A Novel Galantamine-Curcumin Hybrid as a Potential Multi-Target Agent against Neurodegenerative Disorders.

The acetylcholinesterase (AChE) inhibitors are the main drugs for symptomatic treatment of neurodegenerative disorders like Alzheimer's disease. A recently designed, synthesized and tested hybrid compound between the AChE inhibitor galantamine (GAL) and the antioxidant polyphenol curcumin (CU) showed high AChE inhibition in vitro. Here, we describe tests for acute and short-term toxicity in mice as well as antioxidant tests on brain homogenates measured the levels of malondialdehide (MDA) and glutathione (GSH) and in vitro DPPH, ABTS, FRAP and LPO inhibition assays. Hematological and serum biochemical analyses were also performed. In the acute toxicity tests, the novel AChE inhibitor given orally in mice showed LD50 of 49 mg/kg. The short-term administration of 2.5 and 5 mg/kg did not show toxicity. In the ex vivo tests, the GAL-CU hybrid performed better than GAL and CU themselves; in a dose of 5 mg/kg, it demonstrates 25% reduction in AChE activity, as well as a 28% and 73% increase in the levels of MDA and GSH, respectively. No significant changes in blood biochemical data were observed. The antioxidant activity of 4b measured ex vivo was proven in the in vitro tests. In the ABTS assay, 4b showed radical scavenging activity 10 times higher than the positive control butylhydroxy toluol (BHT). The GAL-CU hybrid is a novel non-toxic AChE inhibitor with high antioxidant activity which makes it a prospective multitarget drug candidate for treatment of neurodegenerative disorders.

Synthesis of New Galanthamine-Peptide Derivatives Designed for Prevention and Treatment of Alzheimer's Disease.

BACKGROUND: Although no effective treatment for the Alzheimer's disease currently exist, some drugs acting as Acetylcholinesterase inhibitors, like galanthamine have positively affected such patients. ß- and/or γ-secretase inhibitors are another type of potential drugs. Here we report synthesis of new peptide-galanthamine derivatives, with expected inhibitory activity against both Acetylcholinesterase and ß-secretase. OBJECTIVES: The aim of this work is obtaining new peptide derivatives of galanthamine with decreased toxicity compared to galanthamine. METHODS: Syntheses were conducted in solution using fragment condensation approach. The new derivatives were characterized by melting points, angle of optical rotation, NMR and Mass spectra. Acute toxicity was determined on mice, according to a Standard protocol. All new compounds were tested in vitro for cytotoxic activity in a panel of human (HEP-G2, BV-173) and murine (Neuro-2a) tumor cell lines via a standard MTT-based colorimetric method. RESULTS: New derivatives of galanthamine containing shortened analogues of ß-secretase inhibitor (Boc- Asn-Leu-Ala-Val-OH) and either nicotinic or isonicotinic residue, both connected with a linker (L-Asp) to position 11 of galanthamine were obtained. In vivo toxicity of some new compounds was found up to 1000 mg/kg. Cell toxicity screening against the tumor cell lines showed negligible growth-inhibiting properties of the galanthamine derivatives. CONCLUSION: Synthesis of new galanthamine derivatives comprising peptide moiety and nicotinic acid or isonicotinic acid is reported. Acute toxicity studies reveal they are about 100 times less toxic than galanthamine. This effect is due to the peptide fragment. Cytotoxicity studies show good correlation with low toxicity results. These results are encouraging for the application of this class compounds as medicines.

Norgalanthamine Stimulates Proliferation of Dermal Papilla Cells via Anagen-Activating Signaling Pathways.

Norgalanthamine has been shown to possess hair-growth promoting effects, including increase in hair-fiber length in cultured rat vibrissa follicles and increase in dermal papilla cell (DPC) proliferation. However, the intracellular mechanisms that underlie the action of norgalanthamine in DPCs have not been investigated. In this study, we addressed the ability of norgalanthamine to trigger anagen-activating signaling pathways in DPCs. Norgalanthamine significantly increased extracellular signal-regulated kinase (ERK) 1/2 phosphorylation at 0.1 µM, a concentration at which DPC proliferation was also induced. Furthermore, the increases in norgalanthamine-induced ERK 1/2 activation and subsequent DPC proliferation were suppressed by the mitogen-activated protein kinase/ERK kinase (MEK) 1/2 inhibitor, U0126. A 0.1 µM dose of norgalanthamine also increased phosphorylation of AKT, which was followed by an increase in glycogen synthase kinase 3ß phosphorylation and nuclear translocation of ß-catenin. In addition, LY294002, a phosphatidylinositol 3 kinase (PI3K) inhibitor, blocked the effect of norgalanthamine on DPC proliferation. These results suggest that norgalanthamine can stimulate the anagen phase of the hair cycle in DPCs via activation of the ERK 1/2, PI3K/AKT, and Wnt/ß-catenin pathways.

Docking-based Design of Galantamine Derivatives with Dual-site Binding to Acetylcholinesterase.

The enzyme acetylcholinesterase is a key target in the treatment of Alzheimer's disease because of its ability to hydrolyze acetylcholine via the catalytic binding site and to accelerate the aggregation of amyloid-ß peptide via the peripheral anionic site (PAS). Using docking-based predictions, in the present study we design 20 novel galantamine derivatives with alkylamide spacers of different length ending with aromatic fragments. The galantamine moiety blocks the catalytic site, while the terminal aromatic fragments bind in PAS. The best predicted compounds are synthesized and tested for acetylcholinesterase inhibitory activity. The experimental results confirm the predictions and show that the heptylamide spacer is of optimal length to bridge the galantamine moiety bound in the catalytic site and the aromatic fragments interacting with PAS. Among the tested terminal aromatic fragments, the phenethyl substituent is the most suitable for binding in PAS.

The Chemistry of Galanthamine. Classical Synthetic Methods and Comprehensive Study on its Analogues.

Galanthamine as an Amaryllidaceae alkaloid has an important role in the treatment of Alzheimer's disease. Some efforts were made to elaborate the total synthesis, and hundreds of its derivatives were prepared to find a more effective molecule with advantageous properties. Moreover, almost every part of the tetracycle was changed; in members of the rings, in the nature and position of the heteroatoms, and ring-opened analogues were also synthesized. In this review the basic synthetic works and the most important derivatives and analogues are overviewed.

Galantamine derivatives with indole moiety: Docking, design, synthesis and acetylcholinesterase inhibitory activity.

The inhibitors of acetylcholinesterase are the main therapy against Alzheimer's disease. Among them, galantamine is the best tolerated and the most prescribed drug. In the present study, 41 galantamine derivatives with known acetylcholinesterase inhibitory activities expressed as IC50 were selected from the literature and docked into a recombinant human acetylcholinesterase by GOLD. A linear relationship between GoldScores and pIC50 values was found and used to design and predict novel galantamine derivatives with indole moiety in the side chain. The four best predicted compounds were synthesized and tested for inhibitory activity. All of them were between 11 and 95 times more active than galantamine. The novel galantamine derivatives with indole moiety have dual site binding to the enzyme--the galantamine moiety binds to the catalytic anionic site and the indole moiety binds to peripheral anionic site. Additionally, the indole moiety of one of the novel inhibitors binds in a region, close to the peripheral anionic site of the enzyme, where the Ω-loop of amyloid beta peptide adheres to acetylcholinesterase. This compound emerges as a promising lead compound for multi-target anti-Alzheimer therapy not only because of the strong inhibitory activity, but also because it is able to block the amyloid beta deposition on acetylcholinesterase.

Nasal Application of the Galantamine Pro-drug Memogain Slows Down Plaque Deposition and Ameliorates Behavior in 5X Familial Alzheimer's Disease Mice.

The plant alkaloid galantamine is an established symptomatic drug treatment for Alzheimer's disease (AD), providing cognitive and global relief in human patients. However, as an acetylcholinesterase inhibitor, gastrointestinal side effects limit the dosage and duration of treatment. Memogain (Gln-1062), a pro-drug, liberates galantamine on cleavage by a carboxyesterase in the brain. The possibility to deliver Memogain intranasally may further circumvent side effects, allowing higher dosing compared to galantamine. In this study, the 5X Familial Alzheimer's Disease (5XFAD) mouse model was used to investigate the effect of chronic Memogain treatment on behavior and amyloid-ß (Aß) plaque deposition in the brain. Chronic intranasal dosage of 6 mg/kg body weight twice daily was tolerated well, whereas the double dose caused body weight loss in males and was less effective in some behavioral tests. 8 weeks of chronic treatment resulted in improved performance in behavioral tests, such as open field and light-dark avoidance, and in fear conditioning already at mildly affected stages at the age of 18 weeks compared to untreated controls. Furthermore, after treatment a significantly lower plaque density in the brain, i.e., in the entorhinal cortex (reduction 20% females, 40% males) and the hippocampus (19% females, 31% males) at the age of 18 weeks was observed. These results show that nasal application of Memogain effectively delivers the drug to the brain with the potential to retard plaque deposition and improve behavioral symptoms in AD similar to the approved galantamine.

Molecular Docking Study on Galantamine Derivatives as Cholinesterase Inhibitors.

A training set of 22 synthetic galantamine derivatives binding to acetylcholinesterase was docked by GOLD and the protocol was optimized in terms of scoring function, rigidity/flexibility of the binding site, presence/absence of a water molecule inside and radius of the binding site. A moderate correlation was found between the affinities of compounds expressed as pIC50 values and their docking scores. The optimized docking protocol was validated by an external test set of 11 natural galantamine derivatives and the correlation coefficient between the docking scores and the pIC50 values was 0.800. The derived relationship was used to analyze the interactions between galantamine derivatives and AChE.

Synthesis of a D-ring isomer of galanthamine via a radical-based Smiles rearrangement reaction.

The 1,9-ethanoiminomethano-bridged tetrahydrodibenzo[b,d]-furan 2, a non-natural isomer of the alkaloid (-)-galanthamine (1) varying in the manner in which the D-ring is annulated to the ABC-core, has been prepared in racemic form. The synthetic sequence starts with the cyclopropane 3 and involves intramolecular Heck alkenylation and radical-based Smiles rearrangement reactions as key steps. Unlike natural product 1, but as predicted by docking studies, compound 2 is not a potent inhibitor of acetylcholine esterase.

Antioxidant activity of galantamine and some of its derivatives.

Oxidative stress is implicated in the pathogenesis of different human diseases: Alzheimer, Parkinson, Huntington, amyotrophic lateral sclerosis (Lou Gehrig's disease), Down's syndrome, atherosclerosis, vascular disease, cancer, diabetes mellitus type 1 and type 2, age - related macular degeneration, psoriatic arthritis. The aim of current study is to summarize the scientific evidences for the antioxidant and neuroprotective activity of Galantamine and some of its derivatives. Galantamine is a scavenger of reactive oxygen species and causes neuroprotective effect by lowering the oxidative neuronal damage, through the following pathways: 1) prevention of the activation of P2X7 receptors; 2) protection of mitochondrial membrane potential; 3) pre - vention of the membrane fluidity disturbances. Another mechanism is the decreasing of the overproduction of reactive oxygen species, a result from the increasing of acetylcholine level due to: 1) acethylcholinesterase inhibition; 2) allosteric potentiation of α7 - subtype of nicotinic acetylcholine receptors. A close relationship between acethylcholinesterase inhibition and reduced oxidative injury is observed. Through allosteric potentiation of the α7 - subtype of nicotinic acetylcholine receptors, the drug leads to induction of phosphorylation of serine - threonine protein kinase, stimulates phosphoinositide 3 - kinase and elevates the expression of protective protein Bcl - 2. By activation of these important neuroprotective cascades, Galantamine exerts neuroprotection against a variety of cytotoxic agents (ß- amyloid peptide, glutamate, hydrogen peroxide, oxygen and glucose deprivation). The new trend in therapy of Alzheimer's disease will be the investigation and application of compounds such as Galantamine derivatives, which possess acethylcholinesterase and γ- secretase inhibitory activity and antioxidant properties.

Acetylcholineestarase-inhibiting alkaloids from Lycoris radiata delay paralysis of amyloid beta-expressing transgenic C. elegans CL4176.

The limited symptom relief and side effects of current Alzheimer's disease (AD) medications warrant urgent discovery and study of new anti-AD agents. The "cholinergic hypothesis" of AD prompts us to search for plant-derived acetylcholineesterase (AChE) inhibitors such as galanthamine that has been licensed in Europe for AD treatment. We used the unique amyloid ß-expressing transgenic C. elegans CL4176, which exhibits paralysis when human Aß1-42 is induced, to study two natural benzylphenethylamine alkaloids isolated from Lycoris radiata (L' Her.) Herb, galanthamine and haemanthidine, and their synthetic derivatives 1,2-Di-O-acetyllycorine and 1-O-acetyllycorine for their anti-paralysis effects. Our data indicate that these Lycoris compounds effectively delay the paralysis of CL4176 worms upon temperature up-shift, and prolong the lives of these transgenic worms. Lycoris compounds were shown to significantly inhibit the gene expression of ace-1 and ace-2. Additionally, the Lycoris compounds may modulate inflammatory and stress-related gene expressions to combat the Aß-toxicity in C. elegans.

Galantamine-based hybrid molecules with acetylcholinesterase, butyrylcholinesterase and γ-secretase inhibition activities.

We previously designed novel peptides-containing galantamine analogues. These compounds we analyzed for their putative inhibitory effect towards acetylcholinesterase, butyrylcholinesterase and γ-secretase, three activities of which could be central to various neurodegenerative pathologies including Alzheimer's disease. These pharmacological agents were virtually equipotent on acetylcholinesterase activity but display drastically higher inhibitory activities towards butyrylcholinesterase with several compounds displaying an about 100-fold higher activity than that harboured by galantamine. Strikingly, two of the galantamine amides that displayed low activity towards acetylcholinesterase exhibited the highest inhibitory potency towards butyrylcholinesterase (106 to 133 times more active than galantamine). Interestingly, five compounds show a rather good γ-secretase inhibitory potency while they retain their ability to inhibit AChE and/or BuChE activity. Thus, we have been able to design novel compounds with significant inhibitory activity against several of the enzymes responsible for key dysfunctions taking place in several neurodegenerative diseases. These mixed inhibitors could therefore be envisioned as potential pharmacological tools aimed at circumventing the degenerative processes taking place in these major pathologies.

Alkaloids from Hippeastrum papilio.

Galanthamine, an acetylcholinesterase inhibitor marketed as a hydrobromide salt (Razadyne®, Reminyl®) for the treatment of Alzheimer's disease (AD), is obtained from Amaryllidaceae plants, especially those belonging to the genera Leucojum, Narcissus, Lycoris and Ungernia. The growing demand for galanthamine has prompted searches for new sources of this compound, as well as other bioactive alkaloids for the treatment of AD. In this paper we report the isolation of the new alkaloid 11ß-hydroxygalanthamine, an epimer of the previously isolated alkaloid habranthine, which was identified using NMR techniques. It has been shown that 11ß-hydroxygalanthamine has an important in vitro acetylcholinesterase inhibitory activity. Additionally, Hippeastrum papilio yielded substantial quantities of galanthamine.

Stability-indicating study of the anti-Alzheimer's drug galantamine hydrobromide.

Galantamine hydrobromide was subjected to different stress conditions (acidic, alkaline, thermal, photolytic and oxidative). Degradation was found to occur under acidic, photolytic and oxidative conditions, while the drug was stable under alkaline and elevated temperature conditions. A stability-indicating reversed-phase liquid chromatographic method was developed for the determination of the drug in the presence of its degradation products. The method was validated for linearity, precision, accuracy, specificity, selectivity and intermediate precision. Additionally, the degradation kinetics of the drug was assessed in relevant cases. The kinetics followed a first order behavior in the case of acidic and photolytic degradation, while a two-phase kinetics behavior was found for the oxidative degradation. The degradation products were characterized by mass spectrometry and nuclear magnetic resonance spectroscopy. Dehydration, epimerization and N-oxidation were the main processes observed during the degradation of galantamine. Moreover, if sufficient material could be isolated the inhibitory activity against the target enzyme acetylcholinesterase was also assessed.

Memogain is a galantamine pro-drug having dramatically reduced adverse effects and enhanced efficacy.

Memogain (Gln-1062) is an inactive pro-drug of galantamine, the latter being a plant alkaloid approved for the treatment of mild to moderate Alzheimer's disease. Memogain has more than 15-fold higher bioavailability in the brain than the same doses of galantamine. In the brain, Memogain is enzymatically cleaved to galantamine, thereby regaining its pharmacological activity as a cholinergic enhancer. In animal models of drug-induced amnesia, Memogain produced several fold larger cognitive improvement than the same doses of galantamine, without exhibiting any significant levels of gastrointestinal side effects that are typical for the unmodified drug and other inhibitors of cholinesterases, such as donepezil and rivastigmin. In the ferret, dramatically reduced emetic and behavioral responses were observed when Memogain was administered instead of galantamine. Based on these and other preclinical data, Memogain may represent an advantageous drug treatment for Alzheimer's disease, combining much lesser gastrointestinal side effects and considerably higher potency in enhancing cognition, as compared to presently available drugs.

Promotion effect of norgalanthamine, a component of Crinum asiaticum, on hair growth.

This study was conducted to evaluate the effect of Crinum asiaticum, a plant native to Jeju Island, Korea, on the promotion of hair growth. When rat vibrissa follicles were treated with a 95% ethanol (EtOH) extract of C. asiaticum, the hair-fiber lengths of the vibrissa follicles increased significantly. In addition, after daily topical application of the EtOH extract of C. asiaticum onto the back of C57BL/6 mice, anagen progression of the hair shaft was induced. Moreover, the extract increased the proliferation of immortalized vibrissa dermal papilla cells. When the vibrissa follicles in the anagen phase were treated with the extract, immunohistochemical analysis revealed that the extract was found to increase the expression of proliferating cell nuclear antigen (PCNA) in the bulb region of the 7-day cultured follicles. In particular, norgalanthamine, a principal of the extract, showed activity that increased the hair-fiber lengths of vibrissa follicles and the proliferation of dermal papilla cells. These results suggest that norgalanthamine, a principal of C. asiaticum, has the potential to promote hair growth via the proliferation of dermal papilla.

Synthesis, spectral characterization and biological evaluation of phosphorylated derivatives of galanthamine.

A series of novel phosphorylated derivatives of galanthamine 6-11 and 12-17 were synthesized in two step process with high yields. In the first step galanthamine 1 was reacted with bis (2-chloroethyl) phosphoramidic dichloride 2/4-nitrophenyl phosphorodichloridate 3 in presence of triethylamine (TEA) in dry tetrahydrofuran (THF) yielded the intermediates 4/5. They were further reacted with various compounds like 2-aminoethanol, ethyleneglycol, ethylenediamine, 2-aminoethanethiol, 2-hydroxyethanethiol, monopotassium dihydrogenphosphate to obtain the title compounds 6-11 and 12-17. The title compounds showed promising antimicrobial, antioxidant activities and was greatly influenced by the presence of different bioactive groups.

Synthesis and characterization of new galanthamine derivatives comprising peptide moiety.

New analogues of galanthamine containing peptide fragments either at 6 or 11 position, were synthesized by reaction between galanthamine molecule and dipeptides and tripeptide, derivatives of N-(3,4-dichlorophenyl)-D,L-Ala-OH. The best results according to yields, easily purification of the target products, and simplicity of the scheme realisation was achieved by using of cyanomethyl ester of Boc-Gly-OH as activated compound.

N-Alkylated galanthamine derivatives: Potent acetylcholinesterase inhibitors from Leucojum aestivum.

N-(14-Methylallyl)norgalanthamine, a new natural compound, together with five known alkaloids: N-allylnorgalanthamine, galanthamine, epinorgalanthamine, narwedine, and lycorine were isolated from mother liquors (waste material) obtained after industrial production of galanthamine hydrobromide from Leucojum aestivum leaves. The structures of N-allylnorgalanthamine and N-(14-methylallyl)norgalanthamine were completely determined by (1)H and (13)C NMR spectroscopy and two-dimensional experiments. N-allylnorgalanthamine (IC(50)=0.18microM) and N-(14-methylallyl)norgalanthamine (IC(50)=0.16microM) inhibit AChE considerably more than the approved drug galanthamine (IC(50)=1.82microM).

Recent developments in cholinesterases inhibitors for Alzheimer's disease treatment.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder of the central nervous system (CNS) which is the most common cause of dementia in the elderly. It is characterized by the deficits in the cholinergic system and presence of characteristic hallmarks: neurofibrillary tangles and amyloid plaques. Since the cholinergic system plays an important role in the regulation of learning and memory processes it became a target for the design of anti-alzheimer drugs. Cholinesterase inhibitors enhance cholinergic transmission indirectly, by inhibiting the enzyme which hydrolyses acetylcholine. It has been also demonstrated that acetylcholinesterase (AChE) is involved in the development of amyloid plaques. Therefore, substances which are AChE inhibitors (AChEI) are the only drugs approved for the symptomatic treatment of AD. This review presents the main classes of cholinesterase inhibitors developed recently for the treatment of AD. We have started with the analogues of the existing drugs: tacrine, donepezil, rivastigmine and galantamine which are still of interest for many research groups. Among them there is a very interesting group--dual binding site inhibitors characterized by increased inhibitory potency against AChE and amyloid plaques formation. There is also a group of compounds with additional properties such as: antioxidant activity, affinity to 5-HT(3) receptors, inhibition of N-methyltransferase that metabolize histamine, which can be beneficial for the treatment of AD. Furthermore there are some interesting compounds which belong to different chemical groups also of natural origin. In this review we sum up current research concerned with development of AChEIs which can be more effective in the future treatment of AD.