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WORLD JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES

Srinivas S et al.

World Journal of Pharmacy and Pharmaceutical Sciences

Volume 2, Issue 6, 5842-5851.

Research Article

ISSN 2278 – 4357

DESIGN, SYNTHESIS, BIOLOGICAL EVALUATION AND MOLECULAR DOCKING STUDIES OF NOVEL QUINAZOLINE DERIVATIVES AS GSK-3Β INHIBITORS Srinivas S*, Aparna V. Department of Pharmaceutical Chemistry Ganga Pharmacy College, Nizamabad-503001, A.P, India.

Article Received on 20 August 2013, Revised on 21 Sept 2013, Accepted on 17 November 2013

ABSTRACT A

novel

serial

of

3-[2-(methyl

amino)

methyl]-5-{[(2-

phenylquinazolin-4-yl)oxy]methyl}-1,3,4-oxadiazole-2(3H)-thione and N-4(substituted phenyl)-5-{[(2-phenylquinazolin-4-yl)-oxy]-methyl}1, 3, 4-thiadiazol-2-amine derivatives were synthesized, evaluated for

*Correspondence for

their hypoglycemic activity and molecular docking studies were performed against X-ray crystal structure of GSK-3β, are shown their

Author:

role in the hypoglycemic activity. The molecules are the high potency * Srinivas S,

of compound 6a, 6g, 8a, 8b, and 8e based on glide score and binding

Department of Pharmaceutical Chemistry Ganga Pharmacy

poses of the molecules.

College, Nizamabad-503001,

Key words: GSK-3β inhibitors, quinazolinyl oxadiazoles and kinase

India.

inhibitors.

[email protected]

INTRODUCTION (Glycogen synthase kinase (GSK) is ser/ thr kinase, originally identified as the enzyme responsible for the inactivating phosphorylation of glycogen synthase1. The two isoforms GSK-3α (51 KDa) GSK-3β (47KDa) are known and there is high homology (90%) in their kinase domine2. GSK-3β plays a crucial role in glucose homeostasis, Central Nervous system (CNS) function (via protein tau and β-catenin) and cancer (via angiogenesis, apoptosis and tumor genesis)2. Introduction of gsk-dependent phosphorylation should activate insulin dependent glycogen synthesis; there by mimicking the action of insulin to lower plasma glucose3, thus inhibitors of GSK-3β would afford a novel mode of treating type-II diabeties4,

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along with

World Journal of Pharmacy and Pharmaceutical Sciences

type-II diabetes gsk-3β inhibitors have therapeutic potential for treating

neurodegeneration disease, bipolar disorder, strock, cancer, chronic inflammatory disease and alzemer,s disease, huntingtone,s disease, cardiac ischemia and age related loss of bone4. Keeping in view of this fact, the design and synthesis of newer GSK-3β inhibitors are of immense significance and continue to attract the attention of numerous medicinal chemist and in this present study a novel serice of quinazolinyl oxadiazoles and quinazolinyl thiadiazoles derivatives were synthesized (Scheme-1) and recrystalised by dimethyl formamide as solvent and all synthesized compounds were characterized by IR, H1NMR, a molecular docking studies were performed against X-ray crystal structure of GSK-3β (pdb code; 1q3d). All these molecules were docked in to the ATP binding site of GSK-3β and their docking score represented in table no.1 and evaluated for their hypoglycemic activity. MATERIALS AND METHOD Melting points were recorded on melting apparatus and are uncorrected. IR spectra were recorded on a perkin-Elmer FT-IR 240-c spectrophotometer using KBr optic. H1NMR spectra were recorded on Bruker (Bruker BioScience, USA) AV300 MHz in CDCl3 using TMS as an internal standard. All reactions were monitored by thinlayer chromatography (TLC) on precoated silicagel 60F254 (mesh); spots were visualized with UVlight. The compound (3):(2-phenyl-4-Quinazolinyl)-oxy-acetate were synthesized by reported method5 General procedure for synthesis of (2-phenyl-4-Quinazolinyl)-oxy-acetyl hydrazine (4): Compound 3 (0.01 mol) and hydrazine hydrate (0.01 mol) were taken up in absolute ethanol (15ml) and heated on a water bath at reflux for period of 6hrs. The solid that precipitated on cooling were filtered, dried and purified by column chromatography. The physical data of synthesized compound were represented in table no. 2. IR (KBr, cm-1): 1290(C-O-C), 1616 (C=N), 1762(C=O), 3056, 2918(C-H); H1NMR (CDCl3, ppm) δ: 4.2(s, 2H, OCH2), 4.4 (s, 2H, NH2), 7.4-8.4 (m, 9H, Ar-H), 9.3(s, 1H, N-H). General procedure for synthesis of 5{(2-phenyl quinazolin-4yl)-oxy- methyl}-1, 3, 4 oxadiazole-2(3H)-thione (5): Compound 4 (0.01mol) and carbon disulphide (0.03) were taken in ethanolic solution of KOH (0.1mol) 15 ml, and heated at reflux on a water bath for 20 hrs. Completion of the reaction is monitored by TLC, the solvent was evaporated under reduced pressure and the residue was triturated with cold water, neutralization of the solution with acetic acid yielded a www.wjpps.com

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precipitate which was filtered, dried and purified using column chromatography. The physical data of synthesized compound were represented in table no. 2. IR (KBr, cm-1): 3448 (N-H), 1204 (C=S), 1632 (C=N), 3051, 2851 (C-H); H1NMR (CDCl3, ppm) δ: 4.3 (s, 2H, OCH2) 7.3(s, 1H, NH), 7.4-7.6 (m, 4H, Ar-H), 7.8-7.9 (t, 2H, Ar-H), 8.2-8.3 (t, 2H, Ar-H), 8.5(s, 1H, Ar-H). Synthesis of 3-[2-(methyl amino) methyl]-5-{[(2-phenylquinazolin-4-yl)-oxy]-methyl}-1,3,4oxadiazole-2(3H)-thione(6a-): To a boiling solution of compound 5 (0.01mol) in ethanol (25ml) containing aqueous formaldehyde (40% 1ml) and N-alkyl amines (0.01mol) was added with stirring and the solution heated for 30 mints and left over night. The solid separated was filtered under reduced pressure, washed with pet. ether and recrystalised from ethyl acetate and purified using column chromatography. Similarly the compounds 6b to 6i were synthesized by using different N-alkyl amines and the physical data of synthesized compound were represented in table no. 2. IR (KBr, cm-1): 3063, 2359 (C-H), 2925 (N-H) 1622, 1606 (C=N), 1208 (C=S); H1NMR (CDCl3, ppm) δ: 7.4-8.4 (m, 9H, Ar-H), 4.0 (s, 2H, OCH2), 3.26-3.9 (m 5H CH2CH3), 2.0 (s, 1H, N-H). Synthesis

of

N1-2-phenyl-1,3-quinazolin-4-(yl-oxymethyl)-N4-p-(substituted

phenyl)-

thiosemicarbazide(7): To a boiling solution of compound 4 (0.001mol) in absolute ethanol (25ml) p-substituted phenyl isothiocynate (0.001mol) was added and the solution heated on reflux about 30mints, complete the reaction was monitored by TLC. The solid separated was filtered, recrystalised from DMF and purified using column chromatography. The physical data of synthesized compound were represented in table no. 2. Synthesis of N-phenyl-5-{[(2-phenylquinazolin-4-yl)-oxy]-methyl}-1, 3, 4-thiadiazol-2amine (8a): To the chilled H2SO4 (AR, 25ml) compound 7 (0.001 mol) was added gradually with constant stirring, after the addition was over, poured on crushed ice (100g) with water, recrystalised from (DMF) di methyl formamide and purified using column chromatography. Similarly the compounds 8b to 8e were synthesized and the physical data of synthesized compound were represented in table no. 2. IR (KBr, cm-1): 3195 (N-H), 3061 (C-H), 1627 (C=N); H1NMR (CDCl3, ppm) δ: 10.22 (s, 1H, N-H), 7.4-8.2 (m 13H, Ar-H), 5.20 (s, 2H, OCH2), 1.90 (m, 3H, CH3). www.wjpps.com

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3-[2- (ethyl amino) methyl]-5-{[(2-phenylquinazolin-4-yl)-oxy]-methyl}-1,3,4-oxadiazole2(3H)-thione (6b): H1NMR (CDCl3, ppm) δ: 7.4-8.4 (m, 9H, Ar-H), 4.0 (s, 2H, OCH2), 2.0 (s, 1H, N-H), 2.59-3.26 (m 5H CH2CH3), 3.91 (s, 2H, CH2). 3-[2- (propyl amino) methyl]-5-{[(2-phenylquinazolin-4-yl)-oxy]-methyl}-1,3,4-oxadiazole2(3H)-thione (6c): H1NMR (CDCl3, ppm) δ: 7.4-8.4 (m, 9H, Ar-H), 4.0 (s, 2H, OCH2), 2.0 (s, 1H, N-H), 0.90-1.48 (m 5H CH2CH3), 3.91 (s, 2H, CH2), 2.59-3.26 (m 5H CH2CH3). 3-[2- (butyl amino) methyl]-5-{[(2-phenylquinazolin-4-yl)oxy]methyl}-1,3,4-oxadiazole2(3H)-thione (6d): H1NMR (CDCl3, ppm) δ : 7.4-8.4 (m, 9H, Ar-H), 4.0 (s, 2H, OCH2), 2.0 (s, 1H, N-H), 3.91 (s, 2H, CH2), 2.59-3.26 (m, 5H, CH2CH3). 3-[2-

(morpholinyl

amino)

methyl]-5-{[(2-phenylquinazolin-4-yl)-oxy]-methyl}-1,3,4-

oxadiazole-2(3H)-thione (6e): H1NMR (CDCl3, ppm) δ: 7.4-8.4 (m, 9H, Ar-H), 4.0 (s, 2H, OCH2), 2.0 (s, 1H, N-H), 4.01 (s, 2H, CH2), 2.9 (m, 4H, CH2 CH2), 3.65 (m, 4H, CH2 CH2). 3-[2-

(piperzinyl

amino)

methyl]-5-{[(2-phenylquinazolin-4-yl)oxy]methyl}-1,3,4-

oxadiazole-2(3H)-thione (6f): H1NMR (CDCl3, ppm) δ: 7.4-8.4 (m, 9H, Ar-H), 4.0 (s, 2H, OCH2), 2.0 (s, 1H, N-H), 4.01 (s, 2H, CH2), 2.67 (m, 4H, piperidine). N-(4-methylphenyl)-5-{[(2-phenylquinazolin-4-yl) oxy]-methyl}-1, 3, 4-thiadiazol-2-amine (8b): H1NMR (CDCl3, ppm) δ: 10.22 (s, 1H, N-H), 7.4-8.2 (m 13H, Ar-H), 5.20 (s, 2H, OCH2), 1.90 (m, 3H, CH3). N-(4-methoxy phenyl)-5-{[(2-phenylquinazolin-4-yl) oxy]-methyl}-1, 3, 4-thiadiazol-2amine (8c): H1NMR (CDCl3, ppm) δ: 10.22 (s, 1H, N-H), 7.4-8.2 (m 13H, Ar-H), 5.20 (s, 2H, OCH2), 3.80 (s, 3H, CH3). N-(2-methoxy phenyl)-5-{[(2-phenylquinazolin-4-yl) oxy]-methyl}-1, 3, 4-thiadiazol-2amine (8d): H1NMR (CDCl3, ppm) δ: 10.22 (s, 1H, N-H), 7.4-8.2 (m 13H, Ar-H), 5.20 (s, 2H, OCH2), 3.80 (s, 3H, CH3). N-(2-nitro phenyl)-5-{[(2-phenylquinazolin-4-yl) oxy]-methyl}-1, 3, 4-thiadiazol-2-amine (8e): H1NMR (CDCl3, ppm) δ: 10.22 (s, 1H, N-H), 7.4-8.2 (m 13H, Ar-H), 5.20 (s, 2H, OCH2).

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MOLECULAR DOCKING STUDIES All the derivatives of quinazolinyl oxadiazoles and quinazolinyl thiadiazoles were selected for their molecular docking studies on selected X-ray crystal structure of GSK-3β (pdb code; 1q3d)(3) using glid ( Schrodinger, OPLS-2005 software), the possible binding mode of a GSK-3β inhibitors illustrate in fig.no.1. The compounds (14) were docked in to ATP binding site of GSK. The key interaction for this complex includes hydrogen bonding between portion of amide hydrogen and back bone of carbonyl group with Val135, Asp 200, and Asp 186 respectively. The substituted functional group on phenyl ring to forms another hydrogen bonding with Gln 185, Lys85 and electrostatic interaction of phenyl ring with Tyro140, pyridine nitrogen to forms electrostatic interaction withTyr134, the interaction image of compound 8 with in to ATP binding site of GSK were illustrate in fig.no.2. The docking score calculated by glid score and their docking score of all molecules were shown in table no.1. PHARMACOLOGY All the compounds were screened on albino rats for their hypoglycemic activity following the method of agarwal (6). A group of albino rats were selected randomly assigned into three groups (1-3) of five rats (n=5) each as follows, group 1 received vehicle (Distilled water) and served as control group, group 2 received standard drug (Tolbutamide) and served as standard group. Group 3 received test compounds (100 mg/kg) and served as test- group. All these animals kept on fast for 12 hrs, the fasting blood sugar sample were drawn from the tail veins of the rats and their glucose content was measured. The compounds to be tested were emulsified with 5% gum tragacanth and administered orally at single dose of 100 mg/kg body weight. The blood sample were again drawn from the tail vein at intervals of 1, 2 and 4 hrs and their glucose content were again determined and the percentage Change of glucose level were calculated and results shown in table 2. RESULTS AND DISCUSSION The synthesis of quinazoline oxadiazoles and quinazoline thiadiazole derivatives was achieved following the steps outlined in scheme 1.The antranilic acid underwent cyclization with benzoyl chloride in pyridine at 0-5oC to give 2-phenyl-4(H)-3,1 benzoxazin4-one (1) by benzylation followed by dehydration. Compound 1 up on reaction with foramide results in the more stable 2-phenyl-4(3H)-quinazolinone (2), up on alkylation Compound 2 with ethylchloroacetate in dry acetone over anhydrous potassium carbonate yielded its O-alkylated

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product; ethyl-[(2-phenyl-4-quinazolinyl)-oxy]-acetate (3). Compound 3 was reacted further with hydrazine hydrate (80%) in absolute alcohol to get the corresponding 2-phenyl-4quinazolinyloxy acetyl hydrazide (4). The compound 4 was reacting further with carbon disulphide (CS2) and KOH and p- substituted phenyl isothiocynate yielded 21 thia oxadiazoles (5) and 2phenyl-1, 3 quinazolin-4-oxymethyl-N-p (substituted phenyl) thiosemicarbazide (7). The compound 5 reacts further with formaldehyde 40% and different alkyl amines yields N-methyl 5-(2-phenyl-1, 3 quinazolin 4-yl oxy methyl)-2 thio- 1, 3, 4 oxadiazole(6).The compound 7 undergoes cyclization with H2SO4 (AR) yielded a 2phenyl (1, 3-quinazolin 4-yl oxy methyl)-N-P-(substituted phenyl) 1, 3, 4-thiadiazole derivatives (8). All the synthesized compounds were characterized based on the physical and spectral data; the physical data of all compounds were given in table no.2. The molecular docking studies were performed against crystal structure of GSK-3β and the respective docking scoring functions were given in table no.1. The 6a, 6b, 6c, 6f, 6d, 6g of oxadiazoles and 8a, 8c, 8f, 8b of thiadiazole derivatives are shown to be exhibit good binding its ATP binding sites of GSK-3β by interacting with Val135, Asp186, Asp200 and lys85, the docking results were given in table no.1. All the synthesized compound were evaluated for their hypoglycemic activity on rats and percentage reduction of blood glucose level were calculated, results were given in table no.2. The substituted phenyl on thiazolo- quinazoline is exhibit greater hypoglycemic activity than alkyl substituted oxadiazolo-quinazoline. The compounds 8c, 8b, 8a and 6i, 6f, 6a are shown to be exhibits good hypoglycemic activity. Table. No. 1 Molecular docking studies of all synthesized compounds S.No

class

Entry ID

R1

R2

1

6a

277

H

CH3

-6.06

-6.06

-0.9

2

6b

278

H

C2H5

-5.61

-5.61

-0.8

3 4 5 6 7

6c 6d 6e 6f 6g

279 280 281 282 283

H H H H H

propyl butyl morpholine piperidine p-tolylene

-5.75 -5.96 -5.5 -4.51 -6.49

-5.75 -5.96 -5.5 -4.51 -6.49

-0.9 -0.9 -0.35 0 -0.19

8

6h

284

H

Phenyl piperidine

-4.47

-4.47

0

9

6i

285

H

p-methoxy

-2.3

-2.3

-0.32

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docking XP score GScore

XP HBond

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10 11

8a 8b

293 294

12

8c

295

13

8d

296

14

8e

297

H CH3 pOCH3 OOCH3 O-NO2

piperidine ---

-6.11 -6.11

-6.11 -6.11

-0.7 -0.7

--

-3.93

-3.93

0

--

-5.65

-5.65

-0.7

--

-7.55

-7.55

-0.7

O

COOH

O

a

b

O N

NH 2

c

NH N

COOC 2H 5

II

I

O CONHNH 2 N

d O

N N

III N

g

e

IV

N

NH

O R1 O

O

N

CONHNHCSNH N

S

N

N

V

VII

F R1 N

h

N

N

O N

N

O

N NH

N

R2

S

O

S

N

N

VI VIII R1

Scheme: 1 Synthesis of quinazolinyl oxadiazoles & quinazolinyl thiadiazoles Reagents conditions: a) Benzoyl chloride & pyridine, b) HCONH2, c) Ethyl chloroacetate and dry acetone d) NHNH2.H2O/ ethanol, e) CS2/KOH, f) HCHO, N-alkyl amine, g) psubstituted phenyl isothiocynate, h)H2SO4 (AR).

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Table. No. 2 Physical data and hypoglycemic activity of all synthesized compounds

S.no.

R1

R2

m.p. c

%Yield

6a 6b 6c 6d 6e 6f 6g

H H H H H H H

263-265 182-185 190-192 241-243 150-152 210-212 235-137

70 65 80 75 70 60 55

6h

H

190-192

45

15

6i

H

196-198

50

13

8a 8b

H CH3

CH3 C2H5 propyl butyl morpholine piperidine p-tolylene Phenyl piperidine p-methoxy piperidine ---

Hypoglycemic activity(%reduction blood sugar) 15 13 10 -17 20 15

124-126 140-142

70 65

15 17

8c

p-OCH3 OOCH3 O-NO2

--

146-148

65

20

--

174-176

50

--

--

166-168

55

13

8d 8e

o

Fig.1 Docking image of compound 8 in an X-ray crystal structure of GSK-3β (pdb code; 1q3d)

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Fig.2 Binding interaction of compound 8 with ATP binding site of GSK-3β CONCLUSION Gsk3β represented a promising target and designing gsk3β inhibitors would offer a novel approach to develop potent inhibitors in this class. In the present study we were synthesized quinazolinyl oxadiazoles and quinazolinyl thiadiazoles and molecular docking study were performed against crystal structure of gsk3β and this study results helpful in further detailed study for developing a potential gsk3β inhibitors. ACKNOWLEDGMENT Authors are thankful to the Ganga Educational Society for providing the facilities.

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4. Andrew J, Joyce A, Dickerson H, Dulce G. Novel pyrazolopyrimidine derivatives as GSK-3 inhibitors. Bioorg. Med. chem. Lett, 2004; 14: 2121-2125. 5. Shireesha B, Umasankar K, Srinivas S, Mallareddy V. Synthesis, Antimicrobial Evaluation, and Docking Studies of Novel 4-Substituted Quinazoline Derivatives as DNA-Gyrase Inhibitors. Arch. Phar.chem.lifesci. 2010; 10: 570-576. 6. Agarwal V.R, Nautiyal S.R, Mukerji D.D. Indian drugs, 1986; 23: 458-461. 7. Yasushi M, Yutaka M, Hidenyuki S, masato N, Geoffrey W.M. Rational design of 4amino-5, 6-diaryl-furo[2,3-d] pyrimidines as potent glycogen synthase kinase-3 inhibitors. Bioorg. Med. chem. Lett, 2008; 18: 1967-1971. 8. Nigus D, Prasad V. B, Euro. J.Med.Chem.3D-QSAR studies on antitubercular thymidine monophosphate kinase inhibitors based on different alignment methods 2006; 42: 10141027. 9. Michael J. Alberti, Elizabeth p. Auren, Karen E. Laukey, Discovery and invitro evaluation of potent kinase inhibitors: pyrido[ 1.2: 1.5] pyrazolo[3,4-d] pyrimidines, Bioorg. Med. chem. Lett, 2005; 15: 3778-3781. 10. Jason W, Vincent B, Alessandra G, Neil S. G. 6-Aryl-pyrazolo[3,4-b] pyrimidines: potent inhibitors of glycogen synthase kinase-3 (GSk-3). Bioorg. Med. chem. Lett, 2007; 13: 3055-3057. 11. David G.S, Buffet M, Fenwick AE, David H, Robert J. Macrocyclic bisindolylmaleimides as inhibitors of protein kinase C and glycogen synthase kinase-3. Bioorg. Med. chem. Lett, 2003; 13: 3049-3053.

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