Search for multifunctional agents against Alzheimer’s disease among non-imidazole histamine H3 receptor ligands. In vitro and in vivo pharmacological evaluation and computational studies of piperazine derivatives

In the search for new treatments for complex disorders such as Alzheimer’s disease the Multi-Target-Directed Ligands represent a very promising approach. The aim of the present study was to identify multifunctional compounds among several series of non-imidazole histamine H3 receptor ligands, derivatives of 1-[2-thiazol-5-yl-(2-aminoethyl)]-4-n-propylpiperazine, 1-[2-thiazol-4-yl-(2-aminoethyl)]-4-n-propylpiperazine and 1-phenoxyalkyl-4-(amino)alkylopiperazine using in vitro and in vivo pharmacological evaluation and computational studies. Performed in vitro assays showed moderate potency of tested compounds against acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). Molecular modeling studies have revealed possible interactions between the active compounds and both AChE and BuChE as well as the human H3 histamine receptor. Computational studies showed the high drug-likeness of selected compounds with very good physicochemical profiles. The parallel artificial membrane permeation assay proved outstanding blood–brain barrier penetration in test conditions. The most promising compound, A12, chemically methyl(4-phenylbutyl){2-[2-(4-propylpiperazin-1-yl)-1,3-thiazol-5-yl]ethyl}amine, possesses good balanced multifunctional profile with potency toward studied targets - H3 antagonist activity (pA₂ = 8.27), inhibitory activity against both AChE (IC₅₀ = 13.96 μM), and BuChE (IC₅₀ = 14.62 μM). The in vivo pharmacological studies revealed the anti-amnestic properties of compound A12 in the passive avoidance test on mice.     

Design,synthesis, and pharmacological evaluation of 2-amino-5-nitrothiazole derived semicarbazones as dual inhibitors of monoamine oxidase and cholinesterase: effect of the size of aryl binding site

A series of 2-amino-5-nitrothiazole derived semicarbazones were designed, synthesised and investigated for MAO and ChE inhibition properties. Most of the compounds showed preferential inhibition towards MAO-B. Compound 4, (1-(1-(4-Bromophenyl)ethylidene)-4-(5-nitrothiazol-2-yl)semicarbazide) emerged as lead candidate (IC₅₀?=?0.212?µM, SI?=?331.04) against MAO-B; whereas compounds 21 1-(5-Bromo-2-oxoindolin-3-ylidene)-4-(5-nitrothiazol-2-yl)semicarbazide (IC₅₀?=?0.264?µM) and 17 1-((4-Chlorophenyl) (phenyl)methylene)-4-(5-nitrothiazol-2-yl)semicarbazide (IC₅₀?=?0.024?µM) emerged as lead AChE and BuChE inhibitors respectively; with activity of compound 21 almost equivalent to tacrine. Kinetic studies indicated that compound 4 exhibited competitive and reversible MAO-B inhibition while compounds 21 and 17 showed mixed-type of AChE and BuChE inhibition respectively. Docking studies revealed that these compounds were well-accommodated within MAO-B and ChE active sites through stable hydrogen bonding and/or hydrophobic interactions. This study revealed the requirement of small heteroaryl ring at amino terminal of semicarbazone template for preferential inhibition and selectivity towards MAO-B. Our results suggest that 5-nitrothiazole derived semicarbazones could be further exploited for its multi-targeted role in development of anti-neurodegenerative agents.

A library of 2-amino-5-nitrothiazole derived semicarbazones (4–21) was designed, synthesised and evaluated for in vitro MAO and ChE inhibitory activity. Compounds 4, 21 and 17 (shown) have emerged as lead MAO-B (IC₅₀:0.212?µM, competitive and reversible), AChE (IC₅₀:0.264?µM, mixed and reversible) and BuChE (IC₅₀:0.024?µM, mixed and reversible) inhibitor respectively. SAR studies disclosed several structural aspects significant for potency and selectivity and indicated the role of size of aryl binding site in potency and selectivity towards MAO-B. Antioxidant activity and neurotoxicity screening results further suggested their multifunctional potential for the therapy of neurodegenerative diseases.     

Assay of cholinesterase using a proenhancer of the luminol–H2O2–horseradish peroxidase reaction

2-Naphthyl acetate acts as a pro-enhancer of the luminol–H₂O₂–horseradish peroxidase reaction. Cholinesterase hydrolyses the bound acetyl group and produces 2-naphthol, and this compound is an enhancer of the chemiluminescent reaction. We studied the kinetics of chemiluminescent emission and the influence of 2-naphthyl acetate and cholinesterase enzyme concentration. The cholinesterase concentration versus chemiluminescence intensity maximum was linear for cholinesterase between 0 and 181 μU/mL, with a detection limit of 8 μU/mL and a relative standard deviation of 9.5% (n = 3), for a sample containing 90.67 μU/mL of cholinesterase.     

Insights into (S)-rivastigmine inhibition of butyrylcholinesterase (BuChE): Molecular docking and saturation transfer difference NMR (STD-NMR)

Rivastigmine is a very important drug prescribed for the treatment of Alzheimer’s disease (AD) symptoms. It is a dual inhibitor, in that it inhibits both acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). For our screening program on the discovery of new rivastigmine analogue hits for human butyrylcholinesterase (hBuChE) inhibition, we investigated the interaction of this inhibitor with BuChE using the complimentary approach of the biophysical method, saturation transfer difference (STD)-NMR and molecular docking. This allowed us to obtain essential information on the key binding interactions between the inhibitor and the enzyme to be used for screening of hit compounds. The main conclusions obtained from this integrated study was that the most dominant interactions were (a) H-bonding between the carbamate carbonyl of the inhibitor and the NH group of the imidazole unit of H434, (b) stacking of the aromatic unit of the inhibitor and the W82 aromatic unit in the choline binding pocket via π-π interactions and (c) possible CH/π interactions between the benzylic methyl group and the N-methyl groups of the inhibitor and W82 of the enzyme.     

Synthesis and evaluation of multi-target-directed ligands for the treatment of Alzheimer’s disease based on the fusion of donepezil and melatonin

A novel series of compounds obtained by fusing the acetylcholinesterase (AChE) inhibitor donepezil and the antioxidant melatonin were designed as multi-target-directed ligands for the treatment of Alzheimer’s disease (AD). In vitro assay indicated that most of the target compounds exhibited a significant ability to inhibit acetylcholinesterase (eeAChE and hAChE), butyrylcholinesterase (eqBuChE and hBuChE), and β-amyloid (Aβ) aggregation, and to act as potential antioxidants and biometal chelators. Especially, 4u displayed a good inhibition of AChE (IC₅₀ value of 193 nM for eeAChE and 273 nM for hAChE), strong inhibition of BuChE (IC₅₀ value of 73 nM for eqBuChE and 56 nM for hBuChE), moderate inhibition of Aβ aggregation (56.3% at 20 μM) and good antioxidant activity (3.28 trolox equivalent by ORAC assay). Molecular modeling studies in combination with kinetic analysis revealed that 4u was a mixed-type inhibitor, binding simultaneously to catalytic anionic site (CAS) and the peripheral anionic site (PAS) of AChE. In addition, 4u could chelate metal ions, reduce PC12 cells death induced by oxidative stress and penetrate the blood–brain barrier (BBB). Taken together, these results strongly indicated the hybridization approach is an efficient strategy to identify novel scaffolds with desired bioactivities, and further optimization of 4u may be helpful to develop more potent lead compound for AD treatment.     

Design,synthesis and evaluation of novel 7-aminoalkyl-substituted flavonoid derivatives with improved cholinesterase inhibitory activities

A novel series of 7-aminoalkyl-substituted flavonoid derivatives 5a–5r were designed, synthesized and evaluated as potential cholinesterase inhibitors. The results showed that most of the synthesized compounds exhibited potent acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities at the micromolar range. Compound 2-(naphthalen-1-yl)-7-(8-(pyrrolidin-1-yl)octyloxy)-4H-chromen-4-one (5q) showed the best inhibitory activity (IC₅₀, 0.64 μM for AChE and 0.42 μM for BChE) which were better than our previously reported compounds and the commercially available cholinergic agent Rivastigmine. The results from a Lineweaver–Burk plot indicated a mixed-type inhibition for compound 5q with AChE and BChE. Furthermore, molecular modeling study showed that 5q targeted both the catalytic active site (CAS) and the peripheral anionic site (PAS) of AChE. Besides, these compounds (5a–5r) did not affect PC12 and HepG2 cell viability at the concentration of 10 μM. Consequently, these flavonoid derivatives should be further investigated as multipotent agents for the treatment of Alzheimer’s disease.     

Modification of Bischler-Möhlau indole derivatives through palladium catalyzed Suzuki reaction as effective cholinesterase inhibitors,their kinetic and molecular docking studies

Due to the immense importance of aryl indole nucleus, herein we report the palladium-catalyzed arylation of N-substituted 2-aryl indole utilizing Suzuki-Miyaura cross coupling methodology. The biological screening for cholinesterase inhibition of the resulted biaryl indole moieties was carried out to evaluate their pharmacological potential, expecting to involve the development of new therapeutics for various inflammatory, cardiovascular, gastrointestinal and neurological diseases. This research work also involved the use of utilization of microwave-assisted organic synthesis (MAOS) for the synthesis of Bischler-Möhlau indole which is further biarylated via palladium-catalyzed cross coupling reaction. All the synthetic compounds (3a-n) were tested for cholinesterase inhibition and exhibited high level of AChE inhibitory activities. Interestingly, compounds 3m and 3n were found to be dual inhibitors, however, remaining compound exhibited no inhibitory activity against BChE. The biological potential of the resulted compounds was explained on the basis of molecular docking studies, performed against AChE and BChE, exploring the probable binding modes of most potent inhibitors.     

Alkaloids from Galanthus fosteri

Three new alkaloids, oxoincartine, 3,11-O-diacetyl-9-O-demethylmaritidine and 11-O-acetyl-9-O-demethylmaritidine together with seven known compounds namely, incartine, galanthamine, galanthine, 9-O-methylpseudolycorine, N,O-dimethylnorbelladine, hordenine and vittatine were isolated from Galanthus fosteri Baker (Amaryllidaceae). Their structures were elucidated by spectroscopic analyses (UV, IR, MS, CD and 1D/2D NMR). Cholinesterase inhibitory activity potentials of the compounds were also determined.     

Synthesis and cholinesterase inhibition of cativic acid derivatives

Alzheimer’s disease (AD) is a neurodegenerative disorder associated with memory impairment and cognitive deficit. Most of the drugs currently available for the treatment of AD are acetylcholinesterase (AChE) inhibitors. In a preliminary study, significant AChE inhibition was observed for the ethanolic extract of Grindelia ventanensis (IC₅₀ = 0.79 mg/mL). This result prompted us to isolate the active constituent, a normal labdane diterpenoid identified as 17-hydroxycativic acid (1), through a bioassay guided fractionation. Taking into account that 1 showed moderate inhibition of AChE (IC₅₀ = 21.1 μM), selectivity over butyrylcholinesterase (BChE) (IC₅₀ = 171.1 μM) and that it was easily obtained from the plant extract in a very good yield (0.15% w/w), we decided to prepare semisynthetic derivatives of this natural diterpenoid through simple structural modifications. A set of twenty new cativic acid derivatives (3–6) was prepared from 1 through transformations on the carboxylic group at C-15, introducing a C2–C6 linker and a tertiary amine group. They were tested for their inhibitory activity against AChE and BChE and some structure–activity relationships were outlined. The most active derivative was compound 3c, with an IC₅₀ value of 3.2 μM for AChE. Enzyme kinetic studies and docking modeling revealed that this inhibitor targeted both the catalytic active site and the peripheral anionic site of this enzyme. Furthermore, 3c showed significant inhibition of AChE activity in SH-SY5Y human neuroblastoma cells, and was non-cytotoxic.     

Synthesis of 5-methyl-2,4-dihydro-3H-1,2,4-triazole-3-one’s aryl Schiff base derivatives and investigation of carbonic anhydrase and cholinesterase (AChE,BuChE) inhibitory properties

Carbonic anhydrase enzymes (EC 4.2.1.1, CAs) are metalloenzyme families that catalyze the rapid conversion of H₂O and CO₂ to HCO₃^– and H⁺. CAs are found in different tissues where they participate in various significant biochemical processes such as ion transport, carbon dioxide respiration, ureagenesis, lipogenesis, bone resorption, electrolyte secretion, acid-base balance, and gluconeogenesis. In such processes, many CAs are significant therapeutic targets because of their inhibitory potentials especially in the treatment of some diseases such as edema, glaucoma, obesity, cancer, epilepsy, and osteoporosis. Acetylcholinesterase (AChE) and Butyrylcholinesterase (BuChE) inhibitors are also valuable compounds for different therapeutic applications including Alzheimer’s disease. In this work, we report a fast and effective synthesis of 5-methyl-2,4-dihydro-3H-1,2,4-triazole-3-one’s aryl Schiff base derivatives and also their CA and cholinesterases inhibitory properties. Our findings showed that these Schiff base derivatives, with triazole ring, found as strong CA and cholinesterases inhibitors.