Design,synthesis and evaluation of quinolinone derivatives containing dithiocarbamate moiety as multifunctional AChE inhibitors for the treatment of Alzheimer’s disease

Abstract

A series of novel quinolinone derivatives bearing dithiocarbamate moiety were designed and synthesised as multifunctional AChE inhibitors for the treatment of AD. Most of these compounds exhibited strong and clearly selective inhibition to eeAChE. Among them, compound 4c was identified as the most potent inhibitor to both eeAChE and hAChE (IC₅₀ = 0.22?μM for eeAChE; IC₅₀ = 0.16?μM for hAChE), and it was also the best inhibitor to AChE-induced Aβ aggregation (29.02% at 100?μM) and an efficient inhibitor to self-induced Aβ aggregation (30.67% at 25?μM). Kinetic and molecular modelling studies indicated that compound 4c was a mixed-type inhibitor, which could interact simultaneously with the catalytic anionic site (CAS) and the peripheral anionic site (PAS) of AChE. In addition, 4c had good ability to cross the BBB, showed no toxicity on SH-SY5Y neuroblastoma cells and was well tolerated in mice at doses up to 2500?mg/kg (po).     

N-aryl 2-aryloxyacetamides as a new class of fatty acid amide hydrolase (FAAH) inhibitors

Fatty acid amide hydrolase (FAAH) is a promising target for the development of drugs to treat neurological diseases. In search of new FAAH inhibitors, we identified 2-(4-cyclohexylphenoxy)-N-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide, 4g, with an IC₅₀ of 2.6?µM as a chemical starting point for the development of potent FAAH inhibitors. Preliminary hit-to-lead optimisation resulted in 2-(4-phenylphenoxy)-N-(3-(oxazolo[4,5-b]pyridin-2-yl)phenyl)acetamide, 4i, with an IC₅₀ of 0.35?µM.     

Synthesis,antiviral activity and structure–activity relationship of 1-(1-aryl-4,5-dihydro-1H-imidazoline)-3-chlorosulfonylureas and products of their cyclization

Novel 1-(1-aryl-4,5dihydro-1H-imidazoline)-3-chlorosulfonylourea derivatives 3a–3f were synthesized in the reaction of 1-aryl-4,5-dihydro-1H-imidazol-2-amines with chlorosulfonyl isocyanate. The second series of compounds 4a–4f was prepared from the respective 1-(1-aryl-4,5-dihydro-1H-imidazoline)-3-chlorsulfonylureas 3a–3f and 1,1′-carbonyldiimidazole (CDI). The selected compounds were tested for their activity against Herpes simplex virus and coxsackievirus B3 (CVB3). It was determined that three derivatives, i.e 3d, 4a and 4d are active against Herpes simplex virus (HSV-1). Compounds 3d and 4c are active against CVB3. Their favorable activity can be primarily attributed to their low lipophilicity values. Moreover, the lack of substituent in the phenyl moiety or 4-methoxy substitution can be considered as the most beneficial for the antiviral activity.     

Synthesis of 3-Aminoimidazo[4,5-c]pyrazole Nucleoside via the N-N Bond Formation Strategy as a [5:5] Fused Analog of Adenosine

3-Amino-6-(β-D-ribofuranosyl)imidazo[4,5-c]pyrazole (2) was synthesized via an N-N bond formation strategy by a mononuclear heterocyclic rearrangement (MHR). A series of 5-amino-1-(5-O-tert-butyldimethylsilyl-2,3-O-isopropylidene-β-D-ribofuranosyl-4-(1,2,4-oxadiazol-3-yl)imidaz-oles (6a-d), with different substituents at the 5-position of the 1,2,4-oxadiazole, were synthesized from 5-amino-1-(β-D-ribofuranosyl)imidazole-4-carboxamide (AICA Ribose, 3). It was found that 5-amino-1-(5-O-tert-butyldimethylsilyl-2,3-O-isopropylidene-β-D-ribofuranosyl)-4-(5-methyl-1,2,4-oxadiazol-3-yl)imidazole (6a) underwent the MHR with sodium hydride in DMF or DMSO to afford the corresponding 3-acetamidoimidazo[4,5-c]pyrazole nucleoside(s) (7b and/or 7a) in good yields. A direct removal of the acetyl group from 3-acetamidoimidazo[4,5-c]pyrazoles under numerous conditions was unsuccessful. Subsequent protecting group manipulations afforded the desired 3-amino-6-(β-D-ribofuranosyl)imidazo[4,5-c]pyrazole (2) as a 5:5 fused analog of adenosine (1).     

Design and synthesis of 3-pyrrol-3-yl-3H-isobenzofuran-1-ones as inhibitors of human cytosolic phospholipase A2α

A series of 3-pyrrol-3-yl-3H-isobenzofuran-1-ones was synthesized and assessed for the ability to inhibit cytosolic phospholipase A₂α (cPLA₂α). Several of these compounds were found to be active in both a cell based assay and an isolated enzyme assay. The most potent inhibitor was the thiazolidine-2,4-dione substituted derivative 35. With IC₅₀-values of 0.7 μM and 7.3 μM in the cellular and isolated enzyme assay, respectively, it possesses similar inhibitory potency as the known cPLA₂α inhibitor arachidonyltrifluoromethyl ketone (AACOCF₃). Structure–activity relationship studies revealed that the evaluated isobenzofuran-1-ones seem to exert their cellular activities not only by a direct interaction with the enzyme but also by other as yet unknown mechanisms.     

Analogues of Azepinomycin as Inhibitors of Guanase

Abstract

Synthesis and biochemical screening against guanase of analogues of the naturally occurring guanase inhibitor azepinomycin (2) are reported. Compound e-amino-5,6,7,8,-tetrahydro-4H-imidazo[4,5-e][1,4]diazepine-5,8-dione (3) was synthesized in six steps commencing with 1-benzyl-5-nitroimidazole-4-carboxylic acid (5). Compound 3 and its synthetic precursor 3-benzyl-6-(N-benzyloxycarbonyl)amino-5,6,7,8-tetrahydro-4H-imidazo[4,5-e][1,4]diazepine-5,8-dione (12) were screened against rabbit liver guanase. Both were found to be moderate inhibitors of the enzyme with K₁′s in the range of 10^?4 M.     

α-Glucosidase inhibitory activity and cytotoxic effects of some cyclic urea and carbamate derivatives

The inhibitory activities of selected cyclic urea and carbamate derivatives (1–13) toward α-glucosidase (α-Gls) in in vitro assay were examined in this study. All examined compounds showed higher inhibitory activity (IC₅₀) against α-Gls compared to standard antidiabetic drug acarbose. The most potent was benzyl (3,4,5-trimethoxyphenyl)carbamate (12) with IC₅₀?=?49.85?±?0.10?µM. In vitro cytotoxicity of the investigated compounds was tested on three human cancer cell lines HeLa, A549 and MDA-MB-453 using MTT assay. The best antitumour activity was achieved with compound 2 (trans-5-phenethyl-1-phenylhexahydro-1H-imidazo[4,5-c]pyridin-2(3H)-one) against MDA-MB-453 human breast cancer cell line (IC₅₀?=?83.41?±?1.60?µM). Cyclic ureas and carbamates showed promising anti-α-glucosidase activity and should be further tested as potential antidiabetic drugs. The PLS model of preliminary QSAR study indicated that, in planing the future synthesis of more potent compounds, the newly designed should have the substituents capable of polar interactions with receptor sites in various positions, while avoiding the increase of their lipophilicity.     

Design,synthesis and biological evaluation of 2-aminopyrimidinones and their 6-aza-analogs as a new class of CK2 inhibitors

Abstract

In order to find the new potent CK2 inhibitors the 60 derivatives of 2-aminopyrimidinone and their 6-aza-substituted analogs were synthesized and tested in vitro. Among them, the most efficient inhibitor 2-hydroxy-5-[4-(4-methoxyphehyl)-6-oxo-1,6-dihydropyrimidin-2-ylamino] benzoic acid was identified (IC₅₀?=?1.1?μM). The structure--activity relationship study of newly synthesized derivatives was carried out and their binding mode with adenosine triphosphate-acceptor site of CK2 was proposed.     

Design,synthesis and docking study of novel coumarin ligands as potential selective acetylcholinesterase inhibitors

New coumaryl-thiazole derivatives with the acetamide moiety as a linker between the alkyl chains and/or the heterocycle nucleus were synthesized and in vitro tested as acetylcholinesterase (AChE) inhibitors. 2-(diethylamino)-N-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)acetamide (6c, IC₅₀ value of 43?nM) was the best AChE inhibitor with a selectivity index of 4151.16 over BuChE. Kinetic study of AChE inhibition revealed that 6c was a mixed-type inhibitor. Moreover, the result of H4IIE hepatoma cell toxicity assay for 6c showed negligible cell death. Molecular docking studies were also carried out to clarify the inhibition mode of the more active compounds. Best pose of compound 6c is positioned into the active site with the coumarin ring wedged between the residues of the CAS and catalytic triad of AChE. In addition, the coumarin ring is anchored into the gorge of the enzyme by H-bond with Tyr130.     

4,5-di-O-caffeoylquinic acid methyl ester isolated from Lonicera japonica Thunb. targets the Keap1/Nrf2 pathway to attenuate H2O2-induced liver oxidative damage in HepG2 cells

Background4,5-di-O-caffeoylquinic acid methyl ester (4,5-CQME) is a caffeoylquinic acid (CQA) isolated from Lonicera japonica Thunb., a traditional Chinese medicine. To date, the biological activity of 4,5-CQME has not been fully investigated.PurposeThe aim of the current study was to explore the anti-oxidative activity and the underlying mechanism of 4,5-CQME.MethodsMTT assay was used to evaluate the cytoprotective effect of 4,5-CQME. DCFH-DA was used as a fluorescence probe to detect intracellular ROS. The mitochondrial membrane potential was detected using the fluorescent probe JC-1. MDA and GSH levels were measured using MDA and GSH commercial kits, respectively. Apoptosis assay was performed using the Annexin V-FITC/PI method. The functional mechanism of 4,5-CQME was investigated by analyzing relative signaling pathways through immunofluorescent staining, quantitative PCR and western blot analysis.ResultsHepG2 cells were incubated with different concentrations of 4,5-CQME for 12 h before exposure to 500 μM H₂O₂ for 3 h. 4,5-CQME attenuated H₂O₂-induced oxidative damage and had a higher cytoprotective effect than 3-caffeoylquinic acid, 3-caffeoylquinic acid methyl ester, or 4,5-di-O-caffeoylquinic acid. 4,5-CQME also reduced ROS and MDA levels and rescued GSH depletion. Western blots demonstrated that 4,5-CQME decreased Bax/Bcl-2 and Bak levels. A mechanistic study confirmed that 4,5-CQME significantly suppressed H₂O₂-induced MAPKs phosphorylation but had little effect on MAPKs phosphorylation under normal conditions. By contrast, 4,5-CQME induced AKT phosphorylation in the presence or absence of H₂O₂. 4,5-CQME also regulated the Keap1/Nrf2 signaling pathway and enhanced both the mRNA and protein expressions of HO-1 and NQO1. The anti-oxidative effect of 4,5-CQME was greatly abolished by co-incubation with the Nrf2 inhibitor ML385 or PI3K inhibitor wortmannin.ConclusionsTaken together, these results showed that 4,5-CQME offered significant protection against H₂O₂-induced oxidative stress, and its effect was in part due to the modulation of the Keap1/Nrf2 pathway.     

Synthesis and antibacterial properties of new N4-acetylated hexahydro-2,7-dioxopyrido[2,3-f]quinoxaline-8-carboxylic acids

Direct interaction between 7-chloro-1-cyclopropyl-6-fluoro-8-nitro-4-oxo-1,4-dihydroquinoline-3-carboxylic acid and primary α-amino acids (exemplified by glycine, alanine, and l-valine) in aqueous ethanolic NaHCO₃ at 70–80°C for 24–72?h produced the respective N-(4-oxoquinolin-7-yl)-α-amino acids (6a–c). The latter derivatives underwent reductive lactamization upon treatment with Na₂S₂O₄ in aqueous ethanol to afford moderate yields of the corresponding pyrido[2,3-f]quinoxaline-8-carboxylic acids (8a–c). Acetylation of 8a–c using acetyl chloride afforded N₄-acetylated hexahydro-2,7-dioxopyrido[2,3-f]quinoxaline-8-carboxylic acids (9a–c). The structures, assigned to these new heterocyclic products, are supported by analytical and spectral data. The synthesized compounds (6a–c/9a–c) showed appreciable antibacterial activity as compared with ciprofloxacin.     

A New Series of S-Adenosyl-L-Methionine Synthetase Inhibitors

Abstract

A new series of epithio and epoxy amino acid analogues of L-methionine or L-methoxinine were examined as potential inhibitors of the enzyme S-adenosylmethionine (AdoMet) syn-thetase. The kinetic behaviour of these compounds was studied using recombinant rat liver S-adenosyl-L-methionine sythetase (α-isoform) fractionated from E. coli, transformed with the plasmid pSSRL-T7N. All the compounds tested were competitive inhibitors with respect to L-methionine and the (2S, 4S)-2-amino-4,5-epoxy pentanoic acid was found to be a very potent inhibitor of the enzyme compared to those already reported for AdoMet synthetase from other mammalian tissues.     

Design and Synthesis of Novel Carbocyclic Versions of 2′-spirocyclopropyl Ribonucleosides as Potent Anti-HCV Agents

The discovery of 2′-spirocyclopropyl-ribocytidine as a potent inhibitor of RNA synthesis by NS5B (IC₅₀ = 7.3 μM), the RNA polymerase encoded by hepatitis C virus (HCV), has led to the synthesis and biological evaluation of carbocyclic versions of 2′-spiropropyl-nucleosides from cyclopentenol 6. Spirocyclopropylation of enone 7 was completed by using (2-chloroethyl)-dimethylsulfonium iodide and potassium t-butoxide to form the desired intermediate 9a. The synthesized nucleoside analogues, 18, 19, 26, and 27, were assayed for their ability to inhibit HCV RNA replication in a subgenomic replicon Huh7 cell line. The synthesized cytosine nucleoside 19 showed moderate anti-HCV activity (IC₅₀ = 14.4 μM).     

Urease inhibitors from Hypericum oblongifolium WALL.

The bioassay-guided fractionation of H. oblongifolium has led to the isolation of potent urease inhibitors 1–3. The structures were elucidated by NMR and mass spectroscopic techniques. Compound 2 showed a potent enzyme inhibition activity (IC₅₀ 20.96?±?0.93), which is comparatively higher than that for the standard thiourea (IC₅₀ 21.01?±?0.51 μM). Compounds 1 and 3 also showed a significant activity, with IC₅₀ 37.95?±?1.93 and 138.43?±?1.23 μM, respectively. The sub crude fractions (F1, F2, F3, and F4) were tested in vitro for their urease inhibition activity. Fractions F2 and F4 showed significant activity with IC₅₀ 140.37?±?1.93 and 167.43?±?3.03 μM, respectively.     

Identification of a specific inhibitor of nOGA — a caspase-3 cleaved O-GlcNAcase variant during apoptosis

O-Linked N-acetylglucosamine (O-GlcNAc) modification of serines/threonines on cytoplasmic proteins is a significant signal regulating cellular processes such as cell cycle, cell development, and cell apoptosis. O-GlcNAcase (OGA) is responsible for the removal of O-GlcNAc, and it thus plays a critical role in O-GlcNAc metabolism. Interestingly, OGA can be cleaved by caspase-3 into two fragments during apoptosis, producing an N-terminal fragment (1–413 a.a.), termed nOGA. Here, using 4-MU-GlcNAc (4-methylumbelliferyl 2-acetamido-2-deoxy-β-D-glucopyranoside) as substrate, we found that the nOGA fragment retains high glycosidase activity. To probe the role of nOGA in apoptosis, it is essential to develop a potent and specific nOGA inhibitor. However, many reported inhibitors active at nanomolar concentrations (including PUGNAc, STZ, GlcNAc-statin, and NAG-thiazoline) against full-length OGA were not potent for nOGA. Next, we screened a small triazole-linked carbohydrate library and first identified compound 4 (4-pyridyl-1-(2′-deoxy-2′-acetamido-β-D-glucopyranosyl)-1,2,3-triazole) as a potent and competitive inhibitor for nOGA. This compound shows 15-fold selectivity for nOGA (K _i = 48 μM) over the full-length OGA (K _i = 725 μM) and 10-fold selectivity over human lysosomal β-hexosaminidase A&B (Hex A&B) (K _i = 502 μM). These results reveal that compound 4 can be used as a potent and selective inhibitor for probing the role of nOGA in biological systems.     

Design and synthesis of (5-amino-1, 2, 4-triazin-6-yl)(2-(benzo[d] isoxazol-3-yl) pyrrolidin-1-yl)methanone derivatives as sodium channel blocker and anticonvulsant agents

Abstract

A series of novel (5-amino-3-substituted-1, 2, 4-triazin-6-yl) (2-(6-halo-substituted benzo[d]isoxazol-3-yl) pyrrolidin-1-yl) methanone 5a–5r was synthesized. Their anticonvulsant activities were evaluated by the maximal electroshock (MES) test and neurotoxicity was evaluated by the rotorod test. The MES test showed that (5-amino-3-phenyl-1, 2, 4-triazin-6-yl)(2-(6-fluorobenzo[d]isoxazol-3-yl) pyrrolidin-1-yl) methanone 5c was found to be the most potent compound with ED₅₀ value of 6.20?mg/kg (oral/rat) and a protective index (PI?=?ED₅₀/TD₅₀) value of >48.38, which was much higher than the PI of the reference drug phenytoin. To explain the possible mechanism of action of selected derivatives 5b, 5c, 5i and 5o, their influence on sodium channel was evaluated in vitro.     

Structure and Conformation of 5′-Chlorocyclocytidine,a Potent Inhibitor of Nucleic Acid Synthesis: X-Ray, 1H and 13C NMR Analyses

Abstract

The structure of the hydrochloride of 5′-chlorocyclocytidine, a potent inhibitor of DNA synthesis, was determined by X-ray crystallography. The nucleoside crystallizes in the orthorhombic space group P2₁2₁2₁ with cell dimensions a = 10.413(4), b = 13.236(5), c = 17.064(6) Å and with two independent molecules in the asymmetric unit (Z = 8). Atomic parameters were refined by full-matrix least squares to a final value of R = 0.053 for 2490 observed reflections. In both molecules the furanose ring has a C4′ endo/04′ exo (⁴ T ₀) pucker. In molecule A the orientation of the -CH₂Cl side chain is gauche. In molecule B the side chain is disordered: in 70% of these molecules the orientation is trans and in 30% it is gauche ⁺. ¹H NMR spectra indicate a conformational equilibrium between C4′ exo/04′ endo (₄ T ⁰) and C4′ endo/C3′ exo (⁴ ₃ T) with a population ratio of 38:62. All three side chain rotamers occur in solution, the trans orientation contributing most. ¹J(C, H) values for C1′ and C2′ are significantly higher than normal and can therefore be used as a diagnostic tool for the assignment of bridgehead carbon atoms in cyclonucleosides.     

Synthesis of phosphites and phosphates of neuraminic acid and their glycosyl donor properties — convenient synthesis of GM3

The importance and requirements for catalytic activation of sialyl donors are discussed, leading to the acid sensitive phosphite and phosphate moiety, respectively, as leaving group and nitriles as solvent. Therefore, from readily availableN-acetylneuraminic acid, derivative1 with phosphochloridites2a-f and Huenigs' base sialyl phosphites3a-f were prepared and isolated in high yields. Oxidation of3a, c withtert-butyl-hydroperoxide afforded the corresponding phosphates4a, c. As expected, phosphites3 could be activated in acetonitrile by catalytic amounts of TMSOTf; thus, from3a-e as donors and lactose derivatives8A, B as acceptors the ganglioside building blocks9A and9B, respectively, were obtained in good yields. The best results were obtained with diethyl phosphite derivative3a as sialyl donor, which exceeded by far the reults obtained with the corresponding phosphate derivative4a. Trisaccharide9B was transformed into known9A and into the fullyO-acetylated GM₃-trisaccharide10.     

2,4,5-Triphenylisothiazol-3(2H)-one 1,1-dioxides as inhibitors of human leukocyte elastase

A series of substituted 2,4,5-triphenylisothiazol-3(2H)-one 1,1-dioxides 9 was synthesized and investigated as inhibitors of human leukocyte elastase (HLE). All compounds were found to inhibit HLE in a time-dependent manner and most of them exhibited k_obs/[I] values > 300 M^{? 1}s^{? 1}. The most potent 3-oxosultam of this series was 9l (k_obs/[I] = 2440 M^{? 1}s^{? 1}). Kinetic investigations performed with 9g and different substrate concentrations did not allow to clearly distinguish between a competitive or noncompetitive mode of inhibition. A more complex interaction is supported by the failure of a linear dependency of k_obs values on the inhibitor concentration.     

Synthesis and Antimicrobial Activity of Some S-β-D-Glucosides of 4-Mercaptopyrimidine

5-Acetyl-2-aryl-6-methyl-4-(2,3,4,6-tetra- O -acetyl-β-D-glucopyranosylmercapto)pyramidines 3a–c were obtained by the reaction of 5-acetyl-2-aryl-6-methyl-pyrimidine thiol 1a–c with 2,3,4,6-tetra- O -acetyl-α-D-glucopyranosyl bromide (2) in aq. KOH/acetone. The reaction of 1a–c with peracetylated galactose 5 and peracetylated ribose 8 under MW irradiation gave 5-acetyl-2-aryl-6-methyl-4-(2,3,4,6-tetra- O -acetyl-β-D-galactopyranosylmercapto)pyrimidine 6a–c and 5-acetyl-2-aryl-6-methyl-4-(2,3,5-tri- O -acetyl-β-D-ribofuranosylmercapto)pyrimidines 9a–c. The deprotection of 3a–c, 6a–c, and 9a–c in the presence of methanol and TEA/H₂O yielded the deprotected products 4a–c, 7a–c, and 10a–c. The structures of the compounds were confirmed by using IR, ¹H, ¹³C spectra and microanalysis. Selected members of these compounds were screened for antimicrobial activity.