Engineering iPSC-RBCs for Transfusion

摘要：输血是住院患者最常见的治疗手段。 尽管使用标准捐献者来源的红细胞输血是有效的，但我们相信完全的治疗潜力 输血的数量仍未被开发。最近几个研究小组的研究表明，人类 诱导多能干细胞(IPSCs)可以从少量的外周血中衍生出来，可以 基因组编辑以产生精确的基因变化，并可分化为终末成熟的红细胞 (IPSC-RBCs)，全部在符合cGMP的条件下。虽然IPSC-RBC可能不会取代捐赠者来源的RBC 常规输血在可预见的未来，我们相信IPSC-RBCs已经从基因上 设计以表达新的功能(例如，预防镰状细胞病患者的同种免疫) 可以在目前输血疗法不足的地区找到近期的临床应用。在 合适的治疗利基，工程的IPSC-RBC将是高价值、高影响的产品，可以持续 由于其独特的特点，成本很高。调查小组建议利用最近在以下方面的进展 IPSC和基因组编辑技术，以及他们正在进行的由NHLBI资助的红细胞生物学和 免疫学，追求3个具体目标： (1)优化cGMP级人类外周血细胞的建立、增殖和分化方案 将血液来源的IPSCs转化为成熟红细胞(IPSC-RBCs)； (2A)改造免疫球蛋白干细胞，使所产生的红细胞对多种血型抗原呈阴性，以及(2B) IPSC-RBC输注后体内存活、形态成熟及免疫原性的研究 变成专门的小鼠输血模型；以及 (3)获得FDA Ind和机构IRB批准，并为IPSC进行人自体输血- 研究受者的红细胞存活、成熟和免疫原性。 拟议的研究经过精心设计，以整合最近开发的技术(cGMP IPSC 方法，基因编辑)在Emory(EPIC cGMP血库洁净室，活体)具有专门能力 生物素化红细胞追踪，第一个人类细胞治疗专业知识)，以研究人类IPSC的生物学- 输注给人类受者后衍生的红细胞。成功完成拟议的调查将 开发新的血液产品，以满足输血依赖患者未得到满足的重要临床需求 那些患有镰状细胞病的人。

ABSTRACT: Blood transfusion is the most common therapeutic procedure performed for hospitalized patients. Despite the efficacy of transfusion using standard donor-derived RBCs, we believe the full therapeutic potential of blood transfusion remains untapped. Recent work from several groups has demonstrated that human induced pluripotent stem cells (iPSCs) can be derived from small volumes of peripheral blood, can undergo genome editing to produce precise genetic changes, and can be differentiated into terminally mature RBCs (iPSC-RBCs), all under cGMP-compliant conditions. While iPSC-RBCs may not replace donor-derived RBCs for routine transfusions in the foreseeable future, we believe that iPSC-RBCs that have been genetically engineered to express novel functionalities (eg, prevention of alloimmunization in sickle cell disease patients) could find near term clinical applications in areas where current transfusion therapies are inadequate. In the proper therapeutic niche, engineered iPSC-RBCs would be high-value high-impact products that could sustain high costs due to their unique characteristics. The investigative team proposes to leverage recent advances in iPSC and genomic editing technologies, as well their ongoing NHLBI-funded studies in RBC biology and immunology, to pursue 3 Specific Aims: (1) To optimize cGMP-grade protocols for establishment, propagation, and differentiation of human peripheral blood-derived iPSCs into mature RBCs (iPSC-RBCs); (2A) To engineer iPSCs such that resulting RBCs are negative for multiple blood group antigens, and (2B) To characterize the in vivo survival, morphological maturation, and immunogenicity of iPSC-RBCs when transfused into specialized murine transfusion models; and (3) To obtain FDA IND and institutional IRB approvals, and perform human autologous transfusions of iPSC- RBCs to investigate RBC survival, maturation, and immunogenicity in recipients. The proposed studies have been carefully designed to integrate recently developed techniques (cGMP iPSC methods, gene editing) with specialized capabilities at Emory (EPIC cGMP clean room in the blood bank, in vivo biotinylated RBC tracking, first-in-human cell therapy expertise) to investigate the biology of human iPSC- derived RBCs after transfusion into human recipients. Successful completion of the proposed investigations will lead to novel blood products to meet important unmet clinical needs in transfusion-dependent patients such as those with sickle cell disease.