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Purpose: Leishmaniasis is a major public health problem worldwide in many parts of the world. Current anti-leishmanial drugs have only limited clinical efficacy. Aurothiomalate derivatives are useful for treating rheumatoid arthritis, but have emerged as a promising therapeutic candidate for leishmaniasis. This paper gives a review of the literature about the usefulness of aurothiomalate derivatives against leishmaniasis., Methods: In this study, we reviewed the proposed mechanisms of action of aurothiomalate and related compounds on the metabolism of L. major and collected data by searching relevant articles., Results: Aurothiomalate-based drugs could be effective against leishmaniasis through two direct and indirect mechanisms: first, cytotoxic effects on parasites via thiomalate's false substrate role in the citric acid cycle against malate; and second, immunosuppressive and anti-inflammatory effects of aurothiomalate derivatives with prostaglandin production inhibitory effects., Conclusions: The current study documented that aurothiomalate-based drugs could be effective against leishmaniasis through two direct and indirect mechanisms of action. Gold thiomalate as a promising hit should be evaluated against L. major in vitro and in vivo conditions in the future., (© 2022. The Author(s) under exclusive licence to Witold Stefański Institute of Parasitology, Polish Academy of Sciences.)

Real-time monitoring of drug metabolism in vivo is of great significance to drug development and toxicology research. The purpose of this study is to establish a rapid and visual in vivo detection method for the detection of an intermediate metabolite of the gold (I) drug. Gold (I) drugs such as sodium aurothiomalate (AuTM) have anti-inflammatory effects in the treatment of rheumatoid arthritis. Gold(III) ions (Au 3+ ) are the intermediate metabolite of gold medicine, and they are also the leading factor of side effects in the treatment of patients. However, the rapid reduction of Au 3+ to Au + by thiol proteins in organisms limits the in-depth study of metabolism of gold drugs in vivo. Here we describe a luminescence Au 3+ probe (RA) based on ruthenium (II) complex for detecting Au 3+ in vitro and in vivo. RA with large Stokes shift, good water solubility and biocompatibility was successfully applied to detect Au 3+ in living cells and vivo by luminescence imaging, and to trap the fluctuation of Au 3+ level produced by gold (I) medicine. More importantly, the luminescent probe was used to the detection of the intermediate metabolites of gold (I) drugs for the first time. Overall, this work offers a new detection tool/method for a deeper study of gold (I) drugs metabolite., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Cells of Escherichia coli Rosetta, containing plasmid pET-28a with sequences of DNA of chitinase from Stenotrophomonas maltophilia N4, were used for the efficient synthesis of recombinant chitinolytic enzyme. The objective of this study was to improve thermal stability of the recombinant chitinase by salts and metal nanoparticles (NP). The studied chitinase was thermolabile and largely lost its activity in the first minutes of storage at 50 and 60 °C. The optimum temperature for colloidal chitin hydrolysis by the enzyme was 50 °C. Application of sodium aurothiomalate hydrate and manganese chloride enhanced the activity of the recombinant enzyme. In general, chitinase activity was higher when silver nanoparticles (Ag-NP) were used, but lower for other NP. The thermal stability of chitinase immobilized on Ag-NP and manganese chloride was significantly higher than that of free chitinase. Chitinase thermal stability after gold and manganese oxide nanoparticle application was higher than that of the control at 50 °C. Platinum nanoparticles had no significant effect on thermostability. The Ag-NP had a smaller diameter (from 2 to 20 nm) than Au-NP (from 5 to 70 nm) and Pt-NP (from 4 to 80 nm). The TEM analysis showed that the used NP had a higher affinity for chitinase than for the synthetic substrate. The type, size, and location of the NP on the enzyme played a major role in the thermal stability of chitinase., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Gold(I) complexes are an important tool in the arsenal of established approaches for treating rheumatoid arthritis (RA), while some recent studies have suggested that gold nanoparticles (Au NPs) may also be therapeutically efficacious. These observations prompted the current biological studies involving gold(I) anti-RA agents and Au NPs, which are aimed towards improving our knowledge of how they work. The cytotoxicity of auranofin, aurothiomalate, aurothiosulfate and Au NPs towards RAW264.7 macrophages was evaluated using the MTT assay, with the former compound proving to be the most toxic. The extent of cellular uptake of the various gold agents was determined using graphite furnace atomic absorption spectrometry, while their distribution within macrophages was examined using microprobe synchrotron radiation X-ray fluorescence spectroscopy. The latter technique showed accumulation of gold in discrete regions of the cell, and co-localisation with sulfur in the case of cells treated with aurothiomalate or auranofin. Electrospray ionization mass spectrometry was used to characterize thioredoxin reductase (TrxR) in which the penultimate selenocysteine residue was replaced by cysteine. Mass spectra of solutions of TrxR and aurothiomalate, aurothiosulfate or auranofin showed complexes containing bare gold atoms bound to the protein, or protein adducts containing gold atoms retaining some of their initial ligands. These results support TrxR being an important target of gold(I) drugs used to treat RA, while the finding that Au NPs are incorporated into macrophages, but elicit little toxicity, indicates further exploration of their potential for treatment of RA is warranted., (Copyright © 2014 Elsevier Inc. All rights reserved.)

This study presents a comparative drug-protein, in vitro, binding profile of sodium aurothiomalate and auranofin. It was found that about 40% of total protein-bound gold is attached to albumin after incubation of aurothiomalate with whole blood for 24 h and about 29% of it was with alpha(1)-globulin and the least amount was found with gamma-globulin (6.1%). On the other hand, approximately 84% of the protein-bound auranofin gold attached to globulins of which 51% was found with beta-globulin band. It was almost equally distributed among albumin, alpha(2)-globulin and gamma-globulin, and showed least affinity for alpha(1)-globulin. The gold analyses were performed by standardless instrumental neutron activation method duly validated by use of an established atomic absorption method. The results of this study explain to some extent the difference in, in vivo, pharmacokinetics and pharmacodynamics of the two drugs.

It has been recently demonstrated that small gold compounds could have a potential anti-tumoral activity. Here, we report that aurothiomalate (ATM), a gold compound already used in clinical therapy for the treatment of rheumatoid arthritis, has a pro-apoptotic effect in aggressive prostate cancer (PC3U) cells. In contrast, treatment of human primary epithelial prostate cells (PrEC) with ATM did not cause apoptosis. We demonstrated that ATM is able to disrupt the PKCiota-Par6 complex in PC3U cells and that this disruption leads to the activation of ERK in a dose-dependent manner. Interestingly, we also showed that ERK acts upstream of the activation of caspase 3, leading to apoptosis. ATM treatment also causes activation of p38 and JNK MAP kinases. Moreover we could link ATM treatment to activation of the mitochondrial or so called intrinsic pathway, as we observed release of cytochrome c from mitochondria to cytoplasm, suggesting that the mitochondrial pathway is involved in the pro-apoptotic effect mediated by ATM. Taken together our data suggest that ATM could be a new promising drug for the treatment of advanced prostate cancer.

The aurocyanide anion, Au(CN) (2) (-) , is a human metabolite of several anti-rheumatic gold complexes containing monovalent gold (I) bound to a sulphur ligand. This article reviews some of the chemical and pharmacological properties of this intriguing metabolite, and reports its anti-arthritic and anti-inflammatory activity in rats. Au(CN) (2) (-) is generated from the therapeutic gold complexes by small amounts of hydrogen cyanide, HCN, produced from thiocyanate, SCN(-), by myeloperoxidase (MPO) an enzyme in neutrophils which normally produces hypochlorite, OCl(-). Thus, Au(CN) (2) (-) is formed at sites of inflammation where activated neutrophils are present. This includes atherosclerotic lesions as well as inflamed joints. MPO also oxidises Au(CN) (2) (-) to Au(III) complexes such as Au(CN) (4) (-) .Au(CN) (2) (-) is normally a very stable monovalent gold complex. In a biological context, only low concentrations are ever present at both extracellular and intracellular sites. However, Au(CN) (2) (-) produced locally may facilitate the cellular uptake and hence the therapeutic and toxic effects of gold drugs. Au(CN) (2) (-) may also be involved in a redox cycle where Au(CN) (2) (-) is oxidised to Au(CN) (4) (-) which is, in turn, reduced back to Au(CN) (2) (-) by endogenous thiols. There are still many questions to be resolved concerning Au(CN) (2) (-) including its intrinsic toxicity and the extent to which it may contribute to the overall anti-arthritic activities of the gold-thiolates from which it is formed in vivo.

Zinc finger motifs are present in a wide variety of regulatory proteins and generally function as interaction modules between macromolecules. These functional interactions are controlled by mechanisms of zinc (Zn2+)-binding and release. Besides Zn2+ certain electrophilic metals can potentially react with zinc finger domains and lead to changes in the structure and function of those domains. In these studies, the Cys2His2 zinc finger was chosen as a model for understanding how the gold (I) (Au1+) drug, aurothiomalate (AuTM), interacts mechanistically with the Zn2+ coordination sphere. DNA binding assays were used to analyze functional interactions between AuTM and two model Cys2His2 zinc finger transcription factors, TFIIIA and Sp1; inhibition in the micromolar range of AuTM was observed in both cases. Electrospray ionization mass spectrometry (ESI-MS) was utilized to examine molecular interactions between AuTM and a zinc finger peptide modeled after the third finger of Sp1 (Sp1-3). These experiments demonstrated Au1+ ions can bind the zinc finger structure and trigger the release of the Zn2+ ion. Quantifying the ESI-MS data allowed for a relative affinity value between Zn2+ and Au1+ ions to be calculated and shows Au1+ has a 4-fold higher affinity for Sp1-3 than Zn2+. Mechanistic differences between Zn2+ and Au1+ binding to the model Sp1-3 zinc finger were analyzed at isotopic resolution, and the metal-coordination spheres were probed with small molecules (H+, hydrogen peroxide, glutathione disulfide, and iodoacetamide). Natural isotope cluster analysis suggested the presence of a metal-thiol bond in the Cys2His2 zinc finger structure. Metal exchange reactions between zinc fingers demonstrated Zn2+ ions exchanged more rapidly than Au1+ ions. Circular dichroism (CD) exhibited differences in the secondary structure of the Sp1-3 model peptide when binding Zn2+ or Au1+ ions.

Thioredoxin reductase (TrxR) is a flavoprotein that contains a C-terminal penultimate selenocysteine (Sec) and has an ability to reduce thioredoxin (Trx), which regulates the activity of NF-kappa B. To date, three TrxR isozymes, TrxR1, TrxR2, and TrxR3, have been identified. In the present study, we found that among these isozymes only TrxR1 was induced by tumor necrosis factor-alpha (TNF alpha) in vascular endothelial cells. Furthermore, the overexpression of TrxR1 enhanced TNF alpha-induced DNA-binding activity of NF-kappa B and NF-kappa B-dependent gene expression. The catalytic Sec residue of TrxR1, which is essential for reducing Trx, was required for this NF-kappa B activation, and aurothiomalate, an inhibitor of TrxR, suppressed TNF alpha-induced activation of NF-kappa B and the expression of NF-kappa B-targeted proinflammatory genes such as E-selectin and cyclooxygenase-2. These results suggest that TrxR1 may act as a positive regulator of NF-kappa B and may play an important role in the cellular inflammatory response.

There is considerable evidence that the anti-rheumatic gold complexes are activated by their conversion to aurocyanide. In order to understand the mechanism of production of aurocyanide, we investigated the involvement of myeloperoxidase in the reaction. This haem enzyme of neutrophils and monocytes uses hydrogen peroxide to oxidise chloride and thiocyanate to hypochlorous acid and hypothiocyanite, respectively. When aurothiomalate (10 microM) was incubated with thiocyanate (200 microM), hydrogen peroxide (100 microM) and myeloperoxidase (20 nM), it was transformed to a product that was spectrally identical to authentic aurocyanide. Aurothiomalate was quantitatively converted to aurocyanide in about 10 min at pH 6.0 and in 40 min at pH 7.4. Aurocyanide formation occurred after myeloperoxidase had used all the hydrogen peroxide available to produce hypothiocyanite. Thus, the cyanide must have formed from the slow decomposition of hypothiocyanite. The rate of aurocyanide production was increased in the presence of 100 mM chloride, which indicates that hypochlorous acid accelerates the formation of cyanide. Hypochlorous acid (100 to 400 microM) reacted non-enzymatically with thiocyanate (200 microM) and aurothiomalate (10 microM) to produce aurocyanide. Thus, aurocyanide is produced by two processes, involving both the formation of hypothiocyanite and hypochlorous acid. Aurocyanide is an effective inhibitor of the respiratory burst of neutrophils and monocytes and the proliferation of lymphocytes. Therefore, aurothiomalate may attenuate inflammation by acting as a pro-drug which is reliant on neutrophils and monocytes to produce hypothiocyanite. When the hypothiocyanite decays to hydrogen cyanide, the pro-drug is converted to aurocyanide which then suppresses further oxidant production by these inflammatory cells.

Treponema denticola, Treponema vincentii and Treponema socranskii produce an enzyme that hydrolyses hyaluronic acid (HA) and chondroitin sulphate (CS). The secreted enzyme is specifically inhibited by gold sodium thiomalate and anti-bee-venom antibodies. The use of saturated substrate (HA or CS) transblots allowed the visualization of active enzyme directly from culture supernatants and is a useful tool in clarification of complex polysaccharide-degrading enzyme specificities. The affinity-purified extracellular enzyme of T. denticola contains a single molecular species with a molecular mass of 59 kDa. Since it hydrolyses both HA and CS, it can more appropriately be termed a hyaluronoglucosaminidase (HGase). The HGase has been localized at the cell surface by electron microscopy and may play an active role in the degradation of connective tissue ground substance in the initiation and progression of periodontal disease.

Certain metal ions are known to be potent sensitizers, but the self proteins modified by metal ions and the self peptides recognized by 'metal-specific' T cells are unknown. In humans and mice treatment with gold anti-rheumatic drugs, containing Au(I), may lead to allergic and autoimmune side effects. Human and murine T cells do not react to Au(I), however, but to the reactive metabolite Au(III). Here we show that alteration by Au(III) of a model antigen, bovine ribonuclease (RNase)A, results in T cell sensitization to cryptic peptides of this protein. Upon immunization of mice with Au(III)-pretreated RNase [RNase/Au(III)], CD4+ T cell hybridomas specific for RNase/Au(III) were obtained in addition to those recognizing the immunodominant peptide RNase 74-88; the latter also were obtained after immunization with native RNase. RNase/Au(III)-specific T cell hybridomas reacted against RNase/Au(III) and RNase denatured by S-sulfonation of cysteine residues, but not against native RNase, or RNase pretreated with Au(I), A1(III), Cu(II), Fe(II), Fe(III), Ni(II), Mn(II), or Zn(II). Using a panel of overlapping, synthetic RNase peptides which were devoid of gold or gold-induced modifications, epitope mapping revealed that RNase/Au(III)-specific T cell hybridomas recognized the cryptic peptides 7-21 and 94-108, respectively. Comparison of the proliferative response of bulk CD4+ T cells, prepared from splenocytes after immunization with either RNase/Au(III) or native RNase, revealed that Au(III) pretreatment of RNase led to a markedly enhanced response to the two cryptic peptides while it did not influence the response to the immunodominant peptide. The cryptic peptides were also presented after preincubation of bone marrow-derived macrophages with RNase and Au(I), but not with RNase alone, suggesting that oxidation of Au(I) to Au(III) and subsequent protein alteration by Au(III) can happen in mononuclear phagocytes. We conclude that Au(III) alteration of proteins alters antigen processing and, thus leads to presentation of cryptic peptides. This mechanism may shed light on the development of allergic and autoimmune side effects of Au(I) anti-rheumatic drugs. In addition, it might provide a general mechanism of how metal ions act as T cell sensitizers.

Background: Chrysiasis is a rare blue-gray skin discoloration that occurs in sun-exposed sites of some patients who receive gold salts. A unique case of localized chrysiasis developed immediately after Q-switched ruby laser (694 nm) irradiation for postinflammatory hyperpigmentation secondary to granuloma faciale in a patient who was receiving long-term gold sodium thiomalate therapy for psoriatic arthritis. Skin biopsy specimens showed striking changes in the ultrastructural characteristics of cutaneous gold deposits following laser treatment., Observations: A blue-gray skin discoloration developed immediately after laser exposure and persisted unchanged after 1 year. Transmission electron microscopy of skin biopsy specimens showed electron-dense gold deposits. Before laser irradiation, these deposits were 106 +/- 35 (mean +/- SD) nm in diameter and faceted, consistent with a crystalline structure. Posttreatment deposits were round, smaller, measured 16 +/- 4 nm, and resembled colloidal gold. Identical findings were observed in an area of sun-protected skin treated with the Q-switched ruby laser; irradiation with a pulsed dye laser at 585 nm had no effect., Conclusions: Localized chrysiasis was induced in a patient receiving parenteral gold therapy who underwent treatment with a Q-switched ruby laser. This form of chrysiasis resulted from a structural alteration in dermal gold deposits. A similar physiochemical modification in gold deposits induced by UV light may explain the localization of chrysiasis to sun-exposed skin in affected patients.

1H NMR methods are described which allow direct studies of the Cys34 binding site of albumin in intact human blood plasma in vitro. Antiarthritic gold drugs and the alcohol-aversive drug disulfiram induce a structural transition detectable via H epsilon 1 and H delta 2 resonances of His3 of albumin, and reactions of cystine, glutathione and captopril in plasma have also been investigated. Contrary to most assumptions, little of the albumin in normal plasma appears to be blocked at Cys34 as a cystine disulfide.

The reactions of Myocrisin with albumin and gamma-globulin have been investigated using atomic absorption spectrometry and gel filtration. Albumin rapidly binds gold up to the levels predicted from the concentration of free sulphydryl groups present in the protein. However, in the presence of glutathione, gold incorporation is increased, suggesting that in vivo, free thiols (glutathione, thiomalate) may facilitate gold uptake by the protein. In comparison, gamma-globulin is found to be capable of binding up to one atom of gold per molecule of protein in a slow reaction which requires high Myocrisin concentrations.

The oxidizing capacity of phagocytic cells is suspected to play a major role in the generation of immunogenic drug metabolites, in particular those that cause extrahepatic immunopathological lesions. In the case of the antirheumatic drug gold(I) disodium thiomalate (Na2Au(I)TM), oxidation of the Au(I) ion to Au(III) appears to be responsible for the adverse immune reactions which may develop during gold therapy. Here, we show that the reactive metabolite Au(III) may be generated by mononuclear phagocytes (M phi) exposed to Au(I). The generation of Au(III) was analyzed by means of the adoptive transfer popliteal lymph node assay (PLNA) in mice, using T lymphocytes previously sensitized to Au(III) as a detection probe. Donors of the Au(III)-primed T cells were either directly sensitized to Au(III) by injection of tetrachloroauric acid (HAu(III)Cl4), or indirectly via chronic treatment with Na2Au(I)TM. As donors of peritoneal cells (PC), we used mice which had received weekly i.m. injections of Na2Au(I)TM for 12 weeks and contained increased numbers of activated B cells. The PC of these mice were found to elicit a significant secondary response when used as antigenic material for the restimulation of Au(III)-primed T cells. The immunogenicity of PC obtained from Na2Au(I)TM-treated mice paralleled the total gold content of these cells. Noteworthily, M phi exposed to Au(I) in vitro also proved capable of eliciting a specific secondary response of Au(III)-primed T cells. Hence, M phi exposed to Au(I) generate the reactive intermediate Au(III) which, apparently via oxidation of self proteins, sensitizes T cells. As M phi are constituents of many different organs and, moreover, communicate with T cells, their capacity to generate Au(III) may account for the various extrahepatic adverse immune reactions induced by Au(I) drugs.

Autometallography was used in conjunction with light and electron microscopy to detect traces of gold and mercury in the dorsal root ganglia of rats treated with sodium aurothiomalate and mercuric chloride. In order to differentiate between gold and mercury in tissue sections, the gold accumulations were removed by potassium cyanide, leaving mercury sulphides/selenides as the only possible catalysts for autometallographic development. With this technique, it is now possible to differentiate between all tissue metals capable of initiating the autometallographic process, i.e. gold, vesicular zinc, and sulphides and selenides of mercury and silver.

Gold was characterized in the urine and bile of rats treated with D-penicillamine (D-PEN), 2,3-dimercaptosuccinic acid (DMSA), 2,3-dimercaptopropane sulphonate (DMPS), or N-(2-mercapto-2-methylpropanoyl)-L-cysteine (bucillamine) immediately after gold sodium thiomalate (AuTM) injection by both gel chromatographic and electrophoretic methods. It is suggested that the gold in the urine and bile after AuTM administration was predominantly bound to high molecular weight compounds. The characterization of gold in the urine after administration of AuTM with D-PEN, DMSA, or DMPS showed that most of the gold was bound to the chelating agents. In the treatment with the chelating agents such as D-PEN and DMPS, the gold was mainly excreted as a gold-chelating agent compound in the bile and a minor portion of the gold was present in the form of a gold-L-cysteine compound and high molecular weight compounds. DMSA treatment showed that a major portion of the gold was bound to high molecular weight compounds in the bile and a minor portion of the gold was present in the forms of gold-DMSA and gold-L-cysteine compounds. The administration of AuTM and bucillamine indicated that the gold was mainly present as a gold-Me-bucillamine compound in the urine and a gold-bucillamine compound in the bile.

A sensitive method for the determination of gold-based drugs auranofin, myochrysine, and their metabolites has been developed. These gold-containing compounds were separated by reversed-phase ion-pair chromatography with tetrabutylammonium chloride as the ion-pairing agent. Gold-specific on-line detection utilized inductively coupled plasma mass spectrometry (ICP-MS). The separation conditions of pH, methanol content, concentration of the ion-pairing agent and ionic strength have been investigated. The detection limit for auranofin, the last peak in the chromatogram, was 0.3 ng. These methods were applied to the analysis of gold-containing species in urine from arthritis patients on auranofin, myochrysine or solganol therapy. The recovery of the total gold-containing species from urine was greater than 90%. Dicyanogold(I) anion, [Au(CN)2]-, was detected in the urine of several patients.

The presence of gold was investigated in sections of the adrenal glands from rats which had been exposed to intraperitoneal sodium aurothiomalate (32 to 120 mg). Gold was histochemically detected in cortical endocrine cells, chromaffin cells and in fibroblasts and macrophages of both the cortex and medulla. Invisible traces of gold were silver enhanced by autometallography making them readily visible at both the light and electron microscopic levels. The intracellular staining intensity was dose-dependent. In general, the number as well as the staining intensity of individual cells, were highest in the zona glomerulosa and zona reticularis. In gold-containing cells the silver-amplified deposits were present in lysosomes.

It is postulated that the efficacy of sodium aurothiomalate and d-penicillamine could be partly dependent on the activity of sulphydryl groups. Free thiomalate, the thiol from sodium aurothiomalate was shown to be liberated in vivo and has been detected in the blood and urine of sodium aurothiomalate-treated patients. An increase in intracellular glutathione levels in response to treatment with sodium aurothiomalate and penicillamine was demonstrated and considered to be related to these drugs' properties as thiols. A pilot clinical trial using sodium thiomalate treatment demonstrated clinical improvement in four out of six patients after one course. Two patients relapsed but both improved after a second course and there were no serious adverse reactions. It is concluded that although larger controlled clinical trials are required, the role of gold in "chrysotherapy" must remain in question.

Four normal male volunteers participated in a study designed to examine the disposition of gold given intramuscularly as gold sodium thiomalate. Blood samples were collected for 32 days following the administration of 10 mg of gold sodium thiomalate. Plasma gold concentrations were determined by atomic absorption spectrometry. A triphasic decay pattern in plasma gold concentrations was observed. Terminal log-linear phases corresponded to a mean disposition half-life of 25 days. Apparent total body clearance of gold was 7.0 +/- 0.6 ml kg-1 day-1 and the apparent volume of distribution was 0.26 +/- 0.051 kg-1. These pharmacokinetic data are in contrast to previous data from other investigators who have reported half-lives of approximately 5 days. Data from the current study provide a sound rationale for the currently used empiric dosing regimens.

Inhibition of the activity of delta-aminolevulinic acid dehydratase by heavy metals such as Cu, Cd and Au has been thought to be related to the inhibition of heme bio-synthesis in the bone marrow. This study investigates the interaction between lead and gold on porphyrin metabolism. Gold thiomalate (Au-T) was injected subcutaneously 10 times into the backs of rabbits in doses of 1, 10 and 100 mg per capita for 30 days. The latter two groups of animals showed mild anemia, but they showed neither reticulocytosis nor increase of free erythrocyte protoporphyrin (FEP). In rabbits administered lead only or lead plus gold thiomalate, however, reticulocytosis and an increase in FEP levels were observed. Therefore, the etiology of anemia may be different between rabbits administered gold thiomalate and rabbits administered lead. Rabbits injected with Au-T showed an increase in urinary coproporphyrin (CP-U) excretion. The maximum concentration of CP-U in the gold groups was about 3 times as high as that in the control group, but the increment was lower than that in the lead group. Rabbits injected with Au-T showed a higher concentration of gold in the kidneys than in other tissues. In those injected lead and Au-T together, concentration of gold in all tissues was higher than in those administered Au-T only. Proteinuria was observed in the rabbits injected Au-T. The increase in CP-U in the gold-injected rabbits was assumed to be related to the increase in gold concentration in the kidneys.

1. The subcellular distribution of gold in the livers, kidney cortex and the spleens of rhesus monkeys was determined at 2, 10 and 20 days after the administration of sodium (195Au)-aurothiomalate. The serum gold levels were also determined over the period of study. 2. With the continued decrease in the serum gold levels from day 1 to day 20, a continued increase in the tissue levels of the organs studied was observed. The corresponding subcellular fractions of the tissues also showed continued increases in the radioactive concentrations of gold. 3. Gold was found to localise in all the organelles of the cells in the liver, kidney cortex and the spleen, though in varying amounts. The largest increases in the concentrations of gold from day 2 to day 20 were found in the lysosomal bodies of liver and kidney. The concentrations in the kidney cortex increased from approximately five times those in the liver or the spleen on day 2 to about eight times those on day 20.

The localization of gold in normal, stimulated and activated mouse peritoneal cells exposed in vivo to sodium aurothiomalate was examined by a new histochemical technique based on physical development. Activated macrophages from mice infected with herpes simplex virus type II showed the greatest amount of gold localized within lysosomes. Mast cells contained gold in their granules. Polymorphonuclear leucocytes and lymphocytes, on the other hand, were devoid of gold. Pinocytosis is discussed as the mechanism by which gold is taken up into the cells.

The binding of sodium aurothiomalate, sodium aurothiosulphate and aurothioglucose to plasma proteins has been investigated in vitro. All gold compounds were bound to protein at a similar rate and had similar apparent affinity constants and binding capacities. Penicillamine (10 or 100 microgram/ml) was unable to chelate significant amounts of bound gold from plasma protein.

As used clinically, Myocrisin appears to contain largely polymeric (Autm)n, in which thiomalate (tm) sulphurs bridge between Au(I) ions, together with small amounts of a more reactive gold-bound thiomalate, and free thiomalate, as well as glycerol, unidentified yellow-products from autoclaving and phenylmercury adducts. Solganal usually contains thioglucose sulphinic acid as an impurity. Auranofin, on the other hand, has been crystallised. It is monomeric and Au(I) is almost linearly coordinated by P and S from triethylphosphine and tetraacetyl-beta-D-thioglucose, the latter adopting the chair conformation in the solid state and in solution. The major reaction of auranofin in acidic aqueous solutions appears to be hydrolysis of the sugar acetyl groups but other products arise if methanol is also present in the medium. NMR methods are used to examine in vitro the partition of auranofin between plasma and blood cells. The displacement of the thioglucose ligand and release of PEt3 from gold as OPEt3 are discussed.

A detailed description is given of a method by which gold can be visualized in frozen, paraffin and Epon sections. Histological sections from animals treated with gold compounds are exposed to UV-light from 30 min to several hours. The reduced, metallic gold is then visualized by means of a photographic developer containing silver lactate. Light- and electronmicroscope photographs showing gold in different organs from rats and mice treated with aurothioglucose, aurothiosulfate and aurothiomalate are presented.

Auranofin (AF; ' Ridaura '), an oral chrysotherapeutic agent, parenteral gold sodium thiomalate (GST) and gold thioglucose (GTG) were evaluated in order to compare their preclinical profiles. AF was found to be more effective than GST and GTG in suppressing inflammation and stimulating cell-mediated immunity. In contrast to GST, AF inhibited cellular release of lysosomal enzymes, antibody-dependent cellular cytotoxicity, production of antibodies in adjuvant arthritic rats, and antibodies involved in cytotoxicity reactions. In pharmacokinetic studies, plasma gold in rats following AF administration, exhibited greater cell association than after GST administration. In conclusion, the pharmacological profile of AF is markedly different from those of GST and GTG and this suggests potential for improvements in chrysotherapy.

Double isotope-labelled aurothiomalate (195Au-14C-thiomalate) has been administered to mice, and the excretory fate and tissue distribution have been studied. The results show that the gold and the thiomalate separate in vivo resulting in protein-bound gold and release of free thiomalate. About half of this thiol is excreted in the urine during the first day, and the remaining half is bound to tissue membranes and cells. Although thiomalate penetrates cellular membranes slowly in vitro. the compound is found in all organs, mostly in the liver and the kidneys, after administration of aurothiomalate. Separation of the gold moiety from its thiol carrier also takes place in man. This explains the finding of free thiomalate in the urine of patients receiving aurothiomalate intramuscularly. As thiomalate has now been shown to possess penicillamine-like biological activities it is suggested that at least part of the antirheumatic effects of aurothiomalate may be due to the thiol carrier being released in the body.

Gold concentrations in epidermis, dermis, and whole skin were measured by neutron activation analysis after formation of suction bullae in 8 patients who had received protracted cyrysotherapy. Epidermis contained 3% (median) of the gold content of whole skin. A direct correlation between cumulative gold dose and skin gold level was noted. These findings suggest that apparent gold concentrations in skin are influenced by the depth of the biopsy, that keratinous tissues have little affinity for gold, and that the gold storage capability of skin is not saturated by large cumulative doses of gold. The beneficial effect of gold in pemphigus may not be mediated at the site of blister formation.

Inhibition of adenosine triphosphatase (ATPase) by chlorauric acid (Au3+) and gold sodium thiomalate (Au+) was studied in dog brain and kidney and in human kidney enzyme preparations. Au3+ indiscriminately affected ouabain-sensitive (Na+ + K+-dependent) ATPase and ouabain-insensitive (Mg2+-dependent) ATPase with concentrations for 50% inhibition (I50) approximately 10(-6) M. The I50 of Au3+ for Na+ + K+ ATPase was several-fold higher in homogenates than in microsomal fractions. The enzyme was protected by bovine serum albumin. Although Au3+ and Au+ were equipotent against Mg2+ ATPase, Au+ inhibited Na+ + K+ ATPase 2 to 3 times more effectively than did Au3+. The inhibitory action of Au3+ (but not Au+) was potentiated by ascorbic acid, suggesting reduction of Au3+ to Au+ by ascorbic acid. The fractional inhibition of Na+ + K+ ATPase by Au3+ or Au+ was not affected by changing concentrations of NaCl, KCl, MgCl2, ATP, and MgATP. Decreasing pH from 8.0 to 6.8 enhanced both Au+ and Au3+ inhibition. We conclude that gold is one of the most potent nonspecific of Na+ + K+ ATPase, with characteristics differing from other metallic inhibitors of this enzyme system.

The mechanism of action of auranofin, an oral organic gold compound used in the treatment of rheumatoid arthritis, is probably similar to the previously available parenteral gold compounds. Auranofin affects polymorphonuclear cells and monocytes at lower concentrations than gold sodium thiomalate and generally affects humoral and cell-mediated immunity in the same direction as the latter drug. The pharmacokinetics of auranofin are different from the intramuscular gold compounds. Auranofin is 20-25% orally absorbed and has less total body retention, greater fecal excretion, and less urinary excretion than gold sodium thiomalate. This may be due in part to its differing chemistry, including its lipophilicity and monomeric structure (at least in vitro). While many clinical studies are not yet complete, auranofin (6 mg/day) is clearly more effective than placebo for treating rheumatoid arthritis. Its efficacy relative to gold sodium thiomalate is not clear. Auranofin may be slightly less effective than gold sodium thiomalate, but because it is generally less toxic than intramuscular gold compounds, its therapeutic index may be more favorable.

This study shows that after intra-articular injection, aurothiomalate and colloidal gold of small (200 A) particle size were rapidly absorbed from joints while the larger, 300 A, particle size colloidal radioactive gold could not be found outside them. Larger particle size suspensions seem therefore more likely to remain localised in the joint and its lining synovium after intra-articular injection, the systemic absorption from the joint cavity diminishing with increasing particle size. It was also found that the intra-articular injection of small amounts of aurothiomalate, of colloidal gold and of colloidal radioactive gold produces identical degenerative lesions in the lining cells of the proximal convoluted tubules of the kidneys. These lesions were always found, although gold particles were demonstrated only in sampled kidney tissues of the animals injected with the soluble gold preparation whereas no gold could be detected in the tissues of animals injected with colloidal non-radioactive or radioactive gold. Electron microscopic evidence is presented to suggest the possibility that the mitochondria are the "target" organelles of the gold-induced cellular damage. Mitochondrial damage was demonstrated in liver and spleen in addition to the already described kidney damage. The correlation between structure and function of the mitochondrial changes is not clear, and ionic shifts may be both a cause and a result of damage.

A comparative study of tissue gold levels produced in guinea-pigs after the oral administration of either triethylphosphine gold chloride or Myocrisin (sodium aurothiomalate) or after the injection of Myocrisin intramuscularly is reported. Gold concentrations were determined 5, 24 and 168 hours after administration in stomach, small intestine, large intestine, kidney, liver and spleen and 5 and 24 hours after administration in skin, adrenals, heart, lung and brain. In gastrointestinal tissues, tissue gold concentrations were highest with triethylphosphine gold chloride. The stomach gold level 5 hours after oral administration of triethylphosphine gold chloride is particularly high and, taken in conjunction with the other gastrointestinal gold levels measured, suggests that a stomach rather than an intestinal absorption mechanism may predominate. A more extensive time-course study on kidney and liver is reported and the possible relationship between tissue concentration and toxicity is discussed.

The ultrastructural localization of gold in the anterior pituitary of rats injected intraperitoneally with sodium aurothiomalate has been demonstrated using a histochemical technique that visualizes minute traces of gold. Gold was located intracellularly within lysosome-like bodies of marginal layer cells, follicular cells, and thyrotrophs. Gold was not seen within secretion granules. The possible mechanism by which gold is taken up into the cells is discussed.

The metabolites of gold in the urine of rats given the antiarthritic drug aurothiomalate were investigated by gel permeation chromatography, electrophoresis, and chemical studies. Following a single dose of aurtothiomalate, the excreted gold was protein-bound in the high-molecular-weight (greater than or equal to 150,000 dalton) and serum albumin fractions. Electrophoresis confirmed the presence of albumin, but showed that the other proteins present differ from those in normal or in vitro aurothiomalate-incubated rat sera. The pattern of the proteins establishes that the proteinuria was of the glomerular type. The alterations in the gold distribution produced by incubation of the urine with the low-molecular-weight thiol penicillamine and with exogenously added aurothiomalate indicated the existence of a labile equilibrium of gold among protein binding sites in the urine. Incubation of rat and human sera and commercially prepared serum albumins with aurothiomalate increased the electrophoretic mobility of the albumin. The significance of this change in electrophoretic mobility with respect to two models of gold binding by serum albumin is discussed.

The acetylator phenotype was determined in 59 patients with classical, seropositive and erosive rheumatoid arthritis (RA) treated with sodium-aurothiomalate or d-penicillamine. Patients with adverse drug reactions (ADR) leading to drug withdrawal (n = 29) were compared with a group of patients without ADR (n = 30). The frequency of slow acetylators was significantly (p less than 0.05) increased in all RA patients, irrespective of the presence of ADR, particularly in the male patients, compared with a control population. No association was found between acetylator phenotype and clinical data or secondary Sjögrens's syndrome (SS).

A case of classical RA developed severe interstitial pulmonary disease and respiratory failure while on chrysotherapy. A high concentration of gold was found in lung tissue. Electron microscopic and electron probe examinations confirmed the presence of gold in the interstitial and intra-alveolar macrophages. The clinical course and possible pathogenic mechanism were discussed.

Rat liver and kidney tissue uptake of gold and its localization in the cytosol was studied following various doses of gold sodium thiomalate (GST). The timecourse of gold incorporation into intracellular gold-binding ligands following repeated injections of GST was also investigated (11 injections, one dose/week). Results show that between 30 and 60% of the hepatic and renal gold was localized in the cytosol over a wide range of GST doses. This was also true following repeated doses. In the kidney, the binding of gold to high molecular weight (HMW) proteins was saturated after the third GST dose, while incorporation into the metallothioneins (MT) continued to increase, accounting for as much as 50% of cytosolic gold. On the other hand the binding to hepatic MT was about 10x lower, and the proportion of cytosolic gold incorporated into the MT, decreased from 30% (after first 3 GST injections) to about 15% (following the last 3 injections). The results show that the stimulation of MT biosynthesis in different tissues as a response to the injected GST is not the same and varies within each organ with the dose and/or the duration of repeated exposure. In the liver, the ability of gold to induce MT synthesis was limited and the importance of MT in the cellular uptake and disposition of gold may largely be confined to the kidneys. It is suggested that besides playing a possible role in the detoxification of cellular gold, particularly in the kidney, MT may also contribute towards the retention and localization of gold in the tissues.

The pharmacokinetics of gold sodium thiomalate (GST) and triethylphosphine gold (auranofin; AF) are different. Gold sodium thiomalate (GST) is completely bioavailable while only 15-25% of auranofin (AF) is absorbed. Protein binding of AF occurs to a larger extent to macroglobulins than does GST and total body retention of GST is much greater than AF at six months (30% versus approximately 1%). While terminal serum half-lives are approximately equal, total body half-lives are 250 days for GST and 69 days for AF. In addition, excretory pathways contrast markedly, with 85% of AF appearing in the feces while only 30% of GST is excreted by this route; 15% of AF gold appears in the urine and approximately 70% of GST gold is excreted via this route. With all the above differences one would expect that organ and cellular distribution of these compounds would differ. While gold from both drugs is concentrated in kidney, the percent of the dose found in the kidneys is less for AF than GST, at least in animals (0.4% vs 4.8%). Minute quantities are found in other organs but more study is needed to more clearly define organ distribution of these gold compounds, particularly in man.