A major challenge for the clinical use of human pluripotent stem cells is the development of safe, robust and controlled differentiation protocols. Adaptation of research protocols using reagents designated as research-only to those which are suitable for clinical use, often referred to as good manufacturing practice (GMP) reagents, is a crucial and laborious step in the translational pipeline. However, published protocols to assist this process remain very limited. We adapted research-grade protocols for the derivation and differentiation of long-term neuroepithelial stem cell progenitors (lt-NES) to GMP-grade reagents and factors suitable for clinical applications. We screened the robustness of the protocol with six clinical-grade hESC lines deposited in the UK Stem Cell Bank. Here, we present a new GMP-compliant protocol to derive lt-NES, which are multipotent, bankable and karyotypically stable. This protocol resulted in robust and reproducible differentiation of several clinical-grade embryonic stem cells from which we derived lt-NES. Furthermore, GMP-derived lt-NES demonstrated a high neurogenic potential while retaining the ability to be redirected to several neuronal sub-types. Overall, we report the feasibility of derivation and differentiation of clinical-grade embryonic stem cell lines into lt-NES under GMP-compliant conditions. Our protocols could be used as a flexible tool to speed up translation-to-clinic of pluripotent stem cells for a variety of neurological therapies or regenerative medicine studies.

中文翻译：

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GMP 级神经祖细胞源自临床级人类胚胎干细胞并分化。

人类多能干细胞临床应用的一个主要挑战是开发安全、稳健和受控的分化方案。使用指定为仅供研究使用的试剂对研究方案进行调整，使其适合于临床使用，通常称为良好生产规范 (GMP) 试剂，是转化流程中至关重要且费力的一步。然而，已发布的协助这一过程的协议仍然非常有限。我们将用于长期神经上皮干细胞祖细胞 (lt-NES) 的衍生和分化的研究级方案改编为适合临床应用的 GMP 级试剂和因子。我们用英国干细胞库中存放的 6 个临床级 hESC 系筛选了该方案的稳健性。在这里，我们提出了一种新的符合 GMP 要求的 lt-NES 协议，该协议具有多能性、可存储性且核型稳定。该方案导致了几种临床级胚胎干细胞的稳健且可重复的分化，我们从中衍生出 lt-NES。此外，GMP 衍生的 lt-NES 表现出很高的神经源潜力，同时保留了重定向到几种神经元亚型的能力。总的来说，我们报告了在符合 GMP 的条件下将临床级胚胎干细胞系衍生和分化为 lt-NES 的可行性。我们的方案可以用作灵活的工具，以加快多能干细胞转化为临床的速度，用于各种神经治疗或再生医学研究。