technologyIn vitro maturation of BMP-7-responsive pancraeatic beta cell progenitors by oxygen modulation

技术通过氧调节使 BMP-7 反应性胰腺 β 细胞祖细胞体外成熟

• 批准号: 9344589
• 负责人: Juan Dominguez-Bendala
• 金额: $ 71.09万
• 依托单位: OPHYSIO, INC.
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9344589
• 关键词: Academia; Administrator; Adult; Alpha Cell; Area; Autoimmune Diabetes; Automobile Driving; Award; BMP7 gene; Beta Cell; Biology; Biotechnology; Cell Differentiation process; Cell Lineage; Cell Maturation; Cell Therapy; Cells; Clinical; Clinical Trials; Collaborations; Contracts; Coupled; Cyclic GMP; Data; Development; Device Designs; Devices; Diabetes Mellitus; Embryo; Endocrine; Enhancement Technology; Evolution; Exhibits; Exposure to; FDA approved; Funding; Generations; Glucose; Growth; Human; Immunity; In Situ; In Vitro; Industry; Insulin; Insulin-Dependent Diabetes Mellitus; Islets of Langerhans Transplantation; Knowledge; Lead; Legal patent; Length; Licensing; Mediating; Methods; Mission; Mus; Natural regeneration; Nature; Non-Insulin-Dependent Diabetes Mellitus; Organ; Organ Donor; Oxygen; Pancreas; Pathway interactions; Patients; Phase; Phenotype; Physiological; Pluripotent Stem Cells; Population; Preparation; Procedures; Process; Production; Progress Reports; Protocols documentation; Publications; Publishing; Reporting; Research; Research Institute; Safety; Source; Stem cells; Techniques; Technology; Testing; Therapeutic; Time; Tissues; Traction; Transplantation; United States National Institutes of Health; Universities; Washington; Work; base; cell type; clinical application; design; drug discovery; effective therapy; experience; experimental study; falls; human embryonic stem cell; human pluripotent stem cell; improved; in vivo; islet; mouse model; novel; phase 2 study; progenitor; prospective; receptor; response; scale up; success; tissue preparation

PROJECT SUMMARY Islet transplantation represents the current cell therapy standard for type 1 diabetes (T1D). However, the gap between the availability of donor organs and the clinical demand for them calls for the development of alternative/renewable sources of insulin-producing cells. In addition to this therapeutic need, a steady supply of islets is also needed for research and drug discovery purposes. Human embryonic stem cells (hESc) differentiated into pancreatic -cell precursors are presently the subject of Phase I/II clinical trials. However, the success of this approach hinges on the assumption that the microenvironment that leads to effective maturation in a mouse model will be the same in human patients with autoimmune diabetes. The safety of partially differentiated hESc-derived products, efficacy of the macro- encapsulation devices used to shield them from allo- and auto-immunity, and lag time to functional maturation remain open questions. The use of insulin-producing cells that are mature and functional at the time of transplantation may circumvent some of these problems. However, despite claims to the contrary, there is no current protocol to date that yields -like cells capable of reversing diabetes right after transplantation. In collaboration with our partners at the University of Miami, Ophysio, Inc. has successfully developed a platform to aid in the terminal in vitro differentiation of pancreatic progenitors (PPs) of different origins (hESc and native murine pancreas). This patented technology is based on the accurate targeting of physiological oxygenation throughout cell aggregates in culture –which conventional means of culture fail to achieve. Oxygen tension lies at the crossroads of key pancreatic differentiation pathways, and its evolution throughout development has been conclusively shown to drive cell fate. Here we seek to extend these principles to the terminal maturation of a novel sub-population of PPs that our collaborators have described in human non-endocrine pancreatic tissue (hNEPT), which comprises 98% of the pancreas and is routinely discarded after islet isolation. This sub-population, identified through in vitro lineage-tracing techniques, is characterized by its responsiveness to the FDA- approved bone morphogenetic protein 7 (BMP-7). hNEPT exposure to BMP-7 results in the efficient (up to 15% in preliminary data) generation of endocrine cells that secrete insulin at levels that fall right within the range published for human isolated islets and exhibit robust glucose responsiveness in vitro and in vivo. Our Phase II studies aim at capitalizing on our Phase I data. These include not only the proof of principle that oxygen modulation improves BMP-7-mediated conversion of hNEPT, but also new findings on the phenotype of BMP-7 responsive cells that will allow for their prospective isolation from raw hNEPT preparations. Our specific aims are: (1) To determine whether in vitro targeting of physiological pO2 in PDX1 (P2RY1)+/ALK3+-sorted hNEPT subpopulations results in functional -like cells capable of reversing diabetes in mice; and (2) To scale up the process using an entire organ (10-12 ml of hNEPT pellet after islet isolation) using Ophysio’s new T75 oxygen-modulating devices (designed in the context of our previous award 2R44DK083832-02). In addition to the optimization and scale up of the process, we will simultaneously establish cGMP manufacturing protocols, file for IP protection of the final method and begin licensing contracts with parties for use of the process to obtain the cells for research purposes. We contend that BMP-7-responsive PPs from hNEPT represent a valid alternative to hESc for clinical applications, as this technology capitalizes on current clinical strategies (islet isolation and transplantation) for which there are already well established networks; increased safety of adult cell products vs. hESc-derived ones; and ease of in vitro expansion/differentiation using a single, FDA-approved clinical product. Coupled with Ophysio’s technology for enhanced in vitro maturation, this approach has rapid translational potential for the treatment of diabetes mellitus.

项目摘要 胰岛移植代表了1型糖尿病（T1D）的当前细胞治疗标准。 但是，供体器官的可用性与对其的临床需求之间的差距 需要开发产生胰岛素细胞的替代/可再生能源。在 除了这种治疗需求外，研究还需要稳定的胰岛供应 药物发现目的。人类胚胎干细胞（HESC）分化为胰腺 细胞前体目前是I/II期临床试验的主题。但是，成功 这种方法取决于这样的假设，即导致有效的微环境 在自身免疫性糖尿病的人类患者中，小鼠模型中的成熟将相同。 部分分化的hESC衍生产品的安全性，宏观的效率 封装设备，用于保护它们免受异源和自动免疫的侵害，并滞后时间 功能性成熟仍然是空旷的问题。使用产生胰岛素的细胞的使用 移植时的成熟和功能可能避免其中一些问题。 但是，目的地要求对比，迄今为止没有当前的协议可以产生 能够在移植后立即逆转糖尿病的细胞。 与我们在迈阿密大学Ophysio，Inc。的合作伙伴合作已成功 开发了一个平台，以帮助胰腺祖细胞的体外分化 （pps）不同起源（hESC和本地鼠胰腺）。该专利技术是 基于整个细胞聚集体的物理充氧的准确靶向 文化 - 哪种传统文化手段无法实现。氧张力位于 关键胰腺分化途径的十字路口及其在整个过程中的发展 发育已被确定地证明可以驱动细胞命运。在这里，我们试图扩展这些 我们的合作者的新型PPS的新型子人群的终末成熟原理 已经在人非内分泌胰腺组织（HNEPT）中描述了，该组织占98％ 胰腺并在胰岛隔离后通常会丢弃。该子人群确定 通过体外谱系追踪技术的特征是其对FDA-的反应性 批准的骨形态发生蛋白7（BMP-7）。暴露于BMP-7导致 有效的（初步数据中最多15％）的内分泌细胞产生，秘密胰岛素 恰好属于人类孤立胰岛和暴露稳健的范围内的水平 体外和体内葡萄糖反应能力。我们的第二阶段研究旨在利用我们的 第一阶段数据。这些不仅包括氧气调节改善的原理证明 BMP-7介导的HNEPT的转化，但也是关于BMP-7表型的新发现 响应细胞将允许其从原始的Hnept制剂中进行前瞻性隔离。 我们的具体目的是：（1）确定PDX1中物理PO2的体外靶向 （p2ry1）+/alk3+分级的Hnept亚群会导致功能性类细胞能够 逆转小鼠的糖尿病； （2）使用整个器官（10-12 mL）扩展过程 使用Ophysio的新型T75氧气调节装置后，胰岛隔离后的Hnept颗粒 （在我们以前的奖项2R44DK083832-02的背景下设计）。除了 优化和扩展过程，我们将简单地建立CGMP 制造协议，最终方法的IP保护文件并开始许可合同 与当事方一起使用该过程来获取用于研究目的的细胞。 我们认为，HNEPT的BMP-7响应PPS代表HESC的有效替代方案 对于临床应用，随着该技术利用当前的临床策略（Islet） 隔离和移植）已经建立了良好的网络；增加 成人细胞产品与hESC衍生产品的安全性；并易于体外 使用单一的FDA批准的临床产品扩展/分化。加上 Ophysio的技术增强了体外成熟的技术，这种方法具有快速的翻译 治疗糖尿病的潜力。