Design,synthesis and evaluation of 2,4-disubstituted pyrimidines as cholinesterase inhibitors

A group of 2,4-disubstituted pyrimidine derivatives (7a–e, 8a–e and 9a–d) that possess a variety of C-2 aliphatic five- and six-membered heterocycloalkyl ring in conjunction with a C-4 arylalkylamino substituent were designed, synthesized and evaluated as cholinesterase (ChE) inhibitors. The steric and electronic properties at C-2 and C-4 positions of the pyrimidine ring were varied to investigate their effect on ChE inhibitory potency and selectivity. The structure–activity relationship (SAR) studies identified N-benzyl-2-thiomorpholinopyrimidin-4-amine (7c) as the most potent cholinesterase inhibitor (ChEI) with an IC₅₀ = 0.33 μM (acetylcholinesterase, AChE) and 2.30 μM (butyrylcholinesterase, BuChE). The molecular modeling studies indicate that within the AChE active site, the C-2 thiomorpholine substituent was oriented toward the cationic active site region (Trp84 and Phe330) whereas within the BuChE active site, it was oriented toward a hydrophobic region closer to the active site gorge entrance (Ala277). Accordingly, steric and electronic properties at the C-2 position of the pyrimidine ring play a critical role in ChE inhibition.     

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.     

Novel coumarin derivatives bearing N-benzyl pyridinium moiety: Potent and dual binding site acetylcholinesterase inhibitors

A novel series of coumarin derivatives linked to benzyl pyridinium group were synthesized and biologically evaluated as inhibitors of both acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). The enzyme inhibitory activity of synthesized compounds was measured using colorimetric Ellman’s method. It was revealed that compounds 3e, 3h, 3l, 3r and 3s have shown higher activity compared with donepezil hydrochloride as standard drug. Most of the compounds in these series had nanomolar range IC₅₀ in which compound 3r (IC₅₀ = 0.11 nM) was the most active compound against acetylcholinesterase enzyme.     

New 30-norlupane derivatives through chemical-microbial semi-synthesis of betulinic acid and their neuroprotective effect

In this study, seven 30-norlupane derivatives (2–8) was obtained from the chemical oxidation of betulinic acid followed by biotransformation via Bacillus megaterium CGMCC 1.1741. And metabolites 2–4 and 6–8 were newly identified products. In the first step, betulinic acid was chemically oxidized to platanic acid (1). Following the chemical oxidation, B. megaterium catalyzed the hydroxylation at C-7, C-11, C-15 and C-23 of platanic acid (1) as well as the oxidation of C-3 hydroxyl group. Compared to the labor-intensive isolation from natural plants, this chemical-microbial semi-synthesis is more capable to provide increased structural diversity of oxygenated 30-norlupane. Finally, the potential neuroprotective effect of the derivatives was assessed on neuron-like PC12 cells induced by cobalt chloride (CoCl₂). Metabolite 6 showed a potent neuroprotective activity.     

The first synthesis of 4-phenylbutenone derivative bromophenols including natural products and their inhibition profiles for carbonic anhydrase,acetylcholinesterase and butyrylcholinesterase enzymes

The first synthesis of (E)-4-(3-bromo-4,5-dihydroxyphenyl)but-3-en-2-one (1), (E)-4-(2-bromo-4,5-dihydroxyphenyl)but-3-en-2-one (2), and (E)-4-(2,3-dibromo-4,5-dihydroxyphenyl)but-3-en-2-one (3) was realized as natural bromophenols. Derivatives with mono OMe of 2 and 3 were obtained from the reactions of their derivatives with di OMe with AlCl₃. These novel 4-phenylbutenone derivatives were effective inhibitors of the cytosolic carbonic anhydrase I and II isoenzymes (hCA I and II), acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) with K_i values in the range of 158.07–404.16 pM for hCA I, 107.63–237.40 pM for hCA II, 14.81–33.99 pM for AChE and 5.64–19.30 pM for BChE. The inhibitory effects of the synthesized novel 4-phenylbutenone derivatives were compared to acetazolamide as a clinical hCA I and II isoenzymes inhibitor and tacrine as a clinical AChE and BChE enzymes inhibitor.     

Cytotoxicity and inhibition of DNA topoisomerase I of polyhydroxylated triterpenoids and triterpenoid glycosides

Cytotoxicity and inhibition on human DNA topoisomerase I (TOP1) and II (TOP2) of 74 plant-originated triterpenoids and triterpenoid glycosides were investigated. The cytotoxic compounds are primarily polyhydroxylated oleananes (GI₅₀ of A549: 1.0–10.19 μM). Sixteen cytotoxic aesculiosides isolated from Aesculus pavia inhibited TOP1 catalytic activity by interacting directly with the free enzyme and preventing the formation of the DNA–TOP1 complex. Interestingly, hydrolysis of six active aesculiosides (1, 4, 6, 8, 10, and 23) lost their TOP1 activities but enhanced their cytotoxicities. None of the test compounds showed any activity against TOP2. Structure–activity relationship (SAR) investigations indicated that cytotoxic oleananes required at least one angeloyl moiety at either C-21 or C-22 but the sugar moiety at C-3 may decrease their cytotoxicities. An angeloyl or tigeloyl group at C-21 is required for oleananes to bind the free TOP1 enzyme although the type and length of acyl moiety at C-22 also affects their activity. However, sugar moiety at C-3 is necessary for their TOP1 activities.     

Potential anti-inflammatory phenolic glycosides from the medicinal plant Moringa oleifera fruits

Bioassay-guided isolation and purification of the ethyl acetate extract of Moringa oleifera fruits yielded three new phenolic glycosides; 4-[(2′-O-acetyl-α-l-rhamnosyloxy) benzyl]isothiocyanate (1), 4-[(3′-O-acetyl-α-l-rhamnosyloxy)benzyl]isothiocyanate (2), and S-methyl-N-{4-[(α-l-rhamnosyloxy)benzyl]}thiocarbamate (3), together with five known phenolic glycosides (4–8). The structures of the new metabolites were determined on the basis of spectroscopic analyses including 1D- and 2D-NMR and mass spectrometry. The anti-inflammatory activity of isolated compounds was investigated with the lipopolysaccharide (LPS)-induced murine macrophage RAW 264.7 cell line. It was found that 4-[(2′-O-acetyl-α-l-rhamnosyloxy)benzyl]isothiocyanate (1) possessed potent NO–inhibitory activity with an IC₅₀ value of 1.67 μM, followed by 2 (IC₅₀ = 2.66 μM), 4 (IC₅₀ = 2.71 μM), and 5 (IC₅₀ = 14.4 μM), respectively. Western blots demonstrated these compounds reduced LPS-mediated iNOS expression. In the concentration range of the IC₅₀ values, no significant cytotoxicity was noted. Structure–activity relationships following NO-release indicated: (1) the isothiocyanate group was essential for activity, (2) acetylation of the isothiocyanate derivatives at C-2′ or at C-3′ of rhamnose led to higher activity, (3) un-acetylated isothiocyanate derivatives displayed eight times less activity than the acetylated derivatives, and (4) acetylation of the thiocarbamate derivatives enhanced activity. These data indicate compounds 1, 2, 4 and 5 are responsible for the reported NO-inhibitory effect of Moringa oleifera fruits, and further studies are warranted.     

Design,synthesis and evaluation of novel 4-dimethylamine flavonoid derivatives as potential multi-functional anti-Alzheimer agents

A series of 4-dimethylamine flavonoid derivatives 5a–5r were designed, synthesized and evaluated as potential multi-functional anti-Alzheimer agents. The results showed that most of the synthesized compounds exhibited high acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activity at the micromolar range (IC₅₀, 1.83–33.20 μM for AChE and 0.82–11.45 μM for BChE). A Lineweaver–Burk plot indicated a mixed-type inhibition for compound 5j with AChE, and molecular modeling study showed that 5j targeted both the catalytic active site (CAS) and the peripheral anionic site (PAS) of AChE. Besides, the derivatives showed potent self-induced Aβ aggregation inhibitory activity at 20 μM with percentage from 25% to 48%. In addition, some compounds (5j–5q) showed potent oxygen radical absorbance capacity (ORAC) ranging from 1.5- to 2.6-fold of the Trolox value. These compounds should be further investigated as multi-potent agents for the treatment of Alzheimer’s disease.     

Synthesis of isothiazol-3-one derivatives as inhibitors of histone acetyltransferases (HATs)

High-throughput screening led to the identification of isothiazolones 1 and 2 as inhibitors of histone acetyltransferase (HAT) with IC50s of 3 μM and 5 μM, respectively. Analogues of these hit compounds with variations of the N-phenyl group, and with variety of substituents at C-4, C-5 of the thiazolone ring, were prepared and assayed for inhibition of the HAT enzyme PCAF. Potency is modestly favoured when the N-aryl group is electron deficient (4-pyridyl derivative 10 has IC₅₀ = 1.5 μM); alkyl substitution at C-4 has little effect, whilst similar substitution at C-5 causes a significant drop in potency. The ring–fused compound 38 has activity (IC₅₀ = 6.1 μM) to encourage further exploration of this bicyclic structure. The foregoing SAR is consistent with an inhibitory mechanism involving cleavage of the S–N bond of the isothiazolone ring by a catalytically important thiol residue.     

Microwave assisted synthesis,cholinesterase enzymes inhibitory activities and molecular docking studies of new pyridopyrimidine derivatives

A series of hitherto unreported pyrido-pyrimidine-2-ones/pyrimidine-2-thiones were synthesized under microwave assisted solvent free reaction conditions in excellent yields and evaluated in vitro for their acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes inhibitory activity. Among the pyridopyrimidine derivatives, 7e and 7l displayed 2.5- and 1.5-fold higher enzyme inhibitory activities against AChE as compared to standard drug, galanthamine, with IC₅₀ of 0.80 and 1.37 μM, respectively. Interestingly, all the compounds except 6k, 7j and 7k displayed higher inhibitory potential against BChE enzyme in comparison to standard with IC₅₀ ranging from 1.18 to 18.90 μM. Molecular modeling simulations of 7e and 7l was performed using three-dimensional structure of Torpedo californica AChE (TcAChE) and human butyrylcholinesterase (hBChE) enzymes to disclose binding interaction and orientation of these molecule into the active site gorge of respective receptors.     

Inhibition of neuraminidase activity by polyphenol compounds isolated from the roots of Glycyrrhiza uralensis

We isolated 18 polyphenols with neuraminidase inhibitory activity from methanol extracts of the roots of Glycyrrhiza uralensis. These polyphenols consisted of four chalcones (1–4), nine flavonoids (5–13), four coumarins (14–17), and one phenylbenzofuran (18). When we tested the effects of these individual compounds and analogs thereof on neuraminidase activation, we found that isoliquiritigenin (1, IC₅₀ = 9.0 μM) and glycyrol (14, IC₅₀ = 3.1 μM) had strong inhibitory activity. Structure–activity analysis showed that the furan rings of the polyphenols were essential for neuraminidase inhibitory activity, and that this activity was enhanced by the apioside group on the chalcone and flavanone backbone. In addition, the presence of a five-membered ring between C-4 and C-2′ in coumestan was critical for neuraminidase inhibition. All neuraminidase inhibitors screened were found to be reversible noncompetitive inhibitors.     

Design,synthesis and biological evaluation of new 2,3-diarylquinoline derivatives as selective cyclooxygenase-2 inhibitors

A new group of 2,3-diarylquinoline derivatives possessing a methylsulfonyl COX-2 pharmacophore at the para-position of the C-2 phenyl ring were synthesized and evaluated as selective COX-2 inhibitors. In vitro COX-1/COX-2 structure–activity relationships were determined by varying the substituents on the C-4 quinoline ring. Among the 2,3-diarylquinolines, 2-(4-(methylsulfonyl) phenyl)-3-phenylquinoline-4-carboxylic acid (8) exhibited the highest potency and selectivity for COX-2 inhibitory activity (COX-2 IC₅₀ = 0.07 μM; selectivity index = 687.1) that was more selective than the reference drug celecoxib (COX-2 IC₅₀ = 0.06 μM; selectivity index = 405). A molecular modeling study where 8 was docked in the binding site of COX-2 indicated that the p-MeSO₂ COX-2 pharmacophore group on the C-2 phenyl ring is oriented in the vicinity of the COX-2 secondary pocket (Arg⁵¹³, Phe⁵¹⁸ and Val⁵²³) and the carboxylic acid substituent can interact with Ser⁵³⁰. The structure activity data acquired indicate that the size and nature of the C-4 quinoline substituent are important for COX-2 inhibitory activity.     

Anti-HBV agents. Part 2: Synthesis and in vitro anti-hepatitis B virus activities of alisol A derivatives

Chemical modifications were performed on hydroxyl groups at C-11,23,24,25 positions and C-13(17) double bond of alisol A for structure–activity relationship study. Forty-one derivatives of alisol A were synthesized and assayed for their in vitro anti-hepatitis B virus (HBV) activities and cytotoxicities. Of them, 14 compounds were active against HBV surface antigen (HBsAg) and HBV e antigen (HBeAg) secretion in HepG 2.2.15 cells, and the most promising compound 25 exhibited high activities against secretion of HBsAg (IC₅₀ = 0.028 mM), HBeAg (IC₅₀ = 0.027 mM) and remarkable selective indices (SI_HBsAg >90, SI_HBeAg >93).     

Design,synthesis and in vitro cytotoxicity evaluation of 5-(2-carboxyethenyl)isatin derivatives as anticancer agents

Forty four di- or trisubstituted novel isatin derivatives were designed and synthesized in 5–6 steps in 25–45% overall yields. Their structures were confirmed by ¹H NMR and ¹³C NMR as well as LC–MS. The anticancer activity of these new isatin derivatives against three human tumor cell lines, K562, HepG2 and HT-29, were evaluated by MTT assay in vitro. SAR studies suggested that the combination of 1-benzyl and 5-[trans-2-(methoxycarbonyl)ethen-1-yl] substitution greatly enhance their cytotoxic activity, whereas an intact carbonyl functionality on C-3 as present in the parent ring is required to such a potency. This study leads to the identification of two highly active molecules, compounds 2h (IC₅₀ = 3 nM) and 2k (IC₅₀ = 6 nM), against human leukemia K562 cells.     

Design and synthesis of thiourea derivatives with sulfur-containing heterocyclic scaffolds as potential tyrosinase inhibitors

Tyrosinase is a key enzyme during the production of melanins in plants and animals. A class of novel N-aryl-N′-substituted phenylthiourea derivatives (3a–i, 6a–k) were designed, synthesized and their inhibitory effects on the diphenolase activity of mushroom tyrosinase were evaluated. The results showed some 4,5,6,7-tetrahydro-2-[[(phenylamino)thioxomethyl]amino]-benzo[b]thiophene-3-carboxylic acid derivatives (3a–i) exhibited moderate inhibitory potency on diphenolase activity of tyrosinase. When the scaffold of 4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxylic acid was replaced with 2-(1,3,4-thiadiazol-2-yl)thio acetic acid, the inhibitory activity of compounds (6a–k) against tyrosinase was improved obviously; especially, the inhibitory activity of compound 6h (IC₅₀ = 6.13 μM) is significantly higher than kojic acid (IC₅₀ = 33.3 μM). Moreover, the analysis on inhibition mechanism revealed that compound 6h might plays the role as a noncompetitive inhibitor.     

Synthesis and characterization of 1H-phenanthro[9,10-d]imidazole derivatives as multifunctional agents for treatment of Alzheimer's disease

BackgroundAlzheimer's disease (AD) is a progressive neurodegenerative brain disorder that is characterized by dementia, cognitive impairment, and memory loss. Diverse factors are related to the development of AD, such as increased level of β-amyloid (Aβ), acetylcholine, metal ion deregulation, hyperphosphorylated tau protein, and oxidative stress.MethodsThe following methods were used: organic syntheses of 1H-phenanthro[9,10-d]imidazole derivatives, inhibition of self-mediated and metal-induced Aβ_1–42 aggregation, inhibition studies for acetylcholinesterase and butyrylcholinesterase, anti-oxidation activity studies, CD, MTT assay, transmission electron microscopy, dot plot assay, gel electrophoresis, Western blot, and molecular docking studies.ResultsWe synthesized and characterized a new type of 1H-phenanthro[9,10-d]imidazole derivatives as multifunctional agents for AD treatment. Our results showed that most of these derivatives exhibited strong Aβ aggregation inhibitory activity. Compound 9g had 74% Aβ_1–42 aggregation inhibitory effect at 10 μM concentration with its IC₅₀ value of 6.5 μM for self-induced Aβ_1–42 aggregation. This compound also showed good inhibition of metal-mediated (Cu^2 + and Fe^2 +) and acetylcholinesterase-induced Aβ_1–42 aggregation, as indicated by using thioflavin T assay, transmission electron microscopy, gel electrophoresis, and Western blot. Besides, compound 9g exhibited cholinesterase inhibitory activity, with its IC₅₀ values of 0.86 μM and 0.51 μM for acetylcholinesterase and butyrylcholinesterase, respectively. In addition, compound 9g showed good anti-oxidation effect with oxygen radical absorbance capacity (ORAC) value of 2.29.ConclusionsCompound 9g was found to be a potent multi-target-directed agent for Alzheimer's disease.General significanceCompound 9g could become a lead compound for further development as a multi-target-directed agent 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.     

Microwave assisted synthesis of novel hybrid tacrine-sulfonamide derivatives and investigation of their antioxidant and anticholinesterase activities

A novel series of tacrine derivatives containing sulfonamide group were synthesized and their inhibitory effects on acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) were evaluated. The result showed that all the synthesized tacrine-sulfonamides (VIIIa–o) exhibited inhibitory activity on both cholinesterases. VIIIg showed the highest inhibitory activity on AChE IC₅₀ = 0.009 μM. This value is 220-fold greater than that of galantamine (IC₅₀ = 2.054 μM) and 6-fold greater than tacrine (IC₅₀ = 0.055 μM). VIIIf displayed the strongest inhibition of BuChE (IC₅₀ = 2.250 μM), which is close to donepezil (IC₅₀ = 2.680 μM) and 8-fold greater than that of galantamine (IC₅₀ = 18.130 μM) Furthermore, all of the synthesized tacrine derivatives showed higher inhibition of BuChE than that of galantamine. In addition, the cupric reducing antioxidant capacities (CUPRAC) and ABTS cation radical scavenging abilities of the synthesized compounds were investigated for the antioxidant activity. Among them, VIIIb (IC₅₀ = 94.390 ± 2.310 μM) showed significantly better ABTS cation radical scavenging ability than all of the new synthesized compounds.     

Rational design and synthesis of dihydropyrimidine based dual binding site acetylcholinesterase inhibitors

Based on the pharmacological importance of dihydropyrimidine (DHPM) scaffold, substituted DHPMs linked with acetamide linker to substituted aromatic anilines were synthesized and evaluated for their potency as acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitors. The good AChE inhibitory activity of 4-dihydropyrimidine-2-thione (4a–h) and 2-amino-1,4-dihyropyrimidines (5a–h) series was observed with compound 4a and 4d identified as the most potent compounds with IC₅₀ values of 0.17 ± 0.01 and 0.39 ± 0.04 μM respectively. The inhibition of BChE was found in a broader range of concentrations (2.37–56.32 μM). To explore the binding insights into the enzyme, molecular docking study was carried out using GOLD software. The binding mode analysis indicated that all of these inhibitors are well accommodated in the active site and interact with the key amino acid residues of Catalytic anionic site (CAS) and peripheral anionic site (PAS). Furthermore, in silico ADMET predictions suggest that these compounds are non-AMES toxic with good blood brain barrier (BBB) penetration, human intestinal absorption.     

In vitro and in vivo antimalarial evaluation of semi-synthetic derivatives of gomphostenin

A novel series of semi-synthetic gomphostenin derivatives (1–9) were prepared utilizing C-14 hydroxyl group for the first time and studied for their antimalarial properties. In vitro antiplasmodial activity was evaluated against both the chloroquine sensitive and resistant strains of Plasmodium falciparum. Most of the compounds exhibited superior or comparable antiplasmodial activity compared to parent compound, that is, gomphostenin (GN). Based upon in vitro antiplasmodial activity, compounds with IC₅₀ values less than 10 μM were selected for in vivo antiplasmodial evaluation against Plasmodium berghei infection in mice model. GN derivatives 3 and 5 were found to have curative activity with moderate chemosuppression of 65% and 69%, respectively, at the dose level of 150 mg/kg/day.