Author Report for: Kucukkilinc TT

In this research, a series of coumarin-based scaffolds linked to pyridine derivatives via a flexible aliphatic linkage were synthesized and assessed as multifunctional anti-AD agents. All the compounds showed acceptable acetylcholinesterase (AChE) inhibition activity in the nanomolar range (IC(50) = 2-144 nM) and remarkable butyrylcholinesterase (BuChE) inhibition property (IC(50) = 9-123 nM) compared to donepezil as the standard drug (IC(50) = 14 and 275 nM, respectively). Compound 3f as the best AChE inhibitor (IC(50) = 2 nM) showed acceptable BuChE inhibition activity (IC(50) = 24 nM), 100 times more active than the standard drug. Compound 3f could also significantly protect PC12 and SH-SY5Y cells against H(2)O(2)-induced cell death and amyloid toxicity, respectively, superior to the standard drugs. It could interestingly reduce beta-amyloid self and AChE-induced aggregation, more potent than the standard drug. All the results suggest that compound 3f could be considered as a promising multi-target-directed ligand (MTDL) against AD.

PURPOSE: Alzheimer's disease (AD) is a multifaceted neurodegenerative disease. To target simultaneously multiple pathological processes involved in AD, natural-origin compounds with unique characteristics are promising scaffolds to develop novel multi-target compounds in the treatment of different neurodegenerative disease, especially AD. In this study, novel chromone-lipoic acid hybrids were prepared to find a new multifunctional lead structure for the treatment of AD. METHODS: Chromone-lipoic acid hybrids were prepared through click reaction and their neuroprotection and anticholinesterase activity were fully evaluated. The anti-amyloid aggregation, antioxidant and metal-chelation activities of the best compound were also investigated by standard methods to find a new multi-functional agent against AD. RESULTS: The primary biological screening demonstrated that all compounds had significant neuroprotection activity against H2O2-induced cell damage in PC12 cells. Compound 19 as the most potent butyrylcholinesterase (BuChE) inhibitor (IC50 = 7.55 microM) having significant neuroprotection activity as level as reference drug was selected for further biological evaluations. Docking and kinetic studies revealed non-competitive mixed-type inhibition of BuChE by compound 19. It could significantly reduce formation of the intracellular reactive oxygen species (ROS) and showed excellent reducing power (85.57 mM Fe+2), comparable with quercetin and lipoic acid. It could also moderately inhibit Abeta aggregation and selectively chelate with copper ions in 2:1 M ratio. CONCLUSION: Compound 19 could be considered as a hopeful multifunctional agent for the further development gainst AD owing to the acceptable neuroprotective and anti-BuChE activity, moderate anti-Abeta aggregation activity, outstanding antioxidant activity as well as selective copper chelation ability. A new chromone-lipoic acid hybrid was synthesized as anti-Alzheimer agent with BuChE inhibitory activity, anti-Abeta aggregation, metal-chelation and antioxidant properties.

A novel series of coumarin derivatives linked to the N-benzyl triazole group were synthesized and evaluated against 15-lipoxygenase (15-LOX), and acetyl- and butyrylcholinesterase (AChE and BuChE) to find the most potent derivative against Alzheimer's disease (AD). Most of the compounds showed weak to moderate activity against ChEs. Among the most active BuChE and 15-LOX inhibitors, 8l and 8n exhibited an excellent neuroprotective effect, higher than the standard drug (quercetin) on the PC12 cell model injured by H(2)O(2) and significantly reduced aggregation of amyloid Abeta(1-42), with potencies of 1.44 and 1.79 times higher than donepezil, respectively. Compound 8l also showed more activity than butylated hydroxytoluene (BHT) as the reference antioxidant agent in reducing the levels of H(2)O(2) activated by amyloid beta in BV2 microglial cells. Kinetic and ligand-enzyme docking studies were also performed for better understanding of the mode of interaction between the best BuChE inhibitor and the enzyme. Considering the acceptable BuChE and 15-LOX inhibition activities as well as significant neuroprotection, and anti-amyloid aggregation activities, 8l and 8n could be considered as potential MTDLs for further modification and studies against AD.

Multi-Target approach is particularly promising way to drug discovery against Alzheimer's disease. In the present study, we synthesized a series of compounds comprising the carbazole backbone linked to the benzyl piperazine, benzyl piperidine, pyridine, quinoline, or isoquinoline moiety through an aliphatic linker and evaluated as cholinesterase inhibitors. The synthesized compounds showed IC50 values of 0.11-36.5microM and 0.02-98.6microM against acetyl- and butyrylcholinesterase (AChE and BuChE), respectively. The ligand-protein docking simulations and kinetic studies revealed that compound 3s could bind effectively to the peripheral anionic binding site (PAS) and anionic site of the enzyme with mixed-type inhibition. Compound 3s was the most potent compound against AChE and BuChE and showed acceptable inhibition potency for self- and AChE-induced Abeta1-42 aggregation. Moreover, compound 3s could significantly protect PC12 cells against H2O2-induced toxicity. The results suggested that the compounds 3s could be considered as a promising multi-functional agent for further drug discovery development against Alzheimer's disease.

A series of benzofuran-2-carboxamide-N-benzyl pyridinium halide derivatives (6a-o) are synthesized as new cholinesterase inhibitors. The synthetic pathway involves the reaction of salicylaldehyde derivatives and ethyl bromoacetate, followed by hydrolysis and amidation with 3- and 4-picolyl amine. Subsequently, N-((pyridin-4-yl) methyl) benzofuran-2-carboxamide and substituted N-((pyridin-3-yl) methyl) benzofuran-2-carboxamides reacts with benzyl halides to afford target compounds (6a-o). The chemical structures of all derivatives were confirmed by spectroscopic methods. The studies reveal that some of the synthesized compounds are potent butyrylcholinesterase inhibitors with IC50 values in the range of 0.054-2.7microM. In addition, good inhibitory effects on Abeta self-aggregation are observed for 6h and 6k (33.1 and 46.4% at 100microM, respectively).

Bisphenol A (BPA) is an endocrine disruptor chemical, which is commonly used in everyday products. Adverse effects of its exposure are reported even at picomolar doses. Effects of picomolar and nanomolar concentrations of BPA on cytotoxicity, nitric oxide (NO) levels, acetylcholinesterase (AChE) gene expression and activity, and tumor necrosis factor-alpha (TNF-alpha) and caspase-8 levels were determined in SH-SY5Y cells. The current study reveals that low-dose BPA treatment induced cytotoxicity, NO, and caspase-8 levels in SH-SY5Y cells. We also evaluated the mechanism underlying BPA-induced cell death. Ours is the first report that receptor-interacting serine/threonine-protein kinase 1-mediated necroptosis is induced by nanomolar BPA treatment in SH-SY5Y cells. This effect is mediated by altered AChE and decreased TNF-alpha levels, which result in an apoptosis-necroptosis switch. Moreover, our study reveals that BPA is an activator of AChE.

A novel series of coumarin-lipoic acid conjugates were synthesized via cycloaddition click reaction to find out new multi-target-directed ligands (MTDLs) for treatment of Alzheimer's disease (AD). All of synthesized compounds were screened for neuroprotective and anti-cholinesterase activities. Based on primary screening, two compounds (5 and 11) were subjected to further biological evaluations. In particular, compound 11 which was the most potent AChE inhibitor showed good inhibitory effect on Abeta-aggregation and intracellular ROS (reactive oxygen species) formation, as well as the ability of selective bio-metal chelation and neuroprotection against H2O2- and Abeta1-42-induced cytotoxicity. In the light of these results, the applied hybridization approach introduced new promising lead compound with desired multifunctional properties, being useful in the treatment of Alzheimer's disease.

A novel series of benzylpyridinium-based benzoheterocycles (benzimidazole, benzoxazole or benzothiazole) were designed as potent acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) inhibitors. The title compounds 4a-q were conveniently synthesized via condensation reaction of 1,2-phenylenediamine, 2-aminophenol or 2-aminothiophenol with pyridin-4-carbalehyde, followed by N-benzylation using various benzyl halides. The results of in vitro biological assays revealed that most of them, especially 4c and 4g, had potent anticholinesterase activity comparable or more potent than reference drug, donepezil. The kinetic study demonstrated that the representative compound 4c inhibits AChE in competitive manner. According to the ligand-enzyme docking simulation, compound 4c occupied the active site at the vicinity of catalytic triad. The compounds 4c and 4g were found to be inhibitors of Abeta self-aggregation as well as AChE-induced Abeta aggregation. Meanwhile, these compounds could significantly protect PC12 cells against H2O2-induced injury and showed no toxicity against HepG2 cells. As multi-targeted structures, compounds 4c and 4g could be considered as promising candidate for further lead developments to treat Alzheimer's disease.

The present study describes the synthesis, pharmacological evaluation (BChE/AChE inhibition, Abeta antiaggregation, and neuroprotective effects), and molecular modeling studies of novel 2-[4-(4-substitutedpiperazin-1-yl)phenyl]benzimidazole derivatives. The alkyl-substituted derivatives exhibited selective inhibition on BChE with varying efficiency. Compounds 3b and 3d were found to be the most potent inhibitors of BChE with IC50 values of 5.18 and 5.22muM, respectively. The kinetic studies revealed that 3b is a partial non-competitive BChE inhibitor. Molecular modeling studies also showed that the alkyl-substituted derivatives were able to reach the catalytic anionic site of the BChE. The compounds with an inhibitory effect on BChE were subsequently screened for their Abeta antiaggregating and neuroprotective activities. Compounds 3a and 3b exerted a potential neuroprotective effect against H2O2 and Abeta-induced cytotoxicity in SH-SY5Y cells. Collectively, 3b was found as the most promising compound for the development of multi-target directed ligands against Alzheimer's disease.

In order to find novel cyclooxygenase (COX)-2 inhibitors for treating inflammatory-based diseases such as Alzheimer's disease (AD), an ethyl carboxylate side chain was added to 5-(4-chlorophenyl)-6-(4-(methylsulfonyl)phenyl)-3-(methylthio)-1,2,4-triazine (lead compound II) to maintain residual inhibition of COX-1 through interacting with Arg120. A preliminary molecular docking study on both the COX-1/COX-2 active sites truly confirmed our hypothesis. Accordingly, a series of ethyl 5,6-diaryl-1,2,4-triazine-3-ylthioacetate derivatives were synthesized and their chemical structures were confirmed by NMR, IR and MS spectra. Further in vitro COX-1/COX-2 evaluations revealed that compound 6c (COX-2 IC50 = 10.1 muM, COX-1 IC50 = 88.8 muM) is the most selective COX-2 inhibitor while maintaining residual inhibition of COX-1. In order to evaluate their potential use against AD, an in vitro evaluation of beta-amyloid fibril formation was performed. The results indicated that the prototype compounds 6 are effective beta-amyloid destabilizing agents while compound 6c could inhibit 94% of the beta-amyloid fibril formation after 48 h. Finally, the in silico assessment results of their blood-brain barrier permeability were satisfactory.

To find out new agents for treating inflammatory-involved diseases such as Alzheimer's disease, a series of 1,2-diaryl-2-hydroxyiminoethanones containing vicinal diaryl pharmacophore of COX inhibitors were tested by a set of in vitro, in vivo, and computational studies. The in vivo study of compounds indicated their prominent anti-inflammatory ability at the doses of 10 and 20 mg/kg comparable to celecoxib (10 mg/kg). Further in vitro COX-1/COX-2 evaluations revealed that 4-methoxy derivative 3 had a high selective COX-1 inhibitory activity (COX-1, IC50=0.12 mum, SI>833). To evaluate their potential use against Alzheimer's disease, in vitro evaluation of beta-amyloid fibril formation using Abeta(1-40) and Abeta(1-42) peptides was performed. The evaluation of their antiaggregation ability gave impressive results and comparable to rifampicin and indomethacin. Conformational study of compound 3 and subsequent docking of its restrained analogs on both active sites of COX-1 and COX-2 could provide a proof of its COX-1 selectivity as well as molecular dynamic simulation could elucidate and give more insight into the amyloid disaggregation mechanisms leading to rational design of inhibitors.

To develop new drugs for treatment of Alzheimer's disease, a group of N'-2-(4-Benzylpiperidin-/piperazin-1-yl)acylhydrazones was designed, synthesized and tested for their ability to inhibit acetylcholinesterase, butyrylcholinesterase and aggregation of amyloid beta peptides (1-40, 1-42 and 1-40_1-42). The enzyme inhibition assay results indicated that compounds moderately inhibit both acetylcholinesterase and butyrylcholinesterase. beta-Amyloid aggregation results showed that all compounds exhibited remarkable Abeta fibril aggregation inhibition activity with a nearly similar potential as the reference compound rifampicin, which makes them promising anti-Alzheimer drug candidates. Docking experiments were carried out with the aim to understand the interactions of the most active compounds with the active site of the cholinesterase enzymes.

The cholinesterases, acetylcholinesterase (AChE) and butyrylcholinesterase, are primary targets of organophosphates (OPs). Exposure to OPs can lead to serious cardiovascular complications, respiratory compromise, and death. Current therapy to combat OP poisoning involves an oxime reactivator (2-PAM, obidoxime, TMB4, or HI-6) combined with atropine and on occasion an anticonvulsant. Butyrylcholinesterase, administered in the plasma compartment as a bio-scavenger, has also shown efficacy but is limited by its strict stoichiometric scavenging, slow reactivation, and a propensity for aging. Here, we characterize 10 human (h) AChE mutants that, when coupled with an oxime, give rise to catalytic reactivation and aging resistance of the soman conjugate. With the most efficient human AChE mutant Y337A/F338A, we show enhanced reactivation rates for several OP-hAChE conjugates compared with wild-type hAChE when reactivated with HI-6 (1-(2'-hydroxyiminomethyl-1'-pyridinium)-3-(4'-carbamoyl-1-pyridinium)). In addition, we interrogated an 840-member novel oxime library for reactivation of Y337A/F338A hAChE-OP conjugates to delineate the most efficient oxime-mutant enzyme pairs for catalytic bio-scavenging. Combining the increased accessibility of the Y337A mutation to oximes within the space-impacted active center gorge with the aging resistance of the F338A mutation provides increased substrate diversity in scavenging potential for aging-prone alkyl phosphate inhibitors.

Organophosphates (OPs) exert their toxicity by inhibiting primarily acetylcholinesterase (AChE) and to a lesser extent butyrylcholinesterase (BChE). Binary mixtures of mammalian AChE and oximes of varying structure have been recently considered for treatment of OP poisoning as catalytic bioscavengers. In this study wild type human AChE and human AChE with residue mutations D134H, D134H_E202Q and D134H_F338A were characterized and investigated for inhibition by OPs and consequent oxime reactivation of phosphylated enzymes. The rationale for selecting these substitution positions was based on D134H being a naturally occurring single nucleotide polymorphism (SNP) in humans and that E202Q and F338A mutations slow aging of OP inhibited AChEs. Inhibition of D134H by paraoxon and analogues of cyclosarin was 2-8 times slower than inhibition of wild type (wt), while reactivation of the paraoxon inhibited enzyme by 2PAM was 6 times faster. Both inhibition and reactivation of D134H_E202Q and D134H_F338A double mutants were up to two orders of magnitude slower than the wt indicating that introduction of the active center substitutions abolished fully the effect of the peripherally located D134H. These results indicate that selected residues outside the active center influence inhibition, reactivation and catalysis rates through longer range interactions.

The effects of three cationic triarylmethane dyes--pararosaniline (PR), malachite green (MG), methyl green (MetG)--on electric eel AChE (eAChE) activity were tested at 25 degrees C, in 100 mM MOPS buffer (pH 8) containing 0.125 mM 5-5-dithio-bis(2-nitrobenzoic acid), 20-120 microM acetylthiocholine and 0-20 microM dye. All three dyes caused reversible, linear- or hyperbolic-mixed inhibition of esteratic activity. The respective inhibitory parameters for PR, MG and MetG were K(i)=8.4+/-0.67, 1.9+/-0.51 and 0.27+/-0.017 microM; alpha (competitive coefficient)=5.8+/-2.0, 4.8+/-1.8 and 2.7+/-0.32; beta (noncompetitive coefficient)=0, 0 and 0.20+/-0.011. The data were consistent with ligand binding at the peripheral site and a remote effect on substrate binding and turnover.

Two cationic phenoxazine dyes, meldola blue (MB) and nile blue (NB), and the structurally related phenothiazine, methylene blue (MethB), were found to act as complex inhibitors of human plasma cholinesterase (butyrylcholinesterase, BChE). Studied at 25 degrees C, in 100mM MOPS buffer (pH 8.0), with butyrylthiocholine as substrate, the kinetic pattern of inhibition indicated cooperative I binding at 2 sites. Intrinsic K' values ( identical with[I](0.5)(2) extrapolated to [S]=0) for MB, NB and MethB were 0.64+/-0.05, 0.085+/-0.026 and 0.42+/-0.04 microM, respectively. Under the same experimental conditions the dyes acted as single-occupancy, hyperbolic-mixed inhibitors of electric eel acetylcholinesterase (AChE), with K(i)=0.035+/-0.010, 0.026+/-0.0034 and 0.017+/-0.0063 microM (for MB, NB, MethB); alpha (coefficient of competitive interaction)=1.8-2.4 and beta (coefficient of noncompetitive interaction)=0.15-0.28. The complexity of the BChE inhibitory effect of phenoxazine/phenothiazine dyes contrasted with that of conventional ChE inhibitors which cause single-occupancy (n=1), competitive or mixed inhibition in both AChE and BChE and signaled novel modes of ligand interaction at (or remote from) the active site gorge of the latter enzyme.

The inhibitory effects of the cationic triarylmethane (TAM+) dyes, pararosaniline (PR+), malachite green (MG+), and methyl green (MeG+) on human plasma cholinesterase (BChE) were studied at 25 degrees C in 100 mM Mops, pH 8.0, with butyrylthiocholine as substrate. PR+ and MG+ caused linear mixed inhibition of enzyme activity. The respective inhibitory parameters were K(i) = 1.9 +/- 0.23 microM, alpha = 13 +/- 48, beta = 0 and K(i) = 0.28 +/- 0.037 microM, alpha = 23 +/- 7.4, beta = 0. MeG+ acted as a competitive inhibitor with K(i) = 0.12 +/- 0.017 microM (alpha, infinity, beta, not applicable). The K(i) values were within the same range reported for a number of ChE inhibitors including propidium ion, donepezil, and the phenothiazines, suggesting that TAM+s are active site ligands. On the other hand, the alpha values failed to correlate with values previously reported for a number of ChE inhibitors. It appears that mixed inhibition is the combined result of more than one type of binding and S-I interference. The impact of ligands at the choline-specific and peripheral anionic sites (or, possibly, accessory structural domains) on BChE activity needs to be studied in further detail.