Good manufacturing practice and clinical-grade human embryonic stem cell lines

Abstract

Human embryonic stem cell (hESC) lines, after directed differentiation, hold the greatest potential for cell transplantation treatment in many severe diseases. Good manufacturing practice (GMP) quality, defined by both the European Medicines Agency and the Food and Drug Administration, is a requirement for clinical-grade cells, offering optimal defined quality and safety in cell transplantation. Using animal substance-free culture media, feeder cells or feeder-free matrix in derivation, passaging, expansion and cryopreservation procedures, immune reactions against animal proteins in the cells, and infection risk caused by animal microbes can be avoided. It is also possible to apply GMP to animal components if no better options are available. In recent production of GMP-quality hESC lines, feeder cells had been cultured in fetal bovine serum, and the medium supplemented with an animal protein containing a serum replacement component. Using embryos cultured in a GMP laboratory, isolating the inner cell mass mechanically, deriving lines on human feeder cells originally cultured in xeno-free medium in a GMP laboratory, and using xeno-free media for derivation and culture of hESC lines themselves, GMP-quality xeno-free hESC lines could be established today. Human serum is a xeno-free component available today, but many chemically defined media are under development.

Safety paradigm: genetic evaluation of therapeutic grade human embryonic stem cells

“…The regenerative medicine field aims to translate the tremendous potential of stem cell biology to achieve tissue or organ regeneration in humans. However, if tissue-specific stem cells or hESCs are to be used to treat a wide variety of human diseases, then several formidable challenges are still to be overcome, which include thorough genotyping and phenotyping, well-controlled derivation and differentiation, cell efficacy, immunogenicity, tumourigenicity, appropriate cell-delivery systems, and short-and longterm safety (Civin & Rao 2006;De Sousa et al 2006;Gruen & Grabel 2006;Skottman & Hovatta 2006;Bongso et al 2008;Unger et al 2008).…”

“…Although it is unclear why the enzyme-based methods favour gross chromosomal rearrangements when this is not seen with other cell types such as fibroblasts, which are passaged in a similar way in vitro, the most likely explanation is that it is linked to well-established dynamic reciprocity between architectural integrity through cell adhesion and karyotype stability (Tlsty 1998). hESCs are polarized and express an epithelial plasma membrane protein profile (Krtolica et al 2007;Van Hoof et al 2008). Frequent disruption of cell-cell contacts and polarity may induce karyotype rearrangements and DNA damage that would, in differentiated cells, lead to cell death.…”

Constraining the Pluripotent Fate of Human Embryonic Stem Cells for Tissue Engineering and Cell Therapy – The Turning Point of Cell-Based Regenerative Medicine

“…Establishing defined platform for the long-tern stable maintenance of pluripotent hESCs has overcome some of the major obstacles in translational biology. Good manufacturing practice (GMP) quality, defined by both the European Medicine Agency (EMA) and the Food and Drug Administration (FDA), is a requirement for clinical-grade cells, offering optimal defined quality and safety in cell transplantation [11]. Resolving minimal essential requirements for the maintenance of pluripotent hESCs allows all poorly-characterized and unspecified biological additives, components, and substrates in the culture system, including those derived from animals, to be removed, substituted, or optimized with defined human alternatives for optimal production GMP-quality xeno-free hESC lines and their therapy derivatives [3,12].…”

“…In order to generate a large supply of uniform functional cells for tissue engineering and cell therapy, how to channel the wide differentiation potential of pluripotent hESCs efficiently and predictably to a desired lineage has been a major challenge for clinical translation. In addition, most currently available hESC lines were derived and maintained on animal feeder cells and proteins, therefore, such hESCs have been contaminated with animal biologics and unsuitable for clinical application [2,3,11–13]. Without a practical strategy to convert pluripotent cells direct into a specific lineage, previous studies and profiling of hESC differentiating multi-lineage aggregates have compromised their implications to molecular controls in human embryonic development.…”

Low temperature cell pausing: an alternative short-term preservation method for use in cell therapies including stem cell applications

“…Whilst conventional slow freezing generally yields over 90% viability post-thaw (Kleeberger et al 1999;Kotobuki et al 2005), for certain cell types such as human embryonic stem cells (hESC) grown as colonies, poor survival has been reported (Li et al 2010;Xu et al 2010). Vitrification is an alternative approach for storage of hESC (Li et al 2010), yet is disadvantaged due to higher concentrations of cryoprotective agents (CPAs), smaller storage volumes (typically 1-20µl per cryostraw) (Unger et al 2008;Coopman 2011;Hunt 2011) and the risk of contamination from open cryostraws (Mirabet et al 2012). Diminished functionality of cryopreserved human hepatocytes has also been documented; with cells losing their adherence capabilities and exhibiting an altered metabolic profile post-thaw (Terry et al 2006).…”