Stem Cell Therapies for Primary Immune Deficiency

原发性免疫缺陷的干细胞疗法

• 批准号: 7347251
• 负责人: Gay M Crooks
• 金额: $ 127.56万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-18 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7347251
• 关键词: Stem; Cell; Therapies; Primary; Immune

The overall goal of this Program is to develop novel stem cell transplantation and gene therapy approaches to produce an intact immune system in patients with Severe Combined Immune Deficiency (SCID) and other Primary Immune Deficiencies. The manipulation of stem and progenitor cells in the profoundly immune deficient patient with SCID provides a unique biological and clinical setting to unravel mechanisms of stem cell engraftment, lymphopoiesis and immune reconstitution. The central hypothesis of the Program is that the age of the hematopoietic graft and the host environment influences the ultimate reconstitution of the immune system in the patient with SCID. Each of the Projects will test this hypothesis from a different and complementary perspective. The goal of Project 1 (Crooks) is to delineate the role of the thymic vascular niche in the regulation of homing to the neonatal thymus after bone marrow transplantation (BMT). Project 2 (Dorshkind) will provide new information regarding fetal B cell development and the development of the humoral immune response, with a specific focus on the ontogeny of B-1 B cells. This information will be highly relevant to understanding the restoration of humoral immunity following BMT and gene therapy. The Program brings together an experienced and collaborative team of investigators from UCLA, with interactions facilitated through an Administrative Core. Two scientific cores will support the Projects by providing specialized expertise in cell and tissue isolation and analysis, and in animal models of transplantation. These Projects and Cores will provide synergy and focus for basic and clinical studies in Stem Cell therapies for SCID, the most lethal form of Primary Immune Deficiency. Project 1: Thymic Reconstitution and the Vascular Niche (Gay Crooks, M.D.) Revised Abstract Section (No Change) Studies of age-related changes in thymic function and reconstitution after bone marrow transplant (BMT) have focused on the epithelial component of the thymic microenvironment; little attention has been paid to the influence of the endothelial compartment of the thymus in this process. The goal of this proposal is to delineate the role of the thymic vascular niche in the regulation of homing of bone marrow cells to thymus after transplantation. Clinical data from patients with Severe Combined Immune Deficiency (SCID) and our own data in experimental immune deficient murine models, show that when non-conditioned BMT is performed specifically in the neonatal period, rapid thymic engraftment results in donor lymphopoiesis, with little or no contribution of donor cells to non-lymphoid lineages. These findings demonstrate that although microenvironmental signals for T cell differentiation exist throughout postnatal life, different host mechanisms for thymic engraftment may exist in neonatal and adult hosts. We propose that in the neonatal setting, bone marrow can home to the thymus directly without the need for an intermediate stage of marrow engraftment. We will use the striking dichotomy between neonatal and adult thymic engraftment patterns to define the endothelial mechanisms regulating these differences. Data from murine allogeneic models will be translated to the human setting by testing our findings in a similar xenogeneic BMT model and by analysis of human thymus samples at different stages of postnatal development. Project 2: Identification of Human Cord Blood B Lineage Cells (Kenneth Dorshkind, Ph.D.) Revised Abstract Section Studies of murine B cell development have demonstrated the existence of two distinct waves of cell production. The first initiates during embryogenesis and results in the production of progenitors destined to generate B-1 B cells that are part of the innate immune system and which are particularly efficient at recognizing encapsulated bacteria such as S. pneumoniae. The second major wave generates B-2 B cells that are produced in the bone marrow throughout life and function in adaptive immunity. The central hypothesis of this application continues to be that successful reconstitution of humoral immunity in humans following bone marrow transplantation is dependent upon the recapitulation of these two distinct ontogenic programs. However, other than the fact that B cells emerge in the embryo, nothing is known about human fetal B cell development. Studies to characterize novel human cord blood progenitors that include an intermediate that co-expresses B and myeloid lineage determinants will be performed. The information that will be obtained is relevant to restoration of humoral immunity following bone marrow transplantation, the treatment of immunodeficiency diseases, and the recovery of the immune response following various myeloablative treatments. Administrative Core A Gay Crooks Revised Abstract Section Dr Gay Crooks is the Principal Investigator of the Program Project Grant and Director of the Administrative Core. The Administrative Core will be an essential mechanism to enable all the scientific components of the Program to interact by providing the structure and financial support for regular planning and evaluation of the research. Regular communication between investigators will be critical for the success of the Program. The Core will be responsible for ensuring that the investigators act as a cohesive and interactive group by providing the planning and leadership required for regular and frequent interactions between projects and cores. The Administrative Core will also be responsible for supervising budgetary and regulatory aspects of the Program, and providing the logistical support for transporting samples between institutions. Program interactions and oversight will be facilitated by both an Internal Advisory Board and an External Advisory Board composed of experts in the fields of Primary Immune Deficiency, Bone marrow Transplantation and basic immuno-biology. Core B: Cell Isolation and Analysis Core Director: Linda Baum, MD, PhD Revised Abstract Section The Cell Isolation and Analysis Core (Core B) will consist of two components, flow cytometry and histology, and will provide technical and advisory support for both Projects. The Core will aid investigators in experimental design for cell isolation and in analysis of flow cytometry data from murine and human samples. Core B will be administered by Dr Linda Baum. The core will provide FACS analysis and sorting and preparation of tissue sections and interpretation of specific tissue samples. FACSAria, and BD LSRII flow cytometry machines will be available for cell isolation and analysis. The histology component will support Projects 1 and 2 by preparing whole mounts and tissue sections by cryostat and microtome, and performing staining with standard or immunofluorescent techniques. Core C: Animal Core Director: Donald B. Kohn, MD Revised Abstract Section The focus of the Animal Core Laboratory (ACL) is to provide immunodeficient mice for the Program projects. Chimeric mouse human hybrids have become an important research tool for the evaluation of reconstitution and differentiation potential of hematopoietic and immunologic progenitor populations. The ACL will provide the following services for the projects in the Program: 1) establishment and maintenance of breeding colonies of Immunodeficient mice 3) provision of pups and adult for experiments 3) assistance with transplantation of immune deficient mice with human cells 4) health monitoring and day-to-day care of immune deficient mice, and 5) training of research personnel on the care and techniques used with immunodeficient mice. The Core will be housed within the animal care facility at UCLA under the direction of Dr. Donald Kohn.

该计划的总体目标是开发新型干细胞移植和基因治疗方法，为患有严重联合免疫缺陷（SCID）和其他原发性免疫缺陷的患者产生完整的免疫系统。对严重免疫缺陷 SCID 患者的干细胞和祖细胞进行操作，为揭示干细胞植入、淋巴细胞生成和免疫重建的机制提供了独特的生物学和临床环境。该计划的中心假设是造血移植物的年龄和宿主环境影响 SCID 患者免疫系统的最终重建。每个项目都将从不同且互补的角度检验这一假设。项目 1（Crooks）的目标是描述胸腺血管生态位在骨髓移植（BMT）后新生儿胸腺归巢调节中的作用。项目 2 (Dorshkind) 将提供有关胎儿 B 细胞发育和体液免疫反应发育的新信息，特别关注 B-1 B 细胞的个体发育。这些信息对于了解 BMT 和基因治疗后体液免疫的恢复高度相关。该计划汇集了来自加州大学洛杉矶分校的经验丰富且协作的研究团队，并通过行政核心促进互动。两个科学核心将通过提供细胞和组织分离和分析以及移植动物模型方面的专业知识来支持这些项目。这些项目和核心将为 SCID（最致命的原发性免疫缺陷形式）干细胞疗法的基础和临床研究提供协同作用和重点。 项目 1：胸腺重建和血管生态位（Gay Crooks，医学博士） 修订后的摘要部分 （无变化） 与年龄相关的胸腺功能变化和骨髓移植（BMT）后重建的研究重点关注胸腺微环境的上皮成分。很少有人关注胸腺内皮室在此过程中的影响。该提案的目标是描绘 胸腺血管生态位在移植后调节骨髓细胞归巢中的作用。严重联合免疫缺陷 (SCID) 患者的临床数据和我们自己在实验性免疫缺陷小鼠模型中的数据表明，当专门在新生儿期进行非条件 BMT 时，快速胸腺植入会导致供体淋巴细胞生成，几乎没有或没有供体细胞对非淋巴谱系的贡献。这些发现表明，尽管 T 细胞分化的微环境信号在整个出生后生命中都存在，但新生儿和成人宿主中可能存在不同的胸腺植入机制。我们建议，在新生儿环境中，骨髓可以直接归巢到胸腺，而不需要骨髓移植的中间阶段。我们将利用新生儿和成人胸腺植入模式之间的显着二分法来定义 调节这些差异的内皮机制。通过在类似的异种 BMT 模型中测试我们的发现以及通过分析出生后发育不同阶段的人类胸腺样本，来自小鼠同种异体模型的数据将被转化为人类环境。 项目 2：人类脐带血 B 谱系细胞的鉴定（Kenneth Dorshkind 博士） 修订后的摘要部分 对小鼠 B 细胞发育的研究表明存在两种不同的细胞产生波。第一个过程在胚胎发生期间启动，导致祖细胞的产生，这些祖细胞注定会产生 B-1 B 细胞，这些细胞是先天免疫系统的一部分，并且在识别封装的免疫球蛋白方面特别有效。 细菌，例如肺炎链球菌。第二个主要波产生 B-2 B 细胞，这些细胞在整个生命过程中在骨髓中产生并在适应性免疫中发挥作用。本申请的中心假设仍然是，骨髓移植后人体体液免疫的成功重建取决于这两种不同的个体发生程序的重演。然而，除了 B 细胞在胚胎中出现这一事实之外，我们对人类胎儿 B 细胞的发育一无所知。将进行研究来表征新型人类脐带血祖细胞，其中包括共表达 B 和骨髓谱系决定簇的中间体。将获得的信息与骨髓移植后体液免疫的恢复、免疫缺陷疾病的治疗以及各种清髓治疗后免疫反应的恢复有关。 行政核心A 同性恋骗子 修订后的摘要部分 盖伊·克鲁克斯博士是该计划项目资助的首席研究员和行政核心主任。行政核心将成为一个重要机制，通过为研究的定期规划和评估提供结构和财政支持，使该计划的所有科学组成部分能够相互作用。研究人员之间的定期沟通对于该计划的成功至关重要。核心将负责通过提供项目和核心之间定期和频繁互动所需的规划和领导，确保调查人员成为一个有凝聚力和互动的团队。行政核心还将负责监督该计划的预算和监管方面，并为机构之间运输样本提供后勤支持。由原发性免疫缺陷、骨髓移植和基础免疫生物学领域的专家组成的内部咨询委员会和外部咨询委员会将促进项目互动和监督。 核心 B：细胞分离和分析核心 主任：琳达·鲍姆，医学博士、哲学博士 修订后的摘要部分 细胞分离和分析核心（核心 B）将由流式细胞术和组织学两个部分组成，并将为这两个项目提供技术和咨询支持。该核心将帮助研究人员进行细胞分离的实验设计以及来自小鼠和人类样本的流式细胞术数据分析。核心 B 将由 Linda Baum 博士负责管理。核心将提供FACS分析和排序以及 组织切片的制备和特定组织样本的解释。 FACSAria 和 BD LSRII 流式细胞仪将可用于细胞分离和分析。组织学部分将通过低温恒温器和切片机制备整体切片和组织切片，并使用标准或免疫荧光技术进行染色来支持项目 1 和 2。 核心C：动物核心 主任：医学博士唐纳德·B·科恩 (Donald B. Kohn) 修订后的摘要部分 动物核心实验室（ACL）的重点是为该计划项目提供免疫缺陷小鼠。嵌合小鼠人类杂交体已成为评估造血和免疫祖细胞群的重建和分化潜力的重要研究工具。 ACL 将为该计划中的项目提供以下服务： 1) 建立和维护免疫缺陷小鼠繁殖群 3) 提供幼仔和成年小鼠进行实验 3) 协助免疫缺陷小鼠移植人类细胞 4) 健康监测免疫缺陷小鼠的日常护理，以及 5) 对研究人员进行免疫缺陷小鼠护理和技术培训。该核心将在唐纳德·科恩博士的指导下安置在加州大学洛杉矶分校的动物护理设施内。