Arsenite induces tissue factor synthesis through Nrf2 activation in cultured human aortic smooth muscle cells.

Tissue factor (TF) is the initiator of the coagulation cascade, constitutively expressed in subendothelial cells such as vascular smooth muscle cells and initiating rapid coagulation when the vascular vessel is damaged. TF has been shown to be involved in the development and progression of atherosclerosis. Arsenic, an environmental pollutant, is related to the progression of atherosclerosis, although the pathogenic mechanisms are not fully elucidated. In the present study, we investigated the effect of arsenite on the expression of TF in human aortic smooth muscle cells (HASMCs) and the underlying molecular mechanisms. We found that (1) arsenite stimulated TF synthesis and activated the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway in HASMCs, (2) sulforaphane, an Nrf2 activator, also stimulated TF synthesis in HASMCs, and (3) arsenite-induced upregulation of TF synthesis was prevented by Nrf2 knockdown in HASMCs. These results suggest that arsenite promotes TF synthesis by activating the Nrf2 pathway in HASMCs and that the induction of TF expression by arsenite may be related to the progression of atherosclerosis.

Comparison of the toxicity of sulfur mustard and its oxidation products in vitro.

The molecular toxicology of the chemical warfare agent sulfur mustard (SM) is still not completely understood. It has been suggested that in addition to SM itself also biotransformation products thereof mediate cytotoxicity. In the current study, we assessed this aspect by exposing a human hepatocyte cell line (HepG2) to SM or to its oxidation products sulfur mustard sulfoxide (SMO), sulfur mustard sulfone (SMO2), and divinyl sulfone (DVS). Cytotoxicity, determined with the XTT assay, revealed a significant higher toxicity of SMO2 and DVS compared to SM while SMO had no effect at any concentration. The exact biotransformation of SM leading to SMO, SMO2 and finally DVS is unknown so far. Involvement of the CYP450 system is discussed and was also investigated in the presented study. Modulation of CYP1A2 activity, taken as a model enzyme for CYP450, affected cytotoxicity of SM, SMO2 or DVS significantly. Induction of CYP1A2 with omeprazole led to decreased cytotoxicity for all compounds whereas inhibition with cimetidine resulted in an increased cytotoxicity for SM, but not for SMO2 and DVS. Our results indicate a distinctive role of the CYP450 system in SM poisoning. Future studies should address the metabolic conversion of SM in more detail. Our data may suggest the well-tolerated drug omeprazole as a potential co-treatment after contact to SM.

Design, Synthesis and Antiplasmodial Evaluation of Sulfoximine-triazole Hybrids as Potential Antimalarial Prototypes.

BACKGROUND: Malaria, caused by the deadly Plasmodium falciparum strain, claims the lives of millions of people annually. The emergence of drug-resistant strains of P. falciparum to the artemisinin-based combination therapy (ACT), the last line of defense against malaria, is worrisome and urges for the development of new chemo-types with a new mode of action. In the search of new antimalarial agents, hybrids of triazoles and other known antimalarial drugs have been reported to possess better activity than either of the parent compounds administered individually. Despite their better activity, no hybrid antimalarial drugs have been developed so far. OBJECTIVE: In the hope of developing new antimalarial prototypes, we propose the design, synthesis and antimalarial evaluation of novel sulfoximine-triazole hybrids owing to their interesting biological and physiological properties. METHODS: The sulfoximine part of the hybrid will be synthesized via imidation of the corresponding sulfoxide. Propargylation of the NH moiety of the sulfoximine followed by copper-catalyzed click chemistry with benzyl azide was envisaged to provide the target sulfoximine-triazole hybrids. RESULTS: Five novel sulfoximine-triazole hybrids possessing various substituents on the sulfoximine moiety have been successfully synthesized and evaluated for their antiplasmodial and cytotoxicity activities. The results revealed that the co-presence of the sulfoximine and triazole moieties along with a lipophilic alkyl substituent on the sulfur atom impart significant activity. CONCLUSION: Sulfoximine-triazole hybrids could be used as a prototype for the synthesis of new derivatives with better antiplasmodial activities.

In Vitro Drug-Drug Interaction Potential of Sulfoxide and/or Sulfone Metabolites of Albendazole, Triclabendazole , Aldicarb, Methiocarb, Montelukast and Ziprasidone.

BACKGROUND: The use of polypharmacy in the present day clinical therapy has made the identification of clinical drug-drug interaction risk an important aspect of drug development process. Although many drugs can be metabolized to sulfoxide and/or sulfone metabolites, seldom is known on the CYP inhibition potential and/or the metabolic fate for such metabolites. OBJECTIVE: The key objectives were: a) to evaluate the in vitro CYP inhibition potential of selected parent drugs with sulfoxide/sulfone metabolites; b) to assess the in vitro metabolic fate of the same panel of parent drugs and metabolites. METHODS: In vitro drug-drug interaction potential of test compounds was investigated in two stages; 1) assessment of CYP450 inhibition potential of test compounds using human liver microsomes (HLM); and 2) assessment of test compounds as substrate of Phase I enzymes; including CYP450, FMO, AO and MAO using HLM, recombinant human CYP enzymes (rhCYP), Human Liver Cytosol (HLC) and Human Liver Mitochondrial (HLMit). All samples were analysed by LC-MS-MS method. RESULTS: CYP1A2 was inhibited by methiocarb, triclabendazole, triclabendazole sulfoxide, and ziprasidone sulfone with IC50 of 0.71 µM, 1.07 µM, 4.19 µM, and 17.14 µM, respectively. CYP2C8 was inhibited by montelukast, montelukast sulfoxide, montelukast sulfone, tribendazole, triclabendazole sulfoxide, and triclabendazole sulfone with IC50 of 0.08 µM, 0.05 µM, 0.02 µM, 3.31 µM, 8.95 µM, and 1.05 µM, respectively. CYP2C9 was inhibited by triclabendazole, triclabendazole sulfoxide, triclabendazole sulfone, montelukast, montelukast sulfoxide and montelukast sulfone with IC50 of 1.17 µM, 1.95 µM, 0.69 µM, 1.34 µM, 3.61 µM and 2.15 µM, respectively. CYP2C19 was inhibited by triclabendazole and triclabendazole sulfoxide with IC50 of 0.25 and 0.22, respectively. CYP3A4 was inhibited by montelukast sulfoxide and triclabendazole with IC50 of 9.33 and 15.11, respectively. Amongst the studied sulfoxide/sulfone substrates, the propensity of involvement of CY2C9 and CYP3A4 enzyme was high (approximately 56% of total) in the metabolic fate experiments. CONCLUSION: Based on the findings, a proper risk assessment strategy needs to be factored (i.e., perpetrator and/or victim drug) to overcome any imminent risk of potential clinical drug-drug interaction when sulfoxide/sulfone metabolite(s) generating drugs are coadministered in therapy.

Determination of the impurities in drug products containing montelukast and in silico/in vitro genotoxicological assessments of sulfoxide impurity.

Impurities affecting safety, efficacy, and quality of pharmaceuticals are of increasing concern for regulatory agencies and pharmaceutical industries, since genotoxic impurities are understood to play important role in carcinogenesis. The study aimed to analyse impurities of montelukast chronically used in asthma theraphy and perform genotoxicological assessment considering regulatory approaches. Impurities (sulfoxide, cis-isomer, Michael adducts-I&II, methylketone, methylstyrene) were quantified using RP-HPLC analysis on commercial products available in Turkish market. For sulfoxide impurity, having no toxicity data and found to be above the qualification limit, in silico mutagenicity prediction analysis, miniaturized bacterial gene mutation test, mitotic index determination and in vitro chromosomal aberration test w/wo metabolic activation system were conducted. In the analysis of different batches of 20 commercial drug products from 11 companies, only sulfoxide impurity exceeded qualification limit in pediatric tablets from 2 companies and in adult tablets from 7 companies. Leadscope and ToxTree programs predicted sulfoxide impurity as nonmutagenic. It was also found to be nonmutagenic in Ames MPF Penta I assay. Sulfoxide impurity was dose-dependent cytotoxic in human peripheral lymphocytes, however, it was found to be nongenotoxic. It was concluded that sulfoxide impurity should be considered as nonmutagenic and can be classified as ordinary impurity according to guidelines.

Tetrahydroindazoles as Interleukin-2 Inducible T-Cell Kinase Inhibitors. Part II. Second-Generation Analogues with Enhanced Potency, Selectivity, and Pharmacodynamic Modulation in Vivo.

The medicinal chemistry community has directed considerable efforts toward the discovery of selective inhibitors of interleukin-2 inducible T-cell kinase (ITK), given its role in T-cell signaling downstream of the T-cell receptor (TCR) and the implications of this target for inflammatory disorders such as asthma. We have previously disclosed a structure- and property-guided lead optimization effort which resulted in the discovery of a new series of tetrahydroindazole-containing selective ITK inhibitors. Herein we disclose further optimization of this series that resulted in further potency improvements, reduced off-target receptor binding liabilities, and reduced cytotoxicity. Specifically, we have identified a correlation between the basicity of solubilizing elements in the ITK inhibitors and off-target antiproliferative effects, which was exploited to reduce cytotoxicity while maintaining kinase selectivity. Optimized analogues were shown to reduce IL-2 and IL-13 production in vivo following oral or intraperitoneal dosing in mice.

Assessment of hematological profiles of adult male athletes from two different air pollutant zones of West Bengal, India.

Health effects from air pollution are severe concern of today's world. The study was undertaken to assess the effects of air pollution on hematological profiles of trained and untrained males of West Bengal. The sample consisted of 60 sprinters, 60 footballers, and 120 untrained males, subdivided into two groups from two zones, namely, Tollygunge and Sonarpur. Suspended particulate matter (SPM), respirable particulate matter (RPM), oxides of sulfur (SOx), and oxides of nitrogen (NOx) of ambient air were monitored for both zones. Height and weight of all the subjects were measured. Venous blood sample was drawn from the cubital vein, and the red blood cell count (TC), packed cell volume (PCV), hemoglobin (Hb) concentration, mean corpuscular volume (MCV), and mean corpuscular hemoglobin concentration (MCHC) were determined by standard methods. Results revealed that SPM, RPM, SOx, and NOx concentrations were significantly higher in the Tollygunge area than Sonarpur. TC, PCV, and Hb concentration of untrained males were significantly higher than footballers in both regions but no significant difference were observed when compared with sprinters, except the Hb concentration in the Tollygunge zone. On the other hand, all hematological parameters of both trained and untrained males were significantly higher in the Sonarpur area than Tollygunge. It was concluded that environmental air pollutants might influence hematological profile adversely both in trained and sedentary males. However, further investigation in this area is needed.

Development of PBPK model of molinate and molinate sulfoxide in rats and humans.

Molinate has been widely used as a pre-emergent herbicide in the rice fields of California's Central Valley. In rat studies, the metabolite molinate sulfoxide is suspected of causing testicular toxicity after exposure to molinate. The sulfoxide is generated in the liver and can circulate in the blood, eventually reaching the testis. Man qualitatively produces the same molinate metabolites as the rat. To extrapolate the reproductive risk to man, the present study outlines the development of a preliminary PBPK (physiologically-based pharmacokinetic) model, validation in the rat and extrapolation to man. The preliminary seven-compartment PBPK model for molinate was constructed for the adult, male Sprague-Dawley rat that employed both flow-limited (blood, kidney, liver, rapid-perfused tissues and slowly perfused tissues) and diffusion-limited (fat) rate equations. The systemic circulation connects the various compartments. The simulations predict the molinate blood concentrations of the rat blood and testes compartment favorably with the profiles obtained from 10 and 100mg/kg po or 1.5 and 15mg/kg iv doses. Human physiological parameters were substituted into the oral dosed model and the simulations closely predicted the molinate blood concentration obtained from 5.06mg oral dose. A sensitivity analysis determined for an oral dose that peak blood molinate concentrations were most responsive to the blood flows to kidney and fat compartments while testicular molinate sulfoxide concentrations depended on molinate sulfoxide partition coefficients for the testes compartment and the K(m) for glutathione conjugation of molinate sulfoxide in the liver compartment.

Identification and quantitation of the N-acetyl-L-cysteine S-conjugates of bendamustine and its sulfoxides in human bile after administration of bendamustine hydrochloride.

We recently reported the detection of mercapturic acid pathway metabolites of bendamustine, namely, cysteine S-conjugates in human bile, which are supposed to subsequently undergo further metabolism. In this study, we describe the identification and quantitation of consecutive bendamustine metabolites occurring in human bile using authentic reference standards and the synthesis and structural confirmation of these compounds. Mass spectrometry data along with high-performance liquid chromatography retention data (fluorescence detection) of the synthetic reference standards were consistent with those of the metabolites found in human bile after administration of bendamustine hydrochloride to cancer patients. Analysis of the purified synthetic reference compounds showed a purity of at least 95%. Structural confirmation was achieved by one- and two-dimensional proton as well as carbon-13 NMR spectroscopy and mass spectrometry. A total of 16 bendamustine-related compounds were detected in the bile of patients, 11 of them were recovered as conjugates. Eight conjugates have been structurally confirmed as novel mercapturic acids and sulfoxides. Biliary excretion of the sulfoxides was twice that of the mercapturate precursors. Glutathione S-conjugates of bendamustine have not been detected in bile samples, indicating rapid enzymatic cleavage in humans. Both the lack of glutathione (GSH) conjugates and occurrence of diastereomeric sulfoxides emphasize species-related differences in the GSH conjugation of bendamustine between humans and rats. The total amount recovered in the bile as the sum of all conjugates over the period of 24 h after dosing averaged 5.2% of the administered dose. The question of whether the novel metabolites contribute to urinary excretion should be a target of future investigations.

Formation of three N-acetyl-L-cysteine monoadducts and one diadduct by the reaction of S-(1,2-dichlorovinyl)-L-cysteine sulfoxide with N-acetyl-L-cysteine at physiological conditions: chemical mechanisms and toxicological implications.

Previously, our laboratory has shown that S-(1,2-dichlorovinyl)-L-cysteine sulfoxide (DCVCS), a Michael acceptor produced by a flavin-containing monooxygenase 3 (FMO3)-mediated oxidation of S-(1,2-dichlorovinyl)-L-cysteine (DCVC), is a more potent nephrotoxicant than DCVC. In the present study, we characterized reactions of DCVCS with nucleophilic amino acids. DCVCS incubations with N-acetyl-L-cysteine (NAC) at pH 7.4 and 37 degrees C for 1 h resulted in the formation of three monoadducts and one diadduct characterized by LC/MS, 1H NMR, and 1H-detected heteronuclear single quantum correlation. The formation of all adducts (with relative ratios of 29, 31, 24, and 12%, respectively) was rapid and time-dependent; the half-lives of the two DCVCS diastereomers in the presence of NAC were 13.8 (diastereomer I) and 9.4 min (diastereomer II). Adducts 1 and 2 were determined to be diastereomers of S-[1-chloro-2-(N-acetyl-L-cystein- S-yl)vinyl]-L-cysteine sulfoxide formed by Michael addition of NAC to the terminal vinylic carbon of DCVCS followed by loss of HCl. Adduct 4 was determined to be S-[2-chloro-2-(N-acetyl-L-cystein- S-yl)vinyl]-L-cysteine sulfoxide formed from the initial Michael addition product followed by a less favorable loss of HCl and/or by a rearrangement of adduct 2 through the formation of a cyclic chloronium ion. The addition of another molecule of NAC to monoadducts 1, 2, or 4 resulted in the formation of the novel diadduct, S-[2,2-( N-acetyl-L-cystein-S-yl)vinyl]-L-cysteine sulfoxide (adduct 3), whose detection in relatively large amount suggests that DCVCS could act as a cross-linking agent. DCVCS was not reactive with N-acetyl-L-lysine or L-valinamide at similar incubation conditions. Collectively, the results suggest selective reactivity of DCVCS toward protein sulfhydryl groups. Furthermore, the cross-linking properties of DCVCS may in part explain its high nephrotoxic potency.

S-(1,2,2-trichlorovinyl)-L-cysteine sulfoxide, a reactive metabolite of S-(1,2,2-Trichlorovinyl)-L-cysteine formed in rat liver and kidney microsomes, is a potent nephrotoxicant.

Previously, we have provided evidence that cytochromes P450 (P450s) and flavin-containing monooxygenases (FMOs) are involved in the oxidation of S-(1,2,2-trichlorovinyl)-L-cysteine (TCVC) in rabbit liver microsomes to yield the reactive metabolite TCVC sulfoxide (TCVCS). Because TCVC is a known nephrotoxic metabolite of tetrachloroethylene, the nephrotoxic potential of TCVCS in rats and TCVCS formation in rat liver and kidney microsomes were investigated. At 5 mM TCVC, rat liver microsomes formed TCVCS at a rate nearly 5 times higher than the rate measured with rat kidney microsomes, whereas at 1 mM TCVC only the liver activity was detectable. TCVCS formation in liver and kidney microsomes was dependent upon the presence of NADPH and was inhibited by the addition of methimazole or 1-benzylimidazole, but not superoxide dismutase, catalase, KCN, or deferoxamine, consistent with the involvement of both FMOs and P450s. Rats given TCVCS at 230 micromol/kg i.p. exhibited acute tubular necrosis at 2 and 24 h after treatment, and they had elevated blood urea nitrogen levels at 24 h, whereas TCVC was a much less potent nephrotoxicant than TCVCS. Furthermore, pretreatment with aminooxyacetic acid enhanced TCVC toxicity. In addition, reduced nonprotein thiol concentrations in the kidney were decreased by nearly 50% 2 h after TCVCS treatment compared with saline-treated rats, whereas the equimolar dose of TCVC had no effect on kidney nonprotein thiol status. No significant lesions or changes in nonprotein thiol status were observed in liver with either TCVC or TCVCS. Collectively, the results suggest that TCVCS may play a role in TCVC-induced nephrotoxicity.

Role of cytochrome P4503A in cysteine S-conjugates sulfoxidation and the nephrotoxicity of the sevoflurane degradation product fluoromethyl-2,2-difluoro-1-(trifluoromethyl)vinyl ether (compound A) in rats.

The volatile anesthetic sevoflurane is degraded to fluoromethyl-2,2-difluoro-1-(trifluoromethyl)vinyl ether (FDVE) in anesthesia machines. FDVE is nephrotoxic in rats. FDVE undergoes glutathione conjugation, subsequent conversion to cysteine and mercapturic acid conjugates, and cysteine conjugate metabolism by renal beta-lyase, which is a bioactivation pathway mediating nephrotoxicity in rats. Recent in vitro studies revealed cytochrome P4503A-catalyzed formation of novel sulfoxide metabolites of FDVE cysteine-S and mercapturic acid conjugates in rat liver and kidney microsomes. FDVE-mercapturic acid sulfoxides were more toxic than other FDVE conjugates to renal proximal tubular cells in culture. Nevertheless, the occurrence and toxicological significance of FDVE sulfoxides formation in vivo remain unknown. This investigation determined, in rats in vivo, the existence, role of P4503A, and nephrotoxic consequence of FDVE conjugates sulfoxidation. Rats were pretreated with dexamethasone, phenobarbital, troleandomycin, or nothing (controls) before FDVE, and then, nephrotoxicity, FDVE-mercapturate sulfoxide urinary excretion, and FDVE-mercapturate sulfoxidation by liver microsomes were assessed. The formation of FDVE-mercapturic acid sulfoxide metabolites in vivo and their urinary excretion were unambiguously established by mass spectrometry. Dexamethasone and phenobarbital increased, and troleandomycin decreased (i) liver microsomal FDVE-mercapturic acid sulfoxidation in vitro, (ii) FDVE-mercapturic acid sulfoxide urinary excretion in vivo, and (iii) FDVE nephrotoxicity in vivo assessed by renal histology, blood urea nitrogen concentrations, and urine volume and protein excretion. Urine 3,3,3-trifluoro-2-(fluoromethoxy)propanoic acid, reflecting beta-lyase-dependent FDVE-cysteine S-conjugates metabolism, was minimally affected by the pretreatments. These results demonstrate that FDVE S-conjugates undergo P4503A-catalyzed sulfoxidation in rats in vivo, and this sulfoxidation pathway contributes to nephrotoxicity. FDVE S-conjugates sulfoxidation constitutes a newly discovered mechanism of FDVE bioactivation and toxicification in rats, in addition to beta-lyase-catalyzed metabolism of FDVE-cysteine S-conjugates.

Phototransformation of carboxin in water. Toxicity of the pesticide and its sulfoxide to aquatic organisms.

Sunlight exposure of aqueous suspensions of carboxin (1) causes its phototransformation to sulfoxide 2 and minor components. Similar effects are observed in the presence of humic acid or nitrate or at different pH values. Photoproducts 2-9 were isolated by chromatographic techniques and/or identified by spectroscopic means. Carboxin 1 and its main photoproduct sulfoxide 2 were tested to evaluate acute toxicity to primary consumers typical of the aquatic environment: the rotifer Brachionus calyciflorus and two crustaceans, Daphnia magna and Thamnocephalus platyurus. Chronic tests comprised a producer, the alga Pseudokirchneriella subcapitata, and a consumer, the crustacean Ceriodaphnia dubia.

Fatty acid elongation is important in the activity of thiocarbamate herbicides and in safening by dichlormid.

The thiocarbamates, such as pebulate (S-propyl butyl (ethyl) thiocarbamate) are a well-established class of herbicides. They inhibit fatty acid elongation, which is necessary for the biosynthesis of constituents of surface waxes and suberin and this has been proposed to be important for their toxicity. In this study lipid metabolism was investigated in herbicide-treated barley (Hordeum vulgare) and a pernicious weed, wild oats (Avena ludoviciana), to test the hypothesis that inhibitory effects on fatty acid elongation could be counteracted by the safer, dichlormid. Pebulate and its sulphoxide derivative (thought to be the active metabolite in vivo) were tested against lipid metabolism in barley or wild oat shoots. In both plants there was a significant inhibition of very long chain fatty acid (VLCFA) synthesis at herbicide concentrations > or =25 micro M. The extent to which safener dichlormid could prevent the inhibition of VLCFA synthesis was different in the two species. Previous treatment of barley with dichlormid (N,N-diallyl-2,2-dichloroacetamide) enabled fatty acid elongation in the presence of pebulate or pebulate sulphoxide, but had no effect on wild oats. The effects on fatty acid elongation mimicked the differential safening action of dichlormid observed on shoot elongation and growth in the two species. These data provide further evidence that inhibition of VLCFA formation is important for the mechanism of action of thiocarbamates.

Protection from cytotoxic effects induced by the nitrogen mustard mechlorethamine on human bronchial epithelial cells in vitro.

The present study was undertaken to find potent molecules against the toxicity of nitrogen mustard mechlorethamine (HN2) on respiratory epithelial cells, using a human bronchial epithelial cell line (16HBE14o-) as an in vitro model. The compounds examined included inhibitors of poly(ADP-ribose) polymerase (PARP), sulfhydryl-group donors as nucleophiles, and iron chelators and inhibitors of lipid peroxidation as antioxidants. Their effectiveness was determined upon observance of metabolic dysfunction induced by HN2 following a 4-h exposure, using (3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) reduction and ATP-level assays as indicators. Moreover, the fluorescent probe, monobromobimane (mBBr), and 2',7'-dichlorofluorescin-diacetate (H2DCF-DA) were used to assess intracellular sulfhydryl and peroxide level modifications by flow cytometry, respectively, following a 3-h exposure. At last, cell death was assessed by flow cytometry using the propidium iodide (PI)-dye-exclusion assay following 24-h exposure. PARP inhibitors (niacinamide, 3-aminobenzamide, 6(5H)-phenanthridinone), and two sulfhydryl-group donors (N-acetylcysteine, WR-1065) were found to be effective in preventing HN2-induced metabolic dysfunction when added in immediate or delayed treatment with HN2. Only N-acetylcysteine, however, was found to prevent cell death induced by HN2, though it must be present at the time of the HN2 challenge. Flow cytometric measurements of intracellular sulfhydryl levels strongly suggested that N-acetylcysteine and WR-1065 are preventive in alkylation of cellular compounds, mainly by direct extracellular interaction with HN2. PARP inhibitors prevent secondary deleterious effects induced by HN2, considering metabolism dysfunction as the endpoint. Elsewhere, the oxidative stress appears to be a side effect in HN2 toxicity only upon considering the inefficiency of several antioxidants.

Biotransformation, excretion, and nephrotoxicity of the hexachlorobutadiene metabolite (E)-N-acetyl-S-(1,2,3,4, 4-pentachlorobutadienyl)-L-cysteine sulfoxide.

Hexachlorobuta-1,3-diene (HCBD) is nephrotoxic in rodents. Its toxicity is based upon a multistep bioactivation pathway. Conjugation with glutathione by glutathione S-transferases to form (E)-S-(1,2,3,4,4-pentachlorobutadienyl)-L-glutathione (PCBG), further processing to the corresponding cysteine S-conjugate, and finally processing to a reactive thioketene are thought to be responsible for the observed nephrotoxic effects. A novel metabolite, identified as (E)-N-acetyl-S-(1,2,3,4, 4-pentachlorobutadienyl)-L-cysteine sulfoxide (N-AcPCBC-SO), was described after administration of [14C]HCBD to male Wistar rats. This metabolite is formed by sulfoxidation of N-acetyl-S-(1,2,3,4, 4-pentachlorobutadienyl)-L-cysteine (N-AcPCBC) mediated by cytochrome P450 3A and has been found to be cytotoxic to proximal tubular cells in vitro without activation by beta-lyase. In rats, given HCBD in vivo, only one diastereomer of the sulfoxide is excreted; however, in rat hepatic microsomes two diastereomers, (R)- and (S)-N-AcPCBC-SO, are formed. This study focuses on the mechanisms responsible for this discrepancy and on a possible contribution of N-AcPCBC-SO to the nephrotoxicity of HCBD in vivo. (R,S)-N-AcPCBC-SO (1:1 mixture of both diastereomers) and N-acetyl-alpha-methyl-S-(1,2,3,4,4-pentachlorobutadienyl)-d, L-cysteine sulfoxide (alpha-Me-N-AcPCBC-SO) were administered iv to male and female Wistar rats (20, 40, and 80 micromol/kg of body weight). alpha-Me-N-AcPCBC-SO cannot be cleaved by cysteine conjugate beta-lyase even if alpha-Me-N-AcPCBC-SO is deacetylated by acylases. Excretion of gamma-glutamyltranspeptidase, protein, and glucose in the urine, indicative for kidney damage, and histopathological examination of the kidneys showed marked differences in the renal damage in male and female rats after application of N-AcPCBC-SO and alpha-Me-N-AcPCBC-SO. Necroses of the kidney tubules were only found in male, but not female, rats. Major sex-specific differences were observed in the elimination of sulfoxides; the (R)-isomer was excreted in a 5-10-fold excess to the (S)-isomer after application of (R,S)-N-AcPCBC-SO. After purification, both isomers were administered to male rats resulting in the urinary excretion of (R)-N-AcPCBC-SO after giving the (R)-isomer; treatment with (S)-N-AcPCBC-SO, however, revealed the formation of (S)-N-acetyl-S-(2-glycinylcystein-S-yl-1,3,4, 4-tetrachlorobutadienyl)-L-cysteine. The results show major sex-specific differences in the nephrotoxic potency of N-AcPCBC-SO and alpha-Me-N-AcPCBC-SO. However, both N-AcPCBC-SO and alpha-Me-N-AcPCBC-SO are nephrotoxic in males, suggesting the formation of a vinyl sulfoxide as an additional, beta-lyase-independent mechanism in HCBD-caused nephrotoxicity.

A new antitumor agent: methyl sulfonium perchlorate of echinomycin.

Newly modified-echinomycin such as S-methylated sulfonium perchlorate of echinomycin (1), monosulfoxide (2), disulfoxid (3) and sulfone (4) have been prepared and evaluated for in vitro biological activities of cytotoxicity against P388, B16 and SNU-16 as well as in vivo antitumor activity against murine leukemia P388 and melanoma B16.

Fasciola hepatica: disruption of the vitelline cells in vitro by the sulphoxide metabolite of triclabendazole.

The effects of the active sulphoxide metabolite of the fasciolicide triclabendazole (Fasinex, Ciba-Geigy) on the vitelline cells of Fasciola hepatica were determined in vitro by transmission electron microscopy using both intact flukes and tissue-slice material. At a triclabendazole concentration of 15 micrograms/ml the vitelline cells of intact flukes showed ultrastructural changes only after prolonged incubation periods (12-24 h). The changes observed were a swelling of the granular endoplasmic reticulum (GER) cisternae with decreased ribosomal covering in the intermediate-type cells and condensation of chromatin and disappearance of the nucleolus in the nucleus of the stem cell. Similar changes were evident more quickly (by 6 h) in whole flukes treated at the higher concentration of 50 micrograms/ml. The shell globule clusters were loosely packed in the intermediate type-2 cells, and the number of intermediate type-1 cells declined with more prolonged incubation. Disruption of the nurse-cell cytoplasm was also observed from 12 h onwards. After only 6 h incubation of tissue-slice material at 50 micrograms/ml, intermediate type-1 cells were absent, shell globule clusters in mature cells were loosely packed and the nurse-cell cytoplasm was badly disrupted. By 12 h the vitelline cells were vacuolated and grossly abnormal. The results are discussed in relation to postulated actions of triclabendazole against the microtubule component of the cytoskeleton and against protein synthesis in the fluke.

Roles of cysteine conjugate beta-lyase and S-oxidase in nephrotoxicity: studies with S-(1,2-dichlorovinyl)-L-cysteine and S-(1,2-dichlorovinyl)-L-cysteine sulfoxide.

Effects of S-(1,2-dichlorovinyl)-L-cysteine (DCVC) and its putative metabolite DCVC sulfoxide (DCVCO) on renal function in vivo and in vitro were investigated to assess the role of sulfoxidation in the mechanism of toxicity of cysteine S-conjugates. Both conjugates were potent nephrotoxicants in rats in vivo, but at equimolar doses, DCVCO produced greater renal injury (i.e., increases in blood urea nitrogen levels and anuria and more severe and widespread proximal tubular necrosis) than DCVC. Pretreatment of rats with aminooxyacetic acid (AOAA), a selective cysteine conjugate beta-lyase (beta-lyase) inhibitor, did not protect against DCVCO nephrotoxicity, whereas rats given DCVC and AOAA exhibited partial protection. These results suggest that in addition to cleavage by the beta-lyase, sulfoxidation by the cysteine conjugate S-oxidase (S-oxidase) may play a role in DCVC nephrotoxicity. In isolated rat kidney proximal tubular (PT) and distal tubular (DT) cells, both DCVC and DCVCO produced time- and concentration-dependent increases in the release of lactate dehydrogenase. Because DCVC was generally more toxic in PT cells and DCVCO was more toxic in DT cells, an attempt was made to correlate in vitro cytotoxicity with the cellular distribution of the beta-lyase and S-oxidase. The finding that beta-lyase activity exhibited a 2-fold higher Vmax/Km ratio in PT cells than in DT cells, the greater inhibition of both beta-lyase activity and DCVC toxicity by AOAA in PT cells than in DT cells and the lower (40%) S-oxidase activity in PT cells than in DT cells provide evidence for the importance of the beta-lyase in DCVC toxicity in PT cells. The finding that DCVCO was more toxic in DT cells than in PT cells and the inability of AOAA to protect DT cells from DCVC-induced cytotoxicity, however, provide further evidence for DCVC bioactivation by S-oxidase.(ABSTRACT TRUNCATED AT 250 WORDS)

Activity of a new oxadiazole compound, against experimental infections with Entamoeba histolytica and Giardia lamblia in animal models.

BTI 2286E(+/-)-E-3-(4-methylsulphinylstyryl)-1,2,4-oxadiazole has demonstrated potent amoebicidal activity in a single-dose treatment against Entamoeba histolytica infection in the livers of golden hamsters and the caeca of mice, hamsters and rats. It has intra luminal activity against Entamoeba criceti, a natural infection in golden hamster, and anti-giardial activity against Giardia lamblia infection in suckling mice. BTI 2286E is more potent than metronidazole in extra intestinal, intra luminal amoebiasis models and has significant advantages in that it is non-mutagenic in the Ames test and has a shorter duration of effective treatment.