ISIS and exploding bubbles in the fight against cancer

Cancer Research UK, http://www.cancerresearchuk.org/cancer-info/cancer

Cancer Research UK, http://www.cancerresearchuk.org/cancer-info/cancerstats/, February 2015
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ISIS is playing an increasingly important role in cancer research. Neutron scattering is particularly helpful for examining structures and processes at the molecular and nano level because it enables researchers to see deep into tissues and materials without damaging them. Since Target Station 2 opened in 2009 cancer research has increased by roughly 54%. One particular area of focus is into the nano particles that are being added to the established chemo and radiation therapies. These nanoparticles have either ingenious therapeutic or delivery characteristics and sometimes both.

The scale of the challenge is considerable; in the UK alone 250,000 people are diagnosed each year with cancer. 130,000 of those die as a result of the disease. These numbers are only expected to increase with our ageing population. Due to this, an extra 4.7 million people are expected to die from cancer by 2030.

Toxic chemicals are required to fight cancer, but the toxicity of the drugs not only damages cancerous cells, but healthy tissue too. The cells that are at rest are less likely to be killed by chemotherapy - however the healthy tissues that are constantly growing and dividing such as hair follicles, bone marrow and the lining of the digestive system are more likely to be damaged by chemotherapy. In the human body there are numerous barriers between the drug and its target cells and organs, barriers which can also be raised as the body develops resistance.

Scientists  are working to develop new drugs which are either less toxic to non-cancerous cells or have delivery systems which target drugs more precisely modes of drug delivery to help get the drugs to their desired location. And which can control the rate and exactly where in the body it is released- reducing the number of undesirable side effects.

Exploding bubbles

Francesca Cavalieri from the University of Rome is working to develop sealed microscopic bubbles of medicine to deliver drugs directly to tumours, where their toxicity can be used appropriately. She is developing “microballoons made of protein that could enable drugs to be delivered in a much more targeted way”. They would be effective due to their large surface area to volume ratio and the scope to tune their size and hydrophobicity. Controlled release, bioavailability and elimination are achieved through cooperation between the active molecule, cell and cargo matrix. Francesca Cavalieri has been using IRIS at ISIS and said “I found neutrons a powerful tool to characterize polymeric soft matter.”

She is especially interested in the idea of using targeted sonic waves to burst these delivery systems (the bubbles) when they reached their end destination. "We want to be able to load the balloon with a drug and inject it into the bloodstream. We will be able to target the balloon to specific sites, carried along by the bloodstream and then explode them with an ultrasound when they reach the target area." Dr Cavalieri said.

"If I can functionalise the bubbles there are many potential applications, such as cancer therapy, antimicrobial treatment or clot disruption." Explained Dr Cavalieri.

However little is known about the viability of cargo particles in terms of cytotoxicity and immune response. The implantation of such systems can cause immune responses that jeopardize drug release. And the use of these systems is dependent on maintaining the original conformation of these active agents (the drugs) within the polymer nano particles.

Dr Cavalieri studied the dynamics of water within gel meshes and evaluated these particles as a microdevice for drug delivery. The team tested the use of nano particles from the PVA-MA polymer and its interplay with water using neutrons at ISIS. The loading and release properties (in colon carcinoma cells) of the device were evaluated with the cancer drug doxorubicin in microparticles.

The narrow size distribution of the particles opens up the possibility of administering by injection rather than by drip as the dimensions of the capillaries are comparable to the diameter of the largest microparticles.

Dr Cavalieri and her team looked at the cellular uptake of doxorubicin by the cancerous cells – with the cells taking up about 10% of the drug released in the culture medium.  This gave a >85% reduction of cell numbers in drug treated cultures compared to controls. Overall the study confirmed the possibility of using the PVAMA5S4 “microballoons” for delivering doxorubicin and showed that the drug loaded microparticles fully retain their anticancer activity.

Chemo and endocrine therapy join forces

Polymer-drug conjugates are a new class of antitumour agents. They have a number of advantages over conventional chemotherapy; passive tumour targeting leads to enhanced permeability and retention, there is lower toxicity of bound drug and the potential to bypass drugs resistance.

Maria J. Vicent from Cardiff University carried out a study with the aim to design a polymer conjugate that would for the first time combine endocrine therapy and chemotherapy with the hope of eliciting improved antitumour activity in breast cancer. Hormone therapy is an important treatment in many types of cancer. This is because one of the roles of hormones in our bodies is controlling the growth of cells and organs, but some cancers (such as breast and prostate cancer) use hormones to grow and stopping this can be key in winning the battle against cancer.

The drug tamoxifen is often used to treat breast cancer, acting as an oestrogen receptor antagonist, preventing oestrogen from binding - which is an effective treatment because many breast cancers rely on the hormone oestrogen to grow. Tamoxifen led to the 28% reduction in mortality of breast cancer patients at 5 years. However resistance to tamoxifen can develop which limits it potential. This is because the genetic instability that makes these cells cancerous in the first place helps them to become progressively more resistant to therapies, as they have an abnormally high mutation rate. Aromatase inhibitors are believed to have some ability to stop this resistance forming. These aromatase inhibitors prevent oestrogen biosynthesis by inhibiting the P450 aromatase present in normal tissue and in breast cancer cells of postmenopausal women.

Trials have shown that these aromatase inhibitors are more effective at treating oestrogen receptor positive breast cancer than tamoxifen. The team wanted to give endocrine therapy and chemotherapy attached to the same polymer- this would have significant advantages as they would act synergistically.

The team used neutrons at ISIS to design the polymer drug conjugate “HPMA copolymer–GFLG–AGM–Dox conjugate” which has been shown to act synergistically to give greatly enhanced toxicity. It has already shown activity in clinical trials and underlines the potential importance of these conjugates.

Multitasking nanoparticles

Breast cancer accounts for a quarter of all cancer cases diagnosed in women, making it the most common cancer in the UK. It is highly metastatic, spreading easily to other parts of the body - which leads to an increase in morbidity. However when caught early, treatment is often effective.

Murillo L. Martins from the University of Copenhagen and his team have turned the unfortunate fact that breast cancer cells have a high affinity for hydroxyapatite (a mineral found in bones therefore leading to a high level of bone metastasis) to their advantage and a means to fight cancer. They have developed a multi-tasking nanoparticle to use as a diagnosis tool and drug delivery system which uses this natural affinity. The team produced magnetic nanoparticles, coated in nanocrystals of hydroxyapatite. These nanoparticles will detect and flag the cancer cells by binding to them, which will make cancer cells visible on an MRI scan.

These nanocomposites can not only act diagnostically but they can also treat the cancer, because apatite has been shown to limit the activity of breast cancer cells and reduce metastatic activity. On top of that these nanocomposites could act as drug delivery systems - by carrying anticancer drugs and delivering them to specific tumour sites. Additionally due to the magnetic properties of the nanocomposite they can be directed to the breast cancer tissue using external magnets.

At ISIS the team used the POLARIS instrument to fire neutrons at the nanoparticles and assess the ionic distribution within them. Dr Martins explains “It is very important to tune the magnetic properties of our material depending on its final application. To do so, one of the parameters that must be controlled is the ionic distribution along the magnetic nanoparticle´s microstructure. The neutron powder diffraction data showed that all the Zn ions, which have no magnetic moment, are in the tetrahedral sites of the nanoparticles. It is an important result because such ionic distribution provides the highest magnetic saturation to the nanoparticles and, therefore, a lower amount of material will be required to generate the contrast signal needed for both diagnosis purposes and to carry an anti-tumour drug.”

The next step is to try to encapsulate the antitumour drug into the material. Using neutrons they will be able to see if the drug has really been encapsulated- and if so, whether encapsulation changes it’s 3D structure.

Sugar coating medicine

Research led by Dr Maria Paula Marques at ISIS has shown that coating platinum and palladium based anticancer drugs in a sugary cyclodextrins protects them from being captured by the natural antioxidants in the body. Not only that but it also helps to solubilise the drugs - meaning they can be taken orally rather than injected intravenously.

Cisplatin was the first platinum based anticancer drug discovered and has been successful against many types of cancer - in particular testicular cancer. These metal based drugs bind to DNA in the cancerous cells, therefore supressing cell proliferation. Although the drug is effective in many patients at first, many develop resistance to it. One reason for this is the antioxidant glutathione, which is naturally found in cells- binding to the drugs preventing them from binding to and killing cancerous cells.

Dr Maria Paula explains, “Glutathione is an antioxidant which is needed by us, but unfortunately those drugs that contain a metal, like cisplatin and other platinum agents, bind very strongly to glutathione. This means that when administered, these drugs find glutathione in our cells and they bind to it. This is a big problem as once bound to the glutathione the drugs are not available anymore to bind to the DNA of cancer cells where they should exert their therapeutic effect.”

Using neutrons at ISIS the team have shown that a sugary packaging (the sugar polymeric entities found in cyclodextrins) protects the drugs from binding to glutathione and developing resistance. “We have shown that the drugs are indeed included into the cyclodextrin packet which does not affect the therapeutic agents in any way, simply acting as a carrier.”

“Making these drugs soluble may allow them to be delivered to the patient orally instead of intravenously and that’s a very big step for public health because it avoids several hours for chemotherapy treatment at a health centre. The patient would be able to take the drug at home - patient compliance would therefore be much higher and the economic burden on the government much lower.”

It may be possible to use this method of packaging in other anticancer drugs, selective for other types of cancer. One of the advantages of these cyclodextrins (sugar molecules) is that they are easily modified chemically, in such a way that they recognise only cancer cells and not healthy cells, meaning they could be modified to target cancer cells even more specifically. The next step for the drugs will be to start in vivo testing - they are looking for pharmaceutical companies to invest.

Dr Marques and the team have been users of ISIS for fifteen years, “these kinds of experiments we have been conducting here are something we could only do at ISIS. Maybe they could be carried out at other facilities, but the user support and sample environment at ISIS are exceptional and the reason that has kept us coming back for since our first experiment in 1999.”

Cancer research has wider benefits beyond even the development of new and better cancer therapies. Cancer cells break the most basic rules of cell behaviour - in studying these cancerous cells we discover what the normal rules are and how they are enforced within cells. So in the context of cell biology - cancer has a unique importance and cancer research has profoundly benefited a much wider area of biomedical science then that of cancer alone. It is facilities like ISIS and the development of the soft matter specialising second target station that make this research possible.

Amy Redhead

Research date: February 2015

Further Information

References:

http://www.cancerresearchuk.org/cancer-info/cancerstats/keyfacts/Allcancerscombined/

Exploding bubbles- Cavalieri, F., Chiessi, E., Villa, R., Viganò, L., Zaffaroni, N., Telling, M. and Paradossi, G. (2008). Novel PVA-Based Hydrogel Microparticles for Doxorubicin Delivery. Biomacromolecules, 9(7), pp.1967-1973.

Chemo and endocrine therapy join forces- Vicent, M., Greco, F., Nicholson, R., Paul, A., Griffiths, P. and Duncan, R. (2005). Polymer Therapeutics Designed for a Combination Therapy of Hormone-Dependent Cancer. Angew. Chem., 117(26), pp.4129-4134.

Multitasking nanoparticles- Martins, M., Saeki, M., Telling, M., Parra, J., Landsgesell, S., Smith, R. and Bordallo, H. (2014). Development and characterization of a new bio-nanocomposite (bio-NCP) for diagnosis and treatment of breast cancer. Journal of Alloys and Compounds, 584, pp.514-519.

Sugar coating medicine- paper to be published

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