A Photodynamic and Photochemotherapeutic Platinum-Iridium Charge-Transfer Conjugate for Anticancer Therapy.

The novel hetero-dinuclear complex trans,trans,trans-[PtIV(py)2(N3)2(OH)(µ-OOCCH2CH2CONHCH2-bpyMe)IrIII(ppy)2]Cl (Pt-Ir), exhibits charge transfer between the acceptor photochemotherapeutic Pt(IV) (Pt-OH) and donor photodynamic Ir(III) (Ir-NH2) fragments. It is stable in the dark, but undergoes photodecomposition more rapidly than the Pt(IV) parent complex (Pt-OH) to generate Pt(II) species, an azidyl radical and 1O2. The Ir(III)* excited state, formed after irradiation, can oxidise NADH to NAD⋅ radicals and NAD+. Pt-Ir is highly photocytotoxic towards cancer cells with a high photocytotoxicity index upon irradiation with blue light (465 nm, 4.8 mW/cm2), even with short light-exposure times (10-60 min). In contrast, the mononuclear Pt-OH and Ir-NH2 subunits and their simple mixture are much less potent. Cellular Pt accumulation was higher for Pt-Ir compared to Pt-OH. Irradiation of Pt-Ir in cancer cells damages nuclei and releases chromosomes. Synchrotron-XRF revealed ca. 4× higher levels of intracellular platinum compared to iridium in Pt-Ir treated cells under dark conditions. Luminescent Pt-Ir distributes over the whole cell and generates ROS and 1O2 within 1 h of irradiation. Iridium localises strongly in small compartments, suggestive of complex cleavage and excretion via recycling vesicles (e.g. lysosomes). The combination of PDT and PACT motifs in one molecule, provides Pt-Ir with a novel strategy for multimodal phototherapy.

Tuning the photoactivated anticancer activity of Pt(iv) compounds via distant ferrocene conjugation.

Photoactive prodrugs offer potential for spatially-selective antitumour activity with minimal effects on normal tissues. Excited-state chemistry can induce novel effects on biochemical pathways and combat resistance to conventional drugs. Photoactive metal complexes in particular, have a rich and relatively unexplored photochemistry, especially an ability to undergo facile intersystem crossing and populate triplet states. We have conjugated the photoactive octahedral Pt(iv) complex trans, trans, trans-[Pt(N3)2(OH)2(py)2] to ferrocene to introduce novel features into a candidate photochemotherapeutic drug. The X-ray crystal structure of the conjugate Pt-Fe confirmed the axial coordination of a ferrocene carboxylate, with Pt(iv) and Fe(ii) 6.07 Å apart. The conjugation of ferrocene red-shifted the absorption spectrum and ferrocene behaves as a light antenna allowing charge transfer from iron to platinum, promoting the photoactivation of Pt-Fe with light of longer wavelength. Cancer cellular accumulation is enhanced, and generation of reactive species is catalysed after photoirradiation, introducing ferroptosis as a contribution towards the cell-death mechanism. TDDFT calculations were performed to shed light on the behaviour of Pt-Fe when it is irradiated. Intersystem spin-crossing allows the formation of triplet states centred on both metal atoms. The dissociative nature of triplet states confirms that they can be involved in ligand detachment due to irradiation. The Pt(ii) photoproducts mainly retain the trans-{Pt(py)2}2+fragment. Visible light irradiation gives rise to micromolar activity for Pt-Fe towards ovarian, lung, prostate and bladder cancer cells under both normoxia and hypoxia, and some photoproducts appear to retain Pt(iv)-Fe(ii) conjugation.

In Vivo Trafficking of the Anticancer Drug Tris(8-Quinolinolato) Gallium (III) (KP46) by Gallium-68/67 PET/SPECT Imaging.

KP46 (tris(hydroxyquinolinato)gallium(III)) is an experimental, orally administered anticancer drug. Its absorption, delivery to tumours, and mode of action are poorly understood. We aimed to gain insight into these issues using gallium-67 and gallium-68 as radiotracers with SPECT and PET imaging in mice. [67Ga]KP46 and [68Ga]KP46, compared with [68Ga]gallium acetate, were used for logP measurements, in vitro cell uptake studies in A375 melanoma cells, and in vivo imaging in mice bearing A375 tumour xenografts up to 48 h after intravenous (tracer level) and oral (tracer and bulk) administration. 68Ga was more efficiently accumulated in A375 cells in vitro when presented as [68Ga]KP46 than as [68Ga]gallium acetate, but the reverse was observed when intravenously administered in vivo. After oral administration of [68/67Ga]KP46, absorption of 68Ga and 67Ga from the GI tract and delivery to tumours were poor, with the majority excreted in faeces. By 48 h, low but measurable amounts were accumulated in tumours. The distribution in tissues of absorbed radiogallium and octanol extraction of tissues suggested trafficking as free gallium rather than as KP46. We conclude that KP46 likely acts as a slow releaser of gallium ions which are inefficiently absorbed from the GI tract and trafficked to tissues, including tumour and bone.

Radiometal-Labeled Photoactivatable Pt(IV) Anticancer Complex for Theranostic Phototherapy.

A novel photoactivatable Pt(IV) diazido anticancer agent, Pt-succ-DFO, bearing a pendant deferoxamine (DFO) siderophore for radiometal chelation, has been synthesized for the study of its in vivo behavior with radionuclide imaging. Pt-succ-DFO complexation of Fe(III) and Ga(III) ions yielded new heterobimetallic complexes that maintain the photoactivation properties and photocytotoxicity of the parent Pt complex in human cancer cell lines. Radiolabeled Pt-succ-DFO-68Ga (t1/2 = 68 min, positron emitter) was readily prepared under mild conditions and was stable in the dark upon incubation with human serum. PET imaging of Pt-succ-DFO-68Ga in healthy mice revealed a promising biodistribution profile with rapid renal excretion and limited organ accumulation, implying that little off-target uptake is expected for this class of agents. Overall, this research provides the first in vivo imaging study of the whole-body distribution of a photoactivatable Pt(IV) azido anticancer complex and illustrates the potential of radionuclide imaging as a tool for the preclinical development of novel light-activated agents.

Kinetics and mechanism of sequential ring methyl C-H activation in cyclopentadienyl rhodium(III) complexes.

We have studied activation of the methyl C-H bonds in the cyclopentadienyl ligands of half-sandwich Rh(III) complexes [η5-CpXRh(N,N')Cl]+ by observing the dependence of sequential H/D exchange on variations in CpX = Cp* (complexes 1 and 2), Me4PhCp (CpXPh, 3) or Me4PhPhCp (CpXPhPh, 4), and chelated ligand N,N' (bpy, 1; phen, 2-4). H/D exchange was fastest in d4-MeOD (t1/2 = 10 min, 37 °C, complex 1), no H/D exchange was observed in DMSO/D2O, and d4-MeOD enhanced the rate in CD3CN. The proposed Rh(I)-fulvene intermediate was trapped by [4 + 2] Diels-Alder reactions with conjugated dienes and characterized. The Rh(I) oxidation state was confirmed by X-ray photoelectron spectroscopy (XPS). Influence of solvent on the mechanisms of activation and Diels-Alder adduct formation was modelled using DFT calculations with the CAM-B3LYP functional and CEP-31 g basis set, and influence on the reaction profile of the dimiine ligand and phenyl substituent using the larger qzvp basis set. The Rh(III)-OH intemediate is stabilised by H-bonding with methanol and a Cp* CH3 hydrogen. The Rh(I)(Me4fulvene) species, stabilised by interaction of methanol with a coordinated water, again by two H-bonds H2O-HOMe (1.49 Å) and fulvene CH2 (1.94 Å), arises from synchronous transfer of the methanol OH proton to a Rh(III)-OH ligand and Cp* methyl hydrogen to the methanol oxygen. Additionally, the observed trend in catalytic activity for complexes 1-4 was reproduced by DFT calculations. These complexes form a novel class of catalytic molecular motors with a tunable rate of operation that can be stalled in a given state. They provide a basis for elucidation of the effects of ligand design on the contributions of electronic, rotational and vibrational energies to each step in the reaction pathway at the atomic level, consideration of which will enhance the design principles for the next generation of molecular machines.

Single-Cell Chemistry of Photoactivatable Platinum Anticancer Complexes.

The Pt(IV) prodrug trans, trans, trans-[Pt(pyridine)2(N3)2(OH)2] (Pt1) and its coumarin derivative trans, trans, trans-[Pt(pyridine)2(N3)2(OH)(coumarin-3-carboxylate)] (Pt2) are promising agents for photoactivated chemotherapy. These complexes are inert in the dark but release Pt(II) species and radicals upon visible light irradiation, resulting in photocytotoxicity toward cancer cells. Here, we have used synchrotron techniques to investigate the in-cell behavior of these prodrugs and visualize, for the first time, changes in cellular morphology and Pt localization upon treatment with and without light irradiation. We show that photoactivation of Pt2 induces remarkable cellular damage with extreme alterations to multiple cellular components, including formation of vacuoles, while also significantly increasing the cellular accumulation of Pt species compared to dark conditions. X-ray absorption near-edge structure (XANES) measurements in cells treated with Pt2 indicate only partial reduction of the prodrug upon irradiation, highlighting that phototoxicity in cancer cells may involve not only Pt(II) photoproducts but also photoexcited Pt(IV) species.

Facile protein conjugation of platinum for light-activated cytotoxic payload release.

The novel Pt(iv) complex trans,trans-[Pt(N3)2(Py)2(OH)(OCO-(PEG)2-NHCSNH-Ph-NCS)] (Pt4) conjugates to the side chain of lysine amino acids in proteins under mild conditions. Reaction with myoglobin generated a bioconjugate that was stable in the dark, but released a Pt(iv) prodrug upon visible light irradiation. A similar procedure was used to conjugate Pt4 to the antibody trastuzumab, resulting in the first photoactivatable Pt(iv)-antibody conjugate, demonstrating potential for highly selective cancer phototherapy.

Validation of the plasmid study to relate DNA damaging effects of radionuclides to those from external beam radiotherapy.

INTRODUCTION: The biological consequences of absorbed radiation doses are ill-defined for radiopharmaceuticals, unlike for external beam radiotherapy (EBRT). A reliable assay that assesses the biological consequences of any radionuclide is much needed. Here, we evaluated the cell-free plasmid DNA assay to determine the relative biological effects of radionuclides such as Auger electron-emitting [67Ga]GaCl3 or [111In]InCl3 compared to EBRT. METHODS: Supercoiled pBR322 plasmid DNA (1.25 or 5 ng/µL) was incubated with 0.5 or 1 MBq [67Ga]GaCl3 or [111In]InCl3 for up to 73 h or was exposed to EBRT (137Cs; 5 Gy/min; 0-40 Gy). The induction of relaxed and linear plasmid DNA, representing single and double strand breaks, respectively, was assessed by gel electrophoresis. Chelated forms of 67Ga were also investigated using DOTA and THP. Topological conversion rates for supercoiled-to-relaxed (ksrx) or relaxed-to-linear (krlx) DNA were obtained by fitting a kinetic model. RESULTS: DNA damage increased both with EBRT dose and incubation time for [67Ga]GaCl3 and [111In]InCl3. Damage caused by [67Ga]GaCl3 decreased when chelated. [67Ga]GaCl3 proved more damaging than [111In]InCl3; 1.25 ng/µL DNA incubated with 0.5 MBq [67Ga]GaCl3 for 2 h led to a 70% decrease of intact plasmid DNA as opposed to only a 19% decrease for [111In]InCl3. For both EBRT and radionuclides, conversion rates were slower for 5 ng/µL than 1.25 ng/µL plasmid DNA. DNA damage caused by 1 Gy EBRT was the equivalent to damage caused by 0.5 MBq unchelated [67Ga]GaCl3 and [111In]InCl3 after 2.05 ± 0.36 and 9.3 ± 0.77 h of incubation, respectively. CONCLUSIONS: This work has highlighted the power of the plasmid DNA assay for a rapid determination of the relative biological effects of radionuclides compared to external beam radiotherapy. It is envisaged this approach will enable the systematic assessment of imaging and therapeutic radionuclides, including Auger electron-emitters, to further inform radiopharmaceutical design and application.

Methods and techniques for in vitro subcellular localization of radiopharmaceuticals and radionuclides.

In oncology, the holy grail of radiotherapy is specific radiation dose deposition in tumours with minimal healthy tissue toxicity. If used appropriately, injectable, systemic radionuclide therapies could meet these criteria, even for treatment of micrometastases and single circulating tumour cells. The clinical use of α and ß- particle-emitting molecular radionuclide therapies is rising, however clinical translation of Auger electron-emitting radionuclides is hampered by uncertainty around their exact subcellular localisation, which in turn affects the accuracy of dosimetry. This review aims to discuss and compare the advantages and disadvantages of various subcellular localisation methods available to localise radiopharmaceuticals and radionuclides for in vitro investigations.

DNA-Intercalative Platinum Anticancer Complexes Photoactivated by Visible Light.

Photoactivatable agents offer the prospect of highly selective cancer therapy with low side effects and novel mechanisms of action that can combat current drug resistance. 1,8-Naphthalimides with their extended π system can behave as light-harvesting groups, fluorescent probes and DNA intercalators. We conjugated N-(carboxymethyl)-1,8-naphthalimide (gly-R-Nap) with an R substituent on the naphthyl group to photoactive diazido PtIV complexes to form t,t,t-[Pt(py)2 (N3 )2 (OH)(gly-R-Nap)], R=H (1), 3-NO2 (2) or 4-NMe2 (3). They show enhanced photo-oxidation, cellular accumulation and promising photo-cytotoxicity in human A2780 ovarian, A549 lung and PC3 prostate cancer cells with visible light activation, and low dark cytotoxicity. Complexes 1 and 2 exhibit pre-intercalation into DNA, resulting in enhanced photo-induced DNA crosslinking. Complex 3 has a red-shifted absorption band at 450 nm, allowing photoactivation and photo-cytotoxicity with green light.

Synthesis and in vivo evaluation of PEG-BP-BaYbF5 nanoparticles for computed tomography imaging and their toxicity.

Computed tomography (CT) is one of the most widespread imaging techniques in clinical use worldwide. CT contrast agents are administered to improve soft tissue contrast and highlight blood vessels. However, the range of CT contrast agents available in the clinic is limited and they suffer from short-circulation times and low k-edge values that result in the need for high doses for in vivo applications. Nanomaterials containing a mixture of electron-dense elements, such as BaYbF5 nanoparticles, have shown promise as more efficient CT contrast agents, but they require biocompatible coatings for biomedical applications. Here, we explore the use of a bifunctional PEG polymer (5 kDa) containing a terminal bisphosphonate (BP) anchor for efficient binding to the surface of BaYbF5 nanomaterials. The resulting PEG(5)-BP-BaYbF5 nanoparticles were synthesized and characterized using TEM, DLS, TGA, XRD and Z-potential measurements. Their in vitro stability was verified and their ability to produce CT contrast in a wide range of X-ray energies, covering preclinical and clinical scanners, was demonstrated. In vitro toxicity studies with PEG(5)-BP-BaYbF5 in the phagocytic pro-monocytic human cell line U937 did not identify toxic effects, even at high concentrations (30 mM). In vivo, PEG(5)-BP-BaYbF5 exhibited efficient CT contrast for angiography imaging, highlighting blood vessels and vascular organs, and long circulation times as expected from the PEG coating. However, at late time points (48 h), in vivo toxicity was observed. While the causes could not be completely elucidated, in vitro studies suggest that decomposition and release of Yb3+ and/or Ba2+ metal ions after decomposition of PEG(5)-BP-BaYbF5 may play a role. Overall, despite the promising CT contrast properties, our results suggest that BaYbF5 nanomaterials may suffer from significant long-term toxicities.

Manipulating the In Vivo Behaviour of 68Ga with Tris(Hydroxypyridinone) Chelators: Pretargeting and Blood Clearance.

Pretargeting is widely explored in immunoPET as a strategy to reduce radiation exposure of non-target organs and allow the use of short-lived radionuclides that would not otherwise be compatible with the slow pharmacokinetic profiles of antibodies. Here we investigate a pretargeting strategy based on gallium-68 and the chelator THPMe as a high-affinity pair capable of combining in vivo. After confirming the ability of THPMe to bind 68Ga in vivo at low concentrations, the bifunctional THPMe-NCS was conjugated to a humanised huA33 antibody targeting the A33 glycoprotein. Imaging experiments performed in nude mice bearing A33-positive SW1222 colorectal cancer xenografts compared pretargeting (100 µg of THPMe-NCS-huA33, followed after 24 h by 8-10 MBq of 68Ga3+) with both a directly labelled radioimmunoconjugate (89Zr-DFO-NCS-huA33, 88 µg, 7 MBq) and a 68Ga-only negative control (8-10 MBq of 68Ga3+). Imaging was performed 25 h after antibody administration (1 h after 68Ga3+ administration for negative control). No difference between pretargeting and the negative control was observed, suggesting that pretargeting via metal chelation is not feasible using this model. However, significant accumulation of "unchelated" 68Ga3+ in the tumour was found (12.9 %ID/g) even without prior administration of THPMe-NCS-huA33, though tumour-to-background contrast was impaired by residual activity in the blood. Therefore, the 68Ga-only experiment was repeated using THPMe (20 µg, 1 h after 68Ga3+ administration) to clear circulating 68Ga3+, producing a three-fold improvement of the tumour-to-blood activity concentration ratio. Although preliminary, these results highlight the potential of THPMe as a 68Ga clearing agent in imaging applications with gallium citrate.

Biotinylated photoactive Pt(iv) anticancer complexes.

Novel biotinylated diazido-Pt(iv) complexes exhibit high visible light photocytotoxicity while being stable in the dark. Photocytotoxicity and cellular accumulation of all-trans-[Pt(py)2(N3)2(biotin)(OH)] (2a) were enhanced significantly when bound to avidin; irradiation induced dramatic cellular morphological changes in human ovarian cancer cells treated with 2a.

In vitro cytotoxicity of Auger electron-emitting [67Ga]Ga-trastuzumab.

INTRODUCTION: Molecular radiotherapy exploiting short-range Auger electron-emitting radionuclides has potential for targeted cancer treatment and, in particular, is an attractive option for managing micrometastatic disease. Here, an approach using chelator-trastuzumab conjugates to target radioactivity to breast cancer cells was evaluated as a proof-of-concept to assess the suitability of 67Ga as a therapeutic radionuclide. METHODS: THP-trastuzumab and DOTA-trastuzumab were synthesised and radiolabelled with Auger electron-emitters 67Ga and 111In, respectively. Radiopharmaceuticals were tested for HER2-specific binding and internalisation, and their effects on viability (dye exclusion) and clonogenicity of HER2-positive HCC1954 and HER2-negative MDA-MB-231 cell lines was measured. Labelled cell populations were studied by microautoradiography. RESULTS: Labelling efficiencies for [67Ga]Ga-THP-trastuzumab and [111In]In-DOTA-trastuzumab were 90% and 98%, respectively, giving specific activities 0.52 ± 0.16 and 0.61 ± 0.11 MBq/µg (78-92 GBq/µmol). At 4 nM total antibody concentration and 200 × 103 cells/mL, [67Ga]Ga-THP-trastuzumab showed higher percentage of cell association (10.7 ± 1.3%) than [111In]In-DOTA-trastuzumab (6.2 ± 1.6%; p = 0.01). The proportion of bound activity that was internalised did not differ significantly for the two tracers (62.1 ± 1.4% and 60.8 ± 15.5%, respectively). At 100 nM, percentage cell binding of both radiopharmaceuticals was greatly reduced compared to 4 nM and did not differ significantly between the two (1.2 ± 1.0% [67Ga]Ga-THP-trastuzumab and 0.8 ± 0.9% for [111In]In-DOTA-trastuzumab). Viability and clonogenicity of HER2-positive cells decreased when each radionuclide was incorporated into cells by conjugation with trastuzumab, but not when the same level of radioactivity was confined to the medium by omitting the antibody conjugation, suggesting that 67Ga needs to be cell-bound or internalised for a therapeutic effect. Microautoradiography showed that radioactivity bound to individual cells varied considerably within the population. CONCLUSIONS: [67Ga]Ga-THP-trastuzumab reduced cell viability and clonogenicity only when cell-bound, suggesting 67Ga holds promise as a therapeutic radionuclide as part of a targeted radiopharmaceutical. The causes and consequences of non-homogeneous uptake among the cell population should be explored.

Metallodrugs are unique: opportunities and challenges of discovery and development.

Metals play vital roles in nutrients and medicines and provide chemical functionalities that are not accessible to purely organic compounds. At least 10 metals are essential for human life and about 46 other non-essential metals (including radionuclides) are also used in drug therapies and diagnostic agents. These include platinum drugs (in 50% of cancer chemotherapies), lithium (bipolar disorders), silver (antimicrobials), and bismuth (broad-spectrum antibiotics). While the quest for novel and better drugs is now as urgent as ever, drug discovery and development pipelines established for organic drugs and based on target identification and high-throughput screening of compound libraries are less effective when applied to metallodrugs. Metallodrugs are often prodrugs which undergo activation by ligand substitution or redox reactions, and are multi-targeting, all of which need to be considered when establishing structure-activity relationships. We focus on early-stage in vitro drug discovery, highlighting the challenges of evaluating anticancer, antimicrobial and antiviral metallo-pharmacophores in cultured cells, and identifying their targets. We highlight advances in the application of metal-specific techniques that can assist the preclinical development, including synchrotron X-ray spectro(micro)scopy, luminescence, and mass spectrometry-based methods, combined with proteomic and genomic (metallomic) approaches. A deeper understanding of the behavior of metals and metallodrugs in biological systems is not only key to the design of novel agents with unique mechanisms of action, but also to new understanding of clinically-established drugs.

Strategies for conjugating iridium(III) anticancer complexes to targeting peptides via copper-free click chemistry.

We report the synthesis and characterization of novel pentamethylcyclopentadienyl (Cp*) iridium(III) complexes [(Cp*)Ir(4-methyl-4'-carboxy-2,2'-bipyridine)Cl]PF6 (Ir-I), the product (Ir-II) from amide coupling of Ir-I to dibenzocyclooctyne-amine, and its conjugate (Ir-CP) with the cyclic nona-peptide c(CRWYDENAC). The familiar three-legged 'piano-stool' configuration for complex Ir-I was confirmed by its single crystal X-ray structure. Significantly, copper-free click strategy has been developed for site-specific conjugation of the parent complex Ir-I to the tumour targeting nona-cyclic peptide. The approach consisted of two steps: (i) the carboxylic acid group of the bipyridine ligand in complex Ir-I was first attached to an amine functionalized dibenzocyclooctyne group via amide formation to generate complex Ir-II; and (ii) the alkyne bond of dibenzocyclooctyne in complex Ir-II underwent a subsequent strain-promoted copper-free cycloaddition with the azide group of the modified peptide. Interestingly, while complex Ir-I was inactive towards A2780 human ovarian cancer cells, complex Ir-II exhibited moderate cytotoxic activity. Targeted complexes such as Ir-CP offer scope for enhanced activity and selectivity of this class of anticancer complexes.

Ligand-Controlled Reactivity and Cytotoxicity of Cyclometalated Rhodium(III) Complexes.

We report the synthesis, characterisation and cytotoxicity of six cyclometalated rhodium(III) complexes [CpXRh(C^N)Z]0/+, in which CpX = Cp*, Cpph, or Cpbiph, C^N = benzo[h]quinoline, and Z = chloride or pyridine. Three x-ray crystal structures showing the expected "piano-stool" configurations have been determined. The chlorido complexes hydrolysed faster in aqueous solution, also reacted preferentially with 9-ethyl guanine or glutathione compared to their pyridine analogues. The 1-biphenyl-2,3,4,5,-tetramethylcyclopentadienyl complex [CpbiphRh(benzo[h]quinoline)Cl] (3a) was the most efficient catalyst in coenzyme reduced nicotinamide adenine dinucleotide (NADH) oxidation to NAD+ and induced an elevated level of reactive oxygen species (ROS) in A549 human lung cancer cells. The pyridine complex [CpbiphRh(benzo[h]quinoline)py]+ (3b) was the most potent against A549 lung and A2780 ovarian cancer cell lines, being 5-fold more active than cisplatin towards A549 cells, and acted as a ROS scavenger. This work highlights a ligand-controlled strategy to modulate the reactivity and cytotoxicity of cyclometalated rhodium anticancer complexes.

Axial functionalisation of photoactive diazido platinum(iv) anticancer complexes.

Mono-axial functionalised octahedral diazido Pt(iv) complexes trans, trans, trans-[Pt(py)2(N3)2(OR1)(OR2)] (OR1 = OH and OR2 = anticancer agent coumarin-3 carboxylate (cou, 2a), pyruvate dehydrogenase kinase (PDK) inhibitors 4-phenylbutyrate (PhB, 2b) or dichloroacetate (DCA, 2c)), and their di-axial functionalised analogues with OR1 = DCA and OR2 = cou (3a), PhB (3b), or DCA (3c) have been synthesised and characterised, including the X-ray crystal structures of complexes 2a, 3a, 3b and 3c. These complexes exhibit dark stability and have the potential to generate cytotoxic Pt(ii) species and free radicals selectively in cancer cells when irradiated. Mono-functionalised complexes 2a-2c showed higher aqueous solubility and more negative reduction potentials. Mono- and di-functionalised complexes displayed higher photocytotoxicity with blue light (1 h, 465 nm, 4.8 mW cm-2) than the parent dihydroxido complex 1 (OR1 = OR2 = OH) in A2780 human ovarian (IC50 0.9-2.9 µM for 2a-2c; 0.11-0.39 µM for 3a-3c) and A549 human lung cancer cells (5.4-7.8 µM for 2a-2c; 1.2-2.6 µM for 3a-3c) with satisfactory dark stability. Notably, no apparent dark cytotoxicity was observed in healthy lung MRC-5 fibroblasts for all complexes (IC50 > 20 µM). Significantly higher platinum cellular accumulation and photo-generated ROS levels were observed for the di-functionalised complexes compared with their mono-functionalised analogues when cancer cells were treated under the same concentrations.

Induction of immunogenic cell death in cancer cells by a photoactivated platinum(IV) prodrug.

The platinum(IV) prodrug trans,trans,trans-[Pt(N3)2(OH)2(py)2] (1) is stable and non-toxic in the dark, but potently cytotoxic to cancer cells when irradiated by visible light, including cisplatin-resistant cells. On irradiation with visible light, it generates reactive Pt(II) species which can attack DNA, and produces reactive oxygen species (ROS) and reactive nitrogen species (RNS) which exert unusual effects on biochemical pathways. We now show that its novel mechanism of action includes induction of immunogenic cell death (ICD). Treatment of cancer cells with 1 followed by photoirradiation with visible light induces calreticulin (CRT) expression at the surface of dying cancer cells. This is accompanied by release of high mobility group protein-1B (HMGB1) and the secretion of ATP. Autophagy appears to play a key role in this chemotherapeutically-stimulated ICD. The observed uneven distribution of ecto-CRT promotes phagocytosis, confirmed by the observation of engulfment of photoirradiated CT26 colorectal cancer cells treated with 1 by J774.A1 macrophages. The photoactivatable prodrug 1 has a unique mechanism of action which distinguishes it from other platinum drugs due to its immunomodulating properties, which may enhance its anticancer efficacy.

New Designs for Phototherapeutic Transition Metal Complexes.

In this Minireview, we highlight recent advances in the design of transition metal complexes for photodynamic therapy (PDT) and photoactivated chemotherapy (PACT), and discuss the challenges and opportunities for the translation of such agents into clinical use. New designs for light-activated transition metal complexes offer photoactivatable prodrugs with novel targeted mechanisms of action. Light irradiation can provide spatial and temporal control of drug activation, increasing selectivity and reducing side-effects. The photophysical and photochemical properties of transition metal complexes can be controlled by the appropriate choice of the metal, its oxidation state, the number and types of ligands, and the coordination geometry.