Clubbed thiazoles by MAOS: A novel approach to cyclin-dependent kinase 5/p25 inhibitors as a potential treatment for Alzheimer's disease

DOI: 10.1016/j.bmc.2007.01.043

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Bioorganic and Medicinal Chemistry p. 2601 - 2610 (2007)

Update date:2022-09-26

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Authors:

Shiradkar, Mahendra Ramesh

Akula, Kalyan Chakravarthy

Dasari, Varaprasad

Baru, Vijayakumar

Chiningiri, Bhoomeshwar

Gandhi, Santosh

Kaur, Ranjit

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Article abstract of DOI:10.1016/j.bmc.2007.01.043

A novel clubbed triazolyl thiazole series of cdk5/p25 inhibitors, potentially useful for the treatment of Alzheimer's disease, is disclosed. Evaluation of the SAR of substitution within these series has allowed the identification of a range of compounds w

Full text of DOI:10.1016/j.bmc.2007.01.043

Bioorganic & Medicinal Chemistry 15 (2007) 2601–2610

Clubbed thiazoles by MAOS: A novel approach

to cyclin-dependent kinase 5/p25 inhibitors as a potential

treatment for Alzheimer’s disease

Mahendra Ramesh Shiradkar,^a,*Kalyan Chakravarthy Akula,^aVaraprasad Dasari,^b

Vijayakumar Baru,^bBhoomeshwar Chiningiri,^cSantosh Gandhi^dand Ranjit Kaur^e

^aDr. Reddys Laboratories, 7-1-27, Ameerpet, Hyderabad 16, Andhra Pradesh, India

^bMedicinal Chemistry Laboratory, CKM P.G. College, Desaipet, Warangal 500001, India

^cSai Life Sciences, L.B.Nagar, Hyderabad, Andhra Pradesh, India

^dAISSMS College of Pharmacy, Kennedy Road, Pune 411001, India

^eSt. Johns College of Pharmacy, Vijayanagar, Bangalore 560040, India

Received 7 December 2006; revised 25 January 2007; accepted 26 January 2007

Available online 30 January 2007

Abstract—A novel clubbed triazolyl thiazole series of cdk5/p25 inhibitors, potentially useful for the treatment of Alzheimer’s dis-

ease, is disclosed. Evaluation of the SAR of substitution within these series has allowed the identiﬁcation of a range of compounds

which signiﬁcantly reduce brain cdk5/p25 and thus have potential as possible treatments for Alzheimer’s disease.

1. Introduction

tion. Literature survey reveals 2-aminothiazole deriva-

tives⁶as the potential inhibitors of cdk5/p25 for the

treatment of Alzheimer’s disease and other neurodegen-

erative disorders.^7–13

Alzheimer’s disease (AD) is a progressive neurodegener-

ative disorder accompanied by memory decline, cogni-

tive impairment, and visual-spatial disorientation, for

which no eﬀective treatment exists today. Postmortem

brain analysis of AD patients reveals extensive forma-

tion of neuroﬁbrillary tau protein tangles and amyloid

plaques. The serine/threonine kinase cdk5 along with

its cofactor p25¹(or the longer cofactor, p35) has been

supposed to hyperphosphorylate tau,²leading to the

formation of paired helical ﬁlaments and deposition of

cytotoxic neuroﬁbrillary tangles³and thus responsible

for neurodegenerative disorders such as Alzheimer’s dis-

ease, Parkinson’s disease, stroke, or Huntington’s dis-

ease.⁴cdk5 also phosphorylates Dopamine and Cyclic

AMP-Regulated Phosphorprotein (DARPP-32) at

threonine 75 and is thus indicated in having a role in

dopaminergic neurotransmission.⁵Inhibition of the

anomalous cdk5/p25 complex is, therefore, a viable tar-

get for treating Alzheimer’s disease by preventing tau

hyperphosphorylation and neuroﬁbrillary tangle forma-

Based on this hypothesis, we embarked on a cdk5/p25

inhibitor discovery program to ﬁnd an orally bioavail-

able, high potency compound/s. Screening of an in-

house database provided several hits with modest

cdk5/p25 inhibitory activity, one of which was the

clubbed triazolyl thiazole 1 (IC₅₀= 48 2 nM).

In recent years, environmentally benign synthetic meth-

ods have received considerable attention and solvent

free protocols are reported.^14–16A fast, highly eﬃcient

and eco-friendly solvent-free chemical transformation,

for the synthesis of title compounds, under microwave

irradiation, using acidic alumina is designed Scheme 1.

2. Results and discussion

2.1. Synthesis

Keywords: Thiazole; Triazole; cdk5/p25; Alzheimer’s disease.

Compounds 1a–d, 2e–l, 3e–l, 4e–l, 5e–l, 6e–l, 7m–aj,

8e–l, and 9e–l were synthesized as per the literature.^17,18

Corresponding author. Tel.: +91 9989263660; fax: +91

4023731955; e-mail: rrshiradkar@rediﬀmail.com

doi:10.1016/j.bmc.2007.01.043

2602

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

NHCOR₁

NH₂

R₁COCl

CH₂I₂

2a-l

CH₂

1a-d

3a-l

ClCH₂CN

NHCOR₁

BrCH₂COOCH₃

NHCOR₁

SCH₂CN

SCH₂COOCH₃

NH₂NH₂H₂O

5a-l

4a-l

NHCOR₁

C₆H₅CHO

SCH₂CONHNH₂

6a-l

CS₂/KOH, N₂H_4.H₂O

R₂COCl

NHCOR₁

SCH₂CONHN=CHC₆H₅

CH₂

8a-l

NHCOR₁

NH₂

9a-l

SCH₂CONHNHCOR₂

7a-aj

Scheme 1.

Compounds 1a–d, adsorbed on acidic alumina (alumi-

num oxide, acidic, Brockmann I, ꢀ150 mesh, 58 A CA-

It was observed that there is remarkable loss of product

(44% yield) in the second step for the conversion of 6a–d

to 9a–d when performed in conventional method, while

reaction involving MORE method gave good yield

(71–80%).

MAG 506-C-I, surface area 155 m²/g, pH 6.0), when

treated with 4-chlorobenzoyl chloride at 0 ꢁC to yield

2a–d. The transformed compounds 2a–d on treatment

with diiodomethane in the presence of strong alkali, that

is, sodium hydroxide, gave 3a–d. Title compounds 2a–d

were treated with chloroacetonitrile, which on neutral-

ization with sodium carbonate gave precipitates of com-

pounds 4a–d. Compounds 2a–d, when treated with

methyl bromoacetate in basic condition, produced 5a–d.

Chemical transformation of 5a–d to 6a–d was achieved

by treating it with hydrazine hydrate. While compounds

6a–d, on treatment with appropriate acid chlorides, fur-

nished 7a–l. Schiﬀ bases, the condensation products of

8a–d, were synthesized by treating 6a–d with benzalde-

hyde and conﬁrmed by absence of triplet of NH of

hydrazide. Compounds 6a–d were converted to thiocar-

bazate salts by treatment with carbon disulﬁde and

potassium hydroxide, which on treatment with hydra-

zine hydrate gave 9a–d. The NMR spectra conﬁrmed

formation of triazole derivative from hydrazide, which

shows presence of sulfhydryl proton at d value 12.5.

2.2. Cyclin-dependent kinase 5/p25 inhibiting activity

Kinase inhibition was measured by the use of scintilla-

tion proximity assays (SPA).⁸

The results of the assays are reported in Tables 1–4.

During the preliminary screening compound 1b has

emerged as hit cdk5/p25 (IC₅₀= 48 2 nM), with good

potency and more opportunities for chemical transfor-

mation for the optimization. Testing of 1b against other

cdks revealed that 1b was essentially equipotent at inhib-

iting cdk2/cyclin E (IC₅₀= 50 3 nM), a cancer target.

Thus with an objective to improve cdk5 potency and

minimize cdk2 activity, certain chemical modiﬁcations

have been performed. Variation of the amine side chain

of 1b with MAOS allowed us to rapidly explore the ﬁrst

arm of the pharmacophore. As a ﬁrst step toward lead

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

Table 1. SAR of cdk5/p5 inhibitory screening

2603

R₁

R₂

cdk5 IC₅₀(nM)

NHCOCH₂Cl

NHCOCH₃

NHCOC₆H₅

—

390 89

048 02

630 32

264 27

642 11

462 72

186 12

462 50

761 84

574 65

894 114

674 68

684 78

598 58

614 46

436 28

340 22

382 24

640 34

168 11

674 46

647 84

698 46

587 28

847 86

847 64

764 84

743 46

562 79

062 11

064 04

426 14

657 67

395 64

496 84

487 96

415 78

463 86

574 64

641 78

440 102

380 32

290 41

560 22

674 46

587 76

364 74

648 124

695 118

467 106

598 110

746 148

044 02

072 02

034 01

064 01

—

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–4-ClC₆H₄

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

–CH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–4-ClC₆H₄

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

–CH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–4-ClC₆H₄

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

–CH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–4-ClC₆H₄

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

–CH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–4-ClC₆H₄

NHCOC₆H₅

NHCH₂CH₂COOH

optimization amino group was protected to the corre-

sponding compounds 2a–l however, all of these modiﬁ-

cations resulted in a substantial decrease in activity.

The next structural modiﬁcation made was a dimeric

product of 3a–l but these changes also resulted in a

substantial loss of biological activity.

s-Alkylation with acetonitrile provided the ﬁrst analogs

4b and 4c that demonstrated excellent activity, while

others exhibited moderate to poor activity. Thus it was

decided to modify the structure at SH group. In order

to optimize the sulfhydryl component, compounds 5a–

l were synthesized and investigated, which revealed loss

2604

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

Table 2. SAR of cdk5/p5 inhibitory screening

R₁

R₂

cdk5 IC₅₀(nM)

NHCOCH₂Cl

NHCOCH₃

NHCOC₆H₅

–C₆H₅

—

246 38

197 49

268 78

465 84

201 34

241 36

186 29

943 23

464 64

641 52

340 30

352 24

640 18

534 48

484 43

268 14

224 64

260 44

340 23

180 41

356 64

542 46

564 87

472 46

467 92

421 46

463 75

432 80

530 64

643 84

542 74

462 55

354 43

384 48

643 67

463 47

365 36

436 47

413 81

476 73

462 68

476 74

356 69

264 46

042 01

030 01

246 28

238 34

467 52

485 30

496 54

526 74

536 49

463 86

467 44

635 64

–C₆H₅

—

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–CH₃

—

NHCOC₆H₅

–CH₃

—

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–4-ClC₆H₄

–C₆H₅

CH₃

C₆H₅

CH₂Cl

CH₃

C₆H₅

CH₂Cl

CH₃

C₆H₅

CH₂Cl

—

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

7aa

7ab

7ac

7ad

7ae

7af

7ag

7ah

7ai

7aj

–C₆H₅

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

–CH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–CH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–CH₃

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–4-ClC₆H₄

–C₆H₅

—

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

—

–C₆H₅

—

NHCOC₆H₅

NHCH₂CH₂COOH

NHCOCH₂Cl

NHCOCH₃

–C₆H₅

–CH₃

—

–CH₃

—

NHCOC₆H₅

NHCH₂CH₂COOH

—

of activity. A further modiﬁcation of compounds 5a–l

produced compounds 6a–l. The results of the cdk5/p25

inhibitory activity are quite interesting because four

6a–d of these compounds have shown impressive per-

centage of inhibition. Compounds 6a–l were selected

for further studies as they have a free amino group,

which opened an area for further modiﬁcation at this

point. Compounds 7a–aj were obtained by treatment

with acid chlorides which ultimately showed decreased

activity. Furthermore, compounds 6a–l were converted

to Schiﬀ bases with benzaldehyde, and on investigation,

8a–d have shown promising activity while others re-

mained inactive. Compounds 9a–l were found to be

inactive.

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

Table 3. SAR of cdk5/p5 inhibitory screening

2605

(d) are reported in ppm and coupling constants (J) are

given in Hertz. Signal multiplicities are represented by s

(singlet), d (doublet), t (triplet), ds (double singlet), dd

(double doublet), m (multiplet), and br s (broad singlet).

Mass spectra were recorded on a Finning LCQ mass

spectrometer. Microwave irradiation was carried out in

Raga Scientiﬁc Microwave Systems, Model RG31L at

2450 MHz. Elemental analysis was performed on a Hera-

cus CHN-Rapid Analyser. Analysis indicated by the

symbols of the elements of functions was within 0.4%

of the theoretical values. The purity of the compounds

was checked on silica gel coated Al plates (Merck).

R₁

R₂cdk5 IC₅₀(nM)

NHCOCH₂Cl

NHCOCH₃

NHCOC₆H₅

–4-ClC₆H₄

—

3260 106

7480 114

3180 212

6650 142

6985 164

6537 148

4653 174

4785 156

4635 186

5246 210

6342 200

6412 246

NHCH₂CH₂COOH –4-ClC₆H₄

NHCOCH₂Cl

NHCOCH₃

NHCOC₆H₅

–C₆H₅

NHCH₂CH₂COOH –C₆H₅

NHCOCH₂Cl

NHCOCH₃

NHCOC₆H₅

–CH₃

NHCH₂CH₂COOH –CH₃

Kinase inhibition was measured by use of scintillation

proximity assays (SPA).¹⁰Enzyme activities were as-

sayed as the incorporation of [33P] from the gamma

phosphate of [33P] ATP (Amersham, cat. no. AH-

9968) into biotinylated peptide substrate PKTPK

KAKKL. Reactions were carried out in a buﬀer con-

taining 50 mM Tris–HCI, pH 8.0; 10 mM MgCl₂,

0.1 mM Na₃VO₄, and 1 mM DTT. The ﬁnal concentra-

tion of ATP was 0.5 lM (ﬁnal speciﬁc radioactivity of

4 lCi/nmol), and the ﬁnal concentration of substrate

was 0.75 lM. Reactions, initiated by the addition of

cdk5 and activator protein p25, were carried out at

room temperature for 60 min. Reactions were stopped

by addition of 0.6 volume of buﬀer containing (ﬁnal

concentrations): 2.5 mM EDTA, 0.05% Triton X-100,

100 lM ATP, and 1.25 mg/ml streptavidin coated SPA

beads (Amersham cat. No. RPNQ0007). Radioactivity

associated with the beads was quantiﬁed by scintillation

counting. We have also done cytotoxicity analysis of the

above-synthesized compounds, using neutral red uptake

by using Vero-C-1008 cell line¹⁸at various concentra-

tions (6.25–50 lg/mL), none of them were found toxic.

Hence the activities of the above-synthesized com-

pounds were not due to cytotoxicity of compounds.

Table 4. Selectivity ratio of most active compounds

Compound

cdk5 IC₅₀(nM)

62 11

cdk2 IC₅₀(nM)

Select k2/k5

1140 162

1340 104

890 78

18.3

624 62

468 43

6342 142

8.6

13.8

51 8

512 12

1.2

Attention was then turned to optimization of the 8a–d in

order to gain the selectivity over cdk2. On comparing,

8b aﬀorded improved cdk5 potency that is >17-fold

selectivity versus cdk2. The compound 8a was equally

selective versus cdk2 and had slightly improved cdk5

IC₅₀. Other derivatives were had noticeably decreased

cdk5 activity.

3. Conclusion

4.2. Preparation of N-{4-[(4-amino-5-sulfanyl-4H-1,2,4-

triazol-3-yl)methyl]-1,3-thiazol-2-yl}-2-chloroacetamide

(1a), N-{4-[(4-amino-5-sulfanyl-4H-1,2,4-triazol-3-yl)methyl]-

1,3-thiazol-2-yl}-acetamide (1b), N-{4-[(4-amino-5-sulfanyl-

4H-1,2,4-triazol-3-yl)methyl]-1,3-thiazol-2-yl}-benzamide

(1c), and 3-{N-(4-[(4-amino-5-sulfanyl-4H-1,2,4-triazol-

3-yl)methyl]-1,3-thiazol-2-yl}-amino)} propanoic acid (1d)

In conclusion, a novel series of clubbed triazolyl thiazole

derivatives that inhibit cdk5/p25 has been discovered. It

was found that the potency of the screening hit 1b could

be enhanced ﬁrst by structural transformation to a 2-po-

sition of thiazole core and amino and sulfhydryl groups

in triazole core and subsequently by the introduction of

appropriate substituents on both the heterocyclic rings

leading to the most promising compounds 8a and 8b.

Finally it can be concluded that an ideal cdk5/p25 inhib-

itor with minimal toxicity and potential activity can be

designed using above said compounds as lead molecules.

The said inhibitor can be synthesized using MAOS so as

to get the beneﬁts of this novel technique.

Above titled compounds were prepared as per the

literature.¹⁷

4.3. General preparation of N-[3-({2-[(substituted)amino]-

1,3-thiazol-4-yl}methyl)-5-sulfanyl-4H-1,2,4-triazol-4-yl]-

4-chlorobenzamide

The triazole (1) (1 mmol) in 20 mL of 10% NAOH was

treated dropwise with an equimolar amount of the

4-chlorobenzoyl chloride at 0 ꢁC, which was stirred for

30–45 min. At the end of stirring, precipitate was

observed. It was then ﬁltered, washed thoroughly with

water, and crystallized.

4. Experimental

4.1. General

The melting points were recorded on electrothermal

apparatus and are uncorrected. H NMR spectra on a

Bruker Avance 300 MHz instrument using CDCl₃as sol-

vent using TMS as internal standard; the chemical shifts

4.3.1. N-[3-({2-[(Chloroacetyl)amino]-1,3-thiazol-4-yl}meth-

yl)- 5-sulfanyl-4H-1,2,4-triazol-4-yl]-4-chlorobenzamide (2a).

Yield 71%; mp 241–243 ꢁC; ¹H NMR (300 MHz, CDCl₃):

2606

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

d 3.74 (s, 2H, CH₂), 4.13 (s, 2H, CH₂), 6.21 (s, 1H, Thiazole

CH), 7.12–7.36 (m, 4H, ArH), 8.06 (s, 2H, NH), 12.31 (s,

1H, SH); MS m/z (%) 443 (M⁺, 100), 407 (52), 323

(33.8), 309 (17.4), 248 (14.7), 247 (29.8), 233 (9.8), 220

(10.8), 180 (10.1), 095 (15.1), 86 (9.3); Anal. Calcd for

C₁₅H₁₂Cl₂N₆O₂S₂: C, 40.64; H, 2.73; N, 18.96. Found: C,

40.36; H, 2.48; N, 18.49.

(s, 4H, NH); MS m/z (%) 852 (M⁺, 7.1), 714 (43), 679

(27.5), 622 (5.5), 607 (100), 516 (3.4), 484 (4.7) 453

(8.2), 347 (9.6), 234 (10.3), 185 (13.8), 146 (8.7), 123

(13.2), 104 (10.5), 87 (26.8), 78 (40); Anal. Calcd for

C₃₁H₂₄Cl₄N₁₂O₄S₄: C, 41.43; H, 2.69; N, 18.70. Found:

C, 41.64; H, 2.81; N, 18.96.

4.4.2. N,N⁰-(Methylenebis{sulfanedial-[5-({2-[(acetyl)ami-

no]-1,3-thiazol-4-yl}methyl)-4H-1,2,4-triazole-3,4-di-

al]})di-4-chlorobenzamide (3b). Yield 77%; decomposes

around 262–264 ꢁC; ¹H NMR (300 MHz, CDCl₃): d

2.70 (s, 6H, CH₃), 4.34 (s, 4H, CH₂), 4.64 (s, 2H,

CH₂), 5.93 (s, 2H, thiazole CH), 7.21–7.55 (m, 8H,

ArH), 8.15 (s, 4H, NH); MS m/z (%) 784 (M⁺,35.9),

709 (11.1), 659 (13.2), 667 (23.6), 619 (100), 541 (3.6),

454 (3.7) 419 (9.8), 307 (8.2), 277 (8.1), 254 (11.9), 241

(15.4), 223 (35.8), 216 (24.9), 91 (23.8), 83 (54.2), 69

(25.7); Anal. Calcd for C₃₁H₂₆Cl₂N₁₂O₄S₄: C, 44.87;

H, 3.16; N, 20.26. Found: C, 44.62; H, 3.29; N, 20.04.

4.3.2. N-[3-({2-[(Acetyl)amino]-1,3-thiazol-4-yl}methyl)-

5-sulfanyl-4H-1,2,4-triazol-4-yl]-4-chlorobenzamide (2b).

Yield 77%; mp 250–252 ꢁC; ¹H NMR (300 MHz,

CDCl₃): d 2.12 (s, 3H, CH₃), 3.48 (s, 2H, CH₂), 4.27

(s, 4H, CH₂), 6.11 (s, 1H, Thiazole CH), 7.26–7.41 (m,

4H, ArH), 8.12 (s, 2H, NH), 12.53 (s, 1H, SH); MS

m/z (%) 409 (M⁺, 100), 376 (37), 312 (29.6), 251 (9.8),

223 (15.1), 211 (10.8), 107 (14.7), 087 (10.1), 82 (7.6);

Anal. Calcd for C₁₅H₁₃ClN₆O₂S₂: C, 44.06; H, 3.20;

N, 20.55. Found: C, 44.32; H, 3.51; N, 20.62.

4.3.3. N-[3-({2-[(Benzoyl)amino]-1,3-thiazol-4-yl}methyl)-

5-sulfanyl-4H-1,2,4-triazol-4-yl]-4-chlorobenzamide (2c).

Yield 76%; mp 280–282 ꢁC; ¹H NMR (300 MHz,

CDCl₃): d 3.32 (s, 2H, CH₂), 4.18 (s, 2H, CH₂), 6.17

(s, 1H, thiazole CH), 7.21–7.86 (m, 9H, ArH), 8.13 (s,

2H, NH), 12.13 (s, 1H, SH); MS m/z (%) 471 (M⁺,

80), 436 (21) 306 (28), 292 (8.4), 251 (20), 214 (100),

195 (15), 154 (7), 106 (65); Anal. Calcd for

C₂₀H₁₅ClN₆O₂S₂: C, 51.01; H, 3.21; N, 17.84. Found:

C, 51.17; H, 3.34; N, 17.54.

4.4.3. N,N⁰-(Methylenebis{sulfanedial-[5-({2-[(benzoyl)ami-

no]-1,3-thiazol-4-yl}methyl)-4H-1,2,4-triazole-3,4-dial]}) di-

4-chlorobenzamide (3c). Yield 79%; decomposes around

274–276 ꢁC; H NMR (300 MHz, CDCl₃): d 3.73 (s, 4H,

CH₂), 4.43 (s, 2H, CH₂), 6.27 (s, 2H, thiazole CH),

7.13–7.80 (m, 18H, ArH), 8.16 (s, 4H, NH); MS m/z (%)

908 (M⁺, 14.1), 886 (38), 724 (15.7), 616 (14.3), 601

(100), 542 (23.5), 465 (3.9) 421 (13.2), 312 (5.8), 279

(7.2), 263 (11.0), 257 (11.7), 256 (35.8), 216 (32.8), 91

(22), 83 (27.1), 69 (29.6); Anal. Calcd for

C₄₁H₃₀Cl₂N₁₂O₄S₄: C, 51.62; H, 3.17; N, 17.62. Found:

C, 51.48; H, 3.35; N, 17.82.

4.3.4. 3-{N-[4-[(4-(4-Chlorobenzoylamino)-5-sulfanyl-4H-

1,2,4-triazol-3-yl)methyl]-1,3-thiazol-2-yl}-aminopropa-

noic acid (2d). Yield 69%; mp 271–273 ꢁC; H NMR

(300 MHz, CDCl₃):

2.46–2.51 (t, 2H, CH₂,

4.4.4. 3-[4-(4-(4-Chlorobenzoylamino)-5-[([4-(4-chlorobenzo-

ylamino)-5-(2-[(2-carboxy-ethyl)amino]-1,3-thiazol-4-ylmeth-

yl)-4H-1,2,4-triazol-3-yl]sulfanylmethyl)sulfanyl]-4H-1,2,4-

J = 4.3 Hz), 3.33–3.39 (q, 2H, CH₂, J = 7.6 Hz), 4.01–

4.11 (t, 1H, NH, J = 8.1), 4.32 (s, 2H, CH₂), 6.27 (s,

1H, thiazole CH), 7.43–7.62 (m, 4H, ArH), 8.16 (s,

1H, NH), 10.43 (bs, 1H, OH), 12.43 (s, 1H, SH); MS

m/z (%) 439 (M⁺, 100), 389 (52), 323 (14), 309 (17.1),

280 (6), 134 (35.9); Anal. Calcd for C₁₆H₁₅ClN₆O₃S₂:

C, 43.78; H, 3.44; N, 19.15. Found: C, 43.93; H, 3.27;

N, 19.07.

triazol-3-ylmethyl)-1,3-thiazol-2-yl]aminopropanoic

acid

(3d). Yield 89%; decomposes around 255–257 ꢁC; ¹H

NMR (300 MHz, CDCl₃): d 2.21–2.30 (t, 4H, CH₂,

J = 4.5 Hz), 3.28–3.35 (q, 4H, CH₂, J = 7.4 Hz), 3.76 (s,

4H, CH₂), 4.01–4.12 (t, 2H, NH, J = 7.9 Hz), 4.63 (s,

2H, CH₂), 6.26 (s, 2H, thiazole CH), 7.32–7.76 (m, 8H,

ArH), 8.21 (s, 4H, NH), 10.65 (br s, 2H, OH); MS m/z

(%) 844 (M⁺, 13.6), 791 (100), 725 (40.9), 693 (6), 578

(7.3), 512 (4.1), 472 (13.6), 371 (5), 356 (3.4), 283 (13.7),

269 (6.4), 155 (14.4); Anal. Calcd for C₃₃H₃₀Cl₂N₁₂O₆S₄:

C, 44.54; H, 3.40; N, 18.89. Found: C, 44.71; H, 3.64; N,

18.69.

4.4. General preparation of N,N⁰-(methylenebis{sulfane-

dial-[5-({2-[(2-substituted) amino]-1,3-thiazol-4-yl}meth-

yl)-4H-1,2,4-triazole-3,4-dial]})-di-4-chlorobenzamide

The triazole (2) (1 mmol), diiodomethane (1.5 mmol),

and 5.6 g (1 mmol) potassium hydroxide were dissolved

in 20 mL of dichloromethane. To the said mixture acidic

alumina (20 g) was added. Dichloromethane was evapo-

rated in vacuo, and mixture was kept inside the alumina

bath and irradiated for 5–6 min at the power level of

300 W. The mixture was cooled. The solid thus separat-

ed was dissolved in hot ethanol and ﬁltered. After cool-

ing, the ﬁltrate gave the product as white.

4.5. General preparation of N-{3-({2-[(substituted)amino]-

1,3-thiazol-4-yl}methyl)-5-[(cyanomethyl)sulfanyl]-4H-

1,2,4-triazol-4-yl}-4-chlorobenzamide

The triazole (2) (1 mmol) was mixed with 1.2 mL

(2 mmol) of chloroacetonitrile and dissolved in 25 mL

of water. Neutralization with sodium carbonate gave a

precipitate, which was ﬁltered, washed with cold water

(2· 20 mL), and crystallized.

4.4.1. N,N⁰-(Methylenebis{sulfanedial-[5-({2-[(2-chloro-

acetyl)amino]-1,3-thiazol-4-yl}methyl)-4H-1,2,4-triazole-

3,4-dial]})di-4-chlorobenzamide (3a). Yield 84%; decom-

poses around 278–280 ꢁC; ¹H NMR (300 MHz, CDCl₃):

d 3.34 (s, 4H, CH₂), 4.19 (s, 4H, CH₂), 4.58 (s, 2H, CH₂),

6.27 (s, 2H, thiazole CH), 7.13–7.62 (m, 8H, ArH), 7.84

4.5.1. N-{3-({2-[(2-Chloroacetyl)amino]-1,3-thiazol-4-yl}-

methyl)-5-[(cyanomethyl) sulfanyl]-4H-1,2,4-triazol-4-yl}-

4-chlorobenzamide (4a). Yield 86%; mp 241–243 ꢁC; H

NMR (300 MHz, CDCl₃): d 3.74 (s, 2H, CH₂), 4.17 (s,

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

2607

2H, CH₂), 4.32 (s, 2H, CH₂), 6.26 (s, 2H, thiazole CH),

7.11–7.32 (m, 4H, ArH), 8.24 (s, 2H, NH); MS m/z (%)

482 (M⁺, 100), 428 (21.9), 369 (20.6), 272 (34.2), 270

(40.8), 256 (17.9), 83 (28.9), 69 (16.1), 55 (11.3); Anal.

Calcd for C₁₇H₁₃Cl₂N₇O₂S₂: C, 42.33; H, 2.72; N,

20.33. Found: C, 42.51; H, 2.97; N, 20.06.

2H, NH); MS m/z (%) 515 (M⁺, 100), 457 (14), 407

(12.3), 385 (11.3), 373 (7.2), 316 (7.7), 279 (79), 278

(10), 363 (8.2), 262 (19.5), 248 (7.7), 234 (7.9), 222

(10.5), 220 (5.7), 250 (31.6); Anal. Calcd for

C₁₈H₁₆Cl₂N₆O₄S₂: C, 41.95; H, 3.13; N, 16.31. Found:

C, 41.68; H, 3.42; N, 16.51.

4.5.2. N-{3-({2-[(Acetyl)amino]-1,3-thiazol-4-yl}methyl)-5-

[(cyanomethyl)sulfanyl]-4H-1,2,4-triazol-4-yl}-4-chloroben-

zamide (4b). Yield 82%; mp 264–266 ꢁC; ¹H NMR

(300 MHz, CDCl₃): d 2.31 (s, 3H, COCH₃), 3.62 (s, 2H,

CH₂), 4.04 (s, 2H, CH₂), 6.18 (s, 1H, Thiazole CH),

7.17–7.39 (m, 4H, ArH), 7.91 (s, 2H, NH); MS m/z (%)

448 (M⁺, 100), 407 (21.9), 369 (20.6), 272 (34.2), 270

(40.8), 256 (17.9), 83 (28.9), 69 (16.1), 55 (11.3); Anal.

Calcd for C₁₇H₁₄ClN₇O₂S₂: C, 45.58; H, 3.15; N, 21.89.

Found: C, 45.79; H, 3.36; N, 21.68.

4.6.2. Methyl{[4-(benzoylamino)-5-({2-[(acetyl)amino]-1,3-

thiazol-4-yl}methyl)-4H-1,2,4-triazol-3-yl]sulfanyl}acetate

(5b). Yield 78%; mp 261–263 ꢁC; H NMR (300 MHz,

CDCl₃): d 2.17 (s, 3H, OCH₃), 3.52 (s, 3H, COCH₃),

3.74 (s, 2H, CH₂), 3.92 (s, 2H, SCH₂), 6.46 (s, 1H, CH

of thiazole), 7.17–7.38 (m, 4H, ArH), 8.13 (br, 2H,

NH); MS m/z (%) 481 (M⁺, 9), 426 (31), 367 (1), 323

(2), 309 (1), 273 (100), 272 (8); Anal. Calcd for

C₁₈H₁₇ClN₆O₄S₂: C, 44.95; H, 3.56; N, 17.47. Found:

C, 44.75; H, 3.73; N, 17.69.

4.5.3. N-{3-({2-[(Benzoyl)amino]-1,3-thiazol-4-yl}methyl)-

5-[(cyanomethyl)sulfanyl]-4H-1,2,4-triazol-4-yl}-4-chlorob-

enzamide (4c). Yield 81%; mp 267–269 ꢁC; ¹H NMR

(300 MHz, CDCl₃): d 3.73 (s, 2H, CH₂), 4.14 (s, 2H,

CH₂), 6.24 (s, 1H, thiazole CH), 7.41–7.83 (m, 9H,

ArH), 8.24 (s, 2H, NH); MS m/z (%) 510 (M⁺, 93.3),

485 (10.9), 419 (4.1), 356 (31), 273 (46), 272 (100), 271

(14.3), 256 (9.3), 228 (4.4), 217 (3.2), 189 (3.6), 124

(8.9), 109 (5.8), 81 (4.5), 53 (3); Anal. Calcd for

C₂₂H₁₆ClN₇O₂S₂: C, 51.81; H, 3.16; N, 19.23. Found:

C, 51.68; H, 3.43; N, 19.52.

4.6.3. Methyl{[4-(benzoylamino)-5-({2-[(benzoyl)amino]-1,3-

thiazol-4-yl}methyl)-4H-1,2,4-triazol-3-yl]sulfanyl}acetate

(5c). Yield 75%; mp 226–228 ꢁC; H NMR (300 MHz,

CDCl₃): d 3.32 (s, 3H, OCH₃), 3.62 (s, 2H, CH₂), 3.83

(s, 2H, SCH₂), 6.25 (s, 1H, CH of thiazole), 6.93–7.51

(m, 9H, ArH), 8.22 (br, 2H, NH); MS m/z (%) 543

(M⁺, 69.9), 486 (54), 424 (43), 348 (100), 273 (39), 232

(15); Anal. Calcd for C₂₃H₁₉ClN₆O₄S₂: C, 50.87; H,

3.53; N, 15.48. Found: C, 50.56; H, 3.61; N, 15.73.

4.6.4. 3-{N-[4-(4-(4-Chlorobenzoylamino)-5-[(2-methoxy-2-

oxoethyl)sulfanyl]-4H-1,2,4-triazol-3-ylmethyl)-1,3-thia-

zol-2-yl]amino}propanoic acid (5d). Yield 79%; mp above

4.5.4. 3-{N-[4-(4-(4-Chlorobenzoylamino)-5-[(cyanometh-

yl)sulfanyl]-4H-1,2,4-triazol-3-ylmethyl)-1,3-thiazol-2-yl]-

amino}propanoic acid (4d). Yield 78%; mp 275–277 ꢁC;

¹H NMR (300 MHz, CDCl₃): d 2.43–2.48 (t, 2H, CH₂,

J = 4.1 Hz), 3.31–3.37 (q, 2H, CH₂, J = 7.3 Hz), 3.84

(s, 2H, CH₂), 4.11–4.18 (t, 1H, NH, J = 7.8 Hz), 4.34

(s, 2H, CH₂), 6.26 (s, 1H, thiazole CH), 7.22–7.43 (m,

4H, ArH), 8.25 (s, 1H, NH), 10.84 (br s, 1H, OH); MS

m/z (%) 478 (M⁺, 56.8), 417 (100) 376 (6.9), 349 (16),

348 (12.8), 347 (26.8), 337 (9.7), 331 (12.4), 323 (9.7),

256 (8.8); Anal. Calcd for C₁₈H₁₆ClN₇O₃S₂: C, 45.23;

H, 3.37; N, 20.51. Found: C, 45.37; H, 3.56; N, 20.71.

300 ꢁC; H NMR (300 MHz, CDCl₃): d 2.30–2.41 (q,

2H, CH₂, J = 4.0 Hz), 3.18–3.23 (t, 2H, CH₂,

J = 7.3 Hz), 3.46 (s, 3H, OCH₃), 3.61 (s, 2H, CH₂),

3.93 (s, 2H, SCH₂), 4.11–4.18 (t, 1H, NH, J = 7.9 Hz),

6.26 (s, 1H, CH of thiazole), 7.25–7.61 (m, 4H, ArH),

8.29 (br, 1H, NH), 11.13 (br, 1H, OH); MS m/z (%)

511 (M⁺, 100), 479 (67), 409 (41), 372 (58), 331 (16),

270 (60), 217 (44), 194 (27), 107 (48), 84 (81); Anal.

Calcd for C₁₉H₁₉ClN₆O₅S₂: C, 44.66; H, 3.75; N,

16.45. Found: C, 44.46; H, 3.59; N, 16.24.

4.7. General preparation of N-{3-({2-[(substituted)amino]-

1,3-thiazol-4-yl}methyl)-5-[(2-hydrazino-2-oxoethyl)sul-

fanyl]-4H-1,2,4-triazol-4-yl}-4-chlorobenzamide

4.6. General preparation of methyl{[4-(benzoylamino)-5-({2-

[(substituted) amino]-1,3-thiazol-4-yl}methyl)-4H-1,2,4-tria-

zol-3-yl]sulfanyl}acetate

A solution of (5) (1 mmol) with 5 mL (1 mmol) hydra-

zine hydrate (98%) was prepared in 10 mL ethanol. To

this acidic alumina (10 g) was added. Ethanol then was

evaporated in vacuo, and mixture was kept inside the

alumina bath and irradiated for 5–6 min at the power le-

vel of 300 W. The mixture was cooled and the product

was extracted with ether. Ether was distilled oﬀ and

product thus obtained was crystallized from n-hexane–

carbon tetrachloride mixture.

A solution of triazole (2) (1 mmol), 0.4 g (1 mmol) of

sodium hydroxide, and methyl bromoacetate 1.53 g

(1 mmol) was prepared. To this, acidic alumina was add-

ed in 1:5 equivalent of triazole. The reaction mixture

was mixed, and mixture was kept inside the alumina

bath and irradiated for 4–5 min at the power level of

300 W. The mixture was cooled and poured on ice.

The solid thus separated was extracted with hot ethanol,

ﬁltered. After cooling, ﬁltrate gave almost pure product.

4.7.1.

N-{3-({2-[(2-Chloroacetyl)amino]-1,3-thiazol-4-yl}-

4.6.1. Methyl{[4-(benzoylamino)-5-({2-[(2-chloroacetyl)ami-

no]-1,3-thiazol-4-yl} methyl)-4H-1,2,4-triazol-3-yl]sulfa-

nyl}acetate (5a). Yield 82%; mp 259–251 ꢁC; H NMR

(300 MHz, CDCl₃): d 2.11 (s, 3H, OCH₃), 3.81 (s, 2H,

CH₂), 3.89 (s, 2H, SCH₂), 4.28 (s, 2H, CH₂Cl), 6.32 (s,

1H, CH of thiazole), 7.20–7.46 (m, 4H, ArH), 8.32 (br,

methyl)-5-[(2-hydrazino-2-oxoethyl)sulfanyl]-4H-1,2,4-tria-

zol-4-yl}-4-chlorobenzamide (6a). Yield 83%; mp 245–

247 ꢁC; ¹H NMR (300 MHz, CDCl₃): d 2.13 (d, 2H,

NH₂, J = 6.5 Hz), 3.64 (s, 2H, CH₂Cl), 3.94 (s, 2H,

SCH₂), 4.07–4.15 (t, 1H, NH, J = 4.3 Hz), 6.65 (s, 1H,

CH of thiazole), 7.22–7.48 (m, 4H, ArH), 8.21 (br, 2H,

2608

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

NH); MS m/z (%) 515 (M⁺, 65), 479 (69), 436 (145), 386

(61), 328 (78), 311 (8.4), 269 (24), 235 (100), 201 (13), 184

(18), 156 (53), 124 (25), 89 (49); Anal. Calcd for

C₁₇H₁₆Cl₂N₈O₃S₂: C, 39.62; H, 3.13; N, 21.74. Found:

C, 39.82; H, 3.34; N, 21.53.

4.8.1. N-[3-{[2-(2-Acetylhydrazino)-2-oxoethyl]sulfanyl}-

5-({2-[(acetyl)amino]-1,3-thiazol-4-yl}methyl)-4H-1,2,4-

triazol-4-yl]-4-chlorobenzamide (7b). Yield 71%; mp 227–

229 ꢁC; ¹H NMR (300 MHz, CDCl₃): d 2.42 (s, 6H,

CH₃), 3.74 (s, 2H, CH₂), 3.87 (s, 2H, SCH₂), 4.20–4.28

(dd, 2H, J_NH–NH= 4.35, J_NH–NH= 4.76), 6.27 (s, 1H,

thiazole CH), 7.11–7.32 (m, 4H, ArH), 8.18 (s, 2H,

NH); MS m/z (%) 523 (M⁺, 97), 488 (62), 437 (17.2),

389 (9.1), 327 (74), 297 (54), 241 (8.3), 223 (100), 174

(24), 146 (27); Anal. Calcd for C₁₉H₁₉ClN₈O₄S₂: C,

43.63; H, 3.66; N, 21.43. Found: C, 43.42; H, 3.89; N,

21.51.

4.7.2. N-{3-({2-[(Acetyl)amino]-1,3-thiazol-4-yl}methyl)-

5-[(2-hydrazino-2-oxoethyl) sulfanyl]-4H-1,2,4-triazol-4-

yl}-4-chlorobenzamide (6b). Yield 82%; mp 250–252 ꢁC;

¹H NMR (300 MHz, CDCl₃): d 2.16 (d, 2H, NH₂,

J = 6.5 Hz), 2.32 (s, 3H, CH₃), 3.67 (s, 2H, CH₂), 3.81

(s, 2H, SCH₂), 4.15–4.31 (t, 1H, NH, J = 4.5 Hz), 6.69

(s, 1H, CH of thiazole), 7.45–7.71 (m, 4H, ArH), 8.12

(br, 2H, NH); MS m/z (%) 481 (M⁺, 78), 418 (72), 371

(26.3), 347 (6.3), 265 (18.3), 224 (65.3), 186 (25), 113

(100), 89 (14.3); Anal. Calcd for C₁₇H₁₇ClN₈O₃S₂: C,

42.45; H, 3.56; N, 23.30. Found: C, 42.68; H, 3.36; N,

23.27.

4.8.2.

N-[3-{[2-(2-Benzoylhydrazino)-2-oxoethyl]sulfa-

nyl}-5-({2-[(acetyl)amino]-1,3-thiazol-4-yl}methyl)-4H-

1,2,4-triazol-4-yl]-4-chlorobenzamide (7f). Yield 74%; mp

286–288 ꢁC; ¹H NMR (300 MHz, CDCl₃): d 2.38 (s, 3H,

CH₃), 3.68 (s, 2H, CH₂), 3.71 (s, 2H, SCH₂), 4.13–4.21

(dd, 2H, J_NH–NH= 4.23, J_NH–NH= 4.54), 6.12 (s, 1H,

thiazole CH), 6.94–7.21 (m, 9H, ArH), 8.09 (s, 2H,

NH); MS m/z (%) 584 (M⁺, 86), 526 (71), 481 (14),

427 (78), 379 (38), 325 (41), 287 (17), 241 (35), 167

(100), 109 (37), 98 (19); Anal. Calcd for

C₂₄H₂₁ClN₈O₄S₂: C, 49.27; H, 3.62; N, 19.15. Found:

C, 49.51; H, 3.44; N, 19.34.

4.7.3. N-{3-({2-[(Benzoyl)amino]-1,3-thiazol-4-yl}meth-

yl)-5-[(2-hydrazino-2-oxoethyl) sulfanyl]-4H-1,2,4-tria-

zol-4-yl}-4-chlorobenzamide (6c). Yield 71%; mp 222–

224 ꢁC; H NMR (300 MHz, CDCl₃): d 2.11 (d, 2H,

NH₂, J = 6.5 Hz), 3.38 (s, 2H, CH₂), 3.76 (s, 2H,

SCH₂), 4.12–4.39 (t, 1H, NH, J = 4.1 Hz), 6.11 (s, 1H,

CH of thiazole), 7.13–7.68 (m, 9H, ArH), 8.10 (br, 2H,

NH); MS m/z (%) 543 (M⁺, 89), 486 (31), 421 (60),

378 (14.3), 352 (45), 305 (24), 241 (73), 208 (56), 174

(66), 146 (100), 109 (18), 88 (15); Anal. Calcd for

C₂₂H₁₉ClN₈O₃S₂: C, 48.66; H, 3.53; N, 20.64. Found:

C, 48.71; H, 3.78; N, 20.53.

4.8.3. N-[3-{[2-(2-Chloroacetylhydrazino)-2-oxoethyl]sul-

fanyl}-5-({2-[(acetyl)amino]-1,3-thiazol-4-yl}methyl)-4H-

1,2,4-triazol-4-yl]-4-chlorobenzamide (7j). Yield 57%; mp

166–168 ꢁC; ¹H NMR (300 MHz, CDCl₃): d 2.53 (s, 3H,

CH₃), 3.81 (s, 2H, CH₂), 3.92 (s, 2H, SCH₂), 4.17 (s, 2H,

CH₂Cl), 4.28-4.34 (dd, 2H, J_NH–NH= 4.52, J_NH–

_NH= 4.92), 6.41 (s, 1H, thiazole CH), 7.26–7.46 (m,

4H, ArH), 8.32 (s, 2H, NH); MS m/z (%) 557 (M⁺,

54), 507 (36), 467 (84), 419 (54.2), 384 (9.3), 338 (100),

258 (12), 228 (37), 177 (32); Anal. Calcd for

C₁₉H₁₈Cl₂N₈O₄S₂: C, 40.94; H, 3.25; N, 20.10. Found:

C, 40.76; H, 3.42; N, 20.33.

4.7.4. 3-{N-[4-(4-(4-Chlorobenzoylamino)-5-[(2-hydrazi-

no-2-oxoethyl)sulfanyl]-4H-1,2,4-triazol-3-ylmethyl)-1,3-

thiazol-2-yl]amino}propanoic acid (6d). Yield 84%; mp

243–245 ꢁC; ¹H NMR (300 MHz, CDCl₃): d 2.13 (d,

2H, NH₂, J = 6.1 Hz), 2.27–2.31 (q, 2H, CH₂,

J = 4.2 Hz), 3.16–3.27 (t, 2H, CH₂, J = 7.1 Hz), 3.46 (s,

2H, CH₂), 3.84 (s, 2H, SCH₂), 4.23–4.45 (t, 2H, NH,

J = 4.1 Hz, J = 8.1 Hz), 6.14 (s, 1H, CH of thiazole),

7.34–7.61 (m, 4H, ArH), 8.03 (br, 1H, NH), 11.09 (br,

1H, OH); MS m/z (%) 511 (M⁺, 93.3), 485 (32), 419

(22), 394 (56.4), 316 (28), 247 (64), 217 (100), 147 (83),

108 (71), 79 (10.2); Anal. Calcd for C₁₈H₁₉ClN₈O₄S₂:

C, 42.31; H, 3.75; N, 21.93. Found: C, 42.57; H, 3.63;

N, 21.82.

4.9. General procedure for N-[3-({2-[(2E)-2-benzyli-

denehydrazino]-2-oxoethyl} sulfanyl)-5-({2-[(substituted)ami-

no]-1,3-thiazol-4-yl}methyl)-4H-1,2,4-triazol-4-yl]-4-

chlorobenzamide

A solution of (6) (1 mmol) with benzaldehyde (1 mmol)

was prepared in 10 mL ethanol. To this acidic alumina

(10 g) was added. Ethanol then was evaporated in vac-

uo, and mixture was kept inside the alumina bath and

irradiated for 1 min at the power level of 300 W. The

mixture was cooled and poured on ice. The solid thus

separated was ﬁltered and extracted with ether. Ether

was distilled oﬀ and product thus obtained was crystal-

lized from hot ethanol.

4.8. General preparation of N-[3-{[2-(substituted-hydra-

zino)-2-oxoethyl]sulfanyl}-5-({2-[(substituted)amino]-

1,3-thiazol-4-yl}methyl)-4H-1,2,4-triazol-4-yl]-4-chloro

benzamide

To a solution of (6) (1 mmol) in dichloromethane (ex-

cess amount), appropriate acid chloride (1 mmol) was

added dropwise with constant vigorous stirring. After

25 min of stirring, acidic alumina (10 g) was added.

Dichloromethane then was evaporated in vacuo, and

mixture was kept inside the alumina bath and irradiat-

ed for 5–6 min at the power level of 300 W. The mix-

ture was cooled and the product was extracted with

ether. Ether was distilled oﬀ and product thus obtained

was crystallized from n-hexane–carbon tetrachloride

mixture.

4.9.1. N-[3-({2-[(2E)-2-Benzylidenehydrazino]-2-oxoeth-

yl}sulfanyl)-5-({2-[(2-chloroacetyl)amino]-1,3-thiazol-4-

yl}methyl)-4H-1,2,4-triazol-4-yl]-4-chlorobenzamide (8a).

Yield 81%; decomposed around 222–224 ꢁC; H NMR

(300 MHz, CDCl₃): d 3.77 (s, 2H, CH₂), 4.16 (s, 2H,

SCH₂), 4.13 (s, 2H, CH₂Cl), 6.22 (s, 1H, thiazole CH),

7.30–7.62 (m, 9H, ArH), 8.16 (S, 3H, NH), 8.27 (S,

1H, N@CH); MS m/z (%) 603 (M⁺, 69), 562 (19), 517

(37), 479 (21), 415 (58), 346 (24), 295 (35), 234 (100),

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

2609

157

(11.4),

103

(12.3);

Anal.

Calcd

for

The solid thus separated was ﬁltered, extracted with

ether. Ether was distilled oﬀ and product thus obtained

was crystallized from hot ethanol.

C₂₄H₂₀Cl₂N₈O₃S₂: C, 47.76; H, 3.34; N, 18.57. Found:

C, 47.63; H, 3.57; N, 18.74.

4.9.2. N-[3-({2-[(2E)-2-Benzylidenehydrazino]-2-oxoeth-

yl}sulfanyl)-5-({2-[(acetyl) amino]-1,3-thiazol-4-yl}meth-

yl)-4H-1,2,4-triazol-4-yl]-4-chlorobenzamide (8b). Yield

83%; mp 178–180 ꢁC; H NMR (300 MHz, CDCl₃): d

4.10.1. N-[3-{[(4-Amino-5-thioxo-4,5-dihydro-1H-1,2,4-tria-

zol-3-yl)methyl]sulfanyl}-5-({2-[(2-chloroacetyl)amino]-1,3-

thiazol-4-yl}methyl)-4H-1,2,4-triazol-4-yl]-4-chlorobenzamide

(9a). Yield 72%; mp 228–230 ꢁC; H NMR (300 MHz,

2.32 (s, 3H, CH₃), 3.97 (s, 2H, CH₂), 4.21 (s, 2H,

SCH₂), 6.25 (s, 1H, thiazole CH), 7.31–7.65 (m, 9H,

ArH), 8.26 (s, 3H, NH), 8.36 (s, 1H, N@CH); MS m/z

(%) 569 (M⁺, 94), 519 (41), 487 (9.6), 431 (26), 413

(8.4), 389 (100), 365 (20); Anal. Calcd for

C₂₄H₂₁ClN₈O₃S₂: C, 50.66; H, 3.72; N, 19.69. Found:

C, 50.54; H, 3.88; N, 19.50.

CDCl₃): d 2.16 (s, 2H, NH₂), 3.76 (s, 2H, CH₂), 4.07 (s,

2H, CH₂), 4.25 (s, 2H, CH₂Cl), 6.20 (s, 1H, thiazole CH),

7.23–7.67 (m, 4H, ArH), 8.11 (s, 2H, NH), 12.49 (s, 1H,

SH); MS m/z (%) 571 (M⁺, 82), 517 (31), 461 (26), 384

(31), 326 (13.2), 247 (15), 226 (17), 125 (100); Anal. Calcd

for C₁₈H₁₆Cl₂N₁₀O₂S₃: C, 37.83; H, 2.82; N, 24.51. Found:

C, 37.65; H, 2.59; N, 24.74.

4.9.3. N-[3-({2-[(2E)-2-Benzylidenehydrazino]-2-oxoeth-

yl}sulfanyl)-5-({2-[(benzoyl) amino]-1,3-thiazol-4-

yl}methyl)-4H-1,2,4-triazol-4-yl]-4-chlorobenzamide (8c).

Yield 81%; decomposed around 217–219 ꢁC; H NMR

(300 MHz, CDCl₃): d 3.63 (s, 2H, CH₂), 4.17 (s, 2H,

SCH₂), 6.23 (s, 1H, thiazole CH), 7.21–7.64 (m, 14H,

ArH), 8.16 (s, 3H, NH), 8.41 (s, 1H, N@CH); MS m/z

(%) 631 (M⁺, 79), 571 (25), 515 (52), 463 (63), 418

(9.2), 358 (41), 314 (31), 272 (37), 208 (100), 168

(15.2), 107 (28), 97 (7.4); Anal. Calcd for

C₂₉H₂₃ClN₈O₃S₂: C, 55.19; H, 3.67; N, 17.75. Found:

C, 55.34; H, 3.88; N, 17.53.

4.10.2. N-[3-{[(4-Amino-5-thioxo-4,5-dihydro-1H-1,2,4-

triazol-3-yl)methyl]sulfanyl}-5-({2-[(acetyl)amino]-1,3-

thiazol-4-yl}methyl)-4H-1,2,4-triazol-4-yl]-4-chlorobenz-

amide (9b). Yield 71%; mp 252–254 ꢁC; ¹H NMR

(300 MHz, CDCl₃): d 2.13 (s, 2H, NH₂), 2.35 (s, 3H,

CH₃), 3.79 (s, 2H, CH₂), 4.14 (s, 2H, CH₂), 6.19 (s,

1H, thiazole CH), 7.11–7.35 (m, 4H, ArH), 8.08 (s,

2H, NH), 12.43 (s, 1H, SH); MS m/z (%) 537 (M⁺,

76), 474 (32), 419 (30), 385 (100), 323 (2.93), 272

(12.26), 220 (3.73), 207 (5.86), 192 (7.07); Anal. Calcd

for C₁₈H₁₇ClN₁₀O₂S₃: C, 40.26; H, 3.19; N, 26.08.

Found: C, 40.53; H, 3.46; N, 26.34.

4.9.4. 3-{N-[4-(4-(4-Chlorobenzoylamino)-5-[(2-oxo-2-2-

[(Z)-1-phenylmethylidene] hydrazinoethyl)sulfanyl]-4H-1,2,

4-triazol-3-ylmethyl)-1,3-thiazol-2-yl]amino} propanoic acid

(8d). Yield 78%; mp 120–122 ꢁC; H NMR (300 MHz,

CDCl₃): d 2.65–2.71 (t, 2H, CH₂, J = 4.7 Hz), 3.23–3.31

(q, 2H, CH₂, J = 7.5 Hz), 3.92 (s, 2H, CH₂), 4.17 (s, 2H,

SCH₂), 4.32 (t, 1H, NH, J = 8.3 Hz), 6.23 (s, 1H, Thiazole

CH), 7.17–7.63 (m, 9H, ArH), 8.15 (s, 2H, NH), 8.32 (s,

1H, N@CH), 10.87 (bs, 1H, OH); MS m/z (%) 599 (M⁺,

74), 537 (45), 486 (13), 464 (63), 376 (32), 318 (65), 285

(12), 246 (100), 209 (37), 187 (42); Anal. Calcd for

C₂₅H₂₃ClN₈O₄S₂: C, 50.12; H, 3.87; N, 18.70. Found: C,

50.36; H, 3.69; N, 18.49.

4.10.3. N-[3-{[(4-Amino-5-thioxo-4,5-dihydro-1H-1,2,4-

triazol-3-yl)methyl]sulfanyl}-5-({2-[(benzoyl)amino]-1,3-

thiazol-4-yl}methyl)-4H-1,2,4-triazol-4-yl]-4-chloroben-

zamide (9c). Yield 80%; mp 264–266 ꢁC; ¹H NMR

(300 MHz, CDCl₃): d 2.41 (s, 2H, NH₂), 3.75 (s, 2H,

CH₂), 4.34 (s, 2H, CH₂), 6.23 (s, 1H, thiazole CH),

7.14–7.74 (m, 9H, ArH), 8.17 (s, 2H, NH), 12.63 (s,

1H, SH); MS m/z (%) 599 (M⁺, 93.3), 513 (24), 476

(21), 432 (16), 421 (12), 323 (65), 308 (41), 289 (27),

230 (38), 207 (100), 142 (35.7), 109 (23); Anal. Calcd

for C₂₃H₁₉ClN₁₀O₂S₃: C, 46.11; H, 3.20; N, 23.38.

Found: C, 46.34; H, 3.46; N, 23.51.

4.10.4. 3-[(4-[5-[(4-Amino-5-sulfanyl-4H-1,2,4-triazol-3-

yl)methyl]sulfanyl-4-(4-chlorobenzoylamino)-4H-1,2,4-

triazol-3-yl]methyl-1,3-thiazol-2-yl)amino]propanoic acid

(9d). Yield 77%; mp 237–239 ꢁC; H NMR (300 MHz,

CDCl₃): d 2.13 (s, 2H, NH₂), 2.46–2.49 (t, 2H, CH₂,

J = 4.3 Hz), 3.14–3.20 (q, 2H, CH₂, J = 6.7 Hz), 3.98

(s, 2H, CH₂), 4.23–4.29 (t, 1H, NH, J = 7.3 Hz), 4.36

(s, 2H, CH₂), 6.26 (s, 1H, thiazole CH), 7.27–7.56 (m,

4H, ArH), 8.17 (s, 1H, NH), 10.64 (br s, 1H, OH),

12.35 (s, 1H, SH); MS m/z (%) 567 (M⁺, 84), 497 (23),

450 (47), 438 (38), 371 (25), 370 (75), 354 (10), 235

(100), 220 (22), 207 (68), 192 (70); Anal. Calcd for

C₁₉H₁₉ClN₁₀O₃S₃: C, 40.24; H, 3.38; N, 24.70. Found:

C, 40.41; H, 3.63; N, 24.84.

4.10. General preparation of N-[3-{[(4-amino-5-thioxo-4,5-

dihydro-1H-1,2,4-triazol-3-yl)methyl]sulfanyl}-5-({2-

[(substituted)amino]-1,3-thiazol-4-yl}methyl)-4H-1,2,4-

triazol-4-yl]-4-chlorobenzamide

The (6) (1 mmol) was dissolved in alcoholic potassium

hydroxide (1 mmol) and kept for stirring. Carbon disul-

ﬁde (1.5 mmol) was added dropwise to the solution with

stirring. Thick solid mass was obtained, to which 50 mL

of absolute alcohol was added. Stirring was continued

for 16 h. At the end of 16th hour, dry ether was added

to the mixture. The precipitate (thiocarbazate) obtained

was taken immediately for the next step.

A solution of thiocarbazate (1 mmol) with hydrazine hy-

drate (1 mmol) was prepared in 10 mL ethanol. To this

acidic alumina (10 g) was added. Ethanol then was evap-

orated in vacuo, and mixture was kept inside the alumi-

na bath and irradiated for 5–6 min at the power level of

300 W. The mixture was cooled and poured on ice.

Acknowledgments

Authors are thankful to Dr. K. G. Bothara, Principal,

AISSMS College of Pharmacy, Pune, India, for provid-

ing the facilities.

2610

M. R. Shiradkar et al. / Bioorg. Med. Chem. 15 (2007) 2601–2610

Ahmed, S. Z.; Qian, L.; Chen, B. C.; Zhao, R.; Bednarz,

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