As you remember, our toxicity prioritization index or
ToxPI is made up of of many slices that represent critical
vascular developmental signaling targets, or cellular and
molecular signals that are essential for proper embryonic vascular development.
Over a thousand ToxCast chemicals have been tested
against all of these different targets.
And so each chemical has a unique signature against the putative
vascular disruptor compound ToxPI.
Here you can see the ranking of 1,060 ToxCast chemicals
based on how many of these vascular targets they affected and
to what degree they affected them.
So, a couple of examples here, one is 5HPP-33.
This is a structural analogue thalidomide.
Thalidomide is famous because it's one of the only,
truly known human developmental toxicants due to a tragedy in the mid-20th century,
where thousands of pregnant women took the drug thalidomide to aid morning sickness.
And hundreds, if not thousands of children were born with structural birth defects,
functional deficits, or sadly, were not born at all.
It is known that this particular compound targets vascular development.
And 5HPP-33 is an analogue of thalidomide that was derived specifically
to amplify the anti-angiogenic nature of the parent compound.
It was included in the ToxCast chemical library
as a reference anti-angiogenic compound.
Thankfully, it shows up in the top 5% of theToxCast
chemicals against the vascular disruption signature.
Another example is pyridaben.
This is a mitocide or insecticide also known as
a pesticide that's a mitochondrial respiratory chain uncoupler.
It also has a wide variety of effects on vascular targets in
the vascular disruption signature.
On the other hand, imazamox, which is also a known developmental toxicant in
animal studies, but is not predicted to act via a vascular disruption mechanism,
has almost no activity against this signature, only affecting one out of
the over 25 different targets that are represented here.
It's important to keep in mind that the size of the slice in the ToxPi
represents the potency of the compound against that particular target and
the degree to which that particular target was affected.
So if a compound inhibited the expression of the vascular endothelial
growth factor receptor at a very low concentration, as is the case for
5HPP-33, you see quite a large light blue slice there,
versus inhibition of a particular chemokine expression
which happen at a higher concentration into a lesser degree
results in a smaller orange slice, for example.
To reorient you, I have shown again the adverse outcome pathway for
embryonic vascular disruption in its entirety.
We can adjust this adverse outcome pathway on a chemical-specific
basis to represent the chemical-specific effects on the particular AOP.
Here we can see the predicted AOP for 5HPP-33,
that reference compound that I mentioned.
You can see that 5HPP-33 is affecting the estrogen receptor,
which may control transcription of vascular endothelial growth factor.
It's inhibiting the VEGFR2.
It's inhibiting several signalling molecules in the chemokine pathway,
the extracellular matrix interaction pathway, and vessel remodeling.
This allows us to create a chemical-specific adverse outcome
pathway to try and predict how this particular compound would
disrupt embryonic vascular development and
result in malformations or other types of developmental toxicity.
Most of the targets in the vascular disruptor compound
signature come from the BioMAP system.
This stands for Biologically Multiplexed Activity Profile and
it's a panel of human primary cell systems.
Here you can see the effects of 5HPP-33 and
a whole variety of different cell types.
Interestingly enough,
the four panels of human primary cells to the left of the screen, so the 3C,
the 4H, the LPS, and the SAG, are all vascular-specific cells.
And 5HPP-33 shows a range of effects in those
vascular-specific human primary cells.
It's antiproliferative to endothelial cells, to T-cells.
It inhibits the VEGFR2 receptor as we mentioned.
And it perturbs inflammatory signalling in tissue remodeling.
This is an example of a reference compound where we understand that
the mechanism of action is anti-angiogenesis.
Because it was designed and synthesized for that purpose.
Incidentally, it was synthesized as an anticancer drug to try and
inhibit vascular development in tumors.
To test the putative vascular disruptor compound adverse outcome pathway
with environmental chemicals, we selected a wide range of chemicals
that we know far less about than a reference anti-angiogenic compound.
These 36 chemicals show a range of activities against different
parts of the PVDC signature.
Some of them seem to be targeting specifically the growth factor receptors.
Some seem to be targeting chemokine signalling and
extracellular matrix interactions.
Some only have one slice targeting for example the hypoxia signalling pathway.
And others are completely negative against the signature and
did not affect any of the vascular signalling molecules that we tested.
From those negative compounds, we tried to make sure to include chemicals that were
both nontoxic in animal studies, but also those that were developmentally toxic and
inhibited embryonic and uu development in animal studies.
To try and understand whether or not we could tease apart those
chemicals that were acting specifically via embryonic vascular
disruption as their mechanism for developmental toxicity.
Here on this slide, you can simply see the distribution of those
36 chemicals within in the entire ToxCast chemical library.
So we've tried to cover the range of potential activities and targets.
And we're testing the putative vascular disruptor compound
adverse outcome pathway predictions in a wide range of systems.
We're testing it in small model organisms like a zebra fish.
We're testing it in a virtual tissue platform that allows us to computationally
visualize initial capillary plexus formation in the embryo.
And we're testing it in functional lower throughput angiogenesis
assays that are cocultures of human cells that form tubes.
This concludes our section on testing the PVDC AOP.
Next, we'll be discussing each of the specific testing
platforms that we're using to query our signature and
test whether or not our chemical predictions for
disrupting embryonic vascular development are accurate or not.
Lena SmirnovaResearch Associate
Thomas HartungProfessor
