drug design paper

8/10/2019 Drug Design Paper

1/39

1

Anti-diabetic activities of bioactive compounds

in Euphorbia Hirta Linn to receptors for

diabetes type 2 drug development

Authors: Tram Nguyen, Nghi Nguyen, Luan Vu, Thanh Nguyen, Huong Nguyen

School of Biotechnology, International UniversityVietnam National University in HCMC

Abstract

Objectives: Euphorbia hirta has been lately studied as a potential therapeutic herbal used in

Diabetes type 2 patients. In this research, we examine the antidiabetic activities of bioactive

compounds in Euphorbia hirta Linn to five receptors (11HSD1, PTP1B, -glucosidase,

PPARy, DDP4) to establish treatment for diabetes type 2.

Methods: Firstly, the 3D structures of 29 major bioactive compounds of Euphorbia hirta

were sketched by Gaussview. Secondly, Autodock Tools was applied to build a complete set

of ligands and receptors. Subsequently, a ligand based pharmacophore approach has been

generated for those 29antidiabetic compounds with significance for the development of new

drugs by using LigandScout software.

Result and conclusion:Three pharmacophore features: hydrophobic domain, hydrogen bond

acceptor and hydrogen bond donor were obtained. Also, the top nine affinity binding

compounds (1,3,4-tetra-O-galloyl--D-glucose; -amyrine; campesterol; myricitrin,

quercitrin, pelargonium-3,5-diglucoside; friedelin; taraxerone, taraxerol) give strong

evidence to be good candidates for drug development to diabetes type 2 treatment. Besides,

the pharmacophore models applied the Lipinskis rule to determine if a chemical compound

with a certain pharmacological or biological activity has similar properties that would make

it a likely orally active drug in humans. Moreover, molecular docking is highly

recommended to use to reach the optimum results.

Key words: Diabetes type 2, Euphorbia Hirta Linn, drug design, docking, pharmacophore

modeling.


2/39

2

Introduction

In normal body, the production of insulin stimulates cells to take up glucose from the

bloodstream to manufacture energy for the whole body metabolism. However, when a body

produces little or no insulin, the blood glucose (blood sugar) will increase uncontrollably and

causes serve damage through whole body (heart, eyes, kidney, nerve, teeth, etc.)i. There are

two types of diabetes: type 1 and type 2. Diabetes type 2 is far more common than diabetes

type 1. About 90% of adults with diabetes have type 2, and about 10% have type 1ii. Type 2

diabetes (non-insulin-dependent diabetes) is called insulin resistance the pancreas does

not make enough insulin or the body cannot use the insulin welliii.

Herbal drugs are studied recently due to their less side effects, low cost and higheffectiveness. Therefore, numerous of researches had been processed to test the bioactivities

of some potential herbal candidates. In diabetic treatment, Euphorbia hirta a member of

Euphorbiaceae and genus Euphorbia has been lately studied as a potential therapeutic herbal

used in Type 2 diabetic patients. E. hirta is commonly found in garden paths and wastelands.

Phytochemical analysis of Euphorbia hirta (E. hirta) revealed the presence of alkaloids,

flavonoids, sterols, tannins and triterpenoids in the whole plantiv. Some chemical

components extracted from E. hirta performed biological acitivities, such as: antimalarial,

anti-diarrhoeic, anti-inflammatory, antimicrobial, antibacterial, diuretic, anti-allergic

activities, etc.v,vi,vii,viii,ix,x,xi,xii,xiii,xiv In this research, we examine the antidiabeticactivities of 29

bioactive compounds (belongs to Flavonoids, polyphenols, tannins, triterpenes and

phytosterols)xvin Euphorbia hirta Linn to five receptors: 11HSD1, PTP1B, -glucosidase,

PPARy, DDP4 (directly related to type 2 diabetes) by molecular docking tools and

pharmacophore analysis to2 to determine which compounds are the best candidates for

potential drug design.xvi


3/39

3

Material and Method

1.

Data set collection and receptor-compound preparation

The most important process in pharmacophore model generation is the selection of test set

compounds. Over the last few years, a number of anti-diabetic compounds have been

identified and the Euphorbia Hirta showed a potential bioactivity in anti-diabetes type

2xvii.Therefore, we collect 28 major bioactive compounds in Euphorbia hirta in this current

research. Table 7 shows the 2D structures of 28 bioactive compounds candidates from the

ncbi and Chemspider. The 3D structures of these 28 compounds were sketched by the

Gaussview 5.0xviii

and save in mol2 format. Subsequently, they were imported to Autodock

and ready for docking.

Receptor

11 beta hydroxysteroid dehydrogenase type 1, PTP1B, Alpha-glucosidase, PPARy,

DDP4 are the proteins relating to Diabetes type 2 in humans.xix

11BETA HYDROXYSTEROID DEHYDROGENASE TYPE 1(11B HSD1)

11 Hydroxysteroid dehydrogenase - a microsomal glycoprotein enzyme - belongs

to SDRs (short-chain dehydrogenase /reductase) protein family. 11HSD1 composes

of 282 amino acid and weights 38kDa. It is expressed predominantly in peripheral

tissues such as liver, adipose tissues, skeletal muscles and central nervous system.The

main function of 11 HSD1 is catalysis of cortisone to cortisol active glucocortisoid

conversion process in human. Glucocorticoids have been shown to inhibit a number

of steps in the insulin signaling network through several different mechanismsxx

. Due

to the fact that cortisol plays a critical role in diabetes, 11HSD1 has a potential

therapeutic target for type II diabetes. Many studies have indicated that the high

circulating levels of the active glucorticoid cortisol can also lead to other syndrome

such as central obesity, dyslipidemia and hypertensionxxi. 11 -HSD 1 has both dimer

and tetramer organization. In this study,3D structure of this protein was taken from


4/39

4

Protein Data Bank as the accession number is 1XU7 (tetramer). The further step

(docking and modeling) is worked on chain A of 11 -HSD 1only.

PROTEIN-TYROSINE PHOSPHATASE 1BTyrosine-protein phosphatase non-receptor type 1also known as protein-tyrosine

phosphatase 1B (PTP1B) is an enzyme that is the founding member of the protein

tyrosine phosphatase(PTP) family. In humans it is encoded by the

PTPN1 gene.xxiiPTP1B is a negative regulator of the insulin signaling pathway and is

considered a promising potential therapeutic target, in particular for treatment of type

2 diabetes.xxiii

PTP1B was the first member of the protein tyrosine phosphatase (PTP) superfamily to

be identified and was purified to homogeneity from human placenta as a catalytic

domain of 37 kDa (Tonks et al., 1988). Later, it was characterized as an 50 kDa

protein (435 amino acids), consisting of an N-terminal catalytic domain followed by a

C-terminal segment that serves a regulatory function and anchors the protein at the

cytoplasmic face of the endoplasmic reticulum (ER) membrane (Tonks, 2003).

PTP1B can dephosphorylate the phosphotyrosine residues of the activated insulin

receptorkinase. The phosphatase activity of PTP1B occurs via a two-step mechanism

(Figure 3). In the first step: pTyr substrate is dephosphorylated (a nucleophilic attack

at the phosphocenter by the reduced Cys215 residue, followed by subsequent

protonation by Asp181 to yield the neutral tyrosinephenol). In the second step: The

enzyme intermediates are broken down. The active enzyme is regenerated after the

thiophosphate intermediate is hydrolyzed, which is facilitated by the hydrogen

bondinginteractions of Gln262 and Asp181 that help to position in the water

molecule at the desired site of nucleophillic attack.xxiv

In this study, 3D structure of this protein was taken from Protein Data Bank as the

accession number is 3CWE. The further step (docking and modeling) is worked on

chain A of 3CWE only.
http://www.sciencedirect.com/science/article/pii/S0092867411010063#bib48http://www.sciencedirect.com/science/article/pii/S0092867411010063#bib48http://www.sciencedirect.com/science/article/pii/S0092867411010063#bib48http://www.sciencedirect.com/science/article/pii/S0092867411010063#bib46http://www.sciencedirect.com/science/article/pii/S0092867411010063#bib46http://www.sciencedirect.com/science/article/pii/S0092867411010063#bib46http://www.sciencedirect.com/science/article/pii/S0092867411010063#bib46http://www.sciencedirect.com/science/article/pii/S0092867411010063#bib48


5/39

5

PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR (PPAR-)

Peroxisome proliferator-activated receptor gamma (PPAR-or PPARG), or glitazone

receptor, or NR1C3 (nuclear receptor subfamily 1, group C, member 3) is a type

IInuclear receptor that in humans is encoded by the PPARGgene. It belongs to

the nuclear hormone receptor family.xxvxxvixxvii

In human and mouse, PPARG has been found in 2 isoforms: PPAR-1 (found in

nearly all tissues except muscle) and PPAR-2 (mostly found in adipose tissue and the

intestine).xxviiiDefects in PPARG can lead to type 2 insulin-resistant diabetes and

hypertension. Nuclear receptor that binds peroxisome proliferators such as

hypolipidemic drugs and fatty acids. Once activated by a ligand, the nuclear receptor

binds to DNA specific PPAR response elements (PPRE) and modulates the

transcription of its target genes. Therefore control the regulation of fatty acid storage

and glucose metabolism. Manyinsulin sensitizing drugs (namely,

thethiazolidinediones) used in the treatment of diabetes target PPARG as a means to

lower serum glucose without increasing pancreatic insulin secretion.xxix

DIPEPTIDYL PEPTIDASE-4(DPP4)

DPPIV adenosine deaminasecomplexing protein 2 or CD26 - belongs to the

exopeptidase class of proteolytic enzymes. This antigenic enzyme expressed on the

surface of most cell types and associated with immune regulation, signal

transduction and apoptosis. Exopeptidases that cleave N- terminal and C- terminal

amino acid residues from peptide and protein. DPP4 plays a major role

in glucose metabolism. It is responsible for the degradation of incretions such as GLP-

1 which stimulates insulin release and inhibits glucagon release to lower of blood

glucosexxx.

In this study, 3D structure of this protein was taken from Protein Data Bank as the

accession number is 4J3J (dimer).

ALPHA-GLUCOSIDASE
http://en.wikipedia.org/wiki/Nuclear_receptorhttp://en.wikipedia.org/wiki/Genehttp://en.wikipedia.org/wiki/Anti-diabetic_drug#Sensitizershttp://en.wikipedia.org/wiki/Thiazolidinedioneshttp://en.wikipedia.org/wiki/Diabeteshttp://en.wikipedia.org/wiki/Diabeteshttp://en.wikipedia.org/wiki/Thiazolidinedioneshttp://en.wikipedia.org/wiki/Anti-diabetic_drug#Sensitizershttp://en.wikipedia.org/wiki/Genehttp://en.wikipedia.org/wiki/Nuclear_receptor


6/39

6

Alpha-glucosidase is a glucosidase that acts upon 1,4-alpha-glucosidase bonds

(Figure 5).xxxi

They are in contrast to beta-glucosidase. The main function of alpha-

glucosidase is breaking down starch and disaccharides to glucose. Maltase belongs to

this family.

Figure 5: The effect of alpha-glucosidase on 1,4-alpha bonds

Maltase glucoamylase, the proteinconsist of 875 amino acids, is one of the four

intestinal glycoside hydrolase 31 enzyme activities which respond for the hydrolysis

of terminal starch products into glucose. As the result, an inhibition of the N-terminal

catalytic domain of maltase-glucoamylase (ntMGAM) is necessary for controlling

blood glucose levels in individuals with type 2 diabetesxxxii

. In this research, 3L4T (a

monomer) was taken from Protein Data Bank (PDB) like the representative ofmaltase

glucoamylase.

Bioactive compound in E.hirta

Bioactive compounds in E.hirta is classified into three main families including tannin,

flavonoid and terpenesxxxiii. Tannin and flavonoid are strong antioxidant whose products

have been shown as a key in pathogenesis of diabetes type 1 and 2. Therefore

antioxidants such as tannin and flavonoid are considered to have potential ability for

therapeutic drugs for diabetes treatmnent,. The current study will investigate 28 ofbioactive compounds from all three families to determine how they interact with target

proteins in diabetes type 2.In this research, we will focus on 3 families that mainly

contribute in E. hirta: Flavonoid family (Afzelin, quercetin, quercitrin, quercitol,

rhamnose, rutin, leucocyanidin, myricitrin, cyanidin 3,5- O-diglucoside, kaemferon,


7/39

7

pelargonium 3,5- diglucoside). Tannin family (gallic acid,3,4-di-O-galloyl-quinic acid,

3,4,5-tri-O-galloylquinic acid, 1,2,3,4,6 tetraOgalloyl-b-D-glucose). Triterpenes and

phytosterols family (2,4-methylenecycloartenol, betasitosterol, campesterol, -

stigmasterol, ingenol triacetate, resiniferonol, alpha-amyrine, beta-amyrine, friedelin,taraxerol, taraxerone, cycloartenol, protocatechuic acid).xxxiv xxxv

Many bioactive compounds in E.hirta were found to perform several functions:

antioxidant, anti-inflammatory, antimicrobial, anticancer, cardioprotective,

neuroprotective, antidiabetic, antiosteoporotic, estrogenic/antiestrogenic, anxiolytic,

analgesic,antiallergic activities, etc. (Table 11). However, in this research, we will only

analyze the anti-diabetic activities of those compounds to the target protein receptors

which directly related to diabetes type 2.

Most of the 3D structures of 28 bioactive compounds in E.hirtawere built based on 2D

picture by GaussView 5.0. The 2D and 3D structures of 29 ligands are illustrated in

Table7.

Docking simulation

The docking process was done using Autodock Tools1.5.6 xxxvi

Autodock Tools was applied to build a complete set of ligands and receptors with the file

name pdbqt. Receptor fixation was done by following steps: (1) adding polar hydrogen,

(2) removing water molecule (3) computating of Gasteiger charges and adding charges to

receptors, and (4) locating of Grid box by using Center on ligand (with number of points

in x-dimension/ y-dimension and z-dimension are 40x40x40, spacing is 1 and

exhaustedness is 100). The site of Grid Box is illustrated in Table 9.

The 3D structures of all 29 compounds in pdb or mol2 file type was converted into pdbqt

file type after detect the root to set up the appropriated ligands.


8/39

8

2.

Pharmacophore modeling

LigandScout 3.12xxxvii

will be used to derive the pharmacophore models. LigandScout

software efficiently allows rapidly and transparently generation of 2D and 3D

pharmacophore of data set. It creates the pharmacophore, aligned pharmacophore and

features, aligning of merge pharmacophore of compounds and molecules by reference

points. This tool is scientifically published and based on several years of experience in

pharmacophore generation.xxxviii

In this study, the program was applied to show 3D structure of the receptors, both 2D and

3D structure of ligand in the binding pocket of that receptor with the position, interaction

(types and residues of interaction) as well as ligand properties such as molecular weights,

number of atoms, rings, etc. After identifying pharmacophore of ligands and receptors,

types of bonds were colored and symbol as red arrow, green arrow, red star and orange

bubble are hydrogen bond acceptor, hydrogen bond donor, and negative ionization and

hydrophobic interaction, respectively.

Then in order to evaluate their drugs likeness property, rule of five (Lipinski's rule) was

used, it is a popular rule to evaluate drug like properties or determine if a chemical

compound with a certain pharmacological or biological activity has similar properties

that would make it a likely orally active drug in humansxxxix

The rule is as follows:

There should be less than 5 H-bond donors.

Molecular weight should be less than 500 Daltons.

Partition coefficient (LogP) not over 5 (or MLogP is over 4.15).

There should be less than 10 H-bond acceptors.


9/39

9

Result and discussion

1.

Free energy binding of bioactive compound to receptor related to diabetes

type 2

The results of docking process showed that the absolute value of binding energy ranged

from 4.4 to 11.3 kcal/mol (Table 6). From the observation, there are eight bioactive

compounds shows the strong binding affinity (|binding affinity| >7.0 kcal/mol) to 5

receptors (11HSD1, PTP1B, -glucosidase, PPARy, DDP4). The tannin family had

high binding affinity such as 1,3,4,6 tetraOgalloyl-b-D-glucose. The group of terpenes

including alpha amyrine, beta amyrine, friedelin, taraxerone, campesterol, and

cycloarterol also yield good result. In flavonoid family, three of these members

quercitin, pelargonium 3,5-diglucose and myricitrin were selected for pharmacophore

modeling step. The receptor DDP4 showed highest binding affinities of all bioactive

compounds to the others (20 out of 29 compounds (~70.%) have |binding affinity| >7.0

kcal/mol). Therefore, DDP4 is considered to be the good receptor for diabetes type 2 for

patients that were treated with bioactive compounds in E. hirta.

The absolute value for all of 29 bioactive compounds with five target proteins are

revealed in Graph 1 and Table 10. Top binding affinity includes: Friedelin, alpha-

amyrine, pelargonium-3,5-diglucoside, taraxerone, 1,3,4-tetra-O-galloyl-b-Dglucose with

the absolute binding affinity is higher than 7.5 kcal/mol.


10/39

10

Graph 1:The absolute value for all of 29 bioactive compounds with 5 target proteins

1 = 1,3,4-tetra-O-galloyl-b-Dglucose, 2 = 2,4_methylenecycloartenol, 3 = 3,4 diOgalloyquinic acid, 4=

afzelin, 5 = alpha-amyrin, 6 = beta-amyrin, 7 = beta-sitosterol, 8 = campesterol, 9 = cyaniding-3,5-

diglucoside, 10 = cycloartenol, 11 = friedelin, 12 = gallic acid, 13 = galloylquinic acid, 14 = ingenol

triacetate, 15 = kaempferol, 16 = leucocyanidin, 17 = L-rhamnose, 18 = myricitrin, 19 = tinyatoxin, 20 =

pelargonium-3,5-diglucoside, 21 = protocatechuic acid, 22 = quercetin, 23 = quercitol, 24 = quercitrin, 25 =

resiniferonol, 26 = rutin, 27 = stigmasterol, 28 = taraxerol, 29 = taraxerone.

2. Pharmacophore modeling

11 -HSD1

High binding energy of the ligands to the receptor was explained clearly by interaction

analysis (Table 1). Five molecules (1,3,4-tetra-O-galloyl-b-Dglucose,

24_methylenecycloartenol, alpha amyrin, friedelin, taraxerone) were frequently within

hydrophobic interaction with residues Val 227A, Val 231A, Tyr 177A, Met 179A, Leu

126A and Hydrogen bond donor with residue Tyr 257A. Hydrophobic contacts at

position of methyl group which is non-polar whereas hydrogen bonds contact at steroidalhydroxyl group of the protein. From this observation, six listed residues seemed to play

an essential role in catalytic activity of 11 -HSD 1. Moreover, 1,3,4-tetra-O-galloyl-b-

Dglucose, 24_methylenecycloartenol, alpha amyrin link to the receptor with a high

number of hydrogen bonds and hydrophobic interaction compared with friedelin and

taraxerone. 1,3,4-tetra-O-galloyl-b-Dglucose has all hydrogen bonds at residues Arg

269A, Glu 254A, Asn 270A, Lys 274A, Leu 266A. 24_methylenecycloartenol, alpha

4

6

8

10

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29

11betaHSD1

A-glucosidase

PTP1B

PPARy

DDP4


11/39

11

amyrin, friedelin have all hydrophobic interactions with many identical interactions with

each other. Based on the analysis of ligands and target receptor interaction, the

conclusion of this process is all five selected compounds have good interaction with

receptor. However, 1,3,4-tetra-O-galloyl-b-Dglucose has the molecular weight higher

than 500kDa which will be excluded by L ipinski's rul e of five

xl

.1,3,4-tetra-O-galloyl-b-Dglucose needs to be modified to reduce molecular weight for using as a tempting

compounds for design diabetes type 2 treatment.

PTP1B

The binding affinity of the ligands to the PTP1B receptor is described in the Figure 5.

The top 5 molecules (24_methylenecycloartenol, alpha amyrin, cycloartenol, friedelin,

taraxerone), which show the highest binding affinity to PTP1B receptor (|Binding

affinity| > 7.5 kcal/mol), performed the hydrophobic interaction (yellow bubble) with

residues Thr763A, Tyr520A, Tyr546A, Phe682A, Leu588A, Hydrogen bond donor with

(green arrow) residue Gln762A, Phe862A, Lys541A and Hydrogen bond acceptor (red

arrow) with residue Gln521A. (Table 2.) Those nine named residues worked as emergent

residue in the activity of PTP1B.

Based on the analysis of ligands and target receptor interaction, the conclusion of this

process is all five selected compounds have good interaction with PTP1B receptor.

Therefore, these bioactive compounds are potential candidates for drug development to

diabetes type 2 treatments.

ALPHA-GLUCOSIDASE (MALTASE-GLUCOAMYLASE)The molecular docking (Autodockvina)s result provided that there are 5 molecules:

1,3,4-tetra-O-galloyl-b-Dglucose, Alpha amyrin, Friedelin, Taraxerol, Taraxerone which

have highest affinities with the target protein 3L4T (Table 10). Moreover, hydrophobic is

considered as a main interaction between those molecules with Maltase-Glucoamylase

(3L4T) at PHE450A, TRP406A, THR204A, exclusive of 1,3,4-tetra-O-galloyl-b-

Dglucose (Table 5) when it links with 3L4T at ASP542A, ASP203A,

ARG202A,BJ11001A, TYR605A, GLN603A, TYR299A,ARG334A by hydrogen bond

donor or aceptor. It follows that hydrogen bonds keep molecules tightly bound to proteins

as well as enclosed hydrophobic interactions strengthen the binding of ligands andproteins, therefore hydrogen bonds are important in forming a stable link between

molecules and target protein. In that case, 1,3,4-tetra-O-galloyl-b-Dglucose is best

candidate for drug development among all five selected molecules. However 1,3,4-tetra-

O-galloyl-b-Dglucose weights nearly 795 kDa which violates Lipinski's rule of five. As

consequence, 1,3,4-tetra-O-galloyl-b-Dglucose needs to be remodeled for the purpose of


12/39

12

decreasing molecule weight or be treated as a template for designing a new compounds in

diabetes type 2 treatment.

DPP4

These compounds - 1,3,4-tetra-O-galloyl-b-Dglucose, 24-methylenecycloartenol, beta -amyrin, cycloartenol, taraxeronehave highest binding affinity. Hydrophobic interactions

were showed (in Ligand scout) at TYR547A, TRP627, TRP629A, especially at

TYR547A and hydrogen bond at VAL546A, ASP545A frequently. This analysis proved

that these residues play critical role in DPP4s catalytic activity. These interactions show

compounds ability to interact with receptor DPP4. After comparison (number of

hydrophobic interactions, H bonds), 1,3,4,6-tetra-O-galloyl-b-D-glucose showed strong

interaction to DPP4 receptor. Therefore, its considered to be a potential factor for drug

development.

PPAR

Five compounds that have highest binding affinity are 1,3,4-tetra-O-galloyl-b-Dglucose,

alpha -amyrin, beta - amyrin, cycloartenol, taraxerone are. Hydrophobic interactions were

showed (in Ligand scout) repeatedly at ILE262A, PHE287A . On the other hand,

hydrogen bondsincludeH bond acceptors: GLU259A, ARG280A, SER464A and H bond

donors: LYS275A, OH group, HIS466A. The pharmacoporesshowed that these residues

play critical role in PPARsregulation activity. These interactions show compounds

ability to interact with receptorPPAR. Thus, all the top binding compound showed a

stable and consistent interaction with PPAR. Therefore, its considered to be a potential

factor for drug development. One thing noticed is 1,3,4-tetra-O-galloyl-b-D-glucose

which has hydrogen bonds to keep molecules tightly bound to proteins.


13/39

13

Table 1:Binding modes of selective compounds with 11-HSD1

NAME OF

COMPOUND

IMAGE INTERACTION LIGAND DETAIL

24-

methylene

cycloartenol

Hydrophobic:Val231

A, Leu126A,Met179A, Tyr177A

Formula: C33 H 60O 1Molweight: 472.842

Size & FlexibilityAtoms/Bonds: 94 / 97

Rings: 4Rotatable Bonds: 17

Aromatic Atoms: 0

Polarity & Chemical FeaturescLogP: 9.521TPSA: 20.230

Acceptors: 0Donors: 0

Neg. Ionizable: 0

Pos. Ionizable: 0


14/39

14

Alpha-

amyrine

Hydrophobic:Val231A, Ile230A,

Val227A, Leu126A,

Met179A, Tyr177A.


Size & FlexibilityAtoms/Bonds: 83 / 87Rings: 5

Rotatable Bonds: 9Aromatic Atoms: 0

Polarity & Chemical Features

cLogP: 8.105TPSA: 20.230


Neg. Ionizable: 0

Pos. Ionizable: 0

1,3,4-tetra-

O-galloyl-b-

D-glucose

H bond

donor:Asn270A,

Tyr257A, Leu266A.

H bond acceptor:

Glu254A, Arg269A,

Tyr257A, Lys274A.

Formula: C34 H 36O 22

Molweight: 796.640



Aromatic Atoms: 24

Polarity & Chemical FeaturescLogP: -2.469

TPSA: 390.060Acceptors: 5

Donors: 4

Neg. Ionizable: 0Pos. Ionizable: 0


15/39

15

Friedelin Hydrophobic:Thr265A, Leu262A.

H bond

acceptor:Tyr257A





cLogP: 8.457TPSA: 17.070


Neg. Ionizable: 0

Pos. Ionizable: 0

Taraxerone Hydrophobic:

Tyr177A, Val175A,Leu126A, Val227A,

Val231A

H donor bond:

Ser228A



Rings: 5


Polarity & Chemical FeaturescLogP: 8.249


Donors: 1



16/39

16

Table2: Binding modes of selective compounds with PTP1B

NAME OF

COMPOUND


24-methylene

cycloartenol

Hydrophobic: Phe682A, Thr763A,

Tyr520A

H bond donor:

Gln762A



Rotatable Bonds: 13

Aromatic Atoms: 0


cLogP: 8.818TPSA: 20.230


Neg. Ionizable: 0

Pos. Ionizable: 0


17/39

17

Alpha-amyrine Hydrophobic:Phe682A, Thr763A

H bond donor: Phe682A




Aromatic Atoms: 0



Donors: 1


Cycloartenol Hydrophobic:Tyr546A, Leu588A

H bond donor: Lys541A


Molweight: 430.761



Aromatic Atoms: 0



Donors: 1



18/39

18

Friedelin Hydrophobic: Phe682A Formula: C30 H 50O 1Molweight: 426.729



Aromatic Atoms: 0



Donors: 0


Taraxerone Hydrophobic:

Phe682A, Tyr520A,Thr763A

H bond acceptor: Gln521A




Aromatic Atoms: 0


cLogP: 8.249TPSA: 20.230

Acceptors: 1

Donors: 0Neg. Ionizable: 0

Pos. Ionizable: 0


19/39

19

Table4: Binding modes of selective compounds with DPP4

NAME OF

COMPOUND


1,3,4,6-tetra-O-

galloyl-b-D-

glucose

H bond acceptors:

GLN553A, TYR547A,

LYS122A, ARG125A,

ASN710A, TRP629A

H bond donors:

TYR547A, VAL546A,

ASP545A, TRP629A,

HIS740A, TRP124A,

ASP709A


Molweight: 796.640


Rings: 5Rotatable Bonds: 30Aromatic Atoms: 24

Polarity & Chemical FeaturescLogP: -2.470TPSA: 390.060


Neg. Ionizable: 0

Pos. Ionizable: 0

24-

methylenecyclo

artenol

Hydrophobic

interactions :TYR547A

TRP627A

TRP629A

H bond donor:

VAL546A


Molweight: 416.734

Size & Flexibility

Atoms/Bonds: 82 / 85Rings: 4



cLogP: 7.961TPSA: 20.230




20/39

20

beta-amyrin H bond donor :

TYR752A




Aromatic Atoms: 0


cLogP: 5.416TPSA: 20.230

Acceptors: 0


Pos. Ionizable: 0

Cycloartenol Hydrophobic

interactions:

TYR547A

TRP627A

TRP629A


Molweight: 416.734





Donors: 0



21/39

21

Taraxerone Hydrophobic

interaction:

TYR547A

H bond acceptors:

D3C801A

ARG560ALYS554A

ASN562A

H bond donors: VAL546A

LYS554A

ASP545A




Aromatic Atoms: 16


cLogP: -1.774TPSA: 272.590

Acceptors: 5


Pos. Ionizable: 0


22/39

22

Table5: Binding modes of selective compounds with Alpha-Glucosidase

NAME OF

COMPOUND


Alpha-amyrin Hydrophobic

interaction:PHE450A,

THR204A, TRP406A


Molweight: 428.745





Neg. Ionizable: 0

Pos. Ionizable: 0

1, 3, 4-Tetra-O-

galloyl-b-

Dglucose.

Hydrogen bond donor:

TYR605A, ASP203A,

ASP542A, GLN603A,

TYR299A, BJ110011A

Hydrogen bond

acceptor:TYR605A, ARG202A,

GLN603A, TYR299A,

BJ11001A, ARG334A


Molweight: 794.624

Size & Flexibility

Atoms/Bonds: 90 / 95Rings: 6


Polarity & Chemical FeaturescLogP: -2.716TPSA: 390.060


Neg. Ionizable: 0

Pos. Ionizable: 0


23/39

23

Taraxerol Hydrophobic

interaction:TRP406A,

PHE450A




Aromatic Atoms: 0


cLogP: 8.249TPSA: 20.230

Acceptors: 0


Pos. Ionizable: 0

Friedelin Hydrophobic

interaction:PHE450A




Aromatic Atoms: 0


Acceptors: 0


Pos. Ionizable: 0


24/39

24

Taraxerone Hydrophobic

interaction:PHE450A,

TRP406A




Aromatic Atoms: 0


cLogP: 8.249TPSA: 20.230

Acceptors: 0


Pos. Ionizable: 0


25/39

25

Table3: Binding modes of selective compounds with PPAR

NAME OF

COMPOUNDIMAGE INTERACTION LIGAND DETAIL

1,3,4,6-tetra-O-

galloyl-b-D-

glucose

H bond acceptor:

GLU259A, ARG280A,

SER464A

H bond donor:LYS275A, OH group,

HIS466A


Size & Flexibility

Atoms/Bonds: 90 / 95Rings: 6Rotatable Bonds: 30

Aromatic Atoms: 24

Polarity & Chemical FeaturescLogP: -2.716


Donors: 17



26/39

26

Alpha-amyrine Hydrophobic: ILE262A,

PHE287A

Formula: C30 H 52O1Molweight: 428.745



Aromatic Atoms: 0



Donors: 1


beta-amyrin Hydrophobic: ILE262A,

PHE287A




Aromatic Atoms: 0


Acceptors: 1


Pos. Ionizable: 0


27/39

27

Friedelin Hydrophobic: ILE262A,

PHE287A




Aromatic Atoms: 0


TPSA: 20.230

Acceptors: 1


Pos. Ionizable: 0

Taraxerone Hydrophobic: ILE262A Formula: C30 H 50O 1Molweight: 426.729





Donors: 1



28/39

28

Conclusion

Docking simulation and pharmacophore analysis of 28 bioactive compounds extracted from

Euphorbia hirta has successfully performed the binding modes and gave strong evidence ofmolecular interactions to all five receptors (11HSD1, PTP1B, -glucosidase, PPARy,

DDP4)which directly related to diabetes type 2. In detail, this study showed that flavonoid

and terpenes families including 2,4_methylenecycloartenol, 1,3,4-tetra-O-galloyl-b-

Dglucose, fr iedeli n, amyrine, amyrine and taraxerol , taraxeronehave high binding

affinity to all 5 interested receptors.These binding results consist of a high number of

hydrogen bond and hydrophobic interactions at some similar specific position of each

receptor (showed from table 1 to 5). Moreover, the terpenes families (alpha-amyrine, beta-

amyrine, friedelin, taraxerol, taraxerone, cycloartenol) and the 3,4 di-O-galloyquinicacid of

tannin family show high binding affinity compared to others. However, based on theLipinski's rule of five, themolecular weight of 1,3,4-tetra-O-galloyl-b-Dglucose is greater

than 500Da (~700Da) so that a modification should be applied to decrease molecule weight

or be treated as a template for designing a new compounds in diabetes type 2 treatment.

Since normal docking process (AutodockVina was used in this research) docks thousands of

compounds from free chemical databases which are in freezes, it against the rigid structure

of receptors inaccurate binding affinity between drugs and proteins. Therefore, Molecular

dynamic simulation (MD simulation) is highly recommended to use for further research in

order to reach the optimum and accurate results of hydrophobic interaction and H bond

between ligands and receptors.


29/39

29

Appendix

Table 6. 3D structure of 5 target proteins from NCBI

SpeciesProtein PDBID 3D structure Method

Resolution

(A0)

Referen

Human

(Homos

apien)

11 HSD 1XU7 X-ray

diffraction

1.80 Hosfie

et al, 2

Human(Homos

apien)

PTP1B 3CWE X-ray

diffraction

1.60 Bioorg

Med.

Chem.L

(2008)

Human

(Homos

apien)

PPAR- 4A4V X-ray

diffraction

2.00 Journal(2013)

J.Med.C

m


30/39

30

Human

(Homosapien)

DPP4 4J3J X-ray

diffraction

3.20 Cheme

em ,20

Human

(Homos

apien)

Alpha-

glucosida

se

3L4T X-ray

diffraction

1.90 Sim,, L

2010


31/39

31

Table 7: 2D structures of 28 bioactive compounds candidates suggested from

Chemspider.

1,3,4,6-tetra-O-galloyl-

b-Dglucose

24_methylenecycloarte

nol

3_4 diOgalloyquinic

acid

Afzelin

Alpha amyrin Beta amyrin Beta sitosterol Campesterol

cyanidin 3,5-

diglucoside

Cycloarterol Friedelin Gallic acid

3,4,5-tri-O-

galloylquinic acid

Ingenoltriacetate Kaempferol Leucocyanidin

L_Rhamnose Myricitrin Pelargonium 3_5_

diglucoside


32/39

32

Protocatechuic acid Quercetin Quercetol Quercetrin

Resiniferonol Rutin Stigmasterol Taraxerone

Taraxerol


33/39

33

Table 8: 3D structure of 28 bioactive compounds building from GaussView

,3,4-tetra-O-galloyl-b-D-

glucose

24_methylenecycloartenol 3_4 diOgalloyquinic acid Afzelin

Alpha-amyrin Beta-amyrin Beta sitosterol Campesterol

cyanidin 3,5-diglucoside Cycloarterol Friedelin Gallic acid

Galloylquinic acid Ingenol triacetate Kaempferol Leucocyanidin


34/39

34

Potocatechuic acid Quercetin Quercetol Resiniferonol

Rutin Stigmasterol Taraxerone


35/39

35

Table 9: Position of Grid box center of five target proteins

Protein molecule Protein codeX,Y,Z coordination (Angstroms)

X Y Z

11b HSD1 1XU7 -64.809 -74.388 -13.559

PTP1B 3CWE 23.677 22.779 1.835

PPAR 4A4V -16.666 21.568 -47.791

DPP4 4J3J 5.418 15.755 -23.517

-glucosidase 3L4T 44.964 90.541 34.242

Table 10: Absolute binding energy (kcal/mol) of bioactive compounds in E.hirta

to five receptors.

11betaHSD1 A-glucosidase PTP1B PPARy DDP4

1,3,4-tetra-O-galloyl-b-Dglucose 8.3 7.9 6.9 7.9 9.4

24_methylenecycloartenol 6.4 6.8 6.4 7.4 6.9

3_4 diOgalloyquinic acid 6.9 7.3 7.5 7.5 8

Afzelin 6.5 7.6 7.1 6.9 8

Alpha amyrin 8.4 7.7 8 10 8.6

Beta amyrin 7.4 6.6 7.8 8.6 7.3

Beta sitosterol 6.5 6.4 7 7.3 8.7

Campesterol 7.6 6.8 7 7.2 9.9

cyanidin 3,5-diglucoside 6.4 7.2 7.5 7.9 8.3

cycloarterol 6.9 7.1 7.8 7.9 8

friedelin 8 7.7 8.3 8.8 9.7

gallic acid 5.5 6.4 5.2 5.2 5.7

galloylquinic acid 7.5 7 6.8 7.4 8.6

Ingenol triacetat 6.5 6.2 7.3 7 7.1

kaempferol 7.4 6.2 7 7 7.3

Leucocyanidin 7.5 6.4 6.9 6.9 6.9

L_Rhamnose 5.3 5.5 4.5 5.2 5

myricitrin 7.4 7.8 7.1 7 7.9

tinyatoxin 7.1

pelargonium 3_5_ diglucoside 8.3 7.6 7.9 8.3 9

protocatechuic acid 5.3 6.5 5 5.2 5.8

Quercetin 7.6 6.7 6.8 7.3 7.4

Quercitol 5.3 5.6 4.7 4.9 5.1

Quercitrin 6 7.9 6.9 7.4 7.7


36/39

36

Resiniferono 6.1 6 7.2 6.5 6.7

rutin 7.4 7.4 7.4 7.5 8.1

Stigmasterol 6.4 6.8 7 7.7 8.4

taraxerol 8.4 7.6 7.5 8.8 8.5

taraxerone 8.5 7.6 8.6 8.99.9

Table 11. Function of bioactive compounds in Euphorbia hirta

Family Bioactive

compounds

Biological

functions

References

Flavonoids Afzelin, quercetin,

quercitrin, quercitol,

rhamnose, rutin,

leucocyanidin,myricitrin, cyanidin

3,5- O-diglucoside,

kaemferon,

pelargonium 3,5-

diglucoside

Antioxidation, Anti-

allergy,

antibacterial,

molluscicidal, anti-diarrheal activity.

Mei Fen Shih1 and

Jong Yuh Cherng2

Taiwan 2012; Quy

Trinh, Ly Le, 2013.

Tannin gallic acid, 3,4-di-O-

galloyl-quinic acid,

3,4,5-tri-O-

galloylquinic acid,

1,2,3,4,6

tetraOgalloyl-b-D-

glucose

Antioxidation, anti-

inflammatory

activity.

Sunil Kumar,RashmiMalhotra,andDinesh

Kumar 2010, Yoshida

et al, 1990; Chen

1991.

Triterpenes and

phytosterols

2,4-

methylenecycloartenol,

betasitosterol,

campesterol, -

stigmasterol, ingenol

triacetate,

resiniferonol, alpha-

amyrine, beta-amyrine,

friedelin, taraxerol,taraxerone,

cycloartenol,

tinyatoxin,

protocatechuic acid

Anti-inflammatory,

antiplasmodial

activity.

Mei Fen Shih1 and

Jong Yuh Cherng,

2012; Sandeep

b.Patil, Nilofar

S.Naikwade,

Shandrakant

S.Magdum, 2009;

Quy Trinh, Ly Le,

2013.
http://www.ncbi.nlm.nih.gov/pubmed/?term=Malhotra%20R%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Malhotra%20R%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Malhotra%20R%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Malhotra%20R%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Kumar%20D%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Kumar%20D%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Kumar%20D%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Kumar%20D%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Kumar%20D%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Kumar%20D%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Malhotra%20R%5Bauth%5Dhttp://www.ncbi.nlm.nih.gov/pubmed/?term=Malhotra%20R%5Bauth%5D


37/39

37

Reference

i Curb JD, Rodriguez BL, Burchfiel CM, Abbott RD, Chiu D, Yano K: Sudden death,

impaired glucose tolerance, and diabetes in Japanese American men. Circulation91:2591-2595,1995

ii National Diabetes Data Group: Classification and Diagnosis of Diabetes Mellitus and

Other Categories of Glucose Intolerance.

doi:10.2337/diab.28.12.1039DiabetesDecember 1979vol. 28 no. 12 1039-1057

iii Sanford burnham medical research institute: diabetes type 2

iv Mei Fen Shih and Jong YuhCherng: Potential Applications of Euphorbiahirta in

Pharmacology. Department of Pharmacy, Chia-Nan University of Pharmacy & Science,

Tainan. Department of Chemistry & Biochemistry, National Chung Cheng University,

Chia-Yi, Taiwanv Liu, Y. et al. 2007. Antimalarial Flavonol Glycosides from Euphorbia hirta.

PharmaceuticalBiology. 45:278-81. Abstract

vi Tona, L., et al. 1999. Antimalarial activity of 20 crude extracts from nine African

medicinal plants used in Kinshasa, Congo.Journal of Ethnopharmacology68:193-203

vii Galvez, J. 1993. Antidiarrhoeic activity of Euphorbia hirta extract and isolation of an

active flavonoid constituent.Planta Med. 59: 333-6. Abstract

viii Galvez, J. 1993. Antidiarrhoeic activity of quercitrin in mice and rats. J. Pharm.

Pharmacol. 45:157-9. Abstract

ix Lanhers, M.C. et al. 1991.Analgesic, antipyretic and anti-inflammatory properties of

Euphorbia hirta.Planta Med. 57(3):225-31 Abstract

x Sudhakar, M., et al., 2006, Antimicrobial activity of Caesalpiniapulcherrima, Euphorbia

hirtaand Asystasiagangeticum.Fitoterapia, 77: 37880

xi Vijaya K., Ananthan S., &Nalini R. 1995.Antibacterial effect of theaflavin, polyphenon

60 (Camellia sinensis) and Euphorbia hirta on Shigella spp. -- a cell culture study.J.

Ethanopharmacol, 49(2): 115-8

xii Johnson, P.B. et al. 1999. Euphorbia hirta leaf extracts increase urine output and

electrolytes in rats.J. Ethnopharmacol. 65(1):63-9.xiii

Singh, G. D., et al. 2006. Inhibition of early and late phase allergic reactions by

Euphorbia hirtaL. PhytotherapyRes.20(4): 316-21.

xiv Martinez-Vazquez, M et al. 1999. Anti-inflammatory Active Compounds from the n-

Hexane Extract of Euphorbia hirta. Revista de la SociedadQuimica de Mxico.

43(3,4):103-5


38/39

38

xv Goldie Uppal, Vijay Nigam, Anil Kumar: Antidiabetic activity of ethanolic extract of

Euphorbia hirta Linn SRET College of Pharmacy, Barsar, Hamirpur, India xvi Zhang ZY, Lee SY - PTP1B inhibitors as potential therapeutics in the treatment of type

2 diabetes and obesity - 2003 Feb;12(2):223-33.

xvii Antihyperglycemic, antihyperlipidemic and antioxidant activities of Euphorbia hirta

extract - Sunil Kumar, RashmiMalhotra, Dinesh Kumar. International Research Journal

of Pharmacy (01/2010)

xviii http://www.gaussian.com/g_prod/gv5.htm

xix Review on targeted proteins for Diabetes Drug Desgin Trang D. Ngoc Nguyen, LyT.

Le.

xx New mechanisms of glucocorticoid-induced insulin resistance

xxi Role of glucocorticoids in the physiopathology of excessive fat deposition and insulin

resistance - C Asensio1,2,P Muzzin 2 and F Rohner-Jeanrenaud

xxii Tyrosine-protein phosphatase non-receptor type 1also known as protein-tyrosine

phosphatase 1B(PTP1B) is an enzymethat is the founding member of the protein

tyrosine phosphatase(PTP) family. In humans it is encoded by thePTPN1 gene.

xxiii Combs AP (March 2010). "Recent advances in the discovery of competitive protein

tyrosine phosphatase 1B inhibitors for the treatment of diabetes, obesity, and cancer".J.

Med. Chem. 53(6): 233344.

xxivTonks, NK (2003 Jul 3). "PTP1B: from the sidelines to the front lines!". FEBS

letters 546(1): 1408.

xxv Greene ME et al (1995). "Isolation of the human peroxisome proliferator activated

receptor gamma cDNA: expression in hematopoietic cells and chromosomal

mapping". Gene Expr. 4(45): 28199

xxvi Elbrecht A et al (July 1996). "Molecular cloning, expression and characterization of

human peroxisome proliferator activated receptors gamma 1 and gamma

2".Biochem.Biophys. Res. Commun. 224(2): 4317

xxviiMichalik L et al. (December 2006). "International Union of Pharmacology.LXI.

Peroxisome proliferator-activated receptors".Pharmacol. Rev. 58(4): 72641.

xxviii

Fajas L et al (July 1997)."The organization, promoter analysis, and expression of thehuman PPARgamma gene".J. Biol. Chem. 272(30): 1877989.

xxix Atanasov AG et al "Honokiol: a non- adipogenicPPAR agonist from

nature".Biochim.Biophys.Acta 1830(10):48139.

xxx P.Lalitha* and Shubashini K Sripathi - In silico ligand receptor docking of few

cyclitols for type II diabetes using hex-Department of Chemistry, Avinashilingam

Deemed University for Women, Coimbatore-641043.
http://www.nature.com/ijo/journal/v28/n4s/full/0802856a.html#aff1http://www.nature.com/ijo/journal/v28/n4s/full/0802856a.html#aff2http://www.nature.com/ijo/journal/v28/n4s/full/0802856a.html#aff2http://www.nature.com/ijo/journal/v28/n4s/full/0802856a.html#aff2http://www.nature.com/ijo/journal/v28/n4s/full/0802856a.html#aff2http://www.nature.com/ijo/journal/v28/n4s/full/0802856a.html#aff1


39/39

xxxiAlpha-Glucosidases at the US National Library of Medicine Medical Subject Headings

(MeSH)

xxxiihttp://www.rcsb.org/pdb/explore/explore.do?structureId=3L4T , abstraction

xxxiii

Mohammad et al., 2010, Sandeep et al.; 2011xxxiv REVIEW ON PHYTOCHEMISTRY AND PHARMACOLOGICAL ASPECTS OF EUPHORBIA HIRTA LINN.

SANDEEP B. PATIL*, MRS. NILOFAR S. NAIKWADE, CHANDRAKANT S. MAGDUM

xxxv Potential Applications of Euphorbia hirta in Pharmacology Mei Fen Shih1 and Jong Yuh

Cherng2*xxxviDownloaded at http://vina.scripps.edu/xxxvii

Downloaded at

http://www.rsc.org/chemistryworld/Issues/2006/September/LigandScout.asp

xxxviiiWolber G, Langer T. LigandScout: 3-D pharmacophore derived from protein bound

ligans and their use as virtual screening filters. J Chem Info Model. 2005: 45:160-169

xxxixLipinski CA, Lombardo F, Dominy BW, Feeney PJ Experimental and computational

approaches to estimate solubility and permeability in drug discovery and development

settings. Adv Drug Delivery Rev. 1997; 23:325.

xl L ipinski' s ru le of fi vehttp://www.pharmainformatic.com/html/rule_of_5.html

drug design paper

Documents