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- It's my pleasure to tell you about our CIRM Tran1 Grant
in which we are pursuing a project entitled,
"Human Embryonic Stem Cell-Derived Neural Stem Cells
for Severe Spinal Cord Injury."
And basically I'm gonna tell you the story
of the remarkable biology of neural stem cells
in the context of spinal cord injury,
and how we're trying to leverage that biology
to move this technology along to humans,
and develop a therapy for even severe spinal cord injury.
And I'd like to at the onset
acknowledge the many collaborators
that are part of this project
moving forward both within my own group at UCSD
and our collaborators at a number of other institutions.
So it's really been a multi-institutional effort
supported by many sources
that are listed here at the bottom.
So human spinal cord injury is a area
of great unmet medical need.
There are about 15,000 new injuries per year,
but there are 200,000 to 500,000 people living
with chronic spinal cord injury in the United States.
And tragically, this often affects people early in life,
sometimes at infancy after spina bifida occurs,
but often in younger adults because of activity,
and then there's another wave in older adults
as osteoarthritis of the spine
can cause spinal cord compression.
So there's often functional loss
that is complete below the level of an injury,
and tragically, there are no treatments
to repair the injured spinal cord.
Why doesn't the spinal cord repair itself?
One of the great areas of advanced and modern science
has been an understanding of why the injured spinal cord
doesn't regenerate, and those reasons are listed here.
If you look at the MRI image on the right,
the area of darkness is a spinal cord injury site
in the human several years after injury,
and this injury site is empty.
And regenerating axons need a bridge
onto which they can grow,
and when the spinal injury site is empty,
they have no bridge.
Secondly, the injured spinal cord doesn't produce
new proteins to stimulate growth.
Unlike the inter peripheral nerve which does.
Third, there are actually several proteins
that form around this injury site that block new growth
even if it were to occur.
And fourth, inflammation often worsens the damage.
And then finally, it may be the case that injured cells
of the central nervous system
need a stimulus to wake up again
and actively reenter the process.
And so I'm gonna tell you the story
of the remarkable biology of neural stem cells
and their ability, surprisingly,
to grow a very large numbers of axons
which are the connections of the nervous system
over very long distances after spinal cord injury.
So let me take you through this now.
The concept that we're trying to approach in this work
is to form new relays across the site of injury.
So this is a diagram of a spinal cord
and the red lines are the wires, the connections
of the nervous system that get cut by the injury site
and the injury site itself that results from
the compression of the spinal cord,
often degenerates and there's an empty cyst.
So these axons are stopped at the cyst and can't grow.
And the idea of a stem cell implant
that's shown in green here
is that these stem cells
will survive and fill this injury site,
that they will enable the regeneration of the injured axons
of the person, the host, to enter the stem cells
in the injury site.
Now, the stem cells themselves will extend new connections,
new axons below the injury,
hence splicing the circuit
and forming new relays to restore a function after injury.
That's the concept.
Why do we use neural stem cells?
Well, because they're developing cells
and they're in an intrinsically high growth state,
and they can fill the injury site as shown here in the rats.
So this is a rat spinal cord,
the head is toward the left,
the tail is toward the right and this green thing
occupying the injury site is a stem cell.
And they can indeed fill this injury site
if provided the right substances to do so,
and can reestablish physical continuity
and represent a bridge for growth.
These stem cells are a unique environment
that allows the injured host axons
to regenerate into the injury site,
and these stem cells in turn extend
the new connections below the injury.
So they can bridge the gap,
they can splice the circuit, and hopefully improve function.
So this is the hypothetical concept.
To what extent do we actually see this happen?
I'll show you that in a second.
So let me just go back a step and say, what are stem cells?
So these are the cells that form the brain
and the spinal cord as we are fetuses
developing before we are born.
So stem cells can become all nerve cells, neurons,
as well as the supporting cells
of the nervous system called the glia,
which are also called the astrocytes
and the oligodendrocytes.
So we start with an embryonic stem cell
and some of these cells after many days of development
become committed neural stem cells,
where they will just generate cells of the brain
and the spinal cord.
We are making the cells at that stage
when they are neural stem cells.
We are making them from an embryonic stem cell line
that was first produced at the university of Wisconsin
two decades ago.
And we use these cells because they are generally safe
and they show controlled growth properties.
And these are called the H9 line
of human embryonic stem cells
that were, again, developed decades ago.
We're not generating new stem cells.
We're taking those existing stem cells
and making spinal cord neural stem cells from them.
So we take those cells,
we drive them to become these spinal cord cells.
And they have to be driven to spinal cord stem cells
because their spinal cord properties
are necessary for them to be effective in neuro repair.
So what do they do when we place these cells
in animal models of spinal cord injury?
So if we don't put anything in the injury site,
then it's empty.
So this again is a rat spinal cord,
it's a very thin section of the spinal cord.
The head is to the left, the tail is to the right,
and this is all the spinal cord, and it's been cut.
This is a severe lesion,
this is as severe as it gets
and nothing fills in the injury site.
This is showing the edges
of the cut spinal cord on either side.
So our first job was to get these cells to survive,
in a sense, flourish in the injury site.
And this took years of development
of embedding these cells in a gel
and adding proteins called growth factors
to enable them to survive and fill the lesion site.
And eventually, my partner in this work, Paul Lu,
and our collaborators in our own group and others
were successful in getting these stem cells
to survive in certain injuries.
And when we could do that,
we saw this astonishing biology emerge.
So we made all of our stem cells green,
we genetically made them green by tagging them
with a protein called green fluorescent protein.
So when we introduce these cells into animals,
they stay green and all of the connections
that they grow outward are also green.
So we can see them and we can track them,
we can track their fate.
So we did a complete cut of the rat spinal cord here,
and we implanted human neural stem cells
in the rat spinal cord.
These are immuno deficient animals
so the grafts were not rejected.
And from the injury site here,
you can readily see the output of tens of thousands
in larger animals, hundreds of thousands of new connections,
new axons streaming down the spinal cord
that has lost its inputs because of the connections.
This is absolutely astonishing biology
that we did not anticipate when we began this work.
And if we look at human neural stem cells again,
put into the right side of the spinal cord,
now we're seeing the spinal cord in a cross section.
So the injury is several sites above this.
And if we look four spinal cord levels below the injury,
everything green here is a human axon
growing through the rat spinal cord on the right side.
And you can see that the spinal cord
is almost replaced by human axons.
This amount of growth is, again,
it's just remarkable that this occurs.
And if we look at a higher magnification in the spinal cord,
we see all of these green human axons growing
among the yellow axons of the rat itself.
And here they are branching up
and going into the areas where the cells are,
the gray matter of the spinal cord below the injury.
So what I've shown you so far
is that we can take the stem cells
and plant them in the injury site.
They survive in the injury site
and they extend all the connections below.
But what about the host?
Does it regenerate axons into the stem cell graft?
Because that has to happen too for a relay to occur.
And the answer is yes, it does happen.
So here are the injured axons approaching an injury site,
again, in the rat spinal cord.
So the white dotted line outlines the stem cell graft
we have placed into the spinal cord.
Normally, the connections of the host, these axons,
would stop right here and not grow.
But now in the presence of the graft,
you can see how extensively they actually regenerate
into the injury site.
They regenerate and they form connections,
synapses with the grafted neural stem cells.
The neural stem cells,
as they send the axons down the spinal cord,
also form connections, synapses, with the host neurons
below the spinal cord that lost their inputs.
So we have spliced the circuit.
Does this support functional recovery?
Well, if we do a cervical level injury,
an injury in the neck,
and then we look at the ability of rats
to retrieve food rewards with their hand,
grasping a pellet,
normally with the injury,
the hand is just closed in a fist, and can't do anything.
If the wait over time
and the rats show very little recovery,
this is the number of pellets that they can reach
if they do not have a stem cell graft.
And here, the red line shows their recovery
if they have a stem cell graft.
So it's not complete recovery,
but they're doing about twice as well
as they would without the graft.
And they're recovering roughly half their ability
to use the hand when otherwise they would recover
quite a bit less.
And if we look at the legs
when we do a complete thoracic transection,
so if we cut the spinal cord completely
here in the back as opposed to the neck,
then the arms are fine in rats,
but then their legs are not functioning.
And so when animals that have this complete cut
at the upper thoracic level,
there's very little movement of the legs.
But now if we put in a stem cell graft,
they show much more recovery in the legs.
It's not complete recovery.
This graph goes up to a scale of eight here,
but a normal animal performs at 21 on the scale.
So it's not complete recovery,
but they're able to move each joint of their legs
and that's what this amount of recovery represents.
And so we have anatomical reconstruction,
we have functional improvement.
So we have moved along this technology now
to larger animal models in preparation
for human clinical trials.
So in these larger animals,
we do partial spinal cord injuries
so that the animals, you know, they're larger,
so we want them to be able to move around the cage
and that sort of thing.
So with this kind of partial cervical injury,
they have impairment of just the right hand.
The left hand works,
the legs work and the gallbladder works.
So they're not in pain,
they're not uncomfortable,
but we can measure whether they have recovery of the hand.
And it took again a long time to develop the methods
to successfully implant neural stem cells
into these contusions spinal cord cavities
in larger animals.
But we were successful and now much of the time,
these grafts survive and fill an injury site,
a contusion injury site.
A contusion, again, it's a bang to the spinal cord,
the kind of injury most humans have.
So we can fill the injury site,
and once again, up to hundreds of thousands
of connections emerge from these implants.
And if we look at the spinal cord two sections below,
two levels below the original injury,
there are many of these green human axons
traveling down the larger animal spinal cord.
If we look at a higher magnification,
look at all of these human connections,
these axons traveling down the spinal cord,
the basis for potential functional recovery.
And in larger animals, kind of like the rats,
we see a substantial improvement compared to animals
that don't have a graft in the ability to use their hand.
Again, it's not complete recovery,
they recovered to about 50%,
but that's about twice as good as animals
that don't have a graft.
And for a human, what this might mean
is an ability to now move the arm
and control a motorized wheelchair
when they didn't have that before,
or perhaps if they have a lower injury,
some ability to begin to manipulate a keyboard.
We won't know until we bring this to humans.
So we're now planning to move this technology
to first-in-human clinical trials.
And thank goodness for the CIRM granting mechanism,
because this is providing us the resources we need
to actually manufacture these cells
for a human clinical trial and meet the requirements
of the Food and Drug Administration
to actually start human clinical trials.
So the CIRM Tran1 grant is funding us
to manufacture cells for human clinical trials.
It's allowing us to develop what the FDA requires
to characterize and release the cells for human use.
They're funding us, CIRM is funding us to make
these so-called clinical cell banks,
they're called Master and Working cell banks
that provide the material to expand,
to treat humans with spinal cord injury.
The grant will allow us to show that the cells are safe
and that the newly manufactured cells are effective
and allow us to identify the minimal dose
and the maximum tolerated doses,
the kind of information that the FDA requires
to enable us to begin this work in humans.
And finally, this work will end with a pre-IND meeting
as it's called with the FDA.
So we are very grateful to CIRM for providing these funds.
We've made significant progress to date
on these first two goals
and are continuing to work on the others that will follow.
And we hope that at the end of the day,
we'll be effective in developing a therapy
for even severe human injury
and to benefit the human condition.
Thank you for your attention.
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