Discovery of novel N -β- d -Xylosylindole derivatives as sodium-dependent glucose cotransporter 2 (SGLT2) inhibitors for the management of hyperglycemia in diabetes

Article abstract of DOI:10.1021/jm101072y

A novel series of N-linked β-d-xylosides were synthesized and evaluated for inhibitory activity against sodium-dependent glucose cotransporter 2 (SGLT2) in a cell-based assay. Of these, the 4-chloro-3-(4-cyclopropylbenzyl) -1-(β-d-xylopyranosyl)-1H-indole 19m was found to be the most potent inhibitor, with an EC₅₀ value similar to that of the natural SGLT2 inhibitor phlorizin. Further studies in Sprague-Dawley (SD) rats indicated that 19m significantly increased urine glucose excretion in a dose-dependent manner with oral administration. The antihyperglycemic effect of 19m was also observed in streptozotocin (STZ) induced diabetic SD rats. These results described here are a good starting point for further investigations into N-glycoside SGLT2 inhibitors.

Full text of DOI:10.1021/jm101072y

166 J. Med. Chem. 2011, 54, 166–178
DOI: 10.1021/jm101072y
Discovery of Novel N-β-D-Xylosylindole Derivatives as Sodium-Dependent Glucose Cotransporter 2
(SGLT2) Inhibitors for the Management of Hyperglycemia in Diabetes
Chun-Hsu Yao, Jen-Shin Song, Chiung-Tong Chen, Teng-Kuang Yeh, Ming-Shiu Hung, Chih-Chun Chang, Yu-Wei Liu,
Mao-Chia Yuan, Chieh-Jui Hsieh, Chung-Yu Huang, Min-Hsien Wang, Ching-Hui Chiu, Tsung-Chih Hsieh, Szu-Huei Wu,
Wen-Chi Hsiao, Kuang-Feng Chu, Chi-Hui Tsai, Yu-Sheng Chao, and Jinq-Chyi Lee*
Institute of Biotechnology and Pharmaceutical Research, National Health Research Institutes, No. 35, Keyan Road, Zhunan Town,
Miaoli County 35053, Taiwan, R.O.C.
Received August 18, 2010
A novel series of N-linked β-D-xylosides were synthesized and evaluated for inhibitory activity against
sodium-dependent glucose cotransporter 2 (SGLT2) in a cell-based assay. Of these, the 4-chloro-3-(4-
cyclopropylbenzyl)-1-(β-D-xylopyranosyl)-1H-indole 19m was found to be the most potent inhibitor,
with an EC50 value similar to that of the natural SGLT2 inhibitor phlorizin. Further studies in Sprague-
Dawley (SD) rats indicated that 19m significantly increased urine glucose excretion in a dose-dependent
manner with oral administration. The antihyperglycemic effect of 19m was also observed in strepto-
zotocin (STZ) induced diabetic SD rats. These results described here are a good starting point for further
investigations into N-glycoside SGLT2 inhibitors.
Introduction
and SGLT2. It is estimated that 90% of glomerular filtrate re-
absorption is mediated by high-capacity, low-affinity SGLT2
located on the S1 segment of the proximal tubule; the remain-
ing 10% is accomplished by low-capacity, high-affinity
SGLT1 mainly expressed in the small intestine but also pre-
sented in the S3 segment of the proximal tubule. Inhibition of
SGLT1 and SGLT2 is thus expected to suppress the renal glu-
cose reabsorption and increase the excretion of urinary sugar
and consequentially reduce glycemic levels. However, reports
of the studies on genetic mutations in the SGLT1 gene indi-
cated that glucose-galactose malabsorption syndrome is a
serious consequence for afflicted individuals, and no apparent
glucosuria is observed.8-10 In contrast, persistent renal glucose
excretion is the sole phenotype for those who have SGLT2
mutations with no other adverse effects known to be associated
with the variants.11,12 The observation suggests that SGLT2 is
responsible for the majority of renal glucose reabsorption and
that selective SGLT2 inhibitors would be desired for avoiding
gastrointestinal side effects involved in SGLT1 inhibition.
The natural product β-O-glucoside phlorizin 1 was isolated
from the root bark of the apple tree in 1835 and subsequently
identified as the first SGLT inhibitor (Figure 1).13 There is
evidence that phlorizin can induce renal glucose excretion by
suppressing the reabsorption and therefore reduce plasma
glucose levels, but owing to its lack of selectivity toward SGLT1
and poor metabolic stability in the presence of O-glucosidase, it
was not considered to be an antidiabetic drug candidate. By
use of structure-activity relationships (SARs), several new
compounds have been developed as SGLT2 inhibitors and
studied in clinical trials, including the O-glucosides T-1095 2b,
sergliflozin 3b, and remogliflozin etabonate 4b.14-16 Designed
as prodrugs to avoid degradation by glucosidase within the
gastrointestinal tract, they required relatively high dosage in
the clinic, and all clinical studies have been discontinued because
of poor selectivity, poor efficacy, and lack of competitive
Type 2 or non-insulin dependent diabetes mellitus (NIDDMa)
is a metabolic disorder characterized by both fasting and post-
prandial hyperglycemia. Almost all type 2 diabetes patients dis-
play with varying degrees both insulin resistance and relative
insulin deficiency.1 Untreated hyperglycemics are at risk of
micro- and macrovascular complications including retinop-
athy, nephropathy, neuropathy, stroke, amputation, and myo-
cardial infarction.2 Although a wide range of medications
are prescribed for type 2 diabetes, recent studies suggest that
only about one-third of patients treated for type 2 diabetes
achieved the glycemic target, glycosylated hemoglobin (HbA1c)
of <7%.3,4 Thus, development of novel molecules with new
modes of action to supplement the older therapies for treating
patients with uncontrolled type 2 diabetes is required.
Under normal circumstances, and in healthy individuals,
approximately 180 g of glucose is filtered daily in the kidney
glomerulus and nearly all the filtered plasma glucose (>99%)
is reabsorbed by two sodium-dependent glucose cotransporters
(SGLTs), SGLT1 and SGLT2, in the renal proximal tubules.5,6
The SGLTs are members of the solute carrier family 5A
(SLC5A), which is involved in the transport of sugars, amino
acids, vitamins, iodide, ions, neurotransmitters, and osmolytes
across the renal and intestinal membranes.7 The SLC5A gene
family contains 12 constituents, of which only some transport
glucose and the most important and well studied being SGLT1
*To whom correspondence should be addressed. Phone: þ886-
37-246166, extension 35781. Fax: þ886-37-586456. E-mail: jinqchyi@
nhri.org.tw.
aAbbreviations: AMG, [14C]-labeled R-methyl-D-glucopyranoside;
CHO, Chinese hamster ovary; DDQ, 2,3-dichloro-5,6-dicyano-1,4-ben-
zoquinone; HbA1c, glycosylated hemoglobin; NIDDM, non-insulin depen-
dent diabetes mellitus; SAR, structure-activity relationship; SD, Sprague-
Dawley; SGLT, sodium-dependent glucose cotransporter; SLC5A, solute
carrier family 5A; STZ, streptozotocin.
r
pubs.acs.org/jmc
Published on Web 12/03/2010
2010 American Chemical Society

Article
Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1 167
Figure 1. Structures of known SGLT2 inhibitors.
Chemistry
The novel N-xylosides detailed in this account were synthe-
sized according to the well-established chemistry of Kissei
and Tanabe Seiyaku, with appropriate modifications.24-26 As
shown in Scheme 1, condensation of D-xylose 9 with freshly
prepared or commercially available indolines 10a-r at 50 °C
gave the β-linked D-xylosylindoline, which in turn, without
purification, underwent per-O-acetylation with acetic anhy-
dride in pyridine to furnish the fully protected β-xylosylindo-
line 11a-r. Initially, we took advantage of the strategy des-
cribed by Tanade Seiyaku, namely, treatment of D-xylose 9
with indoline 10a in mixed solvent under reflux conditions.
Although this method worked well for N-glucosides, it failed
to introduce the indoline into xylose, giving instead an intrac-
table mixture of products. Compounds 11a-r were then sub-
jected to oxidation using 2,3-dichloro-5,6-dicyano-1,4-benzoqui-
none (DDQ) to afford the indole derivatives 12a-r, from which
the indole-3-carboxaldehydes 13a-r were obtained through a
Vilsmeier-Haack formylation with POCl3 in DMF.28 Grignard
reaction of compounds 13a-r with phenylmagnesium bromide
to introduce the distal aryl ring followed by reduction of the
resulting alcohols 14a-r with triethylsilane (Et3SiH) and boron
Figure 2. Design of N-indolylxyloside SGLT2 inhibitors.
advantage.17-19 In contrast, C-glucosides are more meta-
bolically stable and combine higher oral bioavailability and
plasma exposure. Dapagliflozin 5 and canagliflozin 6 are the
corresponding C-aryl glucosides, which present good in vitro
and in vivo potencies against SGLT2 and are currently in
phase III clinical trials.20-22 N-Glucosides are another series
of SGLT2 inhibitors expected with greater metabolic stability
compared to O-glucosides, but evaluation of their therapeutic
potentials is difficult because no pharmacokinetic data or clin-
ical results have been presented.23 The absence of advanced
studies left us curious about the role of the N-glycoside SGLT2
inhibitors in the treatment of type 2 diabetes.
trifluoride diethyl etherate (BF3 OEt2) gave the corresponding
3
diarylmethanes 15a-r. Finally, the acetyl groups were removed
ꢀ
under Zemplen conditions to provide the desired β-linked N-
xylosides 16a-r.
Building on the encouraging disclosures of Tanabe Seiyaku
and Kissei,24-26 we initiated our program to explore the N-
glycoside SGLT2 inhibitors 8, in which the aglycone is a
3-substituted indole and the sugar unit is D-xylose instead of
the original D-glucose (Figure 2). D-Xylose is a natural
pentose, obtained from plants cell walls and fiber, which has
the same configuration at C-1, -2, -3, and -4 as D-glucose.
Recently, Lexicon Pharmaceuticals reported a series of SGLT2
inhibitors wherein the glucose was replaced with L-xylose
and the resulting O-xylosides found to be potent in vitro
and in vivo.27 The structural similarity of D-xylose versus
D-glucose and the positive results of O-xylosides suggested
to us that the use of N-linked β-D-xylosides to inhibit SGLT2
would be feasible. Herein, we describe the design, synthesis,
and SAR of the newly developed N-linked β-D-xylosides, as
well as the pharmacokinetic and animal studies for selected
compound.
The same synthetic strategy was applied to prepare 4-chloro-
indolyl-N-xylosides 19a-q over three steps using 13j as star-
ting material (Scheme 2). Addition of various Grignard rea-
gents to 13j introduced the substituted or nonsubstituted aryl,
heteroaryl, and benzofused rings at the distal ring position.
The resulting alcohols 17a-q were then reduced under stan-
dard conditions (Et3SiH and BF3 OEt2) to generate 18a-q,
which were hydrolyzed with sodium methoxide to yield the
desired products 19a-q.
Scheme 3 depicts the synthetic strategy for the installation
of various alkoxy substitutes at the para-position of the distal
phenyl ring. A Lewis acid mediated brominolysis of 18c with
boron tribromide (BBr3) to convert the methoxy group into
the corresponding phenol, followed by Zemplen deacetylation
with sodium methoxide, provided the intermediate 20. Further
alkylation of 20 was performed in the presence of either
3
ꢀ

168 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1
Scheme 1. Synthesis of N-Xylosides 16a-ra
Yao et al.
a Reagents and conditions: (a) (i) indolines 10a-r, EtOH, H2O, 50 °C, 24 h; (ii) Ac2O, pyridine, 0 °C to room temp, 16 h; (b) 2,3-dichloro-5,6-dicyano-
1,4-benzoquinone (DDQ), 1,4-dioxane, room temp, 18 h; (c) POCl3, DMF, 75 °C, 1 h; (d) phenylmagnesium bromide, THF, -78 °C, 1-3 h;
(e) BF3 OEt2, Et3SiH, CH2Cl2, CH3CN, 0 °C, 1-3 h; (f) NaOMe, MeOH, CH2Cl2, 0 °C to room temp, 2 h.
3
Scheme 2. Synthesis of N-Xylosides 19a-qa
a Reagents and conditions: (a) Grignard reagents, THF, -78 °C, 1-3 h; (b) BF3 OEt2, Et3SiH, CH2Cl2, CH3CN, 0 °C, 1-3 h; (c) NaOMe, MeOH,
3
CH2Cl2, 0 °C to room temp, 2 h.
cesium or potassium carbonate and alkyl bromides to afford
the corresponding N-xylosides 21a and 21b. In addition, 21c
was prepared by alkylation of 20 with 2-(2-ethoxyethoxy)-
ethyl mesylate. Purity of all the final compounds was found

Article
Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1 169
Scheme 3. Synthesis of N-Xylosides 20 and 21a-ca
Table 1. In Vitro Inhibitory Activity of N-Xylosides 16a-r on hSGLT2
compd
R
hSGLT2 EC50 (nM)a
16a
16b
16c
16d
16e
16f
16g
16h
16i
16j
16k
16l
16m
16n
16o
16p
16q
16r
1
H
7129 ( 1033
1094 ( 153
40589 ( 10644
3033 ( 911
12956 ( 4319
2237 ( 353
12415 ( 1612
23627 ( 1412
9821 ( 2407
865 ( 266
28217 ( 8178
10279 ( 976
10680 ( 3675
923 ( 92
1819 ( 590
5487 ( 1865
>50000
4-Me
5-Me
6-Me
7-Me
4-F
5-F
6-F
a Reagents and conditions: (a) (i) 1 M solution of BBr3 in CH2Cl2,
CH2Cl2, -78 to 0 °C, 4 h; (ii) NaOMe, MeOH, CH2Cl2, 0 °C to room
temp, 2 h; (b) for 21a, Cs2CO3, 2-phenoxyethyl bromide, DMF, room
temp, 17 h; for 21b, K2CO3, propargyl bromide, DMF, room temp, 21 h;
for 21c, Cs2CO3, CH3CH2(OCH2CH2)2OMs, DMF, room temp, 2 days.
7-F
4-Cl
5-Cl
6-Cl
7-Cl
4-Br
to be more than 95% unless otherwise stated, as measured
by reverse phase HPLC on a C18 column (see Supporting In-
formation).
4-NO2
4-OMe
4-OH
5-F, 6-Cl
8600 ( 2723
108 ( 21b
Results and Discussion
5
3 ( 0.7b
All the synthesized N-xylosides 16a-r, 19a-q, 20, and 21a-c
were evaluated for their in vitro inhibitory activity against
human SGLT2 (hSGLT2); the results are presented in
Tables 1-3. EC50 values were calaulated by measuring inhibition
of the sodium-dependent uptake of [14C]-labeled R-methyl-
D-glucopyranoside (AMG) into Chinese hamster ovary (CHO)
cells stably expressing hSGLT2.29 Because the difference of
protein expression levels of cells will result in the difference of
EC50 values, phlorizin 1 and dapagliflozin 5 were used as
referencecompoundsinthe in vitroactivityevaluation system.
We began with the SAR studies on the effect of the indole
moiety by fixing the phenyl ring in the distal position; the re-
sults are presented in Table 1. The unsubstituted indolyl ana-
logue 16a (EC50=7129 nM) was used as a template, to which
all structural modifications were compared. First, we attemp-
ted to introducean electron donating methyl group (16b-e) at
various positions of the indole moiety. The 4-methyl substitu-
tion (16b) was found to exhibit 6.5 times the potency against
hSGLT2 (EC50=1307 nM) compared to 16a, whereas substi-
tution at the 5-position (16c) resulted in drastic loss of potency
(EC50=40589 nM). When the methyl group was replaced with
an electron-withdrawing fluoro (16f-i) or chloro (16j-m)
group, the same trend was observed: the 4-substituted indole
moiety exhibited enhanced activity toward hSGLT2 compared
with the 5, 6, or 7-substituted moiety. Of the aforementioned
xylosides, the 4-chloro analogue 16j was the most potent
hSGLT2 inhibitor, with an EC50 value of 865 nM. On the basis
of the above results, more functional groups were incorporated
at the 4- position and the potency of these molecules eval-
uated. The inhibition exhibited by 4-bromoindolylxyloside 16n
(EC50 = 923 nM) was found to be the same as that exhibited
by 4-chloro analogue 16j. Substitution at the 4-position with
the strongly electron-withdrawing nitro group (16o) resulted
in 2-fold less potency compared to 16j. Regarding strongly
electron-donating substituents, the 4-methoxy analogue 16p
a The data were obtained from at least two independent experiments.
b The data were obtained from multiple independent experiments.
showed similar activity compared to the unsubstituted in-
dolyl analogue 16a, and the xyloside with a 4-hydroxy group
(16q) was significantly less potent (EC50 > 50 000 nM). The
disubstituted compound 16r was also prepared but proved
to be an ineffective inhibitor (EC50 = 8600 nM).
With the identification of potent compound 16j, we next
turned our attention to the optimization of the distal phenyl
ring by evaluation of substituents at various positions and the
use of benzofused and heteroaryl rings. Encouragingly, it was
found that when the methoxy group was placed in the para-
position, the resulting compound 19c showed significantly
more potency with an EC50 value of 275 nM (Table 2). In
contrast, shifting the methoxy group to the ortho- (19a) and
meta-position (19b) led to a 47-fold and 3-fold loss in hSGLT2
inhibitory activity, respectively, compared to 19c. Incorpora-
tion of the electron-withdrawing fluoro group onto the phenyl
ring gave compounds 19d and 19e, which are inferior inhibi-
tors of hSGLT2 compared with 19c. Retaining the essential
p-methoxy group, and adding an o-methoxy group gave com-
pound 19f which showed a drastic loss of activity (EC50
=
30931 nM). Replacing the o-methoxy group with a m-fluoro
group (19g) led to a slight decrease in activity, indicating that
monosubstitution at para-position of the phenyl ring is opti-
mal. We also replaced the p-methoxyphenyl group with dif-
ferent benzofused rings such as 6-methoxynaphthalene (19o),
benzodioxole (19p), and benzodioxane (19q). It was observed
that compounds 19o and 19p showed 2- to 3-fold loss of
activity compared to 19c, but the benzodioxane analogue 19q
exhibited similar inhibitory activity to 19c, with an EC50 value
of 233 nM. Moreover, decreased activity was observed when
the heteroaromatic group, thiophene (19n), was attached in
place of the p-methoxyphenyl ring. This result clearly reveals

170 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1
Yao et al.
Table 2. In Vitro Inhibitory Activity of N-Xylosides 19a-g and 19n-q
on hSGLT2
Table 3. In Vitro Inhibitory Activity of N-Xylosides 19h-m and 20-22
on hSGLT2
compd
R
hSGLT2 EC50 (nM)a
19c
19h
19i
19j
19k
19l
19m
20
OMe
iPr
275 ( 41
1288 ( 135
1410 ( 177
11137 ( 2277
426 ( 46
297 ( 4
nPr
Ph
SMe
N(CH3)2
cyclopropyl
OH
161 ( 7
255 ( 71
1334 ( 154
530 ( 77
577 ( 49
241 ( 45
108 ( 21b
3 ( 0.7b
21a
21b
21c
22
OCH2CH2OPh
OCH2CtCH
(OCH2CH2)2OEt
1
5
a The data were obtained from at least two independent experiments.
b The data were obtained from multiple independent experiments.
toward hSGLT2. The 4-bromoindolyl-p-cyclopropyl deriva-
tive 22 was hence prepared to evaluate the corresponding bio-
logical activity for comparison purposes. Little difference in
potency was observed between xyloside 19m (EC50 = 161 nM)
and 22 (EC50 = 241 nM).
Compounds with good hSGLT2 inhibition were selected
for further study of their selectivity for hSGLT1 over hSGLT2.
The results are presented in Table 4. By use of the same pro-
tocol for the hSGLT2 inhibition measurement, the EC50
values of hSGLT1 were obtained by measuring inhibition of
the sodium-dependent uptake of [14C]AMG into CHO cells
stably expressing hSGLT1.29,30 Unfortunately, all the SGLT2
inhibitors tested showed no significant selectivity for hSGLT2
versus hSGLT1 (hSGLT1/hSGLT2 = 0.8-2.1). Among them,
compound 19l with a p-N,N-dimethylamino substituent was
found to be the least potent inhibitor in suppressing the reup-
take of hSGLT1 with an EC50 = 628.8 nM. The selectivity for
hSGLT2 over hSGLT1 was 2.1.
Among the aforementioned N-xylosides, the most potent
SGLT2 inhibitor 19m waschosenfor furtherpharmacokinetic
and animal studies.20 Administration of a single 1 mg/kg in-
travenously to rats revealed that 19m has the low clearance
and good oral bioavailability in rats, indicating that 19m is
stable (Table 5). Our in vitro metabolic stability study also
suggested that 19m was resistant upon metabolic incubation
with about 100% remaining after 30 min of rat microsomal
incubation (data not shown). After oral administration of a
1 mg/kg dose of 19m to rats, a Cmax of 1631 ng/mL was attained
at 2 h. The elimination half-life of 19m was 4.1 h in rats.
As shown in Figure 3A, statistically significant dose-depen-
dent efficacy on increasing urine glucose excretion of doses
ranging from 3 to 50 mg/kg of 19m was measured, resulting in
a 12- to 783-fold elevation in glucosuria relative to the vehicle
control. Oral administration of compound 19m in a single
dose of 3, 10, 30, and 50 mg/kg to SD rats induced urine
a The data were obtained from at least two independent experiments.
b The data were obtained from multiple independent experiments.
the importance of the para-substituted nature of the phenyl
ring of N-xylosides for good inhibitory activity. The effect of
substituents on the distal phenyl ring was further explored in
order to identify the optimal substituent at the para-position.
As illustrated in Table 3, replacing the p-methoxy group
with an isopropyl (19h) or n-propyl (19i) group weakened
hSGLT2 inhibition 5-fold. Substitution with the more bulky
4-phenyl group (19j) resulted in drastic loss of potency. When
the methylthio group was substituted for the methoxy group,
the resulting xyloside 19k was a slightly less potent hSGLT2
inhibitor, with an EC50 value of 426 nM. Introduction of
strongly electron-donating groups such as N,N-dimethylami-
no or hydroxyl groups at the para-position led to xylosides 19l
and 20, the potency of which was similar to 19c. Furthermore,
capping of the hydroxyl group with various alkyl chains
(21a-c) led to a decrease in activity, suggesting that a longer
alkyl chain may not be tolerated at the para-position. It is note-
worthy that when the p-cyclopropyl substituent was introduced
(19m), the activity improved slightly and the potency was
similar to that of phlorizin 1 with an EC50 value of 161 nM.
On the other hand, 16n (EC50 = 923 nM) and 16j (EC50 =
865 nM) were identified with the same inhibitory potency

Article
Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1 171
Table 4. In Vitro hSGLT Inhibition and Selectivity Data for N-Xylo-
sides 19c, 19k-m, 19q, 20, and 22
hSGLT2
EC50 (nM)
hSGLT1
EC50 (nM)a
selectivity
hSGLT1/hSGLT2
compd
19c
19k
19l
19m
19q
20
275 ( 41
426 ( 46
297 ( 4
161 ( 7
233 ( 29
255 ( 71
241 ( 45
108 ( 21
3 ( 0.7
229.5 ( 28.8
461.6 ( 53.9
628.8 ( 111.7
208.6 ( 17.2
305.9 ( 87.7
233.9 ( 41.5
327.3 ( 74.5
197.4 ( 69.7b
482.5 ( 116.8b
0.8
1.1
2.1
1.3
1.3
0.9
1.4
1.8
161
22
1
5
a The data were obtained from a single experiment. b The data were
obtained from three independent experiments.
Table 5. Pharmacokinetic Properties of N-Xyloside 19m after Oral and
Intravenous Administration to Rats
parameter
unit
mg/kg
iv
1.0
po
1.0
dose
T1/2
h
3.6
1.2
4.1
clearance
Vss
mL/min/kg
L/kg
0.32
Cmax
Tmax
ng/mL
h
ng h/mL
1631.3
2.0
AUC(0-inf)
F
14787
8370
56.3
3
%
Figure 4. Antihyperglycemic effect of single oral dosing (10 mg/kg)
of 19m and dapagliflozin 5 in STZ-induced diabetic Sprague-Dawley
rats: (A) change in blood glucose levels; (B) suppression of glucose
AUC. Data are expressed as the mean ( SEM (n = 3/group): /, P <
0.05 vs vehicle.
measured in the glucosuria experiment, and the results are
shown in Figure 3B. Compound 19m was found to increase
urine volume in a dose-dependent manner. For assessment of
the antihyperglycemic effect, streptozotocin (STZ) induced
(STZ at 65 mg/kg, ip) diabetic SD rats (blood glucose of
>450 mg/dL) were used to evaluate the capacity of 19m to
lower blood glucose. Blood samples were obtained from the tail
vein at 0, 0.5, 1, 2, 3, 4, and 5 h after a single oral administration
of 19m (10 mg/kg), 5 (10 mg/kg), or vehicle for blood glucose
analysis. Figure 4A illustrates the gradual decrease in blood
glucose level during the observation period of 5 h. The areas
under the glucose levels versus time curve (AUC) were calcu-
lated (Figure 4B); 19m at 10 mg/kg was found to cause a 24%
reduction in blood glucose level compared with the control.
It was found that 19m significantly increased urine glucose
excretion and urine volume. Although 19m increased urine
glucose excretion less than 5, 19m was equally effective at in-
creasing urine volume excretion. Furthermore, the antihyper-
glycemic effects of 19m and 5 at the same dose of 10 mg/kg
were not significantly different. The diarrheogenic activitytest
for 19m was also carried out to explore the possible adverse
effects involved in the lack of selectivity for hSGLT2 versus
hSGLT1. Herein no diarrhea was observed in the 19m-treated
mice within 8 h after the oral administration with a single dose
at a range of 10, 50, and 200 mg/kg.
Conclusions
Figure 3. Dose-dependent response of N-xyloside 19m over 24 h in
normal Sprague-Dawley rats: (A) urine glucose excretion; (B) urine
volume. Data are expressed as the mean ( SEM (n = 4/group): /,
P < 0.05 vs vehicle.
The synthesis and structure-activity relationships of N-linked
β-D-xylosides as SGLT2 inhibitors have been explored.
N-linked glycosides were expected to have greater metabolic
stability compared to O-glucosides, and because the config-
uration at C-1, -2, -3, and -4 of D-xylose and D-glucose is the
same, the use of N-linked β-D-xylosides was considered to be a
glucose excretions of 2, 19, 372, and 701 mg of glucose per 200 g
of body weight over 24 h, respectively. Urine volumes were also

172 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1
Yao et al.
worthy approach for further study on SGLT2 inhibition. Cur-
rent SAR studies show that of the compounds tested, com-
pound 19m, 4-chloroindolyl-N-xyloside with the 4-cyclopro-
pylbenzyl at the distal position, exhibited the best in vitro
inhibitory activity against SGLT2. The pharmacokinetic data
demonstrated that 19m was metabolically stable with a low
clearance and good oral bioavailability in SD rats. In further
efficacy studies, 19m was found to significantly increase 12- to
783-fold urine glucose excretion in normal SD rats and lower
blood glucose levels of STZ-induced diabetic rats. Although
the synthesized N-linked β-D-xylosides are less potent than
dapagliflozin and exhibit almost no selectivity for SGLT2
over SGLT1, these results are a good starting point for further
study in the absence of other information about N-glycoside
SGLT2 inhibitors. Our research is ongoing, and the results of
further investigations will be reported in due course.
General Procedure for the Synthesis of Compounds 11a-r. D-
Xylose 9 (1.5 equiv) was added to a solution of indolines 10a-r
(1.0 equiv) in EtOH/H2O (7/1) at room temperature. The mix-
ture was heated to 50 °C and maintained at this temperature for
24 h. The solvent was removed under reduced pressure and the
crude product used for per-acetylation without further purifica-
tion. This crude product was dissolved in pyridine, cooled to
0 °C, and stirred under nitrogen. Acetic anhydride was added.
The mixture was warmed to room temperature gradually and
stirred overnight (∼16 h). The mixture was cooled to 0 °C and
neutralized with 1 N HCl (aq). The mixture was extracted with
dichloromethane and the organic layer dried over MgSO4, fil-
tered, and concentrated under reduced pressure. The residue
was purified by column chromatography to afford the desired
products 11a-r.
General Procedure for the Synthesis of Compounds 12a-r.
DDQ (2.0 equiv) was added to a solution of 11a-11r (1.0 equiv)
in 1,4-dioxane at room temperature. After stirring for 18 h, the
reaction was quenched by the addition of saturated aqueous
sodium bicarbonate. The mixture was concentrated under re-
duced pressure and then extracted with ethyl acetate. The com-
bined organic layers were washed with saturated aqueous sodium
bicarbonate and brine. The organic phase was dried over MgSO4,
filtered, and concentrated under reduced pressure. The residue
was purified by column chromatography to afford the desired
products 12a-r.
General Procedure for the Synthesis of Compounds 13a-r.
POCl3 (8.0 equiv) was added to a stirred solution of 12a-r (1.0
equiv) in anhydrous DMF at room temperature under nitrogen.
The mixture was warmed to 75 °C and heated for 1 h. The reac-
tion was quenched with water at 0 °C and then extracted with
dichloromethane. The organic phase was dried over MgSO4,
filtered, and concentrated under reduced pressure. The residue
was purified by flash column chromatography to provide 13a-r.
General Procedure for the Synthesis of Compounds 14a-r and
17a-q. A 1 M solution of aryl- or heteroarylmagnesium bro-
mide in THF (4.0 equiv) was added to a solution of 13a-r (1.0
equiv) in THF at -78 °C under argon. After being stirred at -78 °C
for 3 h, the mixture was poured into saturated aqueous ammonium
chloride. The mixture was extracted with dichloromethane
and the organic layer was dried over MgSO4, filtered, and con-
centrated under reduced pressure to give the crude 14a-r or
17a-q. The residue was used for the next step without further
purification.
Experimental Section
General Methods. All chemicals were purchased as reagent
grade and used without further purification unless otherwise
stated. Column chromatography was performed with silica gel
(Merck Kieselgel 60, 230-400 mesh). Reactions were monitored
with thin-layer chromatography (TLC) using Merck 60 F254
silica gel glass-backed plates (5N10 cm2) and visualized under
UV (254 nm) and/or by staining with acidic ceric ammonium
molybdate. 1H NMR spectrawere recorded on a Varian Mercury-
300 (300 MHz) or Mercury-400 (400 MHz) spectrometer. Chemical
shifts (in ppm) are reported relative to the internal standard signal
of CD3OD (δ=3.30 ppm). 13C NMR spectra were obtained with
Varian Mercury-300 (75 MHz) or Mercury-400 (100 MHz) spec-
trometer and reported relative to CD3OD (δ=49.00 ppm). Cou-
pling constants (J) are reported in hertz (Hz). Splitting patterns are
described by using the following abbreviations: s, singlet; d, doublet;
t, triplet; q, quartet; sext, sextet; sept, septet; m, multiplet. LC/MS
data were obtained on an Agilent MSD-1100 ESI-MS/MS system.
High-resolution mass spectra were obtained on a MAT-95XL or
JEOL JMS-700 high resolution mass spectrometer in electron im-
pact (EI) or fast atom bombardment (FAB) ionization modes.
Purity of the final compounds was determined on a Hitachi 2000
series HPLC system with a reverse phase C18 column (Agilent
ZORBAX Eclipse XDB-C18 5 μm, 4.6 mm ꢀ 150 mm, 0.5 mL/min
flow rate). Mobile phase A was acetonitrile. Mobile phase B was
10 mM NH4OAc aqueous solution containing 0.1% formic acid.
The gradient system started from A/B (10%:90%) at 0 min to A/B
(90%:10%) at 45 min. All tested compounds were found to be
>95% pure at 254 nm unless otherwise stated.
General Precedure for the Synthesis of Indolines 10b-e, 10i-l,
and 10p,q. Acetic acid (10 mL/1 mmol indole) was added to a
mixture of indole (1.0 equiv) and NaBH3CN (3.0 equiv) at 0 °C
under nitrogen. The mixture was warmed to room temperature
and stirred for 2-5 h. The reaction mixture was concentrated
under reduced pressure and the residue redissolved in CH2Cl2
and washed with saturated aqueous sodium bicarbonate. The
organic layer was dried over MgSO4, filtered, and concentrated
under reduced pressure. The residue was purified by flash col-
umn chromatography to afford the desired indoline derivatives.
General Precedure for the Synthesis of Indolines 10f-h,
10m-o, and 10r. A solution of indole (1.0 equiv), Et3SiH (2.6
equiv), and TFA (1.5 mL/1 mmol indole) was heated at 50 °C
under nitrogen, with stirring. After 0.5-3 h, the mixture was con-
centrated under reduced pressure. The residue was diluted with
CH2Cl2 and neutralized by the addition of saturated aqueous
sodium bicarbonate. The organic phase was separated and the
aqueous phase extracted with CH2Cl2. The organic layers were
combined, dried over MgSO4, filtered, and concentrated under
reduced pressure. The residue was purified by flash column chro-
matography to provide the desired indoline derivatives.
General Procedure for the Synthesis of Compounds 15a-r and
18a-q. Et3SiH (5.0 equiv) and then BF3 OEt2 (0.5 equiv) were
3
added to a stirred solution of crude 14a-r or 17a-q in CH3CN/
CH2Cl2 (2/1) at 0 °C under argon. After 1 h, the reaction was
quenched by the addition of saturated aqueous sodium bicar-
bonate and extracted with dichloromethane. The organic layer
was dried over MgSO4, filtered, and concentrated under reduced
pressure. The residue was purified by column chromatography
to give 15a-r or 18a-q as a white solid.
General Procedure for the Synthesis of Compounds 16a-r and
19a-q. A 30% solution of NaOMe in MeOH (3.6 equiv) was
added to a solution of 15a-r or 18a-q (1.0 equiv) in MeOH/
CH2Cl2 (2/1) at 0 °C under nitrogen. The mixture was slowly
warmed to room temperature and then neutralized with acidic
resin after 2 h. The reaction mixture was filtered and the filtrate
concentrated under reduced pressure. The residue was purified
by column chromatography to give the desired products 16a-r
or 19a-q as a white solid.
3-Benzyl-1-(β-D-xylopyranosyl)-1H-indole (16a). The title
compound was obtained from D-xylose 9 and 2,3-dihydro-1H-
indole 10a according to the general procedure in 9% overall
yield. 1H NMR (300 MHz, CD3OD) δ 7.46 (d, J = 8.1 Hz, 1H),
7.38 (d, J = 7.8 Hz, 1H), 7.29-7.20 (m, 4H), 7.15-7.10 (m, 3H),
7.01-6.96 (m, 1H), 5.32 (d, J = 9.0 Hz, 1H), 4.06 (s, 2H), 3.95
(dd, J = 10.8, 5.4 Hz, 1H), 3.86 (t, J = 9.0 Hz, 1H), 3.70-3.62
(m, 1H), 3.50 (t, J = 9.0 Hz, 1H), 3.46 (t, J = 10.8 Hz, 1H). 13
C

Article
Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1 173
NMR (100 MHz, CD3OD) δ 142.6, 138.7, 130.1, 129.8, 129.3,
126.9, 124.5, 123.0, 120.7, 120.2, 117.1, 111.4, 87.4, 79.3, 73.5,
71.2, 69.5, 32.6. ESMS m/z: 340 (MHþ), 362 (MNaþ). HRMS
(EI) for C20H21NO4: calcd, 339.1471; found, 339.1468.
3-Benzyl-4-methyl-1-(β-D-xylopyranosyl)-1H-indole (16b). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-4-
methyl-1H-indole 10b according to the general procedure in 7%
overall yield. 1H NMR (300 MHz, CD3OD) δ 7.31-7.11 (m, 6H),
7.02-6.97 (m, 2H), 6.73-6.70 (m, 1H), 5.31 (d, J = 9.0 Hz, 1H),
4.25 (s, 2H), 3.95 (dd, J = 11.1, 5.4 Hz, 1H), 3.85 (t, J = 9.0 Hz,
1H), 3.69-3.61 (m, 1H), 3.50 (t, J = 9.0 Hz, 1H), 3.46 (t, J = 11.1
Hz, 1H), 2.41 (s, 3H). 13C NMR (75 MHz, CD3OD) δ 143.5,
139.4, 132.0, 129.6, 129.4, 128.6, 126.9, 125.6, 123.1, 122.5, 116.9,
109.3, 87.2, 79.3, 73.4, 71.6, 71.2, 69.5, 34.2, 20.4. ESMS m/z:
354 (MHþ), 376 (MNaþ). HRMS (EI) for C21H23NO4: calcd,
353.1627; found, 353.1622.
3-Benzyl-5-methyl-1-(β-D-xylopyranosyl)-1H-indole (16c). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-5-
methyl-1H-indole 10c according to the general procedure in 10%
overall yield. 1H NMR (300 MHz, CD3OD) δ 7.35-7.11 (m, 7H),
7.05 (s, 1H), 6.97 (dd, J = 8.4, 0.9 Hz, 1H), 5.27 (d, J = 9.0 Hz,
1H), 4.02 (s, 2H), 3.94 (dd, J = 10.8, 5.1 Hz, 1H), 3.85 (t, J = 9.0
Hz, 1H), 3.69-3.61 (m, 1H), 3.50 (t, J = 9.0 Hz, 1H), 3.44 (t, J =
10.8 Hz, 1H), 2.34 (s, 3H). 13C NMR (100 MHz, CD3OD) δ 142.7,
137.1, 130.3, 130.0, 129.8, 129.3, 129.3, 124.7, 124.5, 119.9, 116.6,
111.1, 87.5, 79.3, 73.5, 71.2, 69.5, 32.5, 21.6. ESMS m/z: 354
(MHþ), 376 (MNaþ). HRMS (EI) for C21H23NO4: calcd, 353.
1627; found, 353.1625.
3-Benzyl-6-methyl-1-(β-D-xylopyranosyl)-1H-indole (16d). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-6-
methyl-1H-indole 10d according to the general procedure in 13%
overall yield. 1H NMR (300 MHz, CD3OD) δ 7.28-7.10 (m, 7H),
7.02 (s, 1H), 6.83 (d, J = 8.7 Hz, 1H), 5.29 (d, J = 9.0 Hz, 1H),
4.03 (s, 2H), 3.94 (dd, J = 11.1, 5.1 Hz, 1H), 3.88 (t, J = 9.0 Hz,
1H), 3.69-3.61 (m, 1H), 3.50 (t, J = 9.0 Hz, 1H), 3.45 (t, J = 11.1
Hz, 1H), 2.41 (s, 3H). 13C NMR (100 MHz, CD3OD) δ 142.7,
139.2, 132.8, 129.8, 129.3, 128.0, 126.9, 123.9, 122.4, 120.0, 117.1,
111.3, 87.3, 79.3, 73.5, 71.2, 69.5, 32.6, 22.0. ESMS m/z: 354
(MHþ), 376 (MNaþ). HRMS (EI) for C21H23NO4: calcd,
353.1627; found, 353.1624.
3.69-3.63 (m, 1H), 3.50 (t, J = 9.0 Hz, 1H), 3.45 (t, J = 11.2 Hz,
1H). 13C NMR (75 MHz, CD3OD) δ 160.9, 157.8, 142.3, 135.2,
130.5, 130.4, 129.8, 129.4, 127.1, 126.5, 117.1, 117.0, 112.4, 112.3,
111.0, 110.7, 105.1, 104.8, 87.7, 79.2, 73.6, 71.2, 69.6, 32.5. ESMS
m/z: 358 (MHþ), 380 (MNaþ). HRMS (EI) for C20H20FNO4:
calcd, 357.1376; found, 357.1371. HPLC purity 83.6%.
3-Benzyl-6-fluoro-1-(β-D-xylopyranosyl)-1H-indole (16h). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-
6-fluoro-1H-indole10h accordingtothe general procedurein34%
overall yield. 1H NMR (400 MHz, CD3OD) δ 7.31 (dd, J = 8.6,
5.2 Hz, 1H), 7.27-7.11 (m, 6H), 7.11 (s, 1H), 6.79-6.74 (m, 1H),
5.25 (d, J = 9.2 Hz, 1H), 4.04 (s, 2H), 3.96 (dd, J = 11.1, 5.2 Hz,
1H), 3.81 (t, J = 9.2 Hz, 1H), 3.69-3.63 (m, 1H), 3.50 (t, J = 9.2
Hz, 1H), 3.46 (t, J = 11.1Hz, 1H). 13C NMR (100 MHz, CD3OD)
δ162.6, 160.2, 142.4, 138.9, 138.7, 129.8, 129.4, 127.0, 126.6, 125.1,
125.0, 121.2, 121.1, 117.3, 109.1, 108.8, 98.1, 97.8, 87.5, 79.2, 73.6,
71.1, 69.5, 32.5. ESMS m/z: 358 (MHþ), 380 (MNaþ). HRMS
(EI) for C20H20FNO4: calcd, 357.1376; found, 357.1375.
3-Benzyl-7-fluoro-1-(β-D-xylopyranosyl)-1H-indole (16i). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-
7-fluoro-1H-indole 10i according to the general procedure in 14%
overall yield. 1H NMR (400 MHz, CD3OD) δ 7.27-7.12 (m, 7H),
6.95-6.83 (m, 2H), 5.56 (d, J =9.2 Hz, 1H), 4.05 (s,2H), 3.94 (dd,
J = 11.2, 5.6 Hz, 1H), 3.81 (td, J = 9.2, 2.0 Hz, 1H), 3.66-3.60
(m, 1H), 3.47 (t, J = 9.2 Hz, 1H), 3.40 (t, J = 11.2 Hz, 1H). 13
C
NMR (75 MHz, CD3OD) δ 152.9, 149.7, 142.3, 133.9, 129.8,
129.4, 127.1, 125.5, 121.0, 120.9, 118.4, 116.4, 116.3, 109.3, 109.0,
88.7, 79.4, 73.9, 71.1, 69.5, 32.5. ESMS m/z: 358 (MHþ), 380
(MNaþ). HRMS (EI) for C20H20FNO4: calcd, 357.1376; found,
357.1374.
3-Benzyl-4-chloro-1-(β-D-xylopyranosyl)-1H-indole (16j). The
title compound was obtained from D-xylose 9 and 4-chloro-2,3-
dihydro-1H-indole 10j according to the general procedure in
41% overall yield. 1H NMR (400 MHz, CD3OD) δ 7.43 (d, J =
8.0 Hz, 1H), 7.25-7.19 (m, 4H), 7.16-7.11 (m, 2H), 7.07 (t, J =
7.6 Hz, 1H), 7.02-6.98 (m, 2H), 5.31 (d, J = 9.0 Hz, 1H), 4.33 (s,
2H), 3.95 (dd, J = 11.2, 5.6 Hz, 1H), 3.78 (t, J = 9.0 Hz, 1H),
3.67-3.61(m, 1H), 3.49 (t, J = 9.0 Hz, 1H), 3.45 (t, J = 11.2 Hz,
1H). 13C NMR (100 MHz, CD3OD) δ 143.0, 140.2, 129.9, 129.3,
127.4, 126.9, 126.8, 126.5, 123.6, 121.8, 117.1, 110.5, 87.4, 79.2,
73.6, 71.1, 69.6, 33.5. ESMS m/z: 374 (MHþ), 396 (MNaþ).
HRMS (EI) for C20H20ClNO4: calcd, 373.1081; found, 373.
1074.
3-Benzyl-5-chloro-1-(β-D-xylopyranosyl)-1H-indole (16k). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-
5-chloro-1H-indole 10k according to the general procedure in
24% overall yield. 1HNMR(400 MHz, CD3OD) δ7.44 (d, J =8.8
Hz, 1H), 7.33 (d, J = 2.0 Hz, 1H), 7.26-7.22 (m, 4H), 7.17-7.14
(m, 2H), 7.09 (dd, J = 8.8, 2.0 Hz, 1H), 5.29 (d, J = 8.8 Hz, 1H),
4.02 (s, 2H), 3.95 (dd, J = 11.0, 5.2 Hz, 1H), 3.82 (t, J = 8.8 Hz,
1H), 3.69-3.63 (m, 1H), 3.50 (t, J= 8.8 Hz, 1H), 3.45 (t, J=11.0 Hz,
1H). 13C NMR (75 MHz, CD3OD) δ 142.6, 137.0, 131.0, 129.7,
129.4, 127.0, 126.4, 126.2, 123.0, 119.6, 116.7, 112.7, 87.5, 79.1,
73.5, 71.0, 69.5, 32.3. ESMS m/z: 374 (MHþ), 396 (MNaþ).
HRMS (EI) for C20H20ClNO4: calcd, 373.1081; found, 373.1082.
3-Benzyl-6-chloro-1-(β-D-xylopyranosyl)-1H-indole (16l). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-
6-chloro-1H-indole 10j according to the general procedure in 31%
overall yield. 1H NMR (400 MHz, CD3OD) δ 7.50 (d, J = 1.6 Hz,
1H), 7.33 (dd, J = 8.6, 0.8 Hz, 1H), 7.28-7.22 (m, 4H), 7.17-7.15
(m, 2H), 6.99-6.97 (m, 1H), 5.29 (d, J = 8.8Hz, 1H), 4.05 (s, 2H),
3.97 (dd, J = 11.1, 5.6 Hz, 1H), 3.81 (t, J = 8.8 Hz, 1H), 3.70-
3.64 (m, 1H), 3.51 (t, J = 8.8 Hz, 1H), 3.48 (t, J = 11.1 Hz, 1H).
13C NMR (75 MHz, CD3OD) δ 142.1, 138.9, 129.7, 129.3, 128.9,
128.6, 127.0, 125.5, 121.3, 121.2, 117.2, 111.5, 87.3, 79.0, 73.5, 71.0,
69.4, 32.3. ESMS m/z: 374 (MHþ), 396 (MNaþ). HRMS (EI) for
C20H20ClNO4: calcd, 373.1081; found, 373.1076.
3-Benzyl-7-methyl-1-(β-D-xylopyranosyl)-1H-indole (16e). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-7-
methyl-1H-indole 10e according to the general procedure in 3%
overall yield. 1H NMR (300 MHz, CD3OD) δ 7.28-7.10 (m, 7H),
6.90-6.85 (m, 2H), 5.69 (d, J = 8.7 Hz, 1H), 4.04 (s, 2H), 3.91 (dd,
J = 11.0, 5.1Hz, 1H), 3.85 (t, J= 8.7 Hz, 1H), 3.66-3.58 (m, 1H),
3.48 (t, J = 8.7 Hz, 1H), 3.42 (t, J = 11.0 Hz, 1H), 2.68 (s, 3H). 13
C
NMR (100 MHz, CD3OD) δ 142.5, 137.8, 130.7, 129.8, 129.3,
126.9, 126.6, 124.1, 122.1, 120.8, 118.3, 117.4, 87.4, 79.5, 73.6, 71.1,
68.7, 32.5, 20.4. ESMS m/z: (MHþ), 376 (MNaþ). HRMS (EI) for
C21H23NO4: calcd, 353.1627; found, 353.1619.
3-Benzyl-4-fluoro-1-(β-D-xylopyranosyl)-1H-indole (16f). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-
4-fluoro-1H-indole 10f according to the general procedure in
4% overall yield. 1H NMR (300 MHz, CD3OD) δ 7.29-7.02 (m,
8H), 6.71-6.64 (m, 1H), 5.30 (d, J = 9.0 Hz, 1H), 4.14 (s, 2H),
3.95 (dd, J = 10.8, 5.4 Hz, 1H), 3.80 (t, J = 9.0 Hz, 1H), 3.69-
3.61 (m, 1H), 3.49 (t, J = 9.0 Hz, 1H), 3.46 (t, J = 10.8 Hz, 1H).
13C NMR (100 MHz, CD3OD) δ 159.8, 157.4, 142.9, 141.5,
141.4, 129.8, 129.3, 126.9, 125.0, 123.6, 123.5, 118.2, 115.8,
107.9, 107.8, 105.9, 105.7, 87.6, 79.2, 73.6, 71.1, 69.6, 33.5.
ESMS m/z: 358 (MHþ), 380 (MNaþ). HRMS (EI) for
C20H20FNO4: calcd, 357.1376; found, 357.1372.
3-Benzyl-5-fluoro-1-(β-D-xylopyranosyl)-1H-indole (16g). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-5-
fluoro-1H-indole 10g according to the general procedure in 30%
overall yield. 1H NMR (400 MHz, CD3OD) δ 7.43 (dd, J = 8.8,
4.4 Hz, 1H), 7.28-7.13 (m, 6H), 7.00 (dd, J = 9.6, 2.4 Hz, 1H),
6.88 (td, J = 9.2, 2.4 Hz, 1H), 5.28 (d, J = 9.0 Hz, 1H), 4.02 (s,
2H), 3.94 (dd, J = 11.2, 5.6 Hz, 1H), 3.82 (t, J=9.0 Hz, 1H),
3-Benzyl-7-chloro-1-(β-D-xylopyranosyl)-1H-indole (16m). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-7-
chloro-1H-indole 10m according to the general procedure in 12%

174 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1
Yao et al.
overall yield. 1H NMR (300 MHz, CD3OD) δ 7.34 (dd, J = 8.0,
1.2 Hz, 1H), 7.28-7.12 (m, 7H), 6.95 (t, J = 7.8 Hz, 1H), 6.28 (d,
J = 9.6 Hz, 1H), 4.06 (s, 2H), 3.91 (dd, J = 10.9, 5.4 Hz, 1H), 3.85
(dd, J = 9.6, 8.7 Hz, 1H), 3.65-3.57 (m, 1H), 3.48 (t, J = 8.7 Hz,
1H), 3.44 (t, J = 10.9 Hz, 1H). 13C NMR (100 MHz, CD3OD) δ
142.0, 134.1, 133.2, 129.8, 129.4, 127.1, 125.5, 121.5, 119.3, 117.9,
117.7, 79.6, 73.3, 71.2, 68.9, 32.3. ESMS m/z: 374 (MHþ), 396
(MNaþ). HRMS (FAB) for C20H20ClNO4: calcd, 373.1081;
found, 373.1087.
3-Benzyl-4-bromo-1-(β-D-xylopyranosyl)-1H-indole (16n). The
title compound was obtained from D-xylose 9 and 4-bromo-2,3-
dihydro-1H-indole 10n according to the general procedure in
32% overall yield. 1H NMR (400 MHz, CD3OD) δ 7.48 (dd, J =
8.2, 0.8 Hz, 1H), 7.25-7.19 (m, 5H), 7.16-7.11 (m, 1H), 7.01 (s,
1H), 6.99 (t, J = 8.0 Hz, 1H), 5.30 (d, J = 9.2 Hz, 1H), 4.37, 4.34
(ABq, J = 16.4 Hz, 2H), 3.94 (dd, J = 11.2, 5.6 Hz, 1H), 3.78 (t,
J = 9.2Hz, 1H), 3.67-3.61 (m, 1H), 3.50 (t, J = 9.2Hz, 1H), 3.44
(t, J = 11.2 Hz, 1H). 13C NMR (100 MHz, CD3OD) δ 141.6,
138.7, 128.7, 128.0, 126.6, 125.9, 125.6, 124.1, 122.6, 116.2, 113.9,
109.8, 86.0, 77.9, 72.2, 69.8, 68.3, 32.2. ESMS m/z: 418 (MHþ),
440 (MNaþ). HRMS (EI) for C20H20BrNO4: calcd, 417.0576;
found, 417.0581.
3-Benzyl-4-nitro-1-(β-D-xylopyranosyl)-1H-indole (16o). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-
4-nitro-1H-indole 10o according to the general procedure in
12% overall yield. 1H NMR (300 MHz, CD3OD) δ 7.89 (dd, J =
8.1, 0.9 Hz, 1H), 7.65 (dd, J = 8.1, 0.9 Hz, 1H), 7.39 (s, 1H), 7.25
(t, J = 8.1 Hz, 1H), 7.20-7.06 (m, 5H), 5.44 (d, J = 9.0 Hz, 1H),
4.16 (s, 2H), 3.99 (dd, J = 10.8, 5.1 Hz, 1H), 3.81 (t, J = 9.0 Hz,
1H), 3.73-3.64 (m, 1H), 3.53 (t, J = 9.0 Hz, 1H), 3.50 (t, J =10.8
Hz, 1H). 13C NMR (75 MHz, CD3OD) δ 144.8, 142.5, 141.2,
130.4, 129.7, 129.4, 126.9, 121.9, 121.2, 118.3, 117.5, 116.0, 87.4,
79.1, 73.8, 71.0, 69.7, 34.2. ESMS m/z: 385 (MHþ), 407 (MNaþ).
HRMS (EI) for C20H20N2O6: calcd, 384.3826; found, 384.1326.
HPLC purity 94.1%.
3-Benzyl-4-methoxy-1-(β-D-xylopyranosyl)-1H-indole (16p). The
title compound was obtained from D-xylose 9 and 2,3-dihydro-4-
methoxy-1H-indole 10p according to the general procedure in 6%
overall yield. 1H NMR (300 MHz, CD3OD) δ 7.26-7.18 (m, 4H),
7.13-7.07 (m, 3H), 6.85 (s, 1H), 6.52-6.46 (m, 1H), 5.25 (d, J =
9.0 Hz, 1H), 4.19 (s, 2H), 3.93 (dd, J = 10.8, 5.4 Hz, 1H), 3.80 (t,
J = 9.0 Hz, 1H), 3.78 (s, 3H), 3.67-3.59 (m, 1H), 3.48 (t, J = 9.0
Hz, 1H), 3.39 (t, J = 10.8 Hz, 1H). 13C NMR(100 MHz, CD3OD)
δ156.3, 143.9, 140.4, 129.9, 129.1, 126.6, 123.9, 123.2, 119.7, 117.7,
104.7, 101.3, 87.4, 79.2, 73.5, 71.1, 69.5, 55.5, 34.1. ESMS m/z:
370 (MHþ), 392 (MNaþ). HRMS (EI) for C21H23NO5: calcd,
369.1576; found, 369.1579.
for C20H19ClFNO4: calcd, 391.0987; found, 391.0984. HPLC
purity 91.7%.
4-Chloro-3-(2-methoxybenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(19a). The title compound was obtained from 1-(2,3,4-tri-O-
acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and
2-methoxyphenylmagnesium bromide according to the general
1
procedure in 88% yield over three steps. H NMR (300 MHz,
CD3OD) δ 7.44 (d, J = 7.5 Hz, 1H), 7.20-7.14 (m, 1H), 7.08 (t,
J = 7.5 Hz, 1H), 7.01-6.94 (m, 4H), 6.83-6.77 (m, 1H), 5.30 (d,
J = 9.0 Hz, 1H), 4.30, 4.26 (ABq, J = 22.4 Hz, 2H), 3.84 (s, 3H),
3.95 (dd, J = 11.0, 5.3 Hz, 1H), 3.77 (t, J = 9.0 Hz, 1H), 3.68-
3.60 (m, 1H), 3.52-3.42 (m, 2H). 13C NMR (100 MHz, CD3OD)
δ 158.7, 140.2, 131.3, 131.0, 128.2, 127.5, 126.8, 126.6, 123.4,
121.7, 121.5, 116.5, 111.4, 110.5, 87.4, 79.2, 73.6, 71.1, 69.6, 56.0,
27.5. ESMS m/z: 404 (MHþ), 426 (MNaþ). HRMS (EI) for
C21H22ClNO5: calcd, 403.1187; found, 403.1187. HPLC purity
91.6%.
4-Chloro-3-(3-methoxybenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(19b). The title compound was obtained from 1-(2,3,4-tri-O-acetyl-
β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and 3-meth-
oxyphenylmagnesium bromide according to the general procedure
in 64% yield over three steps. 1H NMR (300 MHz, CD3OD) δ7.45
(dd, J = 8.1, 0.9 Hz, 1H), 7.19-6.99 (m, 4H), 6.83-6.71 (m, 3H),
5.32 (d, J = 9.0 Hz, 1H), 4.32 (s, 2H), 3.96 (dd, J = 11.0, 5.4 Hz,
1H), 3.79 (t, J = 9.0 Hz, 1H), 3.73 (s, 3H), 3.70-3.61 (m, 1H), 3.50
(t, J = 9.0 Hz, 1H), 3.46 (t, J = 11.0 Hz, 1H). 13C NMR (100
MHz, CD3OD) δ 159.9, 143.3, 138.8, 128.9, 126.1, 125.5, 125.2,
122.2, 121.0, 120.5, 115.6, 114.3, 110.9, 109.2, 86.1, 77.9, 72.2, 69.8,
68.3, 54.3, 32.2. ESMS m/z: 404 (MHþ), 426 (MNaþ). HRMS (EI)
for C21H22ClNO5: calcd, 403.1187; found, 403.1193.
4-Chloro-3-(4-methoxybenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(19c). The title compound was obtained from 1-(2,3,4-tri-O-acetyl-
β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and 4-meth-
oxyphenylmagnesium bromide according to the general procedure
in 67% yield over three steps. 1H NMR (400 MHz, CD3OD) δ7.42
(dd, J = 8.4, 0.8 Hz, 1H), 7.14-7.12 (m, 2H), 7.07 (t, J = 7.6 Hz,
1H), 7.00-6.98 (m, 2H), 6.82-6.80 (m, 2H), 5.30 (d, J = 9.0 Hz,
1H), 4.26 (s, 2H), 3.94 (dd, J = 11.2, 5.6 Hz, 1H), 3.77 (t, J = 9.0
Hz, 1H), 3.75 (s, 3H), 3.67-3.57 (m, 1H), 3.48 (t, J = 9.0 Hz, 1H),
3.45 (t, J = 11.2 Hz, 1H). 13C NMR (100 MHz, CD3OD) δ 159.4,
140.2, 135.0, 130.8, 127.4, 126.6, 126.5, 123.5, 121.8, 117.7, 114.7,
110.5, 87.4, 79.2, 73.6, 71.1, 69.6, 55.7, 32.7. ESMS m/z: 404
(MHþ). HRMS (EI) for C21H22ClNO5: calcd, 403.1187; found,
403.1181.
4-Chloro-3-(3-fluorobenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(19d). The title compound was obtained from 1-(2,3,4-tri-O-
acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and
3-fluorophenylmagnesium bromide according to the general
3-Benzyl-4-hydroxy-1-(β-D-xylopyranosyl)-1H-indole (16q). The
title compound was obtained from D-xylose 9 and 4-acetyl-2,3-
dihydro-1H-indole 10q according to the general procedure in 18%
overall yield. 1H NMR (300 MHz, CD3OD) δ 7.32-7.29 (m, 2H),
7.23-7.19 (m, 2H), 7.13-7.08 (m, 1H), 6.94-6.88 (m, 2H), 6.76 (s,
1H), 6.40-6.36 (m, 1H), 5.22 (d, J= 9.0 Hz, 1H), 4.24 (s, 2H), 3.92
(dd, J = 10.8, 5.1 Hz, 1H), 3.78 (t, J = 9.0 Hz, 1H), 3.66-3.57 (m,
1H), 3.46 (t, J = 9.0 Hz, 1H), 3.43 (t, J = 10.8 Hz, 1H). 13C NMR
(100 MHz, CD3OD) δ 153.6, 143.9, 140.9, 130.1, 130.0, 129.1,
126.6, 123.9, 122.8, 119.2, 118.0, 105.5, 103.2, 87.4, 79.2, 73.4, 71.2,
69.5, 33.9. ESMS m/z: 356 (MHþ), 378 (MNaþ). HRMS (EI) for
C20H21NO5: calcd, 355.1420; found, 355.1423.
1
procedure in 65% yield over three steps. H NMR (300 MHz,
CD3OD) δ 7.45 (d, J = 7.2 Hz, 1H), 7.27-7.20 (m, 1H), 7.14 (s,
1H), 7.11-6.98 (m, 3H), 6.92-6.83 (m, 2H), 5.34 (d, J = 9.0 Hz,
1H), 4.34 (s, 2H), 3.97 (dd, J = 10.9, 5.1 Hz, 1H), 3.82 (t, J = 9.0
Hz, 1H), 3.70-3.62 (m, 1H), 3.50 (t, J = 9.0 Hz, 1H), 3.47 (t, J =
10.9 Hz, 1H). 13C NMR (100 MHz, CD3OD) δ 165.7, 163.3,
146.4, 146.3, 140.2, 130.9, 130.8, 127.3, 127.1, 126.4, 125.7, 125.6,
123.7, 121.9, 116.4, 116.2, 115.9, 113.5, 113.3, 110.6, 87.4, 79.2,
73.6, 71.1, 69.6, 33.1. ESMS m/z: 392 (MHþ), 414 (MNaþ).
HRMS (FAB) for C20H19ClFNO4: calcd, 391.0987; found, 391.
0983.
3-Benzyl-6-chloro-5-fluoro-1-(β-D-xylopyranosyl)-1H-indole
(16r). The title compound was obtained from D-xylose 9 and
2,3-dihydro-6-chloro-5-fluoro-1H-indole 10r according to the
4-Chloro-3-(4-fluorobenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(19e). The title compound was obtained from 1-(2,3,4-tri-O-
acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and
4-fluorophenylmagnesium bromide according to the general
1
general procedure in 22% overall yield. H NMR (300 MHz,
1
CD3OD) δ 7.58 (d, J = 6.3 Hz, 1H), 7.26-7.13 (m, 7H), 5.28 (d,
J = 9.0 Hz, 1H), 4.02 (s, 2H), 3.97 (dd, J = 10.8, 5.1 Hz, 1H),
3.78 (t, J = 9.0 Hz, 1H), 3.71-3.63 (m, 1H), 3.51 (t, J = 9.0 Hz,
1H), 3.44 (t, J = 10.8 Hz, 1H). 13C NMR (100 MHz, CD3OD) δ
155.4, 153.0, 142.0, 134.9, 129.8, 129.5, 129.0, 128.9, 127.2,
127.1, 117.3, 116.5, 113.0, 106.4, 106.1, 87.7, 79.1, 73.7, 71.1,
69.6, 32.3. ESMS m/z: 392 (MHþ), 414 (MNaþ). HRMS (EI)
procedure in 66% yield over three steps. H NMR (300 MHz,
CD3OD) δ 7.44 (dd, J = 8.2, 0.6 Hz, 1H), 7.23-7.18 (m, 2H),
7.10-7.05 (m, 2H), 7.00-6.92 (m, 3H), 5.32 (d, J = 9.3 Hz, 1H),
4.31 (s, 2H), 3.95 (dd, J=11.1, 5.1 Hz, 1H), 3.79 (t, J=9.3 Hz,
1H), 3.69-3.61 (m, 1H), 3.49 (t, J = 9.3, 1H), 3.46 (t, J = 11.1,
1H). 13C NMR (75 MHz, CD3OD) δ 163.0, 159.8, 138.9, 137.8,
137.7, 130.1, 130.0, 126.0, 125.6, 125.1, 122.3, 120.5, 115.4, 114.6,

Article
Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1 175
114.3, 109.3, 86.1, 77.9, 72.2, 69.8, 68.3, 31.3. ESMS m/z: 392
(MHþ), 414 (MNaþ). HRMS (EI) for C20H19ClFNO4: calcd,
391.0987; found, 391.0983.
126.5, 123.6, 121.8, 116.9, 110.6, 87.4, 79.2, 73.6, 71.1, 69.6,
33.2. ESMS m/z: 450 (MHþ), 472 (MNaþ). HRMS (EI) for
C26H24ClNO4: calcd, 449.1394; found, 449.1397.
4-Chloro-3-(2,4-dimethoxybenzyl)-1-(β-D-xylopyranosyl)-1H-
indole (19f). The title compound was obtained from 1-(2,3,4-tri-
O-acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j
and 2,4-dimethoxyphenylmagnesium bromide according to
4-Chloro-3-[4-(methylsulfanyl)benzyl]-1-(β-D-xylopyranosyl)-
1H-indole (19k). The title compound was obtained from 1-(2,3,
4-tri-O-acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole
13j and 4-thioanisolemagnesium bromide according to the gen-
eral procedure in 65% yield over three steps. 1H NMR (300 MHz,
CD3OD) δ 7.43 (dd, J = 8.1, 0.9 Hz, 1H), 7.15 (s, 1H,), 7.18-6.98
(m, 6H), 5.31 (d, J = 9.0 Hz, 1H), 4.29 (s, 2H), 3.95 (dd, J = 11.2,
5.4 Hz, 1H), 3.78 (t, J = 9.0 Hz, 1H), 3.69-3.60 (m, 1H), 3.49 (t,
J= 9.0 Hz, 1H), 3.45 (t, J= 11.2, 1H), 2.42 (s, 3H). 13CNMR(100
MHz, CD3OD) δ 140.3, 140.2, 136.9, 130.6, 130.5, 130.4, 128.0,
127.4, 126.8, 126.5, 123.6, 121.8, 116.9, 110.6, 87.4, 79.2, 73.5, 71.1,
69.6, 33.0, 16.2. ESMS m/z: 420 (MHþ), 442 (MNaþ). HRMS (EI)
for C21H22ClNO4S: calcd, 419.0958; found, 419.0960.
1
the general procedure in 78% yield over three steps. H NMR
(400 MHz, CD3OD) δ 7.41 (d, J = 8.0 Hz, 1H), 7.05 (t, J = 8.0
Hz, 1H), 6.98 (d, J = 8.0 Hz, 1H), 6.88 (s, 1H), 6.86 (d, J = 8.0
Hz, 1H), 6.51 (s, 1H), 6.35 (d, J = 8.0 Hz, 1H), 5.27 (d, J = 9.0
Hz, 1H), 4.21, 4.18 (ABq, J = 16.8 Hz, 2H), 3.93 (dd, J = 11.0,
5.2 Hz, 1H), 3.78 (s, 3H), 3.78-3.73 (m, 1H), 3.73 (s, 3H), 3.66-
3.59 (m, 1H), 3.48 (t, J = 9.0 Hz, 1H), 3.43 (t, J = 11.0 Hz, 1H).
13C NMR (75 MHz, CD3OD) δ 160.8, 159.4, 140.1, 131.4, 127.5,
126.7, 126.4, 123.5, 123.4, 121.7, 117.0, 110.5, 105.2, 99.3,
87.4, 79.1, 73.5, 71.0, 69.5, 55.9, 55.8, 27.0. HRMS (EI) for
C22H24ClNO6: calcd, 433.1292; found, 433.1289.
4-Chloro-3-(3-fluoro-4-methoxybenzyl)-1-(β-D-xylopyranosyl)-
1H-indole (19g). The title compound was obtained from 1-(2,3,4-
tri-O-acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j
and 3-fluoro-4-methoxyphenylmagnesium bromide according
to the general procedure in 65% yield over three steps. 1H NMR
(300 MHz, CD3OD) δ 7.45 (d, J = 8.1 Hz, 1H), 7.12-6.91 (m,
6H), 5.33 (d, J = 9.0 Hz, 1H), 4.27 (s, 2H), 3.97 (dd, J = 11.0, 5.1
Hz, 1H), 3.83 (s, 3H), 3.81(t, J = 9.0 Hz, 1H), 3.71-3.62 (m, 1H),
3.51 (t, J = 9.0 Hz, 1H), 3.47 (t, J = 11.0 Hz, 1H). 13C NMR (100
MHz, CD3OD) δ 154.9, 152.5, 147.2, 147.1, 140.2, 136.6, 136.5,
127.3, 126.9, 126.4, 125.5, 125.4, 123.7, 121.9, 117.3, 117.1, 116.6,
114.7, 110.6, 87.4, 79.2, 73.6, 71.1, 69.6, 56.9, 32.5. ESMSm/z: 422
(MHþ), 444 (MNaþ). HRMS (EI) for C21H21ClNO5: calcd,
421.1092; found, 421.1083.
4-Chloro-3-(4-isopropylbenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(19h). The title compound was obtained from 1-(2,3,4-tri-O-acetyl-
β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and 4-iso-
propylphenylmagnesium bromide according to the general pro-
cedure in 78% yield over three steps. 1H NMR (400 MHz,
CD3OD) δ 7.43 (dd, J = 8.4, 0.8 Hz, 1H), 7.16-6.99 (m, 7H), 5.30
(d, J = 9.2 Hz, 1H), 4.30 (s, 2H), 3.95 (dd, J = 11.2, 5.2 Hz, 1H),
3.78 (t, J = 9.2 Hz, 1H), 3.68-3.62 (m, 1H), 3.49 (t, J = 9.2 Hz,
1H), 3.46 (t, J = 11.2 Hz, 1H), 2.86 (sept, J = 7.2 Hz, 1H), 1.23 (d,
J = 7.2 Hz, 6H). 13C NMR (100 MHz, CD3OD) δ 147.5, 140.3,
140.2, 129.9, 127.4, 127.3, 126.7, 126.6, 123.5, 121.8, 117.4, 110.5,
87.4, 79.2, 73.5, 71.1, 69.6, 35.1, 33.2, 24.7. ESMS m/z: 416 (MHþ),
438 (MNaþ). HRMS (EI) for C23H26ClNO4: calcd, 415.1550;
found, 415.1546.
4-Chloro-3-(4-n-propylbenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(19i). The title compound was obtained from 1-(2,3,4-tri-O-acetyl-
β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and 4-n-
propylphenylmagnesium bromide according to the general pro-
cedure in 66% yield over three steps. 1H NMR (300 MHz,
CD3OD) δ 7.43 (dd, J = 8.4, 0.9 Hz, 1H), 7.13-6.98 (m, 7H),
5.30 (d, J = 9.0 Hz, 1H), 4.29 (s, 2H), 3.94 (dd, J = 11.0, 5.1 Hz,
1H), 3.77 (t, J = 9.0 Hz, 1H), 3.68-3.59 (m, 1H), 3.48 (t, J = 9.0
Hz, 1H), 3.45 (t, J = 11.0 Hz, 1H), 2.53 (t, J = 7.5 Hz, 2H), 1.61
(sext, J = 7.5 Hz, 2H), 0.92 (t, J = 7.5 Hz, 3H). 13C NMR (100
MHz, CD3OD) δ 141.2, 140.3, 140.2, 129.8, 129.4, 127.4, 126.7,
126.6, 123.5, 121.8, 117.4, 110.5, 87.4, 79.2, 73.6, 71.1, 69.6, 38.8,
33.2, 25.9, 14.2. ESMS m/z: 416 (MHþ), 438 (MNaþ). HRMS
(EI) for C23H26ClNO4: calcd, 415.1550; found, 415.1555.
4-Chloro-3-(4-phenylbenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(19j). The title compound was obtained from 1-(2,3,4-tri-O-
acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and
4-biphenylmagnesium bromide according to the general proce-
dure in 78% yield over three steps. 1H NMR (400 MHz, CD3OD)
δ 7.58-7.28 (m, 10H), 7.10-6.99 (m, 3H), 5.32 (d, J = 9.2 Hz,
1H), 4.37 (s, 2H), 3.95 (dd, J = 11.2, 5.2 Hz, 1H), 3.80 (t, J = 9.2
Hz, 1H), 3.67-3.61 (m, 1H), 3.49 (t, J = 9.2 Hz, 1H), 3.45 (t, J =
11.2 Hz, 1H). 13C NMR (100 MHz, CD3OD) δ 142.5, 142.3,
140.2,140.1, 130.4, 130.0, 129.9, 128.1, 128.0, 127.9, 127.4, 126.9,
4-Chloro-3-[4-(N,N-dimethylamino)benzyl]-1-(β-D-xylopyrano-
syl)-1H-indole (19l). The title compound was obtained from 1-(2,
3,4-tri-O-acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole
13j and 4-(N,N-dimethylamino)phenylmagnesium bromide ac-
cording to the general procedure in 32% yield over three steps.
1H NMR (400 MHz, CD3OD) δ 7.42 (dd, J = 8.0, 0.8 Hz, 1H),
7.10-7.04 (m, 3H), 6.99 (dd, J = 8.0, 0.8 Hz, 1H), 6.93 (s, 1H),
6.76-6.72(m, 2H), 5.29 (d, J = 9.0 Hz, 1H), 4.23 (s, 2H), 3.94 (dd,
J = 11.2, 5.2Hz, 1H), 3.75 (t, J=9.0Hz, 1H), 3.66-3.60(m,1H),
3.48 (t, J =9.0 Hz, 1H), 3.44(t, J = 11.2 Hz, 1H), 2,86 (s, 6H). 13
C
NMR (75 MHz, CD3OD) δ 150.8, 140.2, 131.9, 130.5, 127.5,
126.6, 126.5, 123.5, 121.7, 118.2, 115.0, 110.5, 87.4, 79.2, 73.6,
71.1, 69.6, 41.7, 32.8. ESMS m/z: 417 (MHþ), 439 (MNaþ).
HRMS (EI) for C22H25ClN2O4: calcd, 416.1503; found, 416.1499.
4-Chloro-3-(4-cyclopropylbenzyl)-1-(β-D-xylopyranosyl)-1H-
indole (19m). The title compound was obtained from 1-(2,3,4-tri-
O-acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and
4-cyclopropylphenylmagnesium bromide according to the general
1
procedure in 79% yield over three steps. H NMR (300 MHz,
CD3OD) δ 7.42 (d, J = 7.8 Hz, 1H), 7.10-6.94 (m, 7H), 5.30 (d,
J = 9.0 Hz, 1H), 4.27 (s, 2H), 3.94 (dd, J = 10.8, 5.4 Hz, 1H), 3.76
(t, J = 9.0 Hz, 1H), 3.68-3.59 (m, 1H), 3.48 (t, J = 9.0 Hz, 1H),
3.44 (t, J = 10.8 Hz, 1H), 1.88-1.79 (m, 1H), 0.93-0.86 (m, 2H),
0.64-0.59 (m, 2H). 13C NMR (100 MHz, CD3OD) δ 142.7, 140.2,
139.9, 129.8, 127.4, 126.7, 126.6, 123.5, 121.8, 117.5, 110.5, 87.4,
79.2, 73.5, 71.1, 69.6, 33.2, 15.9, 9.5. ESMS m/z: 414 (MHþ), 436
(MNaþ). HRMS (EI) for C23H24ClNO4: calcd, 413.1394; found,
413.1390.
4-Chloro-3-(2-thienylmethyl)-1-(β-D-xylopyranosyl)-1H-indole
(19n). The title compound was obtained from 1-(2,3,4-tri-O-
acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j and
2-thienylmagnesium bromide according to the general procedure
in 35% yield over three steps. 1H NMR (400 MHz, CD3OD) δ
7.44 (dd, J = 8.4, 0.8 Hz, 1H), 7.20 (s, 1H), 7.15 (dd, J = 5.0, 1.2
Hz, 1H), 7.08 (t, J = 7.6 Hz, 1H), 7.01 (dd, J = 7.6, 1.2 Hz, 1H),
6.88 (dd, J = 5.0, 3.2 Hz, 1H), 6.80-6.78 (m, 1H), 5.33 (d, J =
8.8 Hz, 1H), 4.52, 4.49 (ABq, J = 16.8 Hz, 2H), 3.96 (dd, J =
11.1, 5.2 Hz, 1H), 3.80 (t, J = 8.8 Hz, 1H), 3.69-3.62 (m, 2H),
3.50 (t, J = 8.8 Hz, 1H), 3.46 (t, J = 11.1 Hz, 1H). 13C NMR (75
MHz, CD3OD) δ 146.5, 140.1, 127.7, 127.3, 126.8, 126.1, 125.8,
124.3, 123.6, 121.9, 116.7, 110.6, 87.4, 79.2, 73.6, 71.1, 69.6,
27.8. ESMS m/z: 380 (MHþ), 402 (MNaþ). HRMS (EI) for
C18H18ClNO4S: calcd, 379.0645; found, 379.0642.
4-Chloro-3-(6-methoxy-2-naphthylmethyl)-1-(β-D-xylopyrano-
syl)-1H-indole (19o). The title compound was obtained from
1-(2,3,4-tri-O-acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-
indole 13j and 6-methoxy-2-naphthylmagnesium bromide ac-
cording to the general procedure in 83% yield over three steps.
1H NMR (300 MHz, CD3OD) δ 7.67 (d, J = 8.4 Hz, 1H), 7.60
(d, J = 9.0 Hz, 1H), 7.54 (s, 1H), 7.45 (dd, J = 8.4, 1.2 Hz, 1H),
7.37 (dd, J = 8.4, 1.8 Hz, 1H), 7.18 (d, J = 2.4 Hz, 1H),
7.11-6.98 (m, 4H), 5.32 (d, J = 9.0 Hz, 1H), 4.46 (s, 2H), 3.94
(dd, J = 11.0, 4.8 Hz, 1H), 3.88 (s, 3H), 3.77 (t, J = 9.0 Hz, 1H),
3.66-3.58 (m, 1H), 3.48 (t, J = 9.0 Hz, 1H), 3.45 (t, J = 11.0 Hz,

176 Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1
Yao et al.
1H). 13C NMR (100 MHz, CD3OD) δ 158.8, 140.2, 138.2, 134.7,
130.7, 130.1, 129.3, 127.9, 127.8, 127.5, 126.9, 126.6, 123.9, 123.6,
123.6, 121.8, 119.5, 117.2, 110.6, 106.7, 87.4, 79.2, 73.5, 71.1,
69.6, 55.8, 33.6. ESMS m/z: 454 (MHþ), 476 (MNaþ). HRMS
(EI) for C25H24ClNO5: calcd, 453.1343; found, 453.1342.
3-(1,3-Benzodioxol-5-ylmethyl)-4-chloro-1-(β-D-xylopyranosyl)-
1H-indole (19p). The title compound was obtained from 1-(2,3,4-
tri-O-acetyl-β-D-xylopyranosyl)-4-chloro-3-formyl-1H-indole 13j
and 3,4-(methylenedioxy)phenylmagnesium bromide according
to the general procedure in 47% yield over three steps. 1H NMR
(300 MHz, CD3OD) δ 7.43 (dd, J = 8.2, 0.6 Hz, 1H), 7.10-6.98
(m, 3H), 6.71-6.65 (m, 3H), 5.87 (s, 2H), 5.31 (d, J = 9.0 Hz, 1H),
4.24 (s, 2H), 3.95 (dd, J = 11.1, 5.4 Hz, 1H), 3.79 (t, J = 9.0 Hz,
1H), 3.69-3.61 (m, 1H), 3.49 (t, J = 9.0 Hz, 1H), 3.45 (t, J = 11.1
Hz, 1H). 13C NMR (75 MHz, CD3OD) δ 149.1, 147.1, 140.2,
137.0, 127.4, 126.7, 126.4, 123.6, 122.7, 121.8, 117.3, 110.5, 110.3,
108.9, 102.1, 87.4, 79.2, 73.6, 71.1, 69.6, 33.2. ESMS m/z: 418
(MHþ), 440 (MNaþ). HRMS (FAB) for C21H20ClNO6: calcd,
417.0979; found, 417.0985.
4-Chloro-3-(2,3-dihydro-1,4-benzodioxin-6-ylmethyl)-1-(β-D-
xylopyranosyl)-1H-indole (19q). The title compound was ob-
tained from 1-(2,3,4-tri-O-acetyl-β-D-xylopyranosyl)-4-chloro-
3-formyl-1H-indole 13j and 3,4-(ethylenedioxy)phenylmagnesium
bromide according to the general procedure in 68% yield over
three steps. 1H NMR (300 MHz, CD3OD) δ 7.42 (dd, J = 8.1, 0.6
Hz, 1H), 7.09-6.98 (m, 3H), 6.72-6.66 (m, 3H), 5.31 (d, J = 9.0
Hz, 1H), 4.64 (s, 2H), 4.20-4.17 (m, 4H), 3.95 (dd, J = 10.8, 5.4
Hz, 1H), 3.79 (t, J = 9.0 Hz, 1H), 3.69-3.61 (m, 1H), 3.49 (t, J =
9.0 Hz, 1H), 3.45 (t, J = 10.8 Hz, 1H). 13C NMR (75 MHz,
CD3OD) δ 144.8, 143.2, 140.2, 136.2, 127.4, 126.7, 126.5, 123.5,
122.7, 121.8, 118.5, 117.9, 117.3, 110.5, 87.4, 79.2, 73.5, 71.1, 69.6,
65.7, 65.6, 32.8. ESMS m/z: 432 (MHþ), 454 (MNaþ). HRMS
(FAB) for C22H22ClNO6: calcd, 431.1136; found, 431.1139.
4-Chloro-3-(4-hydroxybenzyl)-1-(β-D-xylopyranosyl)-1H-indole
(20). A 1 M BBr3 solution in CH2Cl2 (5 mL) was added to a stirred
solution of 18c (267 mg, 0.50 mmol) in CH2Cl2 (16 mL) at -78 °C
under nitrogen. The mixture was warmed to 0 °C gradually. After
4 h, the reaction was quenched by the addition of saturated
NaHCO3 (aq) and the solvent was removed under reduced pres-
sure. The residue was redissolved in ethyl acetate and H2O. The
organic phase was separated, and the aqueous phase was extracted
with ethyl acetate twice. The combined organic layers were dried
over MgSO4, filtered, and concentrated under reduced pressure.
The residue was purified by column chromatography (EtOAc/
Hex = 1/3 to 1/2) to give 4-chloro-3-(4-hydroxybenzyl)-1-(2,3,4-
tri-O-acetyl-β-D-xylopyranosyl)-1H-indole (68 mg, 26%).
A 30% solution of NaOMe in MeOH (95 μL) was added to a
solution of 4-chloro-3-(4-hydroxybenzyl)-1-(2,3,4-tri-O-acetyl-
β-D-xylopyranosyl)-1H-indole (68 mg, 0.13 mmol) in MeOH/
CH2Cl2 (2/1, 1.5 mL) at 0 °C under nitrogen. The mixture was
warmed toroomtemperaturegradually andthenneutralizedwith
acidic resin after 2 h. The reaction mixture was filtered, and the
filtratewas concentrated under reducedpressure. The residuewas
purified by column chromatography (MeOH/CH2Cl2 = 1/15 f
1/10) to give the desired product 20 (29 mg, 56%) as a white solid.
1H NMR (300 MHz, CD3OD) δ 7.42 (dd, J = 8.1, 0.9 Hz, 1H),
7.09-6.95 (m, 5H), 6.71-6.67 (m, 2H), 5.30 (d, J = 9.0 Hz, 1H),
4.23 (s, 2H), 3.94 (dd, J = 11.0, 5.4 Hz, 1H), 3.76 (t, J = 9.0 Hz,
1H), 3.68-3.59 (m, 1H), 3.48 (t, J = 9.0 Hz, 1H), 3.45(t, J = 11.0
Hz, 1H). 13C NMR (75 MHz, CD3OD) δ 156.1, 139.9, 133.7,
130.8, 127.3, 126.5, 126.4, 123.5, 121.7, 117.9, 116.0, 110.4, 87.1,
78.9, 73.3, 70.9, 69.3, 32.7. ESMS m/z: 390 (MHþ), 412 (MNaþ).
HRMS (EI) for C20H20ClNO5: calcd, 389.1030; found, 389.1038.
4-Chloro-3-[4-(2-phenoxyethoxy)benzyl]-1-(β-D-xylopyranosyl)-
1H-indole (21a). Cs2CO3 (23 mg, 71 μmol) and 2-phenoxyethyl
bromide (22 mg, 107 μmol) were added to a stirred solution of 20
(14 mg, 35 μmol) in DMF (0.4 mL) at room temperature under
nitrogen. After 17 h, the reaction was quenched by the addition
of H2O and the resulting solution extracted with EtOAc. The orga-
nic phase was dried over MgSO4, filtered, and concentrated under
reduced pressure. The residue was purified by column chroma-
tography (MeOH/CH2Cl2=1/20) to afford the desired product
21a (13.3 mg, 73%) as a white solid. 1H NMR (400 MHz,
CD3OD) δ 7.43 (dd, J = 8.0, 0.8 Hz, 1H), 7.28-7.24 (m, 2H),
7.16-7.13 (m, 2H,), 7.07 (t, J = 8.0 Hz, 1H), 7.00-6.87 (m, 7H),
5.31 (d, J = 9.2 Hz, 1H), 4.27 (s, 6H), 3.94 (dd, J = 10.8, 5.2 Hz,
1H), 3.77 (t, J = 9.2 Hz, 1H), 3.67-3.61 (m, 1H), 3.49 (t, J = 9.2
Hz, 1H), 3.45 (t, J = 10.8 Hz, 1H). 13C NMR (75 MHz, CD3OD)
δ 160.3, 158.5, 140.2, 135.5, 130.9, 130.6, 127.4, 126.7, 126.5, 123.6,
122.1, 117.6, 115.7, 115.6, 110.5, 87.4, 79.2, 73.6, 71.1, 69.6, 68.0,
67.9, 32.8. ESMS m/z: 510 (MHþ), 532 (MNaþ). HRMS (FAB)
for C28H28ClNO6: calcd, 509.1605; found, 509.1603.
4-Chloro-3-[4-(prop-2-yn-1-yloxy)benzyl]-1-(β-D-xylopyranosyl)-
1H-indole (21b). K2CO3 (124 mg, 897 μmol) and propargyl bro-
mide (30 μL, 569 μmol) were added to a stirred solution of 20
(35 mg, 89 μmol) in DMF (0.9 mL) at room temperature under
nitrogen. The mixture was heated to 50 °C and stirred for 21 h. The
reaction was quenched by the addition of H2O, and the resulting
solution was extracted with EtOAc. The organic phase was dried
over MgSO4, filtered, and concentrated under reduced pressure.
The residue was purified by column chromatography (MeOH/
CH2Cl2 = 1/20) to give the desired product 21b (26.8 mg, 70%) as a
white solid. 1H NMR (300 MHz, CD3OD) δ 7.43 (dd, J = 8.1, 0.9
Hz, 1H), 7.17-7.12 (m, 2H), 7.07 (t, J = 8.1 Hz, 1H), 7.01-6.98
(m, 2H), 6.90-6.85 (m, 2H), 5.30 (d, J = 9.0 Hz, 1H), 4.66 (d, J =
2.1 Hz, 2H), 4.72 (s, 2H), 3.99 (dd, J = 11.1, 5.4 Hz, 1H), 3.78 (t,
J = 9.0 Hz, 1H), 3.68-3.60 (m, 1H), 3.49 (t, J = 9.0 Hz, 1H), 3.45
(t, J = 11.1 Hz, 1H), 2.89 (t, J = 2.1 Hz, 1H). 13C NMR (75 MHz,
CD3OD) δ 157.4, 140.2, 135.9, 130.8, 127.4, 126.7, 126.5, 123.6,
121.8, 117.5, 115.8, 110.5, 87.4, 80.2, 79.2, 76.6, 73.5, 71.1, 69.6,
56.8, 32.7. ESMS m/z: 428 (MHþ). HPLC purity 85.6%.
4-Chloro-3-{4-[2-(2-ethoxyethoxy)ethoxy]benzyl}-1-(β-D-xylo-
pyranosyl)-1H-indole (21c). MsCl (0.86 mL, 10.96 mmol) and tri-
ethylamine (3.0 mL, 21.91 mmol) were added to a stirred solution
of diethylene glycol monoethyl ether (0.98 g, 7.30 mmol) in
CH2Cl2 (30 mL) at 0 °C under nitrogen. The mixture was slowly
warmed to room temperature. After 3 days, the mixture was dilu-
ted with CH2Cl2 and concentrated under reduced pressure. The
residue was redissolved in Et2O and washed with 1 N HCl (aq).
The organic phase was dried over MgSO4, filtered, and concen-
trated under reduced pressure. The residue (885 mg, 57%) was
used for alkylation directly without further purification.
To a stirred solution of the aforementioned mesylate residue
(70 mg, 329 μmol) and 20 (17 mg, 43 μmol) in DMF (0.4 mL) at
room temperature under nitrogen was added Cs2CO3 (42 mg,
130 μmol). After 2 days, the reaction was quenched by the addi-
tionofH2O, and the resulting solutionwas extracted withEtOAc.
The organic phase was dried over MgSO4, filtered, and concen-
tratedunderreducedpressure. Theresidue waspurifiedbycolumn
chromatography (MeOH/CH2Cl2 = 1/20 to 1/10) to provide the
1
desired product 21a (15.1 mg, 68%) as a white solid. H NMR
(300 MHz, CD3OD) δ 7.42 (dd, J = 8.2, 0.9 Hz, 1H), 7.14-7.04
(m, 3H), 7.00-6.97 (m, 2H), 6.84-6.81 (m, 2H), 5.30 (d, J = 9.0
Hz, 1H), 4.26 (s, 2H), 4.09-4.05 (m, 2H), 3.94 (dd, J = 11.0, 5.1
Hz, 1H), 3.81-3.77 (m, 3H), 3.68-3.55 (m, 5H), 3.52-3.41 (m,
4H), 1.16 (t, J = 7.2 Hz, 3H). 13C NMR (100 MHz, CD3OD) δ
158.5, 140.2, 135.3, 130.9, 127.4, 126.7, 126.5, 123.5, 121.8, 117.6,
115.5, 110.5, 87.4, 79.2, 73.6, 71.8, 71.2, 71.1, 71.0, 69.6,
68.6, 67.7, 32.7, 15.5. ESMS m/z: 528 (MNaþ). HRMS (EI)
for C26H32ClNO7: calcd, 505.1867; found, 505.1187. HPLC
purity 89.5%.
4-Bromo-3-(4-cyclopropylbenzyl)-1-(β-D-xylopyranosyl)-1H-
indole (22). The title compound was obtained from 1-(2,3,4-tri-
O-acetyl-β-D-xylopyranosyl)-4-bromo-3-formyl-1H-indole 13n
and 4-cyclopropylphenylmagnesium bromide according to the
general procedure for the preparation of 19m in 58% yield over
three steps. 1H NMR (400 MHz, CD3OD) δ 7.48 (d, J = 8.0 Hz,
1H), 7.19 (dd, J =8.0, 0.4Hz, 1H), 7.09-7.02(m, 2H), 7.02-6.95
(m, 4H), 5.30 (d, J = 8.8 Hz, 1H), 4.32, 4.29 (ABq, J = 16.4 Hz,
2H), 3.94 (dd, J=11.2, 5.2 Hz, 1H), 3.75 (t, J = 8.8 Hz, 1H),

Article
Journal of Medicinal Chemistry, 2011, Vol. 54, No. 1 177
3.66-3.60 (m, 1H), 3.48 (t, J = 8.8 Hz, 1H), 3.44 (t, J = 11.2 Hz,
1H), 1.88-1.82 (m, 1H), 0.93-0.88 (m, 2H), 0.64-0.60 (m, 2H).
13C NMR (100 MHz, CD3OD) δ142.7, 139.9, 139.7, 129.9, 127.8,
127.0, 126.5, 125.3, 123.8, 117.9, 115.2, 111.0, 87.3, 79.1, 73.5,
71.0, 69.5, 33.1, 15.9, 9.59. ESMS m/z: 458 (MHþ). HRMS (EI)
for C23H24BrNO4: calcd, 457.0889; found, 457.0898.
a 24 h urine collection, and the experimental animals were
refed at 1 h after glucose challenge. After the urine volumes
were measured and recorded, the urine samples were measured
for glucose levels by using a DRI-CHEM 3500s (FUJI, Tokyo,
Japan). A P value of less than 0.05 was considered statistically
significant.
In Vitro Human SGLT Uptake Assays. Stably transfected
CHO-K1cells wereused for transporterstudies. SGLTwas deter-
mined by uptake of [14C]R-methyl-D-glucopyranoside ([14C]AMG,
specific radioactivity of 310 mCi/mmol) purchased from Perkin-
Elmer (Boston, MA). For the purpose of this study, Krebs-
Ringer-Henseleit (KRH) solution containing 120 mM NaCl,
4.7 mM KCl, 1.2 mM MgCl2, 2.2 mM CaCl2, and 10 mM Hepes
(pH 7.4 with Tris) was used for SGLT uptake assays. All chemi-
cals were purchased from Sigma (Deisenhofen, Germany). Briefly,
hSGLT2/CHO-K1 cells or hSGLT1/CHO-K1 cells were seeded
into a white-walled 96-well culture plate (Corning, NY) at a density
of 30 000 cells/well (hSGLT2) or 20 000 cells/well (hSGLT1) and
incubated for 48 h at 37 °C in a 5% CO2 atmosphere in growth
medium. After 48 h, the culture medium in the wells was removed
and wells were washed three times with 280 μL of KRH solution
and then incubated in KRH solution containing 3 μM [14C]AMG
in the absence or presence of inhibitors for up to 120 min at 37 °C.
At the end of the uptake period, the KRH solution was removed
and the uptake of [14C]AMG was stopped by adding ice-cold KRH
solution (stop solution). The wells were rinsed three times with
150 μL of stop buffer using the microplate washer (TEcan,
Blood Glucose Lowering Effects in the Streptozotocin-Induced
Diabetes Rats.20 Adult male Sprague-Dawley rats (BioLasco,
Ilan, Taiwan) received three intraperitoneal injections of strep-
tozotocin from Sigma (catalog no. S0130, St. Louis, MO) at
65 mg/kg freshly prepared in 0.01 M citrate buffer at one injec-
tion every other day. The animals were then monitored by using
blood glucometer (ACCU-CHEK from Roche, Basel, Switzerland),
for the glucose levels in blood collected via stabbing the tail vein with
25G needle once a week. A streptozotocin-induced diabetes was
confirmed when the blood glucose levels were up more than
450 mg/dL. The diabetic rats were divided into experimental
groups at three rats each and orally administered a single dose of
19m, 5, or vehicle as control. Blood samples were obtained from
the tail vein at 0 (predose), 0.5, 1, 2, 3, 4, and 5 h after the oral
administration and measured for blood glucose levels with the
glucometer. A P value of less than 0.05 was considered statis-
tically significant.
Diarrheogenic Activity Test.31 Male CD-1 (ICR) mice at the
age of 5-6 weeks purchased from BioLASCO (Ilan, Taiwan)
were used in this study. The mice were administrated orally with
19m at doses of 200, 50, 10 mg/kg and vehicle (5% DMSO, 20%
Cremophor EL, and 75% ddH2O). After dosing, all animals were
housed in the metabolism cages (SHINANO, SN-783, Tokyo,
Japan) individually and given diet and water ad libitum during
the experiment. The mice were monitored once hourly within 8 h
after dosing. The bottom of each cage was covered with white
paper for convenient observation of fecal state.
€
Mannedorf, Switzerland). After the third rinse, the stop solution
was completely removed from the wells and the cells were
solubilized by adding 0.1% sodium dodecyl sulfate (Sigma).
After 24 h, the microtiter plate was taken for scintillation
counting of radioactive [14C]AMG using a TopCount (Perkin-
Elmer). The percent of inhibition of inhibitors was calculated by
comparing counts per minute (CPM) in inhibitor-containing
well with CPM in wells containing only DMSO vehicle. Phlor-
izin and dapagliflozin were evaluated in parallel in every assay.
A dose-response curve was fitted to a sigmoidal dose-response
model using GraphPad software to determine the inhibitor
concentration at half-maximal response (EC50).
Acknowledgment. Financial support from National Health
Research Institutes and National Science Council of the Re-
public of China (Grant NSC98-2323-B-400-002) is gratefully
acknowledged.
In Vivo Pharmacokinetics Evaluation of 19m in Rats. The
animal study was approved by Institutional Animal Care and
Use Committee of National Health Research Institutes. A solu-
tion of test compound (1 mg/mL) was prepared by dissolving the
appropriate amount of compound in a mixture of PEG 400/
DMA (20:80, v/v). Male Sprague-Dawley rats, weighing 250-
350 g each (8-10 weeks old), were obtained from BioLASCO,
Ilan, Taiwan. 19m was administered to three male rats each
intravenously by a bolus injection to the jugular vein or orally
at 1 mg/kg dose. At 0 (prior to dosing), 2, 5 (iv only), 15, and
30 min and at 1, 2, 4, 6, 8, and 24 h after dosing, a blood sample
(∼150 μL) was collected from each animal via the jugular-vein
cannula and stored inice (0-4 °C). Plasma was separatedfromthe
blood by centrifugation (14000g for 15 min at 4 °C in a Beckman
model AllegraTM 6R centrifuge) and stored in a freezer (-60 °C).
All samples were analyzed for the test compound by LC-MS/MS
(ABI 4000Q Trap). Data were acquired via multiple reactions
monitoring. Plasma concentration data were analyzed with stan-
dard noncompartmental method with Kinetica (version 4.1.1,
InnaPhase Corporation, PA).
Supporting Information Available: Details of synthetic pro-
cedures for compound 19m and HPLC purity determination
conditions. This material is available free of charge via the
Internet at http://pubs.acs.org.
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