Accreditation of Biosafe Clinical-Grade Human Embryonic Stem Cells According to Chinese Regulations

摘要

class="head no_bottom_margin" id="sec1title">IntroductionHuman embryonic stem cells (hESCs), after directed differentiation, are valuable in regenerative medicine. However, clinical trials using hESC-derived cells remain scarce primarily because hESC lines available worldwide are mostly of nonclinical grade. To generate clinical-grade cells, good manufacturing practices (GMPs), which cover operating procedures and product quality control, must be employed (). GMP is a quality assurance system that requires the traceability of materials and the validation of standard operating procedures (SOPs) (). Stem cells are a type of human cellular and tissue-based product (HCT/P). In many countries, HCT/Ps are regulated under guidelines such as 21 CFR 1270 and 21 CFR 1271 issued by the US Food and Drug Administration (FDA) (). Stem cell-based products must also meet the requirements of other therapeutic products including drugs, medical devices, xeno-transplants, and biological products (). Therefore, a number of countries and professional associations (e.g., International Society for Stem Cell Research) have issued preliminary regulatory policies for the clinical application of stem cells, and GMP serves as the basic requirement for the generation of clinical-grade hESCs (, , ). The existing guidelines incorporate guidelines produced by the British Standards Institute for cell-based clinical application (). Recently, in China, drafts of a stem cell-specific clinical therapy quality control standard and management of stem cell-based clinical experiments were implemented by the China Food and Drug Administration (CFDA) () and National Health and Family Planning Commission of the People’s Republic of China (NHFPC) (). Each of these guidelines focuses on the efficacy, safety, and pharmaceutical quality, which are influenced by the cell sources, manufacturing systems, and specific therapeutic protocols (, ). In addition to validating the biosafety of the hESCs, the CFDA and FDA both require rigorous testing of the donors' eligibility, thus differentiating these guidelines from the present NIH guidelines on human stem cell research (, , ).The generation of hESCs involves numerous reagents, including growth factors, small molecules, and media. To avoid infection by animal-sourced components, mouse feeder cells can be substituted by human fibroblasts (href="#bib9" rid="bib9" class=" bibr popnode">Ellerstrom et al., 2006, href="#bib13" rid="bib13" class=" bibr popnode">Genbacev et al., 2005). Chemically defined hESC culture media such as mTeSR1, mTeSR2, and E8 have been developed to complement future hESC clinical applications (href="#bib6" rid="bib6" class=" bibr popnode">Chen et al., 2011, href="#bib26" rid="bib26" class=" bibr popnode">Ludwig et al., 2006a, href="#bib27" rid="bib27" class=" bibr popnode">Ludwig et al., 2006b). To facilitate and standardize the development of safe and effective clinical-grade hESCs, most clinical hESC studies have been reported as xeno-free or clinical-grade; a summary of current globally available xeno-free hESCs is listed in href="#mmc1" rid="mmc1" class=" supplementary-material">Table S1 (href="#bib7" rid="bib7" class=" bibr popnode">Crook et al., 2007, href="#bib19" rid="bib19" class=" bibr popnode">Ilic et al., 2012, href="#bib22" rid="bib22" class=" bibr popnode">Klimanskaya et al., 2006, href="#bib30" rid="bib30" class=" bibr popnode">Rajala et al., 2010, href="#bib33" rid="bib33" class=" bibr popnode">Schwartz et al., 2012, href="#bib39" rid="bib39" class=" bibr popnode">Tannenbaum et al., 2012). However, studies have only rarely demonstrated the biosafety investigated by an accredited organization. Here, we propose generation of clinical-grade hESCs with the following requirements: (1) donor consent, (2) donor eligibility requirements, (3) the use of completely xeno-free reagents, (4) biosafety tests from authorized organizations, and (5) stability, self-renewal characteristics, and differentiation capability.Two clinical-grade hESC lines (Q-CTS-hESC-1 and Q-CTS-hESC-2) were successfully derived under GMP-controlled conditions in completely xeno-free culture media. Both cell lines were pluripotent and passed biosafety evaluations. For a further validation of their pluripotency and biosafety, both cell lines were reviewed and deemed eligible by the National Institutes for Food and Drug Control (NIFDC). Additionally the cell line Q-CTS-hESC-1 is parthenogenetic. Theoretically, parthenogenetic cells express high level of homologous human leukocyte antigen without recombination during meiosis I; thus, they have low histocompatibility and are applicable to more allograft recipients (href="#bib21" rid="bib21" class=" bibr popnode">Kim et al., 2007). Finally, the cell line Q-CTS-hESC-2 was selected for direct differentiation and was able to differentiate into clinical-grade cell types with representatives of three germ layers (ectoderm: retinal pigment epithelium [RPE] cells and neuronal progenitors; mesoderm: cardiomyocytes; endoderm: hepatocytes). Furthermore, clinical-grade neuronal progenitors differentiated from Q-CTS-hESC-2 cells survived and further differentiated into tyrosine hydroxylase (TH)-positive mature dopamine (DA) neurons after transplantation into Parkinson's disease (PD) rat models. These results demonstrate the value of these two clinical-grade hESC lines as sources for future hESC-based clinical trials or therapies.