The structural elucidation of a novel iridoid derivative from Tachiadenus longiflorus (Gentianaceae) using the LSD programme and quantum chemical computations

Oleanolic acid, scoparone, scopoletin and a novel iridoid derivative, angelone, were isolated from Tachiadenus longiflorus (Gentianaceae). The structure of angelone was determined from NMR data, given as input to the Logic for Structure Determination Programme, and was finally confirmed by comparison of experimental 13C-NMR chemical shifts with those obtained by quantum mechanical calculations.     

A new synthetic approach to 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid (Tic) derivatives via a [2 + 2 + 2] cycloaddition reaction

Tetrahydroisoquinoline-3-carboxylic acid derivatives are prepared via a [2 + 2 + 2] cycloaddition reaction as a key step using Wilkinson's and CpCo(CO)2 catalysts.     

Chloroperoxidase-catalyzed cyclodimerization of methyl (2E)-2,4-pentadienoate: a [4+2] cycloaddition product

The chloroperoxidase (CPO)-catalyzed oxidation of the methyl (2E)-2,4-pentadienoate gives the terminal double bond epoxide (25%) and a cyclodimerization compound (63%) as the major products.     

Potassium- and Carbachol-Evoked Release of [3H]Noradrenaline from Human Neuroblastoma Cells, SH-SY5Y

The human neuroblastoma clone SH-SY5Y expresses potassium-, carbachol-, and calcium ionophore A23187-evoked, calcium-dependent release of [3H]noradrenaline. Release in response to carbachol and potassium was greater than additive. Atropine (Ki = 0.33 nM), hexahydrosiladifenidol (Ki = 18 nM), and pirenzepine (Ki = 1,183 nM) completely inhibited the carbachol-evoked noradrenaline release, an order of potency suggesting that an M3 receptor was linked to release. In contrast, noradrenaline release was only partially inhibited by the M2-selective antagonists methoctramine (10(-4) M) and AFDX-116 (10(-4) M), by approximately 14 and 46%, respectively. The nicotinic antagonist d-tubocurarine (10(-4) M) resulted in a partial inhibition of release, a finding suggesting that a nicotinic receptor may also be involved. SH-SY5Y provides a suitable cell line in which to study the biochemical mechanisms underlying the cholinergic receptor regulation of noradrenaline release.     

Synthesis and preliminary evaluation of a novel glutamine derivative: (2S,4S)4-[18F]FEBGln

We report the preparation of a novel glutamine derivative, (2S,4S)-2,5-diamino-4-(4-(2-fluoroethoxy)benzyl)-5-oxopentanoic acid, (2S, 4S)4-[¹⁸F]FEBGln ([¹⁸F]4), through efficient organic and radiosyntheses. In vitro assays of [¹⁸F]4 using MCF-7 cells showed that it entered cells via multiple amino acid transporter systems including system L and ASC2 transporters but not through the system A transporter. [¹⁸F]4 showed promising properties for tumor imaging and may serve as a lead compound for further optimizing and targeting the system L transporter associated with enhanced glutamine metabolism in cancer cells.     

Polynuclear complexes of a dissociative excited state formed in the [Ru(bpy)2(CN)2]-HgCl2 system.

The complex cis-dicyanobis(2,2'-bipyridine)ruthenium(II) forms various bimetallic complexes with mercury(II)chloride, such as [(NC)Ru(bpy)2(CN)-HgCl2], [Cl2Hg-(NC)Ru(bpy)2(CN)-HgCl2-(NC)Ru(bpy)2(CN)-HgCl2] and [Cl2Hg-(NC)Ru(bpy)2(CN)-(HgCl2)] in CH3CN. These polynuclear complexes of the equilibrium system have been identified and characterized by their formation constants and absorption spectra. Excitation of bimetallic complexes produces the MLCT state localized on [Ru(bpy)2(CN)2] ligand, resulting in the cleavage of the bond formed between the nitrogen atom of the coordinated cyanide ligand and the Hg(II) central atom in ground state. Unlike many photoinduced metal ligand dissociations, the dissociated fragment remains in a luminescent excited state.     

Ruthenium-modified proteins. Reactions of cis-[ Ru(NH3)4(OH2)2]2+ and cis-[ Ru(en)2(OH2)2]2+ with azurin,myoglobin and cytochrome c

Reactions of cis-[Ru(en)₂(OH₂)₂]²⁺ (or cis-[Ru (NH₃)₄(OH₂)₂]²⁺) with Pseudomonas aeruginosa azurin (Az), horse heart myoglobin (Mb^h), and horse heart cytochrome c (cyt c) give Ru-labelled proteins. The ruthenium binding sites in the singly modified derivatives are His-83 (Az), His-81 (Mb^h), and His-33 (cyt c). Spectroscopic and electrochemical measurements indicate that the structures of the proteins are not perturbed by the surface-bound ruthenium complexes. The E°_f values of the Ru(III)/(II) couple in these Ru-modified proteins fall between −0.07 and −0.13 V vs. NHE.     

Effect of Inhibitors of Eicosanoid Metabolism on Release of [3H]Noradrenaline from the Human Neuroblastoma, SH-SY5Y

Abstract: Nordihydroguaiaretic acid (NDGA; a lipoxygenase inhibitor), LY-270766 (an inhibitor of 5-lipoxygenase), and the diacylglycerol lipase inhibitor RG 80267 completely eliminated potassium-evoked release of [³H]noradrenaline ([³H]NA) from the human neuroblastoma clone SH-SY5Y with IC₅₀ values of 10, 15, and 30 μ M , respectively. In contrast, these inhibitors only partially inhibited carbachol-evoked release and had little effect on the calcium ionophore A23187-evoked release of NA in this cell line. Arachidonic acid partially inhibited potassium- and A23187-evoked release but did not reverse the inhibition of potassium-evoked release observed in the presence of RG 80267. These studies suggest that arachidonic acid (or its lipoxygenase products) are not important intermediates in the regulation of exocytosis in SH-SY5Y. This conclusion is strengthened by our studies in which SH-SY5Y cells were grown in medium supplemented with bovine serum albumin-linoleic acid (50 μ M ). Under these conditions there was a selective increase in content of membrane polyunsaturated fatty acids of the ω6 series, including arachidonic acid; however, these changes did not effect potassium-, veratridine-, carbachol-, or calcium ionophoreevoked release of [³H]NA.     

S-[2-(N7-guanyl)ethyl]glutathione, the major DNA adduct formed from 1,2-dibromoethane

The reaction of 1,2-dibromoethane and glutathione with DNA in the presence of glutathione S-transferase results in the formation of a single major DNA adduct, which can be released by thermal hydrolysis at neutral pH and separated by octadecylsilyl and propylamino high-performance liquid chromatography. The same DNA adduct is the only major one formed in livers of rats treated with 1,2-dibromo[1,2-14C]ethane. The DNA adduct was identified as S-[2-(N7-guanyl)ethyl]glutathione: (1) The chromatographic behavior was altered by treatment with gamma-glutamyl transpeptidase or Streptomyces griseus protease. (2) The molecular ions observed in positive and negative mode fast atom bombardment mass spectrometry were those expected for the structure when either glycerol or a mixture of dithiothreitol and dithioerythritol was used as the bombardment matrix. (3) The two-dimensional 1H NMR correlated spectroscopy spectrum of the DNA adduct was compared to the spectra of glutathione, oxidized glutathione, and N7-methylguanine and found to be consistent with the assigned structure. No evidence for in vitro or in vivo opening of the guanyl imidazole ring was observed under these conditions. The structure of the adduct supports a pathway involving enzyme-catalyzed conjugation of 1,2-dibromoethane with glutathione, non-enzymatic dehydrohalogenation of the resulting half-mustard to form a cyclic episulfonium ion, and attack of the N7 nitrogen of DNA guanine on the episulfonium ion to generate this major DNA adduct, which may be related to the carcinogenicity of this chemical.     

18F]azadibenzocyclooctyne ([18F]ADIBO): a biocompatible radioactive labeling synthon for peptides using catalyst free [3+2] cycloaddition

N-Terminally azido-modified peptides were labeled with the novel prosthetic labeling synthon [(18)F]azadibenzocyclooctyne ([(18)F]ADIBO) using copper-free azide-alkyne [3+2]-dipolar cycloaddition in high radiochemical yields (RCYs). (18)F-Labeled [(18)F]ADIBO was prepared by nucleophilic substitution of the corresponding tosylate in 21% overall RCY (EOB) in a fully automated synthesis unit within 55 min. [(18)F]ADIBO was incubated with azide-containing peptides at room temperature in the absence of toxic metal catalysts and the formation of the triazole conjugate was confirmed. Finally, the azide-alkyne [3+2]-dipolar cycloaddition was shown to proceed with 95% radiochemical yield in ethanol within 30 min, allowing for a development of a kit-like peptide labeling approach with [(18)F]ADIBO.     

Induction of G2/M phase arrest and apoptosis by a novel enediyne derivative, THDA, in chronic myeloid leukemia (K562) cells

We studied the effect of 2-(6-(2-thieanisyl)-3(Z)-hexen-1,5-diynyl)aniline(THDA), a newly developed anti-cancer agent, on cell proliferation, cell cycle progression, and induction of apoptosis in K562 cells. THDA was found to inhibit the growth of K562 cells in a time-and dose-dependent manner. Cell cycle analysis showed G2/M phase arrest and apoptosis in K562 cells following 24 h exposure to THDA. During the G2/M arrest, cyclin-dependent kinase inhibitors (CDKIs), p21 and p27 were increased in a time-dependent manner. Analysis of the cell cycle regulatory proteins demonstrated that THDA did not change the steady-state levels of cyclin B1, cyclin D3 and Cdc25C, but decreased the protein levels of Cdk1, Cdk2 and cyclin A. THDA also caused a marked increase in apoptosis, which was associated with activation of caspase-3 and proteolytic cleavage of poly (ADP-ribose) polymerase. These molecular alterations provide an insight into THDA-caused growth inhibition, G2/M arrest and apoptotic death of K562 cells.     

Intramolecular stacking interactions in ternary copper(II) complexes formed by a heteroaromatic amine and 9-[2-(2-phosphonoethoxy)ethyl]adenine, a relative of the antiviral nucleotide analogue 9-[2-(phosphonomethoxy)ethyl]adenine

The stability constants of the mixed-ligand complexes formed between Cu(Arm)2+, where Arm=2,2'-bipyridine (Bpy) or 1,10-phenanthroline (Phen), and the dianions of 9-[2-(2-phosphonoethoxy)ethyl]adenine (PEEA2-) and (2-phosphonoethoxy)ethane (PEE2-), also known as [2-(2-ethoxy)ethyl]phosphonate, were determined by potentiometric pH titrations in aqueous solution (25 degrees C; I=0.1 M, NaNO3). The ternary Cu(Arm)(PEEA) complexes are considerably more stable than the corresponding Cu(Arm)(R-PO3) species, where R-PO3(2-) represents a phosph(on)ate ligand with a group R that is unable to participate in any kind of interaction within the complexes. The increased stability is attributed to intramolecular stack formation in the Cu(Arm)(PEEA) complexes and also, to a smaller extent, to the formation of 6-membered chelates involving the ether oxygen atom present in the -CH2-O-CH2-CH2-PO3(2-) residue of PEEA2-. This latter interaction is separately quantified by studying the ternary Cu(Arm)(PEE) complexes which can form the 6-membered chelates but where no intramolecular ligand-ligand stacking is possible. Application of these results allows a quantitative analysis of the intramolecular equilibria involving three structurally different Cu(Arm)(PEEA) species; e.g., of the Cu(Bpy)(PEEA) system about 11% exist with the metal ion solely coordinated to the phosphonate group, 4% as a 6-membered chelate involving the ether oxygen atom of the -CH2-O-CH2CH2-PO3(2-) residue, and 85% with an intramolecular stack between the adenine moiety of PEEA2- and the aromatic rings of Bpy. In addition, the Cu(Arm)(PEEA) complexes may be protonated, leading to Cu(Arm)(H;PEEA)+ species for which it is concluded that the proton is located at the phosphonate group and that the complexes are mainly formed (50 and 70%) by a stacking adduct between Cu(Arm)2+ and the adenine residue of H(PEEA)-. Finally, the stacking properties of adenosine 5'-monophosphate (AMP2-), of the dianion of 9-[2-(phophonomethoxy)ethyl]adenine (PMEA2-) and of several of its analogues (=PA2-) are compared in their ternary Cu(Arm)(AMP) and Cu(Arm)(PA) systems. Conclusions regarding the antiviral properties of several acyclic nucleoside phosphonates are shortly discussed.     

3-(3-Methoxyphenyl)-6-(3-amino-4-methoxyphenyl)-7H-[1,2,4] triazolo [3,4-b][1,3,4] thiadiazine,a novel tubulin inhibitor,evokes G2/M cell cycle arrest and apoptosis in SGC-7901 and HeLa cells

Gastric cancer and cervical cancer are two major malignant tumors that threaten human health. The novel chemotherapeutic drugs are needed urgently to treat gastric cancer and cervical cancer with high anticancer activity and metabolic stability. Previously we have reported the synthesis, characterization and identification of a novel combretastatin A-4 analog, 3-(3-methoxyphenyl)-6-(3-amino-4- methoxyphenyl) -7H-[1,2,4]triazolo[3,4-b][1,3,4] thiadiazine (XSD-7). In this study, we sought to investigate its anticancer mechanisms in a human gastric cancer cell line (SGC-7901 cells) and human cervical carcinoma cell line (HeLa cells). The 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay showed that XSD-7 induced cytotoxicity in SGC-7901 and HeLa cells with inhibitory concentration 50 values of 0.11 ± 0.03 and 0.12 ± 0.05 µM, respectively. Immunofluorescence studies proved that XSD-7 inhibited microtubule polymerization during cell division in SGC-7901 and HeLa cells. Then, these cells were arrested at G2/M cell cycle and subsequently progressed into apoptosis. In further study, mitochondrial membrane potential analysis and Western blot analysis demonstrated that XSD-7 treatment-induced SGC-7901 cell apoptosis via both the mitochondria-mediated pathway and the death receptor-mediated pathway. In contrast, XSD-7 induced apoptosis in HeLa cells mainly via the mitochondria-mediated pathway. Hence, our data indicate that XSD-7 exerted antiproliferative activity by disrupting microtubule dynamics, leading to cell cycle arrest, and eventually inducing cell apoptosis. XSD-7 with novel structure has the potential to be developed for therapeutic treatment of gastric cancer and cervical cancer.     

Mutagenic activation of 2-amino-3-methylimidazo[4,5-f]-quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline (MeIQ) by subcellular fractions and cells isolated from small intestine,kidney and liver of the rat

The mutagenic activity of the pyrolysis products 2-amino-3-methylimidazo[4,5-f]-quinoline and 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline in Salmonella typhimurium TA98 using rat intestinal and renal subcellular fractions as activation systems was approximately 1 and 5 revertants per nmol, respectively. This was 1,000 times less than the activity with a subcellular fraction from rat liver. The mutagenic activity of both compounds was considerably increased using intestinal, renal and hepatic preparations isolated from PCB (Aroclor 1254)-pretreated rats, compared to preparations from control animals. In addition, both compounds displayed a moderate direct-acting mutagenic activity at concentrations above 10^-5 M. Isolated cells from small intestine, kidney and liver incubated in nucleopore chambers were able to convert both compounds into products which mutated bacteria outside the chambers. The concentrations of chemicals required to yield responses of a similar magnitude were approximately 3 orders of magnitude higher in the intestinal and renal systems compared to the hepatic system. The formation of metabolites mutagenic for Salmonella typhimurium by hepatic subcellular and cellular systems was shown to be superior to the respective intestinal and renal systems.Abbreviations AHH arylhydrocarbon hydroxylase - IQ 2-amino-3-methylimidazo[4,5-f]-quinoline - MC 3-methylcholanthrene - MeIQ 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline - PCB polychlorinated biphenyls (Aroclor 1254) - S9 the 9,000 g supernatant tissue fraction - TCDD 2,3,7,8-tetrachlorodibenzo-p-dioxin     

DNA hybridization sensor utilizing [Os(5,6-dmphen)3]2+ by square wave voltammetry.

DNA hybridization detection utilizing [Os(5,6-dmphen)3]2+ (tris(5,6-dimethyl-phenanthroline) osmium(II/III)) as hybridization indicator was studied, because [Os(5,6-dmphen)3]2+ has the most largest association constant (K2+) and high current density in osmium methyl substituted phenanthroline complexes. As the result, target DNA could be detected selectively ranging from 6.9 x 10(-10) to 6.9 x 10(-5) g/mL using square wave voltammetry.     

Mobilization of Inositol 1,4,5-Trisphosphate-Sensitive Ca2+ Stores Supports Bradykinin- and Muscarinic-Evoked Release of [3H]Noradrenaline from SH-SY5Y Cells

Abstract: The human neuroblastoma cell line SH-SY5Y, maintained at confluence for 14 days, released [³H]-noradrenaline ([³H]NA) when stimulated with either the muscarinic receptor agonist methacholine or bradykinin. The major fraction of release was rapid, occurring in <10 s, whereas nicotine-evoked release was slower. When the extracellular [Ca²⁺] ([Ca²⁺]_e) was buffered to ～50–100 nM, release evoked by nicotine was abolished, whereas that in response to methacholine or bradykinin was reduced by ～50% with EC₅₀ values of ?5.46 ± 0.05 M and ?7.46 ± 0.06 M (log₁₀), respectively. Methacholine and bradykinin also produced rapid elevations of both inositol 1,4,5-trisphosphate [Ins(1,4,5)P₃] and intracellular free [Ca²⁺] ([Ca²⁺]_i). These elevations were reduced at low [Ca²⁺]_e and under these conditions the EC₅₀ values for peak elevation of [Ca²⁺]_i were ?6.00 ± 0.14 M for methacholine and ?7.95 ± 0.34 M for bradykinin (n = 3 for all EC₅₀ determinations). At low [Ca²⁺]_e, depletion of nonmitochondrial intracellular Ca²⁺ stores with the Ca²⁺-ATPase inhibitor thapsigargin produced a transient small elevation of [Ca²⁺]_i and a minor release of [³H]NA. At low [Ca²⁺]_e, thapsigargin abolished elevation of [Ca²⁺]_i in response to methacholine and bradykinin and completely inhibited their stimulation of [³H]NA release. It is proposed, therefore, that Ca²⁺ release from Ins(1,4,5)P₃-sensitive stores is a major trigger of methacholine- and bradykinin-evoked [³H]NA release in SH-SY5Y cells.     

Unusual ligand design: A platinum(II) mediated [2+2] photocycloaddition followed by a nickel(II) induced methoxyactivation

The novel dimer of the composition [Pt₂Cl₄(μ-(κP¹:κP²-o-MeO-trans-dppen))₂] (1) (o-MeO-trans-dppen = 1,2-(bis(o-methoxyphenyl)phosphanyl)ethylene) has been prepared and characterized by a single crystal X-ray structure analysis, NMR spectroscopy, mass spectrometry and elemental analysis. This latter compound undergoes a [2+2] photocycloaddition reaction yielding the tetraphosphane all-trans-1,2,3,4-tetrakis(di(o-methoxyphenyl)phosphanyl)cyclobutane (o-MeO-dppcb). The X-ray structure of the dimeric Ni(II) complex that contains the latter ligand, of the formula [Ni₂Cl₄(μ-(κP¹:κP²:κP³:κP⁴-o-MeO-dppcb))] (2) reveals that the apical coordination sites of both square pyramidal Ni(II) coordination spheres are occupied by methoxy-oxygen atoms of the ligand. As a consequence, this dimeric Ni(II) complex 2 is prone to a thermally induced regio- and diastereoselective metal-assisted methoxy-group cleavage. The stepwise formed new mono- and bis-phenolate complexes [Ni₂Cl₃(μ-(κO¹,κP¹:κP²:κP³:κP⁴-o-MeO-O-dppcb))] (3) and [Ni₂Cl₂(μ-(κO¹,κP¹:κP²:κO²,κP³:κP⁴-o-MeO-O,O′-dppcb))] (4), respectively, contain the novel chiral tetraphosphane ligands all-trans-1,2,3-tris((di-o-methoxyphenyl)phosphano)-4-((o-methoxy-phenyl)(o-phenolate)phosphano)cyclobutane (o-MeO-O-dppcb) and all-trans-1,2-bis((di-o-methoxyphenyl)phosphano)-3,4-bis((o-methoxyphenyl)(o-phenolate)phosphano)cyclobutane (o-MeO-O,O′-dppcb). Compounds 3 and 4 have been synthesized independently and are also fully characterized by both single crystal X-ray structure analyses, NMR spectroscopy, mass spectrometry and elemental analyses. The conversion of 2 into 3 and then further into 4 has been followed by a variable-temperature ³¹P{¹H} NMR experiment with compound 2 in DMF-d⁷, revealing that the cleavage of the second methoxy group is kinetically disfavoured. This is in agreement with the X-ray structure analysis of 3, indicating the lack of any methoxy-oxygen atom coordination that could easily induce a further methoxy-group cleavage. o-MeO-O-dppcb and o-MeO-O,O′-dppcb are rare P-stereogenic tetraphosphine ligands and contribute to the synthetic field of new κ³-P,P,O-coordinating phosphanylphenolate ligands that are believed to be important for the SHOP process (SHOP, Shell Higher Olefin Process).     

Synthesis, characterization and DNA-binding of novel chiral complexes delta- and lambda-[Ru(bpy)2L]2+ (L = o-mopip and p-mopip)

Novel chiral Ru(II) complexes [Ru(bpy)2L]2+ (bpy = 2,2-bipyridine; L: o-mopip = 2-(2-methoxylphenyl)imidazo[4,5-f][1,10]phenanthroline, p-mopip = 2-(4-methoxylphenyl)imidazo[4,5-f][1,10]phenanthroline) containing -OCH3 at different positions on the phenyl ring have been synthesized and characterized. The DNA-binding and DNA-photocleavage properties of the complexes were investigated. The theoretical calculations for these complexes were also carried out applying the density functional theory (DFT) method. The experimental results show that: both these two isomer complexes can bind to DNA in an intercalative mode; the DNA-binding affinity of [Ru(bpy)2(p-mopip)] 2 is greater than that of [Ru(bpy)2(o-mopip)] 1; moreover, the DNA-binding affinities of enantiomers delta-1 and delta-2 are all greater than those of lambda-1 and lambda-2, respectively. In addition, a very interesting finding is experimentally obtained, i.e. under a low [DNA]/[Ru] ratio, the emission intensities of delta-1 and lambda-1 are all weaker than those of delta-2 and lambda-2, however, upon a high [DNA]/[Ru] ratio, the emission intensities of both delta-1 and lambda-1 are stronger than those of delta-2 and lambda-2. Such a difference of the emission spectra can be interpreted by the electric effect of substituent on the intercalative ligand. The difference in DNA-binding affinities of these two isomeric complexes can also be reasonably explained by the DFT calculations.