Hematopoietic Reconstitution Using Ex Vivo Expanded Cord Blood CD34+ Cells

使用离体扩增的脐带血 CD34 细胞进行造血重建

• 批准号: 8315364
• 负责人: Stephen Edward Fischer
• 金额: $ 28.39万
• 依托单位: ARTERIOCYTE, INC.
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-01 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8315364
• 关键词: Address; Adolescent; Adult; Anemia; Blood Banks; Blood Cells; Bone Marrow; Bone Marrow Stem Cell; CD34 gene; Cell Count; Cell Therapy; Cell Transplantation; Cells; Clinical; Collaborations; Core Facility; Dana-Farber Cancer Institute; Development; Disease; Dose; Engraftment; Evaluation; Flow Cytometry; Frequencies; Germ Cells; Goals; Hematologic Neoplasms; Hematopoiesis; Hematopoietic; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Immune; Immunophenotyping; Incidence; Intention; Knockout Mice; Laboratories; Malignant - descriptor; Medical; Minority; Mus; Non-Malignant; Outcome; Patients; Phase; Phenotype; Population; Process; Production; Protocols documentation; Registries; Research; Source; Southern Blotting; Stem cell transplant; Stem cells; System; Technology; Testing; Time; Transplantation; Umbilical Cord Blood; Umbilical Cord Blood Transplantation; Validation; Work; base; cytokine; density; experience; graft vs host disease; in vivo; innovation; mouse model; nanofiber; novel; peripheral blood; pre-clinical; progenitor; reconstitution; scaffold; self-renewal; stem cell niche; success

DESCRIPTION (provided by applicant): Umbilical cord blood (UCB) provides unique advantages over bone marrow (BM) and mobilized peripheral blood (MPB) as a source of hematopoietic stem cells (HSCs) for transplants, as it allows use of partially HLA-matched units without increasing GvHD incidence. Although UCB contains a higher frequency of primitive HSCs than BM and MPB, it contains significantly fewer total HSCs, which has severely limited its use in adult patients. Arteriocyte plans to address this unmet medical need with rapid, high yield ex vivo expansion of UCB cells using our NANEXTM expansion platform. The NANEXTM platform is based on a nano-fiber scaffold that mimics the bone marrow stem cell niche and has been shown to expand UCB total nucleated cells (TNCs) an average of 500-fold and CD34+ cells and average of 150-fold in 10-day culture. NANEXTM is currently available commercially as a Research Use Only product, but ultimately, Arteriocyte aims to develop this platform as a clinical product for use in umbilical cord blood transplant (UCBT). The objective of this Phase I proposal is to investigate the feasibility of using Arteriocyte's NANEXTM scaffold to expand UCB cells ex vivo as a potential means of overcoming the limited cell supply available from UCB for transplantation. We hypothesize that our NANEXTM scaffold will enable efficient production of high doses of UCB cells capable of reconstituting hematopoietic functions in an immune-compromised mouse model. This hypothesis will be tested in the following specific aims: Specific Aim 1: Optimization of Ex Vivo Expansion of UCB Cells on NANEXTM Ex vivo expansion of UCB cells on NANEXTM will be optimized by varying phenotype of the starting cell population, initial cell seeding density, cytokines, and culture time, and characterizing the frequency of CD34+ cells, colony forming cells (CFCs), and long-term culture-initiating cells (LTC-ICs). The optimized NANEXTM-expansion protocol will then be validated in a laboratory with significant experience with HSC transplantation at Dana-Farber Cancer Institute. Specific Aim 2: Evaluation of Engraftment of NANEXTM-Expanded UCB Cells in NOD/SCID/ 2microglobulin null Mice NANEX"- and TCPS-expanded and non-expanded cells will be transplanted in a NOD/SCD/2microglobulinnull mouse model and the level of engraftment for each test groups will be evaluated using immunophenotyping/flow cytometry and Southern blotting (if necessary). The goal is to demonstrate that NANEX" culture significantly increases the number of SCID repopulating cells (SRCs) compared to controls. If feasibility is demonstrated in this Phase I project, Arteriocyte will pursue further development of the NANEXTM system and evaluation in a clinical setting. For this reason, a team of collaborators has been assembled with significant experience in both pre-clinical and clinical UCBT, and the intention is to work together beyond this project. PUBLIC HEALTH RELEVANCE: Arteriocyte proposes to use our novel NANEXTM ex vivo expansion technology for the rapid, high yield production of cord blood-derived cells capable of long-term repopulation in vivo. We aim to test the feasibility of our approach by grafting NANEXTM-expanded cord blood cells in an immune-compromised mouse model. With further development, the NANEXTM system could enable the broader use of cord blood for restoring normal hematopoiesis in adult patients with hematologic cancers, anemias and a variety of other malignant and non- malignant disorders.

描述（由申请人提供）：脐带血（UCB）作为移植用造血干细胞（HSC）的来源，与骨髓（BM）和动员的外周血（MPB）相比具有独特的优势，因为它允许使用部分HLA匹配的单位，而不会增加GvHD的发生率。虽然UCB含有比BM和MPB更高频率的原始HSC，但其含有显著更少的总HSC，这严重限制了其在成人患者中的使用。Arteriocyte计划使用我们的NANEXTM扩增平台，通过快速、高产的UCB细胞体外扩增来解决这一未满足的医疗需求。NANEXTM平台基于纳米纤维支架，模拟骨髓干细胞生态位，在10天培养中可将UCB总有核细胞（TNCs）平均扩增500倍，将CD 34+细胞平均扩增150倍。NANEXTM目前作为仅供研究使用的产品在市场上销售，但最终，Arteriocyte旨在将该平台开发为用于脐带血移植（UCBT）的临床产品。 该I期提案的目的是研究使用Arteriocyte的NANEXTM支架体外扩增UCB细胞的可行性，作为克服UCB移植可用细胞供应有限的潜在手段。我们假设我们的NANEXTM支架将能够在免疫受损的小鼠模型中有效生产高剂量的UCB细胞，这些细胞能够重建造血功能。将在以下具体目标中检验这一假设：具体目标1：UCB细胞在NANEXTM上的离体扩增的优化UCB细胞在NANEXTM上的离体扩增将通过改变起始细胞群的表型、初始细胞接种密度、细胞因子和培养时间，并表征CD 34+细胞、集落形成细胞（CFC）、和长期培养起始细胞（LTC-IC）。然后，优化的NANEXTM扩增方案将在Dana-Farber癌症研究所具有显著HSC移植经验的实验室中进行验证。具体目标二：NANEXTM扩增的UCB细胞在NOD/SCID/2微球蛋白缺失小鼠中的移植评价将NANEXTM和TCPS扩增的和未扩增的细胞移植到NOD/SCD/2微球蛋白缺失小鼠模型中，并使用免疫表型分析/流式细胞术和Southern印迹法（如果需要）评价每个试验组的移植水平。目的是证明与对照组相比，NANEX培养物显著增加了SCID再增殖细胞（SRC）的数量。 如果在该I期项目中证明了可行性，Arteriocyte将进一步开发NANEXTM系统并在临床环境中进行评估。出于这个原因，已经组建了一个在临床前和临床UCBT方面具有丰富经验的合作者团队，目的是在该项目之外共同努力。 公共卫生相关性：Arteriocyte建议使用我们的新型NANEXTM离体扩增技术，快速、高产地生产能够在体内长期再增殖的脐带血衍生细胞。我们的目标是通过在免疫受损小鼠模型中移植NANEXTM扩增的脐带血细胞来测试我们方法的可行性。随着进一步的发展，NANEXTM系统可以使脐带血在患有血液系统癌症、贫血和各种其他恶性和非恶性疾病的成人患者中恢复正常造血的更广泛的使用。