Department of Obstetrics and Gynecology, University Hospital Brno and Masaryk University, Czech Republic.
Department of Histology and Embryology, Faculty of Medicine, Masaryk University, Brno, Czech Republic.
Adv Clin Exp Med. 2023 Aug;32(8):901-907. doi: 10.17219/acem/158777.
Human embryonic stem cells (hESCs) have the unique ability to differentiate into any cell type in the human body and to proliferate indefinitely. Cell therapies involving hESC have shown very promising results for the treatment of certain diseases and confirmed the safety of hESC-derived cells for humans. They are used in cell therapy, mainly in targeted therapy of diseases that are currently incurable.
The aim of this study was the derivation of clinical-grade hESCs usable in drug development, non-native medicine and cell therapy.
Embryos were thawed, cultivated to the blastocyst stage if necessary, and assisted hatching was subsequently performed. Embryoblasts were mechanically isolated using narrow needles. Each line was kept as a separate batch. The derived hESCs were cultured under hypoxic culture conditions (5% O2, 5% CO2, 37°C) in a NutriStem® hPSC XF Medium with a daily medium change.
From January 2018 to July 2020, 138 selected clients were asked for consent to donate embryos, of whom 52 did not respond, 19 terminated the storage of their embryos and 29 extended the storage. Only 38 clients (27.5%) agreed to donate embryos for the derivation of hESCs. At the same time, personal communication with clients took place and another 17 embryo donors were recruited. A total of 160 embryos from 55 donors aged 26-42 years were collected. The embryos were frozen at the blastocyst (33.1%) or morula (46.3%) stage. After the preparation of 64 embryos, embryoblasts were isolated and cultured. Finally, 7 hESC lines were obtained, 4 research-grade and 3 clinical-grade, the first in the Czech Republic.
We established a current good manufacturing practice (cGMP)-defined xeno-free and feeder-free system for the derivation, culture and banking of clinical-grade hESC lines that are suitable for preclinical and clinical trials. The quality control testing with criteria concerning sterility, safety and characterization according to cGMP ensured the clinical-grade quality of hESC lines.
人类胚胎干细胞（hESC）具有分化为人体任何细胞类型并无限增殖的独特能力。涉及 hESC 的细胞疗法已为某些疾病的治疗显示出非常有前景的结果，并证实了 hESC 衍生细胞对人类的安全性。它们被用于细胞疗法，主要用于目前无法治愈的疾病的靶向治疗。
本研究旨在获得可用于药物开发、非本土药物和细胞治疗的临床级 hESC。
胚胎解冻后，如果需要，培养至囊胚阶段，然后进行辅助孵化。胚胎细胞使用细针机械分离。每条线作为一个单独的批次保存。衍生的 hESC 在低氧培养条件（5%O2、5%CO2、37°C）下，在 NutriStem® hPSC XF 培养基中培养，每天更换培养基。
从 2018 年 1 月至 2020 年 7 月，有 138 名选定的客户被要求同意捐赠胚胎，其中 52 名未回复，19 名终止了胚胎储存，29 名延长了储存。只有 38 名客户（27.5%）同意捐赠胚胎以获得 hESC 的衍生。同时，与客户进行了个人沟通，并招募了另外 17 名胚胎供体。共收集了 55 名年龄在 26-42 岁的供体的 160 个胚胎。胚胎在囊胚（33.1%）或桑椹胚（46.3%）阶段冷冻。准备了 64 个胚胎后，分离并培养胚胎细胞。最后，获得了 7 条 hESC 系，其中 4 条为研究级，3 条为临床级，这是捷克共和国的首例。
我们建立了一个符合现行良好生产规范（cGMP）的无动物成分和无饲养细胞的系统，用于衍生、培养和储存临床级 hESC 系，这些系适合临床前和临床试验。通过符合 cGMP 的无菌性、安全性和特性标准的质量控制测试，确保了 hESC 系的临床级质量。
