Shining the beam on anti-cancer drugs

lung cancer cells, Wellcome Library, London

Scientists have been using ISIS to study platinum based anti-cancer drugs. Image shows lung cancer cells. credit: Anne Weston, Wellcome Library, London
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Platinum-based drugs are known for their anticancer ability. Cisplatin, the most famous and most widely prescribed drug of this kind, offers treatment for several types of cancer such as testicular, lung, bladder and ovarian cancers. It works by latching on to the cancer cell’s DNA, preventing cell division, and in doing so marks the cell for destruction. Cisplatin treatment, however, is tough to endure, owing to its significant toxicity to healthy cells in the body, and critically in many cases cancer cells develop resistance to the treatment. In the hope of generating new anti-cancer agents able to overcome this acquired resistance, a group of scientists from Portugal have been using a combination of INS and Raman scattering at ISIS to study cisplatin as well as new platinum and palladium-based complexes with antitumor properties.

Cisplatin was the first big chemotherapy drug discovered in the 1960s and it’s now one of three platinum containing compounds used worldwide to treat a variety of cancers. Administered intravenously it flows in search of its main target - cellular DNA – where it acts to crosslink the DNA, damaging the cells and triggering their death. But how much cisplatin actually reaches the battlefront? Many cancer patients show a high response to the drug initially, but subsequently acquire resistance– several mechanisms for resistance have been suggested, primarily the detoxification of the drug by a natural tripeptide, glutathione, which exists in all our cells as part of their antioxidant defence.

Cisplatin latching on to the cancer cell’s DNA

Cisplatin works by latching on to the cancer cell’s DNA, preventing cell division, and in doing so marks the cell for destruction.
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Scientists use a simultaneous INS and Raman scattering experiments to look at how a natural body antioxidant – glutathione – causes resistance to platinum based anti-cancer drugs.

Dr Maria Paula Marques, Molecular Physical-Chemistry R&D Centre - Coimbra University, Portugal, led the research group, along with Luís Batista de Carvalho. “Cisplatin is toxic to normal cells yielding severe side effects and critically cancer cells often become resistant to it – these are the problems we want to solve in our research. We are looking at platinum and palladium drugs in order to develop new anticancer drugs. The objective of this experiment is to understand how to overcome glutathione-mediated resistance to these types of drugs, which is greatly responsible for the low dosages of compound reaching the biological target. Often only 1/10 of the drug dosage given to the patient reaches the proper destination where the drug really exerts its therapeutic action. This means that lots of the drug is lost during its route to the target. In order to prevent this resistance to the drug we need to get a thorough knowledge of the mechanisms involved and that’s where ISIS comes in.”

“Glutathione is a very potent antioxidant and is essential for us, but glutathione has sulphur atoms,” explains Maria Paula, “Platinum and palladium have a high affinity for sulphur, which means that when glutathione meets these kind of drugs it competes very efficiently for them relative to DNA (where binding occurs to the nitrogens)”.

Platinum(II) is a well-known metal ion for anti-cancer drug compounds such as cisplatin, carboplatin and oxaliplatin. However, the team are also now developing new complexes with platinum(II) and also palladium(II) as they have found that Pd(II) complexes exhibit anti-cancer properties and are often less toxic to healthy cells than cisplatin.

“In our complexes, the Pt(II) or Pd(II) ion is not bound to ammonia as in cisplatin but it is bound to NH2 and NH groups of linear polyamines. These are amines (biogenic polyamines) that exist in every eukaryotic cell being essential for cell growth and differentiation, but when coordinated to platinum and palladium they can act as antineoplastic agents.”

The team have already used ISIS to gain details on the vibrational structure of the newly synthesised compounds as well as on cisplatin´s polymorphic behaviour and on the molecular basis of its interplay with DNA; in this experiment they are using the same experimental technique of Simultaneous Inelastic neutron scattering and Raman scattering on Tosca to take the experiment a step further and study the competition between glutathione and DNA for cisplatin.

“We are adding glutathione to several of our complexes and to cisplatin. We will then use theoretical calculations to help us to interpret the experimental spectra. This will help us to prove that glutathione is binding to the platinum and palladium, where it binds, and what is happening.”

The team also couple their research here with biological assays done back at Coimbra University, where they test their newly synthesised complexes in different types of human cancer cells (e.g. breast cancer, melanoma) to evaluate their cytotoxicity and compare it with cisplatin´s. The compounds are also assessed as to their toxicity towards non-cancer cells.

“Neutron scattering is very helpful because it gives us complementary information to that gathered by infrared and Raman spectroscopies. In addition, being able to perform simultaneous Raman and INS experiments on Tosca allows us to “look” at the same sample, under the same conditions, through two different techniques.”

It’s unusual to be able to solve a problem using only one experimental technique - often multiple techniques are required. Using optical spectroscopies and inelastic neutron spectroscopy on Tosca, however, can provide complementary information on molecular structure and conformation in one shot.

“The capability to make simultaneous neutron and Raman scattering measurements is something that’s unique to ISIS”, explains Dr Stewart Parker, Instrument Scientist on Tosca. “What’s particularly useful from the combining of the two techniques in this experiment is that modes that are strong in one technique may be weak in the other and vice versa. By using two techniques we get a more complete picture. INS is especially useful for the low energy modes which are basically the modes that involve platinum. These are hard to see by infrared and Raman but these are generally quite easy to see by neutrons. These are the motions that tell you directly about the strengths of the interactions between the platinum in various components of the drug itself and in the model system for binding to DNA.”

Cisplatin Sprectra

The Inelastic neutron scattering and Raman spectra for Cisplatin
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A successful collaboration with Diamond Light Source has also allowed to them to obtain a molecular picture of cisplatin-DNA interaction, by EXAFS experiments (at beamline B18) that yielded the local environment of the absorbing Pt(II) center in the cisplatin adducts with DNA (adenine, guanine) and glutathione.

Could sugary armour be the solution to beating detoxification by glutathione?

Initial results from these experiments look promising and the team are coming back to ISIS in March to further develop their experiment. “In our next experiment we are encapsulating our drugs in cyclodextrins - that means that they should, theoretically, be protected from the glutathione.” 

Cyclodextrin a non-toxic sugar that many of us eat every day, as it’s used as a food additive. The team have already been able to encapsulate their complexes in the cyclodextrins back in the Coimbra lab and are preparing these samples for their next experiment on Tosca.

“In this experiment we plan to add glutathione, now not to the free drug, but to the protected drug encapsulated in cyclodextrins. What we wish to see is that there is no change whatsoever in the spectrum of the drug because that means that glutathione is not able to reach the encapsulated drug. We will compare these spectra of the free drug plus glutathione with the spectra of encapsulated drug plus glutathione. From this we hope to prove that cyclodextrins efficiently protect the cisplatin-like drug being a good delivery vehicle for these kinds of cancer drugs. These kind of studies will hopefully provide valuable clues for the rational design of novel cisplatin-like based drugs, displaying a higher efficiency, lower toxicity, decreased acquired resistance and possible oral administration.” Dr Maria Paula Marques.

Felice Laake

Research date: February 2014

Further Information

For Further Information please contact Dr Maria Paula Marques

References 

M. Paula M. Marques, Rosendo Valero, Stewart F. Parker, John Tomkinson, and Luís A. E. Batista de Carvalho. Polymorphism in Cisplatin Anticancer Drug.The Journal of Physical Chemistry B2013117 (21), 6421-6429

Batista de Carvalho, L. A. E., Marques, M. P. M., Martin, C., Parker, S. F. and Tomkinson, J. (2011), Inelastic Neutron Scattering Study of PtII Complexes Displaying Anticancer Properties. ChemPhysChem, 12: 1334–1341. doi: 10.1002/cphc.201001067

M. Adams, S. Parker, F. Fernandez-Alonso, D. Cutler, C. Hodges, and A. King, "Simultaneous Neutron Scattering and Raman Scattering," Appl. Spectrosc. 63, 727-732 (2009).

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