2014 Fiscal Year Annual Research Report
| Project/Area Number |
14F04787
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| Research Institution | Kyoto University |
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Principal Investigator |
山中 伸弥 京都大学, iPS細胞研究所, 教授 (10295694)
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| Co-Investigator(Kenkyū-buntansha) |
JOHANSSON Erik Martin 京都大学, iPS細胞研究所, 外国人特別研究員
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| Project Period (FY) |
2014-04-25 – 2017-03-31
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| Keywords | Glioma / IDH1 / Differentiation / iPSC / DNA methylation / Epigenomics |
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| Outline of Annual Research Achievements |
Mutation of the IDH1 gene is as an early event in low-grade and secondary glioblastoma, preceding other early aberrations such as p53 mutation. Mutant IDH1 catalyzes formation of 2-Hydroxyglutarate, leading to increased DNA and histone methylation and stabilization of the oncogene HIF-1α.
In order to find out more about IDH1 mutant function in glioma we will mimic early glioma development by using iPS cells expressing a Doxycycline-inducible IDH1 mutant. We are planning to differentiate these cells to glial lineages and induce expression of IDH1 mutant protein at different stages of differentiation to see the effect on tumor initiation. We plan to study genome-wide changes in gene expression as well as DNA and histone methylation.
Last year I started differentiation of iPS cells to Neural Stem Cells (NSCs) using several protocols to determine which is the optimal one. After successful differentiation to NSCs I have to further differentiate the cells to astrocytes and oligodendrocytes. Confirmation of successful NSC differentiation was done by analysis of NSC marker expression.
We believe the resulting data from this project will give insight about novel targets of mutant IDH1 and information about aberrant epigenetic events leading to gliomagenesis.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
Since previous reports of in vitro differentiation of pluripotent stem cells to glial lineages are very few, we are currently optimizing a protocol for differentiation of iPS cells to glial cells via neural stem cells. Successful neural stem cell generation from iPS cells has been confirmed by using quantitative PCR and immunocytochemistry to evaluate induced expression of neural stem cell markers (such as Nestin and Pax4) and repressed expression of pluripotency markers (such as Oct4 and Nanog).
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| Strategy for Future Research Activity |
We are now optimizing the second differentiation step: from neural stem cells to glial lineages (astrocytes and oligodendrocytes). Once this has been successfully implemented we will use our established protocol on iPS cells expressing a Doxycycline-inducible IDH1 mutant gene, and induce IDH1 mutant expression at several time points during differentiation in order to study glioma formation as seen by expression of glioma tumor markers and tumorigeneity using both in vitro and in vivo assays. We will use next-generation sequencing to analyze differences, on the single cell level, in global DNA and histone methylation as well as RNA expression during gliomagenesis, depending on the timing of mutant IDH1 induction.
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[Journal Article] TLX activates MMP-2, promotes self-renewal of tumor spheres in neuroblastoma and correlates with poor patient survival2014
Author(s)
Chavali PL, Saini RK, Zhai Q, Vizlin-Hodzic D, Venkatabalasu-bramanian S, Hayashi A, Johansson E, Zeng ZJ, Mohlin S, Påhlman S, Hansford L, Kaplan DR, Funa K
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Journal Title
Cell Death & Disease
Volume: 5
Pages: e1502
DOI
Peer Reviewed / Open Access
