Microcalorimetric and spectroscopic investigation of the antibacterial properties of cationic ytterbium(III)-porphyrin complexes lacking charged peripheral groups

The antibacterial activities towards Escherichia coli of two cationic Yb(III)-monoporphyrin complexes, [Yb(III)(TMP)(H2O)3]Cl (1) and [Yb(III)(TTP)(H2O)3]Cl (2), were investigated at the cellular and sub-cellular levels. The biological effects of the complexes on the growth of E. coli were evaluated by microcalorimetry and by analysis of the resulting metabolic thermogenic curves, from which IC50 values and metabolic parameters such as growth rate and generation time were derived. At the subcellular level, DNA-binding experiments were performed by means of UV/VIS- and fluorescence-titration experiments, as well as by near-infrared (NIR) emission, which revealed that 1 and 2 strongly bind to herring-sperm DNA (HS-DNA), though by different binding modes.     

Toxic effects of mercury(II), cadmium(II) and lead(II) porphyrins on Trypanosoma brucei brucei growth

The effects of free mercury(II), cadmium(II) and lead(II) ions and their metalloporphyrin-derivatives on Trypanosoma brucei brucei growth in culture were studied. All experiments were conducted in the dark. IC(50) values on growth obtained in 24-h time-course experiments were 1.5 x 10(-7), 2.4 x 10(-6), 4.4 x 10(-6) and 2.6 x 10(-5) M for mercury(II) porphyrin, cadmium(II) porphyrin, lead(II) porphyrin and free base porphyrin, respectively. While the IC50 values for Hg2+, Cd2+ and Pb2+ were 3.6 x 10(-6), 1.5 x 10(-5) and 1.6 x 10(-5) M, respectively. These results clearly indicate that the toxicity of the metalloporphyrin complexes of mercury(II), cadmium(II) and lead(II) to T. b. brucei parasites was much higher compared to their free metal ions and free base porphyrin at low concentrations. It was also observed after 8 h incubation that the metalloporphyrins were effective in inhibiting the division of the parasites at concentrations >1.25 x 10(-7) M for mercury(II) porphyrin, concentrations >1.2 x 10(-6) M for cadmium(II) and lead(II) porphyrins and at concentrations >3.6 x 10(-6) M for Hg2+ ion. These observations were not detected in samples treated with the free metal ions and the free base porphyrin at the same concentrations. Interestingly, trypanosomes treated with metalloporphyrin complexes displayed different morphological features from those cells treated with free base porphyrin or metal ions. The chemotherapeutic potential of the metalloporphyrins of H2TMPyP for treatment of African trypanosomiasis is discussed.     

Self-assembly and cytotoxicity study of waterwheel-like dinuclear metal complexes: the first metal complexes appended with multiple free hydroxamic acid groups

Two waterwheel-like dinuclear complexes [M(2)(PHA)(4)(H(2)O)(2)] (M = Cu(II) (1), Zn(II) (2); HPHA = phthal-hydroxamic acid) appended with four free hydroxamic acid groups, namely, free hydroxamic acid metal complexes (FHAMCs) have been synthesized and characterized. The crystal structure of complex 1 was determined by single crystal X-ray diffraction, which adopts the paddlewheel motif with four bidentate carboxylate ligands joining two Cu(II) ions. The relative cytotoxicities of compounds 1 and 2 against SMMC-7721 and HO-8910 cell lines are similar and more predominant than HPHA (IC(50): Cu(II)>Zn(II)>HPHA). The synergic effect of the bound water molecules, multiple free hydroxamic acid groups and dimetal active sites with bridging carboxylate may have significant impacts on their pharmacological activity. As the prototype for a new class of hydroxamic acid derivatives, the self-assembly of FHAMCs presents a promising new strategy in designing multiple hydroxamic acids with remarkable bioactivities.     

Inhibition of [3H]platelet activating factor (PAF) binding by Zn2+: a possible explanation for its specific PAF antiaggregating effects in human platelets

Zinc ions in the micromolar range exhibited a strong inhibitory activity toward platelet activating factor (PAF)-induced human washed platelet activation, if added prior to this lipid chemical mediator. The concentration of Zn2+ required for 50% inhibition of aggregation (IC50) was inversely proportional to the concentration of PAF present. The IC50 values (in microM) for Zn2+ were 8.8 +/- 3.9, 27 +/- 5.8, and 34 +/- 1.7 against 2, 5, and 10 nM PAF, respectively (n = 3-6). Zn2+ exhibited comparable inhibitory effects on [3H]serotonin secretion and the IC50 values (in microM) were 10 +/- 1.2, 18 +/- 3.5, and 35 +/- 0.0 against 2, 5, and 10 nM PAF, respectively (n = 3). Under the same experimental conditions, aggregation and serotonin secretion induced by ADP (5 microM), arachidonic acid (3.3 microM), or thrombin (0.05 U/ml) were not inhibited. Introduction of Zn2+ within 0-2 min after PAF addition not only blocked further platelet aggregation and [3H]serotonin secretion but also caused reversal of aggregation. Analysis of [3H]PAF binding to platelets showed that Zn2+ as well as unlabeled PAF prevented the specific binding of [3H]PAF. The inhibition of [3H]PAF specific binding was proportional to the concentration of Zn2+ and the IC50 value was 18 +/- 2 microM against 1 nM [3H]PAF (n = 3). Other cations, such as Cd2+, Cu2+, and La3+, were ineffective as inhibitors of PAF at concentrations where Zn2+ showed its maximal effects. However, Cd2+ and Cu2+ at high concentrations exhibited a significant inhibition of the aggregation induced by 10 nM PAF with IC50 values being five- and sevenfold higher, respectively, than the IC50 for Zn2+, and with the IC50 values for inhibition of binding of 1 nM [3H]PAF being 5 and 19 times higher, respectively, than the IC50 for Zn2+. The specific inhibition of PAF-induced platelet activation and PAF binding to platelets suggested strongly that Zn2+ interacted with the functional receptor site of PAF or at a contiguous site.     

Interactions Between α-Latrotoxin and Trivalent Cations in Rat Striatal Synaptosomal Preparations

The interactions between alpha-latrotoxin (alpha-LTx), a neurosecretagogue purified from the venom of the black widow spider, and the trivalent cations Al3+, Y3+, La3+, Gd3+, and Yb3+ were investigated in rat striatal synaptosomal preparations. All trivalent cations tested were inhibitors of alpha-LTx-induced [3H]dopamine [( 3H]DA) release (order of potency: Yb3+ greater than Gd3+ approximately Y3+ greater than La3+ greater than Al3+). Only with Al3+ could inhibition of [3H]DA release be attributed to a block of 125I-alpha-LTx specific binding to synaptosomal preparations. The inhibitory effect of trivalent ions was reversible provided synaptosomes were washed with buffer containing EDTA. Trivalent ions also inhibited alpha-LTx-induced [3H]DA release at times when alpha-LTx-stimulated release was already evident. alpha-LTx-induced synaptosomal membrane depolarization was blocked by La3+, but not affected by Gd3+, Y3+, and Yb3+. alpha-LTx-stimulated uptake of 45Ca2+ was inhibited by all trivalent cations tested. These results demonstrate that there exist at least three means by which trivalent cations can inhibit alpha-LTx action in rat striatal synaptosomal preparations: (1) inhibition of alpha-LTx binding (Al3+); (2) inhibition of alpha-LTx-induced depolarization (La3+); and (3) inhibition of alpha-LTx-induced 45Ca2+ uptake (Gd3+, Y3+, Yb3+, La3+).     

ATP receptor. A putative receptor-operated channel in PC-12 cells.

External ATP induces [3H] dopamine [( 3H]DA) release in rat pheochromocytoma cells (PC-12 cells). The ATP-induced release is a saturable process with half-effective concentration of EC50 = 80 microM. ADP is a poor secretagogue of [3H]DA (one-sixth of ATP) and AMP is devoid of secretory capabilities. Adenosine and the non-hydrolyzable analogues of ATP, AppNHp and AppCp are ineffective as inducers of [3H]DA, release, or as inhibitors of the ATP-induced [3H]DA release. The most potent antagonist of ATP-induced release is Coomassie Blue (IC50 = 25 microM), compared to ADP beta S (IC50 = 500 microM). The overall rank order of potency is ATP greater than ADP much greater than AMP greater than adenosine, which is characteristic of the P2-purinergic receptor. ATP-induced secretion is absolutely Ca2+ dependent, indicating an exocytotic process and is independent of Mg2+ (up to 2 mM) suggesting that the active species is not ATP4-. (a) The ATP-induced 45Ca2+ influx into the cells is in good correlation to ATP induction of release (IC50 = 80 and 90 microM, respectively) and is carried over to ADP which has a diminished ability to induce both release and 45Ca2+ influx. (b) Divalent cations (Ba2+ greater than Sr2+ greater than Ln3+ greater than Mn2+) replace Ca2+ and support ATP-induced release similar to their effectiveness in supporting bradykinin- and K+ (50 mM)-induced release in PC-12 cells (Weiss, C., Sela, D., and Atlas, D. (1990) Neurosci. Lett. 119, 241-245). Combined together the absolute requirement of [Ca2+]ex for release, inhibition of release by Gd3+ (IC50 = 100 microM), Ni2+, and Co2+ (IC50 = 1 mM), and support of release by Ba2+, Sr2+, and Mn2+, we suggest that ATP induces Ca2+ entry via ligand-operated Ca2+ channels as previously suggested for ATP in smooth muscle cells (Benham, C.D., Bolton, T.B., Byren, N.G., and Large, W.A. (1987) J. Physiol. (Lond.) 387, 473-488). No significant inhibition by 1 microM verapamil, 10 microM nifedipine, or 2 mM Cd2+ argues against ATP activation of voltage-dependent Ca2+ channels as similarly shown for ATP-induced [3H]noradrenaline release (Inoue, K., Nakazawa, K., Fujimoro, K., and Takanaka, A. (1989) Neurosci. Lett. 106, 294-299). Thus, the widely distributed ATP receptor might play an essential role in Ca2+ homeostasis of the cell by introducing Ca2+ into the cell via specific ligand-gated Ca2+ channels.     

Fluoro-hydrido-oxo and other fluoro-oxo complexes of rhenium (V) containing a triphosphine ligand

The fluoro-hydrido-oxo complex [Re(F)(H)(O)Cyttp]⁺ (3, Cyttp = PhP(CH₂CH₂CH₂PCy₂)₂) was prepared in high yield from [Re(H₂)H₄Cyttp]SbF₆ (1(SbF₆), NaSbF₆ and acetone in toluene at reflux. Reaction chemistry of 3 has been studied and, where appropriate, compared with that of the related dihydrido-oxo complex [ReH₂(O)Cyttp]⁺ (2). Unlike 2, which readily reacts with both CO and SO₂, 3 was found to be inert to these reagents under comparable conditions. However, 3(SbF₆) reacts with NaSbF₆ at elevated temperature to afford the difluoro-oxo complex [ReF₂(O)Cyttp]⁺ (4). 4 undergoes fluoride substitution by Cl⁻ or Br⁻ to yield [Re(X)(F)(O)Cyttp]⁺ (X = Cl (5, Br (6)). 5 can also be obtained by treatment of 6(BPh₄) with LiCl. All of these complexes contain mer-Cyttp, and 3–6 contain trans fluoride and oxide ligands as inferred from spectroscopic data.     

Leishmania plasma membrane Mg2+-ATPase is a H+/K+-antiporter involved in glucose symport. Studies with sealed ghosts and vesicles of opposite polarity

Experiments from other laboratories conducted with Leishmania donovani promastigote cells had earlier indicated that the plasma membrane Mg2+-ATPase of the parasite is an extrusion pump for H+. Taking advantage of the pellicular microtubular structure of the plasma membrane of the organism, we report procedures for obtaining sealed ghost and sealed everted vesicle of defined polarity. Rapid influx of H+ into everted vesicles was found to be dependent on the simultaneous presence of ATP (1 mm) and Mg2+ (1 mm). Excellent correspondence between rate of H+ entry and the enzyme activity clearly demonstrated the Mg2+-ATPase to be a true H+ pump. H+ entry into everted vesicle was strongly inhibited by SCH28080 (IC50 = approximately 40 microm) and by omeprazole (IC50 = approximately 50 microm), both of which are characteristic inhibitors of mammalian gastric H+,K+-ATPase. H+ influx was completely insensitive to ouabain (250 microm), the typical inhibitor of Na+,K+-ATPase. Mg2+-ATPase activity could be partially stimulated with K+ (20 mm) that was inhibitable (>85%) with SCH28080 (50 microm). ATP-dependent rapid efflux of 86Rb+ from preloaded vesicles was completely inhibited by preincubation with omeprazole (150 microm) and by 5,5'-dithiobis-(2-nitrobenzoic acid) (1 mm), an inhibitor of the enzyme. Assuming Rb+ to be a true surrogate for K+, an ATP-dependent, electroneutral stoichiometric exchange of H+ and K+(1:1) was established. Rapid and 10-fold active accumulation of [U-(14)C]2-deoxyglucose in sealed ghosts could be observed when an artificial pH gradient (interior alkaline) was imposed. Rapid efflux of [U-(14)C]d-glucose from preloaded everted vesicles could also be initiated by activating the enzyme, with ATP. Taken together, the plasma membrane Mg2+-ATPase has been identified as an electroneutral H+/K+ antiporter with some properties reminiscent of the gastric H+,K+-ATPase. This enzyme is possibly involved in active accumulation of glucose via a H+-glucose symport system and in K+ accumulation.     

Differential effects of monovalent, divalent and trivalent metal ions on rat brain hexokinase

The effects of monovalent (Li+, Cs+) divalent (Cu2+, Ca2+, Sr2+, Ba2+, Zn2+, Cd2+, Hg2+, Pb2+, Mn2+, Fe2+, Co2+, Ni2+) and trivalent (Cr3+, Fe3+, Al3+) metals ions on hexokinase activity in rat brain cytosol were compared at 500 microM. The rank order of their potency as inhibitors of brain hexokinase was: Cr3+ (IC50 = 1.3 microM) greater than Hg2+ = Al3+ greater than Cu2+ greater than Pb2+ (IC50 = 80 microM) greater than Fe3+ (IC50 = 250 microM) greater than Cd2+ (IC50 = 540 microM) greater than Zn2+ (IC50 = 560 microM). However, at 500 microM Co2+ slightly stimulated brain hexokinase whereas the other metal ions were without effect. That inhibition of brain glucose metabolism may be an important mechanism in the neurotoxicity of metals is suggested.     

Hydrolysis and cytotoxic properties of osmium(II)/(III)-DMSO-azole complexes. Short communication

The antiproliferative properties of the osmium(II) complexes cis,fac-[Os(II)Cl(2)(DMSO)(3)(L)] and trans,cis,cis-[Os(II)Cl(2)(DMSO)(2)(L)(2)] (L = 1H-pyrazole, 1H-imidazole) were studied in three human cancer cell lines, namely 41M (ovary), SK-BR-3 (breast), and SW480 (colon). Their activities were compared with those of osmium(III) and ruthenium(III) NAMI-A type complexes on HT-29 (colon) and SK-BR-3 cancer cell lines. While IC(50) values of all the Os(II) complexes were found to be >1000 microM in all cell lines, Os and Ru-NAMI-A type complexes showed remarkable antiproliferative activity. The marginal in vitro cytotoxicity of the Os(II) compounds is presumably attributed to their resistance to hydrolysis. However, the Os-NAMI-A complexes, which are also kinetically stable in aqueous solution, showed reasonable antiproliferative activity in vitro when compared with the analogous Ru compounds and with the Os(II)-DMSO-azole species, indicating that hydrolysis might be not a prerequisite for the antitumor activity of Os-NAMI-A type complexes.     

A calcium-selective channel from root-Tip endomembranes of garden cress

A Ca2+ channel from root-tip endomembranes of garden cress (Lepidium sativum L.) (LCC1) was characterized using the planar lipid-bilayer technique. Investigation of single-channel recordings revealed that LCC1 is voltage gated and strongly rectifying. In symmetrical 50 mM CaCl2 solutions, the single-channel conductance was 24 picosiemens. LCC1 showed a moderate selectivity for Ca2+ over K+ (9.4:1) and was permeable for a range of divalent cations (Ca2+, Ba2+, and Sr2+). In contrast to Bryonia dioica Ca2+ channel 1, a Ca2+-selective channel from the endoplasmic reticulum of touch-sensitive tendrils, LCC1 showed no bursting channel activity and had a low open probability and mean open time (2.83 ms at 50 mV). Inhibitor studies demonstrated that LCC1 is blocked by micromolar concentrations of erythrosin B (inhibitor concentration for 50% inhibition [IC50] = 1. 8 μM) and the trivalent cations La3+ (IC50 = 5 μM) and Gd3+ (IC50 = 10 μM), whereas verapamil showed no blocking effect. LCC1 may play an important role in the regulation of the cytoplasmic free Ca2+ concentration in root-tip and/or root-cap cells. The question of whether this ion channel is part of the gravitropic signal transduction pathway deserves further investigation.     

Electrophilic abstractions on Os(H)2X(NO)L2: selective access to new unsaturated cationic Os(II) hydrides

Nitrosylation of Os(H)₃ClL₂ (L = P¹Pr₃) affords the known Os(H)₂Cl(NO)L₂ (2). Soft electrophiles (Ag⁻, Na⁻) react with complex 2 by chloride abstraction to ultimately yield truly 16-electron dihydride Os(H)₂(NO)(P¹Pr₃)₂⁻ (4a), characterized by variable-temperature NMR. Complex 4a reversibly binds H₂, forming Os(H)₂(H₂)(NO)(P¹Pr₃)⁻ with an unusually high barrier for intramolecular hydride exchange. Under kinetic conditions, protonation of 2 with strong acids follows the selectivity for chloride abstraction. Thermodynamically, protonation at the hydride is preferred, quantitatively producing cationic OsHCl(NO)L₂⁺, isolated and characterized by X-ray diffraction as the BAr₄^F− salt (7) (Ar^F=3,5−(CF₃)₂C₆H₃). Structures of isoelectronic OsHCl(NO)(PH₃)₂ and OsHCl(CO)(PH₃)₂ were optimized with ab initio DFT (Becke3LYP) methods and compared to show the greater unsaturation of the metal in the cationic species. Both complexes, 4a and 7, are highly electrophilic and reversibly coordinate dichloromethane in solution. The observed reactivity patterns of the synthesized unsaturated hydrides are rationalized in terms of the determining influence of the ‘push-pull’ π-stabilization of the metal center.     

Synthesis and anti-amoebic activity of gold(I), ruthenium(II), and copper(II) complexes of metronidazole

A series of Au, Ru, and Cu complexes of metronidazole (= [1-(2-hydroxyethyl)-2-methyl-5-nitro-1H-imidazole; 1) were prepared as highly potent anti-amoebic drugs. The complexes [Au(PPh3)(1)]PF6 (2), [Ru(1)2(Cl)2(H2O)2] (3), and [Cu(1)2(mu-Cl)(H2O)]2Cl2 (4) were readily synthesized from [Au(PPh3)Cl], RuCl3 x 3 H2O, and CuCl2 x 2 H2O, respectively. All complexes were thoroughly characterized by IR, UV/VIS, 1H-NMR, FAB-MS, elemental and thermogravimetric analyses, and, in the case of 4, also by X-ray crystallography (Fig. 1). All complexes were evaluated in vitro as growth inhibitors of Entamoeba histolytica (HM1:IMSS strain). Their IC50 values were in the range of 0.10-0.51 microM (Table 2), which makes these drugs, especially the Cu(II) complex 4, considerably more potent than uncomplexed metronidazole (1; IC50 = 1.81 microM), the current standard drug for the worldwide treatment of amoebiasis.     

Synthesis and spectroscopic properties of reconstituted zinc-myoglobin appending a DNA-binding platinum(II) complex

A reconstituted zinc-myoglobin (ZnMb) dyad, ZnMb-[Pt(bpy)(en)]2+, has been prepared by incorporating chemically-modified zinc-porphyrin, being capable of DNA-binding of the Pt complex, [Pt(bpy)(en)]2+, where bpy and en are 2,2'-bipyridine and ethylenediamine, respectively. The steady-state fluorescence of the cofactor, [Pt(bpy)(mu-enPP)Zn]Cl2, in MeOH indicates that the excited singlet state of zinc--porphyrin was almost quenched, probably because of the strong hydrophobic and pi-pi stacking interactions between the [Pt(bpy)(mu-enPP)Zn]2+ ions. In the reconstituted ZnMb-[Pt(bpy)(en)]2+, the quenching reaction of 1(ZnMb)* with the [Pt(bpy)(en)]2+ moiety does not occur, indicating apo-Mb matrix is essential. On the other hand, when the [Pt(bpy)(en)]2+ moiety was excited, the enhancement of the fluorescence from ZnMb unit was observed. It is suggested that the energy transfer from (1)([Pt(bpy)(en)]2+)* to ZnMb occurs. The spectroscopic changes of ZnMb-[Pt(bpy)(en)]2+ in the presence of calf-thymus DNA were also provided. Soret band at 428 nm gradually decreased, and isosbestic points at 321, 414, and 432 nm were observed with increasing the DNA concentration. When the Pt(II) moiety was excited at lambda(ex) 321 nm, the fluorescence signal around 600 nm similarly decreased. The synthetic manipulation of ZnMb by using a DNA-binding Pt(II) complex demonstrates sensitive fluorescent signal for DNA and valuable information to study photoinduced electron transfer within a Mb-DNA complex.     

The unexpected reactions of [RuCl3(2mqn)NO] (H2mqn=2-methyl-8-quinolinol) with 2-chloro-8-quinolinol (H2cqn) and of [RuCl(2cqn)(2mqn)NO] on photoirradiation

The reaction of [RuCl₃(2mqn)NO]⁻ (H2mqn=2-methyl-8-quinolinol) with 2-chloro-8-quinolinol (H2cqn) afforded cis-1 [RuCl(2cqn)(2mqn)NO] (the oxygen of 2cqn is trans to the NO) (complex 1), cis-1 [RuCl(2cqn)(2mqn)NO] (the oxygen of 2mqn is trans to the NO) (complex 2) and a 1:1 mixture of cis-2 [RuCl(2cqn)(2mqn)NO] (the oxygen of 2mqn is trans to the NO) and cis-2 [RuCl(2cqn)(2mqn)NO] (the oxygen of 2cqn is trans to the NO) (complex 3). The reaction was compared with that of [RuCl₃(2mqn)NO]⁻ with 8-quinolinol (Hqn) or 5-chloro-8-quinolinol (H5cqn). Photoirradiation reaction of complex 1 at room temperature in deaerated CH₂Cl₂ in the presence of NO gave trans-[RuCl(2cqn)(2mqn)NO] (the Cl is trans to the NO) and complex 2 with recovery of complex 1. The reaction was contrasted with that of cis-1 [RuCl(qn)(2mqn)NO] or cis-1 [RuCl(5cqn)(2mqn)NO]. The crystal structure of complex 1 was determined by X-ray diffraction. The reactions were examined under consideration of atomic charge of the phenolato oxygen in 8-quinolinol and its derivatives calculated at the restricted Hartree-Fock/6-311G** level.     

Zinc potentiation of androgen receptor binding to nuclei in vitro

Zn2+ potentiates binding of the 4.5S [3H]dihydrotestosterone-receptor complex to isolated rat prostate Dunning tumor nuclei in vitro when assayed in the presence of 300 microM ZnCl2, 3 mM MgCl2, 0.25 M sucrose, 5 mM mercaptoethanol, 0.15 M KCl, and 50 mM tris(hydroxymethyl)aminomethane, pH 7.5. In the presence of 5 mM mercaptoethanol, the concentration of 50 microM total Zn2+ required to promote half-maximal receptor binding to nuclei corresponds to a free Zn2+ concentration of 50 nM. The receptor-nuclear interaction appears to be selective for Zn2+; other divalent cations when added at a concentration of 1 mM to a buffer containing 5 mM mercaptoethanol are less effective (Ni2+) or have essentially no effect (Ca2+, Mg2+, Mn2+, Co2+, Cu2+, and Cd2+). Zn2+ does not alter the sedimentation rate of the 4.5S [3H]dihydrotestosterone receptor in the presence of mercaptoethanol; however, in the absence of mercaptoethanol, Zn2+ causes the receptor to aggregate. Zn2+-dependent nuclear binding of the 4.5S [3H]dihydrotestosterone receptor is saturable at 1.4 X 10(-13) mol of receptor sites/mg of DNA, corresponding to approximately 1150 sites/nucleus. In the presence of excess nuclei, up to 60% of added receptor is nuclear bound. An apparent binding constant for the receptor-nuclear interaction of 10(13) M-1 was approximated. Pyridoxal 5'-phosphate (less than or equal to 10 mM), but not 0.4 M KCl, inhibits Zn2+-dependent nuclear binding of the [3H]dihydrotestosterone receptor. Up to 66% of nuclear-bound receptor can be extracted in buffer containing 3 mM ethylenediaminetetraacetic acid plus either 0.4 M KCl or 10 mM pyridoxal 5'-phosphate.(ABSTRACT TRUNCATED AT 250 WORDS)     

Synthesis and in vitro antitumoral activity of novel O,O'-di-2-alkyl-(S,S)-ethylenediamine-N,N'-di-2-propanoate ligands and corresponding platinum(II/IV) complexes

Syntheses of two novel ligand precursors O,O'-diisopropyl- (1a) and O,O'-diisobutyl-(S,S)-ethylenediamine-N,N'-di-2-propanoate dihydrochloride monohydrate (1b) and the corresponding dichloroplatinum(II) (2a and 2b) and tetrachloroplatinum(IV) complexes (3a and 3b) are described here. The substances were characterized by IR, (1)H and (13)C spectroscopy and elemental analysis. Crystal structures were determined for 1a and the corresponding platinum(IV) complex, 3a. In vitro antiproliferative activity was determined against tumor cell lines: human adenocarcinoma HeLa, human myelogenous leukemia K562, human malignant melanoma Fem-x, rested and stimulated normal immunocompetent cells (human peripheral blood mononuclear PBMC cells) using KBR test (Kenacid Blue Dye binding test). The IC(50)(microM) values for the most active compound 3a were: 30.48+/-2.54; 12.26+/-2.60; 13.68+/-3.22; 80.18+/-24.07 and 71.30+/-21.70, respectively.     

Preliminary in vitro studies on two potent, water-soluble trimethoprim analogues with exceptional species selectivity against dihydrofolate reductase from Pneumocystis carinii and Mycobacterium avium

2,4-Diamino-5-[3',4'-dimethoxy-5'-(5-carboxy-1-pentynyl)]benzylpyrimidine (6) and 2,4-diamino-5-[3',4'-dimethoxy-5'-(4-carboxyphenylethynyl)benzylpyrimidine (7) were synthesized from 2,4-diamino-5-(5'-iodo-3',4'-dimethoxybenzyl)pyrimidine (9) via a Sonogashira reaction with appropriate acetylenic esters followed by saponification, and were tested as inhibitors of dihydrofolate reductase (DHFR) from Pneumocystis carinii (Pc), Toxoplasma gondii (Tg), Mycobacterium avium (Ma), and rat in comparison with the widely used antibacterial agent 2,4-diamino-5-(3',4',5'-trimethoxybenzyl)pyrimidine (trimethoprim, TMP). The selectivity index (SI) for each compound was calculated by dividing its 50% inhibitory concentration (IC(50)) against rat DHFR by its IC(50) against Pc, Tg, or Ma DHFR. The IC(50) of 6 against Pc DHFR was 1.0 nM, with an SI of 5000. Compound 7 had an IC(50) of 8.2 nM against Ma DHFR, with an SI of 11000. By comparison, the IC(50) of TMP was 12000 nM against Pc, 300 nM against Ma, and 180000 against rat DHFR. The potency and selectivity values of 6 and 7 were not as high against Tg as they were against Pc or Ma DHFR, but nonetheless exceeded those of TMP. Because of the outstanding selectivity of 6 against Pc and of 7 against Ma DHFR, these novel analogues may be viewed as promising leads for further structure-activity optimization.     

Comparative studies of coordination properties of puromycin and puromycin aminonucleoside towards copper(II) ions

Protonation equilibria of puromycin (PM) and puromycin aminonucleoside (PAN) and their coordination by copper(II) ion were studied in solution by potentiometry, electronic absorption spectroscopy (UV-Vis), circular dichroism (CD), electron paramagnetic resonance (EPR) and mass spectrometry. For puromycin four mononuclear complexes were found, with stoichiometries Cu(PM)2+, CuH(-1)(PM)+, CuH(-2)(PM) and CuH(-3)(PM)(-). In each of them the Cu(II) ion was bound in the peptidic-like manner, the differences of stoichiometries are a consequence of subsequent deprotonations of the sugar C2'-OH group and the coordinated water molecule. The coordination mode for puromycin aminonucleoside was aminosugar-like. Two dimeric complexes, Cu2H(-1)(PAN)2(2+) and Cu2H(-2)(PAN)2+, and one monomeric CuH(-2)(PAN)2 were found. The N6,N6-dimethyladenine moiety of PAN was not involved in the coordination process due to steric hindrance.