Researchers have attained the holy grail of cellular reprogramming: inducing pluripotency without using any DNA-based materials. Using only a cocktail of purified proteins and a chemical additive, investigators have generated induced pluripotent stem (iPS) cells that don't carry the potential burden of unexpected genetic modifications, according to a new study published online today (Apr. 23) in Cell Stem Cell.

iPS cells Image: flickr/CIRM

"This new advancement is both exciting and startling," Huck-Hui Ng, a stem cell researcher at the Genome Institute of Singapore who was not involved in the study, said in an email. "Now, cell reprogrammers are armed with a potentially genome-safe method to make pluripotent stem cells."

Three years ago, Kyoto University's Shinya Yamanaka showed that adult mouse cells could be reprogrammed to embryonic-like stem cells by transfecting the cells with retroviruses containing just four transcription factors. Since then, many researchers have developed other techniques that avoid permanently altering the genome, including using non-integrating adenoviruses, transient plasmids, and a piggyBac transposon system, but all these approaches involve introducing foreign DNA in one form or another. Several investigators have also combined genetic approaches with using small molecules to enhance or simplify the reprogramming process. But now, in a stem cell first, the genetic ingredients have been eliminated altogether, which brings researchers one critical step closer to moving iPS cells into the clinic.

Sheng Ding of the Scripps Research Institute in La Jolla, Calif., and his colleagues used bacteria to grow batches of specially modified versions of Yamanaka's four original reprogramming factors in protein form. The researchers then purified the reprogramming proteins and added them directly to mouse embryonic fibroblast cells in four repeating cycles over the course of a week. The four proteins alone couldn't reprogram the skin cells, but after Ding's team also added valproic acid, a histone deacetylase inhibitor that Harvard University's Douglas Melton showed last year could enhance reprogramming efficiency, they successfully generated stem cells.

Although the technique was much less efficient than virus-based approaches -- 0.006% compared to 0.067% using Yamanaka's original method -- these reprogrammed cells, dubbed "protein-induced pluripotent stem cells," or piPS cells, passed all the benchmarks of pluripotency both in vitro and in vivo. Ding's team also showed that they could do away with one of the proteins, c-Myc, although this further reduced the already poor reprogramming efficiency by about a third.

"This is the first proof of principle demonstration that [protein induction] actually works," Ding told The Scientist. Now, stem cell researchers and protein biochemists "will jump on this and substantially improve" the method. Ding also said that he has unpublished results showing that piPS cells can be generated from adult mouse and human fibroblasts.

Ding's method is "arguably the safest way" to create iPS cells, said Stephen Duncan, a stem cell researcher at the Medical College of Wisconsin in Milwaukee who did not contribute to the research findings. "If you don't have DNA in the first place you're not going to have DNA integrating and possibly mutating."

Ding said that the protein-based technique is simpler and faster than "technical and tedious" genetic approaches. Duncan, however, disagreed. "It's a reasonably elaborate procedure to generate active proteins and it's much easier to [reprogram cells] using plasmid technologies," he said. "If I was given the choice, all else being equal, I would definitely go with plasmids."

Ding's piPS cell technology and specialized cells derived from the cells will be commercially available through a new business partnership announced today between Fate Therapeutics, a San Diego-based biotech company, and Stemgent, a stem cell reagent supplier with dual headquarters in Boston and San Diego. The collaborative program -- called Catalyst -- will also sell iPS cell technology developed by the Whitehead Institute's Rudolf Jaenisch. Both Ding and Jaensich are members of the scientific leadership of the two partnering companies.


Related stories:

Pluripotency via plasmids
[26th March 2009]

Piggybacking to pluripotency
[1st March 2009]

Single-factor stem cells
[5th February 2009]