Development of an automated Umbilical Cord Blood Hematopoietic Stem Cell expansion by endothelial cells for transplantation

 描述(申请人提供)：最初，脐带血移植仅限于儿童，因为输入的细胞剂量较低。在儿科环境中，相关和非相关脐带血移植在治疗各种血液病和代谢性疾病方面都有很高的成功率。近年来，脐带血移植的使用范围扩大了，成人脐带血移植的结果也有所改善，但也存在一些挑战。仅有10%的全球脐带血库存可能是成人和儿童移植的有核细胞总数(TNC)不足。最小的中性粒细胞计数导致患者恢复的严重延迟，使他们容易出现许多致命的并发症。为了充分发挥脐带血对治疗血液系统恶性肿瘤患者的潜力和影响，有必要对脐带血材料进行体外扩增。目前造血干细胞和祖细胞(HSPC)的扩增机制缺乏适当的微环境，这可能是临床试验失败的原因。越来越多的研究小组详细阐述了内皮细胞(ECs)通过分泌大量的生长因子(称为血管分泌因子)在促进HSPC过程中所起的不可或缺的作用。传统的内皮细胞培养机制已不能概括内皮细胞所产生的体内微环境。VeraVec EC平台通过添加Ad5 E4ORF1蛋白消除了这一限制。VeraVec ECs已多次证明了其在共培养条件下以高保真和高可植入性扩增HSPC的能力。临床前数据表明，该平台能够在12天内将UCB扩大1000-1800倍。重要的是，这种扩大的材料保持了HSPC表型的方方面面，包括长期植入、连续移植和与未处理的脐带血相同的多系承诺。然而，HSPC和VeraVec ECs的大规模共培养是劳动密集型的，在后勤方面具有挑战性，并且在符合cGMP的设施中消耗大量试剂。这些障碍使得VeraVec EC平台不太可能被用于HSPC。因此，我们建议将共培养引入TerumoBCT量子中空纤维生物反应器中，以实现自动扩张。最终产品将是自动化、大规模、高保真的扩展，而不会牺牲VeraVec平台的任何好处。

 DESCRIPTION (provided by applicant): Initially, umbilical cord blood (UCB) transplantation was limited to children, given the low cell dose infused. Both related and unrelated cord blood transplants have been performed with high rates of success for a variety of hematologic disorders and metabolic diseases in the pediatric setting. In more recent years the use of UCB transplants has expanded and results for adult umbilical cord blood transplantation have improved, but not without a number of challenges. As little as 10% of the world-wide inventory of UCB may be of insufficient total nucleated cell count (TNC) for both adult and pediatric transplants. The minimal TNC count results in a profound delay in recovery of the patients leaving them susceptible to numerous lethal complications. In order to realize the full potential and impact UCB can have for patients treated for hematological malignancies, ex vivo expansion of the UCB material is necessitated. Current mechanisms for the expansion of Hematopoietic Stem and Progenitor Cells (HSPCs) lack the proper microenvironment, possibly explaining the clinical trial failures. Evidence continues to mount from numerous groups detailing the indispensable role endothelial cells (ECs) have in promoting the HSPC via the secretion of numerous growth factors, termed angiocrine factors. Traditional mechanisms of culturing endothelial cells have not been permissible to recapitulate the in vivo microenvironment generated by ECs. This limitation has been eliminated in ECs by the VeraVec EC platform by the addition of the Ad5 E4ORF1 protein. VeraVec ECs have repeatedly demonstrated their capacity to expand HSPCs with high fidelity and high engraftability in co-culture conditions. Pre-clinical data demonstrates the capacity for the platform to expand UCB 1000-1800 fold over 12 days. Importantly, this expanded material maintains all aspect of the HSPC phenotype including long term engraftment, serial transplant, and multilineage commitment equal to unmanipulated UCB. However, large scale co-cultures of HSPCs and VeraVec ECs are labor intensive, logistically challenging, and consume large quantities of reagents in cGMP compliant facilities. These barriers make the adoption of the VeraVec EC platform for HSPC unlikely. We therefore propose to introduce the co- culture into the TerumoBCT Quantum hollow fiber bioreactor to automate the expansion. The end product will be an automated, large scale, high fidelity expansion without sacrificing any benefits of the VeraVec platform.