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Photoactivatable metal complexes: exciting potential in biotechnology and medicine?

Event

Starts:

June
202012

09:00

Ends:

June
212012

17:00

Location

Kavli Royal Society Centre, Chicheley Hall, Newport Pagnell, Buckinghamshire, MK16 9JJ

Overview

Satellite meeting organised by Professor Peter J Sadler FRS, Professor Akhil R Chakravarty and Dr Nicola J Farrer.

Event details

Photoactivatable metal complexes present relatively unexplored potential for novel applications in biotechnology and medicine, ranging from functional imaging agents, diagnostic agents and biosensors, to target-selective photochemotherapeutic agents with low side-effects. But how can this potential be realised? This meeting will bring together photochemists, photophysicists, biotechnologists, medics and industrialists who will critically assess the prospects for rapid advances in this highly interdisciplinary and emerging new field.

This meeting was preceded by a related discussion meeting Photoactivatable metal complexes: from theory to therapy 18 - 19 June 2012.

Biographies of the organisers and speakers are available below and audio recordings are freely available.

Event organisers

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Schedule of talks

Session 1: Photorelease of small molecules

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Chair

Professor Peter Ford, University of California Santa Barbara, USA

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Design of transition metal complexes for improved ligand photodissication and multiple reaction pathways

Dr Claudia Turro, Ohio State University, USA

Abstract

The presentation will include comparison of quantum yields and ultrafast spectroscopy pointing at different pathways for ligand exchange from the initially generated singlet in Ru(II) complexes.  The population of both the triplet ligand-field (LF) and the triplet metal-to-ligand charge transfer (MLCT) states from the initially populated singlet MLCT is believed to result in a dramatic increase in ligand exchange in some complexes.  The bifurcation from the singlet MLCT  is also explored for potential use in PDT schemes.  Exploration of other transition metals and drug release will also be discussed.

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NO and CO releasing materials

Professor Dr Alexander Schiller, Friedrich-Schiller-University Jena, Germany

Abstract

Nitric oxide (NO) and carbon monoxide (CO) act as important messenger molecules in the human body.[1] NO- and CO-releasing materials (NORMAs & CORMAs) are impor-tant for the development of gasotransmitter delivering devices for therapeutic purposes; toxic metabolites after gas release are kept in the matrix. NO and CO photodonors use light as a convenient non-invasive on/off trigger since it allows the accurate control of site, timing and dosage.[2] Here we report the concept of embedding water-insoluble, photoactive NO and CO metal complexes into nanofibrous polymer non-wovens.[3] NO and CO release into the surrounding medium is performed by light stimulation of the high surface area materials.

For the generation of NORMAs, novel rutheniumnitrosyl complexes {RuNO}6 have been synthesized.[4] For the CORMAs, Mn2(CO)10 and Fe(CO)5 were used.[5] These metal complexes were non-covalently embedded into nanofibrous polylactide non-wovens by electrospinning.[6] The hybrid materials showed differences in their fiber morphologies. The NORMAs exhibited a smooth fiber matrix; the CORMA with Mn2(CO)10 gave highly porous fibers. Identity of the metal complexes was retained in the electrospinning process. Leaching of the metal complexes out of the polymeric matrices into water was negligible due to their water insolubility. Cytotoxicity tests of the NORMA with 3T3 cells (mouse fibroblasts) revealed a very low mortality rate. Irradiation at λ = 366 nm (UV-A) in water, using an adapted fluorescent NO assay, showed a significant phototriggered NO release from the non-wovens.[4] Rapid CO release from CORMAs was detected by the myoglobin assay and vibrational spectroscopy.[5]

[1] R Motterlini, L E Otterbein, Nat Rev Drug Discov 2010, 9, 728. 
[2] D Crespy, K Landfester, U S Schubert, A Schiller, Chem Commun 2010, 46, 6651. 
[3] A Schiller, in Molecules at Work. Selfassembly, Nanomaterials, Molecular Machinery, (Ed: B Pignataro), Wiley-VCH, Weinheim, 2012, 315-338. 
[4] C Bohlender, M Wolfram, H Görls, W Imhof, R Menzel, A Baumgärtel, U S Schubert, U Müller, M Frigge, M Schnabelrauch, R Wyrwa, A Schiller, J Mater Chem 2012, 22, 8785. 
[5] R Wyrwa, M Schnabelrauch, C Altmann, A Schiller, DE10 2012 004 132.2, patent pending. 
[6] A Greiner, J Wendorff, Angew Chem, Int Ed 2007, 46, 5670.

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Photoactivated CO releasing molecules and their bioconjugates

Professor Dr Ulrich Schatzschneider, Julius-Maximilians-Universität, Germany

Abstract

In addition to nitric oxide and hydrogen sulfide, carbon monoxide is now well-established as the third endogenous gasotransmitter in higher organisms, with a delicate balance of cytoprotective vs. cytotoxic activity, depending on the local concentration in tissue. Since CO gas itself is difficult to dose and handle, potential therapeutic applications of carbon monoxide in human medicine will rely on the development of easy-to-handle delivery systems for this gasotransmitter.

A particularly attractive way is the inactivation of carbon monoxide by binding to a transition metal center. Such "caged CO" could then be released from the metal coordination sphere by a proper stimulus at the target site in the organism. Various triggers have been explored in the field of CO releasing molecules (CORMs), including ligand exchange reactions in buffer, enzyme-triggered CO release, and changes in the redox environment of the CORM. In contrast, the Schatzschneider group focuses on the photoactivated release of carbon monoxide from transition metal complexes in PhotoCORMs.

After a short introduction on endogenous CO production by heme oxygenase enzymes, we will discuss the general requirements for therapeutically useful CORMs and present our most recent results to achieve light-triggered carbon monoxide delivery from PhotoCORMs at high excitation wavelengths and incorporation of these systems into active and passive targeting vectors like peptides and nanoparticles.

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Targeted nitric oxide delivery: a novel approach to photochemotherapy for malignancies and infections

Professor Pradip Mascharak, University of California Santa Cruz, USA

Abstract

During the past three decades, the roles of nitric oxide (NO) in various physiological and pathological pathways have been firmly established. This tiny diatomic molecule plays very different roles in biology depending on its concentration. At nanomolar concentrations, it acts as a signaling molecule to control blood pressure and neurotransmission. Quite in contrast, at elevated (micromolar) levels, the same molecule triggers programmed cell death or exhibits strong antimicrobial effects. We have recently designed and synthesized several molecules that can selectively deliver high doses of NO to malignant sites under the total control of light and cause cell destruction.  These unusual NO-donors are ideal candidates for Photodynamic Therapy for skin cancer.  In addition, they can be employed to combat other localized cancers such as prostate and breast cancer.  We have also been successful in eradicating drug-resistant pathogens via NO delivery.  Since bacteria and fungi seldom exhibit resistance to NO (at least for now), treatment with NO offers a new and effective line of treatment for chronic infections. Inclusion of our compounds in biocompatible materials makes the delivery platforms quite safe and target-specific. This talk will include discussions on the fundamentals and potentials of these intriguing treatment modalities.

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Session 2: DNA and targeting

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Chair

Professor Chi-Ming Che, University of Hong Kong, P R China

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Interactions of intercalating photooxidising dipyridophenazine metal complexes with DNA

Professor John Kelly, Trinity College Dublin, Republic of Ireland

Abstract

For many years dipyrido[3,2-a:2’,3’-c]phenazine (dppz) complexes have been extensively studied as probes for DNA. 

[Ru(phen)2(dppz)]3+ (phen = 1,10-phenanthroline) has attracted particular attention as it acts as a DNA-light switch, being non-luminescent in water  but emitting when bound to DNA. By contrast [Ru(TAP)2(dppz)]2+ (TAP = 1,4,5,8-tetraazaphenanthrene) [2] and [Cr(phen)2(dppz)]3+ [3]  are luminescent in aqueous solution but not upon binding to DNA.  This is due to reduction of the metal complex excited state by guanine as has been demonstrated by picosecond transient spectroscopy (with both visible and IR monitoring) for [Ru(TAP)2(dppz)]2+ [2] and also for  [Re(CO)3(dppzF2)]2+ [4].

Although it is accepted that the dppz ligand intercalates between the DNA base-pair, the precise mode of binding deduced from spectroscopic and biophysical measurements has been a matter of debate.  Recent successful crystallisation by the Cardin group of both Ru(TAP)2(dppz)]2+  [5] and  [Ru(phen)2(dppz)]2+  [6] with small defined-sequence DNA has for the first time  provided a wealth of information on the interaction of these complexes with DNA and allowed a comparison with other DNA- intercalating molecules

References
[1] B M Zeglis et al, Chem Comm, 2007, 44, 4565-4579
[2] B Elias  et al, Chemistry – Eur J, 2008, 14, 369-375.
[3] M Wojdyla  et al,  Photochem. Photobiol. Sci, 2010, 9, 1196-1202.
[4] Q Cao et al, Photochem. Photobiol. Sci, 2011, 10, 1355-1364.
[5] J P Hall et al, Proc Natl Acad Sci,  2011, 108, 17610-17614
[6] H Niyazi et al, Nature Chemistry, 2012, in press.

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Metal photocleavage of DNA

Dr Samantha Higgins, Virginia Tech, USA

Abstract

Ru(II) and Os(II) polypyridyl chromophores have been readily studied for applications in DNA photomodification due to their enhanced photophysical and interesting photochemical properties. Coupling a Ru(II) or Os(II) chromophore to a remote bioactive site, such as Pt(II) or Rh(III), through a polyazine bridging ligand provides a supramolecular complex characterized by redox, spectroscopic, and photophysical properties. Studies of the mixed-metal complexes in the presence of DNA show enhanced photoinduced activity using low energy visible light within the therapeutic window (600-900 nm). The supramolecular complexes, [(Ph2phen)2MII(dpp)PtIICl2]Cl2 (Ph2phen = 4,7-diphenyl-1,10-phenanthroline and M = Ru(II) or Os(II)) display unique DNA photobinding and photocleavage properties by utilizing their lowest lying 3metal-to-ligand charge transfer (3MLCT) excited state to facilitate ligand loss at the remote Pt(II) center. Modifying the molecular architecture to [(TL)2MII(dpp)RhIIICl2(TL’)](PF6)3 (TL = terminal ligand on the chromophore, M = Ru(II) or Os(II) and TL’ = terminal ligand on the Rh(III) center) provides for distinctive oxygen independent photobinding and photocleavage features, a result attributed to unit population of the 3MLCT excited state, which is coupled to the lower lying 3metal-to-metal charge transfer (3MMCT) excited state. The varied mechanisms of DNA photocleavage in these supramolecular systems provide a fresh look at mixed-metal systems in photodynamic therapy.

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Multifunctional in cellulo probes

Dr James Thomas, University of Sheffield, UK

Abstract

Ruthenium(II) polypyridyl complexes that interact reversibly with DNA can display high binding affinities and selectivities [1,2] as well as interesting photophysical properties [3]. Although such complexes offer potential as in cellulo probes for luminescence microscopy, poor cellular uptake by live cells restricts the use of such molecules.

In recent studies we have shown that complexes such as 1 and 2 are taken up by both live eukaryotic and prokaryotic cells where they bind to nuclear DNA as evident by both luminescence and transmission electron microscopy[4] suggesting they may find use as cellular specific imaging agents and/or therapeutics [5].

A distinctive property of 2 - revealed by in vitro and in cellulo studies - is that high affinity binding of specific DNA structures leads to distinctive emission signatures [4,6].Herein we present progress in this area, including structural studies aimed at delineating the cause of this complex’s structural binding preferences [7].

Acknowledgments: We thank EPSRC for its Life science DTC and PoC fund.

References
[1] (a) C Metcalfe, J A Thomas, Chem Soc Rev2003, 32, 214. (b) M R Gill, J A Thomas, Chem Soc Rev, in press
[2] P Waywell, V Gonzalez, M R Gill, H A Adams, A J H M Meijer, M P Williamson, J A Thomas, Chem Eur J, 2010, 16, 2407.
[3] (a) S P Foxon, M Towrie, A W Parker, M Webb, J A Thomas, Angew Chem Int Ed,2007, 46, 3686-3689. (b) V G Gonzalez, T Wilson, I Kurihara, A Imai, J A Thomas, J Otsuki, Chem Commun 2008, 168.
[4] M R Gill, J Garcia-Lara, S J Foster, C G W Smythe, G Battaglia, J A Thomas, Nat Chem, 2009, 1,662-667.
[5] M R Gill, H Derat, C G W Smythe, G Battaglia, J A Thomas, ChemBioChem, 2011, 12, 877.
[6] T Wilson, M P Williamson, J A Thomas, Org Biomol Chem, 2010, 8, 2717.
[7] T Wilson, P J Costa, V Felix, M P Williamson, J A Thomas, in preparation.

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Targeted photodynamic therapy

Professor David Phillips CBE, Imperial College London, UK

Abstract

Photodynamic therapy, PDT, is a minimally invasive procedure used in treating a range of cancerous diseases, infections, and in ophthalmology to treat the wet form of age-related macular degeneration. The photodynamic action relies upon the simultaneous interaction between a sensitiser molecule [non-toxic in the absence of light], visible light, and molecular oxygen. The sensitiser, usually a porphryin –related molecule, excited initially to the singlet state, intersystem crosses to a longer-lived triplet state, which then produces reactive oxygen species , ROS, primarily singlet oxygen resulting from electronic energy transfer.

Current photodynamic therapy (PDT) of cancer is limited by inefficiencies involved in specifically targeting photosensitisers to tumours. Although antibodies are being explored as targeting vehicles, they present significant challenges, particularly in terms of pharmacokinetics and drug-coupling. We describe here a novel and effective system to attach covalently multiple photosensitiser molecules (both pre-clinical, pyropheophorbide-a and clinically approved, verteporfin photosensitisers) to single-chain Fvs.  Further, we demonstrate that not only do the resulting photoimmunoconjugates retain photophysical functionality, they are more potent than either free photosensitiser, effectively killing tumour cells in vitro and in vivo. For example, treatment of human breast cancer xenografts with a photo-immunoconjugate comprising an anti Her-2 scFv linked to 8 molecules of pyropheophorbide-a leads to complete tumour regression. These results give an insight into the important features that make scFvs good carriers for PDT drugs, and provide proof of concept of this unique approach to targeted photodynamic therapy (tPDT). This promises to significantly improve upon current photodynamic therapy for the treatment of cancer and paves the way for clinical application.

A second means of introducing selectivity in PDT is through spatial selection , using two-photon excitation of the sensitiser. This has the advantage of using red or near infra-red light, which greatly enhances tissue penetration. We describe the synthesis of porphyrin dimers with very large two-photon cross sections,carried out by Professor Harry Anderson’s group in Oxford,  and their effect in vitro on two cell types, and  in sealing blood vessels in vivo.

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Session 3: Phototherapy

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Professor Colin Hopper, University College London, UK

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Chair

Dr Nicola J Farrer, University of Warwick, UK

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Light-activated drug therapy

Dr Julie Woods, University of Dundee, UK

Abstract

The modality of light-activated chemotherapy is used in the treatment of non-melanoma skin cancer and some internal solid tumours; and also to treat hyperproliferative benign skin disease (eg. psoriasis). However, the range of photosensitisers available is limited, with most being based on a tetrapyrrole-like structure. There is a need for new photosensitisers, and with emerging developments in optical technology a rethink on how this modality can be applied to specific indications. Platinum (IV) prodrugs undergo photoreduction to the more reactive platinum (II) species (amongst other photoproducts) when irradiated. They rapidly kill tumour cells and their cisplatin-resistant derivatives in culture and are more effective than cisplatin tested under similar conditions. The trans- geometry is consistently more effective than the cis- configuration. Nuclear morphology studies and measurement of p53 stabilisation and caspase activity reveal important differences between the mechanism of action of these complexes and cisplatin. Finally, these prodrugs have no or limited toxicity or DNA reactivity in the dark, suggesting that toxicity to surrounding normal tissue could be limited. The complexes can be administered to mice at 10-fold higher doses than cisplatin with no obvious adverse effects. The photoactivated complexes described here are not simply acting as prodrugs of cisplatin. Harnessing the potent properties of platinum drugs selectively at the target site by light-activation could be beneficial for several indications including bladder and oesophageal cancer.

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Synergistic effect of nitric oxide and reactive oxygen species originated from light irradiation of nitrosyl ruthenium complex as potentiation of photodynamic therapy. Photobiological and cytotoxicity studies

Professor Roberto da Silva, University of Sao Paulo, Brazil

Abstract

Photodynamic therapy (PDT) is employed in the clinical treatment of neoplasic tissues and involves activation with light irradiation of a photosensitizing agent in the presence of molecular oxygen. Even though PDT has received increasing acceptance as a clinical approach the technique has been found to fail due low oxygen concentration in tumor region and  challenges concerning the design of effective photosensitizing drugs still have to be faced. In our work we have found that of nitric oxide release following singlet oxygen (1O2) production upon light irradiation may be an important mechanism by which the nitrosyl ruthenium complex exhibits enhanced biological activity in cells. The structural aspects, pharmacological assays, and in vitro photoinduced cytotoxic properties of nitrosyl ruthenium complexes are described. Its biological effect on the B16F10 and MCF-7 cell lines were studied in the presence and absence of visible light irradiation. At comparable irradiation levels nitrosyl ruthenium complexes were much more effective than a compound that produce only 1O2; indeed, enhanced potency was detected when the nitrosyl ruthenium complex was encapsulated in a drug delivery system. Flow cytometry analysis revealed that the photocytotoxic activity was mainly due to apoptosis.

Acknowledgments: 
CNPq, CAPES and FAPESP

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Session 4: Cellular probes, imaging and commercial development

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Dr Arno Wiehe, biolitec research GmbH, Germany

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Chair

Professor Dr Ulrich Schatzschneider, Julius-Maximilians-Universität, Germany

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Challenges during the development of photoactive metal complexes for PDT applications

Professor Dr Volker Albrecht, biolitec research GmbH, Germany

Abstract

Photodynamic therapy (PDT) is a minimal-invasive modality for the treatment of tumorous and other diseases. During PDT, a dye (photosensitizer) is administered that - after enrichment in the tumour - is irradiated with light of a specific wavelength. By interaction of the irradiated photosensitizer-molecule with molecular oxygen cell-toxic substances (especially highly reactive singlet oxygen) are formed which effectively damage the tumour. Numerous compounds have been investigated for their suitability for PDT. The most important among these are dyes from the chemical class of the tetrapyrroles (porphyrins, chlorins, phthalocyanines). Nearly all photosensitizers that are currently authorized for clinical applications are tetrapyrroles (Photofrin, Verteporfin, Temoporfin) or their biosynthetic precursors (ALA, i.e. d-aminolevulinic acid). It will be discussed what requirements metal complexes, in particular tetrapyrrole metal complexes, have to meet to be successfully used in PDT: eg photophysical parameters, factors influencing the uptake in tumour tissue, and pharmaceutical formulation.

Co-author:
Dr A Wiehe, biolitec research GmbH, Germany

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Exploitation of luminescent transition metal complexes as biomolecular and cellular probes

Professor Kenneth Kam-Wing Lo, City University of Hong Kong, Hong Kong, China

Abstract

Many transition metal polypyridine complexes exhibit intense and long-lived emission that is very sensitive to the local environments of the complexes.  This interesting property allows the complexes to serve as useful probes for biomolecules.  Thus, we have attached various reactive functional groups to luminescent transition metal complexes such as cyclometalated iridium(III) and tricarbonylrhenium(I) polypyridine complexes to yield new labels for biomolecules.  Additionally, we have modified these complexes with a range of biological substrates and artificial pendants including indole, estradiol, biotin, lipids, fluorous chains, dendritic skeletons, and poly(ethylene glycol), and utilized the complexes as luminescent biological probes.  Interestingly, these pendants not only perturb the photophysical behavior of the complexes, but also significantly affect their biomolecular recognition and cellular uptake properties.  In this presentation, the molecular design, photophysical properties, and biological properties of a selection of these iridium(III) and rhenium(I) polypyridine complexes will be described.  The cytotoxicity, cellular uptake, and intracellular distribution of these complexes will also be discussed.

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Time-resolved luminescent lanthanide bioprobes

Professor Jean-Claude Bünzli, EPFL, Switzerland and Korea University, Korea

Abstract

Ditopic hexadentate ligands with benzimidazole core self-assemble in water at physiological pH to yield robust and highly luminescent binuclear helicates [Ln2(LCX)3]. These entities are thermodynamically stable, kinetically inert and non-cytotoxic (IC50>500 μM). They enter into live cells by endocytosis and stain the endoplasmatic reticulum as shown by time-resolved luminescence microscopy [1]. The probes are also amenable to NIR excitation by multiphoton processes and can be inserted into nanoparticles [2].

Further derivatization affords helicates which can be bioconjugated to avidin and various monoclonal antibodies. When combined with microfluidic devices, the binuclear bioconjugates are highly efficient in multi detection of biomarkers expressed by cancerous cells and tissues [3].

[1] B Song, C D B Vandevyver, A-S Chauvin, J-C G Bünzli, Org Biomol Chem 2008, 6, 4125
[2] S V Eliseeva, B Song, C D B Vandevyver, A-S Chauvin, J B Wacker, J-C G Bünzli, New J Chem 2010, 34, 2915
[3] V Fernandez-Moreira, B Song, V Sivagnanam, A-S Chauvin, C D B Vandevyver, M A M Gijs, H-A Lehr, J-C G Bünzli, Analyst 2010, 135, 42

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Photoactivatable metal complexes: exciting potential in biotechnology and medicine? Kavli Royal Society Centre, Chicheley Hall Newport Pagnell Buckinghamshire MK16 9JJ