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

Abstract

The use of stem cells for regenerative medicine has captured the imagination of the public, with media attention contributing to rising expectations of clinical benefits. Human embryonic stem cells (hESCs) are the best model for capital investment in stem cell therapy and there is a clear need for their robust genetic characterization before scaling-up cell expansion for that purpose. We have to be certain that the genome of the starting material is stable and normal, but the limited resolution of conventional karyotyping is unable to give us such assurance. Advanced molecular cytogenetic technologies such as array comparative genomic hybridization for identifying chromosomal imbalances, and single nucleotide polymorphism analysis for identifying ethnic background and loss of heterozygosity should be introduced as obligatory diagnostic tests for each newly derived hESC line before it is deposited in national stem cell banks. If this new quality standard becomes a requirement, as we are proposing here, it would facilitate and accelerate the banking process, since end-users would be able to select the most appropriate line for their particular application, thus improving efficiency and streamlining the route to manufacturing therapeutics. The pharmaceutical industry, which may use hESC-derived cells for drug screening, should not ignore their genomic profile as this may risk misinterpretation of results and significant waste of resources.

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

“…47 The low-grade mosaic aneuploidy and the recurrent genomic instability of chromosome 1 that were found in this manuscript illustrate the need for careful monitoring of the cells. Our findings demonstrate the phenomenon of genomic instability in stem cell lines, which increases over time.…”

Induced Pluripotent Stem Cell Differentiation and Three-Dimensional Tissue Formation Attenuate Clonal Epigenetic Differences in Trichohyalin

“…To assess the genomic integrity of our lines, we subjected iKCL004 and iKCL011 to array CGH at two different time points, one at early passages (<10) and the other about 3 months after the lines were adapted to the feeder-free culture. Even though it is widely known that extended periods of cell culture under feeder-free conditions are linked to higher rates of chromosomal abnormalities in pluripotent stem cells, especially iPSC [8,9], we did not detect any copy number changes using array CGH in either iKCL004 or iKCL011 over 3 months of culture on Matrigel in mTeSR1 medium [10], suggesting that the genome of both lines is relatively stable. Next, we subjected iKCL004, iKCL011, and parental BJ fibroblast to exome sequencing.…”

“…Karyotype of both iPSC lines was stable over the extended periods of in vitro culture under feeder-free conditions. Cytogenetic resilience of iKCL004 and iKCL011 lines might arise from particular aspects of cell culture, as well as reflect a genetic predisposition to chromosomal stability/instability, commonly noted with hESC lines [8,9,33,34]. hESC is derived from embryos created by in vitro fertilization, often from donors that have problems with fertility.…”

“…Altered gene expression and distinct differentiation propensity among early passage iPSC clones are likely to be largely attenuated through continuous passaging [7]. In contrast, human pluripotent stem cells, both human ESC (hESC) and especially iPSC, kept for extended period of time in culture are prone to genomic instability [8,9]. It is unknown to what extent the cellular genetic and epigenetic makeup or culture conditions contribute to genomic instability over time.…”

Translation and commercialization of regenerative medicines

“…The commercial use of committed stem cells might be expected to pose fewer risks but in addition the research surrounding them adds significantly to the overall understanding of stem cell lineages. Ilic and colleagues' (Stephenson et al 2010) important contribution emphasizes the need to fully characterize stem cells prior to banking or expansion. Their proposed new quality standard could aid and accelerate the banking process and facilitate appropriate selection of stem cell lines by end-users.…”