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Thursday, November 16, 2006


Session 2: Stem Cell Research Update and Alternative Sources of Pluripotent Stem Cells

Council Discussion

DR. PELLEGRINO:  Floyd, may we turn the meeting over to you?  Would you like to comment from there or up at the  podium?

DR. BLOOM:  I can do it from here just fine.

DR. PELLEGRINO:  Okay.  Thank you.

DR. BLOOM:  I want to start by thanking Dan and Ed for giving me the chance to relive the last three years of the President's Council and go through the enormous literature that you've produced on the controversies in embryonic stem cell research.

And I was reminded in so doing that in 1997 the Thompson paper appeared in Science while I was editor, and we wrote an editorial on publishing controversial research, not realizing at the time how really controversial the entire topic area would be.

I want to congratulate you, Dr. Schöler , on such an intellectual inspiring and graphically advanced presentation.  I had thought from the papers that were sent to us on your work that you were going to emphasize cell fusion.  So in a minute I'll ask you about that, but in fact, what you've done is give us a great introduction to the next hour's worth of work of reexamining the progress that's been made across the field.

My questions for you really start with your own opening slide where you said that you were trying to develop the oocytes to the point where you could do somatic cell nuclear transfer into them.

But then you explored all of the range of options from cell fusion to embryonic cancer cell extracts, to small molecules that can cause de-differentiation.  I find the whole concept that you can de-differentiate a  somatic cell into a pluripotent cell such an astonishing biological result that it is really hard to imagine how that can take place scientifically and under control.

What you've shown us though is that the research is advancing very, very broadly across a wide range of mouse embryonic stem cell opportunities.  And my first question for you is regardless of whether you take fusion or somatic cell nuclear transfer, how much of what you've talked about in the mouse can we imagine in the near future taking place with any of the human embryonic cell lines that exist for which there is the opportunity to do research?

And if so, which are the ones that are most likely to be successful?

DR. SCHÖLER:  I think that the papers that have been published so far show that things that have been developed in the mouse system to a large extent can be transferred, can be also repeated in the human system.

I think Kevin Eggan's paper is a wonderful example for that.  By using human embryonic stem cells, you can reprogram adult somatic cells.

The nuclear transfer that is working now very efficiently in mouse might be a much bigger hurdle in human, and I think there may be at the end intelligent ways of reprogramming by fusion or by using a cocktail of factors, which will be faster, we will see.

We have to see because there's not a lot of things I learned from Woo-Suk Hwang's paper, but one thing I learned:  that he used 2,221 or so oocytes, and still it did not work.  So it's kind of an negative result, but it tells us a lot.  We have to improve the procedure, and I think he had some points that he made that will help future researchers how to in an intelligent way go on with that type of research.

Personally I'm always saying that if he would have worked together with James Thompson to derive embryonic stem cells from clones, a person that knows what he's doing, not repeating something that others have, I think he had the wrong collaborators to derive it.  That's my personal understanding.  He might have had embryonic stem cells at the end.  I might be wrong, but that's what I think.

But nuclear transfer at the end might turn out to be much more difficult.  Maybe Gerald Schatten with his Brevia published in Science at the time was more right than he afterwards believed himself.

But I don't see why the procedure published by Shinya Yamanaka should not work. Maybe you need to exchange one or the other player, but in principle, that procedure I think will work.

From what we and colleagues are doing, we're actually thinking in a different direction that maybe even factors that you get from Drosophila, from Planaria, and so on can do a lot of the job with respect to reprogramming.

This is still something of which I think there are still some parts missing in the picture.  I think that's what Orkin's laboratory with their beautiful paper in Nature is showing, is how complex interactions, protein-protein interactions and so on are.

So if you just take the middle player and put it into a somatic cell, you might have problems with efficiency.  You might have problems with really reprogramming the cell, but this was such an important step into the right direction, I think, that others will — if he's not doing it, others will fill the gaps to get to a more pluripotent state.

And as I don't see why this should not be possible with embryonic stem cells, I don't see why this should not be possible with the existing human embryonic stem cells.

From my perspective, with respect to basic science, understanding the general principles, I think you can get very far with the existing embryonic stem cell lines.  There are certainly problems if you would like to think about therapies, if you want to go into that direction.—

DR. BLOOM:  Maybe we just focus on that because we're going to spend this next hour talking about all of the areas of advances, and the science is clearly advancing.  There's no question about that, but for this Council's purposes, the really important questions have to do with if you have to start with a human embryonic stem cell, we're back where we were.  If you have to start with a human oocyte, we're back in the supply business where we were.

So we're not going to debate the science with you because the science is going to do what it is, but if you had to put your resources into the most likely place of advancing to achieve regenerative medicine potential without the destruction of a human embryo, where today would you put your resources?

DR. SCHÖLER:  Basically, I would try to go along with the three major areas that I've described.  I think at this stage we just don't know which is going to be the most successful.

I think that if you are reprogramming a somatic cell and you have to go back to an intermediate stage, a pluripotential cell is an intermediate stage, it's not going to be as perfect as if you go all the way back and then go again to that intermediate stage because here you would have a full erasure, and then you would come to a stage and the different layers of gene regulation are then established.

If you go back to reprogram a somatic cell to become a pluripotential cell, I think we will find out more and more problems.

So in respect to therapies, if I would bet, I think you have at this stage to use oocytes, and we can discuss the sources for the oocytes, but going back and then to that stage is something that is the only way so far based on the mouse work that is giving pluripotential cells, which have the same quality as embryonic stem cells derived from IVF embryos.

And so all of the other things, is exciting basic science, but in terms of if you ask me where I would fit, I would use oocytes, and that would mean you would need to derive new cell lines.

DR. PELLEGRINO:  Thank you very much.

We are now open to questions from the Council.  Dr. Gazzaniga.

DR. GAZZANIGA:  In your Cdx2 example, was it not the case that you were fusing an oocyte with a male sperm, that first slide you showed?

DR. SCHÖLER:  Yes.

DR. GAZZANIGA:  And then you introduced the micro RNA to stop the processes and develop the two classes.  The reason to be addressing these problems is to try to get around certain moral questions that people have.  Why wouldn't the people that have those concerns not be happy with that approach either?  Because you're basically taking an entity that could be developed into a human, an animal, a mouse in your case, and therefore, it has all of the problems of somatic cell nuclear transfer and all of the rests.

DR. SCHÖLER:  So first of all, I would like to stress that we have been using this specific stage and there's no reason for us to believe that we won't be able to use this approach also as an earlier stage, like the metaphase II stage for a nuclear transfer.

It's a different approach than what we have done in comparison to the one that has been proposed in the ANT procedure, where the nucleus has been changed.  We are introducing this to the oocyte.  So the genetic material has not changed.

We have not done this experiment for the sake of developing an ethically sound way, but we have to do this for the science and we found that there are some parts there that might be interesting with respect to the ethical problems that we have or many people have with that type of research with embryos.

So, first of all, that's something which I don't think is fixed to that specific stage.  The reason why I think this isn't part of the solution is because I don't see it as an embryo.  This is like cell division.  It's like a cell that is dividing.  It's not giving rise to an embryo, and I've tried to make the point that even if you look at the whole genome profile, and you have to do this if you really want to compare one to the other, you see tremendous differences, and they get bigger and bigger because just not having this second lineage.

And if you agree with the fact that an embryo at that stage requires a trophoblast and inner cell mass, from the viewpoint of a development biologist, this is not an embryo.

So if you say that you're manipulating something that ought to be like a fertilized oocyte, ought to be an embryo and you're changing that from what you're doing, you won't convince somebody who has a problem with doing that.

But if you extend this really to the other direction, combine this, for example, with ANT, I mention this type of research because I think it's giving you some ideas about what's happening with the embryo, regardless if you're doing nuclear transfer according to the ANT or if you inject this in the oocyte or in later stages.

Just to give you a complete picture because so far people are just saying these embryos fail, and you have no idea what we're talking about.  Now I think if we ask for this, you get a clearer picture that they're failing because the inner cell mass needs support.  We think ATP is of real importance for this embryo to survive.

And what we have been doing, we have been replacing the trophoblast, which is not present because this is not an embryo, by the feeder layer, and if it only works that you get embryonic stem cells, if you put them on a feeder layer. The feeder layer is nurturing the pluripotential cells, this then gives this nice ball of green cells, and then you can derive very efficiently embryonic stem cells.

That is my understanding what is going on here.  We're trying to or we might have succeeded in supporting what we think maybe is supporting also this ANT procedure.

Did this answer your question?

DR. GAZZANIGA:  It's kind of fascinating because if I recall correctly, basically human embryonic stem cell research cannot go forward in Germany presently, and so you are taking opportunity to advance stem cell biology in various animal models, and one of the unintended consequences is that maybe German biology is actually advancing the understanding of what's actually going on in these incredible phenomena like de-differentiation, where we have sort of slowed down in that within our own country.

DR. SCHÖLER:  May I disagree on this point?   I think that —

DR. GAZZANIGA:  Am I wrong?

DR. SCHÖLER:  I think that the last papers that I've shown, Stuart Orkin, Rudolf Jaenisch, Ihor Lemishka, they are going to the key points of what a pluripotential cell is, and what I think is very interesting, that Stuart Orkin, Ihor Lemishka, and also Irving Weissman, these are people who have been working all their life with hematopoietic stem cells and now putting a lot of their resources into trying to understand embryonic stem cells and trying to understand what pluripotential cells mean.

And with all of their experience that they have developed with human embryonic stem cells, they now kind of dive into that topic in a way that is very, very astonishing, and this is from this country.  It's not from Europe.  You have such a great amount of science here in this country.

So even if they are more than Germany into working with human embryonic stem cells, we still can do work with human embryonic stem cells in Germany.  I also want to make that point clear.  It's just that it's like four, five months later, the cell lines that we can use, and the presidential cell lines were from the first of January 2002, the ones that we can use in Germany.

And we have a discussion now, a big discussion if that date is going to be shifted or going to be dropped.  I have here this what has been presented last Friday from the DFG, the major German funding agency.  Ernst-Ludwig Winnacker has had a press conference where this has been presented, (speaking German) "Stem Cell Research in Germany, Possibilities and Perspectives."

But he has been asking in Germany to drop this date..., not even to push it, and there's a lot of discussion.  I just received an E-mail this morning that our chancellor, Angela Merkel, has been looking very positive into that something has to be changed.

So we have a discussion, a big discussion in Germany.  It still won't be possible that we as scientists would be allowed to derive our own stem cell lines.  We only also in the future would be able to import them.  That's for German scientists.  It's a problem when interacting with scientists from other countries, who say, "Why don't you do it yourself?"  This is a problem, but we have to live with that.

DR. PELLEGRINO:  Professor Schöler has kindly agreed to send us a translation of that paper which we'll distribute to the members of the Council.

Thank you very much.

We have three commentators waiting, Dr. George, Dr. Foster, and Dr. Kass, in that order.

PROF.GEORGE:  Dr. Schöler , I just would like to ask a brief follow-up to Dr. Gazzaniga's question.  From your description, it sounds to me — and I'm asking you to correct me if I'm wrong about this — from the description you give, it sounds as though not only would you not have an embryo, something that would qualify as an embryo, but the lack of integration and self-direction or capacity for self-direction along a developmental trajectory in the direction of maturity such that you wouldn't really be able to say you have an organism of any sort here.  It just isn't an organism.

So my question is is it right to say not only is this not an embryo.  It's not an organism.  Do you see so far as to distinguish between an embryo and say a non-embryonic organism?

DR. SCHÖLER:  So first of all, I would like to apologize that I've been using the term "embryo" in a very loose way.  I'm in a way caught in a situation that whenever I give this structure a name, I'm being accused of trying to hide something.  So if I call this "structure" or if I call this "pluripotency ball," somebody stands up and says, "Why don't you call it embryo?"

Actually I always try to say this is a stage which corresponds to the eight-cell embryo, and during a talk I might lose this and not [always stress that] this is a stage that corresponds to the blastocyst stage or this corresponds to that.

But I hope that you forgive me if I have not in every case done it that very way during my talk, but that's what I meant.  So whenever I had this comparison, I wanted to point to the embryonic stage.

So I don't consider this an embryo.  I don't consider this an organism anyway.  No, I think this is, for me, this is cleavage.  This is cell division of a structure that starts off as an oocyte, but it would not cleave and divide to give rise to an organism.

PROF.GEORGE:  Am I correct that in thinking about what is and is not an embryo, what is and is not an organism the focus should be on whether we have a self-integrating entity that is developing along a trajectory in a direction?  Is that the correct way to think about it?

So that if we see a lack of integration and a lack of self-directed development along a certain trajectory that we associate with the normal developmental trajectory of the species, if we see a lack in these respects, that would be the ground for judging that we have here something that is not an embryo, that is not an organism.

If, on the other hand, we find that there is a degree of integration, that there is self-development along a trajectory,  we would conclude that we have an embryo although in a particular case the embryo may be damaged, the embryo may have defects that will prevent its full manifestation of its potential.  There may be an impossibility of implantation.  There may be an impossibility of survival so that we might not have viability, but we would still have an embryo as something as distinct from something that's not an embryo.

Am I looking at it in the way that you would advise?

DR. SCHÖLER:   I think one would have to look at it exactly the way you have been describing it because otherwise, if you would not be looking at that that way, you could conclude that an embryo that will fail at some stage, like in mouse there's this one mutation, Lim-1, which gives rise to fetuses without a head, and you could say this doesn't have any potential.  The answer would be: why don't we produce these and at the end we can use their organs, which I think would be not in agreement with the view that you have been presenting.

I think the view that you have presented is the view I would see it as well.

PROF.GEORGE:  Thank you.

DR. GAZZANIGA:  But were you here for the presentation?

PROF.GEORGE:  I got here late.

DR. GAZZANIGA:  So he missed the key slide, which is the fusion of the oocyte with the fertilization, and that occurred first and then the intervention of the micro RNA that caused for the two classes of embryo.

So you're starting out with something that is on that trajectory, and you are interfering with it to produce this artifact that can be used in the way that has been demonstrated.

So the question is that I thought would offer you a problem in the final analysis.

PROF.GEORGE:  So then the question would be does Dr. Schöler agree that what he's beginning with is an embryo that is transformed into a non-embryonic condition.

DR. SCHÖLER:  A [pronuclear] zygote to my understanding is not yet an embryo.

DR. ROWLEY:  But it is an oocyte.

DR. HURLBUT: When you gave your presentation, you were talking about the use of pronuclear stage, but in answer to Mike Gazzaniga's previous question, I understood you to say that the same thing would almost certainly work if you did the silencing in the oocyte —

DR. SCHÖLER:  Yes.

DR. HURLBUT: — before nuclear transfer or before whatever procedure, the point being that that would satisfy the full concern about ever creating an embryo.

DR. SCHÖLER:  Yes.

DR. HURLBUT: And some people might argue that a pronuclear stage entity is a embryo.  I know German law says differently, but the point is in America that criterion probably wouldn't be the same.

So to satisfy the American concern, one would have to do the silencing in the oocyte, and you say that you think that's feasible.

DR. SCHÖLER:  That's why we're trying to see if nuclear transfer into oocytes can be [combined with this procedure to imporove embryonic stem cell derivation].  As I tried to explain, we have at this time not done the experiments for getting out of ethical problems, but for scientific reason, and if we would have planned them differently, we would have started at an earlier stage.

DR. PELLEGRINO:  On this point?  I have a list, Bill, of people who are waiting.  You're on this point?

DR. HURLBUT: Yes, on this point.

I just want to clarify that the way we've been using the term "altered nuclear transfer" from the very beginning is that the alteration can be either in the cytoplasm or the nucleus or both.  So the procedure of knocking down the siRNA and the cytoplasm of the metaphase II oocyte would be a form of altered nuclear transfer.  What you've described would be altered nuclear transfer, not just what we would be interested in the nucleus.

DR. PELLEGRINO:  Thank you.

Dr. Foster.

DR. FOSTER:  I didn't have any comment.  I was just glad to get here. 

(Laughter.)

DR. FOSTER:  I was just waving, having waited seven hours to get a flight and then get canceled twice.

DR. PELLEGRINO:  Dr. Kass.

DR. KASS: Well, I want to say hi to Dan Foster and wave, too, and thank Dr. Schöler for really a remarkably exciting and illuminating presentation.

I have really basically two factual questions and then a more theoretical question.  First, these stem cells that you got from the Cdx2 altered cells, have they been tested for pluripotency by the gold standard tests and shown to be pluripotent?  The first question.

And second, neither you nor our colleague Dick Roblin in preparing the materials for this discussion referred to the publicized but not published work of Dr. Verlinsky.  These were fusion experiments done in humans with human embryonic stem cells, and I haven't seen any publications, but I wondered if one knows anything more about this.

Among the things that were striking about that report was that according to his experiments, it was the cytoplasts rather than the nucleoplasts that seem to contain the materials that could successfully produce the reprogramming of the somatic cell with which the cytoplasts were fused.

And then the more searching question has to do with your comment about the superiority of going all the way back to pluripotency, to totipotency, to —

DR. SCHÖLER:  To the oocyte.

DR. KASS: — to the very beginning oocyte and then coming forward.  Since it's clear that for therapeutic reasons one wants to have partial differentiation, as you pointed out at the beginning, to more specialized stem cells so that you don't have the tumorigenic concerns, why if you had a rather controlled process of de-differentiation to some place that was reproducible?  Wouldn't you be — and by these sort of cytoplasmic factors you knew what you were doing.  You knew to what stage you got it .- why does the fact that you haven't gone all the way back to the oocyte present a real liability?

That's a more searching question.  The other two were just questions of fact.

DR. SCHÖLER:  So first of all, I've just briefly scanned over that one slide because I was realizing that I'm talking very long.  So there was one slide that was showing that we have done our homework, that the ES cells derived from the Cdx2 treated pronucleate zygotes, fulfill all of the criteria of pluripotent cells.

The one I've put my head a little bit out of the window was saying that the most important thing is that you show tetraploid aggregation.  That's what we're currently doing.

But all the others, chimerism and so on, that has already been done.  The nice thing about Cdx2 is that it also plays a role for intestine stem cells.  If that would have not been a transient effect, what we have done, the injection into the enucleated zygote and deriving ES cells, if that would have been a more stable effect, there should have been a problem with these intestine cells, and I've only shown you a picture with an intestine where you saw some blue cells.  This was from a ten months old mouse where we waited that long to make sections, to show that this cell compartment is produced by derivatives of these injected cells.  So this is transient, which is the best indication that it's not a long term effect.

And if you think about what's going on is you're reducing Cdx2 in a compartment that might not even give rise to the embryonic stem cells.  It's an open question.  If the outer cells are confused cells because they still carry on material from the oocyte or if they, indeed, can be used to derive embryonic stem cells, we don't know that.

But by all means, we can get germ cells from these after injection and so on.  We think that they don't have a problem.

The second question, Verlinsky's and Strelchenko's work that they have published in the meantime in Reproductive Biomedicine Online — in the meantime means, I think, half a year or so ago — I think if you read the publication, first of all, we initially only knew about that work from, I think, The Scientist, that was citing a patent. I have then the patent.  We didn't get an idea of what was specific about how they did it, and I didn't get it from the paper very well.  So I asked them how they exactly did it [and neither did I get it later from the paper very well].

And the way I understand it is that they are doing this fusion on glass plates where the nuclei have been removed by centrifugation.

DR. KASS: Yes.  They plate, I think, the embryonic stem cells —

DR. SCHÖLER:  Yes.

DR. KASS: — on little glass cover slips.  They centrifuge them upside down.  The nuclei come out.  They're left with the cytoplasts and they fuse the somatic cells if I'm not mistaken.

DR. SCHÖLER:  Yes, but what I couldn't get from the paper is that when they do this and do the fusion, if they can exclude that this on the cover slip is forming something like a syncytium where you have cells fusing, and since the removal of nuclei is not complete, that you could have nuclear factors coming from the leftover nuclei and doing the reprogramming.

And since the efficiency is extremely low, that's something, which is clear from the paper, you don't know what actually did the job.

I think that what we had [originally] published, and I just briefly mentioned our publication, where we have been using nuclei, we were hoping that the cytoplast [would suffice].  That would have been easier.  [Imagine] you send vesicles to the clinics and they put in the nucleus and you get your embryonic stem cells.  That would have been a dream.  Maybe with other processes we can still get there.  We don't know.

But that's what we tried.  We took embryonic stem cells apart and adult stem cells apart and recombined everything, and we could make a big sac out of the cytoplasts in a clean way.  Never did we get any reprogramming.  We needed the nucleus to do this.

And the work from Shinya Yamanaka is kind of confirming this because he needs four nuclear factors.  So I don't think that this will turn out to be a key paper, this cytoplast piece of work.

The other question concerning going back and forth again... This [idea] might turn out to be wrong at the end because somebody is doing a clever experiment to prove me wrong, but my understanding is that if you [need to] make tabula rasa, you [need to] clean up everything and then start building things up again.  It's easier than taking things out, things that at this stage we have no idea what you have to take out, the levels of repression that you need to get expression along one or the other lineage.

If you basically clean the table and then allow the things to develop by itself into the right direction, I think the different layers of gene regulation are then set by the oocyte.  They [the oocytes] are doing the job.

If you push it to that point, since you might have more leftovers of things that are not perfectly reprogrammed, I think at the end that would be more difficult, and so far all the evidence is kind of suggesting that I'm right, because whatever people have obtained by the procedures is not as good as nuclear transfer where an oocyte is involved.

Is that okay?

DR. PELLEGRINO:  Dr. Rowley.

DR. ROWLEY:  I, too, want to thank you for a comprehensive overview of the area and the directions you and others are trying to pursue.

I have a couple of questions, one of which is a follow-on of Mike Gazzaniga's with the comments of Dr. Hurlbut and Robby George, and it does seem to me if you have to start with an oocyte and then you knock down a Cdx2, that the moral problems of using oocyte remains regardless of what you've done, and whether you've added siRNA or new nucleus to that oocyte to my mind isn't different.

And so to form that in the way of a question, is it different?  But I'm not sure.

I have two other questions.  One, is it really correct to call the primitive germ cell unipotent?  Because, in fact, if that cell is fertilized, it goes on to give rise to an embryo which gives rise to all three layers, ectoderm, mesoderm and endoderm.

So in my own view, I don't think of it as unipotent.  So that's a question.  Why do you call it or is it correct to call it unipotent?

And then more not philosophical but scientific question.  In your view why is it so difficult to begin to develop somatic cell nuclear transfer in humans or primates as compared with mice or other mammals?

We have the example of the South Koreans where at least at the present time it said that they use more than 2,000 oocytes and then get a single cell line, and there has to be some difference.  Do you have any insights as to what those differences are?

DR. SCHÖLER:  So first to the statement about that not changing the moral, ethical problems by still using oocytes, is this because of the problems of egg donation or is this from your point of view because of that we still have something which was supposed to give rise to an embryo?

DR. ROWLEY:  Well, I should really let somebody like Robby George answer that because I don't personally have a problem, but it would seem to me that for individuals who do have a problem with using oocytes that, in fact, it should still cause problems.

PROF.GEORGE:  Perhaps I could respond on that then.  I think that people on my side of this debate, and I've been a critic of the destruction of embryos for purposes of this research, are concerned not to destroy living human embryos.  So they're not concerned on that issue with oocytes as such, but rather with embryonic human life.

But then on the question of the use of oocytes, the concern is with how they're obtained and whether they would be obtained in a way that's potentially harmful for women and especially if it might result in the exploitation of women to obtain the larger number of eggs that would be required if therapeutic uses were found for these technologies.

So they're really two distinct questions that I think perhaps have been run together.  So that if there would be a way of obtaining oocytes without exploiting women or subjecting them to danger or harm, and if those oocytes could be used to produce embryonic or embryonic type stem cells without the destruction of embryos, then we would be out of the ethical problem as far as I'm concerned.

DR. ROWLEY:  Well, but then what Robby is sort of suggesting is if you go the way of John Gearhart, which is to use oocytes from fetuses and they are therapeutically aborted fetuses, but which raises another issue, but getting oocytes from them, then he has no problem

I think it's not an easy issue.  Certainly the question of egg donation by healthy women who do have to undergo hormonal treatment in order to release a lot of oocytes, that for me is a separate issue.  I agree from the ethical issue of using oocytes.

DR. SCHÖLER:  So with respect to egg donation, then let's say a combination between the ANT procedure and using what is now currently discussed a lot in Germany because of what the group in Newcastle has asked to do, nuclear transfer into bovine oocytes, with respect to egg donation at least, that should be fine; is that right?

PROF.GEORGE:  That's correct from my point of view, yes.  If I understand what you're saying correctly, where you would not use oocytes taken from female humans —

DR. SCHÖLER:  Yes.

PROF.GEORGE:  — you would rather be using non-human, animal oocytes for the procedure, do I have that correct?

DR. SCHÖLER:  Yes.

PROF.GEORGE:  Yeah, that does not strike me as — as long as we're not then creating an embryo, it doesn't strike me as having an ethical problem.

DR. SCHÖLER:  Okay.  So the first question concerning the primordial germ cells, they are considered to be unipotent because on their own they just give rise to germ cells.  At the end you have an oocyte or an egg, which are terminally differentiated cells.  These are the most exciting cells for me, but they are terminally differentiated.

The exciting thing is that if you bring them together, the clock is set back and now you're getting from two unipotent cells... a totipotent one, but as long as they're on their own, they are considered by scientists to be unipotent.

They're still on what I call the totipotent cycle, because they give rise to an organism, and I call it now germline cycle because people have been confused by having cells, which are unipotent, on the totipotent germline cycle.

So that's what I would like to answer with respect to the potency of these cells.

And why is cloning so inefficient?  I think this has a lot to do with what you're depleting when you remove the chromosomes, and the spindle and so on might be something, which is different between the different species.

So that might be a reason for problems.  This has been  nicely described by Gerald Schatten, why he thinks cloning is so inefficient, and I think from what he has been saying at that time, I think he has been correct.

It was just that Hwang, when he came out with this astonishing result, he [Schatten] thought, oh, maybe I was wrong. But I think he was right.

DR. PELLEGRINO:  Dr. Dresser.

PROF. DRESSER:  Thank you.

I had another question about , SCNT in humans, and I understand that one of the main basic science reasons people want to do it is to create stem cells from cells from patients with genetic disease so that they can learn more about the development of the disease and test drugs and so forth.

So, number one, it looks as though that will be difficult to do in humans.  So I wonder about alternatives to that.  I wonder if any of the procedures you describe offer a way to study that sort of problem.

And the other question I had was whether the difficulties with SCNT would lead researchers to want to try to create embryos with genetic problems through IVF and whether that will be an emerging issue to address.

I suppose you have to have it confirmed on people who have genetic disease to try to make an embryo.  That would be a disease model that would be more efficient than SCNT or perhaps some of these other models.

DR. SCHÖLER:  Yes.  So actually to maybe start with the second question. I think if we come back to the [work] of Verlinsky, he has derived an extensive [set of embryonic stem cell lines] from patients with genetic disease.  These, from what I understand, have been obtained by fertilization.  I think he has done preimplantation diagnosis and then correlated this with the disease.

I think this is, of course, far more efficient than doing nuclear transfer.  The big, big advantage if nuclear transfer would work is that you have the history of a patient documented.  You know the outcome of what you're expecting to understand.  If you're doing it with one of these cell lines, you don't know how much genetics is playing a role in the outcome of what you're investigating.

Of course, there are other problems that we might not see at this stage, but if you know the disease, how the disease develops and you're trying to understand this process in the dish, I think that's the best way of going ahead.  It's like the reverse way of what you do in mice.  You destroy the mutated gene and then you're trying to see what happens. 

In this you know what's happening.  This is a disease that this patient is suffering [from], and if you're taking nuclei of, let's say, a handful of patients and trying to derive stem cell lines [from these], you get to some degree a variation of what you can investigate in the dish.

And I think this will be a focused way of looking at disease.  Of course, you can't get to an understanding of what happens in the whole organism, but by correlating this understanding with the understanding that you have obtained from the patient, I think we will learn a lot.  That's what I think will be very important for the future.

Then the —

PROF. DRESSER:  But I guess my question is:  what if that proves to be too difficult?  Say it takes thousands of eggs or .-

DR. SCHÖLER:  Yes.

PROF. DRESSER:  — it's just not feasible.

DR. SCHÖLER:  That was basically your first question.  So for understanding disease, I couldn't do this approach in Germany anyway, from a practical point of view, but also in terms of the way you can screen for the successful results, the embryonic stem cells are — that's my understanding, again, I might be wrong, but that's my understanding —... the most potent system because you can plate cells.  You can [use] selectable markers... 

And since embryonic stem cells have the power of proliferating in a way that you can get from one to millions and millions of cells, you can screen for rare events, and you can't do yjsy with oocytes.  That's one problem that will remain in the future, that you can't really screen for something where you have a limited number of cells to start off with.

So I, indeed, will be trying to see if we can use the different reprogramming procedures that have been described with embryonic stem cells, if we can get disease this way into the dish, which might not be good enough to use these cells for patients, but may be good enough to understand at least certain aspects of disease.

And then a final point to this is that in science it's very important that different people test different approaches.  This is our approach.  It might not work; it might work.  Others might be more successful with nuclear transfer, and we have to see at the end who is successful.  The patient will win if one of the processes is successful.

DR. PELLEGRINO:  I have Dr. Hurlbut and Dr. McHugh.

DR. HURLBUT: Just a comment on what you were saying.  That fertilization would work for certain dominant alleles, but it wouldn't necessary work as well for a lot of genetic diseases because it's the whole context of the existing genome that counts.

So obviously, the ideal would be to create an identical gene aligned from an identical genome, and this is sort of a question and a comment, just because there's something else I want to ask you.

Even with IVF embryos we don't know if we take the cells at the four-to-five cell stage, we don't know if those genomes can actually produce a full organism yet.  So is it something to consider, that even IVF might not produce perfect cell lines?

Why don't you answer that first?

DR. SCHÖLER:  So if we look how efficient preimplantation development is, well, at the end we are disappointed how inefficient it [actually] is.  I'm not sure exactly about the numbers, but it's in the range of, I think, 50 percent or so that are failing prior to implantation.

So if you take two germ cells and combine them and force these to do something that in vivo they might not have a chance to do, you're not really sure about the outcome of what you have in hand at the end.  So that is an important point.

But if you bring that up, you also have that problem with nuclear transfer because the problem with nuclear transfer is that you're skipping all of the selection that you have along gametogenesis because from my understanding, that if you have a problem with germ cells, germ cells basically don't try to repair what's going wrong.  The [organism] gets rid of such germ cells.  There's a lot of apoptosis. [That has been described.]

If you have a gene that is expressed in germ cells, [that is] during germ cell development, it's often that you see they are driven into apoptosis, active cell death. Because my understanding is that [the organism tries] ... to get rid of something [compromised, instead of]... maintain[ing] it to start the next generation. [However], if you're doing nuclear transfer, you're jumping over this.

So you might at the end have something which, as a germ cell, would have had no chance to get to that point.  That's also — we have to talk about both sides.  And in vitro fertilization, actually the problem would be only a short problem, whereas nuclear transfer would be all the way through.

DR. HURLBUT: Yes, but the fact that you can get successful production of full organisms, in this case mice, using tetraploid complementation does confirm that at least some of the time you're getting a fully functional genome.

DR. SCHÖLER:  Yes, some.

DR. HURLBUT: And, by the way, I'm correct, aren't I, that when he did his experiments with altered nuclear transfer, Rudy Jaenisch did do tetraploid complementation studies?

DR. SCHÖLER:  Yes.

DR. HURLBUT: So those cells really were confirmed, and if I understood you earlier, none of the other methods, reprogramming or any other methods have established that you can get tetraploid complementation to work.

DR. SCHÖLER:  Yes.

DR. HURLBUT: Is that right?

DR. SCHÖLER:  That's correct, but also, if you get to embryonic stem cells, you also have a selection for those embryos that give rise to embryonic stem cells.  It's also something you're — if you would try to force embryonic stem cells at earlier stages which the structures don't have a chance to get to the blastocyst stage, you maybe have more problems with these tetraploid complementation experiments.

I'm just saying that you always have to think about the selection you're doing or not doing in that process, and maybe cloning is inefficient because the genetic material, to some extent, is not as good as it is from germ cells.

So there's so many things we still don't know.

DR. HURLBUT: And finally, could you just say a little more about your statistics of increased efficiency of harvesting ES cell lines using the Cdx2 knock-down, was very interesting.  You got practically a doubling of the efficiency and at the eight cell stage, which is earlier than we had previously thought you could get ES cells.

Could you just expand on that a little bit?

DR. SCHÖLER:  Yes.  I realize at the end I have been too fast on that point.  The one astonishing experiment from Robert Lanza, was that actually he can derive ES cells from single blastomeres from an eight-cell embryo...

So it is to me.

DR. HURLBUT: But his paper indicated he co-cultured the blastomeres.

DR. SCHÖLER:  Yes.

DR. HURLBUT: That's important because they were signaling.

DR. SCHÖLER:  I agree, but that [showed] that it works.  So I wouldn't take it as too much of a surprise that if you take an eight-cell embryo and try to derive stem cells from there using also a feeder here in our case, that experiment, which we did also with the control treated one, but we wouldn't have had to do that because they grow on the outgrowing trophoblast cells, and so that would have been the feeders, but we don't have that in the knockdown experiment.  So to compare both, we had to use feeders in both cases.

If you take these cells, these structures from the Cdx2 knock-down experiment, you had green cells throughout the structure, and we think that the increase of potential candidates to give rise to embryonic stem cells at the end started an increase of about 50 percent in efficiency.

So it means if you take the control treated embryos, compare these with the structures, the Cdx2 treatment didn't worsen the derivation, but improved it.  So our hope is that if you do the Cdx2 siRNA treatment at the metaphase II stage for nuclear transfer, that this would also even result in an increase.

Maybe this way we make nuclear transfer more efficient with respect to embryonic stem cell derivation.  That we have to see.  We don't know.

DR. PELLEGRINO:  Dr. McHugh.

DR. McHUGH:  I, too, want to thank you very much for your presentation, and I'm still thinking about it, and so I'm the slow member of the class trying to ask some simpler questions really simply, in part because of the important issues that you evoke for me, anyway, and for many other people about our existing science, our science base, and the directions we're going.

And so I just wanted to ask simply three or four questions that came out from what you said.  The first thing you said was that, you know, using existing embryonic stem cells we're really quite able to do wonderful things, and you said we have kilograms of these cells now available.

Now, does that mean that, in fact, the cells that were made available really at the birth of this Council by President Bush when he said that he would give federal funding, anyway, to research that based itself on those cells, that in point of fact those cells are adequate for the work that is being done?

Are they adequate in number and in character?  That's the first question.

The second question is, of course, like everyone else I'm very amazed and think it is a wonder, this Cdx2 business.  I want to be really sure that we've answered the question that we're not dealing as we might have been in other situations with essentially a wounded embryo.

I think that's really what we come down to, and from your slide that you began with, you showed that, in fact, in the mouse like in every other creature, the cycle of life begins with a fertilized ovum.  I want to be sure if I understand correctly that in this situation we are not beginning with a fertilized ovum.  We've done the treatment beforehand.

The third thing is that you mentioned the possibility that neurotrophic factors and the like would be helpful in reprogramming things, and of course, that's a very exciting prospect for all of us because if trophic factors are there, they ultimately can be synthesized because they're chemicals, and how close are we to getting to synthesize them?  And then we don't even have to have anything to do with cells.  After all, we won't use the penicillin mold anymore.

And then the last little one, it's a tiny point, but you mentioned that the laws in Germany were going to change in some respects in relationship to this research.  Are they changing in such a fashion that IVF embryos now can accumulate in Germany the way they have accumulated in other countries and be a source of problems for us?

Thank you.

DR. SCHÖLER:  So the adequacy of the cell lines.  So as long as people like Rudolf Jaenisch, Doug Melton, and so on are able to publish in Cell, Nature, Science with these cell lines for basic research, I think you can do quite a lot, and that's why I was trying to distinguish between what you can do to quite some extent in basic research in comparison to applied science.

And if you would, just one scenario, if you would try to use the existing cell lines, which have been cultured for a very long time, have been cultured in the presence of animal cells, sera, and so on, and you would use a primate model; let's say you want to try to cure or at least alter the phenotype of a monkey model for Parkinson's, and you transplant these cells and you have a tumor or the cells are rejected.  At the end you will not be able to know is this because your procedure has not been developed in an appropriate way or is it because the cells were not good enough for the procedure to start off with.

If you have a tumor, is it possible that cells that you differentiated from the embryonic stem cells, let's say you have progenitors for neurons; let's say you have neuronal stem cells.  In embryonic stem cells you might not see that the genes have been mutated over the many passages, but once you get to an intermediate stage or even to the differentiated stage, maybe it's then when they exert their problems.

So if you think about testing this in an animal model, you rather want to start this with genetic material, which is perfect.

DR. McHUGH:  But at the level of the basic science, which is often the complaint about the Bush proposal, that it would block basic science, as far as you say at the basic science study of stem cell they're adequate you're saying.

DR. SCHÖLER:  So for the type of research I am doing —

DR. McHUGH:  Yes.

DR. SCHÖLER:  — basic research I'm doing, I don't have a problem.  The problem I have with the cells now is that in Germany — I'm talking about the problems I have as a scientist — I have a problem with interacting with scientists in other European countries because if we're getting cells, let's say, fromWiCell, we have to sign contracts.  [Other European scientists say:] "We have our own cell lines. [Why can't we use our cells?]"

DR. McHUGH:  Right.

DR. SCHÖLER:  So then it happened with the Sixth Framework that because they wanted to include certain German scientists, they had to use the old cell lines and not the newer ones and said, "But we have our own."

So in the end they say, "Why don't we leave Germans out of... the scheme because we can happily work together?" and so on.  So that's another issue, but that's a German one.

DR. PELLEGRINO:  Thank you very much, Dr. Schöler .  We've been working you very, very hard. 

I'd like to just for a moment ask the Council if they have any comments they want to make on the summarization of alternate procedures that was prepared for us by the staff, Adam Schulman and Dick Roblin, and also if you have any comments on the relationship of these alternatives and also the comments by Professor Schöler to our publication on alternate sources.

That's a little complicated question, but I think any aspect of that we'd appreciate your guidance on.

DR. GAZZANIGA:  Dr. Pellegrino, just before maybe going there, I think this wounded embryo question is central to a lot of the conversation this morning, and maybe he could answer that before we move on to the general.

DR. PELLEGRINO:  All right.

DR. SCHÖLER:  So if you would consider the pronuclear state zygote, the pronuclear zygote as an embryo, then you would say we would be doing something to an embryo, and then it would be in your view a wounded embryo.

So that's why I was saying to address that question one would have to do the treatment earlier, before fertilization, but that's —

DR. GAZZANIGA:  Which is possible to do.

DR. SCHÖLER:  And there is no reason to believe why it's not possible to do, but before you have seen the results, you don't know.

But as I said, we want to go as early as the metaphase II.  That's the point where you would put in the nucleus.  That would be the earliest that we have seen that there's obviously Cdx2 RNA.  So I don't see why it should not work.

DR. ROWLEY:  Well, but I think it's very critical to emphasize that he hasn't done it.

DR. SCHÖLER:  Yes.

DR. ROWLEY:  An it may be that Cdx2 at that point activity or gene expression is critical for the process of fertilization.  Because you have to ask why is it so high.

DR. McHUGH:  That's right.  That's very interesting.

DR. ROWLEY:  And it's not high for no reason, I would suspect, and it's just that we're jumping to the conclusion that you can do it at this stage, and until it's actually done and shown to be successful and as efficient as a later stage, this is an unanswered question.

DR. McHUGH:  Yes, Janet.  I agree with that.  The whole reason for putting forth the concept of wounded embryo is to get us into this discussion of where the science is and what we can depend on and what we can't depend upon.  And that's most helpful; your comment is most helpful in that respect.

DR. SCHÖLER:  If I may add something to this, this experiment I've shown you to give you an idea about the outcome.  So this has not been described, I think, to that extent in the Development paper by Janet Rossant and also not in the Nature paper by Rudolf Jaenisch.  That's the outcome.

But Rudolf Jaenisch has shown a way to address that question not having a wounded embryo, like doing this at a stage, at the metastage II phase to transfer a nucleus...

I assume that we have a very similar outcome, and I think from what is published that we might even have a more severe problem.  So if you can accept what they have done, I think ours is going to be the effect of Cdx2 knock-down is going to be worse even at the pronucleus stage.

So if you do it earlier, we have to wait for the outcome, but I don't see why that should be better.  And if it's not working there, then you say that's unacceptable.  That's the outcome, but we have to see.

DR. HURLBUT: Not worse; better though for our purposes is what you mean, right?

DR. SCHÖLER:  Yes.

DR. HURLBUT: Better in preventing an embryo from coming into being.

DR. SCHÖLER:  Yes.  Thanks for clarifying that.

DR. HURLBUT: Yes.

DR. SCHÖLER:  The procedure works better.

DR. PELLEGRINO:  Other questions or comments?

DR. SCHÖLER:  So I have not yet answered all of those four questions.  Should I?

DR. KASS: Mr. Chairman, I think our guest has one more thing he wanted to add in response to Paul.

DR. PELLEGRINO:  Did I?

DR. KASS: No, I think Dr. Schöler .

DR. SCHÖLER:  [I do not think that such a] change in Germany [will result in]more IVF embryos.  I don't think that this will — first of all, we're not allowed to derive our own embryonic stem cells.  So there's no reason to believe that by any of these procedures that we will increase the numbers of in vitro fertilized embryos.

...I think[that] the numbers that are officially in German fridges are extremely low, but Prof. Autiero might also add something.  I think in Germany they're extremely low.

DR. McHUGH:  That's what we had heard previously, and we just wanted to know or I just wanted to know whether this was going to change and that the number of IVF embryos would now accumulate in Germany like they have here.

DR. SCHÖLER:  No.

DR. McHUGH:  With the implications that they bring to our discourse.

DR. PELLEGRINO:  Leon.

DR. KASS: Well, I think I would like to sort of put on the record my own delight and to some extent astonishment that we are November '06.  The Council's white paper was published in May of '05.  A quick subtraction, 18 months, right?  In 18 months we've had peer reviewed publications showing proof of principle, depending on what you think about the reprogramming studies, how much of proof of principle that we've seen.

But there's been active research in all of these areas, and I don't think from the conversations we've had around here that the doubts and the skepticisms have been completely set aside.  There are large scientific questions that remain, and there are, of course, questions that the people who are keen on working with embryonic stem cells have been too polite to press in the discussion, but from their point of view it's not clear why the research need wait for the development of these alternative sources.  That research goes on.

And I think I appreciate very much your suggestion that the full understanding of what's going to work in this area will depend upon the work that's done with adult stem cells and with pluripotent stem cells derived whether from embryos or from some of these alternative sources.

So I think this research would have gone on I'm sure without the Council's prodding.  I think we owe Bill Hurlbut, I think, a considerable debt of gratitude for having insisted that we somehow lift this possibility that science might find a technical way around an ethical dilemma so that the research, if this is successful, could go forward at a very high level in ways that no one will feel morally compromised.  I think this is very exciting.

Whether there's enough here to warrant our saying anything more other than having this very fine update from you on this occasion, I have my doubts.  I mean, I think it would be very nice if the larger community could have been here to hear this really very elegant presentation and also the summaries that Dick Roblin has prepared on the literature I think are helpful reminders to all of us as to what's going on really in the last two to three years in this area, and maybe there is a way to use our Website to call attention to this presentation and to these publications, but since I guess there was partly a discussion last time as to whether we, the Council, need another publication  at this particular juncture on this subject, I myself don't see it.

DR. PELLEGRINO:  Thank you.

Any further comment on Leon's comment or any other related to the alternative sources?  Yes.

DR. GAZZANIGA:  Ever since the alternative sources have been proposed, there has been no, I don't think, doubt that various kinds of biologic experiments could be applied to those questions.  I don't think that's an issue.

I do think that the concern of some of us is that we're dealing with what the economists think is called lost opportunity cost, that there is a way laid out in the scientific community to go forward, and we are allocating resources, limited as they are, to approaches that for some of us don't seem to really answer the moral question, which were why these ideas came up.

So we just have learned that the Cdx2 experiments, in fact, do have the moral problems that were raised by many on the Council.  Robby George is the most articulate with that view, I think.

And I think the de-differentiation suggestion, if you really think about it has similar problems for those who are concerned about a viable entity that could be a human being.

And one could go through on the other two cases and make other moral arguments.  So I guess I, for one, would be more relaxed about these efforts if I actually thought the people who have these concerns truly are at rest with the moral dimensions of it as they had proposed.

Nonetheless, it was a great review, and we thank you.

DR. SCHÖLER:  Thank you.

DR. PELLEGRINO:  Dr. George.

PROF.GEORGE:  Yes.  I think —

DR. PELLEGRINO:  And then Dr. Meilaender and then Dr. Gómez-Lobo.

PROF.GEORGE:  When you call on them I thought they were going to be —

DR. PELLEGRINO:  No, no.  In the order in which I call them out.

PROF.GEORGE:  Thank you, Dr. Pellegrino.

I think I should just say something in response to Mike Gazzaniga's last comment, and perhaps I misunderstood Mike, but just to be clear about my own position, I found myself reassured by what Dr. Schöler was saying, that in the relevant alternative methods that he was discussing, we are not talking about the creation and destruction of embryos. 

So far from being persuaded that the methods he's discussing have the same moral problems, it seems to me that he was arguing that they don't; that, in fact, we not only don't have an embryo in the research he was discussing, but we don't even have an organism.

So we have, you know, some sort of a tissue culture perhaps, a collection of cells that lack the organization and self-direction of an embryo, but if that's right, then we don't have that moral problem.

Now, we still might have the moral problem with the use of oocyte, but Dr. Schöler has also proposed without going into nearly as much detail that there may be alternative sources of oocytes that will not involve exposing women to super ovulation and the possibility of exploitation.

So have I misunderstood you, Mike?

DR. GAZZANIGA:  Well, no.  There's a little bit of a unclarity with this.  Paul McHugh asked the question driving home the point of whether these were wounded embryos.  I happened to notice you were out of the room, and Dr. Schöler answered, well, if you consider a union of an egg and a sperm the beginnings of life and an entity that's on the trajectory towards whatever organism you're talking about, then, in fact, by introducing the RNAs, we do have a wounded embryo.

So this was the answer to the question which finds me stating the thing that I stated.

PROF.GEORGE:  Well, let me see then if I understand Dr. Schöler 's position, and he can simply clarify this.  Take, for example, the kind of work done by Jaenisch.  In your opinion has Jaenisch created wounded embryos or has Jaenisch created non-embryonic sources of pluripotent cells?

DR. GAZZANIGA:  Just to be clear, we're talking about his experiments, and we're talking about his Cdx2 experiments.

PROF.GEORGE:  Well, maybe you could tell us your opinion in both cases.

DR. SCHÖLER:  So what I tried to show here is the outcome of the Cdx2 experiment under these conditions, and to avoid that one has a wounded embryo according to what you said.  One would have to do that at an earlier stage.  I don't see any reason why the outcome should be any different.  If at all, it would be more severe, even more severe.

PROF.GEORGE:  But would that mean you don't have an embryo at all?

DR. SCHÖLER:  No.

PROF.GEORGE:  No?

DR. SCHÖLER:  If you do it at an earlier stage, you definitely don't have a —

PROF.GEORGE:  Definitely don't have an embryo.

DR. SCHÖLER:  — an embryo, and it's a matter of what — now we caught in different cultures or systems.  And according to the German law, law, not church, law, the pro-nuclear zygote is not considered to be an embryo, whereas Hans Schöler, a developmental biologist, would say as soon as you fertilize the oocyte you're starting the process.  Ergo, I would consider this to be an embryo.

Ergo, you would be right by saying that's a wounded embryo, but if I, Hans Schöler , would say is that something that should be protected or not, I would say no.

But from what you've been saying, this reprogramming, bringing back, you're not even getting close to something that is an embryo.

DR. GAZZANIGA:  Well, just to make my point clear, if we go back to somatic nuclear cell transfer and you have a moving a film forward, you go through stages where you do have cells that could become, if implanted, an animal or a human being, depending on the organism in question. 

So that was a moral concern to many people on this Council, that possibility.   So you have to understand the context of the arguments that have been here.

DR. SCHÖLER:  Absolutely.

DR. GAZZANIGA:  So by dedifferentiating a human cell back to some point, some totipotent place, there you will go again.  You'll have those cells that if implanted could be a human being.

So the de-differentiation technology doesn't answer this deep problem that some people have, and that's my simple point.

DR. SCHÖLER:  These are two different areas.  If you talk about de-differentiation, we are talking about cells.

DR. GAZZANIGA:  Yes, correct.

DR. SCHÖLER:  If you look at transfers, that's not de-differentiation.  That's reprogramming because what you're doing is you're putting a nucleus into an oocyte.  The very second if you say de-differentiation, I'm thinking about a cocktail of factors and pushing it toward the pluripotent state, and I've been answering —

DR. GAZZANIGA:  But you have got those same totipotent cells.

DR. SCHÖLER:  No, you get to a stage which is pluripotent, and what I was saying to the kind of scientific question, that where would I bet my money on is that with respect to therapy, I was saying if you go all the way back and then forward, that would be in my opinion the better way of doing this for of the reasons that I've been mentioning.

And if you can do this the way Dr. Hurlbut has proposed it, then you wouldn't get to something which is an organism embryo.  That's the difference.

DR. McHUGH:  Since I coined the term, can I come back at it then?  I'm concerned ethically with using wounded embryos, but I am not concerned with changing gametes.  Okay?  So that from those gametes we could produce cells.  As I understand it, you're saying, and Janet was talking back and forth with me about how this has to be proven that the Cdx2 could work on the gametes, but it has not been demonstrated yet.  If it was with the gametes, then Robby, me and everyone doesn't think we've got a wounded embryo that we're working with here.

So I gathered from what you said that at least it looks like you can use the Cdx on gametes, and gametes and oocytes are unfertilized, and then go from there.  If that's the case, you don't have an organism and I don't have a problem.

DR. SCHÖLER:  Yes.  So you can be assured that actually when I'm back at my computer I will push that we're going to test even earlier, but that was not the scientific question.

DR. McHUGH:  No, I understand that.  I just want my terms to be clear, why I used the terms I did.

DR. ROWLEY:  Well, wait.  So you're saying it's okay to use a wounded zygote, a wounded oocyte that will then be fertilized by a normal sperm to get a zygote that has the genetic defect.  So you're using a wounded oocyte.

I mean, you know, I don't think this is going anywhere, but I just want to point out that —

DR. McHUGH:  I think it's going somewhere.

DR. ROWLEY:  — the manic morass we've gotten ourselves into.

DR. McHUGH:  Yes, I think we're going somewhere.  I think that at the level of cell reproduction and cell things, if they cannot — if you have a wounded gamete, it cannot be fertilized and turned into an organism it seems to me.

DR. ROWLEY:  Yeah, but if it can't be fertilized, then he can't get an eight cell stage structure to use, and he's betting that, in fact, a wounded oocyte plus a normal sperm will give you this Cdx2 deficient eight cell stage organism or structure that will be useful for cell lines.

DR. SCHÖLER:  I personally have a real, real problem to the terminology "wounded" with respect to a cell.

(Laughter.)

DR. SCHÖLER:  I've seen wounded people, but I think this is creating pictures in our heads, which I don't consider to be absolutely valid.

DR. PELLEGRINO:  The Chair just wants a word for a moment.  We want to give everybody a chance to talk.  There are a lot of hands waving desperately.  I must say I've lost the order a little bit.  The order we had was Meilaender, Gómez-Lobo and Bloom and others who want to get into it put your hand up.  Leon, you had your hand up.  I think, Dr. Foster, you had your hand up, did you?

DR. FOSTER:  I was just going to make one comment without making — the only thing is, you know, I was very encouraged to hear this morning from Robby, for example, for the first time.  Nobody is more concerned about wounded embryos and so forth, if you want to use that, than he is, and he's very encouraged by this.

There are going to be some people who think that if you mutate DNA, you've stopped an embryo.  I thought it was tremendously encouraging for the first time we've been in this Council to say we may be able to go forward along these things.

I think we ought to drop this thing about seeing what cells are wounded.  I mean, it's sort of silly, but there are going to be people in this world that no matter what you do, you'll never do a stem cell research, and you just have to accept that they are going to do it.

So I thought the comments were tremendous from this side of the room and very different from the comments early on because of different circumstances.

Thank you for letting me interrupt.

DR. PELLEGRINO:  Yes, you did.  Thank you.  Thank you for speaking while you were interrupting.

We have Meilaender, Gómez-Lobo, Bloom and anybody else that wants to get on the last time, ask Professor Schöler , who has been very, very kind to agree to wait and not respond until you've all made your comments and then he'll put his response together because I suspect there may be some overlap.

Dr. Meilaender.

PROF. MEILAENDER:  Three comments, each very short.  The first, I suspect that actually not a one of us lacks clarity on what we've been talking about here.  I mean, I don't know anything about cell biology, but we all understand that what Dr. Schöler reported on with respect to his own research is something that might perhaps be called an embryo that had then been disabled, but that he thinks — he's actually fairly confident — that it would be possible to do similar research in a way that didn't do it.

I think we're all clear on that, and I don't see any need to run around that pole indefinitely.

The second point, I want to say a word, Mike Gazzaniga, about your lost opportunity cost point.  I thought the mantra all along had been good science proceeds on as many fronts simultaneously as possible.  Therefore, you don't know; you can't quantify opportunity cost lost or gained until you're at the end of the process.

So it seems to me or at least I've always been told that good science proceeds on different fronts at the same time, and therefore, this is not a case of lost opportunity cost but good science going in various directions.

And then third, just sort of to second what Leon said earlier, I don't know that I see that there's something new to say particularly.  I mean, undoubtedly one could do little riffs on different parts of the white paper, but it seems to me that we did a pretty good job actually on that, and it has stood the test of time quite well.

DR. PELLEGRINO:  I have Gómez-Lobo, Bloom, and, Leon, I have you down here.  I wasn't trying to cut you off.

Anybody else who wants to get on the list, and Bill Hurlbut, but let's go again according to procedure, and we'll ask Professor Schöler to hold off until we've heard from all of you.

Gómez-Lobo.

DR. GÓMEZ-LOBO:  Thank you.

First of all, a question of terminology.  I would drop the word "wounded" altogether.  I think it is more or less standard to talk about mutilated or disabled embryo.

Now, if that's the case, I think we're talking here about the question is whether we have a mutilated embryo or non-embryonic structure at all.  Now, if this is obtained by modifying the gametes, from a moral point of view I don't see any problem.  In fact, I'm a bit disheartened by Mike Gazzaniga's remark because precisely here there has been a convergence.  I mean, there's many of us who are concerned about the intentional destruction of human embryos that are seeing a light here, are seeing a possibility of doing this research in a morally acceptable way.

If there's no embryo and if the de-differentiation does not lead to an embryo, there are no more problems whatsoever.  In fact, I was encouraged even by Bob Lanza's experiments because in principle he was accepting the idea that it made sense to try to derive embryonic stem cells without killing embryos.

Now, he didn't do it, but that was the point.

DR. PELLEGRINO:  Thank you.

Dr. Bloom.

DR. BLOOM:  This is not necessarily for Dr. Schöler 's presentation nor comments, but I wanted to expand the possibility of the list that Dick has been tracking for us to mention the work of Diana Bianchi, which deals with pregnancy associated progenitor cells, fetal cells of origin that persist in the maternal circulation for years and seem to be stem-like cells that can repair injured organs in the mother.

Since they are fetal cells, they will have both maternal and paternal genomes represented, and it seems to me that this is a very underexplored area.  It has been largely in the clinical literature, and it seems to me there is some very exciting cell biology that could be done with this because these are stem cells that don't require the destruction of any kind of embryo at all.

DR. PELLEGRINO:  Thank you very much.

Dr. Hurlbut.

DR. HURLBUT: Hans, I understood you to be saying — I think you used this very term that what is created is not an embryo but a single lineage cell culture if you start with the oocyte; is that correct?  That's the kind of terminology you would accept?

DR. SCHÖLER:  Yes.

DR. HURLBUT: Also, do I understand you correctly saying that even though there are barriers in the procurement of oocytes making SCNT or any kind of ANT work in non-human primates and then later with humans, and — well, suffice it those two, the oocytes and the SCNT.  Those are all barriers right now, but I understood you to say that you thought those were technical problems that by reasonable scientific estimation they could be overcome.  Without knowing for sure, it's reasonable to say that these will —

DR. SCHÖLER:  Could you repeat the last part?

DR. HURLBUT: Well, we will find ways eventually to derive oocytes without having to superovulate women.  You've made that point several times, and also that you thought it was reasonable that we could, not certain, but reasonable that we could make nuclear transfer work in primates.

DR. SCHÖLER:  Yes.

DR. HURLBUT: So given those positions, I just want to underscore what Gil was saying.  There's lost opportunity cost if we don't pursue these methods because whether one agrees or not, there's a political impasse, and pursuing these methods could open up what isn't open now.  We could get stem cell lines that could qualify for federal funding and that would be wonderful because they could be used with good ethical oversight.  They could be used on a national and international level for collaboration, and if the techniques work, we could get unlimited source of genotypes to work with.

So that strikes me as an opportunity that's worth pursuing, but not to mention the fact that how much better would it be for our civilization if we could go forward with this research with social consensus?  Besides the fact that this is a bitter controversy, the fact is that if this ever does come to cell therapies, it would be so much better if every patient who entered the hospital and would partake of these cell therapies felt comfortable morally with the way the therapies were developed.

So on both the social and a personal level, it seems like a very positive thing to see an alternative source of these cells, and from what you've said, it sounds like scientifically there's strong reason to believe that's possible.  Is that all a fair statement?

DR. PELLEGRINO:  Can you hold?

Dr. Kass and then we'll have Dr. Schöler .

DR. KASS: Pass.

DR. PELLEGRINO:  Dr. Kass passes.

Dr. Schöler .

DR. SCHÖLER:  So first of all, for me it is something very important to have social consensus.  That's something that when I came back to Germany from the States about two years ago I had a lot of discussions along these lines... [O]ne person for whom I really, really [have a lot of] respect has been a major player in the Bundestag, Mrs. von Renesse, with whom I discussed this point, that society should not be torn apart by what we scientists are doing, and I think this is very important.

I also think it's for the sake of scientists themselves very important.  I've been in Germany as a student when the "Zornigen Viren," that is, the angry viruses — that's [what] the people were called who burned down labs in Germany.  I remember what Oliver Brüstle had had to suffer, because his name was put in the newspaper in a way that he had to be under police protection.

And so I think there will be certain groups in society who try to take this as an excuse to do harm to others, and for all these reasons, for more reasons, for these reasons and so on, I think as scientists we have a responsibility to take care that society is not torn apart.  I think that's very important. [At least] that is a very important issue for me.

Regardless if I have certain points that differ with respect to other points, I think as a scientist I have to agree to the view that is providing the most consensus on these very critical issues.

And that's why I think going along these lines where people have less, I'm sure that you're never going to satisfy everyone, but by getting the problems solved in a way that more and more people have less of a problem with that type of research I think is extremely important, and that's why if I'm being faced by other scientists who say you're wasting your time because you're not going to please anyone, I think that's wrong.

Some people will try to hold up the stick higher and higher and say, "Now jump over this one," if you see how high it has to be until you stumble.

But I think having said this to the other part of your question, I think a lot of these things are really technical problems.  If you find a drug that disassembles the things that you right now remove together with the chromosomes, that's something that you have to do research on...

I think what science has shown, that normally there are solutions to problems.  It's just like it takes smart scientists together with people who know more about certain issues.  That's a teamwork effort, and we're going to get to solutions.

I'm very optimistic that we're going to find solutions.  If we are patient enough right now to wait until we are through this bottleneck, if we say — that's something that has been discussed a couple of times — that it's important that we move on as many fronts as  possible to see which one is the most successful, and what we, as a society, are willing to accept or not.  Maybe this is more promising, but we don't want to have it.  At the end we have to say if we will see if we were right or wrong what we're doing.

But where I see myself in the dilemma, I think a lot of things are possible here that are not possible even in Germany, and in Germany at the end things will be imported; therapies will be imported from countries where things were developed and Germans did not participate, and we're using the fruits of what has been done.  That's where I personally, Hans Schöler, see a problem.

Did that answer yours?

DR. PELLEGRINO:  Thank you very much, Dr. Schöler , for a very, very comprehensive and stimulating and obviously provocative presentation, which I think has been very useful to all of us.  We're much indebted to you.

Thank you.

DR. SCHÖLER:  Thank you.

(Applause.)

DR. PELLEGRINO:  We recess until two.

(Whereupon, at 12:21 p.m., the meeting was recessed for lunch, to reconvene at 2:00 p.m., the same day.)


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