Mechanism of A beta(1-40) fibril-induced fluorescence of (trans,trans)-1-bromo-2,5-bis(4-hydroxystyryl)benzene (K114)

K114, (trans,trans)-1-bromo-2,5-bis(4-hydroxystyryl)benzene, is a fluorescent Congo Red analogue that binds tightly to amyloid fibrils, but not the monomeric proteins, with a concomitant enhancement in fluorescence. The mechanism for the low aqueous fluorescence and the subsequent enhancement by A beta(1-40) fibrils was investigated by fluorescence spectroscopy and binding analysis. K114's unusually low buffer fluorescence is due to self-quenching in sedimentable aggregates or micelles which upon interacting with amyloid fibrils undergo an enhancement in fluorescence intensity and shifts in the excitation and emission spectra. These spectral changes are suggestive of a stabilization of the phenolate anion, perhaps by hydrogen bonding, rather than an increase in the microenvironment dielectric constant or dye immobilization. 1,4-Bis(4-aminophenylethenyl)-2-methoxybenzene, which lacks the phenol moiety, and X-34, which contains a stabilized phenol (pK approximately 13.4), do not display the phenolate anion fluorescence in the presence of fibrils. The apparent affinity of K114 for fibril binding is 20-30 nM with a stoichiometry of 2.2 mol of K114/mol of A beta(1-40) monomer. Competition studies indicate that K114 and Congo Red share a site, but K114 does not bind to sites on A beta(1-40) fibrils for neutral benzothiazole (BTA-1), cationic thioflavin T, or the hydrophobic (S)-naproxen and (R)-ibuprofen molecules. Comparison of benzothiazole binding stoichiometry which has been suggested to reflect disease-relevant amyloid structures to that of Congo Red analogues which reflect total fibril content may be useful in defining biologically pertinent conformational forms of amyloid.     

Synthesis of (1R,2S)-1-(3'-chloro-4' -methoxyphenyl)-1,2-propanediol (trametol) and (1R,2S)-1-(3',5'-dichloro-4'-methoxyphenyl)-1,2-propanediol,chlorinated fungal metabolites in the natural environment

(1R,2S)-1-(3'-Chloro-4'-methoxyphenyl)-1,2propanediol (Trametol, 3), a metabolite of the fungus Trametes sp. IVP-F640 and Bjerkandera sp. BOS55, was synthesized by employing Sharpless asymmetric dihydroxylation as the key step. Similarly, the (1R,2S)-isomer of 1-(3',5'-dichloro-4'-methoxyphenyl)-1,2-propanediol (4), another metabolite of Bjerkandera sp. BOS55, was synthesized by asymmetric dihydroxylation.     

Synthesis of some new antibacterial active cadmium and mercury complexes of 4-(3-(2-(4-(dimethyl aminophenyl allylidene aminopropyl-imino)prop-1-ethyl)-N,N-dimethyl benzene amine

Abstract

A series of novel cadmium(II) and mercury(II) halide and thiocyanate complexes with an asymmetric Schiff base ligand of 4-(3-(2-(4-(dimethyl aminophenyl allylidene aminopropyl-imino)prop-1-ethyl)-N,N-dimethyl benzene amine has been synthesised and characterised using spectral, physical and analytical data, such as ¹H NMR, UV-Vis and FTIR spectroscopy, melting point, elemental analysis and molar conductivity measurements. The spectral and physical data proposed a pseudo-tetrahedral geometry around the metal centre in the metal complexes. Moreover, the in vitro antibacterial activity of all compounds was assayed against two gram-positive and two gram-negative bacterial strains by a disk diffusion method and the results showed that all compounds have antibacterial characteristics. Also, the minimum inhibitory concentration and minimum bactericidal concentration of each compound were determined.     

2(S)-(Cycloalk-1-enecarbonyl)-1-(4-phenyl-butanoyl)pyrrolidines and 2(S)-(aroyl)-1-(4-phenylbutanoyl)pyrrolidines as prolyl oligopeptidase inhibitors

In order to replace the P2-P1 amide group, different 1-cycloalkenyls and 2-aryls were studied in the place of the P1 pyrrolidine group of a 4-phenylbutanoyl-L-Pro-pyrrolidine structure, which is a well-known prolyl oligopeptidase inhibitor SUAM-1221. The 1-cyclopentenyl and the 2-thienyl groups gave novel compounds, which were equipotent with the corresponding pyrrolidine-analog SUAM-1221. It was shown that the P2-P1 amide group of POP inhibitors can be replaced by an alpha,beta-unsaturated carbonyl group or the aryl conjugated carbonyl group.     

1-(4-Methane(amino)sulfonylphenyl)-3-(4-substituted-phenyl)-5-(4-trifluoromethylphenyl)-1H-2-pyrazolines/pyrazoles as potential anti-inflammatory agents

2-Pyrazolins 14a–l and pyrazoles 15a–l were designed as celecoxib analogs for the evaluation of their in vitro COX-1/COX-2 inhibitory activity and the in vivo anti-inflammatory activity. Compounds 14i, 15a, 15d and 15f were the most COX-2 selective derivatives (S.I. = 5.93, 6.08, 5.03 and 5.27 respectively) while the pyrazoline derivatives 14g and 14i exhibited the highest AI activity (ED₅₀ = 190.5 and 160.1 μmol/kg po, respectively).     

Expression of proteinase-activated receptor 1-4 (PAR 1-4) in human cancer

Proteinase activated receptors (PAR 1-4) are membrane receptors with a unique way of activation by proteinases like thrombin, trypsin and matrix metalloproteinases which lead to a specific cellular response. To evaluate the significance of expression and co-expression of PAR in cancer we performed a survey on published data. A Pubmed literature search on “PAR, thrombin, cancer” was performed and 46 publications were selected for systematic review based on the availability of information on tumor type, material type, detection method and specification of positive cases. PAR-1 was found in 77.3% of malignant samples (n = 678), PAR-2 in 79.5% (n = 592), PAR-3 in 12.6% (n = 87) and PAR-4 in 54.9% (n = 153). PAR-1 and -2 were present in adenocarcinomas, melanomas, osteosarcomas, glioblastomas, meningiomas, leukaemias and squamous cell carcinomas. Presence of PAR-3 was limited to kidney and liver cancer. The data on PAR-4 expression was inconclusive. Those studies analysing PAR-1 and PAR-2 reported coexpression of the two receptors. PAR-1 and -2 are widely expressed in human tumors suggesting an important role in tumorigenesis and providing potential targets for therapy. PAR-3 and PAR-4 are less frequently detectable, their expression and potential role in tumorigenesis require further investigation.     

Triazole analog 1-(1-benzyl-5-(4-chlorophenyl)-1H-1,2,3-triazol-4-yl)-2-(4-bromophenylamino)-1-(4-chlorophenyl)ethanol induces reactive oxygen species and autophagy-dependent apoptosis in both in vitro and in vivo breast cancer models

Autophagy is considered as an important cell death mechanism that closely interacts with other common cell death programs like apoptosis. Critical role of autophagy in cell death makes it a promising, yet challenging therapeutic target for cancer. We identified a series of 1,2,3-triazole analogs having significant breast cancer inhibition property. Therefore, we attempted to study whether autophagy and apoptosis were involved in the process of cancer cell inhibition. The lead molecule, 1-(1-benzyl-5-(4-chlorophenyl)-1H-1,2,3-triazol-4-yl)-2-(4-bromophenylamino)-1-(4-chlorophenyl)ethanol (T-12) induced significant cell cycle arrest, mitochondrial membrane depolarization, apoptosis and autophagy in MCF-7 and MDA-MB-231 cells. T-12 increased reactive oxygen species and its inhibition by N-acetyl-l-cysteine protected breast cancer cells from autophagy and apoptosis. Autophagy inhibitor, 3-methyladenine abolished T-12 induced apoptosis, mitochondrial membrane depolarization and reactive oxygen species generation. This suggested that T-12 induced autophagy facilitated cell death rather than cell survival. Pan-caspase inhibition did not abrogate T-12 induced autophagy, suggesting that autophagy precedes apoptosis. In addition, T-12 inhibited cell survival pathway signaling proteins, Akt, mTOR and Erk1/2. T-12 also induced significant regression of tumor with oral dose of as low as 10 mg/kg bodyweight in rat mammary tumor model without any apparent toxicity. In presence of reactive oxygen species inhibitor (N-acetyl-l-cysteine) and autophagy inhibitor (chloroquine), T-12 induced tumor regression was significantly decreased. In conclusion, T-12 is a potent inducer of autophagy-dependent apoptosis in breast cancer cells both in vitro and in vivo and can serve as an important lead in development of new anti-tumor therapy.     

Synthesis and properties of cationic 2'-O-[N-(4-aminobutyl)carbamoyl] modified oligonucleotides

2'-O-[N-(4-Aminobutylcarbamoyl)]uridine (U(abcm)) was synthesized and incorporated into oligonucleotides. The oligonucleotides incorporating U(abcm) formed more stable duplexes with their complementary and mismatched RNAs than those containing 2'-O-carbamoyluridine (U(cm)). The stability of duplex with a U(abcm)-rG base pair showed higher thermostability than the duplex having unmodified U-rG base pair. The U(abcm) residue showed enhanced resistance to snake venome phosphodiesterase.     

Synthesis and in vitro antitumor activity of novel acylspermidine derivative N-(4-aminobutyl)-N-(3-aminopropyl)-8-hydroxy-dodecanamide (AAHD) against HepG2 cells

Naturally occurring polyamines like Putrescine, Spermidine, and Spermine are polycations which bind to the DNA, hence stabilizing it and promoting the essential cellular processes. Many synthetic polyamine analogues have been synthesized in the past few years, which have shown cytotoxic effects on different tumours. In the present study, we evaluated the antiproliferative effect of a novel, acylspermidine derivative, (N-(4-aminobutyl)-N-(3-aminopropyl)-8-hydroxy-dodecanamide) (AAHD) on HepG2 cells. Fluorescence staining was performed with nuclear stain (Hoechst 33342) and acridine orange/ethidium bromide double staining. Dose and the time-dependent antiproliferative effect were observed by WST-1 assays, and radical scavenging activity was measured by ROS. Morphological changes such as cell shrinkage & blebbing were analyzed by fluorescent microscopy. It was found that AAHD markedly suppressed the growth of HepG2 cells in a dose- and time-dependent manner. It was also noted that the modulation of ROS levels confirmed the radical scavenging activity. In the near future, AAHD can be a promising drug candidate in chalking out a neoplastic strategy to control the proliferation of tumour cells. This study indicated that AAHD induced anti-proliferative and pro-apoptotic activities on HCC. Since AAHD was active at micromolar concentrations without any adverse effects on the healthy cells (Fibroblasts), it is worthy of further clinical investigations.     

Anthelmintic profile of methyl 5(6)-(4-methylpiperidin-1-yl) carbonylbenzimidazole-2-carbamate in experimental helminthiases

Biological evaluation of methyl 5(6)-(4-methylpiperidin-1-yl) carbonylbenzimidazole-2-carbamate against Ancylostoma ceylanicum, Nippostrongylus brasiliensis, Syphacia obvelata, Hymenolepis nana, H. diminuta and Cysticercus fasciolaris in experimental animals is reported. The compound (mg/kg) causes 100% elimination of A. ceylanicum (25 x 1), N. brasiliensis (100 x 1), S. obvelata (50 x 1), H. nana (250 x 3) and C. fasciolaris (50 x 10). It was also effective against the developing larvae (L3, L4 and L5) of A. ceylanicum at a single oral dose of 100 mg/kg. Another study indicated that the compound elicits 100% response within 32 hr of drug administration. The drug is well tolerated and LD50 is greater than 4500 mg/kg.     

In vitro and in vivo pharmacological profile of 4-(4-fluorobenzylidene)-1-[2-[5-(4-fluorophenyl)-1H-pyrazol-4-yl] ethyl] piperidine (NRA0161)

Atypical antipsychotic properties of 4-(4-fluorobenzylidene)-1-[2-[5-(4-fluorophenyl)-1H-pyrazol-4-yl]ethyl] piperidine (NRA0161) were investigated by in vitro receptor affinities, in vivo receptor occupancies and findings were compared with those of risperidone and haloperidol in rodent behavioral studies. In in vitro receptor binding studies, NRA0161 has a high affinity for human cloned dopamine D(4) and 5-HT(2A) receptor with Ki values of 1.00 and 2.52 nM, respectively. NRA0161 had a relatively high affinity for the alpha(1) adrenoceptor (Ki; 10.44 nM) and a low affinity for the dopamine D(2) receptor (Ki; 95.80 nM). In in vivo receptor binding studies, NRA0161 highly occupied the 5-HT(2A) receptor in rat frontal cortex. In contrast, NRA0161 did not occupy the striatal D(2) receptor. In behavioral studies, NRA0161, risperidone and haloperidol antagonized the locomotor hyperactivity in mice, as induced by methamphetamine (MAP). At a higher dosage, NRA0161, risperidone and haloperidol dose-dependently antagonized the MAP-induced stereotyped behavior in mice and NRA0161 dose-dependently and significantly induced catalepsy in rats. The ED(50) value in inhibiting the MAP-induced locomotor hyperactivity was 30 times lower than that inhibiting the MAP-induced stereotyped behavior and 50 times lower than that which induced catalepsy.These findings suggest that NRA0161 may have atypical antipsychotic activities yet without producing extrapyramidal side effects.     

Dual DAT/sigma1 receptor ligands based on 3-(4-(3-(bis(4-fluorophenyl)amino)propyl)piperazin-1-yl)-1-phenylpropan-1-ol

Ester analogs of (+/-)3-(4-(3-(bis(4-fluorophenyl)amino)propyl)piperazin-1-yl)-1-phenylpropan-1-ol were synthesized and evaluated for binding at DAT, SERT, NET, and sigma1 receptors, and compared to GBR 12909 and several known sigma1 receptor ligands. Most of these compounds demonstrated high affinity (K(i)=4.3-51 nM) and selectivity for the DAT among the monoamine transporters. S- and R-1-(4-(3-(bis(4-fluorophenyl)amino)propyl)piperazin-1-yl)-3-phenylpropan-2-ol were also prepared wherein modest enantioselectivity was demonstrated at the DAT. However, no enantioselectivity at sigma1 receptors was observed and most of the ester analogs of the more active S-enantiomer showed comparable binding affinities at both DAT and sigma1 receptors with a maximal 16-fold DAT/sigma1 selectivity.     

Synthesis of novel 3-(1-(1-substituted piperidin-4-yl)-1H-1,2,3-triazol-4-yl)-1,2,4-oxadiazol-5(4H)-one as antifungal agents

A novel series of 1,2,3 triazole compounds possessing 1,2,4 oxadiazole ring were efficiently synthesized. Synthesized compounds were evaluated for their in vitro antifungal activities using standard cup plate method. SAR for the series has been developed by comparing their MIC values with miconazole and fluconazole. Compound 11a from the series was more potent than miconazole against Candida albicans (MIC-20) and Aspergillus flavus (MIC-10) whereas equipotent with miconazole against Fusarium oxysporum (MIC-25) and Aspergillus niger (MIC-12.5). Also compound 11h was more potent than miconazole against Candida albicans (MIC-20) and Aspergillus niger (MIC-10) and equipotent with miconazole against Fusarium oxysporum. Compound 11h was equipotent with fluconazole against Aspergillus niger (MIC-10).     

Synthesis and histamine H3 receptor activity of 4-(n-alkyl)-1H-imidazoles and 4-(omega-phenylalkyl)-1H-imidazoles

The influence of lipophilic moieties attached to a 4-1H-imidazole ring on the histamine H₃ receptor activity was systematically investigated. Series of 4-(n-alkyl)-1H-imidazoles and 4-(ω-phenylalkyl)-1H-imidazoles were prepared, with an alkyl chain varying from 2–9 methylene groups and from 1–9 methylene groups, respectively. The compounds were tested for their activity on the H₃ receptor under in vitro conditions. For the 4-(n-alkyl)-1H-imidazoles the activity is proportional to chain length, ranging from a pA₂ value of 6.3±0.2 for 4-(n-propyl)-1H-imidazole to a pA₂ value of 7.2±0.1 for 4-(n-decyl)-1H-imidazole. For the series 4-(ω-phenylalkyl)-4H-imidazoles an optimum in H₃ activity was found for the pentylene spacer: 4-(ω-phenylpentyl)-1H-imidazole has a pA₂ value of 7.8±0.1.     

Therapeutic effect of IL-13 immunoneutralization during chronic experimental fungal asthma

IL-13 and IL-4 are key contributors to the asthmatic phenotype. The temporal role of these cytokines in airway function, inflammation, and remodeling were assessed in a chronic murine model of Asperigillus fumigatus-induced allergic asthma. IL-13 and IL-4 protein levels were significantly elevated by 30 days after conidia challenge in A. fumigatus-sensitized mice. Furthermore, IL-13Ralpha1 mRNA expression was significantly elevated 7 days after conidia challenge and remained elevated until day 21. In contrast, IL-13Ralpha2 mRNA expression, although constitutively expressed in naive lung, was absent in the lungs of A. fumigatus-sensitized mice both before and after conidia challenge. Membrane-bound IL-4R mRNA expression was significantly elevated 7 days after conidia challenge; however, soluble IL-4R mRNA expression was increased 30 days after conidia challenge. Immunoneutralization of IL-13 between days 14 and 30 or days 30 and 38 after fungal sensitization and challenge significantly attenuated airway hyperresponsiveness, collagen deposition, and goblet cell hyperplasia at day 38 after conidia challenge; however, the effects of IL-4 immunoneutralization during the same time periods were not as marked. IFN-gamma and IL-12 release after Aspergillus Ag restimulation was elevated from spleen cells isolated from mice treated with IL-4 anti-serum compared with IL-13 anti-serum or normal rabbit serum-treated mice. This study demonstrates a pronounced therapeutic effect of IL-13-immunoneutralization at extended time points following the induction of chronic asthma. Most importantly, these therapeutic effects were not reversed following cessation of treatment, and IL-13 anti-serum treatment did not alter the systemic immune response to Ag restimulation, unlike IL-4 immunoneutralization. Therefore, IL-13 provides an attractive therapeutic target in allergic asthma.     

Transient-state kinetics of the reactions of 1-methoxy-4-(methylthio)benzene with horseradish peroxidase compounds I and II

Transient-state reactions of horseradish peroxidase compounds I and II with 1-methoxy-4-(methylthio)benzene (a para-substituted thioanisole) were studied over the pH range from 3.4 to 10.5. The pH-jump technique was applied to the compound II reactions at pH values below 8.6. The reactions of both compound I and compound II with the para-substituted thioanisole consisted predominantly of an initial burst. The burst was followed by a steady-state phase that became more obvious at lower concentrations of the thioanisoles. The burst phase for both compounds I and II can be explained in terms of two independent transient-state reactions with 1-methoxy-4-(methylthio)benzene as follows: (i) a single reaction of compound I (or compound II) with the substrate and (ii) the formation of a complex between compound I or II and the substrate followed by reaction of the productive complex with another molecule of sulfide. The overall rate of reaction path ii is faster than that of path i. The preference for path i or ii is highly dependent upon the concentration of sulfide with step ii favored at higher sulfide concentrations. The experimental results obtained on the overall reaction under both pseudo-first-order and single-turnover conditions indicate that compound II reacts competitively with both the organic sulfide substrate and the sulfur cation radical produced from compound I oxidation of sulfide.     

1-(4-Phenylpiperazin-1-yl)-2-(1H-pyrazol-1-yl)ethanones as novel CCR1 antagonists

A novel series of CCR1 antagonists based on the 1-(4-phenylpiperazin-1-yl)-2-(1H-pyrazol-1-yl)ethanone scaffold was identified by screening a compound library utilizing CCR1-expressing human THP-1 cells. SAR studies led to the discovery of the highly potent and selective CCR1 antagonist 14 (CCR1 binding IC₅₀ = 4 nM using [¹²⁵I]-CCL3 as the chemokine ligand). Compound 14 displayed promising pharmacokinetic and toxicological profiles in preclinical species.     

Discovery of a novel sub-class of ROMK channel inhibitors typified by 5-(2-(4-(2-(4-(1H-Tetrazol-1-yl)phenyl)acetyl)piperazin-1-yl)ethyl)isobenzofuran-1(3H)-one

A sub-class of distinct small molecule ROMK inhibitors were developed from the original lead 1. Medicinal chemistry endeavors led to novel ROMK inhibitors with good ROMK functional potency and improved hERG selectivity. Two of the described ROMK inhibitors were characterized for the first in vivo proof-of-concept biology studies, and results from an acute rat diuresis model confirmed the hypothesis that ROMK inhibitors represent new mechanism diuretic and natriuretic agents.