Novel C-aryl glucoside SGLT2 inhibitors as potential antidiabetic agents: 1,3,4-Thiadiazolylmethylphenyl glucoside congeners

Bioorganic & Medicinal Chemistry 18 (2010) 2178–2194

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Bioorganic & Medicinal Chemistry

journal homepage: w ww.elsevier.com/locate/bmc

Novel C-aryl glucoside SGLT2 inhibitors as potential antidiabetic agents:

1,3,4-Thiadiazolylmethylphenyl glucoside congeners

Junwon Lee ^a, Sung-Han Lee ^a, Hee Jeong Seo ^a, Eun-Jung Son ^a, Suk Ho Lee ^a, Myung Eun Jung ^a,

a,

MinWoo Lee ^a, Ho-Kyun Han ^a, Jeongmin Kim ^a, Jahyo Kang ^b,, Jinhwa Lee

*

^aCentral Research Laboratories, Green Cross Corporation, 303 Bojeong-Dong, Giheung-Gu, Yongin, Gyeonggi-Do 446-770, Republic of Korea

^bDepartment of Chemistry, Sogang University, 1 Shinsu-Dong, Mapo-Gu, Seoul 121-742, Republic of Korea

a r t i c l e i n f o

a b s t r a c t

Article history:

Novel C-aryl glucoside SGLT2 inhibitors containing 1,3,4-thiadiazole moieties were designed and synthe-

sized. Among the compounds tested, biaryl-type compounds containing pyrazine 59, 2-furan 61, and 3-

thiophene 71 showed the best in vitro inhibitory activities to date (IC₅₀= 3.51–7.03 nM) against SGLT2. A

selected compound 61, demonstrated reasonable blood glucose-lowering effects, indicating that the

information obtained from the SAR studies in this 1,3,4-thiadiazolylmethylphenyl glucoside series might

help to design more active SGLT2 inhibitors that are structurally related.

Received 14 December 2009

Revised 22 January 2010

Accepted 23 January 2010

Available online 4 February 2010

Keywords:

SGLT

Dapagliﬂozin

Thiadiazole

Glucoside

Diabetes

1. Introduction

sively at the apical domain of the epithelial cells in the early prox-

imal convoluted tubule. It is estimated that 90% of renal glucose

Diabetes has become an increasing concern to the world’s pop-

ulation. In 2007, approximately 246 million people were consid-

ered diabetic, with an additional 7 million people diagnosed with

the disease every year.¹Diabetes is a chronic metabolic disorder

that is deﬁned by the body’s inability to generate insulin or to re-

spond adequately to circulating insulin. There are two identiﬁed

forms of diabetes: type 1 diabetes is distinguished as an autoim-

mune disease involving pancreatic b-cells, while type 2 diabetes

is deﬁned by b-cell dysfunction and insulin resistance.²Type 2 dia-

betes is the most common disorder of glucose homeostasis,

accounting for nearly 90–95% of all cases of diabetes.

Sodium-dependent glucose cotransporters (SGLTs) couple the

transport of glucose against a concentration gradient with the

simultaneous transport of Na⁺down a concentration gradient.³

Two important SGLT isoforms have been cloned and identiﬁed as

SGLT1 and SGLT2.⁴SGLT1 is located in the gut, kidney, and heart

where its expression regulates cardiac glucose transport.⁵SGLT1

is a high-afﬁnity, low-capacity transporter and therefore accounts

for only a small fraction of renal glucose reabsorption.⁶In contrast,

SGLT2 is a low-afﬁnity, high-capacity transporter located exclu-

reabsorption is facilitated by SGLT2; the remaining 10% is likely

mediated by SGLT1 in the late proximal straight tubule.⁷Since

SGLT2 appears to be responsible for the majority of renal glucose

reabsorption based on human mutation studies,⁸it has become a

target of therapeutic interest. Phlorizin 1, which was isolated form

the root bark of the apple tree, demonstrated antidiabetic activity,

lowering plasma glucose and improving insulin resistance by

increasing renal glucose excretion. However, phlorizin was not

developed as a drug for the treatment of diabetes because the com-

pound was found to be metabolically unstable. This instability is

due to b-glucosidase cleavage in the intestinal tract and prevented

oral administration (Fig. 1). In addition, enzymatic release of the

aglycone phloretin 2, a micromolar inhibitor of sodium-indepen-

dent facilitative glucose transporters (GLUTs), could potentially in-

hibit GLUT-mediated cellular uptake of glucose.^9,15

Subsequently, T-1095 3, by Tanabe Seiyaku, was reported as the

ﬁrst orally absorbable SGLT2 inhibitor, overcoming the disadvan-

tage of phlorizin 1 (Fig. 2).¹⁰3 was absorbed in the intestine and

converted to an active form, T-1095A 4. Following the discovery

of 3, O-glucosides such as sergliﬂozin 5 and remogliﬂozin 7 ad-

vanced furthest in clinical trials. Compound 5 was disclosed by Kis-

sei in 2003 as a potential treatment for diabetes and obesity.¹¹In

2007, Kissei, in collaboration with GlaxoSmithKline, initiated the

development of 7, a pyrazole-based O-glucoside.¹²Again, concern

regarding gut b-glucosidase-mediated degradation, resulted in

* Corresponding authors. Tel.: +82 31 260 9892; fax: +82 31 260 9020 (J.L.), tel.:

+82 2 705 8439; fax: +82 2 701 0967 (J.K.).

E-mail addresses: kangj@sogang.ac.kr (J. Kang), jinhwalee@greencross.com (J.

Lee).

doi:10.1016/j.bmc.2010.01.073

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

2179

HO

O

OH

HO

+

OH

O

OH

O

β-glucosidase

HO

Glucose

OH

1

2

Figure 1. Structure and b-glucosidase-mediated cleavage of phlorizin (1).

Me

O

OH

O

RO

HO

OH

3, T-1095 (R = CO₂Me)

4, T-1095A (R = H)

OH

Me

OMe

O

Me

N

O

RO

HO

RO

HO

OH

5, sergliflozin (R = CO₂Et)

6, sergliflozin A (R = H)

7, remogliflozin (R = CO₂Et)

8, remogliflozin A (R = H)

Cl

OEt

Me

O

HO

F

S

HO

OH

10, canagliflozin

9, dapagliflozin

Figure 2. Structures of SGLT2 inhibitors.

developing sergliﬂozin A 6 and remogliﬂozin A 8 being adminis-

tered as the ethyl carbonate prodrugs 5 and 7, respectively.

Subsequent endeavors to identify SGLT2 inhibitors suitable for

oral administration without the need for a prodrug led to the dis-

covery of C-glucoside-derived SGLT2 inhibitors. C-glucosides ap-

pear to have drug-like properties with enhanced chemical

stability of the glucosidic bond. Extensive SAR studies by Bristol–

Myers Squibb identiﬁed dapagliﬂozin 9, a potent, selective SGLT2

inhibitor for the treatment of type 2 diabetes.^13–15At present,

dapagliﬂozin is the most advanced SGLT2 inhibitor in clinical trials

and is believed to be the ﬁrst SGLT2 inhibitor to go to market.¹⁸On

the other hand, Mitsubishi Tanabe Pharma, in collaboration with

Johnson & Johnson, is developing canagliﬂozin 10, another novel

C-glucoside-derived SGLT2 inhibitor.¹⁶In August 2009, a phase 3

study was reportedly initiated to evaluate the safety and efﬁcacy

of 10 in patients with type 2 diabetes.¹⁷

In the present study, metabolically more stable C-glucosides

bearing a heteroaromatic ring were exploited in order to develop

novel SGLT2 targeting antidiabetic agents. We envisioned that

replacement of the distal ring of dapagliﬂozin 9 with a heterocyclic

ring would be a worthy approach for the improvement of the par-

tition coefﬁcient (log P) value, to potentially decrease plasma pro-

tein binding. For this purpose, the structure of dapagliﬂozin 9 was

modiﬁed into 11, bearing a heterocyclic ring as shown in Figure 3.

Among a variety of heterocycles, we decided to screen thiadiazole

13 as our initial efforts. Herein, we report the design, synthesis and

biological evaluation of 1,3,4-thiadiazolylmethylphenyl glucoside

congeners.

2. Results and discussion

2.1. Chemistry

Synthesis of the target compounds commenced with the prepa-

ration of the lactone 15. Commercially available perbenzylated

gluconolactol 14²⁰was treated with TPAP (tetrapropylammonium

perruthenate) in the presence of NMO (4-methylmorpholine N-

oxide) to provide the requisite perbenzylated gluconolactone 15

in high yield as shown in Scheme 1.

As shown in Scheme 2, the reduction of commercially available

5-bromo-2-chlorobenzoic acid (16) with a borane-dimethyl sulﬁde

complex, and subsequent silylation of the corresponding alcohol

with TIPSCl (triisopropylsilyl chloride) in the presence of imidazole

and DMAP (4-(dimethylamino)pyridine) generated bromide 17 in

95% yield over two steps. Lithium–halogen exchange, followed by

the addition of the nascent lithiated aromatic compound to perb-

enzylated gluconolactone 15, produced a mixture of the corre-

sponding lactols. The lactols were reduced using triethylsilane

and BF₃etherate,¹⁹desilylated and afforded alcohol 18 in 98% yield

over the three steps.

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J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

Cl OEt

R¹

O

Het

O

HO

R²

HO

OH

9

11

R¹

N

R²

O

S

HO

OH

13

OH

Figure 3. Exploration of C-glucoside bearing heteroaryl group at the distal ring.

using hydrazine in the presence of EDCI, HOBt, and DMF in 95%

yield. This hydrazide 26 was coupled with commercially available

5-phenylfuran-2-carboxylic acid (27) in the presence of EDCI,

HOBt, and DMF to produce the corresponding acylhydrazide. Sub-

sequent cyclization using Lawesson’s reagent followed by depro-

tection of the benzyl groups using TMSI yielded the target

compound 65 in 36% yield over three steps as shown in Scheme 6.

The third thiadiazole-containing compound was synthesized as

in Scheme 6. Thus, acid 21 was coupled and concomitantly cyclized

with thiosemicarbazide using phosphorus (III) oxychloride to pro-

vide the corresponding aminothiadiazole. The aminothiadiazole

was converted to chlorothiadiazole 29 using sodium nitrite and a

catalytic amount of copper in the presence of concentrated HCl

and acetic acid in 22% overall yield.²²Treatment of chlorothiadiaz-

ole 29 with NaSMe and subsequent deprotection of the benzyl

groups yielded the target methylthio-thiadiazole 47 in 25% yield

over two steps.

The fourth target compound was prepared as in Scheme 7. Typ-

ical coupling of 21 with the hydrazide 30 and the following cycli-

zation produced the corresponding acylhydrazide, which was

treated with TMSOTf (trimethylsilyl triﬂuoromethanesulfonate)

and acetic anhydride to provide the corresponding tetraacetate.²³

The peracetylated compound was hydrolyzed to yield the target

compound 59 in 10% yield over four steps. This alternative depro-

tection method was frequently employed, especially in the case of

unsatisfactory reactions with TMSI.

O

OH

O

BnO

NMO, TPAP, 4Å MS

OBn

CH₂Cl₂, rt

(86%)

OBn

15

14

Scheme 1. Oxidation of lactol 14 to lactone 15.

The preparation of a key intermediate 21 is described in

Scheme 3. Thus, alcohol 18 was converted to bromide using PBr₃

in the presence of pyridine. The bromide was then treated with

KCN in reﬂuxing aqueous EtOH to generate cyanide 19 in 80% yield

over two steps. A mixture of the two isomers 19 was separated

through recrystallization from ethanol to produce the required

beta-isomer 20 in about 40% yield. Hydrolysis of 20 with sodium

hydroxide in aqueous ethanol generated the carboxylic acid 21 in

quantitative yield. Treatment of 21 with thionyl chloride in reﬂuxing

methanol produced the corresponding methyl ester 22 in 89% yield.

A thiazole-containing C-glucoside 61 was synthesized as shown

in Scheme 4. Coupling of acid 21 and a hydrazide, such as furan-2-

carbohydrazide (23), using EDCI (1-[3-(dimethylamino)propyl]-3-

ethylcarbodiimide), HOBt (1-hydroxybenzotriazole), and NMM

(4-methylmorpholine) in the presence of DMF (N,N-dimethylfora-

mide) yielded the corresponding acylhydrazide 24, which was cy-

clized using Lawesson’s reagent to generate thiadiazole 25 in 81%

over two steps. Deprotection of the four benzyl groups using TMSI

(trimethyliodosilane)²¹followed by peracetylation using acetic

anhydride and puriﬁcation by either column chromatography or

recrystallization generated the corresponding tetraacetate. The tet-

raacetate was hydrolyzed with NaOMe in methanol to yield the

target compound 61 in 31% yield over three steps.

2.2. SAR (Structure–activity relationship) studies

The cell-based SGLT2 AMG (Methyl-a-D-glucopyranoside) inhi-

bition assay was performed to evaluate the inhibitory effects of all

prepared compounds on hSGLT2 activities.^24,25Exploration of the

SAR began by replacing the phenyl moiety at the distal ring posi-

Alternatively, another target compound was prepared as shown

in Scheme 5. Thus, the acid 21 was converted to the hydrazide 26

Cl

Br

CO₂H

16

.

1. BH₃SMe₂/ THF, 0 ºC to rt

2. TIPSCl, Imidazole, DMAP / DMF, rt

(95%, 2 steps)

Cl

OTIPS

O

OH

Br

BnO

1.

, BuLi / THF, -78 ºC

17

OBn

2. Et₃SiH, BF₃.OEt₂/ CH₂Cl₂, -50 to -10 ºC

3. Bu₄NF / THF, rt

OBn

18

15

(98%, 3 steps)

Scheme 2. Synthesis of C-glucoside alcohol intermediate 18.

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

2181

Cl

O

OH

BnO

O

CN

BnO

1. PBr₃, pyridine / ether, rt

(resolution)

OBn

2. KCN / EtOH, H₂O, reflux

(80%, 2 steps)

EtOH

(40%, 3 steps)

OBn

Cl

18

19

SOCl₂

O

CN

O

CO₂H

BnO

NaOH

MeOH

reflux

OBn

BnO

OBn

EtOH, H₂O

OBn

reflux

(98%)

(89%)

21

20

Cl

O

CO₂Me

BnO

OBn

22

Scheme 3. Preparation of key C-glucoside intermediates 21, 22.

Cl

O

H

N

EDCI

HOBt

NMM

DMF

O

CO₂H

+

O

BnO

O

BnO

N

H

CONHNH₂

O

OBn

23

OBn

24

21

Cl

N

Lawesson reagent

THF, reflux

1. TMSI / CH₃CN, 50 ºC

O

S

BnO

O

(81%, 2 steps)

2. Ac₂O, Et₃N, DMAP / CH₂Cl₂, rt

3. NaOMe / MeOH. rt

OBn

25

(31%, 3 steps)

Cl

N

O

S

HO

O

OH

61

Scheme 4. Synthesis of target compound 61.

OH

Cl

O

NH₂NH₂

O

NH₂

O

CO₂H

BnO

N

H

BnO

EDCI

HOBt

DMF

(95%)

OBn

BnO

OBn

Ph

OBn

21

26

Cl

N

O

EDCI

S

HO

Lawesson rgt.

TMSI

O

+

Ph

HO

O

HOBt

NMM

OH

(36%, 3 steps)

O

65

OH

27

Scheme 5. Synthesis of target compound 65.

tion of dapagliﬂozin 9 with a 1,3,4-oxadiazole or a 1,3,4-thiadia-

zole moiety. After it was discovered that the 1,3,4-oxadiazole ser-

ies demonstrated signiﬁcantly lower activity than the

corresponding 1,3,4-thiadiazole series, SAR studies focused mostly

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J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

Cl

N

S

28

Cl

O

CO₂H

S

BnO

H₂N

NHNH₂, POCl₃

BnO

1.

OBn

2. NaNO₂, Cu / c-HCl, AcOH

(22%, 2 steps)

OBn

29

21

Cl

N

SMe

O

1. NaSMe / THF, rt

S

HO

2. TMSI, CH₃CN, 50 ºC

(25%, 2 steps)

OH

47

Scheme 6. Synthesis of target compound 47.

N

30

CONHNH₂,

Cl

1.

N

EDCI, HOBt, NMM

O

CO₂H

S

HO

BnO

N

2, Lawesson reagent

3. TMSOTf / Ac₂O

4. NaOMe / MeOH

OH

OBn

OH

OBn

59

(10%, 4 steps)

21

Scheme 7. Synthesis of target compound 59.

Table 1

on thiadiazoles at the distal ring. The initial results of the mixture

of anomers are shown in Table 1. Straight aliphatic chains on the

In vitro screening data for hSGLT inhibitory activity^a,b

X

N

thiadiazole ring showed weak activity (31, IC₅₀= 6.9

bulky groups such as t-butyl provided a complete loss of activity

(32, IC₅₀>10 M), suggesting that branched aliphatic chains are

lM), while

N

R

O

S

HO

l

not tolerated at this position. Replacement of this moiety with

either phenyl 33 or pyridinyl 34 groups improved the inhibitory

HO

OH

activity against hSGLT2 approximately ﬁvefold (33, IC₅₀= 1.4

lM;

Compound

X

R

IC₅₀(nM)

34, IC₅₀= 1.2 M). The introduction of a small lipophilic substitu-

l

ent, chlorine, at the C4’ position of the C-glucoside proximal phenyl

ring improved the inhibitory activity by an order of magnitude.

Among these compounds, diaryl-types exhibited more favorable

activity (36–39, IC₅₀= 25–103 nM) than carbocycle (35,

6

9

6.25 0.62

0.49 0.04

31

H

6910

32

>10,000

IC₅₀= 542 nM) or benzyl-type moiety (40, IC₅₀= 2.4 lM). Among

substituted phenyls on the thiadiazole ring, 4-propyl (37,

IC₅₀= 24.9 nM) turned out to be more active than the correspond-

ing 4-methyl group (36, IC₅₀= 103 nM), implying the importance

of lipophilicity in this region.

33

34

35

36

37

38

39

H

1420

1230

542

H

N

Table 2 shows the structure–activity relationship upon alter-

ation of the substituent at the distal 1,3,4-thiadiazole ring employ-

ing only the b-anomer. Compounds bearing aliphatic chains at the

distal 1,3,4-thiadiazole ring were prepared and tested. These com-

pounds showed only moderate hSGLT2 inhibitory activities (42–

45, IC₅₀= 122–379 nM). Both triﬂuoromethyl substituted 48 and

triﬂuoroethyl substituted 49 did not improve in vitro activity

against hSGLT2. Substitution with a carbocycle, such as cyclopentyl

50, demonstrated suboptimal activity (IC₅₀= 445 nM). However, as

described in the initial SAR studies (Table 1), the diaryl-type com-

pound containing a phenyl group, 51, showed good in vitro inhib-

itory activity against hSGLT2 (IC₅₀= 16.6 nM). At this juncture, a

variety of different aromatic rings were explored. Five-membered

heterocycles, including 2-furan 61, 3-furan 62, 2-thiophene 70,

and 3-thiophene 71, were shown to be highly active, demonstrat-

ing low nanomolar inhibitory activity against hSGLT2. Compared to

phenyl 51 (IC₅₀= 16.6 nM), the incorporation of 3-furan 62 or 2-

thiophene 70 resulted in similar levels of inhibitory activity (62,

IC₅₀= 17.7 nM; 70, IC₅₀= 14.4 nM). Surprisingly, varying the bond-

ing position, for example, 3-furan 62 to 2-furan 61 or 2-thiophene

Cl

103

24.9

45.3

53.0

N

40

41

Cl

2390

Cl

>10,000

a

The IC₅₀values of 6 and 9 were obtained by in-house assay.

These data were obtained by single determinations.

b

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

2183

Table 2

In vitro screening data for hSGLT inhibitory activity^a,b

Cl

N

R

O

S

HO

OH

Compound

R

IC₅₀(nM)

Compound

R

IC₅₀(nM)

N

6

6.25 0.62

60

190

9

0.49 0.04

214

61

62

7.03

17.7

O

42

O

43

379

63

106

O

44

45

353

122

64

65

58.3

10.6

46

47

OMe

SMe

CF₃

146

66

67

249

257

O

10.9

O

N

48

155

68

119

N

CF₃

49

50

758

445

69

70

3280

14.4

N

S

51

52

16.6

126

71

72

3.51

762

Cl

53

30.3

73

712

S

N

Cl

54

55

638

74

75

18.2

S

97.4

44.5

N

S

(continued on next page)

2184

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

Table 2 (continued)

Compound

R

IC₅₀(nM)

36.4

Compound

R

IC₅₀(nM)

21.8

N

S

56

57

76

N

81.6

77

72.6

S

N

O

58

562

78

79

>10,000

N

O

59

4.06

a

The IC₅₀values of 6 and 9 were obtained by in-house assay.

These data were obtained by single determinations.

b

70 to 3-thiophene 71, improved the inhibitory activity by approx-

imately 2.5-fold or fourfold, respectively (61, IC₅₀= 7.03 nM; 71,

IC₅₀= 3.51 nM). Substitution at the C5 position of 2-furan 61 or

2-thiophene 70 with methyl 64, phenyl 65, or chloride 74 resulted

in a decreased potency. Moreover, addition of substituents at C3 of

2-furan 61 or 2-thiophene 70 resulted in signiﬁcantly lower

hSGLT2 inhibitory activities (63, 72, and 73), likely suggesting that

even slightly increased steric hindrance is not tolerated in the re-

gion. Other heteroaryl groups such as pyrrole (68, IC₅₀= 119 nM),

benzofuran (66, IC₅₀= 249 nM), quinoline (58, IC₅₀= 562 nM), and

pyridazine (60, IC₅₀= 190 nM) were not tolerated. Meanwhile, pyr-

idine 53, isoquinoline 57, benzothiophene 75, and thiazoles 76 and

77 were found to be somewhat tolerable replacements

(IC₅₀= 21.8–81.6 nM), but none appeared to be more potent than

the simple phenyl group. Interestingly, six-membered heteroaryl

compounds with two para heteroatoms, such as pyrazine 59,

proved to have highly potent inhibitory activity against hSGLT2

(IC₅₀= 4.06 nM). Thus, the results indicate that the addition of

small heteroaryl rings with a favorable orientation of the hetero-

atom at the distal ring might be able to further increase hSGLT2

inhibitory activity. It is noteworthy that any substitution at the aryl

or heteroaryl ring connected with the distal thiadiazole ring re-

sulted in products with signiﬁcantly lower hSGLT2 inhibitory activ-

ities, except furanophenyl 65 (IC₅₀= 10.6 nM). Di-substitution at

the heteroaryl ring on the distal thiadiazole moiety further deteri-

orated hSGLT2 inhibitory activities, as shown for compounds 78

and 79, indicating very limited tolerance in the speciﬁc area.

Among the tested compounds, as shown in Table 2, biaryl-type

compounds containing 3-thiophene 71, pyrazine 59, or 2-furan

61 showed the best in vitro inhibitory activities (IC₅₀= 3.51 nM,

4.06 nM, 7.03 nM, respectively). This implies that the 1,3,4-thiadi-

azole heterocycle at the distal ring could become a reasonable sur-

rogate for the phenyl group of dapagliﬂozin 9, and a properly

oriented small heteroaryl substituent at the distal ring might be

capable of augmenting in vitro hSGLT2 inhibitory activity.

cages for baseline urine collection over 24 h. Rats were weighed,

randomized into three groups (n = 3), dosed orally with single doses

of vehicle or drug (9@1 mg/kg, 61@10 mg/kg), and subsequently

dosed orally with 50% aqueous glucose solution (2 g/kg). Immedi-

ately after dosing, rats were returned to the metabolism cages for

24 h urine collection and re-fed at 1 h after the glucose challenge.

The urine glucose and urine volume data were normalized per

200 g of body weight. As shown in Figure 4, a single oral dose of

thiadiazole 61 increased urinary glucose excretion in normal SD

rats, resulting in a 100-fold elevation in glucose disposal relative

to vehicle controls. Urinary glucose excretion of compounds 9 and

61 were 1648 228 mg/200 g body weight and 195 84 mg/200 g

body weight, respectively. Urine volume excreted in normal SD rats

are also shown in Figure 5. Dapagliﬂozin 9 caused increased urine

volume over vehicle by 5.7-fold, while thiadiazole 61 increased ur-

ine volume merely 1.4-fold (vehicle: 3.9 0.23, 9: 22.36 4.18, 61:

5.33 0.67 ml/200 mg body weight, respectively).

For assessment of acute blood glucose effects in diabetic db/db

mice, the animals were weighed and randomized into two groups

(n = 4). Mice were dosed with vehicle or drug (9@1 mg/kg,

61@10 mg/kg), and blood glucose levels were measured immedi-

ately before dosing and at 1, 2, 4, 5, 6, and 24 h post-dose. The ani-

mals were allowed to re-feed after the 6-h time point. In this

experiment, a 72.8% reduction in blood glucose levels versus con-

trols were observed at 4 h after a single 10 mg/kg oral dose of 61

was administered to db/db mice with an initial blood glucose level

of 430 mg/dL. By comparison, a 61.3% reduction in blood glucose

∗

2000

1800

1600

1400

1200

1000

800

A selected promising compound 61 was tested on animal models

for in vivo efﬁcacy. Urinary glucose excretion was evaluated in nor-

mal Sprague-Dawley (SD) rats.^24–26Male SD rats weighing 250–

300 g (7–8 weeks old) were purchased from Orient-Bio Laboratory

Animal Research Center Co. (Gyeonggi-do, Korea). They were

housed in a temperature (25 2 °C) and moisture (55 10%) con-

trolled room, exposed to a controlled 12 h cycle of light and dark-

ness, and allowed free access to food and water. All animals were

acclimated for one week prior to the experiment. For glucosuria

assessment, overnight-fasted SD rats were placed into metabolism

600

400

∗

200

0

Vehicle

Compound 9

Compound 61

Figure 4. Urinary glucose excretion test of vehicle, compound 9 (1 mg/kg) and 61

(10 mg/kg) in normal SD rats. All results are expressed as means S.E.M. The

statistical analysis was performed by one-way ANOVA followed by the Dunnett’s

*

post hoc test. P <0.05 versus vehicle.

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

2185

ring was a worthy approach for the improvement of the partition

coefﬁcient (log P) value and to improve characteristics for develop-

ment, including non-speciﬁc plasma protein binding. For this pur-

pose, the structure of dapagliﬂozin 9 was modiﬁed into a general

structure bearing a heterocycle ring at the distal ring position.

Along these lines, novel C-aryl glucoside SGLT2 inhibitors contain-

ing 1,3,4-thiadiazole moieties were designed and synthesized.

Among the compounds tested, biaryl-type compounds containing

pyrazine 59, 2-furan 61, or 3-thiophene 71 exhibited the best

in vitro inhibitory activities against SGLT2 to date in this series

(IC₅₀= 3.51–7.03 nM). A selected compound 61 demonstrated rea-

sonable urinary glucose excretion and glucosuria in normal SD rats

along with favorable blood glucose-lowering effects in db/db mice,

thereby demonstrating that this 1,3,4-thiadiazolylmethylphenyl

glucoside series possesses both in vitro SGLT2 inhibition and

in vivo efﬁcacy, albeit to a lower degree. Consequently, the infor-

mation acquired from this series of compounds can be utilized as

a quick reference to achieve more advanced series in this ﬁeld.

30

25

20

15

10

5

∗

0

Vehicle

Compound 9

Compound 61

Figure 5. Urine volume of excreted of vehicle, compound 9 (1 mg/kg) and 61

(10 mg/kg) in normal SD rats. All results are expressed as means S.E.M. The

statistical analysis was performed by one-way ANOVA followed by the Dunnett’s

*

post hoc test. P <0.05 versus vehicle.

levels versus controls was observed at 4 h after a single 1 mg/kg

oral dose of 9 was administered to db/db mice having the same ini-

tial blood glucose levels (Fig. 6).

In order to further evaluate this series, the pharmacokinetic

properties of 61 were measured in male SD rats. After oral admin-

4. Experimental

istration of 5 mg/kg of 61 to rats, a C_maxof 0.05 lg/mL was ob-

4.1. General methods

tained at 0.33 h. The elimination half-life of 61 following oral

administration was 0.86 h in rats. Compound 61 showed relatively

low oral bioavailability (F = 7.7%) in rats, suggesting its likely solu-

bility-limited absorption. It is concluded that the decreased in vivo

efﬁcacy of the current SGLT2 inhibitor 61, compared with dapagli-

ﬂozin, could be attributed to the difference in inherent in vitro

potencies (61: IC₅₀= 7.03 nM vs 9: IC₅₀= 0.49 nM) as well as its

pharmacokinetic properties. Thus, replacement of the distal ring

of dapagliﬂozin 9 with a 1,3,4-thiadiazole ring, as in compound

61, appeared to diminish the in vitro activity and oral absorption

as well as the in vivo efﬁcacy in animal models. However, the infor-

mation obtained from the SAR studies in this thiadiazole series

should help to design more active SGLT2 inhibitors that are struc-

turally related.

All reactions are conducted under an inert atmosphere at room

temperature, unless otherwise noted. All reagents were purchased

at the highest commercial quality and used without further puriﬁ-

cation, unless otherwise indicated. Microwave reaction was con-

ducted with a Biotage Initiator microwave reactor. NMR spectra

were obtained on a Varian 400-MR (400 MHz ¹H, 100 MHz ¹³C)

spectrometer. NMR spectra were recorded in ppm (d) relative to

tetramethylsilane (d = 0.00) as an internal standard unless stated

otherwise and are reported as follows: chemical shift, multiplicity

(s = singlet, d = doublet, t = triplet, q = quartet, quint = quintet,

sext = sextet, m = multiplet, and br = broad), coupling constant,

and integration. ¹³C NMR spectra were referenced to the residual

chloroform-d₁(d = 77.0) or DMSO-d₆(d = 39.7). Mass spectra were

obtained with an Agilent 6110 quadruple LC-MSD (ESI+). High res-

olution mass spectra were obtained on a Jeol JMS-700 Mstation

(10 kV, HFAB). Optical rotations were obtained on a Rudolph Auto-

pol III digital polarimeter. Preparative HPLC puriﬁcations were per-

formed on a Gilson puriﬁcation system. For preparative HPLC, ca.

100 mg of a product was injected in 1 mL of methanol onto a Sun-

3. Conclusion

In the present study, metabolically more stable C-glucosides

bearing heteroaromatic rings were exploited in order to develop

novel SGLT2 targeting antidiabetic agents. We envisioned that

replacement of the distal ring of dapagliﬂozin 9 with a heterocyclic

Fire Prep C18 OBD 5 lm 30x100 mm Column with a 30 min gradi-

ent from 5% to 90% acetonitrile in water and a 45 mL/min ﬂow rate.

Biotage SP1 and Isolera puriﬁcation systems were used for normal

phase column chromatography with ethyl acetate and hexane.

Flash chromatography was performed using E. Merck 230–400

mesh silica gel according to the procedure of Still et al. Reactions

were monitored by either thin-layer chromatography (TLC) on

0.25 mm E. Merck silica gel plates (60F-254) using UV light and

p-anisaldehyde solution as visualizing agents or HPLC analysis on

an Agilent 1200 series system.

Acute effect in db/db mice

150

100

50

0

1

2

3

4

5

6

24

-50

-100

-150

-200

-250

-300

-350

∗∗

4.2. Chemistry

4.2.1. 2,3,4,6-Tetra-O-benzyl-D-glucopyranone (15)

To a solution of lactol 14 (125 g, 231 mmol) in dichloromethane

(1 L) was added 4 Å molecular sieve (40 g) and 4-methylmorpho-

line N-oxide (40.7 g, 347 mmol). The solution is stirred for

20 min at room temperature, before adding of tetrapropylammoni-

um perruthenate (2.0 g, 5.6 mmol). After 3 h stirring at ambient

temperature, the solution was ﬁltered through a plug of Celite.

The ﬁltrate was washed with saturated sodium thiosulfate, dried

over anhydrous MgSO₄, ﬁltered and concentrated in vacuo. The

resulting crude residue was puriﬁed by column chromatography

Time (hr)

Compound 61

Vehicle

Compound 9

Figure 6. Blood glucose level in db/db mice following a single oral dose of vehicle

compound 9 (1 mg/kg) or compound 61 (10 mg/kg). All results are expressed as

means S.E.M. The statistical analysis was performed by one-way ANOVA followed

*

by the Dunnett’s post hoc test. P <0.05 versus vehicle.

2186

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

(5–40% EtOAc/hexanes) to yield the title compound (106 g, 86%) as

a colorless oil. ¹H NMR (400 MHz, CDCl₃) d 7.39–7.16 (m, 20H),

4.98 (d, J = 11.2 Hz, 1H), 4.74–4.44 (m, 8H), 4.12 (d, J = 6.4 Hz,

1H), 3.97–3.89 (m, 2H), 3.72 (dd, J = 10.8, 2.4 Hz, 1H), 3.67 (dd,

J = 10.8, 3.2 Hz, 1H).

washed with brine, dried over anhydrous MgSO₄, ﬁltered and con-

centrated in vacuo. The crude residue was puriﬁed on column

chromatography (10–60% EtOAc/hexanes) to yield the title com-

pound (130 g, 98%, ca. 2:1 b:a

) as a white solid. ¹H NMR

(400 MHz, CDCl₃) b-anomer: d 7.50 (s, 1H), 7.38–7.17 (m, 20H),

6.94–6.92 (m, 2H), 4.96–4.46 (m, 9H), 4.23 (d, J = 9.2 Hz, 1H),

3.86 (d, J = 10.4 Hz, 1H), 3.81–3.67(m, 4H), 3.61–3.59 (m, 1H),

4.2.2. (5-Bromo-2-chlorobenzyloxy)triisopropylsilane (17)

To

a

solution 5-bromo-2-chlorobenzoic acid 16 (100 g,

3.44 (t, J = 9.2 Hz, 1H); a-anomer: d 7.78 (s, 1H), 7.63 (dd, J = 8.4,

425 mmol) in THF (500 mL) at 0 °C was added borane-dimethyl

sulﬁde complex (170 mL, 1,700 mmol). The resulting mixture was

stirred with gradual warming to ambient temperature over 12 h,

re-cooled to 0 °C, and quenched with MeOH and then water. The

mixture was extracted with EtOAc. The organic layer was washed

with brine, dried over anhydrous MgSO₄, ﬁltered and evaporated

in vacuo to yield a white solid, which was carried on to the next

step without further puriﬁcation.

1.6 Hz, 1H), 7.38–7.17 (m, 19H), 7.13–7.10 (m, 2H), 5.15 (d,

J = 3.6 Hz, 1H), 4.96–4.46 (m, 8H), 4.01–3.95 (m, 2H), 3.83–3.64

(m, 5H), 3.55–3.51 (m, 1H); MS (ESI) m/z 682 (M+NH₄)⁺, 687

(M+Na)⁺.

4.2.4. 2-(2-Chloro-5-((3S,4R,5R,6R)-3,4,5-tris(benzyloxy)-6-

(benzyloxymethyl)-tetrahydro-2H-pyran-2-

yl)phenyl)acetonitrile (19)

To a solution of the benzyl alcohol in DMF (400 mL) was added

imidazle (57.9 g, 850 mmol), 4-(dimethylamino)pyridine (2.6 g,

21 mmol), and triisopropylsilyl chloride (136 mL, 683 mmol). The

resulting solution was stirred at ambient temperature for 12 h, di-

luted with a saturated ammonium chloride and extracted with

EtOAc. The organic layer was washed with water then brine, dried

over anhydrous MgSO₄, ﬁltered and concentrated in vacuo. The

crude residue was puriﬁed by column chromatography (3–10%

EtOAc/hexanes) to yield the title compound (152 g, 95%) as a color-

less oil. ¹H NMR (400 MHz, CDCl₃) d 7.77 (d, J = 2.4 Hz, 1H), 7.31

(dd, J = 8.4, 2.4 Hz, 1H), 7.16 (d, J = 8.4 Hz, 1H), 4.83 (s, 2H), 1.25–

1.17 (m, 3H), 1.11 (d, J = 6.8 Hz, 18H).

To a solution of 2-chlorobenzyl alcohol 18 (130 g, 195 mmol) in

ether (600 mL) at 0 °C was added pyridine (0.79 mL, 9.8 mmol) and

phosphorus tribromide (6.4 mL, 68 mmol). The reaction was al-

lowed to slowly warm to room temperature over 15 h and reﬂuxed

for 1 h. After cooling to room temperature, the reaction mixture

was diluted with EtOAc and washed with water then brine. The or-

ganic extract was dried over anhydrous MgSO₄, ﬁltered, and evap-

orated in vacuo to yield a yellow solid, which was carried on to the

next step without further puriﬁcation.

To a solution of the 2-chlorobenzyl bromide in ethanol (260 mL)

and water (130 mL) was added potassium cyanide (31.8 g,

488 mmol). The reaction mixture was reﬂuxed overnight. After

cooling to room temperature, the mixture was diluted with water

and extracted with EtOAc. The organic layer was washed with

brine, dried over anhydrous MgSO₄, ﬁltered and evaporated in va-

cuo. Puriﬁcation was accomplished by chromatography to give the

4.2.3. (2-Chloro-5-((3S,4R,5R,6R)-3,4,5-tris(benzyloxy)-6-

(benzyloxymethyl)-tetrahydro-2H-pyran-2-yl)phenyl)methanol

(18)

To a solution of bromide 17 (97 g, 257 mmol) in THF (1 L) at

ꢀ78 °C under an atmosphere of nitrogen was added dropwise n-

butyllithium (2.5 M in hexanes, 103 mL, 257 mmol), and the mix-

ture was stirred for 1.5 h at the same temperature. Then a solution

of lactone 15 (106 g, 198 mmol) in THF (500 mL) was added drop-

wise, and the mixture was stirred for 3 h at the same temperature.

The reaction mixture was quenched by addition of saturated

ammonium chloride. After complete addition, the solution was

gradually raised to room temperature. The organic layer was sepa-

rated and the aqueous layer was extracted with EtOAc. The com-

bined organic layers were washed with brine, dried over

anhydrous MgSO₄, ﬁltered and concentrated in vacuo to yield a

yellow oil, which was carried on to the next step without further

puriﬁcation.

To a stirred ꢀ50 °C solution of the lactols in dichloromethane

(500 mL) was added triethylsilane (63 mL, 396 mmol) followed

by boron triﬂuoride diethyl etherate (50 mL, 396 mmol) at a rate

such that the reaction temperature was maintained between ꢀ40

and ꢀ50 °C. The solution was allowed to warm to ꢀ10 °C over

2 h prior to quenching with saturated potassium carbonate. After

removal of organic volatiles under reduced pressure, the residue

was partitioned between EtOAc and water. Following extraction

of the aqueous layer with ethyl acetate, the combined organic lay-

ers were washed with water prior to drying over anhydrous

MgSO₄. Filtration and concentration under reduced pressure

yielded a yellow oil, which was carried on to the next step without

further puriﬁcation.

title compound (105 g, 80%, ca. 2:1 b:a

) as a colorless oil. ¹H NMR

(400 MHz, CDCl₃) b-anomer: d 7.48 (s, 1H), 7.37–7.17 (m, 20H),

6.93–6.91 (m, 2H), 4.93 (s, 2H), 4.86 (d, J = 10.8 Hz, 1H), 4.63 (d,

J = 10.8 Hz, 1H), 4.62 (d, J = 12.4 Hz, 1H), 4.56 (d, J = 12.4 Hz, 1H),

4.52 (d, J = 10.8 Hz, 1H), 4.22 (d, J = 9.6 Hz, 1H), 3.93 (d,

J = 10.8 Hz, 1H), 3.84–3.68 (m, 6H), 3.62–3.58 (m, 1H), 3.43 (t,

J = 8.8 Hz, 1H);

a-anomer: d 7.83 (d, J = 1.6 Hz, 1H), 7.64 (dd,

J = 8.4, 1.6 Hz, 1H), 7.37–7.19 (m, 19H), 7.13–7.10 (m, 2H), 5.11

(d, J = 2.8 Hz, 1H), 4.93–4.46 (m, 8H), 3.96–3.92 (m, 2H), 3.79–

3.71 (m, 3H), 3.70–3.62 (m, 2H), 3.55–3.51 (m, 1H); MS (ESI) m/z

696 (M+Na)⁺.

4.2.5. 2-(2-Chloro-5-((2S,3S,4R,5R,6R)-3,4,5-tris(benzyloxy)-6-

(benzyloxymethyl)-tetrahydro-2H-pyran-2-

yl)phenyl)acetonitrile (20)

The mixture of cyanides 19 (105 g, 156 mmol, ca. 2:1 b:a) was

slurried in ethanol (1 L) and heated to reﬂux with stirring. The

reaction mixture was held at reﬂux for 1 h to insure that all of solu-

tion had homogenized. It was then cooled evenly at 15 °C/h to

ambient temperature and stirred overnight at this temperature.

The resulting solid was isolated by ﬁltration and dried in vacuo

to yield the title compound (53 g, 51%, ca. 75% separation yield)

as a white solid. ½a _D²¹

ꢁ

ꢀ10.6 (c 1.01, chloroform); ¹H NMR

(400 MHz, CDCl₃) d 7.48 (s, 1H), 7.37–7.17 (m, 20H), 6.93–6.91

(m, 2H), 4.93 (s, 2H), 4.86 (d, J = 10.8 Hz, 1H), 4.63 (d, J = 10.8 Hz,

1H), 4.62 (d, J = 12.4 Hz, 1H), 4.56 (d, J = 12.4 Hz, 1H), 4.52 (d,

J = 10.8 Hz, 1H), 4.22 (d, J = 9.6 Hz, 1H), 3.93 (d, J = 10.8 Hz, 1H),

3.84–3.68 (m, 6H), 3.62–3.58 (m, 1H), 3.43 (t, J = 8.8 Hz, 1H); ¹³C

NMR (100 MHz, CDCl₃) d 129.49, 128.86, 128.45, 128.38, 128.26,

128.08, 128.01, 127.87, 127.82, 127.70, 127.68, 127.67, 127.64,

127.61, 139.09, 138.52, 138.23, 138.07, 137.41, 132.97, 129.49,

128.86, 128.45, 128.38, 128.26, 128.08, 128.01, 127.87, 127.82,

127.70, 127.68, 127.67, 127.64, 127.61, 116.57, 86.87, 83.59,

80.47, 79.41, 78.28, 75.67, 75.16, 74.96, 73.47, 69.07, 22.12; MS

To a solution of the silyl ether in THF (500 mL) was added tet-

rabutylammonium ﬂuoride (1.0 M in THF, 594 mL, 594 mmol)

and the reaction mixture stirred at ambient temperature for 2 h.

After removal of organic volatiles under reduced pressure, the res-

idue was partitioned between EtOAc and saturated ammonium

chloride. The organic layer was separated and the aqueous layer

was extracted with EtOAc. The combined organic layers were

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

2187

(ESI) m/z 696 (M+Na)⁺; HRMS (FAB) m/z 674.2672 [calcd for

MgSO₄, ﬁltered and concentrated in vacuo. The resulting crude res-

idue was puriﬁed by column chromatography (10–60% EtOAc/hex-

anes) to yield the title compound (468 mg, 81%) as a yellow oil. ¹H

NMR (400 MHz, CDCl₃) d 7.50 (d, J = 1.6 Hz, 1H), 7.46 (d, J = 1.6 Hz,

1H), 7.39 (d, J = 8.0 Hz, 1H), 7.35–7.13 (m, 19H), 7.08 (d, J = 3.6 Hz,

1H), 6.90–6.88 (m, 2H), 6.53–6.52 (m, 1H), 4.90 (s, 2H), 4.85(d,

J = 10.8 Hz, 1H), 4.63–4.51 (m, 4H), 4.45 (d, J = 10.8 Hz, 1H), 4.20

(d, J = 9.6 Hz, 1H), 3.88 (d, J = 10.8 Hz, 1H), 3.81–3.72 (m, 5H),

3.60–3.57 (m, 1H), 3.43 (t, J = 8.8 Hz, 1H); MS (ESI) m/z 799

(M+H)⁺, 821 (M+Na)⁺.

C₄₂H₄₁ClNO₅(M+H)⁺674.2673].

4.2.6. 2-(2-Chloro-5-((2S,3S,4R,5R,6R)-3,4,5-tris(benzyloxy)-6-

(benzyloxymethyl)-tetrahydro-2H-pyran-2-yl)phenyl)acetic

acid (21)

To a solution of cyanide 20 (53 g, 79 mmol) in EtOH (300 mL)

was added aq NaOH solution (8.0 N, 300 mL, 2.4 mol). The reaction

mixture was reﬂuxed overnight. After cooling to room tempera-

ture, hydrochloric acid (3.0 N) was added to neutralize the reaction

mixture. The mixture was diluted with water and extracted with

EtOAc. The organic layer was washed with water and brine prior

to drying over anhydrous MgSO₄. Filtration and concentration un-

der reduced pressure yielded the title compound, which was used

4.2.9. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(furan-2-yl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (61)

in the next step without further puriﬁcation. ½a _D²¹

ꢁ

ꢀ4.7 (c 1.10,

To a solution of perbenzylated 25 (468 mg, 0.58 mmol) in aceto-

nitrile (5 mL) was added iodotrimethylsilane (5 mL). The resulting

reaction mixture was heated to 50 °C overnight. After cooling to

0 °C, the reaction was quenched with methanol and concentrated

in vacuo. Peracetylation was achieved by addition of acetic anhy-

dride (3 mL) and DMAP (10 mg, 0.08 mmol) to a solution of this

residue in dichloromethane (10 mL) and triethylamine (3 mL).

After 2 h stirring at ambient temperature, the reaction was

quenched by addition of water, whereupon the resulting mixture

was extracted with dichloromethane. The organic layer was

washed with hydrochloric acid (1.0 N) and brine prior to drying

over anhydrous MgSO₄. After ﬁltration through a plug of silica

gel and concentration under reduced pressure, the residue was car-

ried on to the next step without further puriﬁcation.

chloroform); ¹H NMR (400 MHz, CDCl₃) d 7.37–7.14 (m, 21H),

6.93–6.90 (m, 2H), 4.94 (d, J = 10.8 Hz, 1H), 4.90 (d, J = 10.8 Hz,

1H), 4.86 (d, J = 11.2 Hz, 1H), 4.63 (d, J = 10.8 Hz, 1H), 4.62 (d,

J = 14.0 Hz, 1H), 4.55 (d, J = 12.4 Hz, 1H), 4.43 (d, J = 10.4 Hz, 1H),

4.20 (d, J = 9.6 Hz, 1H), 3.88 (d, J = 10.8 Hz, 1H). 3.81–3.70 (m,

6H), 3.59–3.56 (m, 1H), 3.43 (t, J = 8.8 Hz, 1H); ¹³C NMR

(100 MHz, DMSO-d₆) d 171.84, 139.10, 138.69, 138.66, 138.17,

133.72, 133.49, 131.99, 129.18, 128.68, 128.66, 128.45, 128.26,

128.19, 128.10, 128.00, 127.92, 127.87, 127.84, 86.25, 83.53,

80.14, 78.77, 78.55, 74.97, 74.51, 74.30, 72.75, 69.39, 39.19; MS

(ESI) m/z 710 (M+NH₄)⁺, 715 (M+Na)⁺.

4.2.7. 2-(2-Chloro-5-((2S,3S,4R,5R,6R)-3,4,5-tris-(benzyloxy)-6-

(benzyloxymethyl)-tetrahydro-2H-pyran-2-yl)phenyl)-acetate

(22)

To a solution of the tetraacetate in MeOH (10 mL) was added so-

dium methoxide (25 wt.% in MeOH, 0.5 mL). The reaction mixture

was stirred at ambient temperature for 2 h. Amberlite IR120

hydrogen form was then added to neutralize the reaction mixture.

The resin was ﬁltered off and the ﬁltrate was concentrated in va-

cuo. The resulting crude residue was puriﬁed by reverse phase pre-

parative HPLC to provide the title compound (78 mg, 31%) as a

To a solution of acid 21 (10 g, 14 mmol) in MeOH (140 mL) was

added thionyl chloride (2.1 mL, 29 mmol). The reaction mixture

was reﬂuxed overnight and cooled to room temperature. After re-

moval of organic volatiles under reduced pressure, the residue was

partitioned between EtOAc and water. The organic layer was sep-

arated and the aqueous layer was extracted with EtOAc. The com-

bined organic layers were washed with saturated sodium

bicarbonate and brine, dried over anhydrous MgSO₄, ﬁltered and

concentrated in vacuo. The crude residue was puriﬁed by column

chromatography (5–40% EtOAc/hexanes) to yield the title com-

pound (9.1 g, 89%) as a white solid. ¹H NMR (400 MHz, CDCl₃) d

7.36–7.18 (m, 21H), 6.94–6.92 (m, 2H), 4.96 (d, J = 11.2 Hz, 1H),

4.90 (d, J = 11.2 Hz, 1H), 4.86 (d, J = 10.8 Hz, 1H), 4.63 (d,

J = 11.6 Hz, 2H), 4.55 (d, J = 12.0 Hz, 1H), 4.45 (d, J = 10.8 Hz, 1H),

4.20 (d, J = 9.2 Hz, 1H), 3.90 (d, J = 10.8 Hz, 1H), 3.82–3.67 (m,

6H), 3.67 (s, 3H), 3.60–3.57 (m, 1H), 3.47 (t, J = 8.8 Hz, 1H); MS

(ESI) m/z 729 (M+Na)⁺.

white solid. ½a ²_D¹

ꢁ

+21.4 (c 0.21, methanol); ¹H NMR (400 MHz,

DMSO-d₆) d 7.93 (d, J = 1.6 Hz, 1H), 7.50 (d, J = 2.0 Hz, 1H), 7.42

(d, J = 8.0 Hz, 1H), 7.30 (dd, J = 8.0, 2.0 Hz, 1H), 7.23 (d, J = 3.6 Hz,

1H), 6.70 (dd, J = 3.6, 1.6 Hz, 1H), 5.00 (br, 2H), 4.88 (d, J = 5.6 Hz,

1H), 4.59 (d, J = 16.0 Hz, 1H), 4.53 (d, J = 16.0 Hz, 1H), 4.43 (t,

J = 5.6 Hz, 1H), 4.01 (d, J = 9.6 Hz, 1H), 3.68–3.65 (m, 1H), 3.46–

3.40 (m, 1H), 3.26–3.06 (m, 4H); ¹³C NMR (100 MHz, DMSO-d₆) d

167.62, 158.77, 146.61, 144.94, 140.68, 134.60, 132.39, 131.23,

129.35, 129.26, 113.23, 112.67, 81.67, 80.89, 78.71, 75.19, 70.68,

61.75, 33.77, MS (ESI) m/z 439 (M+H)⁺; HRMS (FAB) m/z

439.0730 [calcd for C₁₉H₂₀ClN₂O₆S (M+H)⁺439.0731]

4.2.10. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(furan-2-yl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (31)

4.2.8. 2-(2-Chloro-5-((2S,3S,4R,5R,6R)-3,4,5-tris(benzyloxy)-6-

(benzyloxymethyl)-tetrahydro-2H-pyran-2-yl)benzyl)-5-(furan-

2-yl)-1,3,4-thiadiazole (25)

Compound 31 was synthesized from de-chlorinated analogue of

To a mixture of acid 21 (500 mg, 0.72 mmol), 2-furoic hydrazide

23 (119 mg, 0.94 mmol), EDCI (207 mg, 1.08 mmol), and HOBt hy-

drate (224 mg, 1.44 mmol) in DMF (10 mL) was added NMM

(0.24 mL, 2.16 mmol). The resulting mixture was stirred at ambient

temperature overnight. After dilution with EtOAc, the organic layer

was subsequently washed with water, hydrochloric acid (1.0 N),

saturated sodium bicarbonate, and brine prior to drying over anhy-

drous MgSO₄. Filtration and removal of the volatiles under reduced

pressure yielded a glassy off-white amorphous solid, which was

carried on to the next step without further puriﬁcation.

To a solution of the acylhydrazide 24 in THF (15 mL) was added

Lawesson reagent (874 mg, 2.16 mmol). The reaction mixture was

reﬂuxed overnight. After cooling to room temperature, the reaction

mixture was diluted with EtOAc and washed with saturated so-

dium bicarbonate. The organic layer was dried over anhydrous

21 (ca. 2:1 (b:a) mixture of anomers) and butyrohydrazide accord-

ing to the procedure used to prepare 61. A white solid. Yield 17%.

¹H NMR (400 MHz, CD₃OD) d 7.40 (s, 1H), 7.35–7.28 (m, 2H),

7.24–7.22 (m, 1H), 4.39 (s, 2H), 4.11 (d, J = 9.2 Hz, 1H), 3.87 (d,

J = 11.2 Hz, 1H), 3.71–3.66 (m, 1H), 3.48–3.30 (m, 4H), 3.00 (t,

J = 7.6 Hz, 2H), 1.75 (sext, J = 7.6 Hz, 2H), 0.97 (t, J = 7.6 Hz, 3H);

MS (ESI) m/z 381 (M+H)⁺.

4.2.11. (3R,4R,5S,6R)-2-(3-((5-tert-Butyl-1,3,4-thiadiazol-2-

yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-2H-pyran-

3,4,5-triol (32)

Compound 32 was synthesized from de-chlorinated analogue of

21 (ca. 2:1 (b:a) mixture of anomers) and pivalohydrazide accord-

ing to the procedure used to prepare 61. A white solid. Yield 20%.

¹H NMR (400 MHz, CD₃OD) d 7.42 (s, 1H), 7.37–7.31 (m, 2H),

2188

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

7.27–7.25 (m, 1H), 4.40 (s, 2H), 4.13 (d, J = 9.6 Hz, 1H), 3.88 (d,

J = 11.6 Hz, 1H), 3.72–3.64 (m, 1H), 3.47–3.32 (m, 4H), 1.43 (s,

9H); MS (ESI) m/z 395 (M+H)⁺, 417 (M+Na)⁺.

4.2.17. (3R,4R,5S,6R)-2-(4-Chloro-3-((5-(pyridin-3-yl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (38)

Compound 38 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:a) and nicotinohydrazide according to the procedure

4.2.12. (2R,3S,4R,5R)-2-(Hydroxymethyl)-6-(3-((5-phenyl-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-tetrahydro-2H-pyran-3,4,5-triol

(33)

Compound 33 was synthesized from de-chlorinated analogue of

21 (ca. 2:1 (b:a) mixture of anomers) and benzoic hydrazide

according to the procedure used to prepare 61. A white solid. Yield

30%. ¹H NMR (400 MHz, CD₃OD) d 7.88 (d, J = 7.6 Hz, 2H), 7.01–7.45

(m, 4H), 7.37–7.29 (m, 3H), 4.47 (br, 1H), 4.13 (d, J = 9.6 Hz, 1H),

3.88–3.85 (m 1H), 3.68 (dd, J = 12.0, 5.2 Hz, 1H), 3.48–3.38 (m,

3H), 3.35–3.30 (m, 2H); MS (ESI) m/z 415 (M+H)⁺, 437 (M+Na)⁺.

used to prepare 61. A white solid. Yield 21%. ¹H NMR (400 MHz,

DMSO-d₆) d 9.07 (d, J = 1.6 Hz, 1H), 8.80 (dd, J = 4.8, 1.6 Hz, 1H),

8.30–8.27 (m, 1H), 7.55–7.51 (m, 2H), 7.43 (d, J = 8.4 Hz, 1H),

7.32 (dd, J = 8.4, 2.0 Hz, 1H), 5.02–4.84 (br, 3H), 4.63 (d,

J = 16.0 Hz, 1H), 4.58 (d, J = 16.0 Hz, 1H), 4.41 (br, 1H), 4.02 (d,

J = 9.6 Hz, 1H), 3.69–3.66 (m, 1H), 3.46–3.41 (m, 1H), 3.23–3.06

(m, 4H);MS (ESI) m/z 450 (M+H)⁺, 472 (M+Na)⁺.

4.2.18. (3R,4R,5S,6R)-2-(4-Chloro-3-((5-(pyridin-3-yl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (39)

4.2.13. (2R,3S,4R,5R)-2-(Hydroxymethyl)-6-(3-((5-(pyridin-3-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-tetrahydro-2H-pyran-

3,4,5-triol (34)

Compound 34 was synthesized from de-chlorinated analogue of

21 (ca. 2:1 (b:a) mixture of anomers) and nicotinohydrazide

according to the procedure used to prepare 61. A white solid. Yield

6.5%. ¹H NMR (400 MHz, CD₃OD) d 9.05 (s, 1H), 8.65 (d, J = 4.8 Hz,

1H), 8.34–8.31 (m, 1H), 7.56 (dd, J = 8.0, 5.2 Hz, 1H), 7.46 (s, 1H),

7.37–7.31 (m, 3H), 4.51 (s, 2H), 4.13 (d, J = 9.6 Hz, 1H), 3.86 (d,

J = 11.6 Hz, 1H), 3.68 (dd, J = 11.6, 5.2 Hz, 1H), 3.48–3.32 (m, 4H);

MS (ESI) m/z 416 (M+H)⁺, 438 (M+Na)⁺.

Compound 39 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:a) and isonicotinohydrazide according to the procedure

used to prepare 61. A white solid. Yield 22%. ¹H NMR (400 MHz,

CD₃OD) d 8.76 (d, J = 6.4 Hz, 2H), 8.12 (d, J = 6.4 Hz, 2H), 7.59 (d,

J = 1.2 Hz, 1H), 7.44–7.39 (m, 2H). 4.68 (s, 2H), 4.14 (d, J = 9.6 Hz,

1H), 3.87 (dd, J = 12.0, 2.0 Hz, 1H), 3.70 (dd, J = 12.0, 5.2 Hz, 1H),

3.48–3.38 (m, 3H), 3.34 (s, 1H), 3.26 (d, J = 8.8 Hz, 1H); MS (ESI)

m/z 450 (M+H)⁺, 472 (M+Na)⁺.

4.2.19. (3R,4R,5S,6R)-2-(4-Chloro-3-((5-(4-chlorobenzyl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (40)

4.2.14. (3R,4R,5S,6R)-2-(4-Chloro-3-((5-cyclohexyl-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (35)

Compound 35 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:a) and cyclohexanecarbohydrazide according to the pro-

Compound 40 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:a) and 2-(4-chlorophenyl)acetohydrazide according to

the procedure used to prepare 61. A white solid. Yield 14%. ¹H

NMR (400 MHz, CD₃OD) d 7.49 (s, 1H), 7.37–7.25 (m, 6H), 4.47 (s,

2H), 4.35 (s, 2H), 4.10 (d, J = 9.6 Hz, 1H), 3.85 (dd, J = 12.0, 1.6 Hz,

1H), 3.68 (dd, J = 12.0, 5.2 Hz, 1H), 3.44–3.29 (m, 3H), 3.24 (t,

J = 9.2 Hz, 1H); MS (ESI) m/z 497 (M+H)⁺, 519 (M+Na)⁺.

cedure used to prepare 61. A white solid. Yield 25%. ¹H NMR

(400 MHz, DMSO-d₆) d 7.45 (d, J = 2.0 Hz, 1H), 7.39 (d, J = 8.4 Hz,

1H), 7.28 (dd, J = 8.4, 2.0 Hz, 1H), 5.07–4.89 (br, 3H), 4.48 (d,

J = 15.6 Hz, 1H), 4.43 (d, J = 15.6 Hz, 1H), 4.42 (br, 1H), 3.99 (d,

J = 9.6 Hz, 1H), 3.69–3.65 (m, 1H), 3.46–3.40 (m, 1H), 3.24–3.00

(m, 5H), 1.99–1.94 (m, 2H), 1.73–1.67 (m, 2H), 1.64–1.59 (m,

1H), 1.46–1.28 (m, 3H), 1.24–1.14 (m, 1H); MS (ESI) m/z 455

(M+H)⁺, 477 (M+Na)⁺.

4.2.20. (3R,4R,5S,6R)-2-(4-Chloro-3-((5-(2-(4-chlorophenyl)

propan-2-yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)-tetrahydro-2H-pyran-3,4,5-triol (41)

Compound 41 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:a) and 2-(4-chlorophenyl)-2-methylpropanehydrazide

according to the procedure used to prepare 61. A white solid. Yield

58%. ¹H NMR (400 MHz, CD₃OD) d 7.49 (s, 1H), 7.38–7.34 (m, 2H),

7.29 (s, 4H), 4.49 (s, 2H), 4.11 (d, J = 9.2 Hz, 1H), 3.85 (dd, J = 12.4,

2.0 Hz, 1H), 3.68 (dd, J = 12.4, 5.6 Hz, 1H), 3.47–3.31 (m, 3H), 3.24

(t, J = 8.8 Hz, 1H), 1.80 (s, 6H); MS (ESI) m/z 525 (M+H)⁺, 547(M+Na)⁺.

4.2.15. (3R,4R,5S,6R)-2-(4-Chloro-3-((5-p-tolyl-1,3,4-thiadiazol-

2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-2H-pyran-

3,4,5-triol (36)

Compound 36 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:a) and 4-methylbenzohydrazide according to the proce-

dure used to prepare 61. A white solid. Yield 34%. ¹H NMR

(400 MHz, CD₃OD) d 7.76 (d, J = 8.0 Hz, 2H), 7.56 (d, J = 2.0 Hz,

1H), 7.43–7.37 (m, 2H), 7.29 (d, J = 8.0 Hz, 2H), 4.58 (s, 2H), 4.14

(d, J = 9.6 Hz, 1H), 3.87 (dd, J = 12.0, 2.0 Hz, 1H), 3.70 (dd, J = 12.0,

5.2 Hz, 1H), 3.48–3.38 (m, 3H), 3.26 (d, J = 9.2 Hz, 1H), 2.37 (s,

3H); MS (ESI) m/z 463 (M+H)⁺, 485 (M+Na)⁺.

4.2.21. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-propyl-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (42)

Compound 42 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:a) and butyrohydrazide according to the procedure used

to prepare 61. The small amount of the

a

-anomer was removed by

reverse phase preparative HPLC. A white solid. Yield 12%. ¹H NMR

(400 MHz, CD₃OD) d 7.51 (d, J = 1.6 Hz, 1H), 7.41–7.36 (m, 2H), 4.52

(s, 2H), 4.12 (d, J = 9.2 Hz, 1H), 3.86 (dd, J = 12.0, 1.6 Hz, 1H), 3.69

(dd, J = 12.0, 5.2 Hz, 1H), 3.47–3.37 (m, 3H), 3.25 (d, J = 8.8 Hz,

1H), 3.01 (t, J = 7.6 Hz, 2H), 1.76 (sext, J = 7.6 Hz, 2H), 0.97 (t,

J = 7.6 Hz, 3H); MS (ESI) m/z 415 (M+H)⁺.

4.2.16. (3R,4R,5S,6R)-2-(4-Chloro-3-((5-(4-propylphenyl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (37)

Compound 37 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:a) and 4-propylbenzohydrazide according to the proce-

dure used to prepare 61. A white solid. Yield 12%. ¹H NMR

(400 MHz, CD₃OD) d 7.79 (d, J = 8.0 Hz, 2H), 7.57 (d, J = 1.6 Hz,

1H), 7.43–7.37 (m, 2H), 7.30 (d, J = 8.0 Hz, 2H), 4.14 (d, J = 9.6 Hz,

1H), 3.87 (dd, J = 12.0, 2.0 Hz, 1H), 3.70 (dd, J = 12.0, 5.2 Hz, 1H),

3.48–3.38 (m, 3H), 3.33 (s, 1H), 3.26 (d, J = 9.2 Hz, 1H), 2.63 (t,

J = 7.6 Hz, 2H), 1.65 (sext, J = 7.6 Hz, 2H), 0.93 (t, J = 7.6 Hz, 3H);

MS (ESI) m/z 491 (M+H)⁺, 513 (M+Na)⁺.

4.2.22. (2S,3R,4R,5S,6R)-2-(3-((5-Butyl-1,3,4-thiadiazol-2-

yl)methyl)-4-chlorophenyl)-6-(hydroxymethyl)-tetrahydro-2H-

pyran-3,4,5-triol (43)

Compound 43 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:

a) and pentanehydrazide according to the procedure

used to prepare 61. The small amount of the

a

-anomer was re-

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

2189

moved by reverse phase preparative HPLC. A white solid. Yield 19%.

4.2.27. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(4-chlorophenyl)-

1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (52)

¹H NMR (400 MHz, DMSO-d₆) d 7.45 (d, J = 2.0 Hz, 1H), 7.39 (d,

J = 8.4 Hz, 1H), 7.28 (dd, J = 8.4, 2.0 Hz, 1H), 5.04 (br, 2H), 4.87 (d,

J = 5.6 Hz, 1H), 4.48 (d, J = 16.0 Hz, 1H), 4.43 (d, J = 16.0 Hz, 1H),

4.43 (br, 1H), 3.99 (d, J = 9.2 Hz, 1H), 3.68–3.65 (m, 1H), 3.46–

3.40 (m, 1H), 3.26–3.04 (m, 4H), 2.97 (t, J = 7.6 Hz, 2H), 1.61 (quint,

J = 7.6 Hz, 2H), 1.29 (sext, J = 7.6 Hz, 2H), 0.85 (t, J = 7.6 Hz, 3H); MS

(ESI) m/z 429 (M+H)⁺, 451 (M+Na)⁺.

Compound 52 was synthesized from an anomeric mixture of

21 (ca. 2:1 b: ) and 4-chlorobenzohydrazide according to the

procedure used to prepare 61. The small amount of the -anomer

a

was removed by reverse phase preparative HPLC. A white solid.

Yield 50%. ¹H NMR (400 MHz, DMSO-d₆) d 7.92 (d, J = 8.4 Hz,

2H), 7.56 (d, J = 8.4 Hz, 2H), 7.51 (d, J = 2.0 Hz, 1H), 7.43 (d,

J = 8.0 Hz, 1H), 7.31 (dd, J = 8.0, 2.0 Hz, 1H), 4.96–4.93 (m, 2H),

4.86 (d, J = 5.6 Hz, 1H), 4.60 (d, J = 16.0 Hz, 1H), 4.55 (d,

J = 16.0 Hz, 1H), 4.43 (t, J = 5.6 Hz, 1H), 4.01 (d, J = 9.6 Hz, 1H),

3.68–3.65 (m, 1H), 3.26–3.07 (m, 4H); MS (ESI) m/z 483 (M+H)⁺,

505 (M+Na)⁺.

4.2.23. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-pentyl-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (44)

Compound 44 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:

to prepare 61. The small amount of the

a

) and hexanehydrazide according to the procedure used

-anomer was removed by

a

reverse phase preparative HPLC. A white solid. Yield 20%. ¹H NMR

(400 MHz, DMSO-d₆) d 7.45 (d, J = 2.0 Hz, 1H), 7.40 (d, J = 8.0 Hz,

1H), 7.28 (dd, J = 8.0, 2.0 Hz, 1H), 4.90 (br, 2H), 4.80 (d, 1H), 4.49

(d, J = 15.6 Hz, 1H), 4.43 (d, J = 15.6 Hz, 1H), 4.39 (t, J = 6.0 Hz,

1H), 4.00 (d, J = 9.2 Hz, 1H), 3.69–3.65 (m, 1H), 3.45–3.40 (m,

1H), 3.24–3.04 (m, 4H), 2.96 (t, J = 7.6 Hz, 2H), 1.67–1.60 (m, 2H),

1.30–1.22 (m, 4H), 0.82 (t, J = 7.6 Hz, 3H); MS (ESI) m/z 443

(M+H)⁺, 465 (M+Na)⁺.

4.2.28. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(pyridin-2-yl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)tetrahydro-

2H-pyran-3,4,5-triol (53)

Compound 53 was synthesized from 21 and picolinohydrazide

according to the procedure used to prepare 61. A white solid. Yield

18%. ¹H NMR (400 MHz, DMSO-d₆) d 8.62 (dd, J = 3.6, 0.8 Hz, 1H),

8.19 (d, J = 7.6 Hz, 1H), 7.98 (td, J = 7.6, 1.6 Hz, 1H), 7.54–7.50 (m,

2H), 7.43 (d, J = 8.4 Hz, 1H), 7.31 (dd, J = 8.0, 1.6 Hz, 1H), 4.94–

4.91 (m, 2H), 4.84 (d, J = 5.6 Hz, 1H), 4.59 (d, J = 16.0 Hz, 1H),

4.54 (d, J = 16.0 Hz, 1H), 4.42 (t, J = 5.6 Hz, 1H), 4.02 (d, J = 9.6 Hz,

1H), 3.69–3.65 (m, 1H), 3.44–3.41 (m, 1H), 3.27–3.07 (m, 4H);

MS (ESI) m/z 450 (M+H)⁺, 472 (M+Na)⁺.

4.2.24. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-heptyl-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (45)

Compound 45 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:

to prepare 61. The small amount of the

a

) and octanehydrazide according to the procedure used

4.2.29. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(6-methylpyridin-3-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (55)

a-anomer was removed by

reverse phase preparative HPLC. A white solid. Yield 38%. ¹H NMR

(400 MHz, CD₃OD) d 7.52 (d, J = 1.6 Hz, 1H), 7.40–7.36 (m, 2H), 4.12

(d, J = 9.2 Hz, 1H), 3.86 (dd, J = 12.0, 2.0 Hz, 1H), 3.69 (dd, J = 12.0,

5.2 Hz, 1H), 3.47–3.38 (m, 3H), 3.25 (d, J = 8.8 Hz, 1H), 3.03 (t,

J = 7.6 Hz, 2H), 1.72 (quint, J = 7.6 Hz, 2H), 1.38–1.24 (m, 8H),

0.87 (t, J = 7.6 Hz, 3H); MS (ESI) m/z 471 (M+H)⁺, 493 (M+Na)⁺.

Compound 55 was synthesized from 21 and 6-meth-

ylnicotinohydrazide according to the procedure used to prepare

61. A white solid. Yield 17%. ¹H NMR (400 MHz, DMSO-d₆) d 8.92

(d, J = 2.0 Hz, 1H), 8.16 (dd, J = 8.0 Hz, 2.4 Hz, 1H), 7.52 (d,

J = 1.6 Hz, 1H), 7.48–7.37 (m, 2H), 7.32 (dd, J = 8.4 Hz, 2.0 Hz, 1H),

4.99–4.90 (m, 2H), 4.86 (d, J = 5.6 Hz, 1H), 4.65–4.53 (m, 2H),

4.43 (t, J = 6.0 Hz, 1H), 4.01 (d, J = 9.2 Hz, 1H), 3.71–3.63 (m, 1H),

3.47–3.05 (m, 5H), 2.50 (s, 3H); MS (ESI) m/z 464 (M+H)⁺.

4.2.25. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-cyclopentyl-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)tetrahydro-

2H-pyran-3,4,5-triol (50)

Compound 50 was synthesized from 21 and cyclopentanecarbo-

hydrazide according to the procedure used to prepare 61. A white

solid. Yield 46%. ¹H NMR (400 MHz, DMSO-d₆) d 7.45 (d, J = 1.6 Hz,

1H), 7.40 (d, J = 8.4 Hz, 1H), 7.29 (dd, J = 8.4, 2.0 Hz, 1H), 4.96–4.93

(m, 2H), 4.85 (d, J = 5.6 Hz, 1H), 4.50–4.41 (m, 3H), 4.00 (d,

J = 9.2 Hz, 1H), 3.69–3.65 (m, 1H), 3.47–3.40 (m, 2H), 3.26–3.03

(m, 4H), 2.09–2.04 (m, 2H), 1.69–1.57 (m, 6H); MS (ESI) m/z 441

(M+H)⁺, 463 (M+Na)⁺.

4.2.30. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(isoquinolin-3-yl)-

1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (57)

Compound 57 was synthesized from 21 and isoquinoline-3-car-

bohydrazide according to the procedure used to prepare 61. A

white solid. Yield 27%. ¹H NMR (400 MHz, DMSO-d₆) d 9.36 (s,

1H), 8.72 (s, 1H), 8.18 (d, J = 8.8 Hz, 2H), 7.86 (td, J = 7.6 Hz,

1.2 Hz, 1H), 7.76 (td, J = 8.0 Hz, 1.2 H, 1H), 7.55 (d, J = 2.0 Hz, 1H),

7.44 (d, J = 8.0 Hz, 1H), 7.32 (dd, J = 8.0 Hz, 2.0 Hz, 1H), 4.66–4.52

(m, 3H), 4.03 (d, J = 9.2 Hz, 1H), 3.68 (d, J = 10.8 Hz, 1H), 3.44 (dd,

J = 11.6 Hz, 5.6 Hz, 2H), 3.33–3.05 (m, 6H); MS (ESI) m/z 500

(M+H)⁺.

4.2.26. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-phenyl-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (51)

Compound 51 was synthesized from an anomeric mixture of 21

(ca. 2:1 b:

a

) and benzoic hydrazide according to the procedure

-anomer was re-

4.2.31. ((2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(quinolin-2-yl)-

1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (58)

used to prepare 61. The small amount of the

a

moved by reverse phase preparative HPLC. A white solid. Yield

27%. ¹H NMR (400 MHz, DMSO-d₆) d 7.89 (dd, J = 7.2, 2.0 Hz, 2H),

7.53–7.42 (m, 5H), 7.32 (dd, J = 8.0, 2.0 Hz, 1H), 5.03 (br, 2H),

4.94 (br, 1H), 4.59 (d, J = 16.0 Hz, 1H), 4.54 (d, J = 16.0 Hz, 1H),

4.50 (br, 1H), 4.03 (d, J = 9.6 Hz, 1H), 3.69 (d, J = 11.2 Hz, 1H),

3.46–3.42 (m, 1H), 3.28–3.13 (m, 3H), 3.09 (t, J = 8.8 Hz, 1H); ¹³C

NMR (100 MHz, DMSO-d₆) d 168.75, 140.69, 134.65, 132.43,

131.72, 131.31, 129.94, 129.87, 129.38, 129.26, 127.99, 81.70,

80.89, 78.68, 75.18, 70.62, 61.71, 34.08; MS (ESI) m/z 449

(M+H)⁺, 471 (M+Na)⁺.

Compound 58 was synthesized from 21 and quinoline-2-carbo-

hydrazide according to the procedure used to prepare 61. A white

solid. Yield 25%. ¹H NMR (400 MHz, DMSO-d₆) d 8.56 (d, J = 8.4 Hz,

1H), 8.31 (d, J = 8.8 Hz, 1H), 8.04 (t, J = 9.4 Hz, 2H), 7.81 (td, J = 8.0,

1.6 Hz, 1H), 7.67 (td, J = 7.2, 1.2 Hz, 1H), 7.56 (d, J = 2.0 Hz, 1H), 7.45

(d, J = 8.4 Hz, 1H), 7.33 (dd, J = 6.4, 2.0 Hz, 1H), 4.99–4.90 (m, 2H),

4.86 (d, J = 5.6 Hz, 1H), 4.67–4.55 (m, 2H), 4.43 (t, J = 5.6 Hz, 1H),

4.04 (d, J = 9.6 Hz, 1H), 3.67 (dd, J = 7.6, 3.6 Hz, 1H), 3.49–3.38 (m,

1H), 3.29–3.07 (m, 4H); MS (ESI) m/z 500 (M+H)⁺.

2190

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

4.2.32. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(furan-3-yl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)tetrahydro-

2H-pyran-3,4,5-triol (62)

4.2.37. ((2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(1-methyl-1H-

pyrrol-2-yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)-tetrahydro-2H-pyran-3,4,5-triol (68)

Compound 68 was synthesized from 21 and 1-methyl-1H-pyr-

role-2-carbohydrazide according to the procedure used to prepare

61. A white solid. Yield 18%. ¹H NMR (400 MHz, DMSO-d₆) d 7.46

(d, J = 2.0 Hz, 1H), 7.42 (d, J = 8.4 Hz, 1H), 7.30 (dd, J = 8.0, 2.0 Hz,

1H), 7.02 (t, J = 2.0 Hz, 1H), 6.65–6.60 (m, 1H), 6.13–6.07 (m, 1H),

4.57–4.42 (m, 4H), 4.01 (d, J = 9.6 Hz, 1H), 3.80 (s, 3H), 3.67 (dd,

J = 11.6, 1.6 Hz, 1H), 3.47–3.38 (m, 1H), 3.27–3.05 (m, 6H); MS

(ESI) m/z 452 (M+H)⁺.

Compound 62 was synthesized from 21 and 3-furoic hydra-

zide according to the procedure used to prepare 61. A white

solid. Yield 32%. ¹H NMR (400 MHz, DMSO-d₆)

d 8.44 (dd,

J = 1.2, 0.4 Hz, 1H), 7.83 (t, J = 2.0 Hz, 1H), 7.50 (d, J = 1.6 Hz,

1H), 7.42 (d, J = 8.4 Hz, 1H), 7.31 (dd, J = 8.4, 2.0 Hz, 1H), 6.96

(dd, J = 2.0, 0.8 Hz, 1H), 4.96–4.93 (m, 2H), 4.85 (d, J = 6.0 Hz,

1H), 4.56 (d, J = 16.0 Hz, 1H), 4.51 (d, J = 16.0 Hz, 1H), 4.44 (t,

J = 6.0 Hz, 1H), 4.01 (d, J = 9.6 Hz, 1H), 3.70–3.65 (m, 1H),

3.45–3.39 (m, 1H), 3.27–3.04 (m, 4H); MS (ESI) m/z 439

(M+H)⁺, 461 (M+Na)⁺.

4.2.38. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(1-methyl-1H-

indazol-3-yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)-tetrahydro-2H-pyran-3,4,5-triol (69)

Compound 69 was synthesized from 21 and 1-methyl-1H-inda-

zole-3-carbohydrazide according to the procedure used to prepare

61. A white solid. Yield 26%. ¹H NMR (400 MHz, DMSO-d₆) d 8.26

(d, J = 8.4 Hz, 1H), 7.74 (d, J = 8.8 Hz, 1H), 7.54–7.49 (m, 2H), 7.43

(d, J = 8.4 Hz, 1H), 7.36–7.30 (m, 2H), 4.94–4.91 (m, 2H), 4.85 (d,

J = 5.6 Hz, 1H), 4.61 (d, J = 15.6 Hz, 1H), 4.56 (d, J = 15.6 Hz, 1H),

4.43 (t, J = 5.6 Hz, 1H), 4.10 (s, 3H), 4.03 (d, J = 9.2 Hz, 1H), 3.70–

3.66 (m, 1H), 3.43 (quint, J = 5.6 Hz, 1H), 3.27–3.06 (m, 4H); MS

(ESI) m/z 503 (M+H)⁺.

4.2.33. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(3-methylfuran-2-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (63)

Compound 63 was synthesized from 21 and 3-methylfuran-2-

carbohydrazide according to the procedure used to prepare 61. A

white solid. Yield 28%. ¹H NMR (400 MHz, DMSO-d₆) d 7.80 (d,

J = 1.2 Hz, 1H), 7.49 (d, J = 2.0 Hz, 1H), 7.41 (d, J = 8.4 Hz, 1H),

7.30 (dd, J = 8.0, 2.0 Hz, 1H), 6.61 (d, J = 1.2 Hz, 1H), 4.97–4.80

(m, 2H), 4.83 (d, J = 5.2 Hz, 1H), 4.60–4.50 (m, 1H), 4.47–4.39

(m, 1H), 4.01 (d, J = 9.6 Hz, 1H), 3.71–3.62 (m, 1H), 3.48–3.37

(m, 1H), 3.33–3.03 (m, 4H), 2.32 (s, 3H); MS (ESI) m/z 453

(M+H)⁺.

4.2.39. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(thiophen-2-yl)-

1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (70)

Compound 70 was synthesized from 21 and thiophene-2-carbo-

hydrazide according to the procedure used to prepare 61. A white

solid. Yield 47%. ¹H NMR (400 MHz, DMSO-d₆) d 7.78 (dd, J = 4.8,

1.2 Hz, 1H), 7.69 (dd, J = 3.6, 1.2 Hz, 1H), 7.51 (d, J = 2.4 Hz, 1H),

7.43 (d, J = 8.0 Hz, 1H), 7.31 (dd, J = 8.0, 2.0 Hz, 1H), 7.16 (dd,

J = 8.0, 4.0 Hz, 1H), 4.97–4.93 (m, 2H), 4.86 (d, J = 6.0 Hz, 1H),

4.57 (d, J = 16.0 Hz, 1H), 4.52 (d, J = 15.6 Hz, 1H), 4.45 (t,

J = 5.6 Hz, 1H), 4.02 (d, J = 9.2 Hz, 1H), 3.70–3.65 (m, 1H), 3.46–

3.39 (m, 1H), 3.27–3.05 (m, 4H); MS (ESI) m/z 455 (M+H)⁺, 477

(M+Na)⁺.

4.2.34. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(5-methylfuran-2-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (64)

Compound 64 was synthesized from 21 and 5-methylfuran-2-

carbohydrazide according to the procedure used to prepare 61. A

white solid. Yield 30%. ¹H NMR (400 MHz, DMSO-d₆) d 7.49 (d,

J = 2.0 Hz, 1H), 7.42 (d, J = 8.4 Hz, 1H), 7.30 (dd, J = 8.4, 2.0 Hz,

1H), 7.11 (d, J = 3.6 Hz, 1H), 6.32 (d, J = 3.6 Hz, 1H), 4.94–4.91 (m,

2H), 4.83 (d, J = 5.6 Hz, 1H), 4.56 (d, J = 16.0 Hz, 1H), 4.51 (d,

J = 16.0 Hz, 1H), 4.42 (t, J = 5.6 Hz, 1H), 4.01 (d, J = 9.6 Hz, 1H),

3.70–3.65 (m, 1H), 3.45–3.40 (m, 1H), 3.27–3.04 (m, 4H), 2.32 (s,

3H); MS (ESI) m/z 453 (M+H)⁺.

4.2.40. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(thiophen-3-yl)-

1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (71)

4.2.35. (2S,3R,4R,5S,6R)-2-(3-((5-(Benzofuran-2-yl)-1,3,4-

thiadiazol-2-yl)methyl)-4-chlorophenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (66)

Compound 71 was synthesized from 21 and thiophene-3-carbo-

hydrazide according to the procedure used to prepare 61. A white

solid. Yield 66%. ¹H NMR (400 MHz, DMSO-d₆) d 8.22 (dd, J = 2.8,

1.2 Hz, 1H), 7.72–7.69 (m, 1H), 7.58 (dd, J = 4.8, 1.2 Hz, 1H), 7.51

(d, J = 1.6 Hz, 1H), 7.42 (d, J = 8.0 Hz, 1H), 7.31 (dd, J = 8.0, 1.6 Hz,

1H), 4.94–4.91 (m, 2H), 4.84 (d, J = 6.0 Hz, 1H), 4.57 (d,

J = 16.0 Hz, 1H), 4.52 (d, J = 16.0 Hz, 1H), 4.42 (t, J = 6.0 Hz, 1H),

4.01 (d, J = 9.6 Hz, 1H), 3.70–3.65 (m, 1H), 3.46–3.40 (m, 1H),

3.25–3.05 (m, 4H); ¹³C NMR (100 MHz, DMSO-d₆) d 166.40,

162.06, 139.16, 133.14, 130.89, 129.85, 129.74, 127.85, 127.68,

127.36, 126.69, 125.08, 80.14, 79.35, 77.19, 73.65, 69.18, 60.24,

32.47; MS (ESI) m/z 455 (M+H)⁺, 477 (M+Na)⁺; HRMS (FAB) m/z

455.0501 [calcd for C₁₉H₂₀ClN₂O₅S₂(M+H)⁺455.0502].

Compound 52 was synthesized from an anomeric mixture of

21 (ca. 2:1 b:a) and benzofuran-2-carbohydrazide according to

the procedure used to prepare 61. The small amount of the

a

-

anomer was removed by reverse phase preparative HPLC. A white

solid. Yield 50%. ¹H NMR (400 MHz, CD₃OD) d 7.71 (d, J = 8.0 Hz,

1H), 7.60–7.52 (m, 3H), 7.44–7.39 (m, 3H), 7.31 (t, J = 7.6 Hz,

2H), 4.15 (d, J = 9.2 Hz, 1H), 3.87 (dd, J = 12.0, 1.6 Hz, 1H), 3.70

(dd, J = 12.0, 5.2 Hz, 3.48–3.38 (m, 3H), 3.33 (s, 2H); MS (ESI)

m/z 489 (M+H)⁺.

4.2.36. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(5-methylisoxazol-3-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (67)

4.2.41. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(3-methylthiophen-

2-yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (72)

Compound 72 was synthesized from 21 and 3-methylthioph-

ene-2-carbohydrazide according to the procedure used to prepare

61. A white solid. Yield 45%. ¹H NMR (400 MHz, DMSO-d₆) d 7.67

(d, J = 5.2 Hz, 1H), 7.50 (d, J = 1.6 Hz, 1H), 7.42 (d, J = 8.4 Hz, 1H),

7.30 (dd, J = 8.4, 1.6 Hz, 1H), 7.06 (d, J = 5.2 Hz, 1H), 4.96(m, 2H),

4.85 (d, J = 5.6 Hz, 1H), 4.59 (d, J = 16.0 Hz, 1H), 4.55 (d,

J = 16.0 Hz, 1H), 4.44 (t, J = 5.6 Hz, 1H), 4.01 (d, J = 9.2 Hz, 1H),

Compound 67 was synthesized from 21 and 5-methylfuran-2-

carbohydrazide according to the procedure used to prepare 61. A

white solid. Yield 16%. ¹H NMR (400 MHz, DMSO-d₆) d 7.52 (d,

J = 2.0 Hz, 1H), 7.43 (d, J = 8.0 Hz, 1H), 7.31 (dd, J = 8.0, 1.6 Hz,

1H), 6.87 (s, 1H), 4.93 (t, J = 4.8 Hz, 2H), 4.85 (d, J = 5.6 Hz, 1H),

4.64 (d, J = 16.0 Hz, 1H), 4.59 (d, J = 16.0 Hz, 1H), 4.42 (t,

J = 6.0 Hz, 1H), 4.02 (d, J = 9.2 Hz, 1H), 3.69–3.65 (m, 1H), 3.46–

3.39 (m, 1H), 3.29–3.05 (m, 4H), 2.48 (s, 3H); MS (ESI) m/z 454

(M+H)⁺, 476 (M+Na)⁺.

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

2191

3.69–3.65 (m, 1H), 3.45–3.39 (m, 1H), 3.27–3.04 (m, 4H), 2.37 (s,

4.2.47. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(3,5-

dimethylisoxazol-4-yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)-tetrahydro-2H-pyran-3,4,5-triol (78)

3H); MS (ESI) m/z 489 (M+H)⁺.

4.2.42. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(3-chlorothiophen-2-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (73)

Compound 78 was synthesized from 21 and 3,5-dimethylisoxa-

zole-4-carbohydrazide according to the procedure used to prepare

61. A white solid. Yield 9.5%. ¹H NMR (400 MHz, DMSO-d₆) d 7.50

(d, J = 2.0 Hz, 1H), 7.42 (d, J = 8.4 Hz, 1H), 7.30 (dd, J = 8.4, 2.0 Hz,

1H), 4.74 (br, 4H), 4.61 (d, J = 16.0 Hz, 1H), 4.56 (d, J = 16.0 Hz,

1H), 4.01 (d, J = 9.6 Hz, 1H), 3.67 (dd, J = 11.6, 1.6 Hz, 1H), 3.44–

3.40 (m, 1H), 3.26–3.19 (m, 2H), 3.16–3.11 (m, 1H), 3.07 (t,

J = 9.2 Hz, 1H), 2.60 (s, 3H), 2.39 (s, 3H); MS (ESI) m/z 468 (M+H)⁺.

Compound 73 was synthesized from 21 and 3-chlorothiophene-

2-carbohydrazide according to the procedure used to prepare 61. A

white solid. Yield 49%. ¹H NMR (400 MHz, DMSO-d₆) d 7.90 (d,

J = 5.2 Hz, 1H), 7.51 (d, J = 2.0 Hz, 1H), 7.43 (d, J = 8.4 Hz, 1H),

7.31 (dd, J = 8.0, 2.0 Hz, 1H), 7.27 (d, J = 5.6 Hz, 1H), 4.92–4.90 (m,

2H), 4.82 (d, J = 5.6 Hz, 1H), 4.62 (d, J = 16.0 Hz, 1H), 4.58 (d,

J = 16.0 Hz, 1H), 4.40 (t, J = 6.0 Hz, 1H), 4.02 (d, J = 9.2 Hz, 1H),

3.70–3.65 (m, 1H), 3.46–3.40 (m, 1H), 3.25–3.05 (m, 4H); MS

(ESI) m/z 489 (M+H)⁺.

4.2.48. ((2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(3-ethyl-5-

methylisoxazol-4-yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)-tetrahydro-2H-pyran-3,4,5-triol (79)

Compound 79 was synthesized from 21 and 3-ethyl-5-methy-

lisoxazole-4-carbohydrazide according to the procedure used to

prepare 61. A white solid. Yield 12%. ¹H NMR (400 MHz, DMSO-

d₆) d 7.50 (d, J = 2.0 Hz, 1H), 7.42 (d, J = 8.4 Hz, 1H), 7.30 (dd,

J = 8.4, 2.0 Hz, 1H), 4.75 (br, 4H), 4.61 (d, J = 16.0 Hz, 1H), 4.57 (d,

J = 16.0 Hz, 1H), 4.01 (d, J = 9.2 Hz, 1H), 3.67 (d, J = 10.0 Hz, 1H),

3.44–3.40 (m, 1H), 3.26–3.19 (m, 2H), 3.16–3.11 (m, 1H), 3.07 (t,

J = 9.2 Hz, 1H), 2.85 (q, J = 7.2 Hz, 2H), 2.59 (s, 3H), 1.75 (t,

J = 7.2 Hz, 3H); MS (ESI) m/z 468 (M+H)⁺.

4.2.43. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(3-chlorothiophen-2-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (74)

Compound 74 was synthesized from 21 and 5-chlorothio-

phene-2-carbohydrazide according to the procedure used to pre-

pare 61. A white solid. Yield 36%. ¹H NMR (400 MHz, DMSO-d₆)

d

7.59 (d, J = 4.0 Hz, 1H), 7.47 (d, J = 2.0 Hz, 1H), 7.39 (d,

J = 8.4 Hz, 1H), 7.27 (dd, J = 8.4, 2.0 Hz, 1H), 7.19 (d, J = 4.0 Hz,

1H), 4.64–4.38 (m, 5H), 3.97 (d, J = 9.6 Hz, 1H), 3.64 (d,

J = 8.0 Hz, 1H), 3.42–3.36 (m, 1H), 3.25–3.10 (m, 5H); MS (ESI)

m/z 489 (M+H)⁺.

4.2.49. 2-(2-Chloro-5-((2S,3S,4R,5R,6R)-3,4,5-tris(benzyloxy)-6-

(benzyloxymethyl)-tetrahydro-2H-pyran-2-

yl)phenyl)acetohydrazide (26)

4.2.44. (2S,3R,4R,5S,6R)-2-(3-((5-(Benzo[b]thiophen-2-yl)-1,3,4-

thiadiazol-2-yl)methyl)-4-chlorophenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (75)

To a mixture of acid 21 (693 mg, 1.0 mmol), EDCI (249 mg,

1.3 mmol), and HOBt hydrate (233 mg, 1.5 mmol) in DMF

(10 mL) was added hydrazine monohydrate (2 mL). The resulting

mixture was stirred at ambient temperature for 12 h and then

poured into water. The resulting solid was isolated by ﬁltration

and dried in vacuo to yield (674 mg, 95%) a white solid. ¹H NMR

(400 MHz, DMSO-d₆) d 9.17 (s, 1H), 7.39–7.14 (m, 21H), 6.88–

6.86 (m, 2H), 4.80 (d, J = 11.2 Hz, 1H). 4.77 (d, J = 11.2 Hz, 1H),

4.70 (d, J = 13.6 Hz, 1H), 4.54 (d, J = 10.8 Hz, 1H), 4.51 (d,

J = 12.4 Hz, 1H), 4.44 (d, J = 12.4 Hz, 1H), 4.32 (d, J = 10.8 Hz, 1H),

4.26 (d, J = 9.6 Hz, 1H), 3.83 (d, J = 10.8 Hz, 1H), 3.73 (t, J = 8.8 Hz,

1H), 3.66–3.45 (m, 7H); MS (ESI) m/z 707 ([M+H]⁺), 729 (M+Na)⁺.

Compound 75 was synthesized from 21 and benzo[b]thio-

phene-2-carbohydrazide according to the procedure used to

prepare 61.

DMSO-d₆)

A

white solid. Yield 36%. ¹H NMR (400 MHz,

8.11 (s, 1H), 8.02 (d, J = 7.2 Hz, 1H), 7.87 (d,

d

J = 7.2 Hz, 1H), 7.54 (d, J = 2.0 Hz, 1H), 7.47–7.39 (m, 3H),

7.32 (dd, J = 8.4, 2.0 Hz, 1H), 4.95 (br, 2H), 4.87 (d, J = 5.6 Hz,

1H), 4.62 (d, J = 16.0 Hz, 1H), 4.57 (d, J = 16.0 Hz), 4.45 (t,

J = 5.6 Hz, 1H), 4.02 (d, J = 9.2 Hz, 1H), 3.70–3.66 (m, 1H),

3.46–3.40 (m, 1H), 3.26–3.06 (m, 4H); MS (ESI) m/z 505

(M+H)⁺.

4.2.50. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(5-phenylfuran-2-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (65)

4.2.45. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(thiazol-4-yl)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)tetrahydro-

2H-pyran-3,4,5-triol (76)

To a mixture of hydrazide 32 (674 mg, 0.95 mmol), 5-phenyl-2-

furoic acid (233 mg, 1.24 mmol), EDCI (273 mg, 1.43 mmol), and

HOBt hydrate (295 mg, 1.90 mmol) in DMF (10 mL) was added

NMM (0.31 mL, 2.85 mmol). The resulting mixture was stirred at

ambient temperature overnight. After dilution with EtOAc, the or-

ganic layer was subsequently washed with water, hydrochloric

acid (1.0 N), saturated sodium bicarbonate, and brine prior to

drying over anhydrous MgSO₄. Filtration and removal of the vola-

tiles under reduced pressure yielded a glassy yellow amorphous

solid, which was carried on to the next step without further

puriﬁcation.

Compound 76 was synthesized from 21 and thiazole-4-carbo-

hydrazide according to the procedure used to prepare 61. A white

solid. Yield 27%. ¹H NMR (400 MHz, DMSO-d₆) d 9.23 (d, J = 2.4 Hz,

1H), 8.49 (d, J = 2.0 Hz, 1H), 7.51 (d, J = 2.0 Hz, 1H), 7.42 (d,

J = 8.4 Hz, 1H), 7.30 (dd, J = 8.0, 2.0 Hz, 1H), 4.93–4.91 (m, 2H),

4.83 (d, J = 5.6 Hz, 1H), 4.59 (d, J = 15.6 Hz, 1H), 4.54 (d,

J = 15.6 Hz, 1H), 4.42 (t, J = 5.6 Hz, 1H), 4.01 (d, J = 9.2 Hz, 1H),

3.69–3.64 (m, 1H), 3.45–3.39 (m, 1H), 3.26–3.05 (m, 4H); MS

(ESI) m/z 456 (M+H)⁺, 478 (M+Na)⁺.

4.2.46. ((2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(2-(pyridin-4-

yl)thiazol-5-yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxylmethyl)-tetrahydro-2H-pyran-3,4,5-triol (77)

Compound 77 was synthesized from 21 and 2-(pyridin-4-yl)thi-

azole-4-carbohydrazide according to the procedure used to pre-

pare 61. A white solid. Yield 19%. ¹H NMR (400 MHz, DMSO-d₆) d

8.73 (d, J = 6.0 Hz, 2H), 8.68 (s, 1H), 7.96 (d, J = 4.4 Hz, 2H), 7.54

(d, J = 2.0 Hz, 1H), 7.44 (d, J = 8.4 Hz, 1H), 7.32 (dd, J = 8.0, 1.6 Hz,

1H), 4.67–4.51 (m, 5H), 4.02 (d, J = 9.2 Hz, 1H), 3.67 (d,

J = 10.4 Hz, 1H), 3.49–3.39 (m, 1H), 3.27–3.07 (m, 5H); MS (ESI)

m/z 533 (M+H)⁺.

To a solution of the acylhydrazide in THF (20 mL) was added

Lawesson reagent (1.2 g, 2.9 mmol). The reaction mixture was re-

ﬂuxed overnight. After cooling to room temperature, the reaction

mixture was diluted with EtOAc and washed with saturated so-

dium bicarbonate. The organic layer was dried over anhydrous

MgSO₄, ﬁltered through a plug of silica gel and concentrated in va-

cuo. The resulting crude residue was carried on to the next step

without further puriﬁcation.

To a solution of the perbenzylated thiadiazole in acetonitrile

(5 mL) was added TMSI (5 mL). The resulting reaction mixture

was heated to 50 °C overnight. After cooling to 0 °C, the

2192

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

reaction quenched with methanol and concentrated in vacuo.

The resulting crude residue was puriﬁed by reverse phase pre-

parative HPLC to provide the title compound (176 mg, 36%) as a

white solid. ¹H NMR (400 MHz, DMSO-d₆) d 7.79–7.77 (m, 2H),

7.52 (d, J = 2.0 Hz, 1H), 7.46–7.31 (m, 6H), 7.20 (d, J = 3.6 Hz,

1H), 4.95 (dd, J = 6.8, 4.8 Hz, 2H), 4.87 (d, J = 5.6 Hz, 1H), 4.61

(d, J = 16.0 Hz, 1H), 4.56 (d, J = 16.0 Hz, 1H), 4.50 (t, J = 6.0 Hz,

1H). 4.02 (d, J = 9.6 Hz, 1H), 3.68–3.62 (m, 1H), 3.43–3.40 (m,

4.2.54. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(methylthio)-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (47)

To a solution of chlorothiadiazole 29 (419 mg, 0.55 mmol in THF

(10 mL) was added sodium methanethiolate (77 mg, 1.1 mmol).

The resulting mixture was stirred at ambient temperature over-

night. After cooling to 0 °C, the reaction was quenched with hydro-

chloric acid (1.0 N) and diluted with EtOAc and water. The organic

layer was separated and the aqueous layer was extracted with

EtOAc. The combined organic layers were washed with brine, dried

over anhydrous MgSO₄, ﬁltered and concentrated in vacuo. The

resulting residue was carried on to the next step without further

puriﬁcation.

1H), 3.27–3.06 (m, 4H); ¹³C NMR (100 MHz, DMSO-d₆)

d

167.63, 158.60, 155.97, 144.51, 143.71, 140.72, 134.63, 132.43,

131.30, 129.52, 129.41, 129.29, 129.14, 124.34, 114.87, 109.08,

81.68, 80.90, 78.73, 75.19, 70.71, 61.78, 33.88; MS (ESI) m/z

515 (M+H)⁺.

To a solution of the perbenzylated thiadiazole in acetonitrile

(5 mL) was added TMSI (5 mL). The resulting reaction mixture

was heated to 50 °C overnight. After cooling to 0 °C, the reaction

quenched with methanol and concentrated in vacuo. The resulting

crude residue was puriﬁed by reverse phase preparative HPLC to

yield the title compound (58 mg, 25%) as a white solid. ¹H NMR

(400 MHz, DMSO-d₆) d 7.45 (d, J = 2.0 Hz, 1H), 7.40 (d, J = 8.0 Hz,

1H), 7.29 (dd, J = 8.0, 2.0 Hz, 1H), 4.49 (d, J = 15.6 Hz, 1H), 4.44 (d,

J = 15.6 z, 1H), 3.99 (d, J = 9.6 Hz, 1H), 3.67 (dd, J = 12.0, 2.0 Hz,

1H), 3.42 (dd, J = 11.6, 5.6 Hz, 1H), 3.25–3.11 (m, 3H), 3.06 (t,

J = 9.2 Hz, 1H), 2.67 (s, 3H); MS (ESI) m/z 419 (M+H)⁺, 441 (M+Na)⁺.

4.2.51. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(triﬂuoromethyl)-

1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol (48)

Compound 48 was synthesized from 26 and triﬂuoroacetic acid

according to the procedure used to prepare 65. A white solid. Yield

56%. ¹H NMR (400 MHz, DMSO-d₆) d 7.53 (d, J = 2.0 Hz, 1H), 7.43 (d,

J = 8.4 Hz, 1H), 7.33 (dd, J = 8.4, 2.4 Hz, 1H), 4.95–4.91 (m, 2H), 4.85

(d, J = 5.6 Hz, 1H), 4.71 (d, J = 16.4 Hz, 1H), 4.67 (d, J = 16.4 Hz, 1H),

4.41 (t, J = 6.0 Hz, 1H), 4.01 (d, J = 9.2 Hz, 1H), 3.69–3.65 (m, 1H),

3.46–3.40 (m, 1H), 3.25–3.04 (m, 4H); MS (ESI) m/z 441 (M+H)⁺,

463 (M+Na)⁺.

4.2.55. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-methoxy-1,3,4-

thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-tetrahydro-

2H-pyran-3,4,5-triol (46)

4.2.52. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(2,2,2-

triﬂuoroethyl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-

(hydroxymethyl)-tetrahydro-2H-pyran-3,4,5-triol (49)

Compound 49 was synthesized from 26 and 3,3,3-triﬂuoro-

propanoic acid according to the procedure used to prepare 65.

A white solid. Yield 61%. ¹H NMR (400 MHz, DMSO-d₆) d 7.50

(d, J = 1.6 Hz, 1H), 7.41 (d, J = 8.4 Hz, 1H), 7.29 (dd, J = 8.4,

1.6 Hz, 1H), 4.97–4.94 (m, 2H), 4.85 (d, J = 6.0 Hz, 1H), 4.56 (d,

J = 16.0 Hz, 1H), 4.51 (d, J = 16.0 Hz, 1H), 4.44 (t, J = 6.0 Hz, 1H),

4.33 (q, J = 10.8 Hz, 2H), 4.00 (d, J = 9.2 Hz, 1H), 3.69–3.63 (m,

1H), 3.45–3.41 (m, 1H), 3.27–3.03 (m, 4H); MS (ESI) m/z 455

(M+H)⁺, 477 (M+Na)⁺.

Compound 46 was synthesized from 29 and sodium methoxide

according to the procedure used to prepare 47. A white solid. Yield

57%. ¹H NMR (400 MHz, DMSO-d₆) d 7.42–7.39 (m, 2H), 7.29 (dd,

J = 8.4, 2.0 Hz, 1H), 4.97–4.94 (m, 2H), 4.85 (d, J = 5.6 Hz, 1H),

4.44 (t, J = 5.6 Hz, 1H), 4.37 (d, J = 15.6 Hz, 1H), 4.32 (d,

J = 15.6 Hz, 1H), 4.01 (d, J = 9.2 Hz, 1H), 3.69–3.65 (m, 1H), 3.45–

3.40 (m, 1H), 3.27–3.03 (m, 4H), 3.39(s, 3H); MS (ESI) m/z 403

(M+H)⁺, 425 (M+Na)⁺.

4.2.56. 2-(2-Chloro-5-((2S,3S,4R,5R,6R)-3,4,5-tris(benzyloxy)-6-

(benzyloxymethyl)-tetrahydro-2H-pyran-2-yl)benzyl)-5-

(pyrazin-2-yl)-1,3,4-thiadiazole (59)

4.2.53. (2-Chloro-5-(2-chloro-5-((2S,3S,4R,5R,6R)-3,4,5-

tris(benzyloxy)-6-(benzyloxymethyl)-tetrahydro-2H-pyran-2-

yl)-benzyl)-1,3,4-thiadiazole (29)

To a solution of acid 21 (2.8 g, 4.0 mmol) in 1,4-dioxane (30 mL)

was added thiosemicarbazide 28 (0.40 g, 4.4 mmol). The solution is

stirred for 1 h at 90 °C, before adding of phosphorus oxychloride

(0.41 mL, 4.4 mmol). After overnight stirring at 110 °C, the reaction

was concentrated in vacuo to yield a glassy off-white amorphous

solid, which was carried on to the next step without further

puriﬁcation.

To a mixture of the thiadiazole amine and copper (0.04 g,

0.62 mmol) in c-hydrochloric acid (6 mL) and acetic acid

(30 mL) at 0 °C was slowly added sodium nitrite (2.8 g, 40 mmol)

while maintaining an internal reaction temperature below 10 °C.

After 1 h stirring at 0 °C, the resulting mixture was stirred with

gradual warming to ambient temperature over 2 h. The mixture

was diluted with water and extracted with EtOAc. The organic

layer was dried over anhydrous MgSO₄, ﬁltered and evaporated

in vacuo. Puriﬁcation was accomplished by column chromatogra-

phy (10–60% EtOAc/hexanes) to yield the title compound (0.66 g,

022%) as a yellow oil. ¹H NMR (400 MHz, CDCl₃) d 7.48 (s, 1H),

7.37–7.17 (m, 20H), 6.93–6.91 (m, 2H), 4.90 (s, 2H), 4.85(d,

J = 10.8 Hz, 1H), 4.63–4.51 (m, 4H), 4.45 (d, J = 10.8 Hz, 1H), 4.20

(d, J = 9.6 Hz, 1H), 3.88 (d, J = 10.8 Hz, 1H), 3.81–3.72 (m, 5H),

3.60–3.57 (m, 1H), 3.43 (t, J = 8.8 Hz, 1H); MS (ESI) m/z 767

(M+H)⁺, 789 (M+Na)⁺.

To a mixture of acid 21 (1.00 g, 1.44 mmol), pyrazine-2-carbo-

hydrazide 30 (259 mg, 1.88 mmol), EDCI (415 mg, 2.16 mmol),

and HOBt hydrate (448 mg, 2.89 mmol) in DMF (10 mL) was added

NMM (0.476 mL, 4.33 mmol). The resulting mixture was stirred at

ambient temperature overnight. After dilution with EtOAc, the or-

ganic layer was subsequently washed with water, hydrochloric

acid (1.0 N), saturated sodium bicarbonate, and brine prior to dry-

ing over anhydrous MgSO₄. Filtration and removal of the volatiles

under reduced pressure yielded a glassy off-white amorphous so-

lid, which was carried on to the next step without further

puriﬁcation.

To a solution of the acylhydrazide in THF (20 mL) was added

Lawesson reagent (874 mg, 2.16 mmol). The reaction mixture

was reﬂuxed overnight. After cooling to room temperature, the

reaction mixture was diluted with EtOAc and washed with satu-

rated sodium bicarbonate. The organic layer was dried over anhy-

drous MgSO₄, ﬁltered through a plug of silica gel and concentrated

in vacuo. The resulting crude residue was carried on to the next

step without any puriﬁcation.

To a solution of perbenzylated pyrazine at ꢀ50 °C in Ac₂O

(10 mL) was slowly added a solution of trimethylsilyl triﬂuoro-

methanesulfonate (1.5 mL, 8.4 mmol) in Ac₂O (5 mL). The resulting

mixture was stirred with gradual warming to ambient temperature

over 12 h, cooled to 0 °C, and quenched with saturated sodium

bicarbonate. After dilution with EtOAc, the organic layer was

washed with sodium bicarbonate and brine prior to drying over

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

2193

anhydrous MgSO₄. Filtration and removal of volatiles under re-

duced pressure yielded peracetylated compound as a yellow amor-

phous solid. Deacetylation was achieved by addition of sodium

methoxide (25 wt.% in methanol, 0.5 mL) to a solution of this resi-

due in methanol (5 mL). The reaction mixture was stirred at ambi-

ent temperature for 2 h. Amberlite IR120 hydrogen form was then

added to neutralize the reaction mixture. The resin was ﬁltered off

and the ﬁltrate was concentrated in vacuo. The resulting crude res-

idue was puriﬁed by reverse phase preparative HPLC to yield the

title compound (47 mg, 10%) as a white solid. ¹H NMR (400 MHz,

DMSO-d₆) d 9.39 (d, J = 1.6 Hz, 1H), 8.78 (d, J = 2.4 Hz, 1H), 8.73

(dd, J = 2.4, 1.6 Hz, 1H), 7.53 (d, J = 1.6 Hz, 1H), 7.43 (d, J = 8.4 Hz,

1H), 7.31 (dd, J = 8.4, 1.6 Hz, 1H), 4.50 (br, 2H), 4.87 (d, J = 5.6 Hz,

1H), 4.64 (d, J = 16.4 Hz, 1H), 4.59 (d, J = 16.4 Hz, 1H), 4.43 (t,

J = 5.6 Hz, 1H), 4.02 (d, J = 9.2 Hz, 1H), 3.69–3.65 (m, 1H), 3.44–

3.41 (m, 1H), 3.26–3.07 (m, 4H); ¹³C NMR (100 MHz, DMSO-d₆) d

171.09, 168.42, 147.00, 145.34, 144.40, 141.88, 140.72, 134.59,

132.40, 131.24, 129.35, 129.32, 81.67, 80.92, 78.71, 75.21, 70.70,

61.76, 34.10, MS (ESI) m/z 451 (M+H)⁺; HRMS (FAB) m/z

451.0842 [calcd for C₁₉H₂₀ClN₄O₅S (M+H)⁺451.0843]

hSGLT2 sequence was cloned into pcDNA3.1(+) for mammalian

expression and were stably transfected into chinese hamster ovary

(CHO) cells. SGLT2-expressing clones were selected based on resis-

tance to G418 antibiotic (Geneticin^Ò, Invitrogen, Carlsbad, CA) and

activity in the ¹⁴C-

assay.

a-methyl-D

-glucopyranoside (¹⁴C-AMG) uptake

4.3.2. Inhibitory effects on human SGLT2 activities

For sodium-dependent glucose transport assay, cells expressing

hSGLT2 were seeded into a 96-well culture plate at a density of

5 ꢂ 10⁴cells/well in RPMI medium 1640 containing 10% fetal bo-

vine serum. The cells were used 1 day after plating. They were

incubated in pretreatment buffer (10 mM HEPES, 5 mM Tris,

140 mM choline chloride, 2 mM KCl, 1 mM CaCl₂, and 1 mM MgCl₂,

pH 7.4) at 37 °C for 10 min. They were then incubated in uptake

buffer (10 mM HEPES, 5 mM Tris, 140 mM NaCl, 2 mM KCl, 1 mM

CaCl₂, 1 mM MgCl₂, and 1 mM ¹⁴C-nonlabeled AMG pH 7.4) con-

taining ¹⁴C-labeled (8

lM) and inhibitor or dimethyl sulfoxide

(DMSO) vehicle at 37 °C for 2 h. Cells were washed twice with

washing buffer (pretreatment buffer containing 10 mM AMG at

room temperature) and then the radioactivity was measured using

a liquid scintillation counter. IC₅₀was determined by nonlinear

regression analysis using GraphPad PRISM.^24,25

4.2.57. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(6-methylpyridin-2-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (54)

Compound 54 was synthesized from 21 and 6-meth-

ylpicolinohydrazide according to the procedure used to prepare

59. A white solid. Yield 31%. ¹H NMR (400 MHz, DMSO-d₆) d 7.99

(d, J = 7.6 Hz, 1H), 7.86 (t, J = 7.6 Hz, 1H), 7.52 (d, J = 1.6 Hz, 1H),

7.43 (d, J = 8.4 Hz, 1H), 7.38 (d, J = 7.6 Hz, 1H), 7.31 (dd, J = 8.4,

1.6 Hz, 1H), 4.96–4.93 (m, 2H), 4.86 (d, J = 5.6 Hz, 1H), 4.58 (d,

J = 15.6 Hz, 1H), 4.53 (d, J = 15.6 Hz, 1H), 4.44 (t, J = 5.6 Hz, 1H),

4.02 (d, J = 9.6 Hz, 1H), 3.69–3.65 (m, 1H), 3.45–3.40 (m, 1H), 3.27–

3.05 (m, 4H), 2.47 (s, 3H); MS (ESI) m/z 464 (M+H)⁺, 486 (M+Na)⁺.

4.4. Normal and diabetic animal studies

4.4.1. Animals

Male SD rats and diabetic db/db mice were purchased by

Charles River Laboratory. All animals were housed at 23 2 °C un-

der a 12-h light/dark cycle (light on 7:00) and were fed a standard

chow and water ad libitum.

4.4.2. Urinary glucose excretion in normal animal^24–26

For glucosuria assessment, overnight-fasted SD rats (5 weeks of

ages) were placed into metabolism cages for baseline urine collec-

tion over 24 h. Rats were weighted, randomized into experimental

groups (n = 4) and orally administered with 50% aqueous glucose

solution (2 g/kg) and drugs. Rats were returned to metabolism

cages for 24 h urine collection. After the urine volume had been

measured, the glucose concentration in the urine was determined

using a LabAssay™ (Wako Pure Chemicals). These data were nor-

malized per 200 g body weight.

4.2.58. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(2-methylpyridin-4-

yl)-1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (56)

Compound 56 was synthesized from 21 and 2-meth-

ylisonicotinohydrazide according to the procedure used to prepare

59. A white solid. Yield 45%. ¹H NMR (400 MHz, DMSO-d₆) d 8.56

(d, J = 4.8 Hz, 1H), 7.74 (s, 1H), 7.67 (d, J = 4.8 Hz, 1H), 7.53 (d,

J = 1.6 Hz, 1H), 7.44 (d, J = 8.0 Hz, 1H), 7.32 (dd, J = 8.0, 1.6 Hz,

1H), 4.98–4.94 (m, 2H), 4.87 (d, J = 6.0 Hz, 1H), 4.64 (d,

J = 16.0 Hz, 1H), 4.53 (d, J = 16.0 Hz, 1H), 4.44 (t, J = 5.6 Hz, 1H),

4.01 (d, J = 9.2 Hz, 1H), 3.68–3.66 (m, 1H), 3.45–3.39 (m, 1H),

3.27–3.05 (m, 4H), 2.56 (s, 3H); MS (ESI) m/z 464 (M+H)⁺, 486

(M+Na)⁺.

4.4.3. Blood glucose effects in diabetic db/db mice^24–26

For assessment of acute blood glucose effects in db/db mice, the

animals were weighed and randomized into two groups (n = 4).

Mice were dosed with vehicle or drug (9@1 mg/kg, 61@10 mg/kg)

and blood glucose level were measured immediately before dosing

and at 1, 2, 4, 5, 6, and 24 h post-dose using blood glucose meter

(Lifescan, a Johnson & Johnson). The animals were allowed to re-

feed after the 6-h time point.

4.2.59. (2S,3R,4R,5S,6R)-2-(4-Chloro-3-((5-(pyridazin-4-yl)-

1,3,4-thiadiazol-2-yl)methyl)phenyl)-6-(hydroxymethyl)-

tetrahydro-2H-pyran-3,4,5-triol (60)

Compound 60 was synthesized from 21 and pyridazine-4-carbo-

hydrazide according to the procedure used to prepare 59. A white

Acknowledgments

solid. Yield 21%. ¹H NMR (400 MHz, DMSO-d₆)

d 9.71 (d,

J = 2.4 Hz, 1H), 9.51 (d, J = 5.6 Hz, 1H), 8.15 (dd, J = 5.6, 2.4 Hz, 1H),

7.54 (d, J = 1.6 Hz, 1H), 7.44 (d, J = 8.4 Hz, 1H), 7.32 (dd, J = 8.4,

1.6 Hz, 1H), 5.03–4.98 (m, 2H), 4.91 (d, J = 5.6 Hz, 1H), 4.68 (d,

J = 16.0 Hz, 1H), 4.63 (d, J = 16.0 Hz, 1H), 4.45 (t, J = 5.6 Hz, 1H),

4.02 (d, J = 9.6 Hz, 1H), 3.69–3.65 (m, 1H), 3.44–3.41 (m, 1H),

3.26–3.06 (m, 4H); MS (ESI) m/z 451 (M+H)⁺, 473 (M+Na)⁺.

This paper is dedicated to the memory of Mr. Young-Sup Huh,

Chairman, CEO, Green Cross Corporation (GCC). We appreciate

Dr. Chong-Hwan Chang for his leadership and dedication during

his tenure of ofﬁce as Head of GCC R&D. Also we are grateful to

Dr. Eun Chul Huh for his leadership and supports during his tenure

as Head of GCC R&D Planning and Coordination.

4.3. In vitro assay

References and notes

4.3.1. Cloning and cell line construction for human SGLT2

Human SGLT2 (hSGLT2) gene was ampliﬁed by PCR from cDNA-

Human Adult Normal Tissue Kidney (Invitrogen, Carlsbad, CA). The

1. International Diabetes Federation Diabetes Atlas, 3rd ed.; International Diabetes

Federation: Brussels, Belgium, 2006.

2. Dwarakanathan, A. J. Insur. Med. 2006, 38, 20.

2194

J. Lee et al. / Bioorg. Med. Chem. 18 (2010) 2178–2194

15. Washburn, W. N. J. Med. Chem. 2009, 52, 1785.

3. Mackenzie, B.; Loo, D. D. J. Biol. Chem. 1996, 20, 32678.

4. Wright, E. M. Am. J. Physiol. 2001, 280, F10.

5. Zhou, L.; Cryan, E. V.; D’Andrea, M. R.; Belkowski, S.; Conway, B. R.; Demarest, K.

T. J. Cell. Biochem. 2003, 90, 339.

6. Evans, S.; Grasset, E.; Heyman, M.; Dumonitier, A. M.; Beau, J. P.; Desjeux, J.-F. J.

Pediatr. Gastroenterol. Nutr. 1985, 4, 878.

7. Moe, O. W.; Berry, C. A.; Rector, F. C. In The Kidney; Brenner, B. M., Rector, F. C.,

Eds., 5th ed.; WB Saunders Co.: Philadelphia, 2000; pp 375–415.

8. van den Heuvel, L. P.; Assink, K.; Willesen, M.; Monnens, L. Hum. Genet. 2002,

111, 544.

16. Nomura, S.; Sakamaki, S.; Hongu, M.; Kawanski, E.; Koga, Y.; Sakamoto, T.;

Yamamoto, Y.; Ueta, K.; Kimata, H.; Nakayama, K.; Tsuda-Tsukimoto, M.

Presented at the 238th American Chemical Society National Meeting and

Exposition, Washington, DC, August 16–20, 2009; Medi-151.

17. Johnson

2009.

& Johnson Pharmaceutical, www.clinicaltrials.gov, NCT00968812,

18. According to a report from Business Wire on October 02, 2009, dapagliﬂozin

study demonstrated signiﬁcantly improved glycemic control and weight

reduction in type 2 diabetes patients inadequately controlled with metformin.

19. Ellsworth, B. A.; Doyle, A. G.; Patel, M.; Caceres-Cortes, J.; Meng, W.;

Deshpande, P. P.; Pullockaran, A.; Washburn, W. N. Tetrahedron: Asymmetry

2003, 14, 3242.

20. (a) Kraus, G. A.; Molina, M. T. J. Org. Chem. 1998, 53, 752; (b) Xie, J.; Durrant, F.;

Valery, J.-M. J. Org. Chem. 2003, 68, 7896.

21. Jung, M. E.; Lyster, M. A. J. Org. Chem. 1997, 42, 3761.

22. Chauvire, G.; Bouteille, B.; Enanga, B.; de Albuquerque, C.; Croft, S. L.; Dumas,

M.; Pri, J. J. Med. Chem. 2003, 46, 427.

23. Angibeaud, P.; Utile, J.-P. Synthesis 1991, 737.

24. Han, S.; Hagan, D. L.; Taylor, J. R.; Xin, L.; Meng, W.; Biller, S. A.; Wetterau, J. R.;

Washburn, W. N.; Whaley, J. M. Diabetes 2008, 57, 1723.

9. Ehrenkranz, J. R. L.; Lewis, N. G.; Kahn, C. R.; Roth, J. Diabetes Metab. Res. Rev.

2005, 21, 31.

10. Tsujihara, K.; Hongu, M.; Saito, K.; Kawanishi, H.; Kuriyama, K.; Matsumoto,

M.; Oku, A.; Ueta, K.; Tsuda, M.; Saito, A. J. Med. Chem. 1999, 42, 5311.

11. Fushimi, N.; Ito, F.; Isaji, M. PCT Int. Appl. WO2003011880, 2004.

12. Fujikura, H.; Fushimi, N.; Nishimura, T.; Nakabayashi, T.; Isaji, M. PCT Int. Appl.

WO2002053573, 2002.

13. Ellsworth, B. A.; Meng, W.; Patel, M.; Girotra, R. N.; Wu, G.; Sher, P.; Hagan, D.;

Obermeier, M.; Humphreys, W. G.; Robertson, J. G.; Wang, A.; Han, S.; Waldron,

T.; Morgan, N. N.; Whaley, J. M.; Washburn, W. N. Bioorg. Med. Chem. Lett. 2008,

18, 4770.

14. Meng, W.; Ellsworth, B. A.; Nirschl, A. A.; McCann, P. J.; Patel, M.; Girotra, R. N.;

Wu, G.; Sher, P. M.; Morrison, E. P.; Biller, A. A.; Zahler, R.; Deshpande, P. P.;

Pullockaran, A.; Hagan, D. L.; Morgan, N.; Taylor, J. R.; Obermeier, M. T.;

Humphreys, W. G.; Khanna, A.; Discenza, L.; Robertson, J. G.; Wang, A.; Han, S.;

Wetterau, J. R.; Janovitz, E. B.; Flint, O. P.; Whaley, J. M.; Washburn, W. N. J.

Med. Chem. 2008, 51, 1145.

25. Katsuno, K.; Fujimori, Y.; Takemura, Y.; Hiratochi, M.; Itoh, F.; Komatsu, Y.;

Fujikura, H.; Isaji, M. J. Pharmacol. Exp. Ther. 2007, 320, 323.

26. Arakawa, K.; Ishihara, T.; Oku, A.; Nawano, M.; Ueta, K.; Kitamura, K.;

Matsumoto, M.; Saito, A. Br. J. Pharmacol. 2001, 132, 578.

Article Doi

Novel C-aryl glucoside SGLT2 inhibitors as potential antidiabetic agents: 1,3,4-Thiadiazolylmethylphenyl glucoside congeners

DOI: 10.1016/j.bmc.2010.01.073

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Full text of DOI:10.1016/j.bmc.2010.01.073

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