Oregon Health & Science University researchers have for the first time generated stable lines of human embryonic stem cells via somatic cell nuclear transfer.1 Whether the platform can carve a niche among existing stem cell-based techniques will hinge on how the cells compare with those generated through other approaches.

Generating embryonic stem cells (ESCs) via somatic cell nuclear transfer (SCNT) involves taking an unfertilized oocyte, removing its nucleus and then transplanting the nucleus of a somatic cell into the enucleated oocyte. The resulting cell is then activated and allowed to divide until a blastocyst is formed. ESCs are collected from this blastocyst and used to establish cell lines.

Researchers in the stem cell space have previously reported on the use of SCNT to generate ESC lines from a range of lab animals,2 including nonhuman primates in a study led by Shoukhrat Mitalipov and published in Nature in 2007.3

Mitalipov is an associate scientist in the Division of Reproductive and Developmental Sciences in the Oregon National Primate Research Center at OHSU.

The next hurdle was to use SCNT to generate ESC lines from human cells. However, early attempts to do so were not successful because human cells generated through SCNT typically stopped dividing after only a few rounds-a phenomenon called early embryonic arrest.4,5

SCNT had been further sidelined owing to the limited supply of donor oocytes and the advent of induced pluripotent stem (iPS) cell technologies.6 The latter are easier to use, more scalable and subject to fewer funding restrictions and ethical considerations than SCNT.

Now, Mitalipov's group at OHSU has reported a protocol that enables the generation of stable human ESC (hESC) lines from cells obtained via SCNT. The researchers used SCNT to fuse fibroblasts from a human cell line with enucleated donor oocytes and then activated the resulting cells.

In culture medium containing caffeine, a subset of the activated cells continued to divide past the early embryonic stage and formed blastocysts. The researchers were able to derive stable ESC lines from these blastocysts. The OHSU group reported in 2007 that adding caffeine to culture medium improved the development of SCNT-generated nonhuman primate cells into blastocysts.7

In the current study, the resulting ESCs expressed known pluripotency markers, formed teratomas when injected into mice and inherited the genome of the donor fibroblast.

Importantly, and contrary to earlier assumptions that deriving an hESC line by SCNT would require unfeasible quantities of donor oocytes,8 the researchers were able to derive at least one ESC line per round of oocyte donation. The result suggests that the SCNT approach could be scalable.

The team's results were published in Cell.

Comparative metrics

The key question is what role SCNT-derived ESCs might have among existing stem cell-based techniques (see "Stem cell types and methods for stem cell generation"). It is probable that the SCNT approach will need to play catch-up with its peers.

iPS cells have already begun to take root in disease modeling and drug screening. For example, Cellular Dynamics International Inc. markets a suite of human iPS cell-derived cell lines and related services for such applications, and iPierian Inc. is using its in-house iPS cell technology platforms to aid the discovery of new therapies.

Researchers will generally want to see whether SCNT-derived hESCs have properties that would make them superior to iPS cells, other hESC lines or tissue-specific stem cells in a particular therapeutic or nontherapeutic setting.

To sort this out, researchers need to first comprehensively characterize SCNT-derived hESCs and compare them with stem cells generated by other approaches.

The New York Stem Cell Foundation CEO Susan Solomon told SciBX that the foundation and its collaborators are doing just that. "We're already conducting comparative cell studies on SCNT-derived ESCs, iPS cells and other ESCs to characterize their similarities and differences," she said. Solomon declined to disclose details about the origin of the SCNT-derived ESCs.

One area in which SCNT-derived hESCs could potentially shine is in the generation of genetically matched tissues for transplantation. Cells derived from current ESC lines are not genetically matched to the patient, which decreases their suitability for use in long-term grafts, in which transplanted cells need to persist and remain functional.

Indeed, Solomon thinks that hematopoietic stem cell transplantation might be an area in which SCNT-derived hESCs could have utility given the high cost and difficulty of finding a genetic match.

"If researchers were able to develop a way to safely derive hematopoietic stem cells from patient-matched ESCs, it would likely result in a less costly and more efficient approach than trying to find a match through a registry," she told SciBX.

Another niche area to consider-and one in which SCNT-derived hESCs could have a potential advantage over iPS cells-would be in patients who have diseases caused by mutations in mitochondrial DNA, said Natalie DeWitt, special projects officer at the California Institute for Regenerative Medicine.

She noted that SCNT-derived ESCs generated from such patients should remain genetically matched but have the oocyte donor's mitochondrial DNA, which would presumably be free of disease-causing mutations. In contrast, iPS cells generated from such patients would retain their mutant mitochondrial DNA.

DeWitt said that she wanted to see studies that involve generating SCNT-derived ESC lines and iPS cell lines from the same individual followed by detailed characterization studies of the cells. She noted that such studies will help the field to better understand the pathways and mechanisms that mediate the reprogramming of cells to a pluripotent state and also provide insights on how to improve iPS cell reprogramming.

She also wanted to see comparisons between the genomic integrity of SCNT-derived ESCs and that of iPS cells.

DeWitt thinks that one of the major barriers to the development of a commercially viable platform for generating SCNT-derived hESCs is the need for large quantities of human donor oocytes.

"Unless a way to create large quantities of human oocytes in vitro is also developed, I think it would be tough to build a commercially viable platform based on nuclear transfer," she told SciBX.

Solomon added that the efficiency of the SCNT approach will also be a key determinant of whether others in this space will want to pick up the technology.

  OHSU has pending patents covering the use of SCNT to generate stem cells for therapeutic application. The technology is available for licensing.

Lou, K.-J. SciBX 6(20); doi:10.1038/scibx.2013.481
Published online May 23, 2013

REFERENCES

1.   Tachibana, M. et al. Cell; published online May 15, 2013; doi:10.1016/j.cell.2013.05.006
Contact: Shoukhrat Mitalipov, Oregon Health & Science University, Portland, Ore.
e-mail: mitalipo@ohsu.edu

2.   Yamanaka, S. & Blau, H.M. Nature 465, 704-712 (2010)

3.   Byrne, J.A. et al. Nature 450, 497-502 (2007)

4.   Egli, D. et al. Nat. Commun. 2, 488 (2011)

5.   Noggle, S. et al. Nature 478, 70-75 (2011)

6.   Patel, M. & Yang, S. Stem Cell Rev. 6, 367-380 (2010)

7.   Mitalipov, S.M. et al. Hum. Reprod. 22, 2232-2242 (2007)

8.   Daley, G.Q. & Solbakk, J.H. Nature 478, 40-41 (2011)

9.   Anokye-Danso, F. et al. Cell Stem Cell 8, 376-388 (2011)

10. Ho, R. et al. J. Cell Phys. 226, 868-878 (2011)

11. Nishimura, K. et al. J. Biol. Chem. 286, 4760-4771 (2011)

12. Stadtfeld, M. & Hochedlinger, K. Genes Dev. 24, 2239-2263 (2010)

COMPANIES AND INSTITUTIONS MENTIONED

California Institute for Regenerative Medicine, San Francisco, Calif.

Cellular Dynamics International Inc., Madison, Wis.

iPierian Inc., South San Francisco, Calif.

The New York Stem Cell Foundation, New York, N.Y.

Oregon Health & Science University, Portland, Ore.

Oregon National Primate Research Center, Portland, Ore.