International Journal of Scientific & Engineering Research Volume 2, Issue 5, May-2011 1

ISSN 2229-5518

Use of Natural Compounds from Plant Sources as AchE Inhibitors for the Treatment of Early Stage

Alzheimer’s disease-An Insilico Approach

Amrendar Kumar, Abhilasha Singh, Biplab Bhattacharjee

Abstract— Traditionally, drugs were discovered by testing compounds manufactured in time consuming multi-step processes against a battery of in vivo biological screens. Promising compounds were then further studied in development, where their pharmacokinetic properties, metabolism and potential toxicity were investigated. Here we present a study on herbal lead compounds and their potential binding affinity to the effectors molecules of major disease like Alzheimer’s disease. Clinical studies demonstrate a positive correlation between the extent of Acetyl cholinesterase enzyme and Alzheimer’s disease. Therefore, identification of effective, well-tolerated acetyl cholinesterase represents a rational chemo preventive strategy. This study has investigated the effects of naturally occurring nonprote- in compounds polygala and Jatrorrhizine that inhibits acetylcholinesterase enzyme. The results reveal that these compounds use less energy to bind to acetylcholinesterase enzyme and inhibit its activity. Their high ligand binding affinity to acetylcholinesterase enzyme introduce the prospect for their use in chemopreventive applications in addition they are freely available natural compounds that can be safely used to prevent Alzheimer’s Disease.

Index Terms— Alzheiemr’s Disease, Acetylcholinesterase, Binding Affinity, Jatrorrhizine, Clinical Studies, Docking, Rational, Toxicity

1 INTRODUCTION

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lzheimer's disease, most common form of de- mentia is incurable, degenerative, and terminal disease mostly diagnosed in people over
65 years of age. The disease advances with symptoms include confusion, irritability and aggression, mood swings, language breakdown, long-term memory loss, and the general withdrawal of the sufferer as their senses decline. Gradually, bodily functions are lost, ultimately leading to death. In advanced stages of the disease, all memory and mental functioning may be lost. The condition predominantly affects the cerebral cortex and hippocampus, which lose mass and shrink (atrophy) as the disease advances. These changes, occurring in the association area of the cerebral cortex, the hippocampus and the middle and temporal lobes, are accompanied by decreased concentrations of the neurotransmitter acetylcho- line.Acetylcholinesterase is also known as AChE. An

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• Author name is Amrendar kumar currently pursuing Bachelors degree program in Biotechnology, Amity University, Indiay, PH-9453167284. E- mail: amrendar2290@gmail.com

• Co-Author name is Abhilasha singh currently pursuing Bachelorss degree

program in Bioinformatics, Amity University, India, PH-9532395556. E- mail: abhilasha7489@gmail.com
acetyl cholinesterase inhibitor or anti-cholinesterase is a chemical that inhibits the cholinesterase enzyme from breaking down acetylcholine, increasing both the level and duration of action of the neurotransmit- ter acetylcholine.

2 METHODOLOGY

Some small molecules were taken as targeting agent which are responsible for inhibiting biological process in AD. The investigation drug Galantamine was used as a reference drug in the studies.
2W9I was taken as targeting protein and structure for the same was taken from PDB.
Then Initial screening is done by Lipinski’s rule of 5. The accepted compounds that were showing better interaction with the target protein and their energy were minimized using Marvin’s Sketch.
Then the selected conformations are saved in three formats:
1. SDF Format
2. PDB Format
3. MOL. Format
Then docking is done between the protein and small
molecules with QUANTUM. The best three results

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obtained were analysed under HEX and ARGUS. IC50 value is taken by QUANTUM.Then graphs are plotted by seeing the values.
After this, ADME TOX analysis was performed. In this analysis ADME features were showing interac- tion
With 2W9I in both Argus and Quantum analysis were predicted under ADME test for toxicity predic- tion.

TABLE 1

Natural compounds and Their Quantum results

TABLE 2

Drugs and Their Quantum results

AMES test is considering for initial screening of the molecule based on their ability to induce mutation. AMES test is used for determining if a chemical is mutagen. The molecules which showing ability to induce mutation were rejected in toxicity based screening.
Using ADME TOX analysis it was found that Ja- trorrhizine was showing lower AMES test values than reference molecules. Further health effects of these molecules in blood, cardiovascular system, ga- strointestinal system, kidney, liver and lungs were
predicted. LD 50 values also were predicted for se- lecting reliable molecule for ADME analysis.
After ADME analysis,a graph was plotted on ADME- TOX values.
After all analysis, one compound Jatrorrhizine was the best molecule.
This molecule is considered as better ligands for
2W9I based on interaction, Pharmacokinetics, and
pharmacodynamic features and can be used for
Chemotherapeutic use.

3 BIOCHEMISTRY OF ALZHEIMER’S

The biochemistry of Alzheimer's disease (AD), one of the most common causes of adult dementia, is as yet not well understood. It has been identified as a pro- tein misfolding disease due to the accumulation of abnormally folded amyloid betaprotein in the brains of AD patients. Amyloid beta, also written A�, is a short peptide that is an abnormal proteolyticbypro- duct of the transmembrane protein amyloid precur- sor protein (APP), whose function is unclear but thought to be involved in neuronal development. The presenilins are components of proteolytic com- plex involved in APP processing and degradation. Amyloid beta monomers are soluble and contain short regions of beta sheet and polyproline II helix secondary structures in solution, though they are largely alpha helical in membranes; however, at suf- ficiently high concentration, they undergo a dramatic conformational change to form a beta sheet-rich ter- tiary structure that aggregates to form amyloid fi- brils. These fibrils deposit outside neurons in dense formations known as senile plaques or neuritic pla- ques, in less dense aggregates as diffuse plaques, and sometimes in the walls of small blood vessels in the brain in a process called amyloid angiopathy orcon- gophilic angiopathy.
AD is also considered a tauopathy due to abnormal aggregation of the tau protein, a microtubule- associated proteinexpressed in neurons that normal- ly acts to stabilize microtubules in the cell cytoskele- ton. Like most microtubule-associated proteins, tau is normally regulated by phosphorylation; however, in AD patients, hyperphosphorylated tau accumulates as paired helical filaments that in turn aggregate into masses inside nerve cell bodies known as neurofibril- lary tangles and as dystrophic neurites associated with amyloid plaques. Although little is known about the process of filament assembly, it has recent- ly been shown that a depletion of a prolyl isomerase protein in the parvulin family accelerates the accu- mulation of abnormal tau.

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4 DISEASE MECHANISM

Although the gross histological features of AD in the brain are well characterized, three major hypotheses have been advanced regarding the primary cause. The oldest hypothesis suggests that deficiency in cholinergic signaling initiates the progression of the disease. Two alternative misfolding hypotheses instead suggest that either tau protein or amyloid beta initiates the cascade. While researchers have not identified a clear causative pathway originating from any of the three molecular hypotheses to explain the gross anatomical changes observed in advanced AD, variants of the amyloid beta hypothesis of molecular initiation have become dominant among the three possibilities.

4.1 THE MECHANISM OF ACTION OF

ACETYLCHOLINESTARASE

Cholinergic nerve transmission is terminated by the enzyme acetylcholinesterase (AchE). AchE is found both on the post-synaptic membrane of cholinergic synapses and in other tissues eg red blood cells. Ace- tylcholine (Ach) binds to AchE and is hydrolysed to acetate and choline. This inactivates the Ach and the nerve impulse is halted. AchE inhibitors (eg rivas- tigmine) prevent the hydrolysis of Ach, which in- creases the concentration of Ach in the synaptic cleft; AchE inhibitors are widely used in the treatment of Alzheimer’s disease.

5 RESULTS AND ANALYSIS

Alzheimer's disease, the most common form of de- mentia, is a progressive disorder characterized by widespread loss of brain cells called neurons, beta- amyloid deposits in the cerebral blood vessels, de- velopment of plaques and the presence of neurofi- brillary tangles. Alzheimer's disease (AD) is an irre- versible, progressive disorder in which brain cells(neurons) deteriorate, resulting in the loss of cognitive functions, primarily memory, judgment and reasoning, movement coordination, and pattern recognition. There is a very high genetic cause for Alzheimer's disease. In addition to genetics as a cause of Alzheimer's disease, many environmental factors including diet are to be considered. Older adults who smoke have an elevated risk of develop- ing Alzheimer's disease. Nerve signals travel across synapses with the help of chemicals known as "neu- rotransmitters," including one called acetylcholine. Nerve cell destruction causes a reduction in acetyl- choline, leading to impaired transmission of nerve signals and poor communication between nerve cells called neurons. In addition to acetylcholine, the brain of Alzheimer’s disease patients have areas of abnor- mal protein called "plaques" and "tangles," the names
reflecting what these abnormalities in the brain look like under the microscope. The underlying cause of Alzheimer's – what actually triggers the changes in the brain – is still not fully known but could partly be due oxidation and damage to nerve cells over time. It is likely that no single factor is responsible, but rather that it is due to a variety of factors, which may differ from person to person.
People whose parents or brothers and sisters develop the disease appear to be at greater risk of developing it themselves, so there may be a genetic component. However, no straightforward pattern of inheritance has been found. It is known that head injury is a risk factor, and also that Alzheimer’s disease often affects people with Down’s syndrome. Some researchers have suggested that people who exercise their brains (for example, doing crosswords and other mental agility exercises) are less likely to develop the dis- ease. And Omega 3 fatty acids, contained in oily fish such as mackerel and salmon may, also help to pre- vent dementia. Australian scientists say they have identified a toxin that may play a key role as a poten- tial cause of Alzheimer's disease. The toxin, called quinolinic acid, kills nerve cells in the brain, leading to dysfunction and death. Quinolinic acid may not be the main cause of Alzheimer's disease, but it plays a key role in its progression. Researchers have shown that a common anesthetic gas can cause fragments of a normal brain protein called amyloid-beta to clump together, which is thought to be the main problem underlying Alzheimer's disease. Intravenous anes- thetics have less of an effect, the team reports in the journal Biochemistry.
Anesthetics used in long surgery, such as inhaled anesthetics isoflurane and halothane, may be another cause of Alzheimer's disease. Scientists conducted a series of lab experiments using nuclear magnetic re- sonance to investigate the reaction of amyloid-beta peptides to the inhaled anesthetic isoflurane and the intravenous anesthetics propofol and thiopental. They found that the peptides aggregated together after 10 to 30 hours' exposure to isoflurane, depend- ing on the concentration of the gas and the size of the protein fragments. The effect was seen with propofol after exposure for 48 hours, but no clumping was seen with thiopental. Biochemistry, January 23, 2007. Having frequent colds or flu or other infections in- creases the risk for this neurological deterioration due to increased overall inflammation in the body and brain.
There are certain natural and synthetic AchE inhibi- tors which will prevent the cause of AD by blocking the Biochemical pathway .Some number of natural compounds which are Inhibiting Ache were taken.

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Docking of these molecules was performed under
QUANTUM 3.3.0.
It was found that the natural compound Polygala,
Sterigmatocystin and Jatrorrhizine were showing
reliable pharmacokinetics and pharmacodynamics
features than the commercial drugs. Hence they were
taken out for work.
After analysing the graphs, it was found that Bulbo-
capnie was the best Acetylcholinesterase Inhibitor.
This molecule is considered as better ligands for Ace-
tylcholinesterase based on ligand-receptor interac-
tion,

Jatrorrhizine is a protoberberine alkaloid isolated

from Enantia chlorantha (Annonaceae) and other spe-
cies. Synonyms that may be encountered include
jateorrhizine, neprotin, jatrochizine, jatrorhizine, or
yatrorizine. It has been reported to have anti inflam-
matory effect, and to improve blood flow and mitotic
activity in thioacetamide-traumatized rat livers. It was found to have antimicrobial and antifungal ac- tivity. It binds and noncompetitively inhibits mo- noamine oxidase (IC50 4 micromolar for MAO-A and
62 for MAO-B).It interferes with multidrug resistance by cancer cells in vitro when exposed to a chemothe- rapeutic agent. Large doses (50-100 mg/kg) reduced blood sugar levels in mice by increasing aerobic gly- colysis.
Fig. 1: Graph showing QUANTUM Results of natural compounds and commercial drugs. Greens are for natural compounds and red is for commercial drugs.


Fig.2: Graph showing Hex results of best two natural compounds and best two commercial drugs based on quantum. Greens are for natural compounds and red is for commercial drugs.

6 CONCLUSION

Alzheimer's disease, the most common form of de- mentia, is a progressive disorder characterized by widespread loss of brain cells called neurons, beta- amyloid deposits in the cerebral blood vessels, de- velopment of plaques and the presence of neurofi- brillary tangles. Alzheimer's disease (AD) is an irre- versible, progressive disorder in which brain cells (neurons) deteriorate, resulting in the loss of cogni- tive functions, primarily memory, judgment and rea- soning, movement coordination, and pattern recogni- tion.There are certain natural and synthetic AchE inhibitors which will prevent the cause of AD by blocking the Biochemical pathway .Some number of natural compounds which are Inhibiting Ache were taken.Docking of these molecules was performed under QUANTUM. It was found that the natural compound POLYGALA, STERIGMATOCYSTIN AND JATRORRHIZINE was showing reliable phar- macokinetics and pharmacodynamics features than the commercial drugs. Hence they were taken out for work.

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Further, it was found that among three natural com- pounds JATRORRHIZINE is the best natural com- pound and better than commercial drugs and hence can be taken as AD chemopreventive agent and it can be effective inhibitors for AchE in AD pathway.

ACKNOWLEDGMENT

The Author wishes to thanks Mr. Ajay Singh and Biplab Bhattacharjee (sponsor and financial support). Researchers that contributed information or assis- tance to the article are Mayank Agarwal, Ajit Kumar pandey.

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