Helen Townley and Xerion | University of Oxford
Chemical synthesis of magnetic nanoparticles
Chemical synthesis of magnetic nanoparticles
Image credit: Shutterstock

Helen Townley and Xerion

I think my personal goal, if it was ever achievable, would be to have chemotherapy with no side effects. There are lots of examples of plants and animals which make ‘poisons’ to protect themselves from predation. But they don’t want to self-poison, so they keep the two components separate (a bit like the two-component glue which only works when you put them together at the point of using them) so they can attack the predator without harming themselves. If we can do the same thing with nanoparticle technology and cancer, to either keep the two components separate until they get to the site of action so that you only get the toxicity in the tumour, or – alternatively – have the drugs in the nanoparticles so that they’re only localising in the tumour and not affecting the rest of the body, we could administer much higher doses, and more aggressively attack the tumour without the systemic toxicity.

We’ve got various uses for nanoparticles: they can be used for carrying cargo, targeting, imaging, or a combination of all three.

In terms of cargo, we make a series of mesoporous silica nanoparticles – they’re about 100 nanometres across (very small), very porous and have a very large surface area, which means they can carry a large load. You can use these particles to deliver, for example, antibodiespeptidesRNAi as well as various drugs.

This student is looking at how to deliver drugs using nanoparticles to get it right to the site of action.

One of my students is looking at the drug LY294002 – it was successful in the laboratory, but turned out to be less successful in the clinic. The problem there was delivery: if you could deliver the drug to the site of action, it could still be as efficacious as it was in the laboratory – it was just going through the body that was the problem. This student is looking at how to deliver the drug using the nanoparticles to get it right to the site of action.

If you’ve got a nanoparticle administered systemically into the bloodstream it goes through the body, but when it gets near a tumour, it falls into it – which is brilliant!

As well as the cargo aspect, you’ve got a targeting aspect. Nanoparticles are very exciting in the sense that they are just the perfect size: a tumour blood vessel has holes in it (fenestrations) which are up to around 350 nanometres. That means if you’ve got a nanoparticle administered systemically into the bloodstream it goes through the body, around the bloodstream, but when it gets near the tumour, it essentially ‘falls out’ through the holes into the tumour – which is brilliant!
Because the tumour is not really an organ, it doesn’t have proper lymphatic drainage (as a typical organ would), which means tumours have a different pressure as well. So not only do these particles come out through these holes, but – because of the pressure – they stay in the tumour. That’s called passive targeting and it’s a function of the size of nanoparticles.

You can also use nanoparticles for imaging. If you dye a nanoparticle (using a fluorophore) you protect the dye from bleaching to a certain extent, and so the dye lasts longer. One of the projects that we were looking at for CRUK was to dye or ‘label’ nanoparticles with near infrared dyes and inject them into a tumour in something like breast cancer and then follow them to see if they drain into the lymph system. One of the big problems with metastasis is that clinicians quite often take out the lymph nodes near the site of the cancer to try and prevent the spread, but removal of the lymph nodes is actually more devastating than you might think, certainly in the groin area – it prevents proper drainage in the leg and you can end up with lymphoedema, which is long-term swelling. So the idea was, if we could see whether there was drainage into the lymph nodes or not, you could make a more considered decision about whether to remove them or not.

The advantage of marking a tumour with a fluorophore-labelled nanoparticle is that the nanoparticle will be retained longer than just free dye.

The advantage of marking a tumour with a fluorophore-labelled nanoparticle is that the nanoparticle will be retained longer than just free dye. This gives the clinicians more flexibility, since it doesn’t matter so much when you do the operation. And once they do come to open the body it’s not normally easy to see the tumour, so having it marked makes it much easier to excise.

That’s mostly everything we’re doing with silica particles. But in addition to that, we’ve got a project using titania particles, which is what our company Xerion Healthcare is based on. Here we’re using the nanoparticle itself, using the intrinsic properties of the nanoparticles to effect the cancer cure. When we started that project, we wanted something that we could just add to current clinical regime, because it’s very difficult to introduce a whole paradigm shift and get it accepted quickly in clinical practice. So we thought, how can we integrate this into what’s currently available and improve it?

We really wanted to be able to use the nanoparticles to access – and somehow be able to treat – much deeper tumours.

There’s an existing technique called photodynamic therapy where you can use photosensitisers (such as titania), where you usually add UV light and excite the particles and it kills the tumour, because they generate reactive oxygen species which damage and kill cells. You can do this on the surface of your body (for example on skin cancers), but UV light doesn’t penetrate very deeply into your skin, so you can’t use this technique for anything subsurface. We really wanted to be able to use the nanoparticles to access – and somehow be able to treat – much deeper tumours. And especially with an eye to those tumours that you can’t remove surgically: a tumour wrapped around your spine, for instance. We needed a way of making those particles usable at depth, and so needed something that was more penetrating to excite those nanoparticles. The most obvious source would be  X-ray, and so could ideally be used with radiotherapy.

To excite the titania we needed a way to capture the X-ray energy and essentially transfer it to the titania so that the titania could then generate the reactive oxygen species. We looked at lots of different elements in the rare earth series that would be able to do this, and in the end used gadoliniumerbium and europium.

Our particles, being around 65 nanometres and spherical, were perfect.

We then designed these particles to be the right size and shape for the job: they’re about 65 nanometres, which is great in terms of the EPR effect. But you also want the nanoparticles to go into the cell, and in order to do that they have to cross the membrane and there’s an optimum size and shape for particles to do that. Cells generally don’t like things that have a high aspect ratio: spherical particles are the most likely to go through the membrane and also (luckily for us) around 50 nanometres is the optimal size. Our particles, being around 65 nanometres and spherical, were perfect. We found that our molecules were being trafficked through the cell and we could see them accumulating around the nucleus in little ‘packets’ essentially. And being close to the nucleus is obviously an advantage in terms of where you want the reactive oxygen species to go. Reactive oxygen species have short diffusion lengths: they’re not going to go very far from where they’re generated. This means you’re less likely to damage healthy tissue which is nearby, because they’re not going to travel very far (although the flip side of that is that you really need a lot of them in the same places for the targeting to work).

In terms of using the nanoparticles clinically there are three main options: 1) direct injection into the tumour (which, having talked to clinicians, is not as difficult as you might think), 2) targeting the nanoparticles to the tumour via the EPR effect (where the nanoparticles, because of their size, tend to accumulate in the tumour more than in the rest of the body) or 3) if you have excised a tumour surgically then, to ensure you’ve got rid of all the cancerous cells, you can then add the nanoparticles into the bed using something like a nanoparticle-gel formulation to mop up all the cancerous cells that haven’t been removed as part of the surgery.

As well as all the nanoparticles work, we’re also doing a big screen on natural products. I’ve always been interested in natural products (I originally trained in botanical biochemistry), and was looking for something to package into our nanoparticles and thought this would be a really interesting area to investigate. I linked up with a group in Brussels who are working on Ophiobolin, which is a fungal derivative with anti-cancer properties. Another student has just finished her DPhil on that and it turned out to be a really interesting molecule – very efficacious against a number of different cancers.

From there, we decided to do our own screen for promising compounds. We linked up with Kiel University in Germany who’ve got an enormous library of compounds. To make the library they sent a message out to say ‘Anyone with any natural products or herbs, please send them to the repository and we will make various extracts of them and create a library for researchers to screen in future.’ It’s a very random library in that sense because it’s just whatever anyone’s sent to them.

We can see from screening the compounds individually that they do have activity, but what we were really interested in was synergy.

We’re screening the compounds individually to look at their activity and screening them against rhabdomyosarcomas (a childhood cancer), as well as breast and brain cancer cell lines. We can see from screening the compounds individually that they do have activity, but what we were really interested in was synergy.

During chemo, they’ll often give you 2 or 3 drugs – the idea is that you're trying to hit different pathways and come at it from all angles. The idea of the synergy between the compounds is that sometimes if you add 2 compounds, instead of just getting an additive effect, you actually end up with much more than a sum of the parts. So the purpose of this screen (of about 250–300 compounds) is to identify what combination of compounds are additive and synergistic and which are antagonistic. And you can’t predict this – if you have 2 drugs in various ratios it may switch from being synergistic to actually being antagonistic! So it’s really important that you find the correct ratios, but the problem you then have in terms of going into the clinic is that different drugs are metabolised at different rates in the body. So you may start out with the correct ratio, but, by the time that they get to the site of action, they might not be in the correct ratio any more and they might actually turn out to be antagonistic.

This is where nanoparticles come in (again!), because if you add the drugs into the nanoparticles in the correct ratio, there’s a much higher chance that they’re going to get to the site of action in the same ratio as they started out. What we’ve found is that many drugs and certainly natural products are stabilised by being in the nanoparticles. Ophiobolin, for example, by itself degrades overnight. But in a nanoparticle, when we release it it’s still in the same state as when we put it in.

 

Could you explain what the broader picture of nanoparticle technology in medicine looks like? How unusual is what you’re doing?
Nanoparticles have mostly been used so far in terms of imaging; that’s where they are most accepted.

Gold or iron oxide nanoparticles were originally used for imaging, but for therapy the amount of gold that you would need in the body for it to be efficacious is enormous and also very expensive. Iron oxide nanoparticles are also interesting because they can be used to cause hyperthermia in cells when a magnetic field is applied. By heating the cells up only a small amount you can cause cell death. This technique is being applied for the treatment of brain cancer.

This is really what we started with: we wanted to use the intrinsic properties of nanoparticles (as in these cases above) and improve upon them firstly by needing far fewer particles to create the reactive oxygen species in a way that was both scalable to make and affordable, and would fit in with the kit that’s already in the clinic.

In terms of delivery of cargo, it’s a difficult one. There are things in clinic like abraxane, which is often cast as a nanoparticle, but it’s not what I would think of as a classical nanoparticle. I think there’s a little way to go before the inorganic nanoparticles, such as silica, are actually accepted for routine use in the clinic.

 

How did you come to have this expertise that sits between medicine and engineering (and botany)?
I started in Plant Sciences and worked on Genetic Modification, and then of course this became very problematic in this country. I decided to move out of that into Zoology on a project about biomimetics. We were looking at how we could use natural products as inspiration for engineering; the project was joint with the Department of Engineering here at Oxford. I was working with Andrew Parker who was looking at butterfly wings. The colour on butterfly wings is not just pigment, it’s actually structure: the colour we see is from the physical interaction of the light with this 3D architecture. We were looking at that as inspiration, since, because it’s a physical structure, you don’t get bleaching (which is pretty significant for things like car paint manufacturers.)

Their silica shells are very highly ornamented – the Victorians used to use them with microscopes as a curio, because they are so beautiful.

We also looked at diatoms. Diatoms are very cool. They’re classified somewhere between plants and animals (experts keep switching where they decide to put them). They’re photosynthetic and can either live in the sea or in fresh water, but they have a silica casing similar to a petri dish – the smaller shell fits inside the bigger shell. They have a really strange way of reproducing. When they part, they take the shells apart but they build the next one inside, so they get smaller and smaller. And they go through this asexual reproduction until they get vanishingly small and then they go through a sexual phase and then go back to the large diatoms again. Nobody quite understands how the sexual phase is initiated, so it was very exciting on the morning that I came into my lab and I suddenly had really big diatoms again! Their silica shells are very highly ornamented – the Victorians used to use them with microscopes as a curio, because they are so beautiful: people would show them at dinner parties under the microscope. The question was: why do these things that are floating around in the sea have these incredibly ornate silica shells?

That was where my interest in silica started – we’d used the shells for attaching antibodies there and I knew it had this porous structure. When that funding finished, I got a professorial fellowship in Engineering which was to use engineering systems for applications in medicine. We were initially looking at microfluidics and that’s when I met Peter Dobson. He’s a big proponent of nanotechnology. The microfluidics project didn't go as far as we’d hoped, so we decided – after getting inspiration from Pete – to change the direction of the project and look at nanotechnology.

What was quite funny, coming from a biochemistry background, was that there was this big deal about nanoparticles, but ultimately – it’s kind of like cooking.

What was quite funny, coming from a biochemistry background, was that there was this big deal about nanoparticles, but ultimately – it’s kind of like cooking. My attitude was always ‘Why not have a go?’ And that’s what I did, I tried various things… and it all seemed to pan out! You still find that: there’s this kind of fear of a different discipline. But once you’ve got the confidence and have the right kit, it’s pretty simple to adapt. I made all the nanoparticles and was largely self-trained with some help along the way from Pete and others.

And I haven’t really looked back since starting in nanotechnology – it was so interesting! It’s absolutely joyous when you go to the TEM (because you can’t see your nanoparticles when you make them) and you see your particles. And something like the mesoporous silica particles: they’re so highly ordered and perfect and beautiful and you think ‘How can we have made this from a test tube?’

I created my lab to really straddle both the biology and engineering sides of it: we can do all the tissue culture, biology, biochemistry but we’re also set up to do our own synthesis so we can make the nanoparticles too. We also have a lot of support from the group at the John Radcliffe who we work with. We get together with paediatric oncologists to try and base things in reality – it’s all very well doing something in the lab, but if you talk to the oncologists and they say there’s no way that we could possibly use that, there’s not much point carrying on in that avenue.

 

What facilities are there in Begbroke? What kit do you need to do your job?
Most of the kit we need for the synthesis is fairly straightforward chemistry kit. But one of the reasons I like Begbroke so much is because there’s the Oxford Materials Characterisation Service – that means we’ve got access to their expertise, which is fabulous, but also they’ve got all the characterisation kit we need: several TEMs, SEMszetasizing - basically all the techniques you need to measure the size, the charge and morphology.

The advantage we have here is that we can do it two ways: you can become trained on the kit if it’s something you’re going to be using regularly. But what’s fantastic is that, if you just want a one-off, you can just go over, ask their advice,  pay a small fee and get that sample run for you.

 

Do you ever feel particularly unusual as a woman in the entrepreneurial space?
It is quite unusual and particularly having been in engineering for a long time, quite often I’m sitting in an enormous lecture theatre and thinking ‘There’s me… and maybe 2 other women… and 400 men.’ And definitely it’s the same at investment conferences: the ratio is incredibly evident. It does give you an opener in some ways, though! Because you can walk up to a woman delegate and say ‘There’s only 3 of us here – can I chat to you?’

 

What’s next?
It’s such a fast-moving field, it’s difficult to say for sure. I would like to do more work on natural products, I think that’s a really interesting area, but I think my personal goal, if it was ever achievable, would be to have chemotherapy with no side effects. There are lots of examples of plants and animals which make ‘poisons’ to  protect themselves from predation. But they don’t want to self-poison, so they keep the two components separate (a bit like the two-component glue which only works when you put them together at the point of using them) so they can attack the predator without harming themselves. If we can do the same thing with nanoparticle technology and cancer, to either keep the two components separate until they get to the site of action so that you only get the toxicity in the tumour, or – alternatively – have the drugs in the nanoparticles so that they’re only localising in the tumour and not affecting the rest of the body, we could administer much higher doses, and more aggressively attack the tumour without the systemic toxicity.

It would be so lovely if you could go to the doctor, be told you have cancer… but it’s ok, it’s treatable. If you think about the change that antibiotics made to medicine, so many things became trivial that were a death sentence before. I don’t think we’re ever going to necessarily cure cancer, but if we can make it so that it’s manageable and not devastating… that would be fantastic. It just takes one small discovery to change the outlook for so many people.

 

What of all your many jobs gives you most job satisfaction?
When I get the data back! It’s so exciting when a student comes back and we have an enormous spreadsheet of data and we’re just sifting through it, the results are dropping out and you can see that something exciting is happening.