Thursday, April 25, 2002
CHAIRMAN KASS: Would the members please rejoin
the meeting. While we are waiting in the hope that our straggling colleagues
will arrive, a couple of matters of business.
Session 2: Stem Cells 2:
Medical Promise of Adult Stem Cell Research (Present and Projected)
Dr. Catherine Verfaillie
If anyone has not turned in a request for a box lunch, please do so
now, and that should be in front of you. We will have lunch in the room
just down the hall where we gathered before.
The photographer who has been around here is doing individual photographs
for the commission and he will want to take individual photos of members,
and we can do that in connection with lunch.
And you will also have in front of you in addition to the materials
that Dr. Gearhart provided us, which by the way is — and the lights were
out and so you couldn't see, but one could recapitulate his talk with
the help of the figures here, as well as checking his article in Nature.
But you also have in front of you a revised version of Bill Hurlbut's
memorandum. This has been updated and corrected, and he would like us
to substitute it for the one that was sent around earlier this week. Is
that correct, Bill?
DR. HURLBUT: Yes.
CHAIRMAN KASS: All right. Well, again, it is a
great pleasure to welcome Dr. Catherine Verfaillie, from the University
of Minnesota. You have her curriculum vitae in the briefing book, which
you can consult.
I won't waste any more of her time by reading from it, and just simply
allow her to help educate us on the prospects of present and projected
of adult stem cells for regenerative medicine.
DR. VERFAILLIE: Good morning. I would also
like to start out and thank Dr. Kass and the council to allow me to present
this information on new findings in adult stem cell biology which have
been received with great excitement, and correctly so. If they are, and
they are actually set upside down, the classical paradigms of biology,
and so to be able to do that you have to have full proof to actually be
able to be in a position like that.
If they are, and they are actually set upside down, the classical paradigms
of biology, and so to be able to do that you have to have full proof to
actually be able to be in a position like that.
As Dr. Gearhart already gave in his previous eloquent description of
what stem cells are and what they can do, and we will get back to that
to some extent at the end, although we are far away from actually being
able to use adult stem cells for clinical applications.
But what I would like to do is give you an overview of the greater potential
of adult stem cells, which has always been termed adult stem cell plasticity,
and what we do know and what we don't know.
And where this may actually lead us. Dr. Gearhart also indicated that
embryonic stem cells in humans are fairly or very much in their infancy,
the same as we are for adult stem cell biology, too, and so I don't think
we are anywhere close to be able to come up with new therapies at this
point in time.
I would also like to reiterate that even though my laboratory and our
group works on adult stem cells, we have actually actively pursued investigators
in embryonic stem cell research, human embryonic stem cells, just so that
within the same institution we would have laboratories that have one cell,
and other laboratories that have the other cell, so we would be in a position
to compare and contrast the potential of the different cell populations,
and I think that is very important.
With that, I will actually start my presentation, and I will point out
that the work was mainly funded through the NIH, since it is all adult
stems that we are working on, and not embryonic stems. And also a number
of foundations and one pharmaceutical company.
Dr. Gearhart already gave you an overview of where embryonic stem cells
come from, and where primordial germ cells or stems come from. And I am
going to reiterate that for you.
I just put up this cartoon that Dr. Weissman published two years ago
in Science to point out a couple of things. During development, cells
in the inner cell mass make sequential decisions, and each of these decisions
is actually accompanied with gain of function, but also loss of function.
The gain of function is that the cells learn how to become a more specified
cell type; and on the other hand, actually lose the potential to become
other cell types.
And so the decision to be made is somatic or germ cell, and within the
somatic lineage doing something that is called gastrulation, cells decide
to become the different parts of our body, whether it is endoderm, which
is the internal organs, mesoderm, which are limbs and soft tissue, and
ectoderm, which really comprise the skin, the central and peripheral nervous
And within each of these groups cells again make decisions and learn
how to become stem cells for specific organs. And the stem cells for specific
organs that has been most well studied is actually the hematopoietic stem
cell, which is currently extensively being used in clinical applications
for bone marrow transplantations or peripheral blood stem cell transplantations,
or cord blood transplantations.
And so that actually has set the paradigm on how we decide what stem
cells are. Aside from hematopoietic stem cells or blood stem cells, we
have a number of investigators who have identified tissue-specific stem
cells in a number of different organs, including for instance the brain,
which we until about 10 or 20 years ago thought was a final product when
we were born.
But it is now clear that there are stem cells in the brain that can
recreate neurons and other components. There is also stem cells in the
liver, and stem cells in the gut, and there is stem cells in the skin,
and so forth.
The reason why I put this slide up is actually to point out that these
arrows have always gone down, and so we have always thought that each
time a cell decided to learn something new that it lost the capability
of doing something else.
And so if we envisioned beforehand that the arrows would be reversed,
we thought that was possible, but we associated that with classical transformation,
or actually cancer-forming cells.
So what do we know about hematopoietic stem cells and that is really
the paradigm to which I am going to try to talk through the whole field
of adult stem cells.
In hematopoietic stem cells, we can actually take a single mouse bone
marrow cell that we characterize by proteins on the cell surface, and
take that single cell, and for instance you can take it from a mouse that
is engineered to fluoresce green under a specific light, and put that
in a regular mouse, and ask whether they can reconstitute the blood elements
of that animal.
And a number of investigators have actually been able to do that. You
can take a single cell, and give it to a mouse that was lethally irradiated
so it has no blood, and this cell can recreate the red cells, the white
cells, platelets, lymphocytes, for the lifetime of that animal.
And that is really the proof that you have a stem cell that can self-renew,
and a single cell can make multiple different things, and it can repopulate
functionally the organ that it needs to repopulate.
And so that is really the criteria that we have to hold ourselves to,
to actually talk about stem cells, and if you talk about plasticity, you
will have to hold us on the same criteria and showing that a single cell
can now make two tissues, and that this cell can make two tissues from
a single cell, and that these new cells can repopulate a tissue functionally
Now, over the last 5 or 6 years, there has been an enormous number —
well, not an enormous number, but probably 40 or 50 papers now that have
come out in the scientific publications that have used the word adult
stem cell plasticity.
And what is meant by that is that you take a cell that was supposed
to be a one cell type. For instance, you take a bone marrow cell, or you
take cells that are enriched for hematopoietic stem cells.
And it appears that some of these cells may acquire characteristics
of cells outside of the organ where they came from. And so it has been
shown for bone marrow cells, or cells enriched for hematopoietic cells,
that if you transplant these into an animal that was irradiated, and you
look in tissues outside of the blood, that you can actually find, for
instance, skeletal-muscle cells, heart muscle cells, or endothelial cells,
that are now derived from this donor hematopoietic cell.
There is also papers that have shown that if you take muscle from an
animal and mix it up in the laboratory, and culture it for a few days,
and then use the muscle tissue to give back to an animal, that you could
reconstitute the blood system in that animal.
Now, if you think in anatomical terms, this is still within one of the
three categories that I gave you at the beginning; mesoderm, endoderm,
and ectoderm, and all of this is still within the mesoderm. So this is
maybe not so hard to understand.
However, there is also papers that two different cells from bone marrow,
hematopoietic cells, and zymogenic cells, which are cells that make bone
and cartilage, can give rise to cells that appear to have neuronal characteristics,
both neurons and glial cells, that support the structure of the brain.
And there is a number of studies that have shown that bone marrow cells
can contribute to liver, skin, lung, gut, and so forth, and so you can
pretty much put arrows in whichever way you want.
You know, people have published data that suggests that indeed this
may be possible. So obviously this goes against our paradigms and this
would say that either something strange is going on, and just something
in the last few years is something that we have actually identified.
Now, if we want to talk about blastocyst, I started out with the paradigm
of stem cells, and so there is multiple different possibilities here.
Either the bone marrow, which seems to be the organ that harbors the
most of these cells, harbors many, many different stem cells, and it harbors
the hematopoietic stem cells, but it also harbors the neuro stem cell,
and the liver stem cell, and so forth.
And which that would not be bad, but that truly would not be a single
stem cell that could be expanded and used to actually transplant patients
with all kinds of different organ diseases.
A second possibility is that somehow the cell can be "de-differentiated"
and redifferentiated, depending on the environment that it is put in,
and that the hematopoietic stem cell can learn how to become a liver if
you put it in the liver, or it can learn how to become a brain if you
put it in the brain.
Or it could be that it is a remnant of embryonic stem cells or the primordial
germ cells that you heard about from Dr. Gearhart that are left around
in the body, and that under specific circumstances can be reactivated
and contribute to tissues.
And the issue of fusion has been brought up because of the two papers
recently in Nature, and the possibility is in theory that what we see
is actually that.
For instance, a hematopoietic stem cell fuses with a liver cell, and
now you actually have something that is a hybrid, but it has actually
The other questions that I am going to try to address, and I don't have
all the answers for this, is this actually clinically relevant? You know,
if you transplant bone marrow into a patient and you find two liver cells
that are derived from the patient, from the donor, it doesn't necessarily
mean that that is going to help anybody down the line.
So the graft has to be robust and persistent, and there has to really
be proved that we don't just see cells that look like a tissue that they
end up in, but they also have to function like a tissue that they end
And then the question that I will bring back up at the end, the first
question, what is plasticity, and will that matter from a clinical standpoint?
And so we started out in this field — I am a hematologist, and I do
bone marrow transplantation as my clinical profession, and I have been
interested in hematopoietic stems in the bone marrow.
And about six years ago somebody in our group asked me whether we could
grow mesenchymal stem cells, which are cells that may grow on cartilage,
to treat children with a specific genetic disease called Hurler's disease.
And when we did this, mesenchymal stems we happened to find, and we
went about trying to create these to be in compliance with GMP qualifications,
meaning we were trying to remove all sera out of the system, and yet we
were trying to use very well defined culture systems.
And so while we were doing this, we came up with a cell that you have
heard Dr. Kass refer to as a multi-potent adult progenitor cell, because
we don't have a much better word for it.
And it will be appreciated as MAPC, and which appears to have a much
greater possibilities than the mesenchymal stem cell possibilities. So
we take these cells from bone marrow from humans, and we can also take
them from mice and from rats.
And you place these in a culture system that is very well defined, and
ingredients, and growth factors, and no serum, and low density, and we
expand the cells as much as we can by splitting the cultures on a regular
And if we do this, we have actually found that these cells appear to
have an enormous growth potential. And so here on the left-hand side would
be bone marrow from an individual, and we start with about 10cc's or a
spoon of bone marrow, deplete all the blood elements from the bone marrow,
and put it in a culture dish, and then grow the cells for long periods
Classical adult cells would actually not expand much more than 50 times
or 60 cell population doublings, just because we have a clock inside the
cell that actually causes the cells to become senescent or old once they
go beyond a certain number of cell divisions.
And so in the human system, as well as in the mouse and the rat system,
we have been able to show that we can create or grow cells that do not
seem to conform to this internal aging clock.
And the cells can go beyond that and the human cells are now close to
a hundred population doublings, and in mouse and rat, over 150 population
If you look at the aging clock itself, which are the telomeres, the
telomeres are long and they do not seem to shorten in culture, which goes
again with the idea that the cells do not senesce in culture.
So in this respect, they have characteristics that are similar to what
you would find in embryonic stem cells, but also this internal clock is
actually not working.
The phenotype of the cell is strange, and it doesn't really fit anything
in particular, but there is definitely no characteristics in these cells.
These cells are blood hematopoietic stem cells, and I am not going to
go through all the details here, but if you do an extensive phenotype
characterization of the cells, they don't look like blood.
They have some characteristics of embryonic stem cells, but there are
a lot of other ones that they do not have. So they have some genes that
are turned on that are present also in embryonic stem cells, which are
the top two here, and then they have on the cell surface antigens that
you really only find on embryonic stem cells, or primordial germ cells.
So in some respects again these cells have some features of embryonic
stem cells, even though we got these from the bone marrow of humans, mice,
We then started trying to test initially all in culture dishes what
these cells could do, and we asked whether they could differentiate in
multiple different cell types.
And because our initial charge was actually to try to grow mesenchymal
cells and make bone and cartilage, that is what we did first. And so what
we showed in the culture dish is that if we switch the culture conditions
around, and actually use ingredients that are no longer supported for
maintaining the stem cells in an undifferentiated state, by actually switch
them such that we hope that we can turn on the genetic programs to make
bone or cartilage, and so forth, we could indeed do this.
And this is no different than the classical mesenchymal stems that have
been described. So we can induce the cells to become bone, and if we say
that they differentiated into bone tissue, it is actually a calcified
tissue at the bottom of a dish.
We can induce the cells to become cartilage that looks like articular
cartilage, even though it isn't very well organized. And you can induce
the cells to become lipid-laden lipocytes, and we can induce them to become
skeletal muscle cells.
And these cells can actually fuse and make long muscle tubes almost,
and we can induce the cells to express a number of muscle markers for
the heart, even though we haven't really seen beating cells.
And so we don't really know whether these cells are heart muscle cells.
So this is still not that strange, because there is this cell in the bone
marrow that has been identified that can do this.
Now, we found three other lineages that are completely outside of the
mesenchymal lineage, and some of this has been published, and most of
it is actually in press currently.
One of the things that we found is that these cells can differentiate
into cells that line blood vessels, which we call endothelial cells. And
we have been able to show that these cells differentiate into cells that
look like endothelial cells, but also function as endothelial cells.
And as shown in this picture here is actually a blood vessel from an
animal that had a tumor underneath the skin, and we actually infused human
endothelial cells derived from human MAPCs in this animal, and showed
that these endothelial cells seek out the tumor and actually help create
new blood vessels in the tumor, which the tumor needs otherwise it can't
And so this proves that these cells that are in the bone marrow can
differentiate into cells that can make endothelium. More surprisingly
is that the cells can differentiate into cells that look like neutrons,
look like astrocytes, and support themselves in the brain, and to some
extent function like these cells in the brain.
And so we show here that they differentiated into cells that look like
neurons and have electrophysiological characteristics like neurons.
And so this is the second major layer of the embryo, and then we also
have been able to show that we can make these cells differentiate into
cells that look like liver cells, and actually function like liver cells
in a culture dish.
And so this would mean that this cell population, these MAPC cells,
can actually differentiate into all of the major components of a human
being, even though we only show a few cell lineages here.
I am not going to go through this in too much detail because it is highly
technical, but essentially we have not been able to use genetic marking
to prove that this could all be derived from a single cell, and we don't
depend on population of cells.
So this fulfills two of the criteria of a stem cell. A single cell can
differentiate and grow for long periods of time, and can differentiate
into multiple different tissue cells.
Two more sets of experiments were done to try to gauge the potential
of these cells. The first one was done in an chimeric animal model, in
which we took the adult cells, and injected even a single adult cell into
the blastocyst of a mouse and asked what would happen in this mouse, and
whether we would see contribution to some tissues, no tissues, or all
So we injected a single cell or we injected 10 to 12 cells, and shown
here are two animals. The top one is obviously and the donor cells here
have a gene that if you stain it correctly the cells turn blue.
So what we did is we let the animals get born, and we looked at the
animals by genetic tools to try to figure out if there were donor cells
in multiple different organs.
And we also then took the mouse and actually cut a thin slice through
the middle of the animal and asked which organs would have blue cells
contributing to the mouse.
The top mouse is an animal that if you looked in the tail by genetic
tools that we couldn't find any donor cells, and the bottom mouse here,
this is its head, and over here would be his tail, and you can see the
spine, and the brain, and all the internal organs.
And you can see that the majority of all the tissues of this animal
actually appear to be derived from a single blue adult cell that we have
put into the blastocyst.
The efficiency isn't a hundred percent, and this is shown on the bottom
here, and so if you look over here, and if you put in one cell per blastocyst,
60 percent of the animals will not be chimeric, but 30 percent or 40 percent
of the animals will be chimeric to varying degrees.
If you increase the cell number the chimericism goes up. So this is
probably not quite as good as embryonic stem cells, but it is a fairly
significant degree of chimericism, and actually the frequency appears
to be one in three cells.
So this would suggest that the cells can probably make under the right
circumstances more cell types than we have be able to prove in a culture
We can also ask if we now take these stem cells and give them to a mouse
that is born, and we give here again cells from the donors' mouse, which
again are blue, and we gave these to an animal that was either not irradiated
or irradiated with a small amount of radiation therapy in the hope that
maybe that would help the cells engraft.
We used an immune-deficient recipient mouse, just because we were worried
that the new genes that are in the blue mouse might actually be a basis
for rejection. So we don't know what would happen in a non-immumodeficient
If we do this, what we found is that we do find engraftment in some
tissues, but not all. So, for instance, in the top panel, we see that
there is engraftment between 3 and 9 percent in the hematopoietic system
of this mouse, and we can find the cells, and the blood we can find in
the bone marrow, and we can bind them in the spleen.
And if we look in these animals, we can also find over here, and what
we did is we actually — the blue color, we used an antibody that is now
green, and co-labeled it with a red stain that stains the specific tissue.
And you can see in the liver that there is areas in the liver where
donor cells appear to be present. And there is areas in the guts, in the
villae of the gut, where donor cells appear to be present.
And there is areas in the lung where donor cells appear to be present.
The presence of these cells can be seen anywhere from four weeks after
transplantation, all the way to 24 weeks, which is about six months, and
the unfortunate thing with the mouse model that we use is that these mice
usually die from lymphomas at an early age because of the deficiency that
So we really have not been able to extend the cultures or have the mouse
experiments beyond 6 months, and so we are actually trying to go further.
We transplant the cells in an animal that is 6 to 8 weeks old, and so
it is not a very young mouse, and it is also not an old mouse. What we
showed is that if you damage certain tissues like the hematopoietic system,
and the gut system, that you have increased engraftment, which is consistent
with the fact that these cells go to places where the repair might be
However, we did not see in this mouse model engraftment in a number
of other tissues, and mind you that we gave these cells IV to an intact
mouse, which actually was not damaged in any way, shape, or form.
And we don't see engraftment in the heart, skeletal muscle, or brain,
and these tissues do not proliferate. We also don't see engraftment in
the skin and the kidney, and so these organs we didn't really see very
However, if you infused the cells directly in the muscle, which causes
damage, and actually done the cells in response to the local cues within
the muscle, appear to be able to differentiate into muscle cells.
So it appears that these cells have the ability and blastocyst experiment
to give rise to many, many different tissue types, if given post-natally,
and we gave them as stem cells, not as differentiated cells.
They appear to be able to respond at least in some respects to cues
that are present in certain organs to differentiate into the cell type
that is specific for that organ.
We have looked carefully at the cells in culture and we do not see a
significant number of gross genetic abnormalities. We have not looked
with a very fine-toothed comb through whether there might be some minor
genetic abnormalities over time and culture, and these studies are ongoing.
If we infused the MAPCs in animals, we really do not see any tumors,
and so far we have not seen that there are tumors that Dr. Gearhart talked
about, and we also have not seen any other tumors.
Obviously if these cells come from bone marrow there is lots of precedent
on bone marrow transplantations, where actually if you do this, actually
you do not cause tumors in patients.
So MAPC that we have identified in our laboratory seems to be a cell
that is not senescing and that can be found in adult tissues of humans,
as well as mouse and rats, and they seem to be capable of giving rise
to cells from the three germ layers, and it can engraft in vitro in a
limited number of tissues.
Now, what I cannot tell is whether these cells actually exist as such
in a person, in a mouse, or in a rat, or whether our culture condition
is actually such that it, quote, reprograms or dedifferentiates the cells
that we take out of the animal, and that then acquire this much more greater
potential, and I will come back to that in just one second.
So we now go back to my initial definition of what is plasticity, which
is really at the bottom of all of the adult stem cell excitement. I mentioned
initially that we would have to show that this is a single cell of a rat,
and I think the majority of papers so far published have actually really
not been able to prove that a single cell could, for instance, give rise
to blood and muscle.
In vitro, we have evidence for that, and in the blastocyst injection,
we took a single cell and actually found multiple different tissues. You
could ask, well, does it matter?
Does it matter if there are multiple different cell types in the bone
marrow, and I think ultimately from an FDA or regulatory standpoint, it
will matter, and we will have to be able to say exactly what cells that
we are using to be able to acquire a certain function in vitro, and so
I think that will be important.
The second question is, is the differentiation or is the remnant ES,
and again you could say, well, it probably doesn't matter. But I think
at this point in time, I don't think anybody in this field knows whether
these are left-over early stem cells like ES cells, or whether these cells
are cells that can be reprogrammed, and redifferentiated, and dedifferentiated
under certain circumstances.
Now, does it matter? Well, you heard from Dr. Gearhart that embryonic
stem cells as such, and not necessarily the differentiated progeny, but
the ES cells themselves can cause teratomas, and even though nobody in
the adult stem cell plasticity era has actually shown teratomas, it doesn't
mean that it might not happen.
If it is dedifferentiation, it means that you reprogram or you change
the genetic material in a cell. But if you do that, currently we have
no proof that we actually change something and actually cause an oncogene
or something like that to be activated, but that is definitely within
the possibilities, and that definitely needs to be looked at carefully.
Is it fusion? All the in vitro work that has been published, including
the data that I have shown to you today, I couldn't prove beyond any doubt
that that is not based on fusion.
Our in vitro data, we have never co-cultured things with anything. So
we have single cells that are deployed that can do multiple different
things, and so we can't really ascribe that to fusion.
However, in vitro, I couldn't prove it to you today, and we are doing
studies to try to address this. I think that fusion might be the reason
why some studies in which a lot of pressure has been put on to the system,
which is essentially what those two papers had to do in vitro.
So we have a lot of pressure exerted to have that one cell survive after
it fuses, and that is a possibility. Also, single cells that are found,
rather than whole colonies, may also be the result of fusion, more so
than experiments where you see huge colonies arise in an in vivo model.
And so I think we currently cannot exclude the possibility that some
of the data is as a result of fusion. Some would say does it matter, and
I think it matters a whole lot, even though some investigators say, well,
if you fuse the cells and it functions properly, it probably doesn't matter.
But I think ultimately that we do need to make sure that we understand
the whole mechanism underlying everything. And is all this plasticity
And so the majority of studies published to date have actually shown
the very low numbers of tissue differentiated cells can be found in multiple
A number of papers have been published, two in particular. The paper
by Lagasse, et al., where they show that they could rescue an animal with
liver failure by bone marrow transplantation, but they have significant
degrees of engraftment.
So that definitely was up to 80 or 90 percent of the liver could be
replaced by bone marrow cells. And a paper by Don Orlic showing that if
they injected stem cells into the heart that was infarcted that a significant
amount of donor cells would be found in the heart.
And in the data that I have shown you, that we have up to 5 to 9 percent
of the differentiated tissue that seems to be derived from the graft.
However, the majority of studies again haven't really addressed the
other question in plasticity, meaning is it in vitro functional differentiation?
And there is really only a single study that has been able to show that,
and it is again the same study by Lagasse, et al., who showed that if
you did bone marrow transplantation in an animal that had a failing liver,
you could rescue the animal and take it off the drugs that kept it alive.
Some studies have shown that there is functional improvement, although
the mechanism for the functional improvement isn't completely known, and
that is to some extent similar to what you heard from Dr. Gearhart.
And so there is a number of studies who have injected cells in adults
in organs and have shown, for instance, that there was improvement in
the neuronal function, and that there was improvement in heart function,
although there is no proof that the cells, per se, were actually responsible
for doing this.
And the question will be is this acceptable from a clinical standpoint,
and if you show only functional improvement without knowing the mechanism
for knowing why we see functional improvement, and in the long term, again,
that is not a tenable situation, and we really have to dig into this much
So what can adult stem cells be used for? Well, I think like embryonic
stem cells, or primordial germ cells as you heard from Dr. Gearhart, the
cells are good tools to study five basic principles in biology.
And we can study self-renewal, and we can study differentiation and
redifferentiation if that is indeed the case, and learn what the implications
for that are.
And actually try to understand how organs are being created, and what
the genetic programs are that you need to turn on. The cells, like other
stem cell populations, could be used for drug discovery, for drug toxicity
Adult stem cells could be used as systemic therapies, and currently
systemic therapies are done with adult stem cells. Bone marrow transplantation
is done every day in many, many institutions around the world, and so
we can infuse these cells if we do not think that they make tumors.
So since adult stem cells don't seem to have that as their side effect,
theoretically, we could genetically correct cells for patients who have
deficiencies of certain enzymes. And the disease, and Hurler's disease
would be one example, and a second possibility would be, for instance,
in hemophilia, where you need to have a cell that produces clotting factors.
Or other congenital diseases, like Alpha-1-Antitrypsin deficiency, or
it could be used for systemic cell therapy, which you would have to treat
in many, many different places in the human being. For instance, muscular
So if you had a stem cell that was able to engraft in most muscles,
and you could genetically correct it, you could correct that disease in
patients with that disease.
Systemic cell therapy may be more complicated with cells that have the
inherent capability of making teratomas just because you would always
run the risk that teratomas might show up.
And then again if this field progresses further, the same diseases that
has been quoted for embryonic stem cell therapies would also be on the
list here, and if indeed the cells can differentiate into functional neuron
cells, they could be used to treat Parkinson's disease and many other
And since the cells can appear to be able to differentiate into functional
liver cells, they could be used either in vivo to replace the liver, but
also would be very useful to make bioartificial livers, for instance.
We have shown, and others have shown, that cells from bone marrow can
contribute to new blood vessels, and so this could be harnessed to create
new blood vessels in vivo, or actually the opposite; lower these cells
with anti-cancer agents, and actually use them in a anti-angiogenesis
approach for treatment of cancer, and then many other diseases.
Again, we are not anywhere close to being able to do this in any way,
shape, or form, and a lot of basic research still needs to go on.
So the first point that was on my previous slide, we really need to
spend a lot of time in trying to understand what these cells are and aren't.
And at the same time, start thinking about how we might be able to scale
these up under GMP conditions that conform with regulatory agencies, and
we will have to ask the question, as with any other stem cell population,
whether we will use the cells as stem cells, or as more mature cells that
have been educated to some extent to become the final product are totally
And then again perform large scale culture systems or develop large
scale culture systems. And then the last question is whether we should
use these cells in an autologous setting or in an allogeneic setting.
Obviously adult stem cells for a number of diseases could be used in
an autologous setting. However, if they were to be capable of repairing
hearts, and you have a heart infarct today, we would not have adult stem
cells sitting around instead of your own to treat you at that moment in
So I think there are some issues, and Dr. Gearhart also brought up the
idea that with diabetes, for instance, in Type-1, is an immune problem,
and again autologous transplantation may not be the way to go.
I think that for adult stem cells, the initial trials may well be autologous,
but that in the long term, to make it more cost effective and more available
to many patients with certain frequent diseases, that it might have to
be an allogeneic therapy, and then we are actually faced with the same
questions that investigators that work with ES cells, and primordial germ
cells are faced with. I think I will stop there. Thank you.
CHAIRMAN KASS: Thank you very much, Dr. Verfaillie,
for a clear, lucid, orderly presentation, and it is very helpful to us.
The floor is open for questions, comments, discussion. Elizabeth Blackburn,
DR. BLACKBURN: Thank you. Could I just
ask a couple of quick clarifications. Dr. Gearhart mentioned in response
to Bill Hurlbut's question the difference between fetally derived human
cells and mouse embryonic stem cells with respect to their teratoma producing
And I could not quite gather whether it is human embryonic stem cells
that are also known to have any teratoma producing properties. Could you
clarify that for me, because you also had mentioned this, and I wasn't
sure if you were referring to the mouse embryonic stem cell work or the
DR. VERFAILLIE: If you use either mouse or
human embryonic stem cells without predifferentiating them into a committed
progenitor cell, and you use the stem cells as such, they will form teratomas,
because it is one of the tools that investigators use that an embryonic
stem cell has that capability. So they will form teratomas.
DR. BLACKBURN: And then post-differentiation?
DR. VERFAILLIE: I think there is very little
data on the human embryonic stem cells, post-differentiation in vivo,
and whether there is still the tendency for these cells to make teratomas.
DR. BLACKBURN: And the second question,
since I promised that I would ask you about, is the fusion issue, and
which of course you have raised in your talk as well, but again a question
of clarification for me, and maybe expanding on your point that you said,
well, fusions are going to be problematic.
I mean, the thing that immediately occurred to me was that these fusions,
as reported from the in vitro culture, and I believe from engraftment
into mice, that they showed aneuploidy, which of course anybody being
a hallmark of tumor cells.
So I wondered if those issues and perhaps others were things you could
tell us a bit more about when you mentioned that you had concerns about
DR. VERFAILLIE: Well, I think it is something
that because of the papers that were published that elegantly showed that
if you took a somatic cell, an adult hematopoietic stem cell or brain
stem cell, and co-cultured it with embryonic stem cells, and then put
quite a bit of selectable pressure on the system in the culture dish,
they proved that an embryonic stem cell quality could be transferred to
the blood brain stem cell.
And initially they interpreted this as being reprogramming of the cell.
But then it turned out that there were four sets of chromosomes, and that
the cells fused.
And they took these fused cells and gave them to — injected them into
a blastocyst as hyperdiploid as cells with four sets of chromosomes. One
group was not able to create chimeric animals, and the second group, under
the direction of Dr. Austin Smith, were able to create chimeras in the
mice that were what he calls unbalanced, meaning that he saw a contribution
to tissues, and that four sets of chromosomes are actually tolerated.
For instance, the liver, where at least 50 percent of the cells, actually
half, have two nuclei. So I think that currently no investigator who has
worked with adult stem cells has set up the right experiment to actually
be able to disprove that it isn't fusion.
I would argue that the data that I showed today in vitro, where single
cells make three layers of the embryo, and these were euploid cells, meaning
that they had a normal set of chromosomes, and which done in human, mouse,
and rat, at the single cell level, we can make the three major layers
of the embryo.
So that would go against the argument that at least in vitro, that all
of it is caused by fusion. In vivo, in our blastocyst experiments, 1 in
3 cells could do it, which is much higher than the one in a million cells
that were quoted in the two papers that were in Nature, but which indicated
that one bone marrow cell out of a million could actually make a fused
And I think one in 50,000 neural stem cells could actually cause fusion.
So that was a very rare event; whereas, our events are higher. We are
in the process of actually going back to these animals — that we have
cryopreserved, to try to identify that since some of the transplants were
done female into male, we should be able to prove that we do not find
the y chromosome in the engrafted areas and in the chimeric areas, which
would get at the question whether it is caused by fusion.
And so I think we really need to set up experiments where we have generic
markers on both sides, meaning the donor and the recipient, so that we
can prove beyond any doubt that the in vivo results would be the results
from a fusion.
DR. BLACKBURN: Yes, I totally agreement
with that. I think the in vitro, and I am very impressed by the in vitro
results, and as you said, there are questions in vivo.
I think in-part my question was addressing this issue, and I was asking
about the tumor forming ability or otherwise, because it was not exactly
4N. It was the median number of chromosomes was different from simply
4N, suggesting that there was aneuploidy, and for example, one might not
find Y chromosomes, for example, because those had been selectively lost.
So one would probably have to do much more extensive genome-wide analysis
of both of those to be sure that there wasn't some genetic contribution
from the recipient cells.
But I certainly am very impressed as you say with the in vitro results,
and they seem quite unequivocal, and I guess which is the question that
you are addressing, and we will find out as the in vitro —
DR. VERFAILLIE: Yes, and I think we need to
set up the experiments where we have on multiple chromosomes genetic markers.
You know, sequences that we can distinguish the donor and recipient between.
So these experiments need to be repeated.
CHAIRMAN KASS: Questions? Janet Rowley, please.
DR. ROWLEY: Well, I would like to ask a question
that will include both Elizabeth, as well as Catherine, because I was
struck in the data that you presented on your human cell lines that you
had passed for more than a hundred generations, that telomerase was still
And I just am curious about that, because many of us do believe that
that is, if you will, the internal clock that limits the number of doublings
that those particular cells can undergo.
And you derive these from adults, presumably young adults in human,
but at least adults, and I am curious as to what you thought about the
mechanism of preserving the telomerase activity, and maybe if Liz would
have any further comments on that, because again one of the critical features
and potential limitations of adult stem cells is the fact that they would
have potentially fewer doublings than would those derived from embryos.
CHAIRMAN KASS: Could I ask as a favor to the non-scientists
in the group if someone would just give an ABCs on the telomerase matter,
and just very, very briefly, so that everybody can understand what the
discussion is about. Elizabeth, or Dr. Verfaillie, if you could just give
the barest —
DR. BLACKBURN: I am the worst person,
because I will fall into expert jargonese and so I will try not to. So,
telomerase keeps the DNA at the ends of chromosomes replenished, and such
replenishment is necessary, because each time one of our cells divides,
the DNA at the end of the chromosome is a little bit whittled away.
So, telomerase keeps putting back a little extra DNA on to the ends
of the chromosomes each time on average a cell divides. So the issue that
Catherine pointed out in her talk was that if you don't have telomerase
after a number of cell multiplications, that whittling away process would
have gone too far, and that sends a signal to cells to cease dividing.
And so many, many normal cells in culture are characterized by the inability
to keep on multiplying. Did that clarify the question? So many cells do
not keep multiplying because they turn the cells' telomerase off as part
of their natural differentiated state.
Cancer cells, on the other hand, have telomerase, almost in a great
majority of the cases, and very up-regulated, and cells of the hematopoietic
system — and I will defer to Catherine on this — have an interesting
intermediate situation, where they have regulated telomerase activity
that is turned on in a natural and regulated way as the cells multiply
in response to signals in the body. Is that fair to say?
DR. ROWLEY: Yes.
DR. BLACKBURN: So I think it is a very
interesting question of why telomerases is turned on in those cells that
are multiplying so well in culture, and has there been a selective event
that has allowed those cells, that for some reason have turned their telomerases
on in the culture conditions.
But those are the cells that are outgrowing perhaps others in the population,
and perhaps that question might be answered by what is the clonal efficiency
with which you get these lines growing out. You may already know this.
DR. ROWLEY: But can I intervene, because you
assured that it was often turned on, and maybe these cells are identified
because they never turned telomerases off.
DR. BLACKBURN: Yes, and I don't know
if that is the typical situation when one puts cells into culture, and
I thought that they more often would turn off and an earlier subset would
keep multiplying, and again I want you to correct me on that cell growth
CHAIRMAN KASS: Thank you.
DR. VERFAILLIE: So currently we do not know
whether it is often turned back on in culture. If we look at the cultures,
for the first 40 population doublings, the cells appear to grow slightly
And then a second wave of cells grows out and it grows slightly slower.
So initially we thought that maybe the more classical senescing cells
were disappearing, and that those were the cells that were growing faster,
and the you then select for the cell that has inherent — you know, has
the system turned on to not be subject to the clock of aging.
The frequency with which we can grow out the cells from human bone marrow
is we believe one in a million bone marrow cells. So it is a very rare
event, and so it will be quite difficult to actually specifically ask
whether it is turned on and then back off, or turned off and then back
on, unless we can actually do some genetic trapping experiments to try
to ask the question.
DR. BLACKBURN: I'm thinking of David
Beaches' experiments in which he was able to show that cells would spontaneously,
if you keep them in culture, turn their telomerases back on, because that
gives them some selective advantage.
DR. VERFAILLIE: Right.
DR. BLACKBURN: And so I was wondering
if such selected advantages occur in your situation?
DR. VERFAILLIE: It could well be, and so the
culture conditions are very particular, and so I didn't go into too much
But if you do anything wrong to the culture conditions, we cannot create
the cell lines, and so it might well be that it is what we call in my
lab a cultural artifact what we see, which would mean that these cells
may not exist really as such, but actually are induced to become this
long-term proliferating cell by the culture conditions that we put them
DR. BLACKBURN: Thank you.
CHAIRMAN KASS: Janet, again, please.
DR. ROWLEY: I have two more questions. One
is a follow-up of a question that I asked you about a year-and-a-half
ago, on whether out of your MAPC cells you can get hematopoietic tissue.
DR. VERFAILLIE: Well, I think I showed you
in vivo that if you infuse the cells into mice that were either not irradiated
or sub-irradiated, that the cells appear to be able to differentiate into
hematopoietic elements that have red cell, and granulocytic markers.
In vitro, we have had more difficulty to try to do that, even though
it appears now that we can at least get for people who don't understand
this, but what would be yolk sac hematopoiesis, even though we haven't
really seen hematopoiesis that would occur in the embryo proper.
But we can find cells that look like the cells that have been created
at the earlier stages of development, where the initial one is made, which
is in the yolk sac.
DR. ROWLEY: And the other question is more
a more practical question. I don't know precisely how many cells would
be required to treat an adult patient with a particular disease, and are
the number of cells required, or what kind of limitations, using your
system, would be faced if you have not one patient, but hundreds or thousands
of patients that could benefit from a particular therapy?
Is this really going to be an applicable strategy?
DR. VERFAILLIE: I think it is a bit too early
currently to really be able to answer that question. We have been able
to take cell populations and have them undergo 80 to a hundred population
doublings, which is really if you were able to do that and not throw cells
away along the way, it is 10 to the 50th cells or something like that.
So it is an enormous number of cells that you can in theory create.
What I didn't go into too much detail on is that the way that we have
to grow these cells is under very low density conditions, meaning that
the cells have to be far away from one another, or otherwise they do not
maintain their undifferentiated state.
Which is quite different from embryonic stem cells, which tend to grow
in tight clusters. From a bioengineering standpoint, meaning scaling it
up to making hundreds of millions of cells, will be a major bioengineering
question of how we can actually adjust the system to be able to do that.
But on theoretical grounds, you know, if you could overcome all the
bioengineering problems, you should be able to create enough cells to
treat multiple individuals, rather than a single individual at a time.
CHAIRMAN KASS: Question. Robert and then Mike.
PROF. GEORGE: Just a very quick question
of clarification in response to Janet's first point. On this question
of whether they were — whether the teleomerases were turned off and then
turned back on in the culture.
If it is not that, and if that's not what is happening, the other possibility
is that they were never off to begin with?
DR. VERFAILLIE: Correct.
CHAIRMAN KASS: Mike Gazzaniga.
DR. GAZZANIGA: Again, thank you for a very
excellent talk and a cautious talk I thought. I thought it would be helpful
for us to understand the new pressures of a biologist like yourself, which
are the following.
Here you have this fantastically interesting finding, and up until 5
or 10 years ago, the normal way that such things would be treated is you
publish the work in peer review, and then you make the stuff, whether
it is reagents, or whether it is cell lines available to others for reproducibility.
And that is a normal sequence of events that we are all familiar with.
And now we have the bio-med inserting itself into these laboratories,
where all of a sudden it becomes proprietary goods from this work.
When the original media picked up on your story, and I guess it was
The New Scientist, there was this cryptic little paragraph in there about
how they had seen the patent on some of this work, which is a very complete
description, and how does that — what is going on here?
How can — and this is where I would like to go obviously, and obviously
it is good for everybody here to get these cell lines that you have out
to other labs, and reproducibility, and then the process goes forward.
Are you constrained in some way, and has life been made complicated
because you didn't have full public funding and you had to use this other
money, or was that your own? What is going on?
DR. VERFAILLIE: So the work was really done
at the university with NIH funds and university funds, and so there was
really no private funds, except for the small amount from the company
that was listed in the beginning, has gone into the work that we have
And because of the possible importance of the observation, the university,
as well as myself, thought we should get some kind of protection, even
though I am not sure that you can truly patent stem cells, because all
of us have them.
But just such that we would be in a position to work with biotech companies
to be able to produce large-scale numbers of cells and things like that,
which is hard to be funded to known private funds.
So there is patents pending on the cell population. Currently, that
really has not precluded us of collaborating with other institutions,
or investigators within the same institution.
So they have collaborations with 10 or 15 different groups within the
U.S., or outside of the U.S., depending upon the expertise that we need,
to try to recreate the cells in other laboratories, and actually use their
expertise, since I am a hematologist, and not a liver physician or a neuro
And to actually be able to use expertise in other people's laboratories
to move the research forward. So there are some minimal ties attached
to working with the cells, but I think it isn't overcomeable, and it really
has not been an issue with other academic investigators to collaborate
And teaching people from those labs to come and to grow the cells, and
at least start working with the cells. But it is a very complicated and
it is a — and I have had myself a lot of problems in trying to find the
right patent between potential biotech interests and academic interests.
DR. GAZZANIGA: Right, and you are not alone
in that dilemma. So are there other MPAC lines at other institutions now
that behave like yours, or is yours still the Golden Grail here?
DR. VERFAILLIE: We have given out the mouse,
and to some extent, human MAPC lines to other investigators who are now
setting the lines back up. We are also explaining and teaching people
how to create them from beginning bone marrow.
And I know that there is one group in Japan who I think pretty much
as the system set back up from human bone marrow. You know, they still
need to do some additional studies to prove that it is really MAPCs, but
we trained a person from there for 3 or 4 months in my lab, and they went
back to Japan, and were able to it appears to recreate them.
CHAIRMAN KASS: Could I ask a couple of sort of
semi-scientific and semi-practical questions? How hard is it — I mean,
you have just indicated that not many people have already been able to
do this, but how hard is it to find these cells?
And by which I mean two pieces, and in how many individuals in which
you look for them can you find them? And how hard is it to find — how
rare are they, and how hard is it to find in any particular individual?
Both of these questions bear upon at least a preliminary assessment
of how useful this might be clinically speaking down the road, although
things could change where you might be able to enhance the yield.
But could you give us a preliminary sense of this?
DR. VERFAILLIE: I think we have studied now
between 70 and 80 normal humans to try to identify the cells. The age
range, the youngest donor was two, and the oldest donor was 55. The majority
are young adults who want some money to donate bone marrow at the universities.
CHAIRMAN KASS: The two-year old?
DR. VERFAILLIE: No, the 20 year olds. The
2 year old actually did a bone marrow donation for a sibling who needed
a bone marrow transplantation. So we have been able to create the cells
I would say in about 70 percent of the individuals that we have looked
Whether that means that the other 30 percent didn't have it, or there
was some technical issue that came about, and we were not able to create
them, we start out with 10 milliliters of bone marrow, and we would usually
find a few clones that can actually grow out.
And so really the frequency is quite low, and it is one in a million,
and that is at least the estimate that we have right now. But there is
lots of bone marrow and so one in a million isn't an impossible task to
CHAIRMAN KASS: And could I also follow up on the
question of these cells and their promise, assuming the best, and the
embryonic stem cells, assuming their best.
This is not a question of whether one should prefer one line of research
or another, or whether we should now go ahead with them. But is there
anything specific that you could imagine could not be done therapeutically
with these MAPC cells that you would then need cells derived from embryonic
tissue to do?
Or is this in the rosiest division, is this really a substitute, and
one that might even have the rejection problem solved if I am dreaming?
And this is not a question about whether the other research should go
forward, but really what is the best promise of this research so that
at least we can think about it?
DR. VERFAILLIE: Well, I think that the data
that we have in vitro suggests that we can create cells of the three germ
layers of the embryo, and so theoretically, you could envision that you
might be able to make more than we have done so far.
We have made liver-like cells, and brain-like cells, and epithelial
cells, and we have not tried all the other ones. In vivo, the blastocyst
experiment, unless that is a fusion event, and if it isn't a fusion event,
would indicate that the cells hold the inherent promise of making all
the different cell types that make up the tissues, the somatic tissues
of an animal.
So again that would suggest that is under — that if we changed culture
conditions further that we might be able to, for instance, create insulin-producing
beta cells, which we haven't done, or create two heart muscle cells that
function like heart muscle cells, and don't just look like it.
So if all these promises hold true, and if we continue the cultures
and they can be expanded even further into 80 or 90 population doublings,
and so there are lots of ifs here, they may be able to be used to treat
a large number of diseases.
The problem at this point in time is that there is so many ifs that
it is a very difficult question to specifically answer.
CHAIRMAN KASS: Of course, and I appreciate that,
and on the question of the longlivedness, or the half-life of these things,
you have gone through — in vitro is what? It is what?
DR. VERFAILLIE: From 80 to 100 population
CHAIRMAN KASS: And it is obviously too early to
say how much longer, and whether those conditions are matched in vivo.
But when the people say that the promise in terms of longevity for cells
derived from the adults is really much less, is there anything to be said
on that question of the basis of knowledge now had?
DR. VERFAILLIE: Human embryonic stem cells
have been kept in culture now for 350 or 400 population doublings. So
that is 3 or 4 times as long as the adult cells. So we are striving to
go there, but we just need time to do that.
Are these cells going to be able to do that? As far as we can tell,
after 80 population doublings, there is no shortening of telomeres, and
so that means that there is at least another 50 or 60 left.
If for some reason telomerase is shut off along the way for reasons
that we currently don't know why that might be, then the longevity would
be less than what has been shown for embryonic stem cells.
Now, for classical adult stem cells, if you take hematopoietic stem
cells that make blood, but not something else, they would not go for 80
So there is something special about these cells, that they can overcome
this senescent block at 60 or 70 population doublings, which is actually
long for any other adult stem cell.
CHAIRMAN KASS: Thank you very much. Questions or
comments? Janet again, please.
DR. ROWLEY: Coming back again to partly the
real world in this iffy situation, and it is a question of the practicality
for treatment for particular individuals.
It seems to me that the notion that you might be able to derive these
MAPC cells from an individual who had some medical problem might have
some limitations because it probably takes 3 to 6 months, or so to get
enough cells to then be able to use them therapeutically in that individual,
and that is always assuming that the individual has some kind of a somatic
disease, and not the basic underlying genetic problem.
So then the way to get around that if it really is 3 to 6 months, and
you don't have that window of time, would be to do somatic cell nuclear
transplant. Now, have you ever tried that in your MAPC cells?
DR. VERFAILLIE: In collaboration with Dr.
Jaenisch, and two weeks from now we will try the mouse MAPC cells in mouse
eggs, and ask whether the efficiency of nuclear transfer would be closer
to what you would see with embryonic stem cells, and where the efficiency
is much, much, much higher than if you use a classical adult cell.
And that might improve efficiency of making cloned embryonic stem cells.
DR. ROWLEY: But I am thinking of the other
experiment. You have a patient who is desperately ill, and so you would
have cells from that patient, and you would want to use the nucleus of
the patient's cells into your MAPC cells, and so that is a different thing.
You have got these cell lines, and how can you make them more compatible
with the patient, and agreeing that you can't get rid of the mitochondrial
problem unless you do additional manipulations and strategies.
But have you ever tried to replace the nucleus in your MAPC cell with
a nucleus from an adult somatic cell?
DR. VERFAILLIE: No, we have not yet.
DR. ROWLEY: Do you plan to?
DR. VERFAILLIE: We might.
CHAIRMAN KASS: We could always get everyone at
the age of 15 to put away a little bit of marrow for the time that we
might need it.
DR. ROWLEY: Another reason to save cord blood.
CHAIRMAN KASS: This is your chance, council members.
This is a wonderful opportunity. Questions?
DR. ROWLEY: Well, I would just be interested
from Catherine's point of view on her answers to some of the questions,
to the two questions that I posed at the end to John Gearhart, and again
give her the option to do this as a written response rather than a direct
response, but I think it may be easier to — and the second question,
which may be very simple to answer in terms of the kinds of restrictions
that you find now in funding.
And I would assume since you are dealing with adult cells that there
aren't any, but I would be interested in your perspective on the funding,
in both government and other agencies.
DR. VERFAILLIE: Well, currently for the work
that is ongoing in my group, which works with adult stem cells, actually
the amount of funding that has become available through the NIH has increased
dramatically over the last few years to support this kind of research.
So that has not been a problem. I have wanted to compare these cells
carefully with embryonic stem cells, and so we are in a position currently
to do this in a mouse, but mice aren't humans.
And so we have really not been able to do that until earlier this year
when human embryonic stem cell research was allowed in academic institutions
under NIH funding.
And as I mentioned, we had actually gone out and tried to recruit an
investigator with that kind of expertise to be in a position to try to
address some of the questions that have come up here, and are these cells
going to be equipotent.
And I think to date, even though they are exciting and they seem to
be quite potent, I can't really say whether that is the case. And so ultimately
we won't be able to answer this question until we can truly compare them
and not across country borders, but actually within the same institution,
where people can look at the two cell populations at the same time.
And so in that respect, I think that the lack of funding for embryonic
stem cell research in humans has made it impossible up until just recently
to be able to do that.
CHAIRMAN KASS: Michael Sandel.
PROF. SANDEL: I wonder if I could put
to you the same question I put to the previous speaker. Given that some
people regard embryonic stem cell research as morally problematic, what
would you think of the idea of imposing a moratorium on embryonic stem
cell research until we could assess what might be achieved by adult stem
DR. VERFAILLIE: I think that my answer is
very much in line with what you heard from Dr. Gearhart. I think that
the main reason why we — to investigate in the field of embryonic stem
cell — human embryonic stem cell research is to be able to compare and
contrast the two cell populations at the same time.
I also think that what we did in our culture dishes to try to differentiate
these MAPCs into liver-like cells or neuronal-like cells is really based
on what has been learned from mainly the mouse embryonic stem cell field,
where investigators have been able to take these cells and drive them
in vitro to become certain cell types, even though that is not a hundred
percent fool-proof, and it is not completely figured out how you should
So I think if you have a number of different cell populations at the
same time, we try to test all these different questions. What we learned
in adult cells might be applicable to embryonic stem cells if they are
the cells that ultimately will be the suitable source for our clinical
applications or the other way around.
And so I think stopping research in one field actually will slow down
research in the other field, and it would be either way. In other words,
if you stop our research in adult cells, or embryonic stems, as I think
what can be learned in the two systems should be translatable in the other
And so I think if you were to ban all embryonic stem cell research,
it would really slow down the insight that could be gained in adult stem
CHAIRMAN KASS: Rebecca Dresser.
PROF. DRESSER: This is unfair, but I am
wondering if you had any ideas about the cost of such a procedure? I mean,
just based on what you have done in mice, and you mentioned at the very
end that to be cost effective that you would probably would have to just
have a number of cell lines and not rely on the patient cell.
Is this going to be a very, very expensive technology, and where we
have to worry about — well, if all these ifs work out, will we have to
worry about who has access, or will it be comparable to the stem cell
transplants that we do now with bone marrow now? Or what do you think?
DR. VERFAILLIE: Well, I think it will be relatively
expensive if you do it on a single person basis, and you will have to
create the cell lines from the beginning, rather than go going to a frozen
stock of cells, where you have a very well-qualified product to start
with and where you expand cells.
And so you might even have already committed cells frozen as well, and
so the cost to get to that point would have to be incurred once rather
than doing this over, and over, and over again.
The costs I think — well, it is hard to say, because I am not sure
how much of the regulatory issues we have actually complied by at this
point in time to actually truly gauge how much it would cost.
But I think that by the time that you do all the quality control tests
for infectious agents and things like that, that amounts to quite a bit
of money for each cell line that you try to establish.
And in the long term I think it would probably be more cost effective
if you would have a therapy for heart infarcts that you could go to a
limited number of cell lines. And to put numbers of them, I don't really
It would probably be in the range of a bone marrow transplantation currently,
which is quite expensive. So it is anywhere between $50,000 and $200,000
If you had qualified cells that were frozen, and then you could expand
them for a short period of time and do a limited number of tests at the
end, the amount of cost incurred would really be all up-front, and then
there would be a relatively small amount per patient.
PROF. DRESSER: I guess the other thing
is that bone marrow transplants work fairly well with some illnesses and
not with others, and would you expect to see those kinds of results with
these kinds of therapies?
DR. VERFAILLIE: I think that would highly
depend on the type of disease that you tried to treat. You know, you are
all well aware of the treatments that have been used for Parkinson's disease,
which the trials that were done in Sweden have made little complications.
But when this was extended in multiple hospitals in the West, there
were a lot more complications if it was done on a larger scale. So I think
that depending on the disease that you go after that it may work better
or worse, and it is really way to early to be able to comment on that.
CHAIRMAN KASS: Bill, do you have a question?
DR. HURLBUT: Well, if we have time, I would
like to ask a couple of scientific questions if that is all right.
CHAIRMAN KASS: Please.
DR. HURLBUT: Do I understand this correctly
that you are saying that your MAPC is put into the blastocyst to perform
more cell lines than do other adult stem cells?
I thought that adult stem cells generally formed lines in a blastocyst?
DR. VERFAILLIE: There are 3 or 4 papers published
on adult stem cells into blastocyst experiments. There is one paper published
by a German group, where they took purified hematopoietic stem cells,
and injected them into the blastocyst, and what they were able to show
was that the cells gave rise to some hematopoietic elements, and that
they actually recapitulated the developmental behavior of hemoglobins,
which switched at different stages of development during embryos, fetuses,
and then adults.
They did not see any contribution outside of the hematopoietic system.
The second paper is a paper from a Swedish group, where they had taken
neural stem cells that have been cultured, and introduced them in the
blastocyst, and as far as I know, they have never had animals been born
And they saw a contribution to a few tissues, but not all tissues of
the mouse fetus. And in the last papers, we did a paper by Austin Smith,
the one that reported on fusion, where they had taken defused cells and
given them to a blastocyst again, and it showed a contribution in one
animal that was born, and that was really only a single animal, to the
liver and a few other tissues.
But it was not quite the amount of contribution that I showed in the
picture here, where every single tissue of the mouse appeared to be having
a fraction of the single MAPC cell.
DR. HURLBUT: That is very, very exciting.
Another question that I think might be of good general interest to our
council, but he question of whether transdifferentiation is occurring,
or even the process of embryonic stem cells just differentiating, it is
always clouded by the question of how do you know when there is actual
differentiation taking place?
In other words, just because you follow one or two gene expressions,
you don't know, and one of the problems with embryonic stem cell therapies
will be to get the target tissues up to speed, like beta cells producing
I know that there are advances being made on this, but can you just
give us a general description of how you identify when you are satisfied
that a tissue has in fact been produced?
And maybe tell us a little bit about the — maybe we need a little education
on messenger RNA assays.
DR. VERFAILLIE: Okay. The criteria to say
that you produced tissue I think needs to include that you turned on the
genetic program that is compatible with the tissue that you want to produce.
You find therefore proteins from the genetic program in the cells, and
the cells have morphological changes consistent with the cells that you
are looking for, and the cells have functional characteristics of the
tissue that you are looking for.
So what happens in a cell is that in an undifferentiated state a number
of gene programs are shut down, meaning there is no transcription to the
messenger RNA, and you will find no protein, and therefore no function.
During a differentiation process, you come in with a growth factor or
a cytokine, or a stimulus from the outside, and you trigger a certain
set of signals that then open up a new genetic program and the first thing
that happens is that you transcribe messenger RNA, that then gets translated
into proteins and/or sugars, that then supposedly give a new function
to the cell.
So what we have been looking for in vitro, and that is where most of
our work has been done initially, is actually taking an undifferentiated
cell and showing that a certain genetic program isn't turned on, meaning
that you don't find mRNA, and you don't find protein, and you don't find
We then switch the culture conditions and add triggers by trial and
error, to a large extent to try to activate certain genetic programs.
And if we do that, we look for protein and mRNA first.
So we look to prove that the genes are turned on, and then we look to
prove that these gene products actually give rise to proteins. We have
gone to the next step also and actually tried to then take the cells that
we believe that are like brain or like liver, and started asking questions.
If it is a liver cell, it should secrete certain things. It should have
the machinery to detoxify blood and things like that. So we have been
able to show that in the liver lineage, for instance, that we do turn
on the programs to make albumin, which is one of the major proteins that
is being secreted in the liver and is present in the blood.
These cells have, for instance, cytochrome p450, which is a massive
machinery in the liver that helps detoxify the blood components. And we
can show that it is there, and it responds in the correct ways as liver
cells would do.
So that is what you do in vitro, and in vivo, it is a bit more complicated,
and you really need to use animal models where there is a disease. So
you would have to show that the cells ingraft and you can find the donor
You would have to show that they turn on RNA and protein, and therefore
have this genetic program turned on. And then function, which means that
if you take an animal that has a failing liver, and you give the liver
cells to this animal, the animal will now live without having drugs that
keeps it alive.
And so that would prove that the cells that you put in have actually
acquired the ability to function like a liver cell. And so for adult stem
cell research, very little proof of the latter is actually present.
For embryonic stem cell research in mice, there is a lot of evidence,
and in the human embryonic stem cells, that evidence is just starting
to become available, just like it is with adult stem cells.
DR. HURLBUT: Could I ask one last little question?
How many genes are we talking about here; like hundreds, or thousands,
and how many do you monitor in fact?
DR. VERFAILLIE: Well, we usually monitor between
— well, there is probably hundreds of thousands that get turned on, and
so using the new technologies, the array technology, and the proteomics
technology, that is one of the things that we are looking at, because
it will give us a much better insight in the whole programs that are being
We just pick and choose the ones that we think are known to be important
at certain stages of the differentiation. So, for instance, if you go
from a stem cell to a liver cell, we know that you have to turn on X number
of genes that happen to be known to be turned on.
So we look at 2 or 3 that are early, and 2 or 3 that are in the middle,
and then 5 or 6 at the end. We have not exhaustively looked at all of
But I think with the human genome being sequenced, we now have the tools
in hand to now take cells created from stem cells and look at the whole
program of genes that is present, and what we created in a culture dish,
compared to what is actually present in real life in vivo, and get a feel
of how closely we actually are getting to the real cell.
DR. HURLBUT: Thank you.
CHAIRMAN KASS: Could I — Robby, did you have a
question? Why don't you go first, because I have a couple of things as
PROF. GEORGE: Actually, I just wanted
to follow up the question that Dr. Kass asked earlier just for clarification,
and I recognize that there is a great deal of uncertainty as to what the
future holds in your area for research, as well as in embryonic stem cell
And estimating or evaluating what the prospects are therapeutically
is a speculative business, but having all of that in mind — and I was
not clear in responding to Dr. Kass whether you identified some areas
in which knowing what we do know now about the differences between embryonic
stem cells and the MAPC cells, it is possible to identify some areas where
we just know that whatever the prospects are for MAPC cells that they
won't be able to do, or our therapies won't be able to be developed based
on them to do certain things.
And that there is at least a prospect of embryonic stem cells being
used to do.
DR. VERFAILLIE: It is so very hard for me
to answer that question, just because embryonic stem cells have been worked
with for so much longer, and so investigators have been able to, for instance,
make cells that secrete insulin to some extent on demand, which has not
been accomplished with adult stem cells.
There is a little bit of evidence from pancreatic tissue itself that
there might be precursors that can do that, but from MAPCs, for instance,
we have not been able to do this yet.
It doesn't mean that we can't. I don't know that answer. So there is
a lot more experience with embryonic stem cells and there is a lot more
— at least in the mouse system, there is a lot more known on how to try
to trigger certain differentiation programs and whether the MAPCs will
respond to the same extent and to the same degree.
And I think that currently I can't really answer that question.
PROF. GEORGE: But asking if you look at
it and not asking what do we know MAPC cells will be able to enable us
to do, and have a prospect of doing, that embryonic stem cells have a
prospect of doing.
But if we simply ask the question as do we know just on the basis of
the facts of what we know about the differences, and that there are in
fact some things that MAPC cells, no matter what, won't be able to do.
Or is the answer that we just don't know?
DR. VERFAILLIE: I think we don't know currently,
and I can't really answer that question, because we just don't know at
this point in time.
DR. FOSTER: I just want to interrupt with
this one point. Those questions are really hard to answer, but there is
another whole area that is going to impact what you are going to use cellular
based therapy for.
And that has to do with good vectors, retroviral gene therapy, and that
you are going to accomplish with other diseases that you don't have to
use cells for at all.
I mean, the most recent thing in severe combined immunodeficiencies
in humans, is you put a retrovirus in, and you put the common gamma chain
in for about five cytokines, you know, for these kids. It was just in
the New England Journal a couple of weeks ago, or three weeks ago, or
something like that.
And they are now two years out, and so there are going to be a whole
lot of diseases that you are not going to have to use regenerative therapy
or cell therapy. You can't predict those things either at this point.
So I think if you try to jump way ahead of what the basic science is
doing, then you are prone to error, and I know everybody wants to know
whether an adult cell is better than an embryonic stem cell, or something
And I don't think you can answer those questions, and one of the things
that we have heard from both the investigators this morning is that they
cross-fertilized with each other.
And so — I mean, that you could not have done what you are doing in
the adult cells without what had already been done with the embryonic
So I just would argue against trying to push investigators to say whether
an adult stem cell can do this or do that at this point, because we have
not even taken into consideration many other approaches to human disease.
I don't mean to be fussy, but I do think that that is an important thing.
PROF. GEORGE: But I was actually asking
— well, I think the question I asked was that it really is about what
we know now. The question is do we know now that there are certain differences,
that as a result of which the prospects for the one area are different
from the prospects of the other. And I got my answer. Thank you.
CHAIRMAN KASS: Let me take the privilege of the
Chair to expand in a way Dan Foster's comment in a direction that he might
not have intended.
DR. FOSTER: That does not surprise me.
CHAIRMAN KASS: Well, I mean, you are a genial sort,
and I think you won't — I mean, one of the things that one has to remember
in this conversation is that wonderful as the stem cell approach is from
whatever source to the treatment of these diseases, that is not the whole
area here also.
And that the gene therapy is not the whole story as well, and there
are preventive measures, and there are all kinds of other things. I mean,
the conversation, because we are taking it up, gives it a certain type
of dramatic focus and concentration.
But for the people who work in clinical medicine, they know that this
is — that there are lots of ways to try to skin this cat. But I wanted
to ask a couple of — to make a comment, and then ask a couple of questions.
You have talked understandably and very welcomely to us about your own
very exciting work. There is a great deal of skepticism about many of
the published works in using adult stem cells.
And unfortunately, for better and for worse, these reports are caught
up in the political controversy that now surrounds us, with people on
both sides having a stake in either making the results on one line of
work seem better than the other, precisely because they are wed to an
Can you, abstracting from all the political considerations, and the
various axes that various people are grinding on these poor cells, can
you say anything at all generally about the kinds of initial reports of
a clinical sort that we have had with alleged adult stem cells?
Because at least according to some accounts, these have been very exciting,
and yet there is a great deal of skepticism about whether these are in
fact stem cells that are producing the results.
Can you tell the council anything at all about how we should at the
moment regard the news that is coming out to us in this area? How should
we receive it?
DR. VERFAILLIE: There have been several publications
that came out over the last 1 or 2 years now, where investigators or clinicians
have looked at individuals who were transplanted with classical bone marrow
transplantations, and looked in tissues outside of the hematopoietic system
to ask whether bone marrow derived of donor-derived cells could be found
in different tissues.
And the reports that have come out have indicated fairly significant
levels of contribution to certain tissues, meaning they have found cells
in the heart, and they have found cells in skin, gut, liver, and so forth.
And we really have not looked in the same situation to see whether we
can confirm these data or not. I know that some clinical groups have put
in doubt to some extent the degree of contribution that has been reported,
and it is not quite clear whether the 5, 10, or 20 percent that has been
quoted in some papers is indeed actually going to hold up over time.
I think there is some contribution, and the question in my mind still
is how clinically important is it what investigators have seen or what
clinicians have seen currently.
If you go strictly by the term of stem cell plasticity, none of these
studies really show that it was a single cell, or it was a blood cell
that gave rise to these tissues, and it might still be that some contaminating
cells were contributing to that.
And really none of these studies have shown that this has had any clinical
impact on what was going on in these patients. And so they didn't really
show that you restored function of the organ that the cells were found
CHAIRMAN KASS: I was thinking of a recent report
on the Parkinson's cases.
DR. VERFAILLIE: Correct. So the Parkinson's
cases were — and that is with fetal brain tissues, and are those the
reports that you are referring to?
CHAIRMAN KASS: Yes.
DR. VERFAILLIE: And so there has been a series
of patients transplanted in Sweden with Parkinson's disease, where one
team of investigators in a non-controlled study, shows that implantation
of the fetal tissue brain — fetal brain tissue into the brain of patients
with Parkinson's disease could rescue patients, and could actually correct
And actually have now done so for some patients for more than 10 years.
Now, these were highly selected patients, and done by a single group of
The same was done in the west in 3 or 4 institutions, and some patients
got better and some patients did not get better. But I think that gets
to the proof of concept that if you have the right cells, and if you can
create the right cells, and if it is from embryonic stem cells, or adult
stem cells, or from tissues itself, that there might be a way of correcting
But there is again — and I think it would behoove us to really look
carefully at exactly what single cell or fused cells that we have to put
into the brains of patients with Parkinson's disease to try to correct
the disease, and not over correct it as it was done in some of the patients
in the U.S., where they had more side effects from the therapy than they
initially had from their Parkinson's disease.
And so even though there is an enormous amount of pressure on all of
us with stem cell research to try to come up with therapies yesterday,
I have been very, very cautious in telling people that do the clinical
work that you can't just go around and take stem cells and put them in
places in the hope that they will work.
Because we will get into situations like the gene therapy field, where
a couple of awful problems have popped up, and have actually halted the
clinical potential of these cells enormously.
CHAIRMAN KASS: Could I follow that up, because
if there had been more time, I would have asked Dr. Gearhart this question
as well, and you are a clinician who deals with patients that are also
— now thanks to your new results, and I am sure that you are getting
lots of calls as well.
There is an ethical dimension to this area that worries not so much
about where you get the cells from, but how we deal with the desperately
sick patients looking for any sort of hope.
And let me say flat out that in-part to fend off the opponents, the
people in the scientific community and medical community, has to some
extent not been adverse to shall I say hyping the benefits here and possibly
even taking rather cruel advantage of these hopes.
And from what I hear from you, and from what I hear from Dr. Gearhart,
these therapies, there are lots of problems to be solved before these
things will be made available.
And that is not to say that there isn't this enormous promise, but what
can you tell us, or what advice would you give us about we could responsibly
speak about this promise without behaving, let me say, unethically in
dealing with the very patients who are coming to us for help?
And I think that's something that you have probably faced directly,
and whatever help you could give us on that would be welcome.
DR. VERFAILLIE: Well, like Dr. Gearhart, my
e-mail and phone have a lot of messages on them from patients locally,
around the country, and around the world who want to bring a child or
a parent with a certain disease, and want us to treat whatever disease
you can come up with.
And we have to speak the truth, and even though we are excited about
the work that we have, and for the work that people do in embryonic stem
cells, at this point it is a promise, and I don't think there is any data
to say that in the next 1 or 2 years we will actually be in clinical trials
with any of this.
So we really have to tell patients, families, and whomever, that currently
we are trying to cure mice, but a lot of mice have been cured with a lot
of different things, and that doesn't necessarily mean that it will translate
And so we need to do the regular science that needs to be done to come
up with a therapy that is both potentially useful and for certain not
And so that the last part of that whole thing is really where everything
sits. And so we could go ahead and do things now, but then run into major,
major complication issues which would make patients way worse off than
they started out.
You could argue that bone marrow transplantation, there was not a whole
lot known when the first bone marrow transplants were done, and that is
before I started in bone marrow transplantation, and probably some patients
didn't fare that well either in the beginning.
But people ultimately still have to learn by doing it in humans, but
we have to learn as much as we can in culture dishes, mice, rats, and
larger animals before we proceed with therapies for things that are not
And so it is not because you are diagnosed with Parkinson's today that
four months from now that you will die from your disease, which is different
if you have a acute leukemia, where there is really no other solutions.
And so I also think it will have to be graded depending on the type
of disease that you start treating.
CHAIRMAN KASS: And I have one last question, and
I don't think we will have another opportunity in this discussion, but
this comes to Rebecca Dresser's question about the costs, and how to think
about this. And also about the applicability.
There was recently a meeting of the major biotech companies in Princeton,
and our scientific director, Dick Roblin, was there, and they were discussing
among other things the question of the solution of the immune rejection
problem from all these various things.
And all of the ones that were present there are putting their research
money not into somatic cell nuclear transfer to deal with the rejection
problem, but into other means, for a reason that would have never have
occurred to me until it came back from this meeting, which is to say that
if you have highly individualized treatments, case by case, that at least
under present regulatory systems, if you call these things products, each
one of them has to be approved independently by the FDA before it can
And so the question is whether or not — and in partly thinking about
the cost and the scalability, and the things that might make things universally
applicable, doesn't it make sense more to be thinking more in terms of
cell lines, whether embryonic or adult, and that could be made universally
applicable, rather than trying to continue to think each person, his or
her own replacement, given these practical problems of scale and product
I am not sure that the question was clearly put, but it bears upon the
efficacy of this in terms of long term clinical use, and the questions
DR. VERFAILLIE: Ideally, it would be personalized
therapy, and so you would create cells that are completely compatible
with the person that you need to treat, except again in situations where
there is an autoimmunity issue, which makes it complicated.
And if it is an autoimmune problem starting out, then cell therapy is
probably not the best way to go about doing this. For instance, Type-1
diabetes would come to mind, where there is really a rejection of your
own islet cells.
I think the costs — and I spoke to that just a little bit before —
of creating everybody's own cell line will in the long term will be extremely
high, and it will not be a therapy that is suitable for acute events.
So if you have an acute stroke, or if you have an acute heart infarct,
and you try to correct that, there is no way that you can clone ESLs to
correct that, or you could create MAPCs to correct that within the next
one or two weeks.
It just takes too much time to try to do this. Then you could argue,
well, I will store our own MAPC cell lines or own ESL lines just in case
we need it, which definitely I don't think is financially tenable.
So even though the ideal situation would be to be able to make everybody's
own cells, and I think in the long term if the cell therapies are proven
to be, for instance, very useful in patients who have a severe MI that
you can actually correct them almost immediately after the MI has occurred,
or within the next few weeks after it has occurred, it almost has to be
done on an allogeneic basis.
And in that case, trying to come up with wise ways of making the cells
acceptable to the vast majority of patients, whether it is multiple cell
lines and a minimum amount of immunosuppression, or establishing partial
chimerism by creating both blood cells and heart muscle cells from the
same cell lines, for instance, would be one way to get around that.
CHAIRMAN KASS: Thank you very much. Thank you very
much for a wonderful presentation, and a very generous and full response
to our questions. If we might take the liberty of just contacting you
with some other things.
I know that your e-mail is full, and we will try to add very little,
but as we go along, we might have some additional things.
DR. VERFAILLIE: That would be great. Thanks.
CHAIRMAN KASS: Thank you very much. Members are
asked to go immediately from here to the other room, where they want a
group photo, and the four or five of us who have not yet posed for our
individual mug shots, are asked to stay. I think lunch will be served
We will reconvene here shortly after 1:30, and let's say about 1:35
(Whereupon, at 12:28 p.m., a luncheon recess was taken.)