Friday, November 16, 2007
Monkey See, Monkey Do:
Primate SCNT Stem Cells Have Arrived - Now What?
BY INSOO HYUN

Last June in Australia, during a last-minute unannounced presentation at the Annual Meeting of the International Society for Stem Cell Research (ISSCR), a U.S. research team declared that they had derived two stem cell lines from cloned rhesus monkey embryos. The conference audience’s reaction at that time could best be described as “cautiously excited.” Fresh from the sting of the Korean stem cell scandal, scientists had taken to heart the old adage that extraordinary claims require extraordinary proof. Attendees were quick to note that the team’s results were unpublished, and that there would, of course, have to be independent confirmation that these primate stem cell lines were indeed derived via somatic cell nuclear transfer (SCNT).

Now, almost exactly five months later, this research team, led by Shoukhrat Mitalipov at the Oregon National Primate Research Center, has published its study online in Nature. In accordance with the Nature editors’ post-Hwanggate recommendation that cloning researchers seek independent verification of their results, Mitalipov and colleagues had their work confirmed by stem cell experts from Australia’s Monash University. Careful genetic analyses of the two primate stem cell lines (named CRES-1 and CRES-2) showed that they were genetically identical to the male cell donor nucleus (a rhesus monkey named Semos) and that the mitochondrial DNA originated from different oocytes. The Monash scientists’ analyses ruled out the possibility that CRES-1 and CRES-2 could have been derived from IVF blastocysts (as was the case with Hwang’s purported “patient-specific” stem cells) or through parthenogenetic oocyte activation (which explained  Hwang’s first “cloned” stem cell line, as George Daley’s team at Harvard recently discovered).

So why did Mitalipov succeed where so many others before him had failed? Part of the explanation is just that Mitalipov is a skilled cloner. Already an expert in the idiosyncrasies of human and nonhuman primate cloning, he refined his SCNT techniques further by working with star cloner Qi Zhou of the Chinese Academy of Sciences. Second, Mitalipov used a new, gentler way of removing the maternal chromosomes from unfertilized eggs. Instead of using a dye and ultraviolet light to locate and remove the eggs’ DNA – conventional techniques that Mitalipov’s group says damages eggs – they used Oosight, a machine that generates dye-free DNA spindle imaging to support rapid and highly efficient enucleation of primate eggs. Thus it was a combination of raw technical know-how, nimble manual dexterity, and refined enucleation techniques that produced the breakthrough.

So now what? Are we on the verge of generating human SCNT stem cells? And has the door swung wide open to the biological possibility of human reproductive cloning? “Why not?” one might say. After all, monkey see, monkey do.

I believe, however, we should resist the temptation to jump to such hasty conclusions. There are two key facts surrounding the Mitalipov team’s research that deserve our careful attention.

First, despite the team’s advance in primate SCNT, the derivation efficiency for primate SCNT stem cells remains very low. Mitalipov used 304 primate eggs to generate CRES-1 and CRES-2. That is an efficiency rate of 0.7%. Assuming that Mitalipov’s modified SCNT techniques are applicable to humans, this low efficiency rate provides good reason to try to improve overall primate SCNT embryo generation and stem cell derivation rates before researchers can justifiably ask women to endure the burdens of providing fresh oocytes for human SCNT attempts.

On a more practical note, efficiency also needs to be improved for the simple reason that the availability of fresh human oocytes for stem cell research is extremely restricted. The United States’ prohibitive “zero tolerance” policies for compensating oocyte providers for research has, to date, resulted in zero “compassionate donations.” Thus it is unlikely that American scientists are on the verge of producing human SCNT stem cell lines using the Mitalipov approach. Without human eggs, American scientists will have to employ other methods to try to produce patient and disease-specific pluripotent stem cells, such as using “Yamanaka factors” to induce pluripotency in human skin cells, or using Kevin Eggan’s nuclear reprogramming technique of transferring somatic cell chromosomes into zygotes or blastomeres that have had their own chromosomes removed. Both of these techniques have worked in mice, but it is unknown whether they will work in humans.

What about the prospect of human reproductive cloning? Are we now any closer to that feared possibility? Apparently not. It should be noted that the Mitalipov team generated 213 cloned rhesus monkey embryos, and 35 of these developed to the blastocyst stage (the developmental stage in which stem cells emerge). However, Mitalipov reported that the primate SCNT blastocysts demonstrated poor morphology and have, so far, failed to produce a single pregnancy despite his team’s considerable efforts at reproductive cloning. Last April the Mitalipov group transferred 77 cloned monkey embryos into monkey surrogates but, again, there were no pregnancies. These results appear to support an argument I made last year in the Hastings Center Report that it might be possible to derive primate (human) SCNT stem cells without the corresponding threat of simultaneously enabling primate (human) reproductive cloning. As Mitalipov and colleagues explain in their current article, the biological requirements for reproductive primate cloning are different from primate research cloning for stem cells in that implantation and a normal trophectoderm are not required for the latter.

As a next step, Mitalipov and colleagues intend to generate disease-specific primate SCNT stem cells by genetically altering the DNA of rhesus monkey somatic cells before transferring them into enucleated monkey eggs. If disease-specific primate stem cells can be generated in this way, maybe the public and some of our elected officials will become more accepting of the prospect of human research cloning for stem cells. Perhaps even a few “compassionate donors” will volunteer to provide oocytes for stem cell research. Only time will tell. But given the rapid pace of primate SCNT, we may not have to wait long. And that’s good news for us monkeys who don’t like waiting.

Insoo Hyun teaches bioethics at Case Western Reserve University School of Medicine. He is chair of the ISSCR Ethics and Public Policy Committee.

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