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Hepatocellular carcinoma (HCC), the predominant type of liver cancer, is an aggressive malignancy with limited therapeutic options. In this study, we assess a collection of newly designed gold(I) phosphine complexes. Remarkably, the compound GC002 exhibits the greatest toxicity to HCC cells and outperforms established medications, such as sorafenib and auranofin, in terms of antitumor efficacy. GC002 triggers irreversible necroptosis in HCC cells by increasing the intracellular accumulation of reactive oxygen species (ROS). Mechanistically, GC002 significantly suppresses the activity of thioredoxin reductase (TrxR), which plays a crucial role in regulating redox homeostasis and is often overexpressed in HCC by binding directly to the enzyme. Our in vivo xenograft study confirms that GC002 possesses remarkable antitumor activity against HCC without severe side effects. These findings not only highlight the novel mechanism of controlling necroptosis via TrxR and ROS but also identify GC002 as a promising candidate for the further development of antitumor agents targeting HCC.

Objective: Cytotoxicity of the antirheumatic drug auranofin (Aur) and the non-steroidal anti-inflammatory drug meclofenamic acid (MA) on several cancer cell lines and isolated mitochondria was examined to assess whether these drugs behave as oxidative phosphorylation inhibitors., Methods: The effect of Aur or MA for 24 h was assayed on metastatic cancer and non-cancer cell proliferation, energy metabolism, mitophagy and metastasis; as well as on oxygen consumption rates of cancer and non-cancer mitochondria., Results: Aur doses in the low micromolar range were required to decrease proliferation of metastatic HeLa and MDA-MB-231 cells, whereas one or two orders of magnitude higher levels were required to affect proliferation of non-cancer cells. MA doses required to affect cancer cell growth were one order of magnitude higher than those of Aur. At the same doses, Aur impaired oxidative phosphorylation in isolated mitochondria and intact cells through mitophagy induction, as well as glycolysis. Consequently, cell migration and invasiveness were severely affected. The combination of Aur with very low cisplatin concentrations promoted that the effects on cellular functions were potentiated., Conclusion: Aur surges as a highly promising anticancer drug, suggesting that efforts to establish this drug in the clinical treatment protocols are warranted and worthy to undertake., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Rodríguez-Enríquez et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Introduction: Auranofin (AF) is a well-established, FDA-approved, antiarthritic gold drug that is currently being reevaluated for a variety of therapeutic indications through drug repurposing. AF has shown great promise as a potential anticancer agent and has been approved for a few clinical trials in cancer. The renewed interest in AF has led to extensive research into the design, preparation and biological evaluation of auranofin analogs, which may have an even better pharmacological profile than the parent drug., Areas Covered: This article reviews the strategies for chemical modification of the AF scaffold. Several auranofin analogs have been prepared and characterized for medical application in the field of cancer treatment over the last 20 years. Some emerging structure-function relationships are proposed and discussed., Expert Opinion: The chemical modification of the AF scaffold has been the subject of intense activity in recent years and this strategy has led to the preparation and evaluation of several AF analogs. The case of iodauranofin is a particularly promising example. The availability of homogeneous biological data for a group of AF derivatives allows some initial structure-function relationships to be proposed, which may inspire the design and synthesis of new and better AF analogs for cancer treatment.

Thioredoxin reductases are frequently overexpressed in various solid tumors as a protective mechanism against heightened oxidative stress. Inhibitors of this system, such as Auranofin, are effective in eradicating cancer cells. However, the clinical significance of thioredoxin reductase 1 (TrxR1) in lung cancer, as well as the potential for its antagonist as a treatment option, necessitated further experimental validation. In this study, we observed significant upregulation of TrxR1 specifically in non-small cell lung cancer (NSCLC), rather than small cell lung cancer. Moreover, TrxR1 expression exhibited associations with survival rate, tumor volume, and histological classification. We developed a novel TrxR1 inhibitor named LW-216 and assessed its antitumor efficacy in NSCLC. Our results revealed that LW-216 is effectively bound with intracellular TrxR1 at sites R371 and G442, facilitating TrxR1 ubiquitination and suppressing TrxR1 expression, while not affecting TrxR2 expression. Treatment of LW-216-induced DNA damage and cell apoptosis in NSCLC cells through the generation of reactive oxygen species (ROS). Importantly, supplementation with N-acetylcysteine (NAC) or ectopic TrxR1 expression reversed LW-216-induced apoptosis. Furthermore, LW-216 displayed potent tumor growth inhibition in NSCLC cell-implanted mice, reducing TrxR1 expression in xenografts. Remarkably, LW-216 exhibited superior antitumor activity compared to Auranofin in vivo. Collectively, our research provides compelling evidence supporting the potential of targeting TrxR1 by LW-216 as a promising therapeutic strategy for NSCLC., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Background: This study explores the repurposing of Auranofin (AF), an anti-rheumatic drug, for treating non-small cell lung cancer (NSCLC) adenocarcinoma and pancreatic ductal adenocarcinoma (PDAC). Drug repurposing in oncology offers a cost-effective and time-efficient approach to developing new cancer therapies. Our research focuses on evaluating AF's selective cytotoxicity against cancer cells, identifying RNAseq-based biomarkers to predict AF response, and finding the most effective co-therapeutic agents for combination with AF., Methods: Our investigation employed a comprehensive drug screening of AF in combination with eleven anticancer agents in cancerous PDAC and NSCLC patient-derived organoids (n = 7), and non-cancerous pulmonary organoids (n = 2). Additionally, we conducted RNA sequencing to identify potential biomarkers for AF sensitivity and experimented with various drug combinations to optimize AF's therapeutic efficacy., Results: The results revealed that AF demonstrates a preferential cytotoxic effect on NSCLC and PDAC cancer cells at clinically relevant concentrations below 1 µM, sparing normal epithelial cells. We identified Carbonic Anhydrase 12 (CA12) as a significant RNAseq-based biomarker, closely associated with the NF-κB survival signaling pathway, which is crucial in cancer cell response to oxidative stress. Our findings suggest that cancer cells with low CA12 expression are more susceptible to AF treatment. Furthermore, the combination of AF with the AKT inhibitor MK2206 was found to be particularly effective, exhibiting potent and selective cytotoxic synergy, especially in tumor organoid models classified as intermediate responders to AF, without adverse effects on healthy organoids., Conclusion: Our research offers valuable insights into the use of AF for treating NSCLC and PDAC. It highlights AF's cancer cell selectivity, establishes CA12 as a predictive biomarker for AF sensitivity, and underscores the enhanced efficacy of AF when combined with MK2206 and other therapeutics. These findings pave the way for further exploration of AF in cancer treatment, particularly in identifying patient populations most likely to benefit from its use and in optimizing combination therapies for improved patient outcomes., (© 2024. The Author(s).)

Gold compounds form a new class of promising anticancer agents with innovative modes of action. It is generally believed that anticancer gold compounds, at variance with clinically established platinum drugs, preferentially target proteins rather than nucleic acids. The reactions of several gold compounds with a few model proteins have been systematically explored in recent years through ESI MS measurements to reveal adduct formation and identify the main features of those reactions. Here, we focus our attention on a group of five gold compounds of remarkable medicinal interest, i.e., Auranofin, Au(NHC)Cl, [Au(NHC) 2 ]PF 6 , Aubipyc, and Auoxo6, and on their reactions with four different biomolecular targets, i.e., the proteins HEWL, hCA I, HSA and the C-terminal dodecapeptide of the enzyme thioredoxin reductase. Complete ESI MS data are available for those reactions due to previous experimental work conducted in our laboratory. From the comparative analysis of the ESI MS reaction profiles, some characteristic trends in the metallodrug-protein reactivity may be identified as detailed below. The main features are described and analyzed in this review. Overall, all these observations are broadly consistent with the concept that cytotoxic gold drugs preferentially target cancer cell proteins, with a remarkable selectivity for the cysteine and selenocysteine proteome. These interactions typically result in severe damage to cancer cell metabolism and profound alterations in the redox state, leading to eventual cancer cell death.

Auranofin, a gold(I)-based complex, is under clinical trials for application as an anticancer agent for the treatment of nonsmall-cell lung cancer and ovarian cancer. In the past years, different derivatives have been developed, modifying gold linear ligands in the search for new gold complexes endowed with a better pharmacological profile. Recently, a panel of four gold(I) complexes, inspired by the clinically established compound auranofin, was reported by our research group. As described, all compounds possess an [Au{P(OMe) 3 }] + cationic moiety, in which the triethylphosphine of the parent compound auranofin was replaced with an oxygen-rich trimethylphosphite ligand. The gold(I) linear coordination geometry was complemented by Cl - , Br - , I - , and the auranofin-like thioglucose tetraacetate ligand. As previously reported, despite their close similarity to auranofin, the panel compounds exhibited some peculiar and distinctive features, such as lower log P values which can induce relevant differences in the overall pharmacokinetic profiles. To get better insight into the P-Au strength and stability, an extensive study was carried out for relevant biological models, including three different vasopressin peptide analogues and cysteine, using 31 P NMR and LC-ESI-MS. A DFT computational study was also carried out for a better understanding of the theoretical fundamentals of the disclosed differences with regard to triethylphosphine parent compounds.

Purpose: Cisplatin resistance is the major obstacle in the clinical treatment of ovarian cancer patients. Molecular mechanisms of cisplatin resistance are multifaceted. Gold(I)-compounds, i.e. N-heterocyclic carbene-gold(I)-complexes (NHC-Au(I)) has been regarded as promising cytotoxic drug candidates. However, their potential to overcome cisplatin resistance has hardly been addressed yet. Here we investigated the activity of the gold(I) drug auranofin and the NHC-Au(I)-compound MC3 in W1CR and A2780cis cisplatin-resistant ovarian cancer cells., Methods: Cytotoxicity of auranofin and MC3 was detected by MTT assay, correlated with intracellular gold(I) content, analyzed by AAS, and with flow cytometric detection of the cell cycle. Insight into cellular redox balance was provided by fluorimetric ROS-formation assay and western blotting thioredoxin (Trx) and Nrf2. The role of ERK was elucidated by using the inhibitor SCH772984 and its impact on cytotoxicity upon co-treatment with cisplatin and Au(I)-compounds, respectively., Results: MC3 overcomes cisplatin resistance in A2780cis and W1CR, and auranofin in W1CR cells completely, which is neither reflected by intracellular gold levels nor cell cycle changes. Upregulated redox balance appears as a basis for resistance. W1CR cells possess higher Trx levels, whereas A2780cis cells display strong Nrf2 expression as anti-oxidative protection. Nevertheless, overcoming redox balance appears not primary mode of activity comparing cisplatin and gold(I)-compounds. pERK emerges as a critical component and thus a promising target for overcoming resistance, regulating apoptosis differently in response to either gold(I) or cisplatin in A2780 cells., Conclusion: These data reflect the complexity of cisplatin resistance in cell models and emphasize NHC-Au(I)-complexes as prospective cytotoxic agents for further investigations in that respect., (© 2023. The Author(s).)

A panel of four novel gold(I) complexes, inspired by the clinically established gold drug auranofin (1-Thio-β-D-glucopyranosatotriethylphosphine gold-2,3,4,6-tetraacetate), was prepared and characterized. All these compounds feature the replacement of the triethylphosphine ligand of the parent compound auranofin with a trimethylphosphite ligand. The linear coordination around the gold(I) center is completed by Cl - , Br - , I - or by the thioglucose tetraacetate ligand (SAtg). The in-solution behavior of these gold compounds as well as their interactions with some representative model proteins were comparatively analyzed through 31 PNMR and ESI-MS measurements. Notably, all panel compounds turned out to be stable in aqueous media, but significant differences with respect to auranofin were disclosed in their interactions with a few leading proteins. In addition, the cytotoxic effects produced by the panel compounds toward A2780, A2780R and SKOV-3 ovarian cancer cells were quantitated and found to be in the low micromolar range, since the IC 50 of all compounds was found to be between 1 μM and 10 μM. Notably, these novel gold complexes showed large and similar inhibition capabilities towards the key enzyme thioredoxin reductase, again comparable to those of auranofin. The implications of these results for the discovery of new and effective gold-based anticancer agents are discussed.

A series of NHC-based selenourea Ag(I) and Au(I) complexes were evaluated for their anticancer potential in vitro, on 2D and 3D human cancer cell systems. All NHC-based selenourea complexes possess an outstanding cytotoxic potency, which was comparable or even better than that of the reference metallodrug auranofin, and were also able to overcome both platinum-based and multi-drug resistances. Intriguingly, their cytotoxic potency did not correlate with solution stability, partition coefficient or cellular uptake. On the other hand, mechanistic studies in cancer cells revealed their ability to strongly and selectively inhibit the redox-regulating enzyme Thioredoxin Reductase (TrxR), being even more effective than auranofin, a well-known TrxR inhibitor, without affecting other redox enzymes such as Glutathione Reductase (GR). The inhibition of TrxR in H157 human cancer cells caused, in turn, the disruption of cellular thiol-redox homeostasis and of mitochondria pathophysiology, ultimately leading to cancer cell death through apoptosis., (© 2022 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

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 ( i.e. human serum albumin and haemoglobin) and with a synthetic dodecapeptide reproducing the C-terminal portion of thioredoxin reductase were comparatively analyzed through 31 P NMR and ESI-MS. Remarkable binding properties toward these biomolecules were disclosed. Moreover, the cytotoxic effects produced by AFETT on two ovarian cancer cell lines (A2780 and A2780 R) and one colorectal cancer cell line (HCT116) were analyzed and found to be strong and nearly superimposable to those of auranofin. Interestingly, for both compounds, the ability to induce downregulation of vimentin expression in A2780 R cells was evidenced. Despite its close similarity to auranofin, AFETT is reported to exhibit some peculiar and distinctive features such as a lower lipophilicity, an increased water solubility and a faster reactivity towards the selected target biomolecules. These differences might confer to AFETT significant pharmaceutical and therapeutic advantages over auranofin itself.

Auranofin, as a thioredoxin reductase (TrxR) inhibitor, has promising anti-cancer activity in several cancer types. However, little is known about the inhibitory effect of auranofin on lung cancer cell growth. We, therefore, investigated the antigrowth effects of auranofin in various lung cancer cells with respect to cell death, reactive oxygen species (ROS), and glutathione (GSH) levels. Treatment with 0~5 µM auranofin decreased cell proliferation and induced cell death in Calu-6, A549, SK-LU-1, NCI-H460, and NCI-H1299 lung cancer cells at 24 h. In addition, 0~5 µM auranofin increased ROS levels, including O 2 •- , and depleted GSH levels in these cells. N-acetyl cysteine (NAC) prevented growth inhibition and mitochondrial membrane potential (MMP, ∆Ψm) loss in 3 and 5 µM auranofin-treated Calu-6 and A549 cells at 24 h, respectively, and decreased ROS levels and GSH depletion in these cells. In contrast, L-buthionine sulfoximine (BSO) enhanced cell death, MMP (∆Ψm) loss, ROS levels, and GSH depletion in auranofin-treated Calu-6 and A549 cells. Treatment with 3 and 5 µM auranofin induced caspase-3 activation and poly (ADP ribose) polymerase (PARP) cleavage in Calu-6 and A549 cells, respectively. Both were prevented by NAC, but enhanced by BSO. Moreover, TrxR activity was reduced in auranofin-treated Calu-6 and A549 cells. That activity was decreased by BSO, but increased by NAC. In conclusion, these findings demonstrate that auranofin-induced cell death is closely related to oxidative stress resulted from increased ROS levels and GSH depletion in lung cancer cells.

The search for highly selective sensitizers with a novel mechanism for tumor targeting therapy is of considerable interest. In this work, we have developed a series of new biotin-targeted Au(I) complexes. Through systematic biological evaluation and comparison, biotinylated Au(I) complex 3a containing a triphenylphosphine ligand was screened, as it realized both prominent efficient inhibition and selective cytotoxicity to cancer cells, and the effect was better than that of popularly used auranofin. Meanwhile, complex 3a , as a potent radiosensitizer, enhances anticancer effects in vitro and in vivo and has sensitization selectivity. From the action mechanism study, we provide evidence that complex 3a could intervene in redox homeostasis through targeted binding and strong suppression of thioredoxin reductase (TrxR) and induce the ferroptosis death process, enabling it to sensitize tumor cells to radiotherapy. Thus, complex 3a has enormous potential as an efficient and specific radiosensitizing agent in cancer therapy.

Purpose: Multiple myeloma (MM) remains an incurable hematologic malignancy which ultimately develops drug resistance and evades treatment. Despite substantial therapeutic advances over the past years, the clinical failure rate of preclinically promising anti-MM drugs remains substantial. More realistic in vitro models are thus required to better predict clinical efficacy of a preclinically active compound., Methods: Here, we report on the establishment of a conical agarose 3D co-culture platform for the preclinical propagation of primary MM cells ex vivo. Cell growth was compared to yet established 2D and liquid overlay systems. MM cell lines (MMCL: RPMI-8226, U266, OPM-2) and primary patient specimens were tested. Drug sensitivity was examined by exploring the cytotoxic effect of bortezomib and the deubiquitinase inhibitor auranofin under various conditions., Results: In contrast to 2D and liquid overlay, cell proliferation in the 3D array followed a sigmoidal curve characterized by an initial growth delay but more durable proliferation of MMCL over 12 days of culture. Primary MM specimens did not expand in ex vivo monoculture, but required co-culture support by a human stromal cell line (HS-5, MSP-1). HS-5 induced a > fivefold increase in cluster volume and maintained long-term viability of primary MM cells for up to 21 days. Bortezomib and auranofin induced less cytotoxicity under 3D vs. 2D condition and in co- vs. monoculture, respectively., Conclusions: This study introduces a novel model that is capable of long-term propagation and drug testing of primary MM specimens ex vivo overcoming some of the pitfalls of currently available in vitro models., (© 2022. The Author(s).)

Auranofin is an oral gold(I) compound, initially developed for the treatment of rheumatoid arthritis. Currently, Auranofin is under investigation for oncological application within a drug repurposing plan due to the relevant antineoplastic activity observed both in vitro and in vivo tumor models. In this review, we analysed studies in which Auranofin was used as a single drug or in combination with other molecules to enhance their anticancer activity or to overcome chemoresistance. The analysis of different targets/pathways affected by this drug in different cancer types has allowed us to highlight several interesting targets and effects of Auranofin besides the already well-known inhibition of thioredoxin reductase. Among these targets, inhibitory-κB kinase, deubiquitinates, protein kinase C iota have been frequently suggested. To rationalize the effects of Auranofin by a system biology-like approach, we exploited transcriptomic data obtained from a wide range of cell models, extrapolating the data deposited in the Connectivity Maps website and we attempted to provide a general conclusion and discussed the major points that need further investigation., (© 2021 The Authors. Medicinal Research Reviews published by Wiley Periodicals LLC.)

In this study, we developed a strategy to determine atto- and femtomolar amounts of metal ions in lysates and mineralizates of cells (human non-small-cell lung carcinoma (NSCLC, A549) and normal lung (MRC-5)) exposed to cytotoxic metallo-drugs: cisplatin and auranofin at concentrations close to the half-maximal inhibitory drug concentrations (IC 50 ). The developed strategy combines data obtained using biological and chemical approaches. Cell density was determined using two independent cell staining assays using trypan blue, calcein AM/propidium iodide. Metal concentrations in lysed and mineralized cells were established employing a mass spectrometer with inductively coupled plasma (ICP-MS) and equipped with a cross-flow nebulizer working in aspiration mode. It allowed for detecting of less than 1 fg of metal per cell. To decrease the required amount of sample material (from 1.5 mL to ~100 µL) without loss of sensitivity, the sample was introduced as a narrow band into a constant stream of liquid (flow-injection analysis). It was noticed that the selectivity of cisplatin accumulation by cells depends on the incubation time. This complex is accumulated by cells at a lower efficiency than auranofin and is found primarily in the lysate representing the cytosol. In contrast, auranofin interacts with water-insoluble compounds. Despite their different mechanism of action, both metallo-drugs increased the accumulation of transition metal ions responsible for oxidative stress.

Cyclic (Alkyl)(Amino)Carbenes (CAACs) have become forceful ligands for gold due to their ability to form very strong ligand-metal bonds. Inspired by the success of Auranofin and other gold complexes as antitumor agents, we have studied the cytotoxicity of bis- and mono-CAAC-gold complexes on different cancer cell lines: HeLa (cervical cancer), A549 (lung cancer), HT1080 (fibrosarcoma) and Caov-3 (ovarian cancer). Further investigations aimed at elucidating their mechanism of action are described. This includes quantification of affinities for TrxR, evaluation of their bioavailability and determination of associated cell death process. Moreover, Transmission Electron Microscopy (TEM) was used to study morphological changes upon exposure. Noticeably, a significant reduction in non-specific binding to serum proteins was observed with CAAC complexes when compared to Auranofin. These results confirm the potential of CAAC-gold complexes in biological environments, which may result in more specific drug-target interactions and decreased side effects., (© 2020 Wiley-VCH GmbH.)

Auranofin is a gold complex used as an anti-rheumatic agent and may act as a potent anticancer drug against breast tumors. Trametinib is a specific mitogen-activated protein kinase inhibitor, approved for the treatment of metastatic melanoma. The aim of this study was to examine the synergistic effects of auranofin and trametinib on apoptosis in MCF-7 human breast cancer cells. The combination treatment inhibited cancer cell proliferation and induced cell cycle arrest at the sub-G1 phase and apoptosis via poly (ADP-ribose) polymerase cleavage and caspase-3/7 activation. It is noteworthy that this treatment significantly increased p38 mitogen-activated protein kinase (MAPK) phosphorylation to induce mitochondrial stress, subsequently promoting cancer cell apoptosis through release of apoptosis-inducing factor. Further data demonstrated that combined treatment significantly induced increase in nuclear translocation of AIF. These results indicated that activation of the p38 MAPK signaling pathway and mitochondrial apoptosis may contribute to the synergistic consequences in MCF-7 cells. Collectively, our data demonstrated that combined treatment with auranofin and trametinib exhibited synergistic breast cancer cell death and this combination might be utilized as a novel therapeutic strategy for breast cancer.

Gold compounds have been employed throughout history to treat various types of disease, from ancient times to the present day. In the year 1985, auranofin, a gold-containing compound, was approved by U.S. Food and Drug Administration (FDA) as a therapeutic agent to target rheumatoid arthritis that would facilitate easy oral drug administration as opposed to conventional intramuscular injection used in treatments. Furthermore, auranofin demonstrates promising results for the treatment of various diseases beyond rheumatoid arthritis, including cancer, neurodegenerative diseases, acquired immune deficiency syndrome, and bacterial and parasitic infections. Various potential novel applications for auranofin have been proposed for treating human diseases. Auranofin has previously been demonstrated to inhibit thioredoxin reductase (TrxR) involved within the thioredoxin (Trx) system that comprises one of the critical cellular redox systems within the body. TrxR comprises the sole known enzyme that catalyzes Trx reduction. With cancers in particular, TrxR inhibition facilitates an increase in cellular oxidative stress and suppresses tumor growth. In this review, we describe the potential of auranofin to serve as an anticancer agent and further drug repurposing to utilize this as a strategy for further appropriate drug developments.

Neglected parasitic diseases remain a major public health issue worldwide, especially in tropical and subtropical areas. Human parasite diversity is very large, ranging from protozoa to worms. In most cases, more effective and new drugs are urgently needed. Previous studies indicated that the gold(I) drug auranofin (Ridaura ® ) is effective against several parasites. Among new gold(I) complexes, the phosphole-containing gold(I) complex {1-phenyl-2,5-di(2-pyridyl)phosphole}AuCl (abbreviated as GoPI) is an irreversible inhibitor of both purified human glutathione and thioredoxin reductases. GoPI-sugar is a novel 1-thio-β-d-glucopyranose 2,3,4,6-tetraacetato- S -derivative that is a chimera of the structures of GoPI and auranofin, designed to improve stability and bioavailability of GoPI. These metal-ligand complexes are of particular interest because of their combined abilities to irreversibly target the essential dithiol/selenol catalytic pair of selenium-dependent thioredoxin reductase activity, and to kill cells from breast and brain tumors. In this work, screening of various parasites-protozoans, trematodes, and nematodes-was undertaken to determine the in vitro killing activity of GoPI-sugar compared to auranofin. GoPI-sugar was found to efficiently kill intramacrophagic Leishmania donovani amastigotes and adult filarial and trematode worms.

Menin‑mixed‑lineage leukemia (MLL) inhibitors have potential for use as therapeutic agents for MLL‑rearranged leukemia. They are also effective against solid cancers, such as breast cancer. The present study demonstrated that menin‑MLL inhibitors, such as MI‑463, unexpectedly induced the ferroptotic cell death of several cancer cell lines. MI‑463 at a double‑digit nM concentration markedly decreased the viable number of OVCAR‑8 ovarian cancer cells for 3 days. Ferrostatin‑1 (a ferroptosis inhibitor) almost completely abrogated the MI‑463‑induced decrease in viable cell numbers. Furthermore, the cancer cell‑killing activity was inhibited by N‑acetylcysteine [a scavenger of reactive oxygen species (ROS)], deferoxamine (DFO, an iron chelator), PD146176 (a specific inhibitor of arachidonate 15‑lipoxygenase), idebenone (a membrane‑permeable analog of CoQ10) and oleic acid [a monounsaturated fatty acid and one of the end products of stearoyl‑CoA desaturase 1 (SCD1)], whereas Z‑VAD‑FMK (an apoptosis inhibitor) had a negligible effect on cell death. It was also found that MI‑463 in combination with auranofin (a thioredoxin reductase inhibitor) synergistically increased cancer the death of breast, ovarian, pancreatic and lung cancer cell lines (88%, 14/16 cell lines). The synergistic induction of cell death was abrogated by ferroptosis inhibitor and DFO. Inhibitors of SCD1, similar to MI‑463, also enhanced cancer cell death synergistically with auranofin, while inhibitors of SCD1 and MI‑463 did not additively induce cell death. Treatment with zinc protoporphyrin‑9, a specific inhibitor of heme oxygenase‑1 (HO‑1), markedly attenuated the cell death induced by MI‑463 plus auranofin. On the whole, these results suggest that the MI‑463‑induced decrease in cell viability may be at least partly associated with the inhibition of SCD1 activity. In addition, the potent induction of HO‑1 contributed to the synergistic effects of MI‑463 plus auranofin. Therefore, menin‑MLL inhibitors, such as MI‑463, in combination with auranofin represent an effective therapeutic approach for several types of cancer via the induction of ferroptosis.

The antiproliferative properties of a series of structurally-related gold(I) and silver(I) linear complexes inspired to the clinically established gold-based drug auranofin were investigated in A2780 ovarian cancer cells and in their auranofin (A2780/AF-R) and cisplatin (A2780/CDDP-R) resistant counterparts. In A2780 cells and in the cisplatin-resistant subline, gold-based analogues manifested a cytotoxicity profile comparable or superior to auranofin, while the silver-based analogues were less active; both gold and silver complexes overcame cisplatin resistance. Yet, a high degree of cross resistance toward gold analogues was noticed in A2780/AF-R cells. In the same cell line cross-resistance for silver analogues was also observed, though lower. All metal complexes were scrutinized for their ability to inhibit thioredoxin reductase (TrxR), the putative primary target for auranofin: overall, gold compounds were more potent TrxR inhibitors than the corresponding silver compounds, probably, as the consequence of the stronger binding of gold to the active site selenocysteine residue. These results highlight that the thiosugar ligand of auranofin is not essential for cytotoxicity while the nature of the metal center (gold/silver) plays a relevant role in its modulation. In addition, a rather clear correlation was found between cytotoxic potency of tested compounds and their ability to inhibit TrxR activity, being gold compounds more effective than silver analogues. However, the residual TrxR activity, measured in A2780 cells treated with the half-maximal inhibitory concentrations of various metal complexes, resulted far higher than expected. These results suggest that additional cytotoxic mechanisms must be operative. The implications of these results are discussed., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Three pseudohalide analogues of the established gold drug auranofin (AF hereafter), of general formula Au(PEt 3 )X, i.e. Au(PEt 3 )CN, Au(PEt 3 )SCN and Au(PEt 3 )N 3 (respectively denoted as AFCN, AFSCN and AFN 3 ), were prepared and characterized. The crystal structure was solved for Au(PEt 3 )SCN highlighting the classical linear geometry of the 2-coordinate gold(I) center. The solution behaviour of the compounds was then comparatively analysed through 31 PNMR 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 31 PNMR. For all the studied gold compounds, the [Au(PEt 3 )] + 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.

Auranofin (AuRF) has been reported to display anticancer activity and has entered several clinical trials; however, its mechanism of action remains largely unknown. In this work, the anticancer mechanism of auranofin was investigated using a proteomics strategy entailing subcellular fractionation prior to mass spectrometric analysis. Bioinformatics analysis of the nuclear sub-proteomes revealed that tumor suppressor p14 ARF is a key regulator of transcription. Through independent analysis, we validated that up-regulation of p14 ARF is associated with E2F-dependent transcription and increased p53 expression. Our analyses further reveal that 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), which is the rate-determining enzyme of the mevalonate pathway, is a novel target of auranofin with half maximal inhibitory concentration at micromolar levels. The auranofin-induced cancer cell death could be partially reverted by the addition of downstream products of the mevalonate pathway (mevalonolactone or geranyleranyl pyrophosphate (GGPP)), implying that auranofin may target the mevalonate pathway to exert its anticancer effect.

Auranofin is a gold (I)-containing compound used for the treatment of rheumatic arthritis. Auranofin has anticancer activity in animal models and is approved for clinical trials for lung and ovarian carcinomas. Both the cytosolic and mitochondrial forms of the selenoprotein thioredoxin reductase (TrxR) are well documented targets of auranofin. Auranofin was recently reported to also inhibit proteasome activity at the level of the proteasome-associated deubiquitinases (DUBs) UCHL5 and USP14. We here set out to re-examine the molecular mechanism underlying auranofin cytotoxicity towards cultured cancer cells. The effects of auranofin on the proteasome were examined in cells and in vitro, effects on DUB activity were assessed using different substrates. The cellular response to auranofin was compared to that of the 20S proteasome inhibitor bortezomib and the 19S DUB inhibitor b-AP15 using proteomics. Auranofin was found to inhibit mitochondrial activity and to an induce oxidative stress response at IC 50 doses. At 2-3-fold higher doses, auranofin inhibits proteasome processing in cells. At such supra-pharmacological concentrations USP14 activity was inhibited. Analysis of protein expression profiles in drug-exposed tumor cells showed that auranofin induces a response distinct from that of the 20S proteasome inhibitor bortezomib and the DUB inhibitor b-AP15, both of which induced similar responses. Our results support the notion that the primary mechanism of action of auranofin is TrxR inhibition and suggest that proteasome DUB inhibition is an off-target effect. Whether proteasome inhibition will contribute to the antineoplastic effect of auranofin in treated patients is unclear but remains a possibility., (Copyright © 2019 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

Gold compounds have a long history of use in medicine. Auranofin was developed more than 30 years ago as an oral therapy for rheumatoid arthritis. Now, however, auranofin is rarely used in clinical practice despite its efficacy for treating rheumatoid arthritis because more novel antirheumatic medications are available. Although its use in clinical practice has decreased, studies on auranofin have continued and it shows promise for the treatment of several different diseases, including cancer and bacterial and parasitic infections. Several potential novel applications of auranofin for treating human disease have been proposed. Auranofin inhibits the activity of thioredoxin reductase (TrxR), an enzyme of the thioredoxin (Trx) system that is important for maintaining the intracellular redox state. Particularly in cancers, TrxR inhibition leads to an increase in cellular oxidative stress and induces apoptosis. TrxR overexpression is associated with aggressive tumor progression and poor survival in patients with breast, ovarian, and lung cancers. The Trx system may represent an attractive target for the development of new cancer treatments. Therefore, the TrxR inhibitor auranofin may be a potent anticancer agent. This review summarizes the current understanding of auranofin for cancer therapy.

Background: The orally available gold complex auranofin (AF) has been used in humans, primarily as an antirheumatic/immunomodulatory agent. It has been safely administered to healthy dogs to establish pharmacokinetic parameters for oral administration, and has also been used as a treatment in some dogs with immune-mediated conditions. Multiple in vitro studies have recently suggested that AF may possess antineoplastic properties. Spontaneous canine lymphoma may be a very useful translational model for the study of human lymphoma, prompting the evaluation of AF in canine lymphoma cells., Methods: We investigated the antineoplastic activity of AF in 4 canine lymphoid tumor derived cell lines through measurements of proliferation, apoptosis, thioredoxin reductase (TrxR) activity and generation of reactive oxygen species (ROS), and detected the effects of AF when combined with conventional cytotoxic drugs using the Chou and Talalay method. We also evaluated the antiproliferative effects of AF in primary canine lymphoma cells using a bioreductive fluorometric assay., Results: At concentrations that appear clinically achievable in humans, AF demonstrated potent antiproliferative and proapoptotic effects in canine lymphoid tumor cell lines. TrxR inhibition and increased ROS production was observed following AF treatment. Moreover, a synergistic antiproliferative effect was observed when AF was combined with lomustine or doxorubicin., Conclusions: Auranofin appears to inhibit the growth and initiate apoptosis in canine lymphoma cells in vitro at clinically achievable concentrations. Therefore, this agent has the potential to have near-term benefit for the treatment of canine lymphoma, as well as a translational model for human lymphoma. Decreased TrxR activity and increasing ROS production may be useful biomarkers of drug exposure.

Dinuclear metal complexes have emerged as a promising class of anticancer compounds with the ability to cross-link biomolecular targets. Here, we describe two novel series of phosphine-linked dinuclear ruthenium(II) p-cymene and gold(I) complexes, in which the length of the connecting poly(ethylene glycol) chain has been systematically modified. The impact of the multinuclearity, lipophilicity, and linker length on the antiproliferative activity of the compounds on tumorigenic (A2780 and A2780cisR) and nontumorigenic (HEK-293) cell lines was assessed. The dinuclear ruthenium(II) complexes were considerably more cytotoxic than their mononuclear counterparts, and a correlation between the lipophilicity of the linker and the cytotoxicity was observed, whereas the cytotoxicity of the gold(I) series is independent of these factors.

New gold(I) thiolate complexes have been synthesized and characterized, and their physicochemical properties and anticancer activity have been tested. The coordination of PTA derivatives provides optimal hydrophilicity/lipophilicity properties to the complexes, which present high solution stability. Moreover, the complexes show a high anticancer activity against Caco-2 cells, comparable to that of auranofin, and a very low cytotoxic activity against enterocyte-like differentiated cells. Their activity has been shown to produce cell death by apoptosis and arrest of the cell cycle because of interaction with the reductase enzymes and consequent reactive oxygen species production. Some of these new complexes are also able to decrease the necessary dose of 5-fluorouracil, a drug used for the treatment of colon cancer, by a synergistic mechanism.

Urokinase-type plasminogen activator (uPA) has been validated as a predictive or prognostic biomarker protein, and mesupron is considered the first-in-class anticancer agent to inhibit uPA activity in human breast cancer. In the present study, we showed that the synergism between mesupron and auranofin, a thioredoxin reductase inhibitor, for inducing of apoptosis in MCF-7 human breast cancer cells. Our results demonstrated that mesupron and auranofin significantly lead to inhibition of the cancer cells proliferation; cell cycle arrest at the G1/S phase of the cell cycle, and apoptosis as indicated by caspase 3 activation, poly(ADP-ribose) polymerase cleavage, and annexin V staining. Isobologram analyses of MCF-7 cells showed a clear synergism between mesupron and auranofin. This combined treatment decreased the levels of mitochondrial anti-apoptotic factors, such as BCL-2, BCL-xL, and MCL-1 and caused nuclear translocation of apoptosis-inducing factor. Mitochondrial membrane potential (Δψ m ) was found to be strongly disrupted in combination-treated cells. In addition, combination treatment significantly enhanced the overproduction of reactive oxygen species, which was rescued by N-acetylcysteine treatment. The combination treatment suppressed phosphorylation of Akt, thus contributing to apoptosis. Taken together, our data suggest that the use of mesupron in combination with auranofin may be important in achieving high anticancer synergy.

Kallikrein-related peptidase 6 (KLK6) is a biomarker of gastric cancer associated with poor prognosis. Mechanisms by which KLK6 could be exploited for chemotherapeutic use are unclear. We evaluated auranofin (AF), a compound with cytotoxic effects, in KLK6-deficient cells, and we investigated whether KLK6 expression induces autophagy and acquisition of drug resistance in gastric cancer. Using cultured human cells and a mouse xenograft model, we investigated how KLK6 affects antitumor-reagent-induced cell death and autophagy. Expression levels of KLK6, p53, and autophagy marker LC3B were determined in gastric cancer tissues. We analyzed the effects of knockdown/overexpression of KLK6, LC3B, and p53 on AF-induced cell death in cancer cells. Increased KLK6 expression in human gastric cancer tissues and cells inhibited AF-induced cell motility due to increased autophagy and p53 levels. p53 dependent induction of KLK6 expression increased autophagy and drug resistance, whereas KLK6 silencing decreased the autophagy level and increased drug sensitivity. During AF-induced cell death, KLK6 and LC3B colocalized to autophagosomes, associated with p53, and were then trafficked to the cytosol. In the xenograft model of gastric cancer, KLK6 expression decreased AF-induced cell death and KLK6-induced autophagy increased AF resistance. Taken together, the data suggest that the induction of autophagic processes through KLK6 expression may increase acquisition of resistance to AF. Our findings may contribute to a new paradigm for tumor therapeutics.

Inhibition of proteasome-associated deubiquitinases (DUBs) is emerging as a novel strategy for cancer therapy. It was recently reported that auranofin (Aur), a gold (I)-containing compound used clinically to treat rheumatoid arthritis, is a proteasome-associated DUB inhibitor. Disulfiram (DSF), an inhibitor of aldehyde dehydrogenase, is currently in clinical use for treating alcoholism. Recent studies have indicated that DSF can also act as an antitumor agent. We investigated the effect of combining DSF and Aur on apoptosis induction and tumor growth in hepatoma cancer cells. Here we report that (i) the combined treatment of Aur and DSF results in synergistic cytotoxicity to hepatoma cells in vitro and in vivo; (ii) Aur and DSF in combination induces caspase activation, endoplasmic reticulum (ER) stress, and reactive oxygen species (ROS) production; (iii) pan-caspase inhibitor z-VAD-FMK could efficiently block apoptosis but not proteasome inhibition induced by Aur and DSF combined treatment, and ROS is not required for Aur+DSF to induce apoptosis. Collectively, we demonstrate a model of synergism between DSF and proteasome-associated DUB inhibitor Aur in the induction of apoptosis in hepatoma cancer cells, identifying a potential novel anticancer strategy for clinical use in the future.

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., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The mammalian thioredoxin reductases (TrxRs) are a family of selenium-containing pyridine nucleotide disulfide oxidoreductases playing a central role in cellular redox homeostasis and signaling pathways. Recently, these selenoproteins have emerged as promising therapeutic targets for anticancer drug development, often being overexpressed in tumor cells and contributing to drug resistance. Herein, we summarize the current knowledge on metal- and semimetal-containing molecules capable of hampering mammalian TrxRs, with an emphasis on compounds reported in the last decade.

Gold phosphine complexes, such as auranofin, have been recognized for decades as antirheumatic agents. Clinical trials are now underway to validate their use in anticancer or anti-HIV treatments. However, their mechanisms of action remain unclear. A challenging question is whether the gold phosphine complex is a prodrug that is administered in an inactive precursor form or rather that the gold atom remains attached to the phosphine ligand during treatment. In this study, we present two novel gold complexes, which we compared to auranofin and to their phosphonium analogue. The chosen ligand is a phosphine-based smart probe, whose strong fluorescence depends on the presence of the gold atom. The in vitro biological action of the gold complexes and the phosphonium derivative were investigated, and a preliminary in vivo study in healthy zebrafish larvae allowed us to evaluate gold complex biodistribution and toxicity. The different analyses carried out showed that these gold complexes were stable and behaved differently from phosphonium and auranofin, both in vitro and in vivo. Two-photon microscopy experiments demonstrated that the cellular targets of these gold complexes are not the same as those of the phosphonium analogue. Moreover, despite similar IC50 values in some cancer cell lines, gold complexes displayed a low toxicity in vivo, in contrast to the phosphonium salt. They are therefore suitable for future in vivo investigations.

Auranofin (Au), an inhibitor of thioredoxin reductase, is a known anti‑cancer drug. In the present study, the anti‑growth effect of Au on HeLa cervical cancer cells was examined in association with levels of reactive oxygen species (ROS) and glutathione (GSH). Au inhibited the growth of HeLa cells with an IC50 of ~2 µM at 24 h. This agent induced apoptosis and necrosis, accompanied by the cleavage of poly (ADP‑ribose) polymerase and loss of mitochondrial membrane potential. The pan‑caspase inhibitor, benzyloxycarbonyl‑Val‑Ala‑Asp‑fluoromethylketone, prevented apoptotic cell death and each of the assessed caspase inhibitors inhibited necrotic cell death induced by Au. With respect to the levels of ROS and GSH, Au increased intracellular O2•- in the HeLa cells and induced GSH depletion. The pan‑caspase inhibitor reduced the levels of O2•- and GSH depletion in Au‑treated HeLa cells. The antioxidant, N‑acetyl cysteine, not only attenuated apoptosis and necrosis in the Au‑treated HeLa cells, but also decreased the levels of O2•- and GSH depletion in the cells. By contrast, L‑buthionine sulfoximine, a GSH synthesis inhibitor, intensified cell death O2•- and GSH depletion in the Au‑treated HeLa cells. In conclusion, Au induced apoptosis and necrosis in HeLa cells via the induction of oxidative stress and the depletion of GSH.

B-cell malignancies are a common type of cancer. One approach to cancer therapy is to either increase oxidative stress or inhibit the stress response systems on which cancer cells rely. In this study, we combined nontoxic concentrations of Auranofin (AUR), an inhibitor of the thioredoxin system, with nontoxic concentrations of buthionine-sulfoximine (BSO), a compound that reduces intracellular glutathione levels, and investigated the effect of this drug combination on multiple pathways critical for malignant B-cell survival. Auranofin interacted synergistically with BSO at low concentrations to trigger death in multiple malignant B-cell lines and primary mantle-cell lymphoma cells. Additionally, there was less toxicity toward normal B cells. Low AUR concentrations inhibited thioredoxin reductase (TrxR) activity, an effect significantly increased by BSO cotreatment. Overexpression of TrxR partially reversed AUR+BSO toxicity. Interestingly, the combination of AUR+BSO inhibited nuclear factor κB (NF-κB) signaling. Moreover, synergistic cell death induced by this regimen was attenuated in cells overexpressing NF-κB proteins, arguing for a functional role for NF-κB inhibition in AUR+BSO-mediated cell death. Together, these findings suggest that AUR+BSO synergistically induces malignant B-cell death, a process mediated by dual inhibition of TrxR and NF-κB, and such an approach warrants further investigation in B-cell malignancies., (Copyright © 2015 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.)

Auranofin is a gold-containing compound classified by the World Health Organization as a clinically established rheumatoid arthritis therapeutic agent. Through drug screening for novel anticancer therapeutics, we unexpectedly identified auranofin as a potent anticancer agent against a p53-null ovarian carcinoma SKOV3 cell line. However, the molecular mechanism underlying auranofin-mediated anticancer activity in ovarian cancer cells is basically unknown. Here, we show that auranofin inhibits proliferation and survival of SKOV3 cells in a dose‑ and time‑dependent manner. Auranofin treatment activates the pro-apoptotic caspase-3, increases protein levels of apoptosis-inducing proteins Bax and Bim and reduces the expression of the anti-apoptotic mediator Bcl-2 in SKOV3 cells. Moreover, auranofin downregulates IκB kinase (IKK)-β and promotes nuclear localization and the activation of FOXO3 tumor suppressor, leading to cellular apoptosis in SKOV3 cells. In contrast, silencing FOXO3 diminishes the pro-apoptotic signaling of auranofin in SKOV3 cells. These results suggest that auranofin may induce caspase-3-mediated apoptosis in a FOXO3-dependent manner. The observed upregulation of pro-apoptotic genes and apoptosis in cancer cells without p53 in response to auranofin suggests a novel p53-independent mechanism underlying auranofin-induced apoptosis in ovarian cancer cells.

Chronic lymphocytic leukemia (CLL) exhibits high remission rates after initial chemoimmunotherapy, but with relapses with treatment, refractory disease is the most common outcome, especially in CLL with the deletion of chromosome 11q or 17p. In addressing the need of treatments for relapsed disease, we report the identification of an existing U.S. Food and Drug Administration-approved small-molecule drug to repurpose for CLL treatment. Auranofin (Ridaura) is approved for use in treating rheumatoid arthritis, but it exhibited preclinical efficacy in CLL cells. By inhibiting thioredoxin reductase activity and increasing intracellular reactive oxygen species levels, auranofin induced a lethal endoplasmic reticulum stress response in cultured and primary CLL cells. In addition, auranofin displayed synergistic lethality with heme oxygenase-1 and glutamate-cysteine ligase inhibitors against CLL cells. Auranofin overcame apoptosis resistance mediated by protective stromal cells, and it also killed primary CLL cells with deletion of chromosome 11q or 17p. In TCL-1 transgenic mice, an in vivo model of CLL, auranofin treatment markedly reduced tumor cell burden and improved mouse survival. Our results provide a rationale to reposition the approved drug auranofin for clinical evaluation in the therapy of CLL., (©2014 AACR.)

It has been a long story of the development of anticancer metallopharmaceuticals since the identification of cisplatin. Advances in metallodrugs discovery during the past 40 years have made it an ever-growing area of research in medicinal inorganic chemistry. Meanwhile, the emerging of N-heterocyclic carbene (NHC) chemistry has stimulated the newly burgeoning interests in the biomedical applications of metal-NHC complexes. This review will detail what have been achieved hitherto in the research of metal-NHC complexes as potential anti-tumor agents coupled with gold, silver, copper, platinum and palladium. Their mechanism of action will also be discussed. All the results obtained indicate that this promising approach is worthy of more focuses and further studies.

Thioredoxin system plays an important role in regulation of intracellular redox balance and various signaling pathways. Thioredoxin reductase (TrxR) is overexpressed in many cancer cells and has been identified as a potential target of anticancer drugs. Auranofin (AF) is potent TrxR inhibitor with novel in vitro and in vivo anticancer activities. Selenocystine (SeC) is a nutritionally available selenoamino acid with selective anticancer effects through induction of apoptosis. In the present study, we demonstrated the synergistic effects and the underlying molecular mechanisms of SeC in combination with AF on MCF-7 human breast cancer cells. The results showed that SeC and AF synergistically inhibited the cancer cell growth through induction of ROS-dependent apoptosis with the involvement of mitochondrial dysfunction. DNA damage-mediated p53 phosphorylation and down-regulation of phosphorylated AKT and ERK also contributed to cell apoptosis. Moreover, we demonstrated the important role of TrxR activity in the synergistic action of SeC and AF. Taken together, our results suggest the strategy to use SeC and AF in combination could be a highly efficient way to achieve anticancer synergism by targeting TrxR.

Gold(I) complexes with 1,3-substituted imidazole-2-ylidene and benzimidazole-2-ylidene ligands of the type NHC-Au-L (NHC = N-heterocyclic carbene L = Cl or 2-mercapto-pyrimidine) have been synthesized and structurally characterized. The compounds were evaluated for their antiproliferative properties in human ovarian cancer cells sensitive and resistant to cisplatin (A2780S/R), as well in the nontumorigenic human embryonic kidney cell line (HEK-293T), showing in some cases important cytotoxic effects. Some of the complexes were comparatively tested as thioredoxin reductase (TrxR) and glutathione reductase (GR) inhibitors, directly against the purified proteins or in cell extracts. The compounds showed potent and selective TrxR inhibition properties in particular in cancer cell lines. Remarkably, the most effective TrxR inhibitors induced extensive oxidation of thioredoxins (Trxs), which was more relevant in the cancerous cells than in HEK-293T cells. Additional biochemical assays on glutathione systems and reactive oxygen species formation evidenced important differences with respect to the classical cytotoxic Au(I)-phosphine compound auranofin.

17-Allylamino-17-demethoxygeldanamycin (17AAG) is an experimental chemotherapeutic agent believed to form free radicals in vivo, and cancer cell resistance to 17AAG is believed to be a thiol-dependent process. Inhibitors of thiol-dependent hydroperoxide metabolism [L-buthionine-S,R-sulfoximine (BSO) and auranofin] were combined with the glucose metabolism inhibitor 2-deoxy-d-glucose (2DG) to determine if 17AAG-mediated cancer cell killing could be enhanced. When 2DG (20mM, 24h), BSO (1mM, 24h), and auranofin (500nM, 3h) were combined with 17AAG, cell killing was significantly enhanced in three human cancer cell lines (PC-3, SUM159, MDA-MB-231). Furthermore, the toxicity of this drug combination was significantly greater in SUM159 human breast cancer cells, relative to HMEC normal human breast epithelial cells. Increases in toxicity seen with this drug combination also correlated with increased glutathione (GSH) and thioredoxin (Trx) oxidation and depletion. Furthermore, treatment with the thiol antioxidant NAC (15mM, 24h) was able to significantly protect from drug-induced toxicity and ameliorate GSH oxidation, Trx oxidation, and Trx depletion. These data strongly support the hypothesis that simultaneous inhibition of GSH- and Trx-dependent metabolism is necessary to sensitize human breast and prostate cancer cells to 2DG+17AAG-mediated killing via enhancement of thiol-dependent oxidative stress. These results suggest that simultaneous targeting of both GSH and Trx metabolism could represent an effective strategy for chemosensitization in human cancer cells., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Complexes [Au(H2Ac4DH)Cl]∙MeOH (1) [Au(H(2)2Ac4Me)Cl]Cl (2) [Au(H(2)2Ac4Ph)Cl]Cl∙2H(2)O (3) and [Au(H(2)2Bz4Ph)Cl]Cl (4) were obtained with 2-acetylpyridine thiosemicarbazone (H2Ac4DH), its N(4)-methyl (H2Ac4Me) and N(4)-phenyl (H2Ac4Ph) derivatives, as well as with N(4)-phenyl 2-benzoylpyridine thiosemicarbazone (H2Bz4Ph). The compounds were cytotoxic to Jurkat (immortalized line of T lymphocyte), HL-60 (acute myeloid leukemia), MCF-7 (human breast adenocarcinoma) and HCT-116 (colorectal carcinoma) tumor cell lines. Jurkat and HL-60 cells were more sensitive than MCF-7 and HCT-116 cells. Upon coordinating to the gold(I) metal centers in complexes (2) and (4), the cytotoxic activity of the H2Ac4Me and H2Bz4Ph ligands increased against the HL-60 and Jurkat tumor cell lines. 2 was more active than auranofin against both leukemia cells. Most of the studied compounds were less toxic than auranofin to peripheral blood mononuclear cells (PBMC). All compounds induced DNA fragmentation in HL-60 and Jurkat cells indicating their pro-apoptotic potential. Complex (2) strongly inhibited the activity of thioredoxin reductase (TrxR), which suggests inhibition of TrxR to be part of its mechanism of action., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Constitutive activation of NF-κB and STAT3 plays an important role in the cellular proliferation and survival of multiple myeloma cells. We first found that auranofin (AF), a coordinated gold compound, induced a significant level of cell cycle arrest at G1 phase and subsequent apoptosis of myeloma cells. Further, AF inhibited constitutive and IL-6-induced activation of JAK2 and phosphorylation of STAT3 followed by the decreased expression of Mcl-1. AF down-regulated the activation of NF-κB, and the combination of AF and a specific NF-κB inhibitor resulted in a marked decrease of Mcl-1 expression. These results suggest that AF inhibits both IL-6 induced-JAK/STAT pathway and NF-κB activation in myeloma cells., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Background and Purpose: Arsenicals have been used medicinally for decades to treat both infectious disease and cancer. Arsenic trioxide (As2O3) is effective for treatment of acute promyelocytic leukaemia (APL), yet the mechanism of action of this drug is still widely debated. Recently, As2O3 was shown to inhibit the activity of the selenoenzyme thioredoxin reductase (TrxR). TrxR has been proposed to be required for selenium metabolism. The effect of inhibitors of TrxR on selenium metabolism has yet to be assessed. This study aims to determine whether chemotherapeutics that target selenocysteine within selenoenzymes may also affect the metabolism of selenium., Experimental Approach: A lung cell line, A549, was used to assess the effect of TrxR inhibitors on selenium metabolism, using 75Se-selenite. The level of mRNA encoding cytosolic TrxR (TrxR1) was determined using real-time reverse transcriptase-PCR. TrxR activity was determined in whole-cell extracts., Key Results: Exposure of cells to As2O3, arsenite or auranofin led to a concentration-dependent reduction of selenium metabolism into selenoproteins. Knockdown of TrxR1, using small inhibitory RNA, did not affect selenium metabolism. Exposure of cells to monomethylarsonic acid, a potent inhibitor of TrxR, did not alter selenium metabolism but did inhibit enzyme activity., Conclusions and Implications: As2O3 and auranofin block the metabolism of selenium in A549 cells. Because As2O3 is used to treat APL, our findings may reveal the mechanism of this therapeutic action and lead to further research targeting selenium metabolism to find novel chemotherapeutic agents for the treatment of APL.

1-O-Octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3) is a synthetic diether phospholipid that is competitive with phosphatidylserine binding to the regulatory domain of protein kinase C (PKC). Our previous studies indicate that the selective inhibition of tumor cell growth by ET-18-OCH3 may be due to altered signal transduction mechanisms, including the inhibition of PKC. To further define the mechanism of action of ET-18-OCH3, we have used it to study the role of PKC in regulation of the transcription factor NF-kappa B, which is activated by diverse stimuli. In the 293.27.2 human kidney cell line, as in hematopoietic cells of all lineages, NF-kappa B is stimulated by 12-O-tetradecanoylphorbol-13-acetate (TPA), tumor necrosis factor-alpha (TNF-alpha), and interleukin-1 alpha (IL-1 alpha). The response to either TNF-alpha or IL-1 alpha is synergistically enhanced by TPA. However, the regulatory mechanisms and signal transduction systems responsible for NF-kappa B activation in response to these different stimuli have not been determined in detail. We have used ET-18-OCH3 and auranofin, which inhibit PKC by different mechanisms, to assess the role of PKC in NF-kappa B activation. ET-18-OCH3 markedly inhibits TPA-induced NF-kappa B activation, as measured by HIV long terminal repeat-directed expression of beta-galactosidase. The IC50 for inhibition by ET-18-OCH3 is approximately 2 microM, a noncytotoxic concentration. Inhibition of TPA-induced NF-kappa B activation was dependent upon preincubation with ET-18-OCH3, and the drug was active at approximately 2 mol% of total cellular phospholipid. ET-18-OCH3 did not inhibit NF-kappa B activation by either TNF-alpha or IL-1 alpha, indicating that there are multiple distinct signal transduction pathways leading to activation of NF-kappa B. We have confirmed these results using auranofin, an antirheumatic drug that is a specific PKC inhibitor interacting with the catalytic domain. Like ET-18-OCH3, auranofin blocked NF-kappa B activation by TPA but not by TNF-alpha or IL-1 alpha. Also like the ether lipid, auranofin only partially blocked the synergy exhibited by TPA and TNF-alpha. To confirm the role of NF-kappa B in this response, we measured NF-kappa B by electrophoretic mobility shift assay. Both ET-18-OCH3 and auranofin inhibited cellular induction of the active NF-kappa B complex in response to TPA but not in response to TNF-alpha.(ABSTRACT TRUNCATED AT 400 WORDS)

SK&F 102912 (mu-[1,2-bis(diphenylphosphino)ethane]bis[(1-thio-beta-D- glucopyranosato-S)gold(I)], [(Autg)2(dppe)]) has shown reproducible and significant activity in transplantable murine tumor models and represents a structurally unique class of antineoplastic agents. A number of in vitro studies were performed to elucidate the cellular pharmacology of this gold-containing complex. [(Autg)2(dppe)] is a potent cytotoxic agent in vitro as demonstrated by its ability to inhibit the clonogenic capacity of a variety of tumor cell lines following a brief exposure to the drug. Cell-cycle analysis using HL-60 cells showed that low concentrations (2 microM) of [(Autg)2(dppe)] induced an S-phase block and higher concentrations induced a secondary block at the G1/S boundary. [(Autg)2(dppe)] had several effects on DNA metabolism and structure including preferential inhibition in cells of DNA synthesis (relative to RNA and protein synthesis) and the production of DNA single- and double-strand breaks as measured by alkaline elution. The cytoxic mechanism of this gold complex appears to be distinct from that of the monophosphine-gold complex auranofin.