0:00
The Glasgow Neurosociety
0:04
in association with SI, or Surgical Neurology International, and SI Digital are happy to present
0:15
the abstracts and discussion of the 10th anniversary Glasgow Neurosociety meeting held in November of 2022 in Glasgow, Scotland.
0:30
Hassan Ishmael is president of the Glasgow Neurosociety at that time. He's from the Wolfson School of Medicine at the University of Glasgow in Scotland and the United Kingdom
0:45
Faculty commentators are Likith L. Akhandi, who's the consultant neurosurgeon at the Queen Elizabeth University Hospital in Glasgow
0:58
And Amy Davidson, a neurologist, also at the University of Glasgow, also at the Institute of Infection, Immunity, and Inflammation in Glasgow, Scotland.
1:11
Other Glasgow neuro hosts were Alidith Middleton, Vice President of Glasgow neuro, and Edica Choudry, another Vice President of Glasgow neuro. Hi, welcome back, everyone So our next stop is
1:27
Andrew Keeneyseid. We'll be discussing Swaridemic Therapy for gliobastoma, focus ultra-mapping, infant throat mitochondria are always in just apoptosis
1:38
and neurosphere models. So they'll pass you over to Andrew.
1:42
Hi, so I'm Andrew, I'm one of the medical students at Dundee, and I've been doing some work with the Center for Medical Engineering and Technology in Dundee as well. So we're interested in looking
1:54
at sonodynamic therapy as an adjuvant therapy.
1:59
glioblastoma treatment. So currently, glioblastoma, it's an extremely malignant brain cancer, which causes patients to
2:10
often pass away within two years, and there's very limited treatment options at the moment. One of the main things is surgery and chemotherapy is standard of care, and there's quite a big movement
2:24
to try and look for additional therapies that can go along with this. One of them, which is quite new, is trying to look at MRI-guided focused ultrasound. There's a number of ways that this new
2:37
technology can be utilized. At the moment, it's being utilized in essential tremor using high-frequency focused ultrasound. We are trying to investigate its use as low frequency, at low intensity
2:50
focused ultrasound to cause apoptosis of tremor cells.
2:56
At the moment. in cancer surgery for glioblastoma. Some people, more often, ALA, five
3:09
fluorescent chemical can be given orally two hours prior to surgery. So what that does is it goes through the blood brain barrier and it goes into cells, it's a precursor to him. And so
3:21
glioblastoma being a very hypoxic tumor, it takes up as much as it can get
3:27
It's metabolised, not quite to him, but to a substrate of that pathway, which is fluorescent. So in surgery, the surgeon can activate a blue light and get a pink fluorescence of
3:46
the tumor to look at margins. We are looking at trying to induce with focused ultrasound habitation of. basically small particles of dissolved gas within the cell. So when those bubbles clap back
4:02
in on themselves, they give off fluorescence, which can cause this, the ALA to substrate to fluoresce, induced roasts, reactive oxygen species, and then go through the normal cell apoptosis
4:20
channels There's a number of big problems that still haven't been fully solved with this therapy. The Barrow Institute in America is trying to do a phase one human trial. Dundee is also looking at
4:38
doing a phase one human trial within the next couple of years. The big unanswered questions are, how does it work?
4:48
And what mechanism does it use to work? And then if you're looking over on a trial, okay, what settings do we need to use? How should we actually try to administer this therapy?
5:00
So what this project is aimed to do is to do three things, really. It's tried to develop a system to test these theories and solve these problems in vitro and in a number of different tumor models.
5:17
We've tried to look at the mechanistic testing using cells. We tried to look at
5:24
the effects of different parameters, so frequency, intensity, duty cycle on both thermal and actual cell death implications that that has. So
5:41
in terms of building the system, you effectively, you have to eliminate air, which is a really difficult thing to do when you're testing cells. Effectively we have an insert that we put into a 96
5:51
well plate that's also merged. We have a transducer in a water bath and The cells sit submerged just into the top of into the top of the water bath in media and Get ultra sounded from underneath what
6:06
we've done is how we have a automated system that we've developed which allows the Plate to get moved and allows for rapid testing Our main findings from this have been that we used a number of
6:24
different fluorescent dyes to look at the production of reactive oxygen species, so does it produce ross and Then to look at apoptosis. So if it creates ross, does it cause apoptosis? So we
6:39
ran the tests using the using 90 millisecond pulses 10 duty cycle and 55 watts. which is roughly what they've used in successful animal models in rats, but there's been no human cell testing of
6:59
anti-tumor effects. So this is a new area, so we're not quite sure what we should be using. So this is our best guess.
7:08
So we found that it produces Ross selectively, and you have to have both given five ALA to the cells and have exposed them to ultrasound to get any Ross production.
7:21
We then tested it again with apoptosis. We found the same selective induction of apoptosis of GBM cells. So this kills tumor cells in vitro. There was no necrosis. So it's not that it's calling
7:35
the cells to break down and release tumor factors into the extracellular environment. It's causing controlled cell death, which is an important thing to look at. Another thing we looked at is we
7:50
gave a Ross scavenger and then ran this whole thing again. So effectively what a Ross scavenger does is you give it prior to doing any therapy that would cause Ross and it takes it all in.
8:06
We ran the tests and we ran it with and without the scavenger. If you don't, if you scavenge Ross and you take Ross away, you do not get apoptosis Therefore, it is the Ross that causes apoptosis.
8:18
So we have a better idea of what actually causes the cell death in sonatodynamic therapy for gluoblastoma.
8:28
Going on from this, what we looked at doing is using simulations and we had a number of different field mapping simulations to try and figure out, right, if we're trying to improve this system, we
8:40
can always just use a hydrofoam, which is effectively like a measuring device goes into a well, a measure. and map all the field. We have to be able to find a way to simulate it, which also
8:52
pushes us into more rapid determinations of different setups for more accurate dose determination. 'Cause that's one of the big things is, if we're going to try and translate this to practice, we
9:04
need to know what we're giving in vitro so we can know what to expect in people.
9:10
We tried neurospheres, so we've looked at both formation and form dassays, and I'll explain what those are. Neurospheres are 3D-organoid cultures, so you dissolve, well not dissolve, but you
9:25
trypsonize and split up
9:28
a cell culture into single cells that don't aggregate together. You plate them in stem cell media because they are stem cells, and they grow their own mini tumors. And so they're extremely good
9:44
models at looking at GBM, One of the problems with GBM is it's a very hypoxic tumor and it has a hypoxic and necrotic core. In a normal cell culture, you don't actually get that necrotic core, you
9:55
don't know how that interacts. We will use these mini tumors to actually look at how does aerodynamic therapy work with almost a micro tumor environment with the actual same structure as a human GBM
10:13
in situ
10:14
So we've given the ALA, we've given the FUS, and we are going to do one of two things. We're either going to plate the individual cell, treat them and wait and see how they grow after three weeks.
10:27
That's a formation assay. So in that case, synodemic therapy reduced the diameter of those small tumors by 83 and that was significant. And that was selective to synodemic therapy So that's early
10:42
and focused off to some together.
10:45
We then looked at - A formed assay, so where you've plated the cells, you let them grow for three weeks. You then treat them and you continue to watch them for another three, that reduced their
10:56
continued growth by 53, so more than half. And that again was significant and selective for sonodynamic therapy.
11:06
What this all tells us is that we have a way to simulate
11:13
sonodynamic therapy quickly We can change the parameters and run, say, 10 different parameters a day, so we can find out more rapidly what parameters we should be using for upcoming human trials.
11:25
We know
11:28
that mitochondrial raw, so we looked at confocal microscopy with our fluorescent dies and we could localise those to mitochondria, which is where this heme metabolism is occurring and where, put
11:42
before nine, the fluorescent substrate is located. that's where the Ross is being produced and that's also where apoptosis is being triggered from. We've been able to look at why is apoptosis
11:57
caused and the fact that
11:59
we've now sort of figured out that it is because of this Ross. We've looked at tumor models, so in with
12:08
the new chronic cores that we've got, we know that GBM
12:14
cells are susceptible to sonodemic therapy and human stem cells as well. We tested in a variety of different types, so from everything from borderline cells to necrotic cells to peripheral cells,
12:28
all of them had selective apoptosis Going on to sort of
12:35
look at, okay, this is all great. How does this actually change anything? We were able to effectively run mock.
12:49
examples of different field parameters, so we know that if we give a cell X dose, we get a certain response. We can run effectively almost many trials of different doses to know more quickly what
13:08
we should be giving in humans, rather than having to do a whole year cycle of a specific dose tweak it a whole year cycle tweak that and run it again. We can do it in maybe a week rather than a year.
13:22
And we can do far more in-depth assays as well, so we looked at western plotting, we looked at a variety of things I won't bore you with that, but
13:35
we basically got an effective rapid way of looking at how sonodynamic therapy can be used. We know it works and we know that it can be. an alternative to treat GBM without surgery, without
13:50
chemotherapy, or without radiation dose. And that's me, thank you.
14:01
Thanks very much Andrew, that was, that was really interesting. Now I'm a neurologist to trade so apologies Mr. Alexander will be much more knowledgeable about this, so I'm going to ask probably
14:10
some fairly straightforward questions and you're going to be like, but um, so when we're talking about this in the glioblastomas, um, and you talked about it being an adjuvant therapy. So are you
14:22
talking about we would do this beforehand to shrink the tumor down before going into reception or something if we've tried to debug rather than completely resect, would they then go in and I guess an
14:33
additional to that. So I obviously have a tiny bit of experience about the essential tremor cycle, the high frequency, but do you still obviously, if you need to see if it's, if it's lighting up,
14:44
still needing a crane automate, still going to be a fairly invasive procedure, even if we're using ultrasound. Sorry, that was two questions at once, but take your time Okay, no, no, so
14:54
effectively what there is, is,
14:58
We'd be suggesting this as a treatment that could be used anytime in the patient's journey. However, I think the initial thought is to use it on tumor margins. So it's not possible to achieve
15:14
complete resection of every single cell in
15:20
a resection of a GBM. Also a very long time ago, the biopsied both left and right hemispheres. So the lateral and contralateral hemispheres from a tumor and they found that small seed cells had
15:36
permeated their way through both hemispheres. So it's not necessarily a surgically curable disease alone. So we'd suggest looking at it
15:53
initially after surgery.
15:57
with your second point about comparing it to a central tremor. So that high frequency looks to ablate it. Or we use far, far lower intensities. And we actually have a thermal effect of about 02
16:04
degrees rather than 20 degrees. So it's far less uncomfortable for the patient to actually go through. You mentioned the craniotomy to see if it's
16:23
lighting up. We don't do that. So it would be completely transcranial We wouldn't have any surgery. You'd have to get haloin as with an essential tremor. You'd still have that big fluid cap. So
16:35
you'd still have to have your head shaved. But it's as non-invasive as possible to kill cells. And it would have probably less risk than radiation therapy or chemotherapy, my cytotoxic chemotherapy
16:54
And so I take it, it's minute like. minimally damaging then to normal cells. I mean, there'll be a small uptake of your fluorescence throughout normal cells with, you know, metabolism, but you
17:06
would be aiming to run the ultrasound over a large field. So not just the tumor area, you were trying to reach kind of seeding. So not quite as phallic, but, you know, is it with the AMB that
17:17
you covered more than just the area that you suspected the tumor to be in against this matter? So initially what we'd think is we'd aim for tumor margins So where we thought there might be tumor
17:27
cells. But the thing about sonodynamic therapy is it's almost double selective. So not only can we aim within the millimeter where we want the focused ultrasound to actually be, where we want the
17:40
dose to sort of distribute itself, the ALA is only up taken by the GBM because of two factors. The GBM itself disrupts the blood brain barrier.
17:54
And so that allows it to permeate through. And also it's hypoxic nature helps it actually gives it active uptake through an additional channel, which is a mutation. So we saw effectively just even
18:09
so our controls were actually other tumor cells. So we compared it to, so giving ALA alone to a tumor cell, it makes it a bit unhappy, but it's not gonna kill it. Giving focused on the lung does
18:22
nothing So the focused ultrasound is inert unless they've already been exposed to five ALA. GBM also, it has a enzyme deficiency, which stops it actually clearing proof before in nine. So it's got
18:41
a small natural accumulation that we're basically just bolstering by giving it oral or IV ALA
18:49
prior to practice onodynamic therapy
18:53
Thank you very much, that would be really helpful.
18:57
Okay, can I ask, so
19:00
basically it's a very interesting concept, thanks a
19:05
lot for that. My question, the first I've asked two parts, the first part is more of a practical point of view. Okay. As a surgeon, that's what we do and that's what you're interested in
19:17
basically So in real life, first of all, this question of biopsy, you wouldn't want to do something with the radiological diagnosis, you need a tissue diagnosis, you need to have a biopsy. So
19:31
operation, you know, until you can get excellent MR spectroscopy to get
19:39
the gene, but the molecular diagnosis, we'll still need some tissue diagnosis Secondly, we know in normal practice 5LAs, the uptake of 5LAs in
19:50
GBM is not uniform. There are some parts of GBM where you actually don't see any expression of 5A and no fluorescence at all and there are some where your intensity of the pinkness, you know, it
20:03
decreases as we go. This is for tumor cells which are infiltrative. So obviously there is a variation in the level of expression of this 5A So from
20:18
your ultrasound point of view, how do we know first of all if the tumor is expressed or is sticking up viral clavs in several tumors which don't have any fluorescence at all. And if it is doing,
20:32
for example, you know, even if the patient is taking it orally, it may not be absorbed or it may have enough time to release the tumor or it may not So how do we know that we need ultrasound, you
20:46
know, like a focus therapy?
20:49
Did you, did you get the question? Sorry. My internet may not be there.
20:53
So, let us part one of the question and we'll come back with it on to discuss this bit here. Sure, so first you mentioned tissue diagnosis. Initially, any trials would be post-surgery, so we
21:07
would have a tissue diagnosis and that would, I think, be required for enrollment. In
21:13
terms of the different expressions and different cell lines expressing or having different uptake of 5A late,
21:22
that is true. So we did most of our testing in GBM22, which is a fairly resistant cell line. So we basically gave it almost a steel man-type scenario where we tested our experiments against the
21:41
most resistant cell type. One of the issues we actually ended up running into is that our dose was a bit high for some of the other cell lines, The tumors, even not exposed to FUS were a bit
21:56
unhappy.
21:59
But what we actually think was, this was down to is that weren't able to clear metabolites, but in humans with circulation, that wouldn't be the case. So that would be fine.
22:10
There is a difference, you're all right. And that's one of the reasons we use neurospheres because not only do when they become a hypoxic and necrotic does it change how the LA circulates through
22:22
the cells, but when cells are able to aggregate together the share waste products and they have a way of basically trying to spread the burden of reactive oxygen.
22:39
We think and because we tested them in the neurosphere models that that would be fine. And we tested them across a wide in cell lines alone GCE both. across
22:52
gin and standard GBM cell lines.
22:58
So everything from fairly central cells to quite peripheral.
23:05
I think that's a valid thing in a neurosphere, it's definitely better than cell lines. But when you come to real in vivo, there is going to be a big hurdle because it behaves very differently.
23:17
That's a good thing. The second question is more of the signs behind it Now, the rosus are part of, you know, are required for many cellular functions, not just tumors, isn't it? You know,
23:29
they can be in edema, the edema, there is necrosis, there is some ischemia. So there is various, you know, the ros involvement. So what is the effect of this focus on ultrasound on these kind
23:42
of cells, which are part of a cancer, you know, a tumor? And like you mentioned, the tumor is not a surgical, The garb-bastama is not a surgical disease, it's a disease of the. of the brain,
23:52
so there's nothing to cut out. And what you're proposing is, in fact, as well as you're removing bits of cells, it's not re-conditioning the whole brain. So the question is, the rawest part of a
24:06
general metabolism is not just a tumor thing. It can happen in ischemia, edema, infarction, et cetera. So will that be very specific for this? So it is So Ross, you're right, is a general
24:21
metabolite, and it's involved in a number of different pathways. What we looked at is, we did actually look at some of these pathways. So we looked at things like cell signaling pathways, either
24:33
24
24:36
hours before or two hours, sorry, 24 hours after or two hours after treatment. So even in, you're all right that in, if you just give ALA or if you expose it to FUS, There is a slight increase
24:50
in cell signaling. terms of self-stress, from changes to Ross metabolism in other cells, sort of other tumor cells who haven't been exposed to both ALA and FUS, either control ALA alone or FUS
25:08
alone. That resolves within 24 hours and there's no sort of actual damage to those cell cultures. The only ones that cause apoptosis, and it causes apoptosis alone, it doesn't cause necrosis, and
25:22
it doesn't actually cause just complete lysing of the cell either. It is selective for apoptosis, and the other cells, though they may have a transient disturbance in metabolism, these would only
25:42
be tumor cells which are actively uptake, and the actively uptake
25:46
in 5LA, unable to clear Put before 9. and are also disrupting the blood brain barrier. So it's all these different factors, select for. Yeah, okay. That's a good idea. Yeah, you're selecting
26:00
it, but it's a method. So yeah, I won't take up much, but this is my area. So one more final question. So have you thought about using it
26:13
intraoperatively? For example, you know, you've done a five-elegant resection and I look at the bed and there is a little faint hints of
26:21
being kind of ultrasound to target these.
26:26
So is that a possibility? It's a great mechanism in what you have proposed is a really good study and I congratulate you and your team on that. So for that sort of scenario, we wouldn't be looking
26:35
at sonodynamic therapy, we'd be looking at photodynamic therapy, where instead of - No, I'm asking specifically about sonodynamic but therapy intraoperatively instead of the light So why the
26:37
requires.
26:49
or the current systems which use focused option are MRI-guided unless it's it
26:56
be very difficult to clear or use intra-operative diagnostic ultrasound to look for tumor residue, put a probe inside the tumor cavity. So it's an idea, but this, you know, a thought process for
27:09
you to have. Anyway, I think that's the end of my question, yeah Yeah, the different, the current system, it's not one probe. I think
27:20
the proposed system for human use is 1200 ultrasound transducers, extremely high power, much more than what we'd be using for diagnostic use. That's why we're having to worry about thermal effect,
27:36
which it's not an issue for us, but it is for higher frequencies, higher intensities than in that system. But it. an intraoperative thing would be looking at the same mechanism, which also we've
27:51
looked at in the same models. So we know that, again, look through this research that Ross has produced an apoptosis of produced because once you activate protein before nine, we're in a common
28:03
pathway between sonodynamic and photodynamic therapy. Interoperative photodynamic therapy is a proposed new addition which would fit the scenario that you're discussing But that's been tried in the
28:16
past and it's not been grown breaking to say the least, but anyway, so yeah, well done. That's a good paper you've done.
28:27
Thank you.
28:30
Thank you very much. And I think on this, there's any other questions any other questions from the panelists?
28:37
Well, and I think that in that case, that concludes talk to band. We hope you enjoy these presentations.
28:46
The information provided in this program is for informational purposes and is not intended for use as diagnosis or treatment of a health problem or as a substitute for consulting a licensed medical
29:02
profession
29:06
Please fill out your evaluation of this video to attain CME credit.
29:14
This recorded session is available free on snidigitalorg.
29:21
Send your questions or comments to
29:27
osmondsnidigitalorg
29:30
This program has been supported by the James I. and Carolyn R. Osman Educational Foundation, owner of SNI and SNI Digital and the Waymaster Corporation, producers of the Leading Gen Television
29:47
Series, silent majority speaks and role models and the new Medical News Network
30:15
Thank you.
View Transcript
Listen to Podcast