Testing the Therapeutic Potential of iPS Cells for Inherited Skin Diseases

项目摘要 大疱性表皮病（EB）是一组罕见的遗传性水疱， 这些疾病可能是毁灭性的，在某些情况下是致命的。这项技术突破让成年人 将皮肤细胞重新编程为诱导多能干细胞（iPSC）现在提供了一种可能性， 开发EB的永久性矫正疗法。治疗方法依赖于产生 患者特异性的iPSC，然后经过遗传校正和分化成皮肤细胞， 移植从需要治疗的同一患者产生iPSC不仅可能 避免了免疫排斥的并发症，但也提供了无限和可扩展的患者来源， 适合移植的特定细胞。对于由母公司R 01资助的初始研究，我们专注于 为患有EB的Dowling-Meara亚型的患者队列开发基于iPSC的疗法 单纯型（EBS-DM），他们在角蛋白14（Krt 14）蛋白中共享相同的R125 C“热点突变”， 导致70%的EBS-DM病例。这项EBS-DM研究使我们能够解决所有主要的 这是开发临床相关的基于iPSC的EBS疗法所需的障碍和先决条件。 具体来说，我们已经开发了一种新的，有效的方法，用于产生临床相关的整合- 免费的iPSCs我们还使用了非整合选择标记的基因编辑技术， 纠正从EBS患者产生的iPSC中的遗传缺陷。此外，我们的高效率 最近，重编程使我们能够将基因编辑和EBS成纤维细胞的重编程结合起来，形成一个联合收割机。 一步程序。此次修订申请提出扩大一步组合基因编辑， 重编程的方法，以更严重的形式EB，隐性营养不良EB（RDEB），并推进我们的 临床试验的结果。鉴于需要几个团体之间的协作努力， 为了确保将复杂的iPSC疗法成功转化为临床，我们最近进入了 与斯坦福大学和哥伦比亚大学结成联盟， 用于RDEB处理的遗传校正的iPSC衍生的细胞产物的制造方案。因此，我们建议， 该提议的目的1是提高基于iPSC的电子设备的安全性并降低其制造复杂性。 通过将重编程和基因编辑结合到一步程序中来治疗RDEB;目标2和3是 生产符合cGMP标准的重编程和基因编辑试剂， 为FDA生成研究性新药（IND）支持数据的联盟。EB iPS细胞 联盟成功获得RDEB临床试验的批准将扩大治疗潜力 用于治疗影响其他内脏器官的各种单基因疾病，其中 由于难以监测基于iPSC的治疗的不良反应，它们不太可能成为第一目标。

Project Summary No effective treatments are available for epidermolysis bullosa (EB), a group of rare inherited blistering disorders that can be devastating and in some cases lethal. The technological breakthrough that allows adult skin cells to be reprogrammed into induced pluripotent stem cells (iPSCs) now offers the possibility of developing a permanent corrective therapy for EB. The therapeutic approach relies on the generation of patient-specific iPSCs, which then undergo genetic correction and differentiation into skin cells suitable for transplantation. The generation of iPSCs from the same patient in need of treatment would not only potentially avoid the complication of immune rejection, but also provide an unlimited and scalable source of patient- specific cells suitable for transplantation. For the initial studies funded by the parent R01, we focused on developing an iPSC-based therapy for a cohort of patients suffering from the Dowling-Meara subtype of EB simplex (EBS-DM), who share the same R125C “hot spot mutation” in the keratin 14 (Krt14) protein that is responsible for 70% of the EBS-DM cases. This EBS-DM study has allowed us to address all of the major obstacles and pre-requisites required for the development of a clinically relevant iPSC-based therapy for EBS. Specifically, we have developed a novel, efficient method for the generation of clinically relevant integration- free iPSCs. We have also used non-integrating selection marker-free gene editing technology to successfully correct the genetic defect in iPSCs generated from an EBS patient. In addition, our high-efficiency of reprogramming has recently allowed us to combine gene editing and reprogramming of EBS fibroblasts into a one-step procedure. This revision application is proposing to expand the one-step combined gene editing and reprogramming approach to a more severe form of EB, recessive dystrophic EB (RDEB) and to advance our findings toward a clinical trial. Given that the collaborative efforts among several groups will be required to ensure the successful translation of complex iPSC-based therapies into the clinic, we have recently entered into a consortium with Stanford and Columbia Universities to facilitate the development of an optimal manufacturing protocol for genetically corrected iPSC-derived cell products for RDEB treatment. Therefore, Aim 1 of this proposal is to improve the safety and reduce the manufacturing complexity of an iPSC-based therapy for RDEB by combining reprogramming and gene editing into a one-step procedure; Aims 2 and 3 are to manufacture cGMP-compliant reagents for reprogramming and gene editing respectively to assist the Consortium in generating Investigational New Drug (IND) - enabling data for the FDA. The EB iPS Cell Consortium’s success in obtaining approval for a clinical trial for RDEB would expand the therapeutic potential of iPSCs for the treatment of a variety of monogenic diseases affecting other internal organs, where the difficulty in monitoring adverse effects of an iPSC-based therapy would make them unlikely first targets.