Why most cells fail to reprogram

The ability to drive somatic, or fully differentiated, human cells back to a pluripotent or ‘stem cell’ state would overcome many of the scientific and social challenges to the use of embryo-derived stem cells and help realise the promise of regenerative medicine.

Recent research with mouse and human cells has demonstrated that such a transformation is possible, although the current process is inefficient and — when it does work — poorly understood. But now, thanks to the application of powerful new integrative genomic tools, a cross-disciplinary US research team from Harvard University, Whitehead Institute and the Broad Institute of MIT and Harvard has uncovered critical molecular events that underlie the direct reprogramming process.

Their findings are published online in the journal Nature.

“We used a genomic approach to identify key obstacles to the reprogramming process and to understand why most cells fail to reprogram,” said research leader Alexander Meissner, assistant professor at Harvard University’s Department of Stem Cell and Regenerative Biology and associate member of the Broad Institute.

“Currently, reprogramming requires infecting somatic cells with engineered viruses. This approach may be unsuitable for generating stem cells that can be used in regenerative medicine. Our work provides critical insights that might ultimately lead to a more refined approach.”

Previous work had demonstrated that four transcription factors could drive fully differentiated cells, such as skin or blood cells, into induced pluripotent stem (iPS) cells. With that as a basis, the researchers examined both successfully and unsuccessfully reprogrammed cells to better understand the complex process.

“Interestingly, the response of most cells appears to be activation of normal fail-safe mechanisms,” said Tarjei Mikkelsen, a graduate student at the Broad Institute and first author of the Nature paper.

“Improving the low efficiency of the reprogramming process will require circumventing these mechanisms without disabling them permanently.”

The researchers used next-generation sequencing technologies to generate genome-wide maps of epigenetic modifications — which control how DNA is packaged and accessed within cells — and integrated this with gene expression profiling, to monitor how cells change during the reprogramming process.

Their key findings include:

1. Fully reprogrammed iPS cells demonstrate gene expression and epigenetic modifications that are strikingly similar, although not necessarily identical, to embryonic stem cells.
2. Cells that escape their initial fail-safe mechanisms can still become stuck in partially reprogrammed states.
3. By identifying characteristic differences in the epigenetic maps and expression profiles of these partially reprogrammed cells, the researchers designed treatments using chemicals or RNA interference (RNAi) that were sufficient to drive them to a fully reprogrammed state.
4. One of these treatments, involving the chemotherapeutic 5-azacytidine, could improve the overall efficiency of the reprogramming process by several hundred per cent.

“A key advance facilitating this work was the isolation of partially reprogrammed cells,” said co-author Jacob Hanna, a postdoctoral fellow at the Whitehead Institute.

“We expect that further characterisation of partially programmed cells, along with the discovery and use of other small molecules, will make cellular reprogramming even more efficient and eventually safe for use in regenerative medicine.”