N-Indolylglycosides bearing modifications at the glucose C6-position as sodium-dependent glucose co-transporter 2 inhibitors

Article abstract of DOI:10.1016/j.bmc.2016.03.058

Suppression of glucose reabsorption through the inhibition of sodium-dependent glucose co-transporter 2 (SGLT2) is a promising therapeutic approach for the treatment of type 2 diabetes. To investigate the effect of C6-substitution on inhibition of SGLT2 b

Full text of DOI:10.1016/j.bmc.2016.03.058

Accepted Manuscript
N-Indolylglycosides Bearing Modifications at the Glucose C6-Position as So-
dium-Dependent Glucose Co-transporter 2 Inhibitors
Kuang-Feng Chu, Chun-Hsu Yao, Jen-Shin Song, Chiung-Tong Chen, Teng-
Kuang Yeh, Tsung-Chih Hsieh, Chung-Yu Huang, Min-Hsien Wang, Szu-Huei
Wu, Wei-En Chang, Yu-Sheng Chao, Jinq-Chyi Lee
PII:
S0968-0896(16)30215-2
DOI:
http://dx.doi.org/10.1016/j.bmc.2016.03.058
BMC 12907
Reference:
Bioorganic & Medicinal Chemistry
To appear in:
Received Date:
Revised Date:
Accepted Date:
4 February 2016
29 March 2016
29 March 2016
Please cite this article as: Chu, K-F., Yao, C-H., Song, J-S., Chen, C-T., Yeh, T-K., Hsieh, T-C., Huang, C-Y.,
Wang, M-H., Wu, S-H., Chang, W-E., Chao, Y-S., Lee, J-C., N-Indolylglycosides Bearing Modifications at the
Glucose C6-Position as Sodium-Dependent Glucose Co-transporter 2 Inhibitors, Bioorganic & Medicinal
Chemistry (2016), doi: http://dx.doi.org/10.1016/j.bmc.2016.03.058
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N-Indolylglycosides Bearing Modifications at the Glucose C6-Position as
Sodium-Dependent Glucose Co-transporter 2 Inhibitors
Kuang-Feng Chu†, Chun-Hsu Yao†, Jen-Shin Song, Chiung-Tong Chen, Teng-Kuang Yeh,
Tsung-Chih Hsieh, Chung-Yu Huang, Min-Hsien Wang, Szu-Huei Wu, Wei-En Chang, Yu-Sheng
Chao‡, Jinq-Chyi Lee*
Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, 35
Keyan Road, Zhunan Town, Miaoli County 35053, Taiwan
*Corresponding author:
Jinq-Chyi Lee, Ph.D.
Tel.: +886 37 246166 ext. 35781
fax: +886 37 586456
E-mail addresses: jinqchyi@nhri.org.tw (J.-C. Lee)
Institute of Biotechnology and Pharmaceutical Research
National Health Research Institutes
35 Keyan Road, Zhunan Town
Miaoli County 35053, Taiwan
† These authors contributed equally to this work.
‡ Present address: Diamond BioFund INC., 5F., No. 3, Sec. 1, Dunhua S. Rd., Taipei City 105,
Taiwan, R. O. C.

Abstract:
Suppression of glucose reabsorption through the inhibition of sodium-dependent glucose
co-transporter 2 (SGLT2) is a promising therapeutic approach for the treatment of type 2 diabetes.
To investigate the effect of C6-substitution on inhibition of SGLT2 by N-indolylglucosides, a small
library of 6-triazole, 6-amide, 6-urea, and 6-thiourea N-indolylglycosides were synthesized and
tested. A detailed structure-activity relationship (SAR) study culminated in the identification of
6-amide derivatives 6a and 6o as potent SGLT2 inhibitors, which were further tested for inhibitory
activity against SGLT1. The data obtained indicated that 6a and 6o are mildly to moderately
selective for SGLT2 over SGLT1. Both compounds were also evaluated in a urinary glucose
excretion test and pharmacokinetic study; 6a was found capable of inducing urinary glucose
excretion in normal SD rats.
Keywords
N-indolylglycosides, sodium-dependent glucose co-transporter, structure-activity relationship, type
2 diabetes mellitus

1. Introduction
Inhibitors of sodium-dependent glucose co-transporter 2 (SGLT2) are a uniquely attractive
method of type 2 diabetes treatment because of their distinct mechanism of action, in which blood
glucose levels are reduced independently of insulin secretion.1 SGLT2, one of twelve constituents
of the solute carrier family 5A (SLC5A), is located on the S1 segment of the proximal tubule in the
kidney and mediates the reabsorption of the majority (>90%) of renal glucose filtered by the kidney
glomeruli.2-5 Reabsorption of the remainder is accomplished by SGLT1, another member of the
SLC5A gene family, which is mainly expressed in the small intestine but also presented in the
S2/S3 segment of the proximal tubule. The reabsorbed glucose is then delivered into bloodstream,
which is hazardous to people with diabetes. Accordingly, SGLT2 inhibition is a promising new
therapeutic approach for the control of blood glucose levels by suppressing the reuptake of glucose,
which leads to glucose excretion in urine and hence reduces blood glucose levels. SGLT2 inhibitors
could also increase the caloric output in urine due to the urinary glucose excretion, suggesting that
their use would not result in the weight gain often seen with other anti-hyperglycemic agents
currently prescribed.6 A modest reduction in blood pressure has also been observed in patients
following treatment with SGLT2 inhibitors, another potential benefit.
To avoid the gastrointestinal side effects associated with SGLT1 inhibition,7-9 inhibitors selective
for SGLT2 are sought. Several selective SGLT2 inhibitors have been developed by structural
modification of phlorizin,10 the first known SGLT inhibitor, and C-linked β-glycosides are at
advanced stages of development because of their metabolic stability, high oral bioavailability, and
plasma exposure. Of these, dapagliflozin,11 canagliflozin,12 ipragliflozin,13 tofogliflozin,14
luseogliflozin15 and empagliflozin16 have been approved for the treatment of type 2 diabetes
mellitus (T2DM) recently (Figure 1); however, long-term follow-up of patients was recommended
due to various unexpected side-effects, including cancers, liver injury, cardiovascular diseases and
ketoacidosis.6,17,18 Also, clinical efficacy studies revealed that current agents can only induce <50%

of filtered glucose load in humans into the urine, perhaps because inhibition of SGLT2 forces
SGLT1 to reabsorb glucose as its maximum capacity.19,20 Accordingly, novel, safer SGLT2
inhibitors also able to partially inhibit SGLT1 without gastrointestinal side effects are sought. Also,
the optimal degree of selectivity for SGLT2 over SGLT1 needs to be determined.
Et
dapagliflozin
O
O
1
2
R
= Cl, R
=
OH
OH
OEt
HO
ipragliflozin
1
R
S
OH
2
1
2
R
R
= F, R
=
tofogliflozin
canagliflozin
Me
OH
OH
O
S
1
2
R
= Me, R
=
HO
F
MeO
OEt
OH
empagliflozin
OH
S
1
2
HO
R
= Cl, R =
O
OH
OH
O
luseogliflozin
Figure 1. SGLT2 inhibitors in clinical use.
In our previous study, a series of N-indolylxyloside derivatives were synthesized and their
inhibitory potency against hSGLT2 demonstrated. This work culminated in the identification of a
potent SGLT2 inhibitor 4-chloro-3-(4-cyclopropylbenzyl)-1-(β-D-xylopyranosyl)-1H-indole 1.21
Pharmacokinetic studies of 1 found it to be metabolically stable with low clearance and good oral
bioavailability (56.3%) in Sprague-Dawley (SD) rats. In further efficacy studies, 1 was found to
significantly increase urine glucose excretion and lower blood glucose levels in streptozotocin
(STZ)-induced
diabetic
rats.
Comparison
of
N-indolylglucoside
4-chloro-3-(4-cyclopropylbenzyl)-1-(β-D-glucopyranosyl)-1H-indole 222-24 with N-indolylxyloside 1
showed that 2 possessed better SGLT2 inhibition when the same aglycone moiety was installed at
the C1 position (1: EC50 = 203 nM; 2: EC50 = 14 nM). This result suggested that the C6 position of
the sugar moiety may play a critical role in the suppression of SGLT2. Therefore, in this article, lead
optimization of N-glycosides proceeded via modification at the C6 position only, the aglycone unit
4-chloro-3-(4-cyclopropylbenzyl)-1H-indole being fixed (Figure 2). A detailed description of the
design, synthesis, and structure-activity relationship (SAR) studies of these newly developed

C6-substituted N-glycosides is presented.
Cl
Ar
R
N
N
OH
OH
OH
OH
O
O
H
R
Z
OH
OH
N-indolylxylosides: Z = H
N-indolylglucosides: Z = CH OH
2
Figure 2. Design of N-indolylglycoside SGLT2 inhibitors with C6-modification on the glucose
moiety.
2. Results and discussion
2.1 Chemistry
The test compounds of Table 1 were prepared according to a general synthetic method depicted in
Scheme 1, using N-indolylglucoside 2 as the starting material. Treatment of 2 with methanesulfonyl
chloride (MsCl) in pyridine gave the corresponding 6-OMs N-glycoside, which was sequentially
reacted with sodium azide (NaN3) to afford the 6-azido compound 3 in 77% yield over 2 steps.
Copper-catalyzed azide-alkyne click chemistry was used to prepare the 1,2,3-triazole derivatives
4a–4d in yields of 76-87%.25,26 Hydrolysis of 4d gave the carboxylic acid 4e in 69% yield. Next,
the synthesis of amide, urea, or thiourea derivatives was carried out via the intermediate amine 5,
generated by the reduction of azido group in 3 with Zn/HOAc in THF (91%). Amine 5 underwent
direct amide bond formation with a variety of acyl chlorides to furnish 6-amide derivatives 6a–6q
in fair to good yields (29–82%). The carboxylic acid 6r was then obtained successfully in 86%
yield via the hydrolysis of methyl ester 6o. In addition, reaction of 5 with isocyanate or
isothiocyanate afforded the desired ureas 7a–7d (48–59%) and thioureas 8a–8e (50–71%),
respectively.

Cl
O
Cl
O
Cl
O
N
N
N
a
b
OH
OH
OH
OH
OH
OH
N
N
N
R
HO
N
3
OH
OH
OH
2
3
4a-4d
4d: R = CO Me
2
c
4e: R = CO H
d
2
Cl
O
Cl
O
Cl
O
N
N
N
f
e
OH
OH
OH
OH
OH
H
N
H
N
1
2
H N
2
R
R N
OH
OH
OH
O
X
OH
5
6a-6q
1
7a-7d: X = O
8a-8e: X = S
6o: R = CH CO Me
2
2
c
1
6r:
R = CH CO H
2 2
Scheme 1. The General Synthetic Route towards C6-Modified N-Indolylglycosides. Reagents and
conditions: (a) (i) MsCl, pyridine, 0 °C to rt; (ii) NaN3, DMF, 80 °C, 77% over 2 steps; (b) RC≡CH,
CuSO4·5H2O, sodium ascorbate, saturated NaHCO3(aq), tBuOH, 50 °C, 76-87%; (c) NaOMe, MeOH,
H2O, rt, 4e: 69%, 6r: 86%; (d) Zn, HOAc, THF, rt, 91%; (e) R1COCl, K2CO3, THF, rt, 29-82%; (f)
for 7a-7d, R2NCO, K2CO3, THF, or R2NCO, pyridine, 48-59%; for 8a-8e, R2NCS, K2CO3, THF, or
R2NCS, pyridine, 50-71%.
2.2 Biological evaluation
All the N-glycosides synthesized were evaluated for their inhibitory activity against hSGLT2. The
EC50 values were calculated by measuring inhibition of the sodium-dependent uptake of
[14C]-labeled α-methyl-D-glucopyranoside (AMG) into Chinese hamster ovary (CHO) cells stably
expressing human SGLT2 (hSGLT2).27 Phlorizin, the first known SGLT inhibitor, was used as
reference compound in this in vitro activity evaluation system.
Table 1 shows the structure-activity relationships elucidated upon the alteration of only the
C6-substituent of glucose. First, the SAR was explored by introducing a 4-substituted triazolyl
group (4a–4e) at the C6-position of the glucose moiety of N-indolylglucoside 2. Of these,
compound 4d bearing a 4-carboxylic acid methyl ester-triazolyl group was found to be the most
potent SGLT2 inhibitor, with an EC50 value of 119 nM. Its hydrolyzed acid product 4e showed a
23-fold decrease in hSGLT2 inhibitory activity (EC50 = 2723 nM). This result suggests that a

Table 1
Effect of C6-modified N-indolylglycosides on hSGLT inhibitory activity and selectivity
Cl
N
OH
O
R
OH
OH
EC50 (nM)a,b
hSGLT2
EC50 [nM]a,c
hSGLT1
EC50 (nM)a,b
hSGLT2
EC50 [nM]a,c
hSGLT1
Cpd
R
Sel.d
Cpd
R
Sel.d
H
N
N
N
MeO2C
EtO2C
－
－
359 ± 83
1585 ± 320
655 ± 90
119 ± 32
2723 ± 918
237 ± 216
42 ± 23
284 ± 36
249 ± 76
39 ± 7
5758 ± 498
4692 ± 663
5424 ± 1357
3813 ± 602
－
20
19
6m
6n
6o
6p
6q
6r
4a
4b
4c
4d
4e
5
N
O
O
O
O
O
O
O
O
H
N
N
N
－
－
－
16
－
9
N
H
N
N
N
N
N
－
139
32
MeO2C
EtO2C
N
EtO
H
N
N
1859 ± 221
－
118 ± 15
588 ± 96
3441 ± 1185
598 ± 125
151 ± 33
N
MeO C
2
H
N
N
EtO C
2
－
N
HO C
2
H
N
H2N
－
－
2027 ± 218
1412 ± 241
3049 ± 708
HO2C
H
N
H
N
H
N
－
－
34
27
7a
7b
6a
6b
Cl
O
H
N
H
N
H
N
Cl
113 ± 10
874 ± 154
6
Cl
O
Cl
H
N
H
N
H
N
EtO C
2
－
－
－
－
184 ± 17
515 ± 55
7c
6c
Cl
O
O
H
N
H
N
H
N
Cl
－
－
－
－
－
－
－
－
－
－
－
－
－
3
160 ± 44
151 ± 10
649 ± 209
552 ± 155
1004 ± 24
315 ± 28
716 ± 99
286 ± 22
7d
8a
8b
8c
6d
6e
6f
EtO C
2
O
O
S
S
S
H
N
H
N
H
N
Br
O
H
N
H
N
H
N
－
O
H
N
H
N
H
N
EtO C
2
845 ± 188
6g
O
MeO
H
H
N
H
N
MeO
N
－
－
－
－
－
－
－
－
－
－
2
2367 ± 373
2591 ± 1371
387 ± 73
586 ± 118
485 ± 17
203 ± 29
8d
8e
1
6h
6i
S
O
H
H
Cl
H
N
N
N
S
O
Cl
H
N
N
S
488 ± 182
6j
O
O
H
N
－
－
－
－
499 ± 14
14 ± 1
27 ± 19
2
1
2
6k
6l
O
H
N
PZNd
2091 ± 547
123 ± 11
153 ± 44
S
O
aData obtained by at least two independent experiments, each experiment performed in triplicate.
bInhibition of uptake of [14C]-AMG in CHO-K1 cells stably transfected with human SGLT2.
cInhibition of uptake of [14C]-AMG in CHO-K1 cells stably transfected with human SGLT1.
dSel: Selectivity values were calculated by EC50 hSGLT1/EC50 hSGLT2.
ePZN: phlorizin.

4-substituted triazolyl group with relatively high hydrophilicity is less well tolerated for hSGLT2
inhibition; although, when an ethoxy group was attached in place of the methyl ester group, the
inhibitory activity decreased (4c, EC50 = 655 nM). On the other hand, when the rigid and compact
cyclopropyl substituent was attached at the 4-position of the triazole unit, the EC50 value of the
resulting glycoside 4a was just 359 nM, making it 3 times less potent than 4d; and indicating that
increased steric bulk in this region is disfavored. This effect was even more evident when a
cyclopentyl group was introduced at this position; the resulting product, 4b, was found to have an
even higher EC50 of 1585 nM.
A series of 6-amide derivatives (6a–6r) were also synthesized and their inhibitory activity against
hSGLT2 evaluated. Initially, compound 5 bearing a free amino group was examined for its ability to
suppress the reuptake of hSGLT2, and found to be moderately active (EC50 = 237 nM). With this
promising data, we then evaluated the inhibitory effect of 6-amide N-indolylglycosides on hSGLT2.
Encouragingly, it was found that when the amino group was acylated with acetyl chloride, the
resulting compound 6a was a more potent inhibitor of SGLT2, with an EC50 value of 42 nM. We
then attempted to replace the side chain with other functional groups without changing the amido
bond. When the methyl group was modified to a monochloromethyl (6b) or dichloromethyl (6c)
group, a loss in inhibitory potency resulted (EC50 = 113 nM for 6b and 184 nM for 6c). Extension
of the spacer to two methylene units gave rise to 6d (EC50 = 160 nM) and 6e (EC50 = 151 nM), both
with similar potencies to 6b. In addition, there was no improvement in hSCLT2 inhibition using
isopropyl (6f, EC50 = 649 nM) or cyclopropyl (6g, EC50 = 552 nM) substitution, indicating that a
hydrocarbon moiety bulkier than methyl group is not tolerated at this site. The decreased inhibitory
activity was also noted when substituted phenyl groups (6h, EC50 = 2367 nM; 6i, EC50 = 2591 nM)
or isoxazolyl (6j, EC50 = 387 nM) were introduced at this position. Also, the potency was not
recovered as indicated by 2-methylthiophene 6k (EC50 = 499 nM) and methylsulfanylbenzene 6l
(EC50 = 2091 nM). Accordingly, the amido substituent was restricted to sterically small groups for

further exploration of potential inhibitors. As expected, replacement of the bulky group with a
methyl or ethyl ester to give compounds 6m and 6n improved the potency (EC50 = 284 nM and 249
nM, respectively). Even more encouragingly, it was found that when a methyl acetyl group was
introduced in place of the methyl ester group, the corresponding N-glycoside 6o exhibited a
substantial improvement in inhibitory activity with an EC50 of 39 nM. A similar structural
modification was also extended to 6n, leading to the identification of 6p (EC50 = 118 nM) with
modestly enhanced on hSGLT2 inhibition. The hydrolyzed acid product 6r was found to be a
greatly inferior inhibitor (EC50 = 3441 nM). Moreover, decreased potency was also observed when
the spacer was lengthened to two methylenes (6q, EC50 = 588 nM), compared to 6p.
Finally, urea (7a–7d) and thiourea (8a–8e) functionality was installed at the C6-position of the
sugar moiety. In the urea analogue series, 7b bearing a 3-chloropropyl group was found to be the
most potent inhibitor of hSGLT2, with an EC50 value of 151 nM. Changing the length of the
methylene spacer of 7b from three to two (7a, EC50 = 598 nM), or substitution of the side chain
with the ethyl acetyl (7c, EC50 = 515 nM) or ethyl ester (7d, EC50 = 1004 nM) resulted in a 4-, 3-,
and 7-fold loss in potency, respectively. The decreased inhibitory activity against hSGLT2 was also
found in the series N-glycosides bearing 6-thiourea (8a-8e), with EC50 values ranging from 286 to
716 nM.
N-indolylglycosides with good hSGLT2 inhibition in each series were selected for further
evaluation of their selectivity for hSGLT2 over hSGLT1. Of these, compound 6m was the least
potent inhibitor, suppressing the reuptake of hSGLT1 with an EC50 = 5758 nM, and exhibiting a
20-fold selectivity over hSGLT1. In contrast, the hSGLT1 inhibitory activity of the most potent
SGLT2 inhibitors 6a and 6o were 1412 nM and 5424 nM, respectively; these were also the most
selective inhibitors, with the ratio of hSGLT1/2 = 34 and 139, respectively. Compounds 6a and 6o
were found to be more potent and more selective SGLT2 inhibitors than our previously reported
N-indolylxyloside 1 (EC50 = 203 nM; hSGLT1/2 = 2), and yet less potent but more selective

inhibitors compared to N-indolylglucoside 2 (EC50 = 14 nM; hSGLT1/2 = 2). Although the
synthesized 6-amide N-indolylglycosides 6a and 6o are less potent than 2, their mild and moderate
selectivity with regard to SGLT1 has come to our attention for new-generation SGLT2 inhibitors
evaluation.
2.3 In Vivo Evaluation of compounds 6a and 6o
Potent N-indolylglycoside SGLT2 inhibitors 6a and 6o with mild and moderate selectivity over
hSGLT1were further studied to evaluate their ability to induce urinary glucose excretion in normal
SD rats. After oral administration of 6a in single doses of 10 and 50 mg/kg to SD rats, only
increased glucosuria was observed at 50 mg/kg of 101 mg glucose per 200 g body weight (BW)
over 24 h compared with that of the control (0.23 mg glucose/200 g BW) (Figure 3). Blood glucose
levels were also evaluated over a 5 h period after the oral glucose challenge. Although marginal
reduction in glucose levels at 0.5 h and 1 h compared with the control, no statistically significant
reduction in the areas under the blood glucose levels versus time curve (AUC) was observed at 50
mg/kg (data not shown). These results could be attributed to the fact that 6a has unfavorable
pharmacokinetic properties (Table 2). Administration of a single 1.1 mg/kg intravenous dose to the
rats revealed that 6a has a low total body clearance (18.8 mL/min/kg), suggesting it to be a
metabolically stable compound; however, the low plasma concentration (AUC0–inf. = 99 ng/mL*h)
of 6a observed after oral administration indicated poor absorption, reflected in low oral
bioavailability (F = 10%). In the case of 6o, no increased urinary glucose excretion was observed at
oral doses up to 50 mg/kg (data not shown). The pharmacokinetic evaluation of 6o showed the only
signal corresponded to the hydrolyzed acid product 6r; 6o could not be detected regardless of the
mode of administration (intravenous or oral administration) proving it to be a metabolically
unstable compound. Compound 6r has already been proven to be a poor SGLT2 inhibitor (EC50 =
3441 nM) in vitro.

Figure 3. Urine glucose excretion of N-indolylglycoside 6a over 24 h in normal Sprague-Dawley
rats. *P < 0.05 vs vehicle.
Table 2
Pharmacokinetic Properties of N-indolylglycoside 6a after Oral and Intravenous Administration to
Rats
iv
po
1.1
3.2
18.8
2.7
1.1
2.5
dose (mg/kg)
t1/2 (h)
Cl (mL/min/kg)
Vss (L/kg)
Cmax (ng/mL)
tmax (h)
25.2
1.2
99
AUC(0-inf.) (ng/mL*h)
F (%)
996
10
3. Conclusion
Structural modifications of 6-triazole, 6-amide, 6-urea, and 6-thiourea N-indolylglycosides have
been conducted to identify potential SGLT2 inhibitors. Among the compounds assayed, 6-amide
N-indolylglycosides 6a and 6o were found to show the best inhibitory activity against hSGLT2 with
EC50 values of 42 nM and 39 nM, respectively. Moreover, the mild and moderate selectivity over
SGLT1 made their SGLT2 inhibition particularly interesting. Although their poor pharmacokinetic

properties resulted in an unfavorable outcome in a glucosuria study in rats, the in vivo efficacy of
6a was proven, as shown by the increased glucose excretion in the urine. Further structural
modifications and in vivo efficacy studies on the series will be reported in due course.
4. Experimental methods
4.1. Chemistry
The general information of test compounds used in this study are published as supplementary
material.
4.1.1. 1-(6-Azido-6-deoxy-β-D-glucopyranosyl)-4-chloro-3-(4-cyclopropylbenzyl)-1H-indole (3).
MsCl (56 L, 0.72 mmol) was added to a stirred solution of 2 (267 mg, 0.60 mmol) in pyridine (5.0
mL) at 0 °C under nitrogen. The reaction was warmed to room temperature and stirred for 3 h. The
reaction was quenched by the addition of H2O and extracted with EtOAc. The organic layer was
dried over MgSO4, filtered, and concentrated under reduced pressure. The residue was purified by
column chromatography (MeOH/CH2Cl2 = 1/30) to provide 6-OMs N-glucoside (263 mg, 84%).
Sodium azide (328 mg, 5.04 mmol) was added to a stirred solution of 6-OMs N-glucoside (263
mg, 0.50 mmol) in DMF (5.0 mL) at room temperature. The reaction was warmed to 80 °C and
heated overnight. The reaction was quenched by the addition of H2O and extracted with EtOAc. The
organic layer was dried over MgSO4, filtered, and concentrated under reduced pressure. The residue
1
was purified by column chromatography (MeOH/CH2Cl2 = 1/10) to provide 3 (217 mg, 92%). H
NMR (400 MHz, CD3OD):  7.45 (d, J = 8.0 Hz, 1H), 7.10-6.93 (m, 7H), 5.41 (d, J = 9.2 Hz, 1H),
4.27 (s, 2H), 3.79 (t, J = 9.2 Hz, 1H), 3.72-3.67 (m, 1H), 3.62-3.46 (m, 3H), 3.40 (dd, J = 13.2, 5.2
Hz, 1H), 1.88-1.81 (m, 1H), 0.92-0.85 (m, 2H), 0.63-0.59 (m, 2H); 13C NMR (100 MHz, CD3OD):
 142.7, 140.2, 139.9, 129.9, 127.4, 126.7, 126.6, 123.6, 121.8, 117.5, 110.7, 86.7, 79.1, 78.8, 73.7,
71.9, 52.5, 33.2, 15.9, 9.5; ESI-MS m/z: 469 (MH+), 491 (MNa+); HPLC purity 95.6%.

4.1.2. General Procedure for the Synthesis of Compound 4a-4d
A solution of CuSO4‧5H2O (0.4 equiv) and sodium ascorbate (1.0 equiv) in H2O was added to a
stirred solution of 3 (1.0 equiv) and 1-alkyne (2.5 equiv) in t-BuOH (a mixed solvent system
comprising equal volumes of t-BuOH and saturated NaHCO3(aq) was used for alkynes bearing an
ester group) at 50 °C. After 2 h, the reaction was diluted with H2O and extracted with EtOAc. The
organic layer was dried over MgSO4, filtered, and concentrated under reduced pressure. The residue
was purified by column chromatography to afford the desired products 4a–4d.
4.1.2.1.
4-Chloro-3-(4-cyclopropylbenzyl)-1-[6-(4-cyclopropyl-1H-1,2,3-triazol-1-yl)-6-
deoxy-β-D-glucopyranosyl]-1H-indole (4a). The title compound was obtained from 3 with
1
cyclopropyl acethylene according to the general procedure in 76% yield. H NMR (400 MHz,
CD3OD): δ 7.30 (s, 1H), 7.13 (dd, J = 7.6, 1.6 Hz, 1H), 7.10–6.94 (m, 7H), 5.36 (d, J = 9.2 Hz, 1H),
4.74 (dd, J = 14.4, 2.4 Hz, 1H), 4.45 (dd, J = 14.4, 7.2 Hz, 1H), 4.27 (s, 2H), 3.86 (ddd, J = 9.6, 7.2,
2.4 Hz, 1H), 3.79 (t, J = 9.2 Hz, 1H), 3.57 (dd, J = 9.2, 8.8 Hz, 1H), 3.25 (dd, J = 9.6, 8.8 Hz, 1H),
13
1.87–1.79 (m, 2H), 0.92–0.85 (m, 4H), 0.64–0.58 (m, 4H); C NMR (75 MHz, CD3OD): δ 142.8,
140.1, 139.9, 129.9, 127.5, 126.8, 126.6, 123.8, 122.0, 117.5, 110.6, 86.5, 78.9, 78.0, 72.9, 72.3,
52.1, 33.2, 16.0, 9.6, 8.4, 8.3, 7.4; ESI-MS m/z: 535 (MH+), 557 (MNa+); HPLC purity 100%.
4.1.2.2. 4-Chloro-1-[6-(4-cyclopentyl-1H-1,2,3-triazol-1-yl)-6-deoxy-β-D-glucopyranosyl]-3-(4-
cyclo-propylbenzyl)-1H-indole (4b). The title compound was obtained from 3 with cyclopentyl
acethylene according to the general procedure in 83% yield. 1H NMR (400 MHz, CD3OD): δ 7.34 (s,
1H), 7.18–7.14 (m, 1H), 7.08–6.93 (m, 7H), 5.38 (d, J = 9.2 Hz, 1H), 4.77 (dd, J = 14.4, 2.4 Hz,
1H), 4.44 (dd, J = 14.4, 7.2 Hz, 1H), 4.28, 4.26 (ABq, J = 15.6 Hz, 2H), 3.86 (ddd, J = 9.6, 7.2, 2.4
Hz, 1H), 3.81 (dd, J = 9.2, 8.8 Hz, 1H), 3.58 (dd, J = 9.2, 8.8 Hz, 1H), 3.27 (dd, J = 9.6, 9.2 Hz, 1H),
3.01 (quint, J = 8.0 Hz, 1H), 1.96–1.91 (m, 2H), 1.87–1.80 (m, 1H), 1.65–1.56 (m, 4H), 1.49–1.40
(m, 2H), 0.92–0.87 (m, 2H), 0.63–0.59 (m, 2H); 13C NMR (75 MHz, CD3OD): δ 142.8, 140.3,

139.9, 129.8, 127.6, 126.8, 126.7, 126.6, 123.8, 122.0, 117.6, 110.4, 86.3, 78.9, 77.9, 72.8, 72.4,
52.2, 37.9, 34.3, 34.2, 33.1, 26.1, 16.0, 9.6; ESI-MS m/z: 563 (MH+), 585 (MNa+); HPLC purity
100%.
4.1.2.3. 4-Chloro-3-(4-cyclopropylbenzyl)-1-[6-deoxy-6-(4-ethoxy-1H-1,2,3-triazol-1-yl)-β-D-
gluco-pyranosyl]-1H-indole (4c). The title compound was obtained from 3 with 1-ethoxyacetylene
1
according to the general procedure in 87% yield. H NMR (400 MHz, CD3OD): δ 7.17 (dd, J = 6.8,
2.4 Hz, 1H), 7.09–6.94 (m, 8H), 5.39 (d, J = 9.2 Hz, 1H), 4.71 (dd, J = 14.4, 2.4 Hz, 1H), 4.42 (dd,
J = 14.4, 7.2 Hz, 1H), 4.27 (s, 2H), 3.93–3.79 (m, 4H), 3.58 (dd, J = 9.2, 8.8 Hz, 1H), 3.28 (dd, J =
9.6, 8.8 Hz, 1H), 1.86–1.82 (m, 1H), 1.26 (t, J = 7.2 Hz, 3H), 0.93–0.88 (m, 2H), 0.64–0.60 (m, 2H);
13C NMR (75 MHz, CD3OD): δ 162.2, 142.8, 140.2, 139.9, 129.8, 127.6, 126.8, 126.6, 123.7, 122.0,
117.6, 110.6, 109.4, 86.4, 78.9, 78.0, 72.9, 72.3, 67.9, 53.0, 33.2, 16.0, 15.1, 9.6; ESI-MS m/z: 539
(MH+), 561 (MNa+); HPLC purity 97.3%.
4.1.2.4. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[4-(methoxycarbonyl)-1H-1,2,3-tri-
azol-1-yl]-β-D-glucopyranosyl}-1H-indole (4d). The title compound was obtained from 3 with
methyl propiolate according to the general procedure in 79% yield. 1H NMR (400 MHz, CD3OD): δ
8.04 (s, 1H), 7.07–7.03 (m, 3H), 7.02–6.92 (m, 5H), 5.35 (d, J = 9.2 Hz, 1H), 4.83 (dd, J = 14.4, 2.8
Hz, 1H), 4.49 (dd, J = 14.4, 7.2 Hz, 1H), 4.24 (s, 2H), 3.90 (ddd, J = 9.6, 7.2, 2.8 Hz, 1H),
3.82–3.76 (m, 1H), 3.80 (s, 3H), 3.58 (t, J = 8.8 Hz, 1H), 3.28 (dd, J = 9.6, 8.8 Hz, 1H), 1.85–1.79
13
(m, 1H), 0.91–0.87 (m, 2H), 0.63–0.59 (m, 2H); C NMR (100 MHz, CD3OD): δ 162.4, 142.8,
140.3, 140.0, 139.7, 131.2, 129.9, 127.5, 126.8, 126.7, 126.6, 123.8, 122.0, 117.6, 110.6, 86.6, 78.9,
77.5, 72.8, 72.3, 52.6, 52.4, 33.2, 15.9, 9.6; ESI-MS m/z: 553 (MH+), 575 (MNa+); HPLC purity
97.8%.
4.1.3. 1-[6-(4-Carboxy-1H-1,2,3-triazol-1-yl)-6-deoxy-β-D-glucopyranosyl]-4-chloro-3-(4-
cyclopropylbenzyl)-1H-indole (4e).

A 30% solution of NaOMe in MeOH (10 μL), and water were sequentially added to a solution of
4d (20 mg) in MeOH (1.0 mL) at room temperature. After being stirred overnight, the reaction was
diluted with MeOH and neutralized with acidic resin Dowex 50 W2-200. The resulting mixture was
filtered and the filtrate was concentrated under reduced pressure. The residue was purified by
column chromatography (MeOH/CH2Cl2 = 1/5) to provide 4e (13.5 mg, 69%). 1H NMR (400 MHz,
CD3OD): δ 8.12 (s, 1H), 7.09–7.05 (m, 3H), 7.01–6.93 (m, 5H), 5.37 (d, J = 9.2 Hz, 1H), 4.84 (dd, J
= 14.4, 2.8 Hz, 1H), 4.59 (dd, J = 14.4, 7.2 Hz, 1H), 4.25 (s, 2H), 3.93 (ddd, J = 9.6, 7.2, 2.8 Hz,
1H), 3.78 (dd, J = 9.2, 8.8 Hz, 1H), 3.58 (dd, J = 9.2, 8.8 Hz, 1H), 3.30–3.25 (m, 1H), 1.87–1.81 (m,
13
1H), 0.92–0.83 (m, 2H), 0.64–0.60 (m, 2H); C NMR (100 MHz, CD3OD): δ 141.5, 138.7, 138.5,
129.8, 128.6, 126.2, 125.42, 125.37, 125.3, 122.5, 120.7, 116.3, 109.2, 85.3, 77.6, 76.3, 71.6, 70.9,
51.0, 31.9, 14.6, 8.3; ESI-MS m/z: 539 (MH+), 561 (MNa+); HPLC purity 91.8%.
4.1.4.
(5).
1-(6-Amino-6-deoxy-β-D-glucopyranosyl)-4-chloro-3-(4-cyclopropylbenzyl)-1H-indole
A mixture of 3 (447 mg, 1.01 mmol), Zn (1.06 g, 16.14 mmol), and acetic acid (1.1 mL) in THF
(10 mL) was stirred at room temperature under nitrogen overnight. The reaction was filtered
through a pad of Celite and the filtrate was concentrated under reduced pressure. The residue was
dissolved in EtOAc, washed with H2O, dried over MgSO4, filtered, and concentrated under reduced
pressure. The residue was purified by column chromatography (MeOH/CH2Cl2 = 1/5 to 1/1) to
1
provide the desired product 5 (406 mg, 91%) as a yellowish solid. H NMR (300 MHz, CD3OD): δ
7.47 (dd, J = 8.1, 0.9 Hz, 1H), 7.13–7.07 (m, 3H), 7.05–7.01 (m, 2H), 6.98–6.95 (m, 2H), 5.44 (d, J
= 9.0 Hz, 1H), 4.29 (s, 2H), 3.85 (t, J = 9.0 Hz, 1H), 3.66 (ddd, J = 10.5, 8.4, 3.0 Hz, 1H), 3.57 (t, J
= 9.0 Hz, 1H), 3.38–3.34 (m, 1H), 3.26 (dd, J = 13.5, 3.0 Hz, 1H), 2.91 (dd, J = 13.5, 8.4 Hz, 1H),
13
1.89–1.83 (m, 1H), 0.95–0.88 (m, 2H), 0.66–0.60 (m, 2H); C NMR (75 MHz, CD3OD): δ 142.8,
140.2, 139.9, 129.9, 127.5, 126.8, 126.6, 123.7, 121.9, 117.6, 110.6, 86.8, 80.0, 79.0, 73.6, 73.2,
43.7, 33.2, 15.9, 9.6; ESI-MS m/z: 443 (MH+), 465 (MNa+); HPLC purity 95.8%.

4.1.5. General Procedure for the Synthesis of Compounds 6a–6q
Acyl chloride (1.1 equiv) was added to a suspension of 5 (1.0 equiv) and potassium carbonate (10
equiv) in THF at room temperature under nitrogen. After 2 h, the reaction was quenched by the
addition of H2O and the resulting mixture was extracted with EtOAc. The organic layer was dried
over MgSO4, filtered, and concentrated under reduced pressure. The residue was purified by column
chromatography to provide the desired products 6a–6q.
4.1.5.1. 1-[6-(Acetylamino)-6-deoxy-β-D-glucopyranosyl]-4-chloro-3-(4-cyclopropylbenzyl)-1H-
indole (6a). The title compound was obtained from 5 with acetyl chloride according to the general
1
procedure in 55% yield. H NMR (400 MHz, CD3OD): δ 7.43 (d, J = 8.4 Hz, 1H), 7.10–7.06 (m,
3H), 7.01–6.95 (m, 4H), 5.38 (d, J = 8.8 Hz, 1H), 4.28 (s, 2H), 3.80 (t, J = 8.8 Hz, 1H), 3.67 (dd, J
= 14.4, 2.8 Hz, 1H), 3.59–3.55 (m, 1H), 3.55 (t, J = 8.8 Hz, 1H), 3.33–3.25 (m, 2H), 1.88 (s, 3H),
13
1.88–1.81 (m, 1H), 0.92–0.88 (m, 2H), 0.64–0.60 (m, 2H); C NMR (75 MHz, CD3OD): δ 174.1,
142.8, 140.2, 140.0, 129.9, 127.5, 126.8, 126.61, 126.58, 123.6, 121.9, 117.4, 110.7, 86.7, 78.7,
78.6, 73.6, 72.8, 41.9, 33.2, 22.5, 15.9, 9.5; ESI-MS m/z: 485 (MH+), 507 (MNa+); HPLC purity
95.1%.
4.1.5.2. 4-Chloro-1-{6-[(chloroacetyl)amino]-6-deoxy-β-D-glucopyranosyl}-3-(4-cyclopropyl-
benzyl)-1H-indole (6b). The title compound was obtained from 5 with chloroacetyl chloride
1
according to the general procedure in 50% yield. H NMR (400 MHz, CD3OD): δ 7.43 (d, J = 8.0
Hz, 1H), 7.10–6.94 (m, 7H), 5.39 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H), 3.98 (s, 2H), 3.82 (t, J = 9.2 Hz,
1H), 3.75 (dd, J = 14.0, 2.8 Hz, 1H), 3.62 (ddd, J = 9.6, 6.8, 2.8 Hz, 1H), 3.56 (t, J = 9.2 Hz, 1H),
3.37-3.28 (m, 2H), 1.88–1.81 (m, 1H), 0.93–0.88 (m, 2H), 0.64–0.60 (m, 2H); 13C NMR (75 MHz,
CD3OD): δ 169.8, 142.8, 140.2, 139.9, 129.9, 127.5, 126.7, 126.6, 123.7, 121.9, 117.5, 110.7, 86.7,
78.7, 78.3, 73.5, 72.9, 43.2, 42.1, 33.2, 15.9, 9.6; ESI-MS m/z: 519 (MH+), 541 (MNa+); HPLC
purity 93.9%.

4.1.5.3. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(dichloroacetyl)amino]-β-D-gluco-
pyranosyl}-1H-indole (6c). The title compound was obtained from 5 with dichloroacetyl chloride
according to the general procedure in 52% yield. 1H NMR (400 MHz, CD3OD): δ 7.42 (dd, J = 8.0,
0.8 Hz, 1H), 7.10–6.95 (m, 7H), 6.19 (s, 1H), 5.40 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H), 3.85–3.80 (m,
2H), 3.64 (ddd, J = 9.6, 7.2, 2.4 Hz, 1H), 3.56 (t, J = 9.2 Hz, 1H), 3.35–3.26 (m, 2H), 1.89–1.82 (m,
13
1H), 0.93–0.88 (m, 2H), 0.67–0.61 (m, 2H); C NMR (100 MHz, CD3OD): δ 167.1, 142.8, 140.3,
140.0, 129.9, 127.5, 126.7, 126.62, 126.58, 123.7, 121.9, 117.5, 110.7, 86.6, 78.8, 78.2, 73.4, 73.1,
67.6, 42.5, 33.2, 15.9, 9.5; ESI-MS m/z: 575 (MNa+); HPLC purity 99.0%.
4.1.5.4. 4-Chloro-1-{6-[(3-chloropropanoyl)amino]-6-deoxy-β-D-glucopyranosyl}-3-(4-cyclo-
propylbenzyl)-1H-indole (6d). The title compound was obtained from 5 with 3-chloropropanoyl
chloride according to the general procedure in 75% yield. 1H NMR (400 MHz, CD3OD): δ 7.44 (dd,
J = 8.0, 0.8 Hz, 1H), 7.11–7.06 (m, 3H), 7.02–6.96 (m, 4H), 5.38 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H),
3.81 (t, J = 9.2 Hz, 1H), 3.76–3.69 (m, 3H), 3.61–3.54 (m, 2H), 3.37–3.29 (m, 2H), 2.60 (t, J = 6.4
Hz, 2H), 1.89–1.83 (m, 1H), 0.94–0.89 (m, 2H), 0.65–0.61 (m, 2H); 13C NMR (100 MHz, CD3OD):
δ 173.3, 142.8, 140.2, 140.0, 129.9, 127.5, 126.8, 126.62, 126.58, 123.6, 121.9, 117.4, 110.7, 86.7,
78.7, 78.6, 73.6, 72.8, 41.8, 41.3, 39.9, 33.2, 15.9, 9.5; ESI-MS m/z: 533 (MH+), 555 (MNa+); HPLC
purity 92.7%.
4.1.5.5. 1-{6-[(3-bromopropanoyl)amino]-6-deoxy-β-D-glucopyranosyl}-4-chloro-3-(4-cyclo-
propylbenzyl)-1H-indole (6e). The title compound was obtained from 5 with 3-bromopropanoyl
chloride according to the general procedure in 43% yield. 1H NMR (400 MHz, CD3OD): δ 7.44 (dd,
J = 8.0, 0.8 Hz, 1H), 7.11–6.95 (m, 7H), 5.38 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H), 3.81 (t, J = 9.2 Hz,
1H), 3.70 (dd, J = 14.0, 2.8 Hz, 1H), 3.62–3.54 (m, 4H), 3.37–3.30 (m, 2H), 2.72 (t, J = 6.4 Hz,
13
2H), 1.89–1.82 (m, 1H), 0.93–0.88 (m, 2H), 0.65–0.61 (m, 2H); C NMR (75 MHz, CD3OD): δ
173.5, 142.8, 140.2, 139.9, 129.9, 127.5, 126.8, 126.63, 126.57, 123.6, 121.9, 117.5, 110.7, 86.7,

78.7, 78.6, 73.6, 72.8, 41.8, 40.0, 33.2, 28.6, 16.0, 9.6; ESI-MS m/z: 599 (MNa+); HPLC purity
83.8%.
4.1.5.6. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(2-methylpropanoyl)amino]-β-D-
glucopyranosyl}-1H-indole (6f). The title compound was obtained from 5 with 2-methylpropanoyl
chloride according to the general procedure in 66% yield. 1H NMR (300 MHz, CD3OD): δ 7.43 (dd,
J = 8.1, 0.9 Hz, 1H), 7.11–6.95 (m, 7H), 5.39 (d, J = 9.0 Hz, 1H), 4.28 (s, 2H), 3.81 (t, J = 9.0 Hz,
1H), 3.67–3.54 (m, 3H), 3.36–3.23 (m, 2H), 2.39 (sept, J = 6.6 Hz, 1H), 1.90–1.81 (m, 1H), 1.054
(d, J = 6.6 Hz, 3H), 1.046 (d, J = 6.6 Hz, 3H), 0.94–0.88 (m, 2H), 0.65–0.60 (m, 2H); 13C NMR (75
MHz, CD3OD): δ 181.0, 142.8, 140.3, 140.0, 129.9, 127.5, 126.7, 126.6, 123.6, 121.9, 117.5, 110.6,
86.6, 78.65, 78.58, 73.5, 72.9, 41.6, 36.2, 33.2, 20.1, 20.0, 15.9, 9.6; ESI-MS m/z: 513 (MH+), 535
(MNa+), HPLC purity 98.8%.
4.1.5.7. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-[(cyclopropylcarbonyl)amino]-6-deoxy-β-D-
glucopyranosyl}-1H-indole (6g). The title compound was obtained from
5
with
1
cyclopropanecarbonyl chloride according to the general procedure in 56% yield. H NMR (400
MHz, CD3OD): δ 7.45 (dd, J = 8.0, 0.8 Hz, 1H), 7.11–6.94 (m, 7H), 5.39 (d, J = 9.2 Hz, 1H), 4.28
(s, 2H), 3.82 (t, J = 9.2 Hz, 1H), 3.68 (dd, J = 14.4, 2.4 Hz, 1H), 3.59–3.54 (m, 2H), 3.38–3.25 (m,
2H), 1.87–1.81 (m, 1H), 1.55–1.49 (m, 1H), 0.93–0.88 (m, 2H), 0.84–0.80 (m, 2H), 0.73–0.68 (m,
2H), 0.64–0.60 (m, 2H); 13C NMR (100 MHz, CD3OD): δ 177.4, 142.7, 140.2, 140.0, 129.8, 127.5,
126.8, 126.60, 126.57, 86.7, 78.8, 78.5, 73.6, 72.7, 41.9, 33.1, 15.9, 14.8, 9.6, 9.5, 7.6, 7.5; ESI-MS
m/z: 511 (MH+); HPLC purity 98.1%.
4.1.5.8. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(4-methoxybenzoyl)amino]-β-D-gluco-
pyranosyl}-1H-indole (6h). The title compound was obtained from 5 with methyl
1
4-methoxybenzoyl chloride according to the general procedure in 44% yield. H NMR (400 MHz,

CD3OD): δ 7.73–7.69 (m, 2H), 7.42–7.37 (m, 1H), 7.09–6.91 (m, 9H), 5.42 (d, J = 9.2 Hz, 1H),
4.28, 4.25 (ABq, J = 16 Hz, 2H), 3.87–3.81 (m, 2H), 3.82 (s, 3H), 3.70 (ddd, J = 9.6, 6.8, 2.8 Hz,
1H), 3.60 (t, J = 9.2 Hz, 1H), 3.54 (dd, J = 14.4, 6.8 Hz, 1H), 3.35–3.30 (m, 1H), 1.87–1.80 (m,
13
1H), 0.92–0.87 (m, 2H), 0.63–0.58 (m, 2H); C NMR (75 MHz, CD3OD): δ 170.5, 164.1, 142.7,
140.3, 140.0, 130.4, 129.8, 127.5, 127.4, 126.7, 126.61, 126.57, 123.6, 121.9, 86.6, 78.7, 78.6, 73.4,
73.0, 56.1, 42.2, 33.1, 15.9, 9.6; ESI-MS m/z: 577 (MH+), 599 (MNa+); HPLC purity 95.0%.
4.1.5.9. 4-Chloro-1-(6-{[4-(chloromethyl)benzoyl]amino}-6-deoxy-β-D-glucopyranosyl)-3-(4-
cyclopropylbenzyl)-1H-indole (6i). The title compound was obtained from 5 with
1
4-(chloromethyl)benzoyl chloride according to the general procedure in 60% yield. H NMR (400
MHz, CD3OD): δ 7.73–7.70 (m, 2H), 7.45–7.42 (m, 2H), 7.38 (dd, J = 7.6, 1.6 Hz, 1H), 7.09–7.04
(m, 3H), 7.00–6.92 (m, 4H), 5.42 (d, J = 9.2 Hz, 1H), 4.66 (s, 2H), 4.28, 4.25 (ABq, J = 16.4 Hz,
2H), 3.88 (dd, J = 14.0, 2.8 Hz, 1H), 3.85 (dd, J = 9.2, 8.8 Hz, 1H), 3.72 (ddd, J = 9.6, 6.8, 2.8 Hz,
1H), 3.60 (t, J = 8.8 Hz, 1H), 3.51 (dd, J = 14.0, 6.8 Hz, 1H), 3.36–3.30 (m, 1H), 1.87–1.80 (m, 1H),
0.92–0.87 (m, 2H), 0.63–0.58 (m, 2H); 13C NMR (75 MHz, CD3OD): δ 170.3, 143.1, 142.7, 140.3,
140.0, 135.3, 129.9, 129.8, 128.9, 127.5, 126.7, 126.62, 126.58, 123.6, 121.9, 117.4, 110.6, 86.6,
78.7, 78.5, 73.4, 73.1, 46.2, 42.4, 33.1, 15.9, 9.6; ESI-MS m/z: 595 (MH+), 617 (MNa+); HPLC
purity 89.0%.
4.1.5.10. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(1,2-oxazol-5-ylcarbonyl)amino]-β-
D-glucopyranosyl}-1H-indole (6j). The title compound was obtained from 5 with
isoxazole-5-carbonyl chloride according to the general procedure in 74% yield. 1H NMR (400 MHz,
CD3OD): δ 8.45 (d, J = 1.6 Hz, 1H), 7.40–7.37 (m, 1H), 7.09–6.93 (m, 7H), 6.86 (d, J = 1.6 Hz,
1H), 5.40 (d, J = 9.2 Hz, 1H), 4.27 (s, 2H), 3.89 (dd, J = 14.0, 2.8 Hz, 1H), 3.85 (dd, J = 9.2, 8.8
Hz, 1H), 3.73 (ddd, J = 10.0, 7.2, 2.8 Hz, 1H), 3.58 (dd, J = 9.2, 8.8 Hz, 1H), 3.48 (dd, J = 14.0, 7.2
13
Hz, 1H), 3.34 (t, J = 9.2 Hz, 1H), 1.87–1.80 (m, 1H), 0.92–0.87 (m, 2H), 0.64–0.59 (m, 2H); C

NMR (100 MHz, CD3OD): δ 164.0, 158.6, 152.4, 142.7, 140.2, 139.9, 129.9, 127.5, 126.8, 126.6,
123.6, 121.9, 117.4, 110.7, 107.3, 86.7, 78.8, 78.2, 73.4, 73.2, 42.0, 33.2, 15.9, 9.6; ESI-MS m/z:
538 (MH+), 560 (MNa+); HPLC purity 98.0%.
4.1.5.11. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(thiophen-2-ylacetyl)amino]-β-D-
glucopyranosyl}-1H-indole (6k). The title compound was obtained from 5 with 2-thiopheneacetyl
chloride according to the general procedure in 62% yield. 1H NMR(400 MHz, CD3OD): δ 7.41 (dd,
J = 8.0, 1.2 Hz, 1H), 7.13–7.07 (m, 4H), 7.02 (dd, J = 7.6, 1.2 Hz, 1H), 6.99–6.95 (m, 3H),
6.83–6.78 (m, 2H), 5.36 (d, J = 9.2 Hz, 1H), 4.29 (s, 2H), 3.77 (dd, J = 9.2, 8.8 Hz, 1H), 3.69 (dd, J
= 14.0, 2.8 Hz, 1H), 3.65 (d, J = 0.8 Hz, 2H), 3.59–3.52 (m, 2H), 3.36–3.30 (m, 1H), 3.25 (dd, J =
13
9.6, 8.8 Hz, 1H), 1.89–1.82 (m, 1H), 0.93–0.88 (m, 2H), 0.65–0.60 (m, 2H); C NMR(75 MHz,
CD3OD): δ 173.6, 142.8, 140.3, 140.0, 138.1, 129.9, 128.0, 127.8, 127.5, 126.7, 126.64, 126.56,
125.9, 123.7, 121.9, 117.5, 110.7, 86.6, 78.6, 78.5, 73.6, 72.9, 41.8, 37.9, 33.2, 16.0, 9.6; ESI-MS
m/z: 589 (MNa+); HPLC purity 95.7%.
4.1.5.12. 4-Chloro-3-(4-cyclopropylbenzyl)-1-(6-deoxy-6-{[(phenylsulfanyl)acetyl]amino}-β-D-
glucopyranosyl)-1H-indole (6l). The title compound was obtained from 5 with (phenylthio)acetyl
chloride according to the general procedure in 65% yield. 1H NMR (400 MHz, CD3OD): δ 7.40 (dd,
J = 8.0, 1.2 Hz, 1H), 7.25–7.22 (m, 2H), 7.16–7.05 (m, 6H), 7.01 (dd, J = 7.6, 1.2 Hz, 1H),
6.99–6.97 (m, 2H), 6.90 (s, 1H), 5.31 (d, J = 9.2 Hz, 1H), 4.30 (s, 2H), 3.65 (dd, J = 9.2, 8.8 Hz,
1H), 3.62–3.58 (m, 1H), 3.58 (s, 2H), 3.53–3.48 (m, 2H), 3.39 (dd, J = 14.0, 6.4 Hz, 1H), 3.13 (dd,
J = 9.6, 9.2 Hz, 1H), 1.90–1.82 (m, 1H), 0.92–0.88 (m, 2H), 0.64–0.60 (m, 2H);13C NMR (75 MHz,
CD3OD): δ 172.1, 142.8, 140.2, 139.9, 136.4, 130.6, 130.4, 129.9, 128.0, 127.5, 126.7, 126.6, 126.5,
123.7, 121.9, 117.6, 110.7, 86.6, 78.43, 78.40, 73.5, 72.4, 41.8, 38.5, 33.3, 16.0, 9.6; ESI-MS m/z:
593 (MH+), 615 (MNa+); HPLC purity 100%.
4.1.5.13. 4-Chloro-3-(4-cyclopropylbenzyl)-1-(6-deoxy-6-{[methoxy(oxo)acetyl]amino}-β-D-

glucopyranosyl)-1H-indole (6m). The title compound was obtained from 5 with methyl oxalyl
1
chloride according to the general procedure in 57% yield. H NMR (400 MHz, CD3OD): δ 7.41 (dd,
J = 8.4, 1.2 Hz, 1H), 7.11–7.05 (m, 3H), 7.02–6.95 (m, 4H), 5.39 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H),
3.82 (t, J = 9.2 Hz, 1H), 3.80 (s, 3H), 3.79 (dd, J = 14.0, 2.8 Hz, 1H), 3.68 (ddd, J = 10.0, 7.6, 2.8
Hz, 1H), 3.55 (t, J = 9.2 Hz, 1H), 3.36 (dd, J = 14.0, 7.6 Hz, 1H), 3.35–3.28 (m, 1H), 1.89–1.82 (m,
13
1H), 0.93–0.87 (m, 2H), 0.65–0.61 (m, 2H); C NMR(100 MHz, CD3OD): δ 161.9, 159.4, 142.8,
140.2, 139.9, 129.9, 127.5, 126.8, 126.6, 123.7, 121.9, 117.5, 110.7, 86.7, 78.7, 78.0, 73.4, 73.2,
42.3, 33.2, 16.0, 9.6; ESI-MS m/z: 529 (MH+), 551 (MNa+); HPLC purity 95.4%.
4.1.5.14. 4-Chloro-3-(4-cyclopropylbenzyl)-1-(6-deoxy-6-{[ethoxy(oxo)acetyl]amino}-β-D-
glucopyranosyl)-1H-indole (6n). The title compound was obtained from 5 with ethyl oxalyl
chloride according to the general procedure in 29% yield. 1H NMR(400 MHz, CD3OD): δ 7.41 (dd,
J = 8.4, 1.2 Hz, 1H), 7.11–6.95 (m, 7H), 5.39 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H), 4.25 (q, J = 7.2 Hz,
2H), 3.85–3.76 (m, 2H), 3.68 (ddd, J = 10.0, 7.6, 2.8 Hz, 1H), 3.56 (t, J = 9.2 Hz, 1H), 3.38–3.28
13
(m, 2H), 1.88–1.82 (m, 1H), 1.27 (t, J = 7.2 Hz, 3H), 0.93–0.88 (m, 2H), 0.64–0.60 (m, 2H); C
NMR (100 MHz, CD3OD): δ 161.9, 159.4, 142.8, 140.2, 139.9, 129.9, 127.5, 126.8, 126.6, 123.7,
121.9, 117.5, 110.7, 86.7, 78.7, 78.0, 73.4, 73.2, 58.5, 42.3, 33.2, 18.5, 16.0, 9.6; ESI-MS m/z: 565
(MNa+); HPLC purity 92.6%.
4.1.5.15. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(3-methoxy-3-oxopropanoyl)amino]-
β-D-glucopyranosyl}-1H-indole (6o). The title compound was obtained from 5 with methyl
1
3-chloro-3-oxopropanoate according to the general procedure in 46% yield. H NMR (400 MHz,
CD3OD): δ 7.46 (dd, J = 8.4, 0.8 Hz, 1H), 7.11–6.96 (m, 7H), 5.40 (d, J = 9.2 Hz, 1H), 4.29 (s, 2H),
3.81 (t, J = 9.2 Hz, 1H), 3.73 (dd, J = 14.0, 2.4 Hz, 1H), 3.65 (s, 3H), 3.63–3.54 (m, 2H), 3.38–3.25
(m, 4H), 1.90–1.84 (m, 1H), 0.94–0.89 (m, 2H), 0.65–0.61 (m, 2H); 13C NMR (100 MHz, CD3OD):
δ 170.2, 169.1, 142.8, 140.2, 140.0, 129.9, 127.4, 126.8, 126.60, 126.57, 123.6, 121.9, 117.4, 110.8,

86.7, 78.59, 78.57, 73.6, 72.7, 52.9, 41.9, 33.2, 15.9, 9.6; ESI-MS m/z: 543 (MH+), 565 (MNa+);
HPLC purity 100%.
4.1.5.16. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(3-ethoxy-3-oxopropanoyl)amino]-β-
D-glucopyranosyl}-1H-indole (6p). The title compound was obtained from 5 with ethyl
1
3-chloro-3-oxopropanoate according to the general procedure in 38% yield. H NMR (400 MHz,
CD3OD): δ 7.46 (dd, J = 8.4, 0.8 Hz, 1H), 7.11–6.95 (m, 7H), 5.39 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H),
4.13–4.08 (q, J = 7.2 Hz, 2H), 3.82 (t, J = 9.2 Hz, 1H), 3.72 (dd, J = 14.4, 2.8 Hz, 1H), 3.60 (ddd, J
= 9.6, 6.8, 2.8 Hz, 1H), 3.56 (t, J = 9.2 Hz, 1H), 3.39–3.24 (m, 4H), 1.88–1.82 (m, 1H), 1.20 (t, J =
7.2 Hz, 3H), 0.93–0.89 (m, 2H), 0.64–0.61 (m, 2H); 13C NMR (100 MHz, CD3OD): δ 169.8, 169.2,
142.8, 140.2, 140.0, 129.9, 127.4, 126.8, 126.6, 123.6, 121.9, 117.4, 110.8, 86.7, 78.6, 73.6, 72.7,
62.5, 41.9, 33.2, 16.0, 14.6, 9.6; ESI-MS m/z: 557 (MH+), 579 (MNa+); HPLC purity 96.9%.
4.1.5.17. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(4-ethoxy-4-oxobutanoyl)amino]-β-
D-glucopyranosyl}-1H-indole (6q). The title compound was obtained from 5 with ethyl succinyl
chloride according to the general procedure in 56% yield.1H NMR (400 MHz, CD3OD): δ 7.47 (dd,
J = 8.0, 0.8 Hz, 1H), 7.11–6.94 (m, 7H), 5.38 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H), 4.07 (q, J = 7.2 Hz,
2H), 3.82 (t, J = 9.2 Hz, 1H), 3.65 (dd, J = 14.0, 2.4 Hz, 1H), 3.58–3.53 (m, 2H), 3.36–3.26 (m, 2H),
2.56–2.53 (m, 2H), 2.43–2.39 (m, 2H), 1.88–1.82 (m, 1H), 1.20 (t, J = 7.2 Hz, 3H), 0.93–0.88 (m,
2H), 0.64–0.60 (m, 2H); 13C NMR (100 MHz, CD3OD): δ 175.2, 174.5, 142.7, 140.2, 140.0, 129.9,
127.4, 126.8, 126.6, 126.5, 123.6, 121.9, 117.4, 110.8, 86.7, 78.7, 78.5, 73.6, 72.7, 61.8, 41.7, 33.2,
31.4, 30.6, 15.9, 14.7, 9.6; ESI-MS m/z: 593 (MNa+); HPLC purity 89.8%.
4.1.6. 1-{6-[(carboxyacetyl)amino]-6-deoxy-β-D-glucopyranosyl}-4-chloro-3-(4-cyclopropyl-
benzyl)-1H-indole (6r). A 30% solution of NaOMe in MeOH (5 μL) and water were sequentially
added to a solution of 6o (30 mg, 0.055 mmol) in MeOH (1.0 mL) at room temperature. After being
stirred 1 h, the reaction was diluted with MeOH and neutralized with acidic resin Dowex 50

W2-200. The resulting mixture was filtered and the filtrate was concentrated under reduced pressure.
The residue was purified by column chromatography (MeOH/CH2Cl2 = 1/3) to provide 6r (25 mg,
86%). 1H NMR (400 MHz, CD3OD): δ 7.46 (dd, J = 8.4, 0.8 Hz, 1H), 7.12–6.95 (m, 7H), 5.39 (d, J
= 8.8 Hz, 1H), 4.28 (s, 2H), 3.82 (dd, J = 9.2, 8.8 Hz, 1H), 3.74 (dd, J = 14.4, 2.8 Hz, 1H), 3.61
(ddd, J = 9.6, 6.8, 2.8 Hz, 1H), 3.56 (t, J = 9.2 Hz, 1H), 3.37 (dd, J = 14.4, 6.8 Hz, 1H), 3.35 (t, J =
9.6, 9.2 Hz, 1H), 3.23 (s, 2H), 1.89–1.82 (m, 1H), 0.93–0.88 (m, 2H), 0.64–0.61 (m, 2H); 13C NMR
(75 MHz, CD3OD): δ 171.6, 169.8, 142.7, 140.2, 140.0, 129.8, 127.4, 126.9, 126.6, 123.7, 121.9,
117.3, 110.8, 86.8, 78.6, 73.6, 72.7, 41.8, 33.1, 15.9, 9.5; ESI-MS m/z: 529 (MH+), 551 (MNa+);
HPLC purity 96.0%.
4.1.7. General Procedure for the Synthesis of Compounds 7a–7d
7a and 7d: Isocyanate (1.2 equiv) was added to a suspension solution of 5 (1.0 equiv) and
potassium carbonate (5 equiv) in THF at room temperature. After 3 h, the reaction was quenched by
the addition of H2O and the resulting mixture was extracted with EtOAc. The organic layer was
dried over MgSO4, filtered, and concentrated under reduced pressure. The residue was purified by
column chromatography to provide the desired products 7a and 7d.
7b and 7c: Isocyanate (1.2 equiv) was added to a suspension solution of 5 (1.0 equiv) in pyridine
at room temperature. After 3~4 h, the reaction was concentrated under reduced pressure and the
residue was purified by column chromatography to provide the desired products 7b and 7c.
4.1.7.1. 4-Chloro-1-(6-{[(2-chloroethyl)carbamoyl]amino}-6-deoxy- β-D-glucopyranosyl)-3-(4-
cyclopropylbenzyl)-1H-indole (7a). The title compound was obtained from 5 with 2-chloroethyl
1
isocyanate according to the general procedure in 53% yield. H NMR (300 MHz, CD3OD) : δ 7.45
(dd, J = 8.4, 0.9 Hz, 1H), 7.11–6.95 (m, 7H), 5.40 (d, J = 9.0 Hz, 1H), 4.29 (s, 2H), 3.81 (t, J = 9.0
Hz, 1H), 3.64–3.46 (m, 5H), 3.37–3.26 (m, 4H), 1.88–1.81 (m, 1H), 0.94–0.88 (m, 2H), 0.65–0.60
13
(m, 2H); C NMR (75 MHz, CD3OD): δ 161.3, 142.8, 140.2, 140.0, 129.9, 127.5, 126.8, 126.63,

126.59, 123.7, 121.9, 117.4, 110.6, 86.7, 79.4, 78.6, 73.7, 72.6, 44.9, 43.2, 42.3, 33.2, 16.0, 9.5;
ESI-MS m/z: 548 (MH+), 570 (MNa+); HPLC purity 92.7%.
4.1.7.2. 4-Chloro-1-(6-{[(3-chloropropyl)carbamoyl]amino}-6-deoxy- β-D-glucopyranosyl)-3-
(4-cyclopropylbenzyl)-1H-indole (7b). The title compound was obtained from 5 with
1
3-chloropropyl isocyanate according to the general procedure in 59% yield. H NMR (300 MHz,
CD3OD) : δ 7.44 (dd, J = 8.4, 1.2 Hz, 1H), 7.11–6.95 (m, 7H), 5.40 (d, J = 9.0 Hz, 1H), 4.29 (s, 2H),
3.81 (t, J = 9.0 Hz, 1H), 3.63–3.46 (m, 5H), 3.32–3.26 (m, 2H), 3.17 (t, J = 6.6 Hz, 2H), 1.95–1.75
(m, 3H), 0.94–0.88 (m, 2H), 0.65–0.60 (m, 2H); 13C NMR (100 MHz, CD3OD): δ 161.6, 142.8,
140.2, 140.0, 129.9, 127.5, 126.8, 126.62, 126.59, 123.7, 121.9, 117.5, 110.6, 86.7, 79.5, 78.6, 73.6,
72.6, 43.3, 42.4, 38.4, 34.2, 33.2, 15.9, 9.6; ESI-MS m/z: 562 (MH+), 584 (MNa+); HPLC purity
94.3%.
4.1.7.3. 4-Chloro-3-(4-cyclopropylbenzyl)-1-(6-deoxy-6-{[(ethoxycarbonylmethyl)carbamoyl]-
amino}-β-D-glucopyranosyl)-1H-indole (7c). The title compound was obtained from 5 with ethyl
isocyanatoacetate according to the general procedure in 56% yield. 1H NMR (400 MHz, CD3OD): δ
7.46 (dd, J = 8.4, 0.8 Hz, 1H), 7.11–7.03 (m, 4H), 7.01 (dd, J = 7.6, 0.8 Hz, 1H), 6.97–6.94 (m, 2H),
5.39 (d, J = 9.2 Hz, 1H), 4.29 (s, 2H), 4.13 (q, J = 7.2 Hz, 2H), 3.82 (dd, J = 9.2, 8.8 Hz, 1H), 3.79
(s, 2H), 3.61 (dd, J = 14.0, 2.8 Hz, 1H), 3.57 (t, J = 8.8 Hz, 1H), 3.56–3.52 (m, 1H), 3.37–3.28 (m,
2H), 1.88–1.82 (m, 1H), 1.23 (t, J = 7.2 Hz, 3H), 0.93–0.88 (m, 2H), 0.64–0.60 (m, 2H); 13C NMR
(100 MHz, CD3OD): δ 172.8, 161.4, 142.7, 140.2, 140.0, 129.8, 127.4, 126.8, 126.6, 126.5, 123.7,
121.8, 117.4, 110.7, 86.8, 79.4, 78.5, 73.7, 72.5, 62.3, 42.9, 42.3, 33.1, 15.9, 14.6, 9.6; ESI-MS m/z:
594 (MNa+); HPLC purity 90.5%.
4.1.7.4. 4-Chloro-3-(4-cyclopropylbenzyl)-1-(6-deoxy-6-{[(ethoxycarbonyl)carbamoyl]amino}-
β-D-glucopyranosyl)-1H-indole (7d). The title compound was obtained from 5 with ethyl
1
isocyanatoformate according to the general procedure in 48% yield. H NMR (400 MHz, CD3OD):

δ 7.42 (dd, J = 8.0, 0.8 Hz, 1H), 7.10–6.94 (m, 7H), 5.36 (d, J = 9.2 Hz, 1H), 4.27 (s, 2H), 4.01 (q, J
= 7.2 Hz, 2H), 3.80 (dd, J = 9.2, 8.8 Hz, 1H), 3.59–3.51 (m, 3H), 3.33–3.21 (m, 2H), 1.87–1.81 (m,
1H), 1.16 (t, J = 7.2 Hz, 3H), 0.92–0.83 (m, 2H), 0.63–0.59 (m, 2H); APCI-MS m/z: 556 (M-H)-;
HPLC purity 100%.
4.1.8. General Procedure for the Synthesis of Compounds 8a–8e
8a: Isothiocyanate (1.2 equiv) was added to a suspension solution of 5 (1.0 equiv) and potassium
carbonate (5 equiv) in THF at room temperature. After 3 h, the reaction was quenched by the
addition of H2O and the resulting mixture was extracted with EtOAc. The organic layer was dried
over MgSO4, filtered, and concentrated under reduced pressure. The residue was purified by column
chromatography to provide the desired product 8a.
8b8e: Isocyanate (1.2 equiv) was added to a suspension solution of 5 (1.0 equiv) in pyridine at
room temperature. After 3~6 h, the reaction was concentrated under reduced pressure and the
residue was purified by column chromatography to provide the desired products 8b8e.
4.1.8.1. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(methylcarbamothioyl)amino]-β-D-
glucopyranosyl}-1H-indole (8a). The title compound was obtained from 5 with methyl
isothiocyanate according to the general procedure in 69% yield. 1H NMR(400 MHz, CD3OD): δ
7.44 (dd, J = 8.4, 0.8, Hz, 1H), 7.11–7.00 (m, 5H), 6.97–6.94 (m, 2H), 5.41 (d, J = 9.2 Hz, 1H),
4.29 (s, 2H), 4.00–3.56 (m, 3H), 3.82 (t, J = 9.2 Hz, 1H), 3.58 (t, J = 9.2 Hz, 1H), 3.36 (t, J = 9.2 Hz,
1H), 2.81 (brs, 3H), 1.89–1.82 (m, 1H), 0.93–0.88 (m, 2H), 0.64–0.60 (m, 2H); ESI-MS m/z: 516
(MH+), 538 (MNa+); HPLC purity 100%.
4.1.8.2. 4-Chloro-3-(4-cyclopropylbenzyl)-1-{6-deoxy-6-[(ethylcarbamothioyl)amino]-β-D-
glucopyranosyl}-1H-indole (8b). The title compound was obtained from 5 with ethyl
1
isothiocyanate according to the general procedure in 71% yield. H NMR (400 MHz, CD3OD) : δ

7.44 (dd, J = 8.0, 0.8, Hz, 1H), 7.11–7.04 (m, 4H), 7.02 (dd, J = 7.6, 0.8 Hz, 1H), 6.98–6.94 (m,
2H), 5.42 (d, J = 9.2 Hz, 1H), 4.29 (s, 2H), 4.00–3.57 (m, 3H), 3.83 (t, J = 9.2 Hz, 1H), 3.59 (t, J =
9.2 Hz, 1H), 3.40–3.30 (m, 2H), 3.36 (t, J = 9.2 Hz, 1H), 1.88–1.82 (m, 1H), 0.99 (brs, 3H),
0.93–0.88 (m, 2H), 0.64–0.60 (m, 2H); ESI-MS m/z: 530 (MH+), 552 (MNa+); HPLC purity 99.2%.
4.1.8.3. 4-Chloro-3-(4-cyclopropylbenzyl)-1-(6-deoxy-6-{[(ethoxycarbonylmethyl)carbamo-
thioyl]amino}-β-D-glucopyranosyl)-1H-indole (8c). The title compound was obtained from 5 with
1
ethyl isothiocyanatoacetate according to the general procedure in 58% yield. H NMR (400 MHz,
CD3OD): δ 7.45 (d, J = 8.4 Hz, 1H), 7.11–7.05 (m, 4H), 7.01 (dd, J = 7.6, 0.8, Hz, 1H), 6.97–6.95
(m, 2H), 5.42 (d, J = 9.2 Hz, 1H), 4.28 (s, 2H), 4.26–4.00 (m, 5H), 3.83 (t, J = 9.2 Hz, 1H),
3.80–3.71 (m, 1H), 3.69–3.65 (m, 1H), 3.58 (t, J = 9.2 Hz, 1H), 3.38 (t, J = 9.2 Hz, 1H), 1.88–1.82
(m, 1H), 1.22 (t, J = 6.8 Hz, 3H), 0.93–0.88 (m, 2H), 0.64–0.61 (m, 2H); ESI-MS m/z: 588 (MH+),
610 (MNa+); HPLC purity 95.8%.
4.1.8.4. 4-Chloro-3-(4-cyclopropylbenzyl)-1-(6-deoxy-6-{[(3-methoxypropyl)carbamothioyl]-
amino}-β-D-glucopyranosyl)-1H-indole (8d). The title compound was obtained from 5 with
1
3-methoxypropyl isothiocyanate according to the general procedure in 50% yield. H NMR (300
MHz, CD3OD)  7.43 (dd, J = 8.4, 0.9 Hz, 1H), 7.11–6.94 (m, 7H), 5.42 (d, J = 9.0 Hz, 1H), 4.28 (s,
2H), 4.00–3.54 (m, 3H), 3.82 (t, J = 9.0 Hz, 1H), 3.57 (t, J = 9.0 Hz, 1H), 3.37–3.30 (m, 1H), 3.33
(s, 3H), 3.20 (brs, 4H), 1.90–1.84 (m, 1H), 1.59 (brs, 2H), 0.93–0.86 (m, 2H), 0.67–0.61 (m, 2H);
ESI-MS m/z: 596 (MNa+); HPLC purity 98.1%.
4.1.8.5. 4-Chloro-3-(4-cyclopropylbenzyl)-1-(6-deoxy-6-{[(4-methoxyphenyl)carbamothioyl]-
amino}-β-D-glucopyranosyl)-1H-indole (8e). The title compound was obtained from 5 with
4-chlorophenyl isothiocyanate according to the general procedure in 61% yield. 1H NMR (400 MHz,
CD3OD): δ 7.47 (dd, J = 8.0, 0.8 Hz, 1H), 7.12–7.01 (m, 7H), 6.97–6.92 (m, 4H), 5.46 (d, J = 9.2
Hz, 1H), 4.30 (s, 2H), 4.40-4.20 (m, 1H), 3.87 (t, J = 8.0 Hz, 1H), 3.78 (ddd, J = 10.0, 7.2, 2.8 Hz,

1H), 3.61 (t, J = 9.2 Hz, 1H), 3.64–3.57 (m, 1H), 3.37 (t, J = 9.2 Hz, 1H), 1.86–1.80 (m, 1H),
0.91–0.86 (m, 2H), 0.62–0.58 (m, 2H); ESI-MS m/z: 612 (MH+), 634 (MNa+); HPLC purity 100%.
4.2. In Vitro Human SGLT Inhibition Assays21,27,28
The transporter assays were performed according to the method from Castaneda and Kinne with
some modification, which are published as supplementary material.
4.3. In Vivo Efficacy and Pharmacokinetics Evaluations of 6a and 6o in Rats21,29
The assessments of urinary glucose excretion and pharmacokinetics were performed using
previously reported procedures. For the Oral Glucose Tolerance Test of 6a, blood glucose levels of
rats were monitored at 0, 0.5, 1, 1.5, 2, 3, 4 and 5 h after the oral glucose challenge. The detailed
description of these experiments is published as supplementary material.
Acknowledgements
We are grateful to the National Health Research Institutes and the Ministry of Science and
Technology of Taiwan (NSC 99-2323-B-400-003) for financial support.
Supplementary data
Supplementary data associated with this article can be found, in the online version, at
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