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Reprogramming: what’s next?

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The search for the ideal reprogramming method has been ongoing in many stem cell laboratories since the famous Yamanaka paper was published in 2007 on reprogramming human somatic cells to iPSCs using retroviral methods. Although this method was a breakthrough for the stem cell field, it also showed a low reprogramming efficiency and gave rise to potential medical safety problems with the random integration of oncogenes into the genome. Since then, many different non-integrative approaches have been used to replace the viral factors, but none of them have been able to do this with a high efficiency.

This year, an in-depth paper was published in Cell Stem Cell (Warren et al., 2010) that raised the possibility of an efficient and truly non-integrative system to reprogram somatic cells using synthetically modified mRNA. There were many pitfalls in using mRNAs to reprogram somatic cells including the instability of unmodified mRNA, the short half-life of mRNAs in the cell, and the intracellular anti-viral responses caused by transient introduction of these mRNAs into the cells.

This paper describes the problems that occur when attempting to use synthetic mRNAs for reprogramming and explains the different approaches used to overcome these hurdles. The result is an efficient method for reprogramming somatic cells to induced pluripotent stem cells that takes advantage of using hypoxic conditions, prevents the cells’ antiviral response using a recombinant protein for interferon inhibition, and uses a five-factor reprogramming cocktail of modified mRNAs. This modified RNA protocol was able to reach a maximum efficiency of approximately 36 fold higher than their achieved efficiency using the Yamanaka retroviral factors alone. The researchers also used their newly modified mRNA protocol to direct differentiation of their RNA-derived iPSC lines into the terminally differentiated myogenic cell fate. The experiments performed in this paper demonstrate the potential of modified mRNAs in reprogramming and in directed differentiation. As is always the case with a novel method, other labs will need to repeat this approach and see if it is robust enough to catch on as a widely used technique.

In order to take the scientific breakthroughs in stem cell research to the clinical level, it is imperative to have reprogramming and differentiation methods that do not cause any serious genomic abnormalities. The methods developed by Dr. Rossi and his lab appear to be the closest that stem cell researchers have come to achieving the ideal reprogramming method for use of the resulting iPS cells in the clinical setting for regenerative medicine.

 

References

Warren, L., Manos, P.D., Ahfeldt, T et al. (2010). Highly efficient reprogramming to pluripotency and directed differentiation of human cells with synthetic modified mRNA. Cell Stem Cell 7, 618-630.

Also see related article ‘Reprogramming the methods that induce pluripotency