Synthesis and Evaluation of Structurally Diverse C-2-Substituted Thienopyrimidine-Based Inhibitors of the Human Geranylgeranyl Pyrophosphate Synthase.

Novel analogues of C-2-substituted thienopyrimidine-based bisphosphonates (C2-ThP-BPs) are described that are potent inhibitors of the human geranylgeranyl pyrophosphate synthase (hGGPPS). Members of this class of compounds induce target-selective apoptosis of multiple myeloma (MM) cells and exhibit antimyeloma activity in vivo. A key structural element of these inhibitors is a linker moiety that connects their (((2-phenylthieno[2,3-d]pyrimidin-4-yl)amino)methylene)bisphosphonic acid core to various side chains. The structural diversity of this linker moiety, as well as the side chains attached to it, was investigated and found to significantly impact the toxicity of these compounds in MM cells. The most potent inhibitor identified was evaluated in mouse and rat for liver toxicity and systemic exposure, respectively, providing further optimism for the potential value of such compounds as human therapeutics.

Spatiotemporal evaluation of the mouse embryonic redox environment and histiotrophic nutrition following treatment with valproic acid and 1,2-dithiole-3-thione during early organogenesis.

Redox regulation during metazoan development ensures that coordinated metabolic reprogramming and developmental signaling are orchestrated with high fidelity in the hypoxic embryonic environment. Valproic acid (VPA), an anti-seizure medication, is known to increase markers of oxidation and also increase the risk of neural tube defects (NTDs) when taken during pregnancy. It is unknown, however, whether oxidation plays a direct role in failed neural tube closure (NTC). Spatial and temporal fluctuations in total glutathione (GSH) and total cysteine (Cys) redox steady states were seen during a 24 h period of CD-1 mouse organogenesis in untreated conceptuses and following exposure to VPA and the Nrf2 antioxidant pathway inducer, 1,2-dithiole-3-thione (D3T). Glutathione, glutathione disulfide (GSSG), and Cys, cystine (CySS) concentrations, measured in conceptal tissues (embryo/visceral yolk sac) and fluids (yolk sac fluid/amniotic fluid) showed that VPA did not cause extensive and prolonged oxidation during the period of NTC, but instead produced transient periods of oxidation, as assessed by GSH:GSSG redox potentials, which revealed oxidation in all four conceptal compartments at 4, 10, and 14 h, corresponding to the period of heartbeat activation and NTC. Other changes were tissue and time specific. VPA treatment also reduced total FITC-Ab clearance from the medium over 3 h, indicating potential disruption of nutritive amino acid supply. Overall, these results indicated that VPA's ability to affect cellular redox status may be limited to tissue-specific windows of sensitivity during the period of NTC. The safety evaluation of drugs used during pregnancy should consider time and tissue specific redox factors.

Effects of dibenzothiophene, a sulfur-containing heterocyclic aromatic hydrocarbon, and its alkylated congener, 2,4,7-trimethyldibenzothiophene, on placental trophoblast cell function.

Worldwide demand for petroleum products has resulted in increased oil and gas activities in many countries. Conventional and unconventional oil and gas extraction, production, and transport lead to increased levels of petroleum-derived polycyclic aromatic hydrocarbons (PAHs) in the environment. PAH exposure has profound effects on reproduction by affecting pathways involved in placental trophoblast cell function and impairing normal placental development and function-key contributors to reproductive success. However, other components found in petroleum and wastewaters from oil and gas extraction, including the sulfur-containing heterocyclic aromatic compounds such as dibenzothiophene (DBT) and its alkylated derivatives, may also impact reproductive success. The goal of this study was to examine the effect of exposure to DBT, a compound commonly detected in the environment, and one of its alkylated analogues, 2,4,7-trimethyldibenzothiophene (2,4,7-DBT), on steroidogenic and angiogenic pathways critical for mammalian development in placental trophoblast cells (HTR-8/SVneo cells). 2,4,7-DBT but not DBT increased estradiol output in association with increased tube-like formation (surrogate for angiogenesis). These changes in angiogenesis did not appear to be related to altered expression of the key placental angiogenic gene targets (ANGPTL4, VEGFA, and PGF). Neither compound showed a concentration related effect on progesterone synthesis or its receptor expression. Our results suggest that 2,4,7-DBT can disrupt key pathways important for placental trophoblast function and highlight the importance of determining the impact of exposure to both parent and alkylated compounds. Further, these data suggest that exposure to sulfur-containing heterocyclic aromatic compounds may lead to placental dysfunction and impact reproductive success at environmentally relevant levels.

Potential of the zebrafish model for the forensic toxicology screening of NPS: A comparative study of the effects of APINAC and methiopropamine on the behavior of zebrafish larvae and mice.

The increased diffusion of the so-called novel psychoactive substances (NPS) and their continuous change in structure andconceivably activity has led to the need of a rapid screening method to detect their biological effects as early as possible after their appearance in the market. This problem is very felt in forensic pathology and toxicology, so the preclinical study is fundamental in the approach to clinical and autopsy cases of difficult interpretation intoxication. Zebrafish is a high-throughput suitable model to rapidly hypothesize potential aversive or beneficial effects of novel molecules. In the present study, we measured and compared the behavioral responses to two novel neuroactive drugs, namely APINAC, a new cannabimimetic drug, and methiopropamine (MPA), a methamphetamine-like compound, on zebrafish larvae (ZL) and adult mice. By using an innovative statistical approach (general additive models), it was found that the spontaneous locomotor activity was impaired by the two drugs in both species: the disruption extent varied in a dose-dependent and time-dependent manner. Sensorimotor function was also altered: i) the visual object response was reduced in mice treated with APINAC, whereas it was not after exposure to MPA; ii) the visual placing responses were reduced after treatment with both NPS in mice. Furthermore, the visual motor response detected in ZL showed a reduction after treatment with APINAC during light-dark and dark-light transition. The same pattern was found in the MPA exposed groups only at the dark-light transition, while at the transition from light to dark, the individuals showed an increased response. In conclusion, the present study highlighted the impairment of spontaneous motor and sensorimotor behavior induced by MPA and APINAC administration in both species, thus confirming the usefulness of ZL as a model for a rapid behavioural-based drug screening.

D-A polymers for fluorescence/photoacoustic imaging and characterization of their photothermal properties.

NIR-II fluorescence imaging has great potential in diagnosis, but the quantum efficiency of contrast agents is an urgent problem to be solved. We synthesized two new multifunctional polymers, P-TT and P-DPP, with a tetrahedral C (sp3) and branched alkyl chains in the main chain, which were beneficial to obtain high quantum efficiency. P-TT and P-DPP showed absorption peaks of 686 nm and 763 nm, respectively, and fluorescence emission peaks of 1071 nm and 1066 nm, respectively. The photothermal effect of P-DPP can reach 52 °C, and the quantum yield reaches 1.5%, which was three times higher than that of nanotube fluorophores (quantum yield 0.4%). P-DPP is used for stable fluorescence imaging of blood vessels and photoacoustic imaging of nude mice, and successfully applied to phototherapy of nude mouse tumours.

Measured and predicted affinities of binding and relative potencies to activate the AhR of PAHs and their alkylated analogues.

Polycyclic aromatic hydrocarbons (PAHs) and their alkylated forms are important components of crude oil. Both groups of PAHs have been reported to cause dioxin-like responses, mediated by aryl hydrocarbon receptor (AhR). Thus, characterization of binding affinity to the AhR of unsubstituted or alkylated PAHs is important to understand the toxicological consequences of oil contamination on ecosystems. We investigated the potencies of major PAHs of crude oil, e.g., chrysene, phenanthrene and dibenzothiophene, and their alkylated forms (n=17) to upregulate expression of AhR-mediated processes by use of the H4IIE-luc transactivation bioassay. In addition, molecular descriptors of different AhR activation potencies among PAHs were investigated by use of computational molecular docking models. Based on responses of the H4IIE-luc in vitro assay, it was shown that potencies of PAHs were determined by alkylation in addition to the number and conformation of rings. Potencies of AhR-mediated processes were generally greater when a chrysene group was substituted, especially in 1-methyl-chrysene. Significant negative correlations were observed between the in vitro dioxin-like potency measured in H4IIE-luc cells and the binding distance estimated from the in silico modeling. The difference in relative potency for AhR activation observed among PAHs and their alkylated forms could be explained by differences among binding distances in the ligand binding domain of the AhR caused by alkylation. The docking model developed in the present study may have utility in predicting risks of environmental contaminants of which toxicities are mediated by AhR binding.

Toxicity of thiophenes from Echinops transiliensis (Asteraceae) against Aedes aegypti (Diptera: Culicidae) larvae.

Structureactivity relationships of nine thiophenes, 2,2': 5',2″-terthiophene (1), 2-chloro-4-[5-(penta-1,3-diyn-1-yl)thiophen-2-yl]but-3-yn-1-yl acetate (2), 4-(2,2'-bithiophen-5-yl)but-3-yne-1,2-diyl diacetate (3), 4-[5-(penta-1,3-diyn-1-yl)thiophen-2-yl]but-3-yne-1,2-diyl diacetate (4), 4-(2,2'-bithiophen-5-yl)-2-hydroxybut-3-yn-1-yl acetate (5), 2-hydroxy-4-[5-(penta-1,3-diyn-1-yl)thiophen-2-yl]but-3-yn-1-yl acetate (6), 1-hydroxy-4-[5-(penta-1,3-diyn-1-yl)thiophen-2-yl]but-3-yn-2-yl acetate (7), 4-(2,2'-bithiophen-5-yl)but-3-yne-1,2-diol (8), and 4-[5-(penta-1,3-diyn-1-yl)thiophen-2-yl]but-3-yne-1,2-diol (9), isolated from the roots of Echinops transiliensis, were studied as larvicides against Aedes aegypti. Structural differences among compounds 3, 5, and 8 consisted in differing AcO and OH groups attached to C(3″) and C(4″), and resulted in variations in efficacy. Terthiophene 1 showed the highest activity (LC50 , 0.16 µg/ml) among compounds 1-9, followed by bithiophene compounds 3 (LC50 , 4.22 µg/ml), 5 (LC50 , 7.45 µg/ml), and 8 (LC50 , 9.89 µg/ml), and monothiophene compounds 9 (LC50 , 12.45 µg/ml), 2 (LC50 , 14.71 µg/ml), 4 (LC50 , 17.95 µg/ml), 6 (LC50 , 18.55 µg/ml), and 7 (LC50 , 19.97 µg/ml). These data indicated that A. aegypti larvicidal activities of thiophenes increase with increasing number of thiophene rings, and the most important active site in the structure of thiophenes could be the tetrahydro-thiophene moiety. In bithiophenes, 3, 5, and 8, A. aegypti larvicidal activity increased with increasing number of AcO groups attached to C(3″) or C(4″), indicating that AcO groups may play an important role in the larvicidal activity.

An overview of the preclinical toxicity and potential carcinogenicity of sitaxentan (Thelin®), a potent endothelin receptor antagonist developed for pulmonary arterial hypertension.

Sitaxentan (Thelin®), an endothelin receptor antagonist with a long duration of action and high specificity for the endothelin receptor A subtype, was used to treat pulmonary arterial hypertension. It was withdrawn from the market due to an idiosyncratic risk of drug-induced liver injury identified from emerging clinical trial data and clinical case reports. The preclinical safety profile of sitaxentan is presented, including single- and repeat-dose toxicity in mice, rats, and dogs and carcinogenicity in mice and rats. Sitaxentan-related adverse effects included coagulopathy in rats and dogs, increased serum alkaline phosphatase activity in mice and dogs, and hepatic hypertrophy in all species. Decreased albumin, erythrocyte count, hemoglobin concentration and hematocrit, and increased coagulation times and liver weight were also noted. These effects generally occurred at systemic exposures (AUC(0-24)) that were substantially greater than those seen in humans. Twice-daily (vs. once daily) dosing resulted in increased toxicity, which correlated with increased trough plasma sitaxentan concentrations. Sitaxentan appeared to have a low potential for testicular and hepatic toxicity and was not carcinogenic. These studies suggested that sitaxentan would have a reasonable margin of safety when used as directed in humans and supported a positive benefit:risk assessment at the time of marketing approval.

Hepatic effects of duloxetine-I: non-clinical and clinical trial data.

OBJECTIVE: Review nonclinical and clinical trial data for hepatic effects of duloxetine. METHODS: Review studies of toxicology, metabolism, mitochondrial effects, and clinical trials. RESULTS: Nonclinical studies revealed no treatment-related transaminase elevations and no effects of duloxetine on mitochondrial beta-oxidation in rat hepatocytes. In patients with a normal baseline alanine transaminase (ALT), duloxetine was associated with elevated transaminases >3X ULN in about 1% of patients. ALT and aspartate transaminase values peaked at 8 weeks, alkaline phosphatase steadily increased to maximum value at Week 52 and mean total bilirubin values were not increased. Hepatic-related treatment-emergent adverse events were uncommon. Seven of 23,000 duloxetine- and 2/6000 placebo-treated patients met criteria for modified Hy's rule (significant elevation of both ALT and total bilirubin) but were complicated by contributing factors such as excessive alcohol consumption (n=3), gall stones, common bile duct calculus, hepatitis C, and liver adenocarcinoma (n=1 each). CONCLUSIONS: Duloxetine has an effect on the liver, manifested by transient, self-limiting transaminase elevations. Rare events characterized as hepatocellular injury, cholestatic injury, or mixed type of hepatic injury have been reported. The pattern of liver effects was different from that in laboratory animals.

Treatment of proximal deep-vein thrombosis with the oral direct factor Xa inhibitor rivaroxaban (BAY 59-7939): the ODIXa-DVT (Oral Direct Factor Xa Inhibitor BAY 59-7939 in Patients With Acute Symptomatic Deep-Vein Thrombosis) study.

BACKGROUND: An effective and safe oral anticoagulant that needs no monitoring for dose adjustment is urgently needed for the treatment of diseases that require long-term anticoagulation. Rivaroxaban (BAY 59-7939) is an oral direct factor Xa inhibitor currently under clinical development. METHODS AND RESULTS: This randomized, parallel-group phase II trial in patients with proximal deep-vein thrombosis explored the efficacy and safety of rivaroxaban 10, 20, or 30 mg BID or 40 mg once daily compared with enoxaparin 1 mg/kg BID followed by vitamin K antagonist. Each treatment was administered for 12 weeks. The primary efficacy end point was an improvement in thrombotic burden at day 21 (assessed by quantitative compression ultrasonography; > or = 4-point improvement in thrombus score) without recurrent symptomatic venous thromboembolism or venous thromboembolism-related death. The primary safety end point was major bleeding during 12 weeks of treatment. Outcomes were adjudicated centrally without knowledge of treatment allocation. The primary efficacy end point was achieved in 53 (53.0%) of 100, 58 (59.2%) of 98, 62 (56.9%) of 109, and 49 (43.8%) of 112 patients receiving rivaroxaban 10, 20, or 30 mg BID or 40 mg once daily, respectively, compared with 50 (45.9%) of 109 patients treated with enoxaparin/vitamin K antagonist. There was no significant trend in the dose-response relationship between rivaroxaban BID and the primary efficacy end point (P=0.67). Major bleeding was observed in 1.7%, 1.7%, 3.3%, and 1.7% of patients receiving rivaroxaban 10, 20, or 30 mg BID or 40 mg once daily, respectively. There were no major bleeding events with enoxaparin/vitamin K antagonist. CONCLUSIONS: Results of this proof-of-concept and dose-finding study support phase III evaluation of the orally active direct factor Xa inhibitor rivaroxaban, because efficacy and safety were apparent in the treatment of proximal deep-vein thrombosis across a 3-fold range of fixed daily dosing.

Antifeedant and toxicity effects of thiophenes from four Echinops species against the Formosan subterranean termite, Coptotermes formosanus.

Over 220 crude extracts from repositories generated from plants native to Greece and Kazakhstan were evaluated for termiticidal activity against the Formosan subterranean termite, Coptotermes formosanus Shiraki (Isoptera: Rhinotermitidae). Emerging from this screening effort were bioactive extracts from two Greek species (Echinops ritro L. and Echinops spinosissimus Turra subsp. spinosissimus) and extracts from two Kazakhstan species (Echinops albicaulis Kar. & Kir. and Echinops transiliensis Golosh.). Fractionation and isolation of constituents from the most active extracts from each of the four species has been completed, resulting in the isolation of eight thiophenes possessing varying degrees of termiticidal activity. 2,2':5',2"-Terthiophene and 5'-(3-buten-1-ynyl)-2,2'-bithiophene demonstrated 100% mortality against C. formosanus within 9 days at 1 and 2 wt% concentrations respectively. In addition, all but two of the eight compounds tested were significantly different from the solvent controls in the filter paper consumption bioassay.

Preclinical assessment of gastrooesophageal tolerance of the new antiosteoporotic drug strontium ranelate: an endoscopic study in monkeys.

This study was aimed at evaluating the digestive tolerance of the new antiosteoporotic drug, strontium ranelate, and to compare it to that of another strontium salt, strontium chloride (SrCl2). Strontium ranelate, SrCl2, or placebo were administered orally (capsules) to 3 groups of 2 male and 2 female cynomolgus monkeys (Macaca fascicularis) once a day for 7 days at a dose of 2 g/day, which is the recommended therapeutic dose in man. Endoscopic examination of the oesophagus, the stomach and the first part of the duodenum was performed on fasted animals approximately 3 hr after the first (Day 1) and last dosing (Day 7), and, on Day 8 and Day 14 in case of lesions on Day 7. Strontium ranelate did not induce any acute or subchronic toxic effect on the gastric mucosa, the oesophagus and the first part of the duodenum. On the contrary, acute and superficial damages were noted on all animals receiving SrCl2 such as haemorrhagic and erosive lesions (formation of an ulcer in one male and a marked congestive antritis in one female). These effects were reversible after cessation of treatment. The microscopic examination of biopsies sampled at the site of gastric lesions revealed moderate granulocyte infiltration, indicating a local irritating origin of the lesions. Strontium ranelate by oral route is safe for the gastric mucosa while SrCl2 induced superficial and reversible lesions.

Phototoxic activity of a thiophene polyacetylene from Leuzea carthamoides.

The thiophene polyacetylene (E)-1-[5-(hept-5-en-1,3-diynyl)-2-thienyl]ethan-1,2-diol, isolated from the roots of Leuzea carthamoides, showed phototoxic activity in the assay systems of histidine photo-oxidation, Artemia and Tubifex assays. The effects were compared with the standard photosensitizer xanthotoxin.

Comparison of the dark and light-induced toxicity of thio and seleno analogues of the thiopyrylium dye AA1.

2,6-Bis(4-anilino)-4-(4-N,N-dimethylanilino)thiopyrylium chloride (AA1) and -selenopyrylium chloride (AA1-Se) and 2,6-bis(4-anilino)-4-(4-N-morpholinophenyl)thiopyrylium chloride (1) and -selenopyrylium chloride (2) were prepared via the addition of 4-N,N-dimethylanilino magnesium bromide and 4-N-morpholinophenyl magnesium bromide to chalcogenopyranones 3 followed by treatment with HCl gas then water. Cellular uptake of these dyes varied from 12+/-3fmol/cell for AA1 to 150+/-40 fmol/cell for AA1-Se. upon exposure to 5 x 10(-5) M solutions of the dyes for 3 h. Exposure of cell cultures to 1.8 J/cm2) of 360-750-nm light following incubation with 1 x 10(-6) M of either AA1, 1, or 2 for 24h resulted in no significant additional phototoxicity while AA1-Se showed a significant (p < 0.05) reduction in cell viability from 81% to 46%. Thiopyrylium dyes AA1 and 1 showed significant dark toxicity relative to selenopyrylium dyes AA1-Se and 2, respectively. AA1 was the only one of the four dyes to show inhibition of whole-cell mitochondrial cytochrome c oxidase activity in the dark. Irradiation of whole cells or mitochondrial suspensions treated with AA1, AA1-Se, or 2 gave inhibition of mitochondrial cytochrome c oxidase activity. Studies of JC-1-efflux indicated that all four cationic dyes accelerated the loss of JC-1 from the mitochondria, which suggests that all four dyes target the mitochondria.

Effects of 3H-1,2-dithiole-3-thione, 1,4-phenylenebis(methylene)selenocyanate, and selenium-enriched yeast individually and in combination on benzo[a]pyrene-induced mutagenesis in oral tissue and esophagus in lacZ mice.

We have studied the effects of three chemopreventive agents alone or in binary combinations on benzo[a]pyrene (BaP)-induced mutagenesis in the oral cavity and esophagus of lacZ mice using galE(-) selection. The mice were fed diets supplemented with 1,4-phenylenebis(methylene)selenocyanate (p-XSC) at 2.5 and 10 ppm Se, selenium-enriched yeast (SeY) at 2.5 and 10 ppm Se, and 3H-1,2-dithiole-3-thione (D3T) at 65 and 250 ppm, for 6 weeks. Two weeks after the start of the dietary regimen, mice were gavaged with five doses of 125 mg/kg BaP over 2 weeks, and the experiment was terminated 2 weeks later. Mutagenesis was measured in tongue, other pooled oral tissues (OTs), and esophagus. In mice treated with BaP alone, mutagenesis in the above tissues was in the range of 21-32 mutants/10(5)pfu (ca. 6-10 background levels for the corresponding tissues). p-XSC modestly inhibited mutagenesis (10-33% inhibition) in all tissues, but statistical significance was only observed at the low dose in esophagus, and pooled OT. SeY was not inhibitory alone. Greater inhibitory effects were observed with D3T, and inhibition was statistically significant at the high dose in tongue and esophagus (ca. 33%). Two combinations of low doses of the inhibitors were tested, and the D3T + SeY mix was most effective, leading to statistically significant inhibition in all three tissues (ca. 30-40% inhibition). The mixture D3T + p-XSC was of similar effectiveness as the low dose of D3T alone. This study combined with those previously done in our laboratory demonstrates effectiveness of D3T and to a lesser extent, p-XSC in the inhibition of mutagenesis, and provides support for the use of certain combinations of inhibitors as a means to increase effectiveness and reduce the dose of chemopreventive agents.

Subchronic toxicity of benzothiophene on rats following dietary exposure.

The systemic and neurobehavioral effects of benzo[b]thiophene (routinely referred to as benzothiophene) were studied in rats following 13-wk oral exposure. Male (170 +/- 16 g) and female (146 +/- 12 g) Sprague-Dawley rats (10 animals per group) were fed diet containing 0.5, 5, 50, or 500 ppm benzothiophene for 13 wk. Control animals were given rat feed plus vehicle (corn oil) only. No clinical signs of toxicity and neurobehavioral effects were observed using screening tests that included cage-side observations, righting reflex, open field activities, and forelimb and hindlimb grip strength. Elevated serum aspartate aminotransferase activity and bilirubin level were observed in highest dose females. Except for a statistically significant decrease in hematocrit in the highest dose males, benzothiophene exerted no marked effects on hematological parameters. Benzothiophene exposure did not result in alterations in hepatic alkaline phosphatase activity, or the typical hepatic phase I (aniline hydroxylase, ethoxyresorufin O-deethylase, pentoxyresorufin O-dealkylase, aminopyrine N-demethylase) and phase II (UDP-glucuronosyltransferase, glutathione S-transferase) drug-metabolizing enzyme activities. No significant elevation in urinary ascorbic acid, protein, and N-acetylglucosaminidase activity was detected in the treated animals. Peribiliary fibrosis was the most significant histological change and occurred in the liver of females in the 50 and 500 ppm groups. Mild epithelial hyperplasia in the renal pelvis was detected in the majority of 5 and 50 ppm females, with epithelial hyperplasia in the urinary bladder observed in the 50 ppm females. In males, increased incidence and severity of mild binucleation of hepatocytes and mild thickening of the basement membrane in kidney cortex were observed at 500 ppm. Benzothiophene was not detected in the urine of high-dose animals at the termination of the experiment. Based on the kidney, hepatic, and hematocrit changes, the no-observed-adverse-effect level (NOAEL) in the diet was determined to be 0.5 ppm (0.04 mg/kg/d) for females and 50 ppm (3.51 mg/kg/d) for males.

A review of the occurrence, toxicity, and biodegradation of condensed thiophenes found in petroleum.

Condensed thiophenes comprise a significant portion of the organosulfur compounds in petroleum and in other products from fossil fuels. Dibenzothiophene (DBT) has served as a model compound in biodegradation studies for over two decades. However, until quite recently, few other organosulfur compounds were studied, and their fates in petroleum-contaminated environments are largely unknown. This paper presents a review of the types of organosulfur compounds found in petroleum and summarizes the scant literature on toxicity studies with condensed thiophenes. Reports on the biodegradation of benzothiophene, alkylbenzothiophenes, DBT, alkylDBTs, and naphthothiophenes are reviewed with a focus on the identification of metabolites detected in laboratory cultures. In addition, recent reports on quantitative studies with DBT and naphtho[2,1-b]thiophene indicate the existence of polar sulfur-containing metabolites that have escaped detection and identification.

Metal binding and resultant loss of phototoxicity of alpha-terthienyl: metal detoxification versus alpha-terthienyl inactivation.

The members of the plant family asteraceae are noted for their cosmopolitan distribution and their versatility is attributed to be largely due to their morphological adaptations. Interestingly members of this family are endowed with rich levels of secondary plant metabolites, many of which are photochemically active (Bakker et al. 1979; Kagan et al. 1989). The secondary plant metabolite alpha-terthienyl derived from the plant family asteraceae is among the new class of light activated insecticide. The photobiocidal effects associated with alpha-terthienyl in presence of sunlight and ultraviolet light (300-400 nm), has stimulated a great deal of interest in its toxic mechanism of action against a number of organisms including phytopathogenic fungi, nematodes and mosquito larvae. Trials under tropical conditions indicate a very high level of activity as a larvicide to mosquito. There is no cross resistance to this compound in malathion resistant mosquito larvae (Arnason et al. 1989). Even though many researchers feel that the phototoxicity of secondary plant metabolites has arisen independently many times in evolution as a defense mechanism, the physiological impact of such biologically active compounds in the plant producing them also should be addressed (Arnason et al. 1987). Moreover, the accumulation of secondary plant metabolites in the roots of many asteraceae members as is the case with alpha-terthienyl in marigold (Tagetes sp.) roots, hints towards their functional divergence. In this study, we demonstrate a potent heavy metal quenching activity of alpha-terthienyl.

Functional aspects of membrane folate receptors in human breast cancer cells with transport-related resistance to methotrexate.

Two methotrexate (MTX)-resistant human breast-cancer cell lines with impaired transport via the reduced folate carrier (RFC), one established in vitro (MTX(R)-ZR-75-1) and another inherently resistant (MDA-231), were adapted to grow in medium containing 2 nM folic acid. This induced the expression of previously undetectable membrane folate receptors (MFR) to levels of 8.2 and 2.3 pmol/10(7) cells, respectively. Polymerase chain reaction (PCR) quantitation revealed that MFR messenger-RNA levels of the isoform first described in human nasopharyngeal carcinoma KB cells (MFR-alpha) were increased in low-folate-adapted MTX(R)-ZR-75-1 cells, whereas placental transcripts (MFR-beta) coincided with MFR-alpha expression in low-folate (LF)-adapted MDA-231 cells. These cell lines were used to study the role of MFR in the uptake and growth-inhibitory effects of five different antifolates with varying affinities for MFR: N10-propargyl-5, 8-dideazafolic acid (CB3717) > 5,10-dideazatetra-hydrofolic acid (DDATHF) > N-5-[N-(3,4-dihydro-2-methyl-4-oxoquinazolin-6-methyl) -N-methyl-amino]-2-theonyl}-glutamic acid (ZD1694) >> MTX > edatrexate (EDX). Expression of MFR only slightly decreased the resistant phenotype for MTX, EDX, and ZD1694, suggesting that these drugs are not transported intracellularly to cytotoxic concentrations at these levels of MFR expression. On the other hand, both cell lines became from at least 180- to 400-fold more sensitive to growth inhibition by CB3717 and DDATHF, which may be correlated with their high affinity for MFR. These sensitivity/resistance profiles were largely similar following cell culture in medium containing 1 nM L-leucovorin, a folate with an affinity for MFR 10-fold lower than that of folic acid, the one exception being the increased sensitivity for ZD1694 seen in the LF-adapted cells with the highest level of MFR expression (MTX(R)-ZR-75-1). These results illustrate that the efficacy of MFR in mediating antifolate transport and cytotoxicity depends on their affinity for the folate antagonist, their degree of expression, and the levels of competing folates.