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Breakthrough of the year

Breakthrough of the year. Induced pluripotent stem cells (iPS cells). Definitions.

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Breakthrough of the year

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  1. Breakthrough of the year Induced pluripotent stem cells (iPS cells)

  2. Definitions • iPS are cells created from adult differentiated cells by a simple genetic trick - the genes coding four transcription factors are cloned into viral vector, and simply adding the vectors to a culture of adult cells (e.g. fibroblasts) under right conditions results in cells reprogrammed to embryonic “stemness”

  3. Induced pluripotent stem cells (iPS cells) • Reprogramming of somatic cells by transcription factors • Indistinguishable from ES cells • Germ line competent • Both mouse- and human-derived lines were obtained • Reprogramming with c-MYC oncogene • Retroviral transduction Shinya Yamanaka (Kyoto University) ! +++ Rudolf Jaenisch (MIT) Differentiation Differentiation Differentiation RNA Pol II Oct-4 SOX2 Myc Klf4 Growth factors Differentiation Differentiation Polycomb Differentiation Differentiation Differentiation

  4. Derivation of iPS cells using an inducible lentiviral system -the two step process 1. Oct-4 locus or Nanog locus Oct-4 locus or Nanog locus GFP GFP Dox Fibroblast (GFP-) iPS cell (GFP+) 2. Infection Oct-4 Sox2 5’LTR tetO 3’LTR Klf4 c-Myc Brambrink et al. Cell Stem Cell, February 2008

  5. iPS cells make germ line chimeras Oct-4 GFP iPS cells Nanog GFP iPS cells Oct-4 GFP fibroblasts Injection chimera (white and brown coat color) Oct-4 GFP iPS chimera Nanog GFP iPS chimera Brambrink et al. Cell Stem Cell, February 2008

  6. Kinetics of fibroblast to iPS cell reprogramming-not too long way to go back stable reprogramming Time in days 0 4 8 12 Thy1 (and other fibroblast genes) Fibroblasts iPS cells SSEA-1 retroviral activity pluripotency genes telomerase silent X chromosome Stadtfeld et al. Cell Stem Cell, March 2008

  7. A reprogramming rush since 2006 August 2006, Cell iPS cells from mouse embryonic and adult fibroblasts July 2007, Nature germline-competent iPS cells August 2007, Nat Biotech isolation of “iPS” cells by stringent selection for activation of a neomycin-resistance gene inserted into the endogenous Oct-4 November 2007, Science iPS cells without c-Myc from mouse fibroblasts November 2007, Nat Biotech iPS cells without c-Myc from human fibroblasts December 2007, Science treatment of sicle-cell anaemia mouse model with iPS cells February 2008, Science iPS cells from adult mouse liver and stomach cells April 2008, Cell iPS cells from mature mouse B lymphocytes April 2008, PNAS neurons from mouse iPS cells improve symptoms of rats with PD May 2008, Stem Cells differentiation of mouse iPS into cardiovascular and hematopoietic lineages

  8. iPS cells versus hESCs(Induced pluripotent stem cells versus human embryonic stem cells) • Progress toward socially beneficial applications of stem cell science would be indefensibly delayed if iPS cell research is pursued at the expense of further hESC research (e.g. unsolved question of tumorigenicity and safety - c-Myc and retroviral promoters) • iPS cell research might not translate to clinics (e.g. limited differentiation, unexpected deregulation of genes) • hESCs will have to be used as important control to examine the safety and abilities of human iPS cells (hESCs represent the only pluripotent cells that are genetically unmodified) • There are invaluable avenues of research that may not be easily pursued with reprogrammed somatic cells (e.g. developmental studies)

  9. 5 things to know before jumping on the iPS playground • Four familiar genes do the trick • But as simple as this procedure might seem, iPS cells are not easy to make • Low efficiency of derivation; expertise in hESC culture is absolutely critical • Nevertheless, iPS expertise is becoming common - “the whole word is doing it now” Anyone can do it! Status: Fact (mostly) Adapted from Nature, March 2008

  10. 5 things to know before jumping on the iPS playground • Alternative system for therapeutic cloning (?) • Some of the viral vectors, as well as some of the genes themselves, may cause cancer • The use of custom-made cells would take a ridiculous amount of money (~1M US$) and long time (~2 years; April 2008 - a national library of therapy-ready cell lines from placental and cord blood in Japan) 2. Everyone can have their own custom-tailored pluripotent cells! Status: Fiction (unless you are rich) Adapted from Nature, March 2008

  11. 5 things to know before jumping on the iPS playground • iPS cells will certainly provide models for disease first - easy access to patient’s cells (big biotech companies are already interested!) • UCLA and Harvard U are discussing plans to start iPS bank for the purpose of disease modeling • iPS for model sicle-cell anaemia and PD (Kyoto and MIT) • The first trial of hESC-based treatment will start in the middle of this year (Geron, CA; oligodendrocytes and patients with spinal-cord injuries) • Application of iPS cells largely depends on how above trial goes (unpublished results - iPS cell-based treatment of spinal-cord injury in mice) 3. The cures are on their way! Status: Too soon to tell Adapted from Nature, March 2008

  12. 5 things to know before jumping on the iPS playground • Based on morphology, chromosome profile and gene expression there are no major differences yet • Difficult task as markers don’t mean anything! (e.g. some cancer cells express protein markers of pluripotency as well) • Some iPS cells express just a few markers of pluripotency, some express all - different cells? Different tumorigenic potential, differentiation potential, migratory potential? • Each line of iPS cells will require rigorous testing as it is ongoing with hESCs. • Thomson: “If there will be no differences between hESCs and iPS cells, hESCs will probably turn out to be a historical anomaly!” 4. Embryonic stem cells are the same as iPS cells! Status: Fact (so far, anyway) Adapted from Nature, March 2008

  13. 5 things to know before jumping on the iPS playground • iPS cells represent scientific advancement within ethical boundaries (G.Bush day after Yamanaka and Thomson announced the creation of human iPS cells) • Yamanaka: “many people can do it - and without telling anybody. Someone might use iPS cells to derive gametes, eggs and sperm from one iPS cell line!” • iPS cell “adventurers” might try to create a live, cloned human (the first cloned mouse by using iPS cells was created) • February 21, 2008 - a notification specifically forbidding “the imlantation of embryos made with iPS cells into human or animal wombs, the production of an individual in any other way from iPS cells, the introduction of iPS cells into embryo or fetus, and the production of germ cells from iPS cells” in Japan 5. iPS cells have no ethical issues! Status: Fiction (depends on what you want to do) Adapted from Nature, March 2008

  14. Good luck with the tests!

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