High-resolution DNA analysis of human embryonic stem cell lines reveals culture-induced copy number changes and loss of heterozygosity

Abstract

Prolonged culture of human embryonic stem cells (hESCs) can lead to adaptation and the acquisition of chromosomal abnormalities, underscoring the need for rigorous genetic analysis of these cells. Here we report the highest-resolution study of hESCs to date using an Affymetrix SNP 6.0 array containing 906,600 probes for single nucleotide polymorphisms (SNPs) and 946,000 probes for copy number variations (CNVs). Analysis of 17 different hESC lines maintained in different laboratories identified 843 CNVs of 50 kb-3 Mb in size. We identified, on average, 24% of the loss of heterozygosity (LOH) sites and 66% of the CNVs changed in culture between early and late passages of the same lines. Thirty percent of the genes detected within CNV sites had altered expression compared to samples with normal copy number states, of which >44% were functionally linked to cancer. Furthermore, LOH of the q arm of chromosome 16, which has not been observed previously in hESCs, was detected.

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

“…A recent study compared early and late passages of hESC lines and reported about high rate of copy number variations and loss of heterozygocity changes. 37 Taken together, these findings emphasize the need of tight surveillance of genomic integrity during long-term culture and the preference of low passages for clinical applications.…”

Concise Review: Induced Pluripotent Stem Cells Versus Embryonic Stem Cells: Close Enough or Yet Too Far Apart?

“…Overall, even though iPSC obviously need to pass through one additional selection process, reprogramming, iPSC and ESC are not drastically different considering the mutational load. In addition, there is some correlation between mutations found in late passage human ESC, iPSC, and cancers cells [75][76][77]. Thus, pluripotent cells cultured in vitro are by definition proliferating and can acquire specific aberrations that support growth advantage, similar to tumor progression, that eventually take over the population.…”

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

“…Though, among 17 lines analysed by Närvä et al (2010) one, FES 21, had identical q arms on chromosome 16, whereas all others had a heterozygous set of chromosomes.…”

“…Despite wide interest in defining the properties of hESCs, comprehensive characterization has been done with only a subset of lines. The popularity of some of the earliest lines derived, such as H1 and H9, both for research and for the generation of clinical grade banks, arises not necessarily because they are superior lines, as these cells were derived on mouse feeders in the presence of serum, but because by default (Adewumi et al 2007;Allegrucci & Young 2007;Lefort et al 2008;Närvä et al 2010). Although many of such studies have been useful in identifying 'master' genes of pluripotency, no consistent clustering could be ascribed to variations in chromosomal stability or differentiation propensity of the hESC lines.…”

“…This might be particularly useful since S684 Review. Genetic evaluation of therapeutic hESC E. Stephenson et al hESCs in culture have been shown to acquire small chromosomal amplifications or deletions (Maitra et al 2005;Lefort et al 2008;Spits et al 2008;Hovatta et al 2010;Närvä et al 2010). UPD, a condition in which both alleles have originated from a single parent, occurs as heterodisomy and isodisomy (Engel 1980;Robinson 2000).…”