Novel 6-alkyl-bridged 4-arylalkylpiperazin-1-yl derivatives of azepino[4,3-b]indol-1(2H)-one as potent BChE-selective inhibitors showing protective effects against neurodegenerative insults.

Due to the putative role of butyrylcholinesterase (BChE) in regulation of acetylcholine levels and functions in the late stages of the Alzheimer's disease (AD), the potential of selective inhibitors (BChEIs) has been envisaged as an alternative to administration of acetylcholinesterase inhibitors (AChEIs). Starting from our recent findings, herein the synthesis and in vitro evaluation of cholinesterase (ChE) inhibition of a novel series of some twenty 3,4,5,6-tetrahydroazepino[4,3-b]indol-1(2H)-one derivatives, bearing at the indole nitrogen diverse alkyl-bridged 4-arylalkylpiperazin-1-yl chains, are reported. The length of the spacers, as well as the type of arylalkyl group affected the enzyme inhibition potency and BChE/AChE selectivity. Two compounds, namely 14c (IC50 = 163 nM) and 14d (IC50 = 65 nM), bearing at the nitrogen atom in position 6 a n-pentyl- or n-heptyl-bridged 4-phenethylpiperazin-1-yl chains, respectively, proved to be highly potent mixed-type inhibitors of both equine and human BChE isoforms, showing more than two order magnitude of selectivity over AChE. The study of binding kinetics through surface plasmon resonance (SPR) highlighted differences in their BChE residence times (8 and 47 s for 14c and 14d, respectively). Moreover, 14c and 14d proved to hit other mechanisms known to trigger neurodegeneration underlying AD and other CNS disorders. Unlike 14c, compound 14d proved also capable of inhibiting by more than 60% the in vitro self-induced aggregation of neurotoxic amyloid-ß (Aß) peptide at 100 µM concentration. On the other hand, 14c was slightly better than 14d in counteracting, at 1 and 10 µM concentration, glutamate excitotoxicity, due to over-excitation of NMDA receptors, and hydrogen peroxide-induced oxidative stress assessed in neuroblastoma cell line SH-SY5Y. This paper is dedicated to Prof. Marcello Ferappi, former dean of the Faculty of Pharmacy of the University of Bari, in the occasion of his 90th birthday.

Coumarin-azasugar-benzyl conjugates as non-neurotoxic dual inhibitors of butyrylcholinesterase and cancer cell growth.

We have applied the copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction to prepare a library of ten coumarin-azasugar-benzyl conjugates and two phthalimide-azasugar-benzyl conjugates with potential anti-Alzheimer and anti-cancer properties. The compounds were evaluated as cholinesterase inhibitors, demonstrating a general preference, of up to 676-fold, for the inhibition of butyrylcholinesterase (BuChE) over acetylcholinesterase (AChE). Nine of the compounds behaved as stronger BuChE inhibitors than galantamine, one of the few drugs in clinical use against Alzheimer's disease. The most potent BuChE inhibitor (IC50 = 74 nM) was found to exhibit dual activities, as it also showed high activity (GI50 = 5.6 ± 1.1 µM) for inhibiting the growth of WiDr (colon cancer cells). In vitro studies on this dual-activity compound on Cerebellar Granule Neurons (CGNs) demonstrated that it displays no neurotoxicity.

Environmental exposure to glyphosate does not inhibit human acetylcholinesterase and butyrylcholinesterase.

Glyphosate has remained the leading herbicide on the global market to date, despite the continuous debate between consumers, scientific community, and regulatory agencies over its carcinogenicity, genotoxicity, environmental persistence, and the role in the development of neurodegenerative disorders. Chemically, glyphosate belongs to a large family of organophosphorus pesticides, which exert a neurotoxic effect by inhibiting acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), enzymes of the cholinergic system essential for maintaining neurotransmission. Although research shows that glyphosate is a weak cholinesterase inhibitor in fish and mammals compared to other OP compounds, no conclusive data exist concerning the inhibition of human AChE and BChE. In our study we analysed its inhibitory potency on human AChE and BChE, by establishing its IC50 and reversible inhibition in terms of dissociation inhibition constants. Glyphosate concentration of 40 mmol/L caused near total inhibition of enzyme activity (approx. 10 % activity remaining). Inhibition dissociation constants (K i) of glyphosate-AChE and -BChE complexes were 28.4±2.7 mmol/L and 19.3±1.8 mmol/L, respectively. In conclusion, glyphosate shows a slight binding preference for BChE but exhibits inhibition only in a high concentration range. Our results are in line with studies reporting that its neurotoxic effect is not primarily linked to the cholinergic system.

Discovery, Structure-Based Modification, In Vitro, In Vivo, and In Silico Exploration of m-Sulfamoyl Benzoamide Derivatives as Selective Butyrylcholinesterase Inhibitors for Treating Alzheimer's Disease.

For the potential therapy of Alzheimer's disease (AD), butyrylcholinesterase (BChE) has gradually gained worldwide interest in the progression of AD. This study used a pharmacophore-based virtual screening (VS) approach to identify Z32439948 as a new BChE inhibitor. Aiding by molecular docking and molecular dynamics, essential binding information was disclosed. Specifically, a subpocket was found and structure-guided design of a series of novel compounds was conducted. Derivatives were evaluated in vitro for cholinesterase inhibition and physicochemical properties (BBB, log P, and solubility). The investigation involved docking, molecular dynamics, enzyme kinetics, and surface plasmon resonance as well. The study highlighted compounds 27a (hBChE IC50 = 0.078 ± 0.03 µM) and (R)-37a (hBChE IC50 = 0.005 ± 0.001 µM) as the top-ranked BChE inhibitors. These compounds showed anti-inflammatory activity and no apparent cytotoxicity against the human neuroblastoma (SH-SY5Y) and mouse microglia (BV2) cell lines. The most active compounds exhibited the ability to improve cognition in both scopolamine- and Aß1-42 peptide-induced cognitive deficit models. They can be promising lead compounds with potential implications for treating the late stage of AD.

Variability of Isoquinoline Alkaloid Profiles and Anticholinesterase Activities with Binding-Mode Predictions of Glaucium flavum Population.

In this study, phytochemical and biological activity studies supported by docking were carried out on a species of the genus Glaucium, a repository of isoquinoline alkaloids. The GC-MS (Gas Chromatography-Mass Spectrometry) method is used to characterize the isoquinoline alkaloids of Glaucium flavum Crantz. (Papaveraceae). G. flavum was collected from seven different regions of Türkiye (Antalya, Urla-Izmir, Mordogan-Izmir, Mugla, Assos-Canakkale, Karabiga-Canakkale, Giresun) and totally 17 compounds were detected by GC-MS. Glaucine was found to be the major constituent in the sample collected from Mugla, whereas isocorydine was recorded to be the principal alkaloid in other samples. Further fractionation studies on G. flavum collected from Antalya province in Southwestern Türkiye, yielded five major alkaloids (isocorydine 1, dihydrosanguinarine 2, glaucine 3, dehydroglaucine 4, protopine 5) which were characterized by spectroscopic methods. Anticholinesterase activities of the extracts and isolated alkaloids were also tested by in vitro Ellman method. The isolated compounds were also analyzed by a molecular docking technique to determine the binding orientations in the gorge of the active site of acetylcholinesterase (AChE) and a homology model of butyrylcholinesterase (BuChE). This is the first comparative investigation of the phytochemical composition and biodiversity of Glaucium flavum species growing in Türkiye.

Novel Dual-Acting Hybrids Targeting Type-2 Cannabinoid Receptors and Cholinesterase Activity Show Neuroprotective Effects In Vitro and Amelioration of Cognitive Impairment In Vivo.

Alzheimer's disease (AD) is a neurodegenerative form of dementia characterized by the loss of synapses and a progressive decline in cognitive abilities. Among current treatments for AD, acetylcholinesterase (AChE) inhibitors have efficacy limited to symptom relief, with significant side effects and poor compliance. Pharmacological agents that modulate the activity of type-2 cannabinoid receptors (CB2R) of the endocannabinoid system by activating or blocking them have also been shown to be effective against neuroinflammation. Herein, we describe the design, synthesis, and pharmacological effects in vitro and in vivo of dual-acting compounds that inhibit AChE and butyrylcholinesterase (BChE) and target CB2R. Within the investigated series, compound 4g proved to be the most promising. It achieved IC50 values in the low micromolar to submicromolar range against both human cholinesterase isoforms while antagonizing CB2R with Ki of 31 nM. Interestingly, 4g showed neuroprotective effects on the SH-SY5Y cell line thanks to its ability to prevent oxidative stress-induced cell toxicity and reverse scopolamine-induced amnesia in the Y-maze forced alternation test in vivo.

Optical imaging probes for selective detection of butyrylcholinesterase.

Butyrylcholinesterase (BChE), a member of the human serine hydrolase family, is an essential enzyme for cholinergic neurotransmission as it catalyzes the hydrolysis of acetylcholine. It also plays central roles in apoptosis, lipid metabolism, and xenobiotic detoxification. On the other side, abnormal levels of BChE are directly associated with the formation of pathogenic states such as neurodegenerative diseases, psychiatric and cardiovascular disorders, liver damage, diabetes, and cancer. Thus, selective and sensitive detection of BChE level in living organisms is highly crucial and is of great importance to further understand the roles of BChE in both physiological and pathological processes. However, it is a very complicated task due to the potential interference of acetylcholinesterase (AChE), the other human cholinesterase, as these two enzymes share a very similar substrate scope. To this end, optical imaging probes have attracted immense attention in recent years as they have modular structures, which can be tuned precisely to satisfy high selectivity toward BChE, and at the same time they offer real time and nondestructive imaging opportunities with a high spatial and temporal resolution. Here, we summarize BChE selective imaging probes by discussing the critical milestones achieved during the development process of these molecular sensors over the years. We put a special emphasis on design principles and biological applications of highly promising new generation activity-based probes. We also give a comprehensive outlook for the future of BChE-responsive probes and highlight the ongoing challenges. This collection marks the first review article on BChE-responsive imaging agents.

Design, synthesis, and activity evaluation of novel multitargeted l-tryptophan derivatives with powerful antioxidant activity against Alzheimer's disease.

Alzheimer's disease (AD) is a multifactorial neurological disease, and the multitarget directed ligand (MTDL) strategy may be an effective approach to delay its progression. Based on this strategy, 27 derivatives of l-tryptophan, 3a-1-3d-1, were designed, synthesized, and evaluated for their biological activity. Among them, IC50 (inhibitor concentration resulting in 50% inhibitory activity) values of compounds 3a-18 and 3b-1 were 0.58 and 0.44 µM for human serum butyrylcholinesterase (hBuChE), respectively, and both of them exhibited more than 30-fold selectivity for human serum acetylcholinesterase. Enzyme kinetics studies showed that these two compounds were mixed inhibitors of hBuChE. In addition, these two derivatives possessed extraordinary antioxidant activity in OH radical scavenging and oxygen radical absorption capacity fluorescein assays. Meanwhile, these compounds could also prevent ß-amyloid (Aß) self-aggregation and possessed low toxicity on PC12 and AML12 cells. Molecular modeling studies revealed that these two compounds could interact with the choline binding site, acetyl binding site, and peripheral anionic site to exert submicromolar BuChE inhibitory activity. In the vitro blood-brain barrier permeation assay, compounds 3a-18 and 3b-1 showed enough blood-brain barrier permeability. In drug-likeness prediction, compounds 3a-18 and 3b-1 showed good gastrointestinal absorption and a low risk of human ether-a-go-go-related gene toxicity. Therefore, compounds 3a-18 and 3b-1 are potential multitarget anti-AD lead compounds, which could work as powerful antioxidants with submicromolar selective inhibitory activity for hBuChE as well as prevent Aß self-aggregation.

Preliminary studies of 1,5-benzoxazepine derivatives as potential histamine H3 receptor antagonists.

Aims: Our research aimed to evaluate how the rigidification of the characteristic 3-aminopropyloxy linker by incorporating it into 1,5-benzoxazepines affects the potency of histamine H3 receptor (H3R) antagonists/inverse agonists. This research constitutes a starting point for the full characterization of the pharmacological properties of this group of compounds. Materials & methods: Several 1,5-benzoxazepine derivatives were synthesized and pharmacologically tested as potential H3R antagonist/inverse agonists. In a addition, the effect of the derivatives on acetylcholinesterase and butyrylcholinesterase inhibition and cytotoxicity were tested. Results: The studies indicated 1,5-benzoxazepine containing three carbon side chains as a compound for further modification. Conclusion: Further optimization of the lead structure is necessary, which will favorably affect biological targets.

DNA protection, molecular docking, antioxidant, antibacterial, enzyme inhibition, and enzyme kinetic studies for parietin, isolated from Xanthoria parietina (L.) Th. Fr.

Parietin was isolated from Xanthoria parietina (L.) Th. Fr.' (methanol:chloroform) extract, using a silica column. 13 C NMR and 1H NMR were used to confirm the structure of the isolated parietin. For the first time, parietin was investigated for its antioxidant, antibacterial and DNA protective activities. Molecular docking was carried out to determine the binding affinity and interactions between the enzymes and our molecule. Inhibition and kinetic mechanism studies for the action of the enzymes were performed too. Parietin exhibited high metal chelating activity. The MIC values of parietin were sufficient to inhibit different bacterial strains; E. coli, P. aeruginosa, K. pneumoniae and S. aureus. Molecular docking applications exhibited that acetylcholinesterase (AChE), butyrylcholinesterase (BChE), lipase, and tyrosinase have high potential for binding with the parietin. Especially, the parietin's highest binding affinity was recorded with AChE and tyrosinase. These results were confirmed by the inhibition and kinetics results, where, parietin observed a potent inhibition with an IC50 values between 0.013-0.003 µM. Moreover, parietin acts' as a non-competitive inhibitor against AChE, BChE, and lipase, and as a competitive inhibitor against tyrosinase with a high rate of inhibition stability. The promising biological properties of parietin revealed its effectiveness in terms of suitability in the food and pharmaceutical industries.Communicated by Ramaswamy H. Sarma.

Protective effect of bromelain on some metabolic enzyme activities in tyloxapol-induced hyperlipidemic rats.

Elevation of one or more plasma lipids, such as phospholipids, cholesterol esters, cholesterol, and triglycerides, is known as hyperlipidemia. In humans and experimental animals, bromelain, the primary active ingredient isolated from pineapple stems, has several positive effects, including anti-tumor growth, anticoagulation, and anti-inflammation. Hence, the purpose of this study was to determine the possible protective impact of bromelain on some metabolic enzymes (paraoxonase-1, glutathione S-transferase, glutathione reductase, sorbitol dehydrogenase [SDH], aldose reductase [AR], butyrylcholinesterase [BChE], and acetylcholinesterase [AChE]), activity in the heart, kidney, and liver of rats with tyloxapol-induced hyperlipidemia. Rats were divided into three groups: control group, HL-control group (tyloxapol 400 mg/kg, i.p. administered group), and HL+bromelain (group receiving bromelain 250 mg/kg/o.d. prior to administration of tyloxapol 400 mg/kg, i.p.). BChE, SDH, and AR enzyme activities were significantly increased in all tissues in HL-control compared to the control, whereas the activity of other studied enzymes was significantly decreased. Bromelain had a regulatory effect on all tissues and enzyme activities. In conclusion, these results prove that bromelain is a new mediator that decreases hyperlipidemia.

A biochemistry-oriented drug design: synthesis, anticancer activity, enzymes inhibition, molecular docking studies of novel 1,2,4-triazole derivatives.

In this study, we researched the reactions of 5-(5-bromofuran-2-yl)-4-methyl-1,2,4-triazole-3-thiol and 5-thiophene-(3-ylmethyl)-4R-1,2,4-triazole-3-thiols with some halogen-containing compounds, a number of new compounds were synthesized (1.1-1.5 and 2.1-2.8). These compounds showed excellent to good inhibitory activities on acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes. For obtaining the effects of these compounds on AChE and BChE enzymes were determined spectrophotometrically according to Ellman. IC50 values of these enzymes were ranging between 1.63 and 17.68 nM for AChE and 8.71 and 84.02 nM for BChE. After, prostate cancer is the second leading cause of cancer-related mortality for men over the age of 65 in developed countries. Current treatment options remain limited in the treatment of advanced-stage prostate cancer leading to biochemical recurrence in almost 40% of the patients. Therefore, there is an urgent need for development of novel therapeutic tools for treatment of prostate cancer patients. In this study, we aimed at analyzing the potential of all compounds against prostate cancer cells. We found that, of the tested compounds, 2.1, 2.2 and 2.3 showed significant cytotoxic activities against PC3 prostate cancer cells, although their effect on the viability of normal prostate cells was limited. These findings suggest their selective targeting potential for prostate cancer cells and offer them as candidate therapeutic agents against prostate cancer. The inhibitory activities of some chemical compounds, such as (1.1-1.5 and 2.1-2.8) were assessed by performing the molecular docking study in the presence of AChE, BChE and prostate cancer protein. MM/GBSA methods are calculated binding free energy. Finally, ADME/T analysis was performed to examine the drug properties of the 13 studied molecules.Communicated by Ramaswamy H. Sarma.

Synthesis and biological evaluation of novel salicylidene uracils: Cytotoxic activity on human cancer cell lines and inhibitory action on enzymatic activity.

A series of salicylidene uracil (1-18) derived from 5-aminouracil and substituted salicylaldehydes were analyzed for cytotoxic activity and enzyme inhibitory potency. Nine out of eighteen derivatives (6-8, 10, 12-15, 18) are novel molecules synthesized for the first time in this work, and other derivatives were previously synthesized by our group. The compounds were characterized by Proton nuclear magnetic resonance, carbon nuclear magnetic resonance, fourier transform infrared spectroscopy, and elemental analysis. All compounds were tested for their in vitro cytotoxicity against PC-3 (human prostate adenocarcinoma), A549 (human alveolar adenocarcinoma), and SHSY-5Y (human neuroblastoma) cancer cell lines and the nontumorigenic HEK293 (human embryonic kidney cells) cell line. The 3,5-di-tert-butylsalicylaldehyde derived compound (8) was toxic to PC-3 human prostate adenocarcinoma cells, showing a promising IC50 value at 7.05 ± 0.76 µM. The present study also aimed to evaluate the inhibitory effects of the compounds against several key enzymes, namely carbonic anhydrase I and II (CA I and CA II), acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and glutathione reductase (GR), which are implicated in various global disorders, such as Alzheimer's disease, epilepsy, cancer, malaria, diabetes, and glaucoma. The inhibitory profiles of the tested compounds were assessed by determining their Ki values, which ranged from 2.96 to 9.24 nM for AChE, 3.78 to 12.57 nM for BChE, 8.42 to 25.74 nM for CA I, 7.24 to 19.74 nM for CA II, and 0.541 to 1.124 µM for GR. Molecular docking studies were also performed for all compounds. Most derivatives exhibited much more effective inhibitory action compared with clinically used standards. Thus, our findings indicate that the salicylidene derivatives presented in this study are promising drug candidates that need further evaluation.

Synthesis and molecular docking simulation of new benzimidazole-thiazole hybrids as cholinesterase inhibitors.

Dementia is a cognitive disturbance that is generally correlated with central nervous system diseases, especially Alzheimer's disease. The limited number of medications available is insufficient to improve the lifestyle of the patients suffering from this disease. Thus, new benzimidazole-thiazole hybrids (3-10) were designed and synthesized as acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory agents. The in vitro evaluation displayed that the derivatives 4b, 4d, 5b, 6a, 7a, and 8b demonstrated dual inhibitory efficiency against both AChE with IC50 ranging from 4.55 to 8.62 µM and BChE with IC50 ranging from 3.50 to 8.32 µM. By analyzing the Lineweaver-Burk plot, an uncompetitive form of inhibition was determined for the highly active compound 4d, revealing its inhibition type. The human telomerase reverse transcriptase-immortalized retinal pigment epithelial cell line was used to ensure the safety of the most potent cholinesterase inhibitors. Furthermore, compounds 4b, 4d, 5b, 6a, 7a, and 8b were evaluated for their neuroprotective and antioxidant properties, as well as their ability to suppress COX-2. The results demonstrated that compounds 4d, 5b, and 8b presented significant neuroprotection efficiency against H2 O2 -induced damage in SH-SY5Y cells with % cell viability of 67.42 ± 7.90%, 62.51 ± 6.71%, and 72.61 ± 8.10%, respectively, while the tested candidates did not reveal significant antioxidant activity. Otherwise, compounds 4b, 6a, 7a, and 8b displayed outstanding COX-2 inhibition effects with IC50 ranging from 0.050 to 0.080 µM relative to celecoxib (IC50 = 0.050 µM). In addition, molecular docking was carried out for the potent benzimidazole-thiazole hybrids with the active sites of both AChE (PDB ID: 4EY7) and BChE (PDB code: 1P0P). The tested candidates fit well in the active sites of both portions, with docking scores ranging from -8.65 to -6.64 kcal/mol (for AChE) and -8.71 to -7.73 kcal/mol (for BChE). In silico results show that the synthesized benzimidazole-thiazole hybrids have good physicochemical and pharmacokinetic properties with no Lipinski rule violations. The preceding results exhibited that compound 4d could be used as a new template for developing more significant cholinesterase inhibitors in the future.

A novel near-infrared fluorescent probe for high-sensitivity detection of butyrylcholinesterase in various pathological states.

Butyrylcholinesterase (BChE) is a crucial hydrolytic enzyme predominantly synthesized in the liver, playing a significant role in conditions like liver disorders, diabetes, Alzheimer's disease, and fat metabolism regulation. This study aims to address the current limitations in visualizing BChE activity in diseases at various states by introducing an ultra-sensitive near-infrared fluorescent probe, FDCM-BChE. The probe was engineered to have several properties, such as a large Stokes shift, rapid response time, high stability, excellent selectivity, and low detection limits. We validated the efficacy of FDCM-BChE in quantifying BChE activity in human serum and leveraged its low cytotoxicity for cellular imaging. The study revealed the downregulation of BChE activity in liver cancer and hepatic injury and the upregulation in diabetes. Thus, FDCM-BChE shows promise as a tool for specific applications, providing insights into diseases associated with BChE activity.

Piperazine-2-carboxylic acid derivatives as MTDLs anti-Alzheimer agents: Anticholinesterase activity, mechanistic aspect, and molecular modeling studies.

Development of Multitarget-Directed Ligands (MTDLs) is a promising approach to combat the complex etiologies of Alzheimer's disease (AD). Herein we report the design, synthesis, and characterization of a new series of 1,4-bisbenzylpiperazine-2-carboxylic acid derivatives 3-5(a-g), 7a-f, 8a-s, and their piperazine-2-yl-1,3,4-oxadiazole analogs 6a-g. In vitro inhibitory effect against Electrophorus electricus acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) from Equine serum was evaluated using modified Ellman's method, considering donepezil and tacrine as reference drugs. Lineweaver-Burk plot analysis of the results proved competitive inhibition of AChE and BChE with Ki values, in low micromolar range. The free carboxylic acid series 4a-g showed enhanced selectivity for AChE. Hence, 4c, 1,4-bis (4-chlorobenzyl)-piperazinyl-2-carboxylic acid), was the most active member of this series (Ki (AChE) = 10.18 ± 1.00 µM) with clear selectivity for AChE (SI âˆ¼ 17.90). However, the hydroxamic acids 7a-f and carboxamides 8a-s congeners were more potent and selective inhibitors of BChE (SI âˆ¼ 5.38 - 21862.5). Extraordinarily, 1,4-bis (2-chlorobenzyl)-piperazinyl-2-hydroxamic acid 7b showed promising inhibitory activity against BChE enzyme (Ki = 1.6 ± 0.08 nM, SI = 21862.5), that was significantly superior to that elicited by donepezil (Ki = 12.5 ± 2.6 µM) and tacrine (Ki = 17.3 ± 2.3 nM). Cytotoxicity assessment of 4c and 7b, on human neuroblastoma (SH-SY5Y) cell lines, revealed lower toxicity than staurosporine and was nearly comparable to that of donepezil. Molecular docking and molecular dynamics simulation afforded unblemished insights into the structure-activity relationships for AChE and BChE inhibition. The results showed stable binding with fair H-bonding, hydrophobic and/or ionic interactions to the catalytic and peripheral anionic sites of the enzymes. In silico predicted ADME and physicochemical properties of conjugates showed good CNS bioavailability and safety parameters. In this regard, compound (7b) might be considered as a promising inhibitor of BChE with an innovative donepezil-based anti-Alzheimer activity. Further assessments of the most potent AChE and BChE inhibitors as potential MTDLs anti-Alzheimer's agents are under investigation with our research group and will be published later.

Serum oxidant-antioxidant status and butyrylcholinesterase activity in dogs with idiopathic epilepsy - A pilot study.

Oxidative stress plays an important role in pathogenesis of idiopathic epilepsy (IE). Although IE is the most common neurological condition, oxidant-antioxidant status in epileptic dogs is still unknown. The aim of this study is to evaluate the serum oxidant-antioxidant status in dogs with newly diagnosed IE. The status in 15 dogs with IE and 15 healthy dogs is estimated through spectrophotometric determination of two oxidant markers: advanced oxidation protein products-albumin index (AOPP) and thiobarbituric acid reactive substances (TBARS); and three antioxidant markers: total thiols (R-SH) level, glutathione (GSH) level, and paraoxonase-1 (PON-1) activity. Also, butyrylcholinesterase (BChE) activity is assessed in both groups of dogs. Higher AOPP is observed in the dogs with newly diagnosed IE, while TBARS level shows no difference when compared to the healthy dogs. In contrast, lower levels of antioxidants (R-SH, GSH, and PON-1) and BChE activity are found in the dogs with IE. No significant differences are observed in the oxidant and antioxidant markers and BChE activity across the investigated IE cases with focal and generalized seizures. Our findings provide evidence that dogs with IE are characterized by an impaired serum oxidant-antioxidant balance and lower BChE activity, which may contribute to a better understanding of IE pathogenesis.

In-vitro and in-silico studies of annelated 1,4,7,8-tetrahydroazocine ester derivatives as nanomolar selective inhibitors of human butyrylcholinesterase.

Based on previous finding showing 2,3,6,11-tetrahydro-1H-azocino[4,5-b]indole as suitable scaffold of novel inhibitors of acetylcholinesterase (AChE), a main target of drugs for the treatment of Alzheimer's disease and related dementias, herein we investigated diverse newly and previously synthesized ß-enamino esters (and ketones) derivatives of 1,4,7,8-tetrahydroazocines (and some azonines) fused with benzene, 1H-indole, 4H-chromen-4-one and pyrimidin-4(3H)-one. Twenty derivatives of diversely annelated eight-to-nine-membered azaheterocyclic ring, prepared through domino reaction of the respective tetrahydropyridine and azepine with activated alkynes, were assayed for the inhibitory activity against AChE and butyrylcholinesterase (BChE). As a major outcome, compound 7c, an alkylamino derivative of tetrahydropyrimido[4,5-d]azocine, was found to be a highly potent BChE-selective inhibitor, which showed a noncompetitive/mixed-type inhibition mechanism against human BChE with single digit nanomolar inhibition constant (Ki = 7.8 ± 0.2 nM). The four-order magnitude BChE-selectivity of 7c clearly reflects the effect of lipophilicity upon binding to the BChE binding cavity. The ChEs' inhibition data, interpreted by chemoinformatic tools and an in-depth in-silico study (molecular docking combined with molecular dynamics calculations), not only highlighted key structural factors enhancing inhibition potency and selectivity toward BChE, but also shed light on subtle differences distinguishing the binding sites of equine BChE from the recombinant human BChE. Compound 7c inhibited P-glycoprotein with IC50 of 0.27 µM, which may support its ability to permeate blood-brain barrier, and proved to be no cytotoxic in human liver cancer cell line (HepG2) at the BChE bioactive concentrations. Overall, the biological profile allows us to envision 7c as a promising template to improve design and development of BChE-selective ligands of pharmaceutical interest, including inhibitors and fluorogenic probes.

Phenyl-quinoline derivatives as lead structure of cholinesterase inhibitors with potency to reduce the GSK-3ß level targeting Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder that leads to cognitive decline and memory loss. Unfortunately, there is no effective treatment for this condition, so there is a growing interest in developing new anti-AD agents. In this research project, a series of phenyl-quinoline derivatives were designed as potential anti-AD agents. These derivatives were substituted at two different positions on benzyl and phenyl rings. The structures of the derivatives were characterized using techniques such as IR spectroscopy, 1H NMR, 13C NMR, and elemental analysis. During the in vitro screening, the derivatives were tested against both acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). It was observed that most of the derivatives showed higher selectivity against BChE compared to AChE. Among the derivatives, analog 7n (with a methoxy group at R1 and a 4-bromine substituent at R2 exhibited the highest potency, with a 75-fold improvement in the activity compared to the positive control. Importantly, this potent analog demonstrated no toxicity at the tested concentration on SH-SY5Y cells, indicating its potential as a safe anti-AD agent. The level of GSK-3ß was also reduced after treatments with 7n at 50 µM. Overall, this study highlights the design and evaluation of phenyl-quinoline derivatives as promising candidates for developing novel anti-AD agents.

Evaluation of 4-aminoquinoline derivatives with an n-octylamino spacer as potential multi-targeting ligands for the treatment of Alzheimer's disease.

The most successful therapeutic strategy in the treatment of Alzheimer's disease (AD) is directed toward increasing levels of the neurotransmitter acetylcholine (ACh) by inhibition of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), the enzymes responsible for its hydrolysis. In this paper, we extended our study on 4-aminoquinolines as human cholinesterase inhibitors on twenty-six new 4-aminoquinolines containing an n-octylamino spacer on C(4) and different substituents on the terminal amino group. We evaluated the potency of new derivatives to act as multi-targeted ligands by determining their inhibition potency towards human AChE and BChE, ability to chelate biometals Fe, Cu and Zn, ability to inhibit the action of ß-secretase 1 (BACE1) and their antioxidant capacity. All of the tested derivatives were very potent inhibitors of human AChE and BChE with inhibition constants (Ki) ranging from 0.0023 to 1.6 µM. Most of the compounds were estimated to be able to cross the blood-brain barrier (BBB) by passive transport and were nontoxic to human neuronal, kidney and liver cells in concentrations in which they inhibit cholinesterases. Generally, newly synthesised compounds were weak reductants compared to standard antioxidants, but all possessed a certain amount of antioxidant activity compared to tacrine. Of the eleven most potent cholinesterase inhibitors, eight compounds also inhibited BACE1 activity at 10-18%. Based on our overall results, compounds 8 with 3-fluorobenzyl, 11 with 3-chlorobenzyl and 17 with 3-metoxy benzyl substituents on the terminal amino group stood out as the most promising for the treatment of AD; they strongly inhibited AChE and BChE, were non-toxic on HepG2, HEK293 and SH-SY5Y cells, had the potential to cross the BBB and possessed the ability to chelate biometals and/or inhibit the activity of BACE1 within a range close to the therapeutically desired degree of inhibition.