Evaluation of Pluripotent Stem Cell-Derived Blood Cells in Nonhuman Primate Model

描述(申请人提供)：异基因造血干细胞和祖细胞移植(HSPC)具有治愈血液病的潜力。然而，许多患者没有匹配的供者，移植物抗宿主病是一个重要的问题。自体患者的HSPC可以通过基因矫正来治愈疾病，但自体HSPC的低产量和体外操作会导致“茎”的丧失，从而减少植入率。因此，从患者特定的诱导多能干细胞(IPSCs)中生产HSPC将解决这些问题，并代表着一个无限的细胞来源。为了促进IPSC治疗药物的临床翻译，我们提出了一种新的策略来扩大IPSC来源的HSPC用于造血移植。具体地说，我们建议在临床相关的非人灵长类动物模型中利用内皮细胞(ECs)从猪尾猕猴(MN)IPSCs中产生、扩增和植入假定的HSPC。在与Shahin Rafii博士的合作中，我们开发了一个有效的新平台，通过与Akt激活的人内皮细胞(E4+ECs)共培养，将猕猴CD34+LT-HSCs扩增至25倍。我们最近发现，在E4+ECs上扩增的IPSC-HSPC在NSG小鼠体内的植入率很高(高达50%的CD45+细胞)。这一证据证实了我们通过与E4+ECs特有的血管分泌/造血信号直接接触培养来改变IPSC-HSPC生物学的新方法。这项拟议的研究将把这些发现转化为非人类灵长类动物，从而为生产具有造血重建和纠正遗传疾病能力的IPSC-HSPC迈出了重要的一步。

DESCRIPTION (provided by applicant): Allogeneic hematopoietic stem and progenitor cell (HSPC) transplantation has the potential to cure hematologic disease. However, many patients do not have an HLA matched donor, and graft-versus-host disease is a significant problem. Autologous patient HSPCs can be genetically corrected to cure the disease, but low yields of autologous HSPCs and ex vivo manipulation cause a loss of "stemness" leading to reduced engraftment. Thus, HSPC production from patient-specific induced pluripotent stem cells (iPSCs) would solve these problems and represent an unlimited cell source. To advance clinical translation of iPSC therapeutics, we propose a novel strategy to expand iPSC-derived HSPCs for hematopoietic transplantation. Specifically, we propose to engineer endothelial cells (ECs) for generation, expansion, and engraftment of putative HSPCs from pigtail macaque (Mn)iPSCs in the clinically relevant nonhuman primate model. In a promising collaboration with Dr. Shahin Rafii, we developed an effective, novel platform to expand macaque CD34+ LT- HSCs up to 25-fold by co-culture with Akt-activated human endothelial cells (E4+ECs). We recently found that iPSC-HSPCs expanded on E4+ECs have high levels of engraftment in NSG mice (up to 50% CD45+ cells). This evidence substantiates our novel approach to alter iPSC-HSPC biology through direct contact culture with angiocrine/hematopoietic signals unique to E4+ECs. The proposed studies will translate these findings to nonhuman primates and thus provide a major step toward producing iPSC-HSPCs with the capacity for hematopoietic reconstitution and correction of genetic diseases.