This is the colorful slide that George
Daley was talking about, and it is a
thank you slide as much as a
conflict-of-interest slide. Thank you
all of you, and everybody that helps turn
our sometimes half-baked or crazy ideas
into things that actually have an impact
in society. We've been fortunate
that many of the things we work
on turn out to be exponential
technologies, and what that means is it
can change by two to tenfold per year, so
they're going exponentially, improving
in quality and price, faster than
electronics, which is already a
mind-boggling rate of change. Then this
includes reading, writing, and editing of
DNA - a 10 million-fold change in price and
somehow people still managed to make
money on it even after it's brought the
price down 10 million fold. And so we
brought the $3-billion genome,
which everybody talks about as the Genome Project I'm embarrassed by - I was I was
not a big, you know, started it in 1984 but
I wanted to bring the price down. It did
eventually come down to less than a
thousand dollars about four years ago
and Veritas Genomics was one of the
companies that provided genetic
counseling, but now we think we've
got a business model where we can give
away genomes for zero dollars and
this, like many things I'll mention today,
may seem science fiction ... but
it is, many of these things are past or
present tense. They're in, think of
this like maps and search on the
internet, those are free to the consumer - 
somebody has to pay for it -
but with genetics there are a few ways
that we've discovered I think are
plausible so these are just technically,
these were all done not by filling a
roomful of machines or technicians, but
by reducing it, so that one tube, one
pipette, one of whatever you have
now does billions of reactions with the
same effort of doing one, so it doesn't,
you can scale that up if you want to the
size of the room, but the important thing
is we scale it down
to billions of reactions in a single
small tube. Now the themes that I just
like to develop here, arguments in a
certain sense, is there's an
immediately accessible revolution in
genetics we don't have to wait for, which
is monogenic or Mendelian diseases. These
are so-called simple - nothing's simple - but
these can be done by orphan drugs, by
gene therapy, and I would, I constantly
encourage us to think about genetic
counseling as a cost-effective
alternative to therapy even though a
huge fraction of my research is on gene
therapy. But you can bring down the cost
in ways we'll talk about a moment. Then
there's the alternative, which is
multigenic, multifactorial, environmental
and these are sometimes called common or
complex or chronic diseases and I
think many of these should be or are
being rebranded as diseases of
aging. Ninety percent of us in industrialized
nations will die of of these kind of
diseases but it doesn't necessarily mean
it's a complex treatment, and I'll give
you an example of that. And then there's
a question of whether we're doing
prevention or they're doing reversal,
partial reversal, or full-time cure. We're
mostly focusing on reversal, as you'll
see in a moment, and whether this is
disease specific or taking one aging
disease at a time or whether coming up with a
general set of aging-specific... and we can
do this with cell therapies or gene
therapies. Now here's an example of a
textbook classic multigenic trait, multi-
environmental trait, tens of thousands of
single-nucleotide polymorphisms, and it
seems hopelessly complex, both
diagnostically and therapeutically, but
there are some genes that have very
large effects and these turn out, these
can be turned into therapies, most
notably human growth hormone
somatotropin, which is a single gene,
single gene product protein that works
on all these different diseases, used
clinically. So you might think this is a
silly question for me to be asking, 
is human genomics useful today? But I will
ask for audience participation: How many
people here have their
full human genome sequence available to 
them today? That's a truly remarkably
high number and disappointingly low
number. Its typically less than 1% in the
audience, but I will argue this
is useful, but it's useful enough in a
curious way that we'll get to. The
cost for one of these Mendelian
diseases, and there are about 7,000
of them, has been estimated somewhere,
depending on whether it's orphan drugs, it can be
a hundred thousand to a million dollars
per year per patient, so a lifetime bill
might be as high as $20 million, and
the number is very, I'll give a citation
here, from a Canadian reference, but
the point is it's high. And this can
be recovered if we can avoid Mendelian
diseases through genetic counseling or
other means. This can be preconception; it
could be even premarital. And the reason
I'll say this is quirky is I think it's
analogous to seatbelts. Because we had
seatbelts, they were essentially free they
were standard equipment, they were
required by law, there was ad campaigns.
Nevertheless people did not buckle up,
until this little piece of technology
was it was added to the seat belt, which the
seatbelt wasn't just a passive lock, it
actually triggered a circuit and
stopped an annoying sound. So we need the
equivalent of an annoying sound for
human genetics, which like seatbelts, only
one percent of people will extremely
benefit, but that one percent is
such a big deal that they should do it,
and you don't know that you're gonna be
safe, that you won't be one of the
million people a year that dies of car
accidents or Mendelian diseases. 
Now when we do gene therapy, we have options of
addition/subtraction,
precise editing and epigenetic, meaning
regulatory. There's been a lot of
attention recently to CRISPR which is
mostly subtractive, but I think we need
to think about the whole gamut of
possible interventions and for aging
we've heard a beautiful description of
some and shown some of
the data behind these nine or ten
different pathways that are critical for
longevity and/or aging reversal, but
we have such deep knowledge of these, 
so you can look at it half-empty/half-full, but we
have such deep knowledge I think that we
can start turning these into therapies
quite easily, and the route
that we've taken is gene therapy as
you'll see in a moment. This includes
you know reduction of cells that have
genomic damage or are senescent, the
telomeres, the protein regulation, caloric
restriction, you've heard of many of
these already. So I'm going to go through
very, very quickly. I just want you to see eight or nine examples here
and look for the word "reversal" in here
because I just, I wanted to say that
this is past tense. We have examples
already shown in animal models and now
we want to convert these into therapies
that are suitable, safe, and effective for
humans. So here's a couple of
examples of small molecules the
rapamycin, metformin, I think we'll
be talking about these later on. Here's an
example where there's four transcription
factors, more about transcription factors in
a moment, that regulate processes and
and these have now been turned from
something used in cell biology
in the tissue culture plates
into an experiment that has been
done in mice where you get amelioration
of many age-associated hallmarks.
Telomerase can do reversal and in
particular cases if you have cancer-
resistant mice messing around with tumor
suppressor genes, you can also have
telomerase have its impact without much
risk. We've heard about the senolytic
methodology, the p16 pathway, there are
actually the dark matter of the
genome, the repetitive elements
have been implicated in in this sort of
senescence, and we have a lot of work on
almost every major category of
repetitive elements. These are the parts
that most genome people don't want you
to talk about. It's like the bad secret.
Limb regeneration happens in some
animals, and there's efforts to see if
this can be translated into various
regenerative methods in humans. There is
this heterochronic parabiosis which is
jargon for fusing the circulatory
systems of two animals and/or just
taking the blood itself, and this has
been shown to be valuable for reversing
cardiac and skeletal muscle, neuronal
function in bone, another senolytic... Any time you can count the number
of atoms, that means it's a small molecule, but
our emphasis is on larger molecules.
This is work from David Sinclair, with whom
we collaborate, and it's a segue
into the first of our large molecules.
This is a mitochondrial pathway, which George Daley mentioned,
the Haigis group is working on
as well as David's, and this is
nicotinamide, one way of
restoring the inevitable decline in a
number of small and large molecules that
drop between age six and twenty two
months in mice
and corresponding aging in humans. So we
tackled one step in this the, TFAM down
at the bottom there which results in
loss of mitochondrial homeostasis and we
decided we would tackle it directly,
now using CRISPR not in its normal role
which is "hatchet man," where it just kills
genes, but in this case we knocked out
the nuclease and attached it to a
regulatory domain so it can now imitate
transcriptional activator regulators and
targeted for four different sites
upstream from the TFAM, which itself is a
transcription factor regulator. Sorry if
there's too many levels of regulation
here, but it's it's all good because we
managed to restore - there's a two-fold
decrease in mitochondrial function
via this pathway - we restored this up to
eight fold, which is more than we need, and
so we have some working room for
modulating this. One of the
advantages I should mention, one of
the reasons we keep getting attracted to
to gene therapy is that it is something
that unlike small molecules or even
protein therapies or orphan drugs, this
is something you can do once in your
lifetime and it can have a lasting
effect. It's something that you can
target very specifically to specific
cells or even within a cell, and you
can have feedback systems. It's a much
more sophisticated drug, and I think of
it as like the difference between a
simple transistor and a modern
supercomputer. So what we've done to
generalize that a bit, and here we're
restoring protein functions as
they decline with age for the most part
just by boosting them up. And so we look
through all of the genes that
have these sorts of properties, that have
experiments that had been done in a
variety of animals that indicate that
there's evidence for either
longevity extension, sometimes up to two
fold or more of the life span, or aging
reversal in various animals, and we then,
they're systematically turning those
into gene therapies, which is remarkably
easy to get a proof-of-concept. Noah
Davidsohn pictured here, is a postdoctoral
fellow who almost single-handedly did
45 gene therapies
and then using them in various
combinations to see, because we want to
get all of those nine or ten pathways of
aging, because we think that it's naive
to think that just one of those pathways
will be sufficient, but it might not be
that hard to get nine of them or the
small number of genes, so using two or
three at a time we have now, Noah has
found a combinatorial gene therapy
which is under review at Nature
Biotechnology right now, which hits five
out of five of these age-related
diseases, which is indicative of, but is
not quite proof, that this is getting at
some fundamental aging process. This is
high-fat diet based obesity's were sort
of inducing accelerated aging in many of
these cases, type 2 diabetes,
osteoarthritis, cardiac damage model
recovery, and kidney damage. Now part of
the reason those those gene therapies
were restricted to a subset of
the genes in Pedro Magalhães’
gene age database - he was a previous
postdoc in my lab - it was restricted to
the subsets which are so called non cell
autonomous meaning that they they had
actions that spread to adjacent cells or
throughout the body as a small subset
and it's because adeno-associated virus is not
yet fully tamed. It is among the best
delivery systems that we have. It is the
most approved method of gene therapy
delivery, but it has room for improvement,
and we have now combined machine
learning, artificial intelligence and
synthetic biology loop that allows us to
engineer now over a quarter of a million
different viral designs. These are not
random designs, these are
machine learning based designs where
we've made specific, many specific
changes to the AV virus, looking for
improved resistance to immune problems
and to tissue distribution, so we can
inject this mixture of engineered
viruses into an animal and then take
samples from throughout the body and
find which tissues it's going to and
develop new viruses. And this is a
work of Pierce Ogden who is a graduate
student, Eric Kelsic, who is a postdoc in
in transition.
The hypothesis that we have, one of the
hypotheses that we're using in this work,
is that we can go from almost any cell
type and age to almost any other, notably
from pluripotent stem cells to any
other tissue, and we have examples where
you can go from 70-80 year-old tissue to
almost embryonic and in the other
direction, and the most dramatic case of
this is Bruce Yankner's a paper on
which I was a co-author that just came
out a few days ago (George Daley
mentioned Bruce's work on a different
subject) where we could take
people with Alzheimer's disease, late
onset and so-called sporadic, and healthy
age-matched controls and establish
stem cells for each them and
show that their RNAs are almost
identical, which is a nice control for
the next step, where we turn them into
neuronal progenitor cells and neuronal
networks, and now they're dramatically
different. So even though sporadic means
that by definition they don't have the
classic genetic markers, we can
nevertheless classify these into people
that will get Alzheimer's and those that
don't. And more important than just the
diagnostic component here is that we can
use this for testing new
therapies. So here's example of
differentiating human pluripotent stem
cells. These actually are from my body.
It's more ethical to do experiments
on me than for me to do experiments on
my students. Anyway this is Alex Ng
Parastoo Khoshakhlagh, who are recipients of Blavatnik
support, early adopters in this
Accelerator - very grateful - and here is
producing endothelial cells and bipolar
neurons, initially in separate media and
and then later combined together. This
was actually harder than it looks.
All those little red lines are very thin
capillaries, five microns, just like the
capillaries in your body. There's almost equal number of capillaries
and neurons in your brain ... 86 billion of each and that's what
you're seeing here, is you have these big
nuclei
for the neurons and the smaller red
capillaries floating through them, the
blue nuclei for the neurons. 
That's an intimate connection between
two different cell types that we
developed from a single cell type and
here's two other ones where the
oligodendrocytes wrap the neurons with
myelin. You may have heard of this as
speeding up the conductances through the
nerves that go through the white matter
in your brain, so you can go long
distances at high speed. This is
beautiful wrapping that looks just like
natural. So that one of the examples of a
success story for gene therapy is CAR-T
cells - hopefully you've heard of this.
This is not only a way for T-cells to
fight a variety of cancers but it is, it
kind of shows off how gene therapy, sorry,
how genetic editing can help with gene
therapy in making them much
more immune compatible so you can
actually transplant from any person to
any other person's, sometimes called
allogeinic or universal cells. And we
generalize this even further from any
person/any person to any animal to any
person, and we're in the process of
testing this now. We've made dozens of
changes in the pigs to make them
humanized and missing endogenous
retroviruses so they're less of a risk,
and this is now in preclinical trials
in non-human primates at MGH and
other places, using these pigs. And to do
this we had to overcome natural
inclination for cells to die when
they're cut multiple times with CRISPR.
We made so many changes to the cells we
had to develop ways to stabilize them
and make them happy with that number of
edits, and we've now set the record at 62
and a new record is now at 15,000 edits
in a single cell. It's not published yet -
you heard it here first. And finally I
just want to say that part of the
attraction for engineering organs is not
just to deal with the incredible
worldwide shortage where millions of
people could benefit;
it's the opportunity to do
something that's very hard to do in
humans whether the human organ donors
are just humans in a preventive medicine
sense, which is to make us resistant to
multiple pathogens, to make us cancer-resistent, senescence-resistant, immunity - that
we've been talking about and even
cryopreservation if you want to bank
away organs. This is very hard to do, but
we know how to do it in animals, so what
if we could transplant it from animals
to humans and here's some examples of
some animals that can survive through
full body freezing. So that's all I want
to say. Thanks again not just to the
students and postdocs I mentioned along
the way but here's Bruce Yankner who's helped us with a number of these
neurogenerative diseases and Jennifer Lewis
who's helped with the organs. So thank you
very much and I'll open it up for questions.
Thank you.
