Hormonal action of plant derived and anthropogenic non-steroidal estrogenic compounds: phytoestrogens and xenoestrogens.

Herbivorous and omnivorous vertebrates have evolved in the presence of a variety of phytoestrogens, i.e., plant-derived compounds that can mimic, modulate or disrupt the actions of endogenous estrogens. Since the discovery of the estrus-inducing effects of some plant products in 1926, considerable effort has been devoted to the isolation and structural and pharmacological characterization of phytoestrogens. Recently, agricultural and industrial pollution has added anthropogenic estrogenic compounds to the list of environmental estrogens. Unlike phytoestrogens, these xenoestrogens tend to accumulate and persist in adipose tissue for decades and may cause long-lasting, adverse endocrine effects. Here we review the endocrine effects of known phytoestrogens and xenoestrogens with special emphasis on molecular structure-activity relationships. Phytoestrogens include flavonoids, isoflavonoids, chalcons, coumestans, stilbenes, lignans, ginsenosides and other saponins, as well as the recently discovered tetrahydrofurandiols. Fungal estrogenic compounds may enter the food chain via infested crops. Since some phytoestrogens have been shown to display organ-specific actions, pharmaceutical estrogen analogues with similar properties (selective estrogen receptor modulators, SERMs) are also discussed. Xenoestrogens include dichlorodiphenyltrichloroethane (DDT) and its metabolites, bisphenols, alkylphenols, dichlorophenols, methoxychlor, chlordecone, polychlorinated benzol derivatives (PCBs), and dioxins. While most of these compounds act through estrogen receptors alpha and beta, some of their effects may be mediated by other nuclear or membrane-bound receptors or receptor-independent mechanisms. Some might also interfere with the production and metabolism of ovarian estrogens. Better understanding of the molecular pharmacology of phyto- and xenoestrogens may result in the development of novel compounds with therapeutic utility and improved environmental protection.

Comparative characterization of experimental and calculated lipophilicity and anti-tumour activity of isochromanone derivatives.

Compound lipophilicity connected to ADME(T)(a) has great importance in drug development and it has to be evaluated by the generally used drug developmental process. In addition to the importance of lipophilicity in ADMET, recently it has been reported that lipophilicity of small molecules correlates with their antiproliferative activity because of certain specific hydrophobic and lipophilic interactions. Due to the complexity of ADME(T) parameters an efficient and fast method is needed to characterize the many promising candidate lead molecules as a preselection in order not to be rejected from the latter phase of drug development. In the present paper we provide an overview of the importance of lipophilicity of drug candidates for biological action and for ADME(T) and describe a novel approach for drug-likeness characterization of a molecular library using correlation study between lipophilicity and biological activity. Lipophilicity and molecular characteristics have been measured, predicted and optimized for a diverse library from which the best members have been selected to describe their biological, chemical and drug-likeness properties. Molecules were selected from the family of alpha,beta-unsaturated ketones and thorough HPLC characterization for lipophilicity and morphological, antiproliferative and flow cytometric studies were carried out on them. Based on the results 17 member isochromanone library including E and Z geometric isomers were selected for further characterization. In this focused library linear correlation has been found between the calculated and measured lipophilicity and significant parabolic correlation was found between the antiproliferative effect and lipophilicity. Using our efficient and fast method, from a diverse library, we identified an outstandingly effective inhibitor of A431 tumour cell growth via a PARP(a) cleavage dependent apoptosis. In summary the optimized HPLC analyses of lipophilicity combined with the cell-culture assay, introduced above, resulted in the determination of an optimal lipophilicity range. This optimized lipophilicity range should be used in designing novel antiproliferative compounds.

Characterization of lipophilicity and antiproliferative activity of E-2-arylmethylene-1-tetralones and their heteroanalogues.

A molecular library based on E-2-arylmethylene-1-tetralone has been designed and synthesized. A reversed phase high performance liquid chromatographic (RP-HPLC) method has been developed and applied to separate them and to characterize their lipophilicity. The chromatographic method applied here was suitable to separate the structural (ortho and para) isomers of compounds and was sensitive enough to differentiate their lipophilicities. The measured (k') and computer calculated (CLOGP) lipophilicity values has been compared. Good linear correlation has been found in the case of these structurally related molecules. In vitro biological assay has been performed with Methylene blue dying to investigate the antiproliferative potency of the compounds synthesized in this work. The measured (k') and calculated (CLOGP) lipophilicities of the compounds were compared with the antiproliferative activities and an optimum value of lipophilicity has been found for these compounds.

Determination of the basicity of Mannich ketones by capillary electrophoresis.

Capillary electrophoretic (CE) method to characterise Mannich ketones (MKs) containing morpholine moiety was developed. Basicity (pKa,exp) of the MKs was characterised by measured data derived from the electrophoretic mobility values obtained in the CE separation. The MKs were found to be weaker bases than the parent morpholine molecule itself and the experimentally determined basicity values proved to be dependent on the chemical structure of the MKs. Since the basicity of the MKs has an influence on their reactivity and biological activity it seems to be useful to determine experimentally the pKa,exp values of the newly synthesised compounds to support rational drug design or screening of the molecule libraries.

Synthesis and antibacterial study of unsaturated Mannich ketones.

Several Mannich ketones of 2-arylmethylenecycloalkanones were synthesised using the classical acid-catalysed Mannich reaction. Antibacterial activity of these new water-soluble compounds was reported against Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Staphylococcus saprophyticus, Micrococcus luteus and Bacillus subtilis standard strains. Human cell line cytotoxicity of our new compounds was evaluated against HeLa cell lines. Some compounds showed low cytotoxicity (41.52 nM mL(-1) for 14 and 46.60 nM mL(-1) for 18) and proved to be efficient antibacterial agents against the Gram-positive strains. Minimum inhibitory concentrations varied from 1.56 to 100 microg mL(-1). The mechanism of action was examined, too.

23Na NMR shift reagents enhance cardiac staircase effect in isolated perfused rat hearts.

The effects of the currently used (23)Na NMR shift reagents, dysprosium bis-triphosphate [Dy(PPP)(2)], dysprosium triethylenetriamine hexaacetate [Dy(TTHA)] and thulium 1,4,7, 10-tetraazacyclododecane-N,N',N",N"'-tetra(methylenephosphonate) [Tm(DOTP)] were studied in the rat heart cardiac staircase model. Rat hearts were perfused with low or normal extracellular free calcium ([Ca(o)](f)). At low [Ca(o)](f) (0.34 +/- 0.05 mM), hearts were perfused with Dy(PPP)(2) (group I), Dy(TTHA) (group II) or no shift reagent (group III), while at normal [Ca(o)](f) (1.25 +/- 0.15 mM), hearts were perfused with Tm(DOTP) (group IV), Dy(TTHA) (group V) or no shift reagent (group VI). Left ventricular developed pressure (LVDP) values in group I were significantly higher than in groups II and III (p < 0.01), while no significant differences were found between groups II and III. LVDP values in group IV were significantly higher than in groups V and VI (p < 0.05), while the LVDP values in groups V and VI were almost identical. Also, a positive correlation between pacing rate and intracellular sodium ([Na(i)]) was evident. The [Na(i)] values at high [Ca(o)](f) were significantly lower than at low [Ca(o)](f) at each pacing level (p <0.01), indicating a negative correlation between [Na(i)] and [Ca(o)](f). No statistical differences were found in [Na(i)] between groups I vs II and IV vs V, showing that determination of [Na(i)] is not affected by any of these shift reagents. Thus the different LVDP responses in groups I vs II and IV vs V were not mirrored in [Na(i)] changes. We hypothesize that a direct, sarcolemmal Ca-Dy(PPP)(2)-, or Ca-Tm(DOTP)-induced positive inotropic effect could be responsible for these Na(i)-independent LVDP increases in groups I and IV.

Synthesis and in vivo evaluation of new contrast agents for cardiac MRI.

Analogues 2-6 of N(3),N(6)-bis(2'-myristoyloxyethyl)-1, 8-dioxotriethylenetetraamine-N,N,N',N'-tetraacetic acid (BME-DTTA) (1), which like 1 are also based on the DTTA structure but contain shorter fatty acyl chains, were synthesized to improve the water solubility of the corresponding gadolinium complexes. The gadolinium complexes of 1 and 3-5 have very low solubility in water. Thus liposomal preparations are necessary for their in vivo MRI application. These liposomal preparations retain high in vitro relaxivities (27.1, 21.57, 20.32, 23.1 s(-1) mM(-1), respectively) and induce sustained MRI signal intensity enhancements (67.2, 38.4, 52.1, 41.7 in arbitrary units, respectively). The gadolinium complex of 2 is quite soluble in water. Its lifetime in the blood stream, however, is short. The gadolinium complex of analogue 6, N-(2-butyryloxyethyl)-N'-(2-ethyloxyethyl)-N,N'-bis[N' ',N' '-bis(carboxymethyl)acetamido]-1,2-ethanediamine (ABE-DTTA), has demonstrated its potential as a water-soluble, cardiac-specific, MRI contrast agent. It is completely soluble in water at a 25 mM concentration, allowing the preparation of an injectable dose. The in vitro relaxivity of the complex is 16.24 s(-1) mM(-1). The agent shows a specific accumulation in the heart tissue reaching its maximum within 15 min after administration, inducing a sustained MRI signal intensity enhancement of 43.6%. This enhancement lasts for at least 3 h, thus indicating a reasonably long lifetime of this contrast agent in the myocardium without deleterious effects on heart function parameters.

2-Substituted indazoles. Synthesis and antimicrobial activity.

2-Isothiocarbamoyl substituted fused pyrazolines and their S-alkyl derivatives were prepared as potentially antimicrobial agents. Conventional methods were used to synthesize the novel derivatives starting from cyclic unsaturated ketones and thiosemicarbazide under acidic catalyst. These cyclizations yielded only one diastereoisomer of 3-H, 3a-H cis. The alkylations were performed applying alkyl halides. The structures of the new compounds, including configurations and conformations, were elucidated by NMR spectroscopy, also making use of 2D-HSC, DEPT and DNOE measurements. The S-alkyl derivatives were evaluated for activity against Gram-negative and Gram-positive bacteria and their in vitro toxicity was determined on HeLa cells. The structure-activity relationship was also studied.

The modulation of pacing-induced changes in intracellular sodium levels by extracellular Ca2+ in isolated perfused rat hearts.

This study demonstrates the inverse relationship between extracellular free calcium ([Ca(o)]f) and intracellular sodium ([Na(i)]) in isolated perfused rat hearts and thus supports the role of [Na(i)] in the "calcium paradox". It also shows that the extent of the increase in [Na(i)] (delta[Na(i)]), and the extent of the decrease in left ventricular developed pressure (deltaLVDP) in isolated perfused rat hearts, induced by pacing, is modulated by [Ca(o)]f. At low (0.24 mM) as well as normal (1.15 mM) [Ca(o)]f, [Na(i)] increased with pacing, progressively and significantly (P<0.01 and P<0.05, respectively), reaching a maximum of 12.56 +/- 0.46 and 9.22 +/- 0.16 mM at 500 beats/min, respectively. At high [Ca(o)]f (2.2 mM), however, no pacing-induced increase in [Na(i)] was observed. Simultaneously, within the pacing range of 250-500 beats/min, the interval-force relationship was negative for all [Ca(o)]f. With decreasing [Ca(o)]f, a gradually increasing delta[Na(i)] was induced. We hypothesise that a [Ca(o)]f-dependent Na-Ca exchanger activity modulates Na+ uptake, and thus baseline [Na(i)]. During incremental pacing, the increase in pacing rate induces a [Ca(o)]f-dependent delta[Na(i)], which may interact further with the sarcolemmal Na-Ca exchanger activity. As a result, both baseline [Na(i)] and the pacing-induced, [Ca(o)]f-dependent delta[Na(i)] modulate the net Ca2+ uptake, and thus SR Ca, in a manner that results in a modulated left ventricular force development.

Human epidermal blister: a convenient tissue for toxicological and genetic studies of benzo(a)pyrene metabolism.

Benzo(a)pyrene (BP) metabolism has been studied in epidermal blisters maintained in a culture medium for 24 h and 48 h. The viability of the cells has been assayed by [3H]proline incorporation into proteins and by [14C]BP metabolism into unconjugated metabolites. A screen of BP metabolism in 19 individuals shows a great variation of basal epidermal activity. Induction of BP metabolism by the application of coal tar 24 h before the epidermal blister sampling, resulted in two- to eight-fold increase in BP metabolism. This induction is not increased when the coal tar application is repeated.

[Potentially bioactive pyrimidine derivatives. 1. 2-Amino-4-aryl-8-arylidene-3,4,5,6,7,8-hexahydroquinazoline]. / Potentiell bioaktive Pyrimidinderivate. 1. Mitteilung: 2-Amino-4-aryl-8-aryliden-3,4,5,6,7,8-hexahydrochinazoline.

Six condensed 2-amino-4-arylpyrimidines (1a-f) were synthesized by the base catalyzed reaction of some monoarylidenecycloalkanones and guanidine. The reaction of 2,6-diarylidenecyclohexanones with guanidine yielded 2-amino-4-aryl-3,4,5,6,7,8-hexahydroquinazolines which were stable only as salts 3a-j. The oxidation of 3a-j led to 2-amino-4-aryl-8-arylidene-5,6,7,8-tetrahydroquinazolines 4a-j.

Treatment of generalized pustular psoriasis with Ro-10-9359.

4 patients suffering from generalized pustular psoriasis, von Zumbusch type, were treated with the aromatic retinoid Ro-10-9359. Lesions were completely cleared after 4 weeks of therapy. A maintenance dose was usually required to prevent relapses. Side-effects were often observed but always remained very mild. Ro-10-9359 represents the best available treatment for generalized pustular psoriasis.

A comparative study between interdermal tests with phytohemagglutinin and delayed hypersensitivity reactions elicited by tuberculin.

Intradermal (i.d.) tests with phytohemagglutinin (PHA) and tuberculin are used in clinical practice to evaluate the cell-mediated immunity. The biologic significance of the skin response to PHA, clinically evaluated by the extent of erythema and induration, was studied histologically after incorporation of tritiated thymidine. It was compared to tuberculin tests. The rate of recruitment and the respective amount of inflammatory cells varied between patients and in time. In PHA tests the inflammatory reaction occurred in two phases, the first one consisting in the predominance of polymorphonuclears, the second one being characterized by chemotactism of lymphoblasts many of which synthesized DNA. The "in vivo" reaction to PHA is therefore complex; the same clinical evaluation corresponds to markedly different stages and events in the biological reaction. The histological grading of the PHA test is therefore desirable in the evaluation of the cell-mediated immunity because only the blastic proliferation is of importance in its rating and not the edema and the accumulation of polymorphonuclears.