid in developing treatments for hemoglobinopathies based on generating hematopoietic allochimerism.