Susan J. Berners-Price

Mitochondria-Targeted Chemotherapeutics: The Rational Design of Gold(I) N-Heterocyclic Carbene Complexes That Are Selectively Toxic to Cancer Cells and Target Protein Selenols in Preference to Thiols

A family of lipophilic, cationic Au(I) complexes of N-heterocyclic carbenes (NHCs) have been designed as new mitochondria-targeted antitumor agents that combine both selective mitochondrial accumulation and selective thioredoxin reductase inhibition properties within a single molecule. Two-step ligand exchange reactions with cysteine (Cys) and selenocysteine (Sec) occur with release of the NHC ligands. At physiological pH the rate constants for the reactions with Sec are 20- to 80-fold higher than those with Cys. The complexes are selectively toxic to two highly tumorigenic breast cancer cell lines and not to normal breast cells, and the degree of selectivity and potency are optimized by modification of the substituent on the simple imidazolium salt precursor. The lead compound is shown to accumulate in mitochondria of cancer cells, to cause cell death through a mitochondrial apoptotic pathway and to inhibit the activity of thioredoxin reductase (TrxR) but not the closely related and Se-free enzyme glutathione reductase.

Gold compounds as therapeutic agents for human diseases

The application of gold in medicine is traceable for several thousand years and Au(i) compounds have been used clinically to treat rheumatoid arthritis since the last century. Recently research into gold-based drugs for a range of human diseases has seen a renaissance. Old as well as new Au(i) and Au(iii) compounds have been used and designed with an aim of targeting cellular components that are implicated in the onset or progression of cancers, rheumatoid arthiritis, viral and parasitic diseases. In addition, new disease targets have been found for gold compounds that have given insight into the mechanism of action of these compounds, as well as in the molecular pathophysiology of human diseases. Here we discuss the rationale for the design and use of gold compounds that have specific and selective targets in cells to alleviate the symptoms of a range of human diseases. We summarise the most recent findings in this research and our own discoveries to show that gold compounds can be developed to become versatile and powerful drugs for diseases caused by dysfunction of selenol and thiol containing proteins.

Mechanisms of cytotoxicity and antitumor activity of gold(I) phosphine complexes: the possible role of mitochondria

Abstract Well known for their clinical anti-arthritic properties, gold-based drugs have also attracted interest as potential antitumor agents with gold(I) phosphine derivatives being among the most active in vivo. Auranofin, a linear tetraacetylthioglucose gold(I) phosphine complex, increased the life span of mice inoculated with P388 leukaemia, inhibited DNA polymerases and was preferentially cytotoxic to cells with altered mitochondria. Triethylphosphine gold(I) chloride inhibited tumor colony formation in vitro, reacted with DNA, and inhibited oxidative phosphorylation, ATP production and the viability of isolated rat hepatocytes. Bis[1,2-bis(diphenylphosphino)ethane]gold(I) chloride ([Au(dppe)2]Cl) had reproducible and significant antitumor activity in a number of murine tumor models in vivo. [Au(dppe)2]Cl also inhibited tumor colony formation in vitro, formed DNA strand beaks, induced DNA-protein cross links and had antimitochondrial effects on P388 leukemia cells and isolated hepatocytes. Tetrahedral Au(I)complexes of bidentate pyridyl phosphines have shown promising in vitro and in vivo antitumor properties that are determined by their drug lipophilicity. Although the exact intracellular targets responsible for their antitumor activity are unclear, gold(I) phosphines are directly cytotoxic and many appear to have antimitochondrial activity. Optimization of their hydrophilic–lipophilic balance may be key to improving their selectivity for tumor mitochondria versus oxidative phosphorylation pathways of normal cells.

Role of lipophilicity in determining cellular uptake and antitumour activity of gold phosphine complexes

Purpose: The lipophilic cation [Au(I)(dppe)2]+ [where dppe is 1,2-bis(diphenylphosphino)ethane] has previously demonstrated potent in vitro antitumour activity. We wished to determine the physicochemical basis for the cellular uptake of this drug, as well as of analogues including the 1:2 adducts of Au(I) with 1,2-bis(di-n-pyridylphosphino)ethane (dnpype; n=2, 3 and 4), and to compare in vitro and in vivo antitumour activity. Methods and results: Logarithmic IC50 values for the CH-1 cell line bore a parabolic dependence on drug lipophilicity, as measured either by high-performance liquid chromatography or by n-octanol-water partition. Cellular uptake of drug, as measured by inductively coupled plasma mass spectrometry, varied by over three orders of magnitude over the series. Logarithmic uptake had a parabolic dependence on drug lipophilicity but a linear relationship to logarithmic IC50 values. Free drug concentrations were determined under the culture conditions and logarithmic free drug IC50 values and uptake rates were linearly related to lipophilicity. Uptake of drug in vivo in tissue from murine colon 38 tumours was approximately proportional to the dose administered. Host toxicity varied according to lipophilicity with the most selective compound having an intermediate value. This compound was also the most active of those tested in vivo, giving a growth delay of 9 days following daily intraperitoneal dosing (10 days) at 4 μmol kg−1 day−1. It was also significantly more active than another lipophilic cation, MKT-077. Conclusions: Alteration of lipophilicity of aromatic cationic antitumour drugs greatly affects cellular uptake and binding to plasma proteins. Changes in lipophilicity also affect host toxicity, and optimal lipophilicity may be a critical factor in the design of analogues with high antitumour activity.

Coordination chemistry of metallodrugs: insights into biological speciation from NMR spectroscopy

Abstract There is much current interest in the design of metal compounds as drugs and diagnostic agents and in understanding the molecular mechanisms of action of metallopharmaceuticals already in clinical use. Central to progress in this area is investigation of the speciation of metal compounds, especially in biological media such as cells, body fluids and cell culture media. Modern multinuclear NMR approaches are very powerful for investigation of the thermodynamics and kinetics of reactions of metal compounds with both small and large biomolecules and it is possible to study the coordination chemistry of metallodrugs under physiologically relevant conditions. For example [ 1 H, 15 N] inverse detection methods allow studies of intermediates in the pathways of DNA platination by anticancer drugs and the direct detection of sulphur adducts of platinum drugs in urine. Other applications which are discussed include ligand exchange reactions of gold antiarthritic drugs, copper, silver, gold and ruthenium anticancer agents and bismuth antiulcer drugs. Resolution enhancement, specific isotopic labelling of amino acid side-chains and high field NMR studies of metal nuclei provide insight into the uptake and release of metallopharmaceuticals from the blood plasma proteins albumin (66 kDa) and transferrin (80 kDa). The use of 31 P cross-polarization magic angle spinning NMR spectroscopy to investigate the structures of bioactive metal phosphine complexes in the solid state is also described.

Dinuclear gold(I) complexes of bridging bidentate carbene ligands: synthesis, structure and spectroscopic characterisation

Eight dinuclear Au(i)-carbene complexes have been synthesized from various imidazolium-linked cyclophanes and related acyclic bis(imidazolium) salts, by treatment of the imidazolium salts with [Au(i)(SMe(2))Cl] in the presence of a carboxylate base. Single crystal structural studies showed that the Au(i)-carbene compounds contain dinuclear (AuL)(2) cations in which a pair of gold(i) centres are linked by a pair of bridging dicarbenoid ligands. Interestingly, the structural studies revealed short AuAu contacts of 3.0485(3)[Angstrom] and 3.5425(6)[Angstrom] in two of these complexes. NMR studies showed that the (AuL)(2) cations constructed from the cyclophane-based ligands retain a relatively rigid structure in solution, whilst those of the non-cyclophane ligand systems are fluxional in solution. The electronic absorption and emission spectra of the complexes in solution at room temperature were recorded and the complex with the shortest AuAu contact was found to emit intensely at 400 nm and more weakly at 780 nm upon excitation at 260 nm. The compounds with longer AuAu separations were not emissive under these conditions.

Structural and Solution Chemistry of gold(I) and Silver(I) complexes of bidentate pyridyl phosphines: selective antitumour agents

Abstract The 1:2 adducts of Ag(I) and Au(I) with 1,2-bis(di- n -pyridylphosphino)ethane (d n pype) for n =2, 3 and 4 have been synthesised and solution properties characterised by multinuclear NMR spectroscopy. The complexes are hydrophilic analogs of the lipophilic Au(I) antitumour complex [Au(dppe) 2 ] + and the degree of hydrophilicity depends critically on the position of the N atom in the pyridyl ring. The complexes of d3pype and d4pype are simple monomeric [M(d3pype) 2 ] + and [M(d4pype) 2 ] + species which have a much higher water solubility than the 2-pyridyl complexes which crystallise in the solid state as dimeric [{M(d2pype) 2 } 2 ] 2+ . In solution these 1:2 M:d2pype species exist as equilibrium mixtures of monomeric, dimeric and trimeric (Ag) or tetrameric (Au) clusters. The Au(I) and Ag(I)pyridyl phosphine complexes have been evaluated for antitumour activity against a panel of cultured human ovarian carcinoma cell lines. The results show both potent and selective activity for the compounds with IC 50 values ranging from 0.18 to 1500 μM. There is a correlation between the degree of antitumour selectivity and the octanol/water partition coefficients with the greatest selectivity (500-fold range) found for the most hydrophilic complex [Au(d4pype) 2 ]Cl. Clinical development of the parent compound [Au(dppe) 2 ] + was halted by liver toxicity and the hydrophilic pyridylphosphine analogs are significantly less toxic than [Au(dppe) 2 ] + when exposed to isolated rat hepatocytes. Convenient synthetic routes to the bidentate pyridyl phosphines d2pype, d3pype and d4pype are also described.

Activating Platinum Anticancer Complexes with Visible Light

oxaliplatin alone are expected to reach US

Recent advances in the application of 13C and 15N NMR spectroscopy to soil organic matter studies.

3 billion in total within the next 2 years. Despite this success, there are good reasons to attempt to improve their design. These existing platinum drugs are not active against all types of cancer, they can have adverse side effects, and resistance to therapy can develop. If relatively nontoxic platinum complexes could be designed that could be selectively activated in tumor cells, they might find widespread use in the clinic. The report by Sadler and co-workers of complexes with these characteristics is therefore notable. Cisplatin kills cells by binding to GG sequences in DNA, kinking the DNA, and triggering apoptosis. Carboplatin and oxaliplatin can form similar lesions, except that the latter has a 1,2-diaminocyclohexane ligand in place of two NH3 ligands. This different ligand arrangement distorts the DNA structure differently and affects recognition by mismatch-repair and damage-recognition proteins. The type of nitrogen ligand on Pt can therefore affect the activity of the complex. Platinum(IV) complexes are known to be less reactive and less toxic than Pt complexes, so Sadler and co-workers based their design on Pt prodrugs. To enable light activation, they incorporated azido ligands so as to introduce intense ligandto-metal charge-transfer absorption bands. Early attempts by Bednarski and co-workers to achieve this aim by using iodido ligands were promising but resulted in complexes which reacted too readily with glutathione (GSH), the abundant intracellular reducing agent, and so were not dark-stable. The cis-diazido Pt complex 1 is dark-stable and forms Pt–GG cross-links similar to cisplatin on DNA, but only when irradiated with light by a mechanism involving electron transfer from N3 to Pt and the formation of N2. If 1 is merely a prodrug for cisplatin, then the trans complex 2 might be expected to be inactive, as is transplatin itself. However, this is not the case. When irradiated with UVA light 2 is as active as cisplatin under conditions which might be used for photochemotherapy (short treatment and irradiation times of less than 1 h). Moreover, when the geometry is changed from cis to trans, the ligand-to-metal charge-transfer (LMCT) band is shifted to a longer wavelength. Shortwavelength UVA light (365 nm) penetrates tissues to a depth of approximately 1 mm and could be useful for surface cancers, such as bladder and oesophageal cancer, but visible light penetrates more deeply (e.g. green to 3 mm and red to 5 mm). In fact, although trans-diamine complexes were not thought to be active on account of the inactivity of transplatin, in the last few years many active trans complexes have been discovered by the research groups of Farrell, Natile, NavarroRanninger, Gibson, and others. Indeed, trans-[PtCl2((E)imino ether)2] complexes can be more active than their cis isomers, and trans-diamine complexes can have quite different properties to those of cis complexes. For example, complexes such as trans-[Pt(acetate)2(pyridine)2] are relatively inert towards hydrolysis and yet cytotoxic to cisplatinand oxaliplatin-resistant cancer cells. The potency of 2 as a photoactivated agent can be greatly increased by replacing one of the NH3 ligands with pyridine: complex 3 attacks DNA rapidly and causes lesions which are different from those caused by cisplatin, and more difficult to [*] Prof. Dr. S. J. Berners-Price Institute for Glycomics, Griffith University Gold Coast Campus, Qld 4222 (Australia) Fax: (+ 61)7-5552-8220 E-mail: s.berners-price@griffith.edu.au

Reactions of cisplatin hydrolytes with methionine, cysteine, and plasma ultrafiltrate studied by a combination of HPLC and NMR techniques.

Nuclear magnetic resonance (NMR) spectroscopy has been applied to many studies in soil science, geochemistry, and environmental science. In recent years, the study of soil organic matter (SOM) using NMR techniques has progressed rapidly. NMR spectroscopy has been used to study chemical changes of SOM during decomposition, and also of soil extract fractions such as humic acid and fulvic acid. NMR spectroscopy of soils has improved rapidly in recent years with the introduction of pre-treatment and particle-size fractionation. In addition to routine liquid- and solid-state 13C NMR applications, 15N NMR spectra of natural abundant samples have been reported, but 15N-enriched material is more convenient to use due to the low natural abundance of 15N. Some newly developed NMR techniques have also been utilised, such as 2-dimensional NMR spectroscopy and improved 1H NMR techniques. These are reviewed and commented on in this paper.