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Biologics are emerging as the most important class of drugs and are used to treat a large variety of pathologies. Most of biologics are proteins administered in large amounts, either by intramuscular injection or by intravenous infusion. Asparaginase is a large tetrameric protein assembly, currently used against acute lymphoblastic leukemia. Here, a gadolinium(III)-DOTA derivative has been conjugated to asparaginase, and its relaxation properties have been investigated to assess its efficiency as a possible theranostic agent. The field-dependent

The globally widespread perfluorooctanoic acid (PFOA) is a concerning environmental contaminant, with a possible toxic long-term effects on the environment and human health The development of sensible, rapid, and low-cost detection systems is a current change in modern environmental chemistry. In this context, two triamine-based chemosensors, L1 and L2, containing a fluorescent pyrene unit, and their Zn(II) complexes are proposed as fluorescent probes for the detection of PFOA in aqueous media. Binding studies carried out by means of fluorescence and NMR titrations highlight that protonated forms of the receptors can interact with the carboxylate group of PFOA, thanks to salt bridge formation with the ammonium groups of the aliphatic chain. This interaction induces a decrease in the fluorescence emission of pyrene at neutral and slightly acidic pH values. Similarly, emission quenching has also been observed upon coordination of PFOA by the Zn(II) complexes of the receptors. These results evidence that simple polyamine-based molecular receptors can be employed for the optical recognition of harmful pollutant molecules, such as PFOA, in aqueous media.

Pt-Based drugs play a very important role in current cancer treatments; yet, their cellular and mechanistic aspects are not fully understood. NMR metabolomics provides a powerful tool to investigate the metabolic perturbations induced by Pt drugs in cancer cells and decipher their meaning in relation to the presumed molecular mechanisms. We have carried out a systematic and comparative

Pd compounds form a promising class of experimental anticancer drug candidates whose mechanism of action is still largely unknown; in particular, a few organopalladium compounds seem very attractive. To gain mechanistic insight into medicinal palladium compounds, we have explored here - through ESI MS analysis - the interactions of four organopalladium agents (1-4) - showing remarkable

A novel gold(I) complex inspired by the known medicinal inorganic compounds auranofin and thimerosal, namely ethylthiosalicylate(triethylphosphine)gold(I) (AFETT hereafter), was synthesized and characterised and its structure was resolved through X-ray diffraction. The solution behavior of AFETT and its interactions with two biologically relevant proteins (

The reaction of the cytotoxic compound dirhodium tetraacetate with a B-DNA double helical dodecamer was studied by X-ray crystallography and mass spectrometry.

The synthesis of three new polyamine receptors based on a common 1,4,8,11-tetraazacyclododecane (cyclen) platform with appended, via a methylene linker, heteroaromatic fluorophores, a single quinoline (Q) or an 8-hydroxy-quinoline (8-OH-Q) moiety (L1 and HL2) or a Q and an 8-OH-Q unit (HL3), is reported. The proton binding features of these receptors, together with those of two similar receptors containing two equal Q moieties (L4) and two Q and two acetate groups (H2L5), have been studied by means of potentiometric, 1H NMR, UV-vis absorption and fluorescence emission measurements in aqueous solution at different pH values. While cyclen is a commonly used binding unit, Q and 8-OH-Q represent classical examples of signalling units in conjugated fluorescent chemosensors and their protonation features can strongly influence their ability in metal cations, and anion coordination and sensing. This study reveals that the structural features of these receptors are controlled by a subtle balance between hydrogen bonding and electrostatic repulsions occurring in their protonated species. At the same time, their emission properties are tuned by photoinduced electron and/or proton transfer processes taking place in the excited state.

NMR metabolomics represents a powerful tool to characterize the cellular effects of drugs and gain detailed insight into their mode of action. Here, we have exploited NMR metabolomics to illustrate the changes in the metabolic profile of A2780 ovarian cancer cells elicited by auranofin (AF), a clinically approved gold drug now repurposed as an anticancer agent. An early and large increase in intracellular glutathione is highlighted as the main effect of the treatment accompanied by small but significant changes in the levels of a few additional metabolites; the general implications of these findings are discussed in the frame of the current mechanistic knowledge of AF.

International audience; The reactions of the medicinal gold(i) compound auranofin and its close analogues with vasopressin and the diselenide analogue were comparatively investigated by LC-electrospray MS/MS. Evidence is gained of the possible cleavage of the S-S and Se-Se bridges induced by Au(i). Notably, we found that, in the absence of reducing agents, the sulfur and selenium atoms are metallated only at high temperature (70 °C) with the preferential binding of gold to selenium. The reaction with the S-S bridge can take place at physiological temperature (37 °C) under reducing conditions. The implications of these results are discussed in the general frame of the reactivity of biologically relevant soft Lewis acids with peptides and proteins.

Arsenoplatin-1 (AP-1) is a dual-action anticancer metallodrug with a promising pharmacological profile that features the simultaneous presence of a cisplatin-like center and an arsenite center. We investigated its interactions with proteins through a joint experimental and theoretical approach. The reactivity of AP-1 with a variety of proteins, including carbonic anhydrase (CA), superoxide dismutase (SOD), myoglobin (Mb), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and human serum albumin (HSA), was analyzed by means of electrospray ionization mass spectrometry (ESI MS) measurements. In accordance with previous observations, ESI MS experiments revealed that the obtained metallodrug-protein adducts originated from the binding of the [(AP-1)-Cl]

Purpose Ovarian cancer is the fifth leading cause of cancer-related deaths in women. Standard treatment consists of tumor debulking surgery followed by platinum and paclitaxel chemotherapy; yet, despite the initial response, about 70–75% of patients develop resistance to chemotherapy. Gold compounds represent a family of very promising anticancer drugs. Among them, we previously investigated the cytotoxic and pro-apoptotic properties of Au(NHC) and Au(NHC)2PF6, i.e., a monocarbene gold(I) complex and the corresponding bis(carbene) complex. Gold compounds are known to alter the redox state of cells interacting with free cysteine and selenocysteine residues of several proteins. Herein, a redox proteomic study has been carried out to elucidate the mechanisms of cytotoxicity in A2780 human ovarian cancer cells. Methods A biotinylated iodoacetamide labeling method coupled with mass spectrometry was used to identify oxidation-sensitive protein cysteines. Results Gold carbene complexes cause extensive oxidation of several cellular proteins; many affected proteins belong to two major functional classes: carbohydrate metabolism, and cytoskeleton organization/cell adhesion. Among the affected proteins, Glyceraldehyde-3-phosphate dehydrogenase inhibition was proved by enzymatic assays and by ESI–MS studies. We also found that Au(NHC)2PF6 inhibits mitochondrial respiration impairing complex I function. Concerning the oxidized cytoskeletal proteins, gold binding to the free cysteines of actin was demonstrated by ESI–MS analysis. Notably, both gold compounds affected cell migration and invasion. Conclusions In this study, we deepened the mode of action of Au(NHC) and Au(NHC)2PF6, identifying common cellular targets but confirming their different influence on the mitochondrial function.

Human cytoplasmic ferritins are heteropolymers of H and L subunits containing a catalytic ferroxidase center and a nucleation site for iron biomineralization, respectively. Here, ESI-MS successfully detected labile metal–protein interactions revealing the formation of tetra- and octa-iron clusters bound to L subunits, as previously underscored by X-ray crystallography.

Three novel platinum(II) complexes bearing N-heterocyclic ligands, i.e., Pt2c, Pt-IV and Pt-VIII, were previously prepared and characterized. They manifested promising in vitro anticancer properties associated with non-conventional modes of action. To gain further mechanistic insight, we have explored here the reactions of these Pt compounds with a few model proteins, i.e., hen egg white lysozyme (HEWL), bovine pancreatic ribonuclease (RNase A), horse heart cytochrome c (Cyt-c) and human serum albumin (HSA), primarily through ESI MS analysis. Characteristic and variegate patterns of reactivity were highlighted in the various cases that appear to depend both on the nature of the Pt complex and of the interacting protein. The protein-bound Pt fragments were identified. In the case of the complex Pt2c, the adducts formed upon reaction with HEWL and RNase A were further characterized by solving the respective crystal structures: this allowed us to determine the exact location of the various Pt binding sites. The implications of the obtained results are discussed in relation to the possible mechanisms of action of these innovative anticancer Pt complexes.

Au2phen ((2,9-dimethyl-1,10-phenanthroline)2Au2(µ-O)2)(PF6)2 and Auoxo6 ((6,6′-dimethyl-2,2′-bipyridine)2Au2(µ-O)2)(PF6)2 are two structurally related gold(III) complexes that were previously reported to display relevant and promising anticancer properties in vitro toward a large number of human cancer cell lines. To expand the knowledge on the molecular mechanisms through which these gold(III) complexes trigger apoptosis in cancer cells, further studies have been performed using A2780 ovarian cancer cells as reference models. For comparative purposes, parallel studies were carried out on the gold(III) complex AuL12 (dibromo(ethylsarcosinedithiocarbamate)gold(III)), whose proapoptotic profile had been earlier characterized in several cancer cell lines. Our results pointed out that all these gold(III) compounds manifest a significant degree of similarity in their cellular and proapoptotic effects; the main observed perturbations consist of potent thioredoxin reductase inhibition, disruption of the cell redox balance, impairment of the mitochondrial membrane potential, and induction of associated metabolic changes. In addition, evidence was gained of the remarkable contribution of ASK1 (apoptosis-signal-regulating kinase-1) and AKT pathways to gold(III)-induced apoptotic signaling. Overall, the observed effects may be traced back to gold(III) reduction and subsequent formation and release of gold(I) species that are able to bind and inhibit several enzymes responsible for the intracellular redox homeostasis, in particular the selenoenzyme thioredoxin reductase.

Auranofin (AF) and its iodido analog, i.e., Au(PEt3) I (AFI), were reported to exhibit very promising anticancer properties both in vitro and in vivo. However, both these gold compounds have a scarce aqueous solubility that hampers their pharmaceutical use. Here, we explore whether encapsulation of these metallodrugs inside hydroxypropyl-beta–cyclodextrin (HPβ–CD) may lead to an improved biopharmaceutical profile for the resulting adducts. Phase solubility studies, performed at 25 °C in an aqueous buffer, revealed, in both cases, the formation of a 1:1 drug to cyclodextrin complex; a far greater apparent stability constant (K1:1) was measured for AFI compared to AF (331 M−1 versus ca. 30 M−1). NMR studies conducted on the AFI/HPβ–CD system confirmed the formation of a stable 1:1 adduct. Then, binary systems of AF and AFI with HPβ–CD were prepared by colyophilization and characterized by DSC and PXRD. The results revealed the occurrence of drug complexation and/or amorphization for the AFI/HPβ–CD binary system. Afterwards, the antiproliferative properties of the two cyclodextrin adducts and of the corresponding free drugs were comparatively evaluated in vitro in three representative ovarian cancer cell lines, i.e., A2780, SKOV3, and IGROV-1. The results, in all cases, point out that CD complexation of the two gold drugs does not substantially affect their biological activity. The implications of these findings are discussed in the frame of the current knowledge of AF and its analogs.

Chloro(triethylphosphine)gold(I), (Et3PAuCl hereafter), is an Auranofin (AF)-related compound showing very similar biological and pharmacological properties. Like AF, Et3PAuCl exhibits potent antiproliferative properties in vitro toward a variety of cancer cell lines and is a promising anticancer drug candidate. We wondered whether Et3PAuCl encapsulation might lead to an improved pharmacological profile also considering the likely reduction of unwanted side-reactions that are responsible for adverse effects and for drug inactivation. Et3PAuCl was encapsulated in biocompatible PLGA–PEG nanoparticles (NPs) and the new formulation evaluated in colorectal HCT-116 cancer cells in comparison to the free gold complex. Notably, encapsulated Et3PAuCl (nano-Et3PAuCl hereafter) mostly retains the cellular properties of the free gold complex and elicits even greater cytotoxic effects in colorectal cancer (CRC) cells, mediated by apoptosis and autophagy. Moreover, a remarkable inhibition of two crucial signaling pathways, i.e. ERK and AKT, by nano-Et3PAuCl, was clearly documented. The implications of these findings are discussed.

A few gold compounds were recently found to show antimicrobial properties in vitro, holding great promise for the discovery of new drugs to overcome antibiotic resistance. Here, the inhibition of the bacterial virulence factor urease by four Au(I)-compounds, namely Au(PEt3)Cl, Au(PEt3)Br, Au(PEt3)I and [Au(PEt3)2]Cl, obtained from the antiarthritic Au(I)-drug Auranofin and earlier reported to act as antimicrobials, is investigated. The three monophosphino Au(I) complexes showed IC50 values in the 30–100 nM range, while the diphosphino Au(I) complex, though being less active, still showed a IC50 value of 7 μM. The structural basis for this inhibition was provided by solving the crystal structures of urease co-crystallized with Au(PEt3)I and [Au(PEt3)2]Cl: at least two Au(I) ions bind the enzyme in a flap domain involved in the catalysis, thus obliterating enzyme activity. Peculiar changes observed in the two structures reveal implications for the mechanism of soft metal binding and enzyme inactivation.

A group of triethylphosphine gold(I) and silver(I) complexes, structurally related to auranofin, were prepared and investigated as potential anticancer drug candidates. The antiproliferative properties of these metal compounds were assessed against two leukemia cell lines, i.e., CCRF-CEM and its multidrug-resistant counterpart, CEM/ADR5000. Interestingly, potent cytotoxic effects were disclosed for both series of compounds against leukemia cells, with IC50 values generally falling in the low-micromolar range, the gold derivatives being on the whole more effective than the silver analogues. Some initial structure-function relationships were drawn. Subsequently, the ability of the study compounds to inhibit the three main catalytic activities of the proteasome was investigated. Different patterns of enzyme inhibition emerged for the various metal complexes. Notably, gold compounds were able to inhibit effectively both the trypsin-like and chymotrypsin-like proteasome activities, being less effective toward the caspase-like catalytic activity. In most cases, a significant selectivity of the study compounds toward the proteasome proteolytic activities was detected when compared to other proteases. The implications of the obtained results are discussed.

Electrospray ionization mass spectrometry (ESI MS) is a powerful investigative tool to analyze the reactions of metallodrugs with proteins and peptides and characterize the resulting adducts. Here, we have applied this type of approach to four experimental anticancer gold(III) compounds for which extensive biological and mechanistic data had previously been gathered, namely, Auoxo6, Au2phen, AuL12, and Aubipyc. These gold(III) compounds were reacted with two representative proteins, i.e., human serum albumin (HSA) and human carbonic anhydrase I (hCA I), and with the C-terminal dodecapeptide of thioredoxin reductase. ESI MS analysis allowed us to elucidate the nature of the resulting metal-protein adducts from which the main features of the occurring metallodrug-protein reactions can be inferred. In selected cases, MS data were integrated and supported by independent 1HNMR and UV-Vis absorption measurements to gain an overall description of the occurring processes. From data analysis, it emerges that most of the investigated gold(III) complexes, endowed with an appreciable oxidizing character, undergo quite facile reduction to gold(I); the resulting gold(I) species tightly associate with the above proteins/peptides with a remarkable selectivity for free cysteine residues. In contrast, in the case of the less-oxidizing Aubipyc complex, the gold(III) oxidation state is conserved, and a gold(III) fragment still containing the original ligand is found to be associated with the target proteins. It is notable that the C-terminal dodecapeptide of thioredoxin reductase containing the characteristic -Gly-Cys-Sec-Gly metal-binding motif is able in all cases to trigger gold(III)-to-gold(I) reduction. Our investigation allowed us to identify in detail the nature of the gold fragments that ultimately bind the protein targets and determine the exact binding stoichiometry; some insight on the reaction kinetics was also gained. Notably, a few clear correlations could be established between the structure of the metal complexes and the nature of the resulting protein adducts. The mechanistic implications of these findings are analyzed and thoroughly discussed. Overall, the present results set the stage to better understand the real target biomolecules of these gold compounds and elucidate at the atomic level their interaction modes with proteins and peptides.

Gold compounds form an attractive class of cytotoxic metal compounds of potential application as anticancer agents. Notably, the mode of action of cytotoxic gold compounds appears to differ from that of the widely used anticancer Pt drugs -to which they were initially inspired- and to be basically DNA-independent. However, mechanistic details are still largely lacking for this class of metal-based drugs. To shed light on these issues we have developed a proteomic strategy that is capable of highlighting the perturbations in protein expression elicited by gold drugs in a selected cancer cell line with the final aim to disclose the underlying molecular mechanisms. In recent years, this type of strategy has been systematically applied, in our laboratory, to a representative panel of gold compounds including seven outstanding cytotoxic agents, i.e. six experimental gold(III) and gold(I) compounds and the clinical gold(I) drug, auranofin. A2780 human ovarian cancer cells were used as the standard cellular model for these studies. Proteins differentially expressed upon treatment were separated by 2-DE and identified by MALDI TOF and their meaning tentatively assessed through bioinformatic analysis. The occurrence of various and often overlapping molecular mechanisms was revealed. The affected proteins were found to belong -in most cases- to redox control systems and/or to the proteasome machinery implying that the severe cellular damage induced by gold compounds predominantly originates at these two distinct levels. However, for one Au(III) and one Au(I) compound, i.e. [(bipydmb-H)Au(OH)][PF6] (bipydmb-H = deprotonated 6-(1,1-dimethylbenzyl)-2,2′-bipyridine) (Aubipyc) and the bis(1-butyl-3-methyl-imidazole-2-ylidene) gold(I) [Au(NHC)2]PF6, a substantially greater number of proteomic alterations were detected pointing out, in both cases, to glucose metabolism as an additional target process of the cytotoxic action. The results that were obtained with the seven gold complexes are discussed in the frame of the available knowledge on anticancer gold drugs and their mechanisms. In general, our studies underscore the large amount of information that proteomic measurements may provide on the mode of action of metal-based drugs at the cellular level and delineate a very effective methodology for the identification of the respective cytotoxic mechanisms. We propose that the interpretation of the proteomic data in terms of the main affected cellular processes is further supported and validated through the implementation of complementary metabolomics and metallomics experiments.

// Ida Landini 1 , Andrea Lapucci 1 , Alessandro Pratesi 2 , Lara Massai 2 , Cristina Napoli 3 , Gabriele Perrone 1 , Pamela Pinzani 4 , Luigi Messori 2, * , Enrico Mini 1, * and Stefania Nobili 3, * 1 Department of Experimental and Clinical Medicine, University of Florence, Firenze, Italy 2 Department of Chemistry “Ugo Schiff”, University of Florence, Firenze, Italy 3 Department of Health Sciences, University of Florence, Firenze, Italy 4 Department of Experimental and Clinical Biomedical Sciences, University of Florence, Firenze, Italy * These authors have contributed equally to this work Correspondence to: Enrico Mini, email: enrico.mini@unifi.it Stefania Nobili, email: stefania.nobili@unifi.it Luigi Messori, email: luigi.messori@unifi.it Keywords: auranofin; tumour drug resistance; human tumour cell lines; drug effects; gene expression Received: March 24, 2017 Accepted: August 08, 2017 Published: October 09, 2017 ABSTRACT The anti-arthritic drug auranofin exerts also potent antitumour activity in in vitro and in vivo models, whose mechanisms are not yet well defined. From an auranofin-sensitive human ovarian cancer cell line A2780, a highly resistant (>20-fold) subline (A2780/AF-R) was developed and characterized. Marked reduction of gold accumulation occurred in auranofin-resistant A2780 cells. Also, moderately higher thioredoxin reductase activity in A2780/AF-R cells was observed while no changes in intracellular glutathione content occurred. Resistance to auranofin was associated with a low level of cross-resistance to some investigational gold compounds as well as to oxaliplatin and other anticancer drugs with different mode of action (i.e. melphalan, vinblastine, doxorubicin, etoposide, and paclitaxel). Reduced gold accumulation was associated to substantial gene expression changes in various influx (e.g. SLC22A1, SLC47A1, SLCO1B1 ) and efflux (e.g. ABCB1, ABCC2, ABCC3 ) transporters. The expression levels of selected proteins (i.e. SLC22A1, SLC47A1, P-gp) were also changed accordingly. These data provide evidence that multiple drug transporters may act as mediators of transport of auranofin and other gold compounds in cancer cells. Further investigation into the molecular mechanisms mediating transport of auranofin and new gold complexes in view of their potential clinical application in the treatment of cancer is warranted.

Na/K-ATPase is emerging as an important target for a variety of anticancer metal-based drugs. The interactions of Na/K-ATPase (in its E1 state) with three representative and structurally related cytotoxic gold(III) complexes, i.e. [Au(bipy)(OH)(2)][PF6], bipy = 2,2'-bipyridine; [Au(py(dmb)-H)(CH3COO)(2)], py(dmb)-H = deprotonated 6-(1,1-dimethylbenzyl)-pyridine and [Au(bipy(dmb)-H)(OH)][PF6], bipy(c)-H = deprotonated 6-(1,1-dimethylbenzyl)-2,20-bipyridine, are investigated here in depth using a variety of spectroscopic methods, in combination with docking studies. Detailed information is gained on the conformational and structural changes experienced by the enzyme upon binding of these gold(III) complexes. The quenching constants of intrinsic enzyme fluorescence, the fraction of Trp residues accessible to gold(III) complexes and the reaction stoichiometries were determined in various cases. Specific hypotheses are made concerning the binding mode of these gold(III) complexes to the enzyme and the likely binding sites. Differences in their binding behaviour toward Na/K-ATPase are explained on the ground of their distinctive structural features. The present results offer further support to the view that Na/K-ATPase may be a relevant biomolecular target for cytotoxic gold(III) compounds of medicinal interest and may thus be involved in their overall mode of action.

Citrate-capped gold nanoparticles (AuNPs) were functionalized with three distinct antitumor gold(III) complexes, e.g., [Au(N,N)(OH)2][PF6], where (N,N)=2,2&prime, bipyridine, [Au(C,N)(AcO)2], where (C,N)=deprotonated 6-(1,1-dimethylbenzyl)-pyridine, [Au(C,N,N)(OH)][PF6], where (C,N,N)=deprotonated 6-(1,1-dimethylbenzyl)-2,2&prime, bipyridine, to assess the chance of tracking their subcellular distribution by atomic force microscopy (AFM), and surface enhanced Raman spectroscopy (SERS) techniques. An extensive physicochemical characterization of the formed conjugates was, thus, carried out by applying a variety of methods (density functional theory&mdash, DFT, UV/Vis spectrophotometry, AFM, Raman spectroscopy, and SERS). The resulting gold(III) complexes/AuNPs conjugates turned out to be pretty stable. Interestingly, they exhibited a dramatically increased resonance intensity in the Raman spectra induced by AuNPs. For testing the use of the functionalized AuNPs for biosensing, their distribution in the nuclear, cytosolic, and membrane cell fractions obtained from human lymphocytes was investigated by AFM and SERS. The conjugates were detected in the membrane and nuclear cell fractions but not in the cytosol. The AFM method confirmed that conjugates induced changes in the morphology and nanostructure of the membrane and nuclear fractions. The obtained results point out that the conjugates formed between AuNPs and gold(III) complexes may be used as a tool for tracking metallodrug distribution in the different cell fractions.

Three pseudohalide analogues of the established gold drug auranofin (AF hereafter), of general formula Au(PEt3)X, i.e. Au(PEt3)CN, Au(PEt3)SCN and Au(PEt3)N3 (respectively denoted as AFCN, AFSCN and AFN3), were prepared and characterized. The crystal structure was solved for Au(PEt3)SCN highlighting the classical linear geometry of the 2-coordinate gold(I) center. The solution behaviour of the compounds was then comparatively analysed through 31PNMR providing evidence for an acceptable stability under physiological-like conditions. Afterward, the reaction of these gold compounds with bovine serum albumin (BSA) and consequent adduct formation was investigated by 31PNMR. For all the studied gold compounds, the [Au(PEt3)]+ moiety was identified as the reactive species in metal/protein adducts formation. The cytotoxic effects of the complexes were subsequently measured in comparison to AF against a representative colorectal cancer cell line and found to be still relevant and roughly similar in the three cases though far weaker than those of AF. These results show that the nature of the anionic ligand can modulate importantly the pharmacological action of the gold-triethylphosphine moiety, affecting the cytotoxic potency. These aspects may be further explored to improve the pharmacological profiles of this family of metal complexes.

The study of metal-based drugs represents an important branch of modern bioinorganic chemistry. The growing importance of this field is linked to the large success in medicine of a few metal based drugs, either in clinical use or still experimental. Moreover, these metal-based drugs are frequently used as reference compounds to assess comparatively the behavior of newly synthesized metallodrugs. For the convenience of researchers working in this area we report here a compilation of the relevant analytical and spectroscopic data of ten representative metallodrugs based on Platinum, Ruthenium, Gold and Mercury. The selected compounds, namely Cisplatin, Oxaliplatin, Carboplatin, Auranofin, Sodium Aurothiomalate, NAMI-A, KP1019, Thimerosal, Merbromin and Phenylmercury Acetate, were chosen owing to their importance in the field. We believe that this compilation may turn very helpful to researchers as these data are difficult to find and generally scattered over a large number of (old) publications.

[Image: see text] In recent years, a few successful attempts were made to repurpose the clinically approved antiarthritic gold drug, Auranofin (AF), as an anticancer agent. The present study shows that the iodido(triethylphosphine)gold(I) complex, (Et(3)PAuI hereafter)—an AF analogue where the thiosugar ligand is simply replaced by one iodide ligand—manifests a solution chemistry resembling that of AF and exerts similar cytotoxic and proapoptotic effects on A2780 human ovarian cancer cells in vitro. However, when evaluated in a preclinical orthotopic model of ovarian cancer, Et(3)PAuI produces a far superior anticancer action than AF inducing a nearly complete tumor remission. The highly promising in vivo performances here documented for Et(3)PAuI warrant its further evaluation as a drug candidate for ovarian cancer treatment.

The reactions with a few model proteins of two structurally related gold carbene compounds, namely the gold(I) monocarbene complex Au(NHC)Cl and the corresponding bis-carbene complex [Au(NHC)2]PF6 (where NHC is an N-heterocyclic carbene ligand), were comparatively studied by ESI MS measurements. The investigated proteins were: human serum albumin, human carbonic anhydrase and bovine superoxide dismutase; in addition, the reactions of the two gold carbenes with the C-terminal synthetic dodecapeptide of thioredoxin reductase were analyzed. Formation of metallodrug-protein adducts was observed in all cases made exception for the reactions of [Au(NHC)2]PF6 with carbonic anhydrase and superoxide dismutase. Notably, in line with expectations, the monocarbene gold complex turned out to be more effective than its dicarbene counterpart in forming protein adducts. The reactivity of these gold carbene complexes with model proteins is compared to that of a few other gold(III) and gold(I) complexes whose reactions with model proteins had been previously investigated with the same methodology; it emerges that the two gold carbenes react more selectively with proteins at well-defined anchoring sites. The implications of these results are discussed in the light of the current knowledge on medicinal gold compounds.

A micro-electrochemical reaction cell was coupled to an electrospray mass spectrometer in order to track redox transformations for two representative medicinal gold compounds - i.e. [(2,3,4,6-tetra-O-acetyl-1-thio-β-D-glucopyranosato-S)(triethylphosphine)gold(I)] and [Au(bipy

A series of palladium(II) (1-3) and platinum(II) chloride complexes (4 and 5) with 2,2':6',2″-terpyridine (terpy) derivatives substituted at the 4' position, was synthesized and fully characterized. Single crystal X-ray diffraction analysis of complexes 2, 3 and 5 showed tridentate coordination of the 4'-substituted terpyridine (terpy) ligands to the metal center. Moreover, in vitro cytotoxic activity of these complexes toward a panel of human cancer cell lines (lung cancer A549, colorectal cancer HCT116, ovarian cancer IGROV-1) and toward normal cell line HDF (dermal fibroblast) was determined by Trypan Blue exclusion assay. Overall, the tested compounds manifested a relevant cytotoxicity for the selected cancer cell lines with complex 4 also showing a modest cytotoxicity on the normal cell lines. To better understand the mode of action of these metal complexes, their reactivity with three model proteins, i.e. hen egg white lysozyme (HEWL), cytochrome c (cyt c) and ribonuclease A (RNase A) were comparatively investigated through ESI-MS analysis. The results highlighted a different behavior between the two series of complexes being platinum compounds more reactive toward RNase and cyt c than palladium compounds. Based on the obtained results, it is proposed that in presence of RNase A and cyt c, the platinum complexes undergo activation through release of labile ligands followed by binding to the protein. In contrast, palladium complexes revealed a far lower reactivity implying the likely occurrence of a different mechanism of action.

Aurothiomalate (AuTm) is an old, clinically established, antiarthritic gold drug that is currently being reconsidered as a candidate drug for cancer treatment and for other therapeutic indications within a more general drug repositioning program. As the biological effects of gold drugs seem to be mediated, mainly, by their interactions with protein targets we have analyzed here, in detail, the metalation patterns produced by aurothiomalate in a few model proteins. In particular, the reactions of aurothiomalate with the small proteins ribonuclease A, cytochrome c and lysozyme were explored through ESI MS (electrospray ionization mass spectrometry) analysis. Notably, characteristic and rather constant features emerged in the protein metalation patterns induced by AuTm that are markedly distinct from those caused by auranofin; a non-covalent interaction mode is invoked for AuTm binding to the mentioned proteins. The affinity constants of AuTm toward the three mentioned proteins were also initially assessed. The implications of the present findings are discussed.

Auranofin is a gold based drug in clinical use since 1985 for the treatment of rheumatoid arthritis. Beyond its antinflammatory properties, auranofin exhibits other attractive biological and pharmacological actions such as a potent in vitro cytotoxicity and relevant antimicrobial and antiparasitic effects that make it amenable for new therapeutic indications. For instance, auranofin is currently tested as an anticancer agent in four independent clinical trials; yet, its mode of action is highly controversial. With the present study, we explore the effects of auranofin in Saccharomyces cerevisiae and its likely mechanism. Notably, auranofin is reported to induce remarkable yeast growth inhibition. Solid evidence is provided that growth inhibition is the consequence of a direct cytotoxic insult occurring at the mitochondrial level; a profound depression of cell respiration is indeed clearly documented as the main cause of cell death while induction of ROS plays only a secondary role. More in detail, the mitochondrial NADH kinase Pos5 is identified as a primary target for auranofin. The implications of these results are discussed in the frame of current mechanistic knowledge on the cellular effects of auranofin and of its role as a prospective anticancer drug.

The interaction of the anticancer Ru(iii) complex AziRu - in comparison with its analogue NAMI-A, currently in advanced clinical trials as an antimetastatic agent - with DNA model systems, both single stranded and duplex oligonucleotides, was investigated using a combined approach, including absorption UV-vis spectroscopy, circular dichroism (CD) and electrospray mass spectrometry (ESI-MS) techniques. UV-vis absorption spectra of the Ru complexes were recorded at different times in a pseudo-physiological solution, to monitor the ligand exchange processes in the absence and in the presence of the examined oligonucleotides. CD experiments provided information on the overall conformational changes of the DNA model systems induced by these metal complexes. UV- and CD-monitored thermal denaturation studies were performed to analyse the effects of AziRu and NAMI-A on the stability of the duplex structures. ESI-MS experiments, carried out on the oligonucleotide/metal complex mixtures under investigation, allowed us to detect the formation of stable adducts between the guanine-containing oligomers and the ruthenium complexes. These data unambiguously demonstrate that both AziRu and NAMI-A can interact with the DNA model systems. Although very similar in their structures, the two metal compounds manifest a markedly different reactivity with the examined sequences, respectively, with either a naked Ru(3+) ion or a Ru(Im)(3+) (Im = imidazole) fragment being incorporated into the oligonucleotide structure via stable linkages.

International audience; Physicochemical methods have been used to investigate interactions occurring in solution between the dicarbene gold(I) complex [Au(9‐methylcaffein‐8‐ylidene)2]BF4 (AuNHC) and a human telomeric DNA sequence, namely Tel23. Circular dichroism measurements allow identification of the conformational changes experienced by Tel23 upon interaction with AuNHC, and the respective binding stoichiometries and constants were determined. Computational studies provide a good link between previous crystallographic results of the same system and the present solution data, offering an exhaustive description of the inherent noncovalent metallodrug–DNA interactions. Remarkably, we found that a preformed AuNHC/Tel23 adduct is capable of producing strong and selective inhibition of the enzyme telomerase. The latter feature is mechanistically relevant and might account for the conspicuous in vitro anticancer properties of the investigated dicarbene gold(I) complex.

// Francesca Magherini 1 , Tania Fiaschi 1 , Elisa Valocchia 1 , Matteo Becatti 1 , Alessandro Pratesi 2 , Tiziano Marzo 2, 3 , Lara Massai 2 , Chiara Gabbiani 3 , Ida Landini 4 , Stefania Nobili 5 , Enrico Mini 5 , Luigi Messori 2 , Alessandra Modesti 1 and Tania Gamberi 1 1 Department of Experimental and Clinical Biomedical Sciences “Mario Serio”, University of Florence, Florence, Italy 2 Department of Chemistry “Ugo Schiff”, University of Florence, Florence, Italy 3 Department of Chemistry and Industrial Chemistry, University of Pisa, Pisa, Italy 4 Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy 5 Department of Health Sciences, University of Florence, Florence, Italy Correspondence to: Tania Gamberi, email: tania.gamberi@unifi.it Luigi Messori, email: luigi.messori@unifi.it Keywords: gold(I)-N-heterocyclic carbene complexes; A2780 human ovarian cancer cells; proteomics; mitochondrial metabolism; thioredoxin reductase Received: August 13, 2017 Accepted: May 19, 2018 Published: June 15, 2018 ABSTRACT Au(NHC) and Au(NHC) 2 , i.e. a monocarbene gold(I) complex and the corresponding bis(carbene) complex, are two structurally related compounds, endowed with cytotoxic properties against several cancer cell lines. Herein, we explore the molecular and cellular mechanisms at the basis of their cytotoxicity in A2780 human ovarian cancer cells. Through a comparative proteomic analysis, we demonstrated that the number of modulated proteins is far larger in Au(NHC) 2 -treated than in Au(NHC)-treated A2780 cells. Both gold compounds mainly affected proteins belonging to the following functional classes: protein synthesis, metabolism, cytoskeleton and stress response and chaperones. Particularly, Au(NHC) 2 gave rise to an evident upregulation of several glycolytic enzymes. Moreover, only Au(NHC) 2 triggered a net impairment of respiration and a metabolic shift towards glycolysis, suggesting that mitochondria are relevant cellular targets. We also found that both carbenes, similarly to the gold(I) compound auranofin, caused a strong inhibition of the seleno-enzyme thioredoxin reductase (TrxR). In conclusion, we highlighted that coordination of two carbene ligands to the same gold(I) center greatly enhances the antiproliferative effects of the resulting compound in comparison to the monocarbene derivative. Moreover, TrxR inhibition and metabolic impairment seem to play a major role in the Au(NHC) 2 cytotoxicity. Overall, these antiproliferative effects were also confirmed on other two human ovarian cancer cell lines ( i.e. SKOV3 and IGROV1).

AuL10 [AuIIICl2DMDT] (DMDT: N,N-dimethyldithiocarbamate) and AuL12 [AuIIIBr2ESDT] (ESDT: ethylsarcosine dithiocarbamate) are two extremely promising gold-based anticancer compounds. Their mechanisms of action are still largely unknown although some specific hypotheses were put forward. To shed light on reactivity with probable biological targets we report here on their interactions with serum albuminand with calf thymus DNA taken as model biomolecules. Quite unexpectedly, spectrophotometric investigations revealed substantially different patterns of interaction for these two gold complexes with the mentioned biomolecules, probably arising from differences in redox chemistry. Afterwards, AuL12 was tested against A549 human non-small-cell lung carcinoma cells, evaluating its real-time profile of cell-growth inhibition by the xCELLigence Real-Time Cell Analysis (RTCA) system. Measures of the impedance at the base of each well by non-invasive gold microelectrodes point out that this gold complex inhibits cell proliferation after only 4 hours of treatment, producing its cytotoxic effects probably at the membrane level.

Gold compounds form a new class of promising metal-based drugs with a number of potential therapeutic applications, particularly in the fields of anticancer and antimicrobial treatments. Previous research revealed that a group of structurally diverse gold compounds cause conspicuous inhibition of the protease activities of the human proteasome. Given the pharmacological importance of protease inhibition, the present study further explored whether these gold compounds might inhibit a few other proteases that are accepted druggable targets for disease treatment. In particular, four distinct cysteine proteases were considered here: cathepsin B and L that play a primary role in tumor-cell invasion and metastasis; rhodesain, the major cathepsin L-like cysteine protease of Trypanosoma brucei rhodesiense and CPB2.8 Delta CTE, a Leishmania mexicana mature cysteine protease. Based on the encouraging results obtained for some of the tested gold compounds on the two parasitic cysteine proteases, especially against CPB2.8 Delta CTE, with IC50s in the micromolar range, we next evaluated whether those gold compounds might contrast effectively the growth of the respective protozoa and indeed important antiprotozoal properties were disclosed; on the other hand a certain lack of selectivity was highlighted. Also, no direct or clear correlation could be established between the in vitro antiprotozoal properties and the level of protease inhibition. The implications of these results are discussed in relation to possible pharmaceutical applications.

Aubipyc is an organogold(III) compound endowed with encouraging anti-proliferative properties in vitro that is being evaluated pre-clinically as a prospective anticancer agent. A classical proteomic approach is exploited here to elucidate the mechanisms of its biological actions in A2780 human ovarian cancer cells. Based on 2-D gel electrophoresis separation and subsequent mass spectrometry identification, a considerable number of differentially expressed proteins were highlighted in A2780 cancer cells treated with Aubipyc. Bioinformatic analysis of the groups of up-regulated and down-regulated proteins pointed out that Aubipyc primarily perturbs mitochondrial processes and the glycolytic pathway. Notably, some major alterations in the glycolytic pathway were validated through Western blot and metabolic investigations. Biological significance This is the first proteomic analysis regarding Aubipyc cytotoxicity in A2780/S ovarian cancer cell line. Aubipyc is a promising gold(III) compound which manifests an appreciable cytotoxicity toward the cell line A2780, being able to overcome resistance to platinum. The proteomic study revealed for Aubipyc different cellular alterations with respect to cisplatin as well as to other gold compound such as auranofin. Remarkably, the bioinformatic analysis of proteomic data pointed out that Aubipyc treatment affected, directly or indirectly, several glycolytic enzymes. These data suggest a new mechanism of action for this gold drug and might have an impact on the use of gold-based drug in cancer treatment.

The perspective of introducing plasmonic particles for applications in biomedical optics is receiving much interest. However, their translation into clinical practices is delayed by various factors, which include a poor definition of their biological interactions. Here, we describe the preparation and the biological profiles of gold nanorods belonging to five different size classes with average effective radii between similar to 5 and 20 nm and coated with polyethylene glycol (PEG). All these particles exhibit decent stability in the presence of representative proteins, low cytotoxicity and satisfactory compatibility with intravenous administration, in terms of their interference with blood tissue. However, the suspension begins to become unstable after a few days of exposure to blood proteins. Moreover, the cytotoxicity is a little worse for smaller particles, probably because their purification is more critical, while undesirable interactions with the mononuclear phagocyte system are minimal in the intermediate size range. Overall, these findings hold implications of practical relevance and suggest that PEGylated gold nanorods may be a versatile platform for a variety of biomedical applications.