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    IPSC Reprogramming: Genomic Hardware-Friendly But Marred By Epigenetic Software Bugs
    By Jennifer Wong | January 28th 2013 09:30 PM | Print | E-mail | Track Comments
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    My column covers the latest primary research discoveries in the life-science discipline. Much of what is reported here are considered discoveries...

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    In light of ethical concerns with using human embryonic stem (hES) cells for research and therapeutic development, scientists have since developed technologies to allow one to reprogram somatic cells into pluripotent stem cells that are akin to hES cells.

    With the advent of induced pluripotent stem cell (iPSC) technology in 2006, scientists can now easily reprogram somatic cells into pluripotent stem cells. Although an incredible achievement, there still remains considerable skepticism with regards to whether iPSCs truly behave like ES cells, whether information garnered from iPSCs in disease models could be reliable, and more importantly whether iPSCs could have cancer-causing potential that could seriously impact the therapeutic safety of iPSCs.

    The above skepticism soon became a true concern as scientists uncovered disturbing molecular flaws in iPSCs. Studies by Chin et al 2009 (Cell Stem Cell 5: 11–123) demonstrated profound gene expression differences between iPSCs and ES cells. This was followed by a troublesome discovery by Stadtfeld et al 2010 (Nature 465, 175–181), who revealed that iPSCs carry epigenetic memories that contribute to the inability of iPSCs to support embryonic development.

    Moreover, these aberrant epigenetic memories were further demonstrated to be associated with cancer (Ohm et al., 2010, Cancer Res, 70(19):7662-7673).  In 2011, a series of publications in Nature revealed profound flaws in the iPSC genomic landscape including: somatic coding mutations with the average of 6 coding point mutations per exon (Gore et al, 2011, Nature 471, 63–67), de novo copy number variations specific to iPSCs (Hussein et al, 2011, Nature 471, 58–62), as well as aberrant DNA methylation (epigenetic) patterns in up to 69% of methylation hotspots through which gene expression programs are regulated. 

    With these unsettling findings, iPSC technology quickly gained the notorious reputation as the “flawed” representative of nature’s pluripotent embryonic stem cells.

    Less than a year later, iPSC technology made an incredible comeback with exciting discoveries suggesting that the reprogramming technique itself is not the cause of genomic aberrations observed in iPSCs. For example, studies by Cheng et al, 2012 (Cell Stem Cell 10, 337–344) and Young et al 2012 (Cell Stem Cell 10,  570-582),  reported compelling deep-sequencing evidence suggesting that somatic mutations and copy number variations in the iPSC genome are not caused by reprogramming itself.

    Specifically, they find that both episomal and lentiviral reprogramming techniques generated iPSC clones that individually harbored unique genomic mutations distinct from each other- indicating that mutations are likely not from reprogramming itself. Furthermore, each iPSC clone shared at least 50% of their genomic aberrations with the parental somatic cells from which they were generated, strongly suggesting the parental origin of these mutations. The remaining 50% of the mutations were shown to be non-clustered, suggesting that they are likely caused by random mutational events during iPSC expansion in culture.

    The good news is that none of these mutations fall into the cancer-causing category, suggesting that these mutations most likely reflect the normal phenomenon of cells passaged in culture.

    The above insights from deep-sequencing indicate that reprogramming itself does not cause any changes in the genomic hardware of iPSCs. Rather, the problem lies in the pre-existing genomic hardware of the somatic cells whose flaws are captured during the reprogramming process, and consequently re-emerge in the iPSC genomic landscape; a problem that can be easily fixed by setting standard selection criteria for parental cells. Moreover, de novo mutations that emerge in iPSCs likely reflect random benign mutations that typically occur in cells passaged in culture- a problem that can be minimized by standardization of culture techniques.

    However, the same cannot be said for the influences of iPSC reprogramming on the epigenetic software governing global gene expression. Although there is evidence that iPSCs display the somatic epigenetic program of their parental cells (Ohi et al., 2011, Nature Cell Biology 13(5): 541-549), iPSCs also carry cancer-causing epigenetic aberrations that arise from transcription factors responsible for reprogramming (Ohm et al., 2010, Cancer Res, 70(19):7662-7673).

    A recently discovered epigenetic abnormality in iPSCs is the progressive erosion of the epigenetic software behind X-chromosome inactivation, which can seriously impact the ability of iPSCs to accurately model X-linked diseases (Mekhoubad et al., 2012, Cell Stem Cell, 10, 595-609). 

    It is therefore apparent that although reprogramming technology has little impact on the integrity of the genomic hardware in iPSCs, there still remains a few epigenetic software bugs that may still be cause for concern.