Goldyne Savad Institute of Gene Therapy, Hadassah Hebrew University Medical Center, Jerusalem, Israel.
PLoS One. 2012;7(6):e35325. doi: 10.1371/journal.pone.0035325. Epub 2012 Jun 20.
Clinically compliant human embryonic stem cells (hESCs) should be developed in adherence to ethical standards, without risk of contamination by adventitious agents. Here we developed for the first time animal-component free and good manufacturing practice (GMP)-compliant hESCs. After vendor and raw material qualification, we derived xeno-free, GMP-grade feeders from umbilical cord tissue, and utilized them within a novel, xeno-free hESC culture system. We derived and characterized three hESC lines in adherence to regulations for embryo procurement, and good tissue, manufacturing and laboratory practices. To minimize freezing and thawing, we continuously expanded the lines from initial outgrowths and samples were cryopreserved as early stocks and banks. Batch release criteria included DNA-fingerprinting and HLA-typing for identity, characterization of pluripotency-associated marker expression, proliferation, karyotyping and differentiation in-vitro and in-vivo. These hESCs may be valuable for regenerative therapy. The ethical, scientific and regulatory methodology presented here may serve for development of additional clinical-grade hESCs.
临床合规的人类胚胎干细胞（hESC）应在符合伦理标准的情况下开发，且不会有外源因子污染的风险。在此，我们首次开发了无动物源成分且符合良好生产规范（GMP）的 hESC。在供应商和原材料资格认证后，我们从脐带组织中获得了无动物源、GMP 级别的饲养层细胞，并在一种新颖的、无动物源的 hESC 培养系统中使用它们。我们按照胚胎获取、良好组织、制造和实验室实践的规定，衍生和表征了三种 hESC 系。为了尽量减少冻融，我们从最初的外植体开始连续扩大细胞系，并将样本作为早期库存和银行进行冷冻保存。批次放行标准包括用于身份鉴定的 DNA 指纹分析和 HLA 分型、多能性相关标志物表达的特征分析、体外和体内增殖、核型分析和分化。这些 hESC 可能对再生疗法有价值。这里提出的伦理、科学和监管方法可用于开发其他临床级别的 hESC。
