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Tumorigenicity-associated characteristics of human iPS cell lines.

ABSTRACT: Human induced pluripotent stem cells (hiPSCs) represent promising raw materials of human cell-based therapeutic products (hCTPs). As undifferentiated hiPSCs exhibit intrinsic tumorigenicity properties that enable them to form teratomas, hCTPs containing residual undifferentiated hiPSCs may cause tumor formation following transplantation. We first established quantitative and sensitive tumorigenicity testing of hiPSCs dissociated into single cells using NOD/Shi-scid IL2R?null (NOG) mice by inhibiting apoptosis of hiPSCs with a Rho kinase inhibitor. To examine different features in tumorigenicity of various hiPSCs, 10 commonly available hiPSC lines were subjected to in vivo tumorigenicity testing. Transplanted hiPSC lines showed remarkable variation in tumor incidence, formation latency, and volumes. Most of the tumors formed were classified as immature teratomas. However, no signs of malignancies, such as carcinoma and sarcoma, were recognized in the tumors. Characteristics associated tumorigenicity of hiPSCs were investigated with microarray analysis, karyotype analysis, and whole exome sequencing. Gene expression profiling and pathway analysis supported different features of hiPSC lines in tumorigenicity. hiPSC lines showed chromosomal abnormalities in some lines and 61-77 variants of cancer-related genes carrying effective nonsynonymous mutations, which were confirmed in the COSMIC databases. In this study, the chromosomal abnormalities and cancer-related gene mutations observed in hiPSC lines did not lead to the malignancy of tumors derived from hiPSCs. Our results suggest that the potential tumorigenicity risk of hCTPs containing residual undifferentiated hiPSCs is dependent on not only amounts of undifferentiated hiPSCs but also features of the cell lines used as raw materials, a finding that should be considered from the perspective of quality of hCTPs used.

SUBMITTER: Yasuda S

PROVIDER: S-EPMC6171902 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Tumorigenicity-associated characteristics of human iPS cell lines.

Yasuda Satoshi S Kusakawa Shinji S Kuroda Takuya T Miura Takumi T Tano Keiko K Takada Nozomi N Matsuyama Satoko S Matsuyama Akifumi A Nasu Michiyo M Umezawa Akihiro A Hayakawa Takao T Tsutsumi Hideki H Sato Yoji Y

PloS one 20181004 10

Human induced pluripotent stem cells (hiPSCs) represent promising raw materials of human cell-based therapeutic products (hCTPs). As undifferentiated hiPSCs exhibit intrinsic tumorigenicity properties that enable them to form teratomas, hCTPs containing residual undifferentiated hiPSCs may cause tumor formation following transplantation. We first established quantitative and sensitive tumorigenicity testing of hiPSCs dissociated into single cells using NOD/Shi-scid IL2Rγnull (NOG) mice by inhibi ...[more]

PMID: 30286143

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Project description:BACKGROUND:hTERT/cdk4 immortalized myogenic human cell lines represent an important tool for skeletal muscle research, being used as therapeutically pertinent models of various neuromuscular disorders and in numerous fundamental studies of muscle cell function. However, the cell cycle is linked to other cellular processes such as integrin regulation, the PI3K/Akt pathway, and microtubule stability, raising the question as to whether genetic modification related to the cell cycle results in secondary effects that could undermine the validity of these cell models. RESULTS:Here we subjected five healthy and disease muscle cell isolates to transcriptomic analysis, comparing immortalized lines with their parent primary populations in both differentiated and undifferentiated states, and testing their myogenic character by comparison with non-myogenic (CD56-negative) cells. Principal component analysis of global gene expression showed tight clustering of immortalized myoblasts to their parent primary populations, with clean separation from the non-myogenic reference. Comparison was made to publicly available transcriptomic data from studies of muscle human pathology, cell, and animal models, including to derive a consensus set of genes previously shown to have altered regulation during myoblast differentiation. Hierarchical clustering of samples based on gene expression of this consensus set showed that immortalized lines retained the myogenic expression patterns of their parent primary populations. Of 2784 canonical pathways and gene ontology terms tested by gene set enrichment analysis, none were significantly enriched in immortalized compared to primary cell populations. We observed, at the whole transcriptome level, a strong signature of cell cycle shutdown associated with senescence in one primary myoblast population, whereas its immortalized clone was protected. CONCLUSIONS:Immortalization had no observed effect on the myogenic cascade or on any other cellular processes, and it was protective against the systems level effects of senescence that are observed at higher division counts of primary cells.

Project description:The reprogramming of human fibroblasts to generate induced pluripotent stem cells (hiPSCs) has been achieved through the expression of only a few exotic factors1-8, which is morphologically and molecularly verified in outer cellular states by characteristic markers, due to the remodeling of the somatic cell transcription programs in inner cellular states to the ES-like condition. Transcription factor-induced reprogramming to self-renewal and pluripotency raises the question as to how the exotic factors act to bring about these changes in the two cellular states9-11. Here, we applied RNA profiling to uncover gene expression changes and glycan profiling12 to survey structural changes in glycans, to compare hiPSCs and parental somatic cells, in total 51 cells, which were originally cultured and established, and the changes were analyzed by the combination of standard statistical techniques and a network approach13. We fist found a gene expression signature of 2502 genes with significant difference between iPSCs and SCs, and by the following network analysis by considering the expression signature, we found a network signature of 28 regulatory networks of 76 genes, which were related to the glycan biosynthetic pathways including 3 glycosyltransferase, in addition to well known signal pathways and cell-cell interaction pathways. Concomitantly, we found a glycan signature of six glycan structures characterized 16 lectins on lectin microarray by the correspondence with 12 glycosyltransferases in expression signature. In particular, the correspondence detected between the three expression signatures revealed 14 candidate glycosyltransferase, which are responsible for glycan transfer related to known epitopes for the differentiation such as SSEA epitope family in glycan biosynthesis pathway, based on characteristic changes in the cellular surface states of the hiPSCs. This sheds new light on a possible linkage between the inner and outer cellular states for reprogramming to self-renewal and pluripotency in hiPSC.

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Tumorigenicity-associated characteristics of human iPS cell lines.

Publications

Tumorigenicity-associated characteristics of human iPS cell lines.

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