The SmartBio System for the Improved Preservation of Human Hematopoietic Stem Cells

摘要 冷冻保存(CP)的最终目标是开发一种能够导致解冻后的生物库建立过程 细胞活性和功能水平与未保存的姐妹种群相同。 临床水平测定人造血干细胞及相关干细胞CP的常用方法 然而，细胞由于解冻后的坏死和凋亡而导致较差的恢复，从而产生劣质细胞 给病人输液的产品。鉴于临床上超过一半或更多的干细胞产品 许多研究表明，今天的试验，包括hHSCs，都是在10%DMSO中冷冻保存的，DMSO是剧毒的 研究了改善CP的方法以及替代低温保护剂(CPA)，如糖 避免已知的DMSO毒性和对HHSC分化的影响。然而，到目前为止，还没有新的 工业上已开发出消除DMSO的HHSC CP工艺。CPSI生物技术假说 必须达到多个里程碑才能产生最佳的HHSC CP工艺 干细胞患者的最佳结局，同时最大限度地减少或消除对DMSO的需求。第一, 必须开发一种独特的高通量冷冻设备，以便HHSC特定的冷冻剖面可以 被定义和执行。其次，必须用可以防止形成的药剂取代二甲基亚砜 以及在CP过程中致命冰的再结晶。第三，必须有独特的干融系统 开发将消除潜在的样品污染，服从文件和大多数 重要的是，可以提供与HHSC匹配的解冻剖面，这可以通过各种阵列实现 把瓶子和袋子放进生物库。最后，解冻后恢复调理剂和输液介质 必须开发出能够防止解冻后延迟性细胞死亡和改善移植的 移植过程中的HHSC。该第二阶段计划旨在解决以下每个里程碑 以下具体目标：SA1a-开发StemFreeze-一种独特的高通量自动冷冻 与当前系统相比，操作员和集装箱具有更大灵活性的系统，从而与HHSC匹配 可以开发冷冻配置文件；SA1b-开发StemThw-SmartThw系统的衍生产品 由母公司第一阶段项目支持，该项目将明确设计用于优化解冻 HHSCs和其他干细胞的过程导致解冻后活性和功能的改善；SA2a- 开发StemCP-一种无毒的CPA冷冻鸡尾酒，含有冰重结晶抑制剂(IRIS)以 更换可添加到任何载体介质中的DMSO，以提高hHSCs的CP 不含DMSO；SA2b-配方StemRevive和StemInsion-解冻后一流 用于改善解冻后细胞凋亡/坏死增加的调理剂和稀释液 干细胞输注效率和SA3-评估干细胞特异性SmartBio平台的优化 体外和体内培养的HSCs的CP。总而言之，这个多组分系统将包括一个干细胞 CPSI SmartBio平台下的特定产品线。总而言之，CPSI假设解决所有 五个挑战将共同简化生物库过程，并推动HHSC CP科学更近一步 到实现相当于非冷冻hHSCs的冷冻保存产品的最终里程碑。 这一成就将改善接受干细胞移植的患者的结局。

Abstract The ultimate goal of cryopreservation (CP) is to develop a biobanking process that results in a post-thaw level of cell viability and function identical to a sister population that has not undergone preservation. Most methods used for the clinical level CP of human hematopoietic stem cells (hHSCs) and related stem cells, however, result in poor recovery due to post-thaw necrosis and apoptosis yielding an inferior cell product which is infused into patients. Given that over half or more of all stem cell products in clinical trials today, including hHSCs, are cryopreserved in 10% DMSO which is highly toxic, many studies have investigated ways to improve CP as well as alternative cryoprotective agents (CPAs) such as sugar alcohols to avoid the known toxicity and impact of DMSO on hHSC differentiation. Yet, to date no new hHSC CP process that eliminates DMSO has been developed commercially. CPSI Biotech hypothesizes that multiple milestones must be achieved to generate the best possible hHSC CP process to yield optimal outcome for the stem cell patient while minimizing or eliminating the requirement for DMSO. First, a unique high throughput freezing device must be developed so that hHSC-specific freezing profiles can be defined and executed. Second, DMSO must be replaced with agents that can prevent the formation and re-crystallization of lethal ice during the CP process. Third, a unique dry thaw system must be developed that will eliminate potential sample contamination, be amenable to documentation and most importantly can provide for hHSC-matched thaw profiles that can be achieved with a diverse array of biobanking vials and bags. Finally, a post-thaw recovery conditioning reagent and transfusion medium must be developed that can prevent post-thaw delayed onset cell death and improve engraftment of hHSC during transplant. This Phase II program is designed to address each of these milestones under the following specific aims: SA1a – Develop StemFreeze – a unique high throughput automated freezing system that has more operator and container flexibility than current systems such that hHSC-matched freezing profiles can be developed; SA1b - Develop StemThaw – a derivative of the SmartThaw system supported by the parent Phase I project that will be expressly designed for optimizing the thawing process of hHSCs and other stem cells resulting in improved post-thaw viability and function; SA2a - Develop StemCP - a non-toxic CPA cryo-cocktail containing Ice Recrystallization Inhibitors (IRIs) to replace DMSO which can be added to any carrier medium of choice to improve CP of hHSCs in the absence of DMSO; SA2b - Formulate StemRevive and StemInfusion – a first in class post-thaw conditioning reagent and dilution media designed to ameliorate post-thaw apoptosis/necrosis increasing stem cell transfusion efficacy and SA3 – Evaluate the stem cell specific SmartBio platform for optimized CP of HSCs in vitro and in vivo. Collectively this multicomponent system will comprise a stem cell specific product line under CPSI’s SmartBio platform. In summary, CPSI hypothesizes that addressing all five challenges together will streamline the biobanking process and move the hHSC CP sciences closer to the ultimate milestone of achieving a cryopreserved product that is equivalent to non-frozen hHSCs. This achievement will result in improved outcome for patients receiving stem cell transplants.