Synthesis, enzyme kinetics and computational evaluation of N-(β-d-glucopyranosyl) oxadiazolecarboxamides as glycogen phosphorylase inhibitors

Article abstract of DOI:10.1016/j.bmc.2013.07.024

All possible isomers of N-β-d-glucopyranosyl aryl-substituted oxadiazolecarboxamides were synthesised. O-Peracetylated N-cyanocarbonyl-β- d-glucopyranosylamine was transformed into the corresponding N-glucosyl tetrazole-5-carboxamide, which upon acylation

Full text of DOI:10.1016/j.bmc.2013.07.024

Bioorganic & Medicinal Chemistry 21 (2013) 5738–5747
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis, enzyme kinetics and computational evaluation
of N-(b-D-glucopyranosyl) oxadiazolecarboxamides
as glycogen phosphorylase inhibitors
b
b
Mária Polyák a, Gergely Varga a, Bence Szilágyi a, László Juhász a, , Tibor Docsa , Pál Gergely ,
⇑
Jaida Begum c, Joseph M. Hayes c, László Somsák a,
⇑
a Department of Organic Chemistry, University of Debrecen, PO Box 20, H-4010 Debrecen, Hungary
b Department of Medical Chemistry, Medical and Health Science Centre, University of Debrecen, Egyetem tér 1, H-4032 Debrecen, Hungary
c Centre for Materials Science, Division of Chemistry, University of Central Lancashire, Preston PR1 2HE, United Kingdom
a r t i c l e i n f o
a b s t r a c t
Article history:
All possible isomers of N-b-
D-glucopyranosyl aryl-substituted oxadiazolecarboxamides were synthesised.
Received 6 May 2013
Revised 8 July 2013
Accepted 10 July 2013
Available online 25 July 2013
O-Peracetylated N-cyanocarbonyl-b-
D
-glucopyranosylamine was transformed into the corresponding
N-glucosyl tetrazole-5-carboxamide, which upon acylation gave N-glucosyl 5-aryl-1,3,4-oxadiazole-2-
carboxamides. The nitrile group of the N-cyanocarbonyl derivative was converted to amidoxime which
was ring closed by acylation to N-glucosyl 5-aryl-1,2,4-oxadiazole-3-carboxamides. A one-pot reaction
of protected b-D-glucopyranosylamine with oxalyl chloride and then with arenecarboxamidoximes fur-
nished N-glucosyl 3-aryl-1,2,4-oxadiazole-5-carboxamides. Removal of the O-acetyl protecting groups
by the Zemplén method produced test compounds which were evaluated as inhibitors of glycogen phos-
Keywords:
1,2,4-Oxadiazole
1,3,4-Oxadiazole
b-d-Glucopyranosyl derivatives
Glycogen phosphorylase
Inhibitor
phorylase. Best inhibitors of these series were N-(b-
3-carboxamide (Ki = 30 M), N-(b- -glucopyranosyl) 5-(naphth-2-yl)-1,3,4-oxadiazol-2-carboxamide
(Ki = 33 M), and N-(b- -glucopyranosyl) 3-phenyl-1,2,4-oxadiazol-5-carboxamide (Ki = 104 M).
D-glucopyranosyl) 5-(naphth-1-yl)-1,2,4-oxadiazol-
l
D
l
D
l
ADMET
ADMET property predictions revealed these compounds to have promising oral drug-like properties with-
out any toxicity.
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
based inhibitors targeting the catalytic site are the most exten-
sively investigated derivatives14–17 and a glucopyranosylidene-
Glycogen phosphorylase (GP), the main regulatory enzyme of
the glycogen metabolism pathway, is a validated target to control
hepatic glucose output in non-insulin dependent or type 2 diabetes
mellitus (T2DM).1–3 T2DM is a major concern to public health4,5
with several long term complications6 such as cardiovascular dis-
ease, neuropathy, retinopathy and nephropathy. Biochemical and
pharmacological aspects of T2DM have been amply reviewed and
for a detailed rationalization of the possible use of GP inhibitors
(GPIs) as antidiabetics, the reader is kindly referred to those arti-
cles.1–3 Besides T2DM, inhibition of GP has also been studied in
connection with diseases caused by abnormalities in glycogen
metabolism,7,8 such as myocardial ischemia9, cerebral ischemia10
and tumors.11,12
spiro-thiohydantoin has been shown to have appreciable in vivo
hypoglycaemic effects.18
N-Acyl-b-
and N-acyl-N0-b-
D
-glucopyranosylamines19–21 (Chart 1, I), N-aryl-13
D
-glucopyranosyl ureas (II)13,22,23 are among the
best glucose analogue inhibitors of GP discovered to date, which
are efficient in or below the low micromolar range.
Bioisosteric replacement is widely used in medicinal chemistry
to design new drug molecules by systematic modification of lead
compounds.29 Nonclassical bioisosteric replacements of the NHCO
moiety in I by heterocyclic linkers A–D (Chart 1) resulted in inhib-
itors of varying efficiency.24–27 While 1,2,3-triazoles IA proved
equipotent with the amides I, among oxadiazoles IB–D the consti-
tution of the ring was decisive for the effect, and only ID was of
similar efficiency as I. In these compounds the presence of a large
hydrophobic aromatic ring was very advantageous for the inhibi-
tion, and derivatives with a 2-naphthyl group were the best inhib-
itors in each series. Replacements of the ‘second’ amide moiety in II
(highlighted in Chart 1) with heterocycles E and F were detrimen-
tal for the binding, but revealed that compounds with a phenyl
Several types of compounds have been shown to inhibit this
enzyme under in vitro conditions.13 Among them the glucose
⇑
Corresponding authors. Tel.: +36 52512900x22474; fax: +36 52512744 (L.J.);
tel.: +36 52512900x22348 (L.S.).
E-mail addresses: juhasz.laszlo@science.unideb.hu, dr.juhasz.m.laszlo@gmail.
com (L. Juhász), somsak.laszlo@science.unideb.hu (L. Somsák).
0968-0896/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2013.07.024

M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
5739
linkers to replace the highlighted NHCO moieties
N
N
N O
O N
N
N
N
O
N
N
A
B
C
D
linker
OH
H
N
O
HO
HO
R
1624
3625
10%
3827
1226
HO
at 625 µM26
O
I 1020
O
N
N N
N
E
F
linker
OH
H
N
H
N
O
HO
HO
No inh.
No inh.
R
HO
at 625 µM28 at 625 µM28
O
O
target compounds of this study:
IIB-D
17228
7528
(R = Ph)
(R = Ph)
II 0.3513
Chart 1. Selected inhibitors of GP and their efficiency against rabbit muscle GPb (Ki [lM] for R = 2-naphthyl).
OPG
route A
amide
OPG
O
H
N
O
formation
PGO
PGO
PGO
PGO
X
linker B/C/D
R
route A
OPG
X: NH2, N3
OPG
O
route B
+
route B
heterocycle formation
linker B/C/D
Y
R
OPG
O
O
H
Y: OH, Cl
PGO
PGO
N
Z
OPG
O
Z: group for heterocyclization (CN, C(NH2)NOH, COCl)
Scheme 1. Retrosynthetic analysis of the target compounds.
group could bind stronger than those with
substituent.28
a
2-naphthyl
an oxadiazolecarboxylic acid or acid chloride, or equivalently, by
acylation of glucopyranosylamine (route A), or heterocyclisation
of suitable functional groups of the corresponding N-acylated
glucopyranosylamine derivatives (route B).
As a continuation of our systematic structure–activity relation-
ship studies25,26,28,30 on bioisosteric replacements of NHCO moie-
ties in inhibitors
I
and II, we have now synthesised new
To study synthetic routes A, preparation of the necessary
oxadiazolecarboxylic acids was first attempted. Following a litera-
ture protocol, commercially available benzhydrazide (1) was
ethoxalylated to give 2 (Scheme 2) which was closed to ethyl
5-phenyl-1,3,4-oxadiazole-2-carboxylate (3) in 47% yield.31 Hydro-
lysis of 3 gave carboxylic acid 4 in 70% yield.32 Synthesis of 1,2,4-
oxadiazole-carboxylic acids was also tried from the corresponding
ethyl esters,33 however, we were unable to reproduce the reported
hydrolytic step34 because of opening of the heterocycle.
oxadiazolecarboxamide derivatives IIB–D and evaluated through
kinetic experiments their potential for GP inhibition. In addition,
prediction of absorption, distribution, metabolism, excretion and
toxicity (ADMET) properties was also performed to evaluate the
drug-like potential of these derivatives.
2. Results and discussion
2.1. Syntheses
Acylation of the glycosylimino-trimethylphosphorane obtained
from 2,3,4,6-penta-O-acetyl-b-D-glucopyranosyl azide (6) with car-
boxylic acid 4 failed, however, oxadiazolecarboxamide derivative
10a could be prepared in 10% yield by using acid chloride 5.
In view of the low yield of this transformation and the failure
of getting other oxadiazolecarboxylic acids, we turned to the
Preparation of the target compounds was envisaged and inves-
tigated by two possible routes (Scheme 1): formation of the amide
C–N bond in a direct transformation of protected glucopyranosyl
azide via acylation of the in situ generated iminophosphorane with

5740
M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
O
O
N
N
H
N
i
ii
PhCONHNH2
Ph
COOR
iv
EtO
N
H
Ph
O
O
2
3 R = Et
4 R = H
5 R = Cl
1
iii
O
N
O
TsO
O
OAc
O
OAc
O
OAc
H
N
OAc
O
vi
O
AcO
AcO
AcO
AcO
AcO
v
CN
NH2
N3
OAc
AcO
OAc
O
6
7
8
vii
N
NH
N
OR
N
OAc
O
N
H
N
viii
H
a:
b:
c:
O
Ar = phenyl
Ar
RO
RO
AcO
AcO
N
Ar = 2-naphthyl
Ar = 1-naphthyl
O
N
OR
OAc
O
O
9
10a - c: R = Ac
11a - c:
ix
R = H
Scheme 2. (i) EtOCOCOCl, 3 equiv Et3N, CH2Cl2, 0 °C to rt and in the same pot (ii) TsCl, rt, overnight; (iii) LiOH in THF/H2O = 1:1; (iv) SOCl2, reflux; (v) PMe3; CH2Cl2; (vi) dry
toluene, 50 °C; (vii) Me3SiN3, Bu2SnO; dry toluene 80 °C; (viii) ArCOCl, dry toluene, reflux; (ix) NaOMe (cat.), MeOH, rt.
at elevated temperature37 in moderate to good yields (10a: 80%;
10b: 74%, 10c: 30%). O-Deacetylations were performed by the
Zemplén protocol to give excellent yields (up to 87%) of unpro-
tected 1,3,4-oxadiazole derivatives 11a–c.
OAc
O
OAc
O
NH2
N
H
N
H
N
i
ii
AcO
AcO
AcO
AcO
CN
OH
OAc
OAc
O
O
8
Next, we turned to the synthesis of N-glucopyranosyl 5-aryl-
1,2,4-oxadiazole-3-carboxamides 15a–c (Scheme 3). In a continu-
ous operation, N-cyanocarbonyl derivative 8 was transformed into
amidoxime 12 by NH2OH, followed by acylation with aroyl
chlorides and ring closure in the presence TBAF to the desired
1,2,4-oxadiazole derivatives 14a–c in moderate overall yields
(22–24%). Synthesis of 14a was also carried out via the isolated
but unpurified 12 and fully characterized 13a in a two steps proce-
dure, but the overall yield was similar (28% as compared to 24% by
the one-pot reaction). O-Deacetylations by the Zemplén protocol
gave the unprotected 5-aryl-1,2,4-oxadiazole derivatives 15a–c in
acceptable yields (49–64%).
Synthesis of N-glucopyranosyl 3-aryl-1,2,4-oxadiazole-5-car-
boxamides 20 was attempted by cycloaddition of nitrile-oxides
to the CN group of 8, however, this resulted in an unseparable mul-
ticomponent product mixture. To get 20 in a less direct but one-pot
procedure, acylation of glucopyranosylamine 7 was achieved with
(COCl)2 in dry THF (Scheme 4) to give the N-substituted oxamidoyl
chloride 16. Next, freshly prepared arenecarboxamidoximes 18a–
c,38–40 obtained from the corresponding nitriles 17a–c, were
acylated by 16 to furnish 19a–c which underwent immediate ring
closure to the desired 3-aryl-1,2,4-oxadiazole-5-carboxamides
20a–c in low to acceptable yields (15–55%). O-Deacetylations were
12
O
OR
N
OAc
NH2
N
H
H
N
O
Ar
O
RO
RO
N
AcO
AcO
iii
O
Ar
N
OR
OAc
O
O
O
13a-c
14a - c: R = Ac
15a - c: R = H
iv
a: Ar = phenyl
b: Ar = 2-naphthyl
c: Ar = 1-naphthyl
Scheme 3. (i) NH2OHꢀHCl; dry pyridine; 50 °C; (ii) 1. ArCOCl, dry pyridine–toluene,
110 °C; (iii) TBAF in THF-dry toluene, 110 °C; (iv) NaOMe (cat.), dry MeOH.
preparation of our target compounds by route B (Scheme 1). N-
Cyanocarbonyl-2,3,4,6-penta-O-acetyl-b-D-glucopyranosylamine
(8, Scheme 2) was the key intermediate of this pathway, which was
prepared from glucosylamine 7 in 57% yield by using Renslo’s
method.35 Transformation of 8 into N-glucopyranosyl tetrazol-5-
carboxamide 9 was achieved with Me3SiN3–Bu2SnO36 in 88% yield.
The necessary 5-aryl-1,3,4-oxadiazole derivatives 10a–c were ob-
tained from 9 by the corresponding aroyl chloride in dry toluene
Ar
NH2
OAc
O
OAc
O
OAc
O
O
O
H
N
H
N
ii
i
AcO
AcO
AcO
AcO
AcO
AcO
N
NH2
O
Cl
OAc
OAc
OAc
O
O
19a - c
7
16
NH2
N
N
OR
O
iv
H
N
O
OH
Ar
RO
RO
Ar CN
17a - c
Ar
N
OR
18a - c
O
a: Ar = phenyl
20a - c:
R = Ac
21a - c: R = H
iii
b: Ar = 2-naphthyl
c: Ar = 1-naphthyl
Scheme 4. (i) (COCl)2 in dry THF at 0 °C; (ii) amidoximes 18a–c, THF, rt; (iii) NaOMe in dry MeOH; (iv) NH2OHꢀHCl, dry pyridine, 50 °C.

M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
5741
performed by the Zemplén protocol to give good (62–79%) yields of
2.3. ADMET properties calculations
unprotected 3-aryl-1,2,4-oxadiazole derivatives 21a–c.
Unfavourable pharmacokinetic properties can in many cases
lead to the clinical trials failure of otherwise potentially successful
drug candidates. Their evaluation, therefore, at an earlier stage is
desired.43,44 Accordingly, we have predicted ADMET properties of
our inhibitors using the QikProp 3.5 program (Schrodinger, LLC)
which estimates both physically significant descriptors and phar-
maceutically relevant properties. Considering the reported accu-
racy of ALOGPS45 in comparison with other programs,46 log S
(aqueous solubilities) and log P(o/w) (partition of ligands in an oct-
anol/water mixture) values from ALOGPS 2.1 are also reported.
Meanwhile, toxicity is the leading cause of drug attrition in clinical
trials, together with lack of efficacy.47 Therefore, toxicity structural
warnings for our inhibitors were also probed using the FAF-Drugs2
server.48
The results of our calculations are given in Table 2. Property
predictions that are outside the range observed for 95% of known
drugs (QikProp, version 3.5, User Manual) are flagged with an
asterisk (⁄), while violations of Lipinski’s ‘rule of five’49 and Jorgen-
sen’s ‘rule of three’50,51 (QikProp 3.5 User Manual) for oral bioavail-
ability are highlighted in italics. The property values should be
treated as approximate but serve as a useful guideline for future li-
gand studies, of particular relevance here.
As a first test of the drug-likeness of the ligands, we applied
Lipinski’s ‘rule of five’ requiring candidates to have no more than
5 and 10 hydrogen bond donors (HBDs) and acceptors (HBAs),
respectively, molecular weights (MW) less than 500 amu, and log
P(o/w) values less than 5. An orally active compound/drug should
have no more than one violation of these rules. No violations of
these rules was observed for any of our inhibitors. Furthermore,
the properties were within the range of values for 95% of known
drugs. Satisfactory agreement between QikProp and ‘consensus’
ALOGPS45 logP(o/w) values was obtained, with a root-mean-square
deviation (RMSD; Eq. (1)) of 1.1 and a maximum absolute differ-
ence of 1.3 units.
2.2. Enzyme inhibition studies
The kinetic parameters of the deprotected compounds (inhibi-
tion constants (Ki) against rabbit muscle glycogen phosphorylase
b (RMGPb)) were determined according to the protocol described
earlier.41 The results are summarized in Table 1, together with
the Ki-s of relevant reference compounds.
The new N-glucopyranosyl oxadiazole-carboxamides showed
inhibitory properties in a very broad range from inactive com-
pounds to low micromolar inhibitors. The most remarkable obser-
vation was that the best compounds in the different series did not
have the same aromatic moiety as it could have been expected
from previous experiences. Thus, from 1,3,4-oxadiazoles 11, from
5-aryl-1,2,4-oxadiazoles 15, and from 3-aryl-1,2,4-oxadiazoles 21,
the 2-naphthyl 11b, the 1-naphthyl 15c, and the phenyl 21a deriv-
atives, respectively, were the best inhibitors. Since the size of the
heterocycles must be very similar, this phenomenon might be
due to variations of interactions between the oxadiazole rings
and the enzyme as well as to the probably different orientations
of the aromatic substituents. It is also worth noting that in two ser-
ies the 2-naphthyl derivatives 15b and 21b were inactive, although
this was similar to the cases of isoxazoles 22 and 1,2,3-triazoles 23.
In comparison to the homoaromatic N-glucopyranosyl arenecarb-
oxamides 24 the inhibition of the oxadiazolecarboxamides was
generally weaker for the phenyl (a) and the 2-naphthyl (b) deriva-
tives, while stronger for the 1-naphthyl (c) compounds. This may
be attributed to the different size of the molecules and the orienta-
tion of the aromatic rings. Finally, a comparison to the ‘parent’
molecules 25 used as lead for the bioisosteric replacement showed
a significant loss of the activity with each of the aromatic substit-
uents. Molecular dockings to get a better insight in the structural
details of the binding peculiarities of these and other heterocyclic
N-glucopyranosyl carboxamides as well as to predict more efficient
structures are in progress.
Jorgensen’s ‘rule of three’ considers a Caco-2 cell permeability
53
value >22 nm sꢁ1 (used as a model for gut–blood barrier),
a
Table 1
log S value >ꢁ5.7 and number of primary metabolites (NPM) <7
to be characteristic of potential drugs with better oral bioavailabil-
ity, more ‘drug-like’ molecules having fewer violations. The log S
and NPM criteria are satisfied for all ligands. The RMSD (Eq. (1)) be-
tween QikProp and ALOGPS log S values was 0.6, with a maximum
absolute difference of 0.7 units. The Caco-2 cell permeabilities (18–
22 nm sꢁ1), however, are generally borderline the Jorgensen limit
and outside the desirable range of 95% of known drugs
(>25 nm sꢁ1). This is consistent with inhibitor polar surface areas
(PSAs; ꢂ170 Å2) exceeding Veber et al.,52 suggested limit (PSA
<140 Å2), but contrary to the PSAs lying within the range for 95%
of known drugs (7–200 Å2). It is clear, however, that the sensitive
balance between adequate lipophilicity and solubility properties
will need attention in ‘lead optimization’ of any heterocylic deriv-
ative conjugated to glucose found to have attractive GP inhibitory
Inhibition of rabbit muscle glycogen phosphorylase
compounds and selected glucose derivatives Ki (lM)
b
(RMGPb) by the new
OH
H
N
O
HO
HO
linker Ar
Ar
HO
O
Linker
a
b
c
N
N
11 545a
15 136a
21 104
30
172a
O
N O
No inhibitionb 33
N
O
N
No inhibitionb 145a
N
potential (low lM activity or better).
O N
No
22 17228
—
The degree of plasma protein binding also affects the amount
of bioavailable drug. LogKhsa is the prediction of the degree of
binding to human serum albumin (hsa) and is satisfactory for
all ligands (ꢂ1), within the range for 95% of known drugs
(ꢁ1.5–1.5). Likewise, the predicted blood–brain barrier co-effi-
cients (logBB values: ꢁ3.0 to –2.7) are within the desirable lim-
its (ꢁ3.0–1.2). Finally, a complete lack of toxicity structural
warnings from FAF-Drugs2 is encouraging with respect to the
toxicity profiles for conjugates of glucose and substituted
heterocycles.48
inhibitionb28
N
N
N
No
23 7528
24 8120
—
inhibitionb28
none
1020
44420
H
N
25 4.613
0.3513
1513
O
a
Calculated from the IC50 values by the Cheng–Prusoff equation: Ki = IC50
/
(1 + [S]/Km).42
b
No inhibition at 625 lM.

5742
M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
Table 2
Results of ADMET property predictions for the inhibitors studied in this worka
d
Inhibitor
Lipinskis’s rule of five and violations (V)b
V
Jorgensen’s rule of three and violations (V)b
V
PSAc (Å2)
LogKhsa
LogBBe
TSWf
Mr (Da)
(<500)
351.3
HBDg
(65)
5
HBAh
(610)
10
LogP (o/w)i
Caco-2j (nm sꢁ1
)
LogSi
NMPk
(<7)
6
(<5)
(>22)
(>ꢁ5.7)
(<140 Å2)
11a
11b
11c
ꢁ1.4
0
0
0
0
0
0
0
0
0
—
22.3⁄
ꢁ2.6
0
0
1
1
1
1
1
1
1
—
170.7
ꢁ1.04
ꢁ0.85
ꢁ0.85
ꢁ1.10
ꢁ0.91
ꢁ0.92
ꢁ1.10
ꢁ0.91
ꢁ0.92
ꢁ2.7
ꢁ2.8
ꢁ2.8
ꢁ2.8
ꢁ2.9
ꢁ2.8
ꢁ2.8
ꢁ3.0
ꢁ2.9
ꢁ3.0–1.2
—
—
—
—
—
—
—
—
—
—
(ꢁ0.8 0.7)
ꢁ0.6
(ꢁ2.0)
ꢁ3.4
401.4
401.4
351.3
401.4
401.4
351.3
401.4
401.4
130–725
5
10
22.3⁄
21.8⁄
18.5⁄
21.1⁄
21.9⁄
19.4⁄
18.1⁄
20.0⁄
6
170.7
171.2
171.9
170.4
170.5
171.3
172.4
171.4
7–200
(0.3 0.7)
ꢁ0.7
(ꢁ2.7)
ꢁ3.3
5
10
6
(0.3 0.7)
ꢁ1.6
(ꢁ2.7)
ꢁ2.6
15a
15b
15c
5
10
5
(ꢁ0.7 0.5)
ꢁ0.8
(ꢁ2.0)
ꢁ3.3
5
10
5
(0.4 0.5)
ꢁ0.9
(ꢁ2.7)
ꢁ3.3
5
10
5
(0.4 0.5)
ꢁ1.6
(ꢁ2.7)
ꢁ2.6
21a
21b
21c
5
10
5
(ꢁ0.7 0.5)
ꢁ0.9
(ꢁ2.0)
ꢁ3.4
5
10
5
(0.4 0.5)
ꢁ0.9
(ꢁ2.7)
ꢁ3.3
5
10
5
(0.4 0.5)
ꢁ2.0–6.5
(ꢁ2.6)
Rangel
0–6
2–20
<25 poor; >500 great
ꢁ6.5–0.5
1–8
ꢁ1.5–1.5
a
b
c
d
e
f
ADMET data were calculated as described in the text using Qikprop 3.5; predicted properties outside the range for 95% of known drugs are indicated with an asterisk (⁄).
Rules as listed in the columns, with any violations of the rules highlighted in italics.
52
PSA represents the van der Waals (polar) surface areas of N and O atoms with a recommended PSA <140 Å2
LogKhsa: predicted binding to human serum albumin.
LogBB: the predicted blood–brain barrier co-efficient.
Toxicity structural warnings from FAF-Drugs2.
.
g
h
i
Number of hydrogen bond donors.
Number of hydrogen bond acceptors.
Values calculated with ALOGPS are given in parentheses (a ‘consensus’ value standard deviation in the case of logP(o/w)).
Caco-2 cell permeability.
Number of primary metabolites.
j
k
l
Range for 95% of known drugs; reference: QikProp version 3.5 User’s Manual.
plates were visualized under UV light, and by gentle heating. For
column chromatography Kieselgel 60 (Merck, particle size
(0.063–0.200 mm) was applied.
3. Conclusions
Synthetic procedures were elaborated for all possible isomers of
N-b-D-glucopyranosyl aryl-substituted-oxadiazolecarboxamides.
4.2. 5-Phenyl-1,3,4-oxadiazole-2-carboxylic acid (4)
The compounds with phenyl, 1- and 2-naphthyl substituents were
assayed against rabbit muscle GPb to show low micromolar
efficiency for the best inhibitors. Both the constitution of the oxa-
diazole ring and the type of the aryl substituent had a strong bear-
ing on the inhibition, and the best compounds of the different
To the solution of ethyl 5-phenyl-1,3,4-oxadiazole-2-carboxyl-
ate (3) (1.5 g, 6.91 mmol) in the 1:1 mixture of THF/water
(78 mL) LiOH (248 mg; 10 mmol) was added and stirred at room
temperature for 30 min. Then the mixture was acidified with c
H2SO4 and the precipitated product was filtered off, washed with
water and dried on air. The product is light yellow crystals (900
mg, 68%, mp: decomposed over 230 °C). 1H NMR (DMSO-d6): d
(ppm) = 7.5–7.7 (m, 3H, aromatic), 8.0–8.1 (m, 2H, aromatic), 9.1
(s, 1H, COOH). 13C NMR (DMSO-d6): d (ppm): 124.5, 128.23,
130.5, 133.5, 166.15 (aromatic carbons), 178.5 (COOH). Anal. Calcd
for C9H6N2O3 (190.04): C, 56.85; H, 3.18; N, 14.73; Found: C, 56.94;
H, 14.85.
series were 11b (Ki = 30
lM), 15c (Ki = 33 lM), and 21a (Ki = 104 -
lM). ADMET property predictions revealed all ligands to have oral
‘drug-like’ properties based on Lipinski’s ‘rule of 5’. Apart from po-
tential permeability issues which would require ‘optimization’
based on Jorgensens ‘rule of three’, the inhibitors had satisfactory
pharmacokinetic profiles and were devoid of any toxicity struc-
tural warnings. The consideration of heterocyclic substitutions in
N-b-D-glucopyranosyl carboxamides is thus justified and further
syntheses and computational evaluation of such compounds will
be reported in due course.
4.3. 5-Phenyl-1,3,4-oxadiazole-2-carbonyl chloride (5)
4. Experimental
5-Phenyl-1,3,4-oxadiazole-2-carboxylic acid (4) (400 mg, 2.11
mmol) was dissolved in thionyl chloride (4 mL) and refluxed for
four hours. Then the mixture was concentrated in vacuum and
used without any purification.
4.1. General synthetic methods
Melting points were measured in open capillary tubes or on a
Kofler hot-stage and are uncorrected. Optical rotations were deter-
mined with a Perkin–Elmer 241 polarimeter at room temperature.
NMR spectra were recorded with Bruker 360 (360/90 MHz for
1H/13C) spectrometer. Chemical shifts are referenced to TMS as
the internal reference (1H), or to the residual solvent signals.
Microanalyses were performed on an Elementar vario Micro cube.
TLC was performed on DC-Alurolle Kieselgel 60 F254 (Merck). TLC
4.4. N-Cyanocarbonyl-2,3,4,6-tetra-O-acetyl-b-D-glucopyranosyl-
amine (8)
To
1,3-dioxane-4,6-dione35 (2.19 g, 6.7 mmol) in dry toluene
(50 mL) 2,3,4,6-tetra-O-acetyl-b- -glucopyranosylamine (7) (2.35 g,
a stirred solution of 2,2-dimethyl-5-(p-tosyloxyimino)-
D

M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
5743
6.8 mmol) was added and the mixture was stirred for 2 days at 50
°C. Subsequently the mixture was concentrated in vacuum, and the
residue purified by column chromatography (eluent: hexane/
ethyl-acetate = 1:1) to give 8 as white crystals. (1.54 g, 57%;
J = 9.8, 4.6 and 2.1 Hz, H-5), 4.16 (1H, dd, J = 12.6 and 4.6 Hz, H-
6A), 4.28 (1H, dd, J = 12.6 and 2.1 Hz, H-6B), 5.13, 5.18, 5.36 (3 ꢃ
1H, pseudo t, J = 9.8, 9.4 and 9.4 Hz, H-2, H-3, H-4), 5.44 (1H, d,
J = 9.4 Hz, H-1), 7.50 (2H, t, J = 7.3 Hz, aromatic), 7.56 (1H, m, aro-
matic), 8.15 (2H, d, J = 7.3 Hz, aromatic). 13C NMR (CDCl3) d
(ppm): 20.68 (COCH3), 20.79 (3 ꢃ COCH3), 61.74 (C-6), 68.09,
70.38, 72.90, 73.95 (C-2, C-3, C-4, C-5), 78.12 (C-1), 122.69,
127.68, 129.35, 132.95 (phenyl), 153.68, 157.67 (1,3,4-oxadiazole),
166.89 (NHCO), 169.64, 170.11, 170.63, 170.76, (COCH3). Anal.
Calcd for C23H25N3O11 (519.15): C, 53.18; H, 4.85; N, 8.09. Found:
C, 53.25; H, 4.96; N, 8.18.
[a]
D = 9.33 in CHCl3; c = 0.51); mp.: 125–145 °C). 1H NMR (CDCl3):
d (ppm): 2.02 (3H, s, COCH3), 2.04 (3H, s, COCH3), 2.08 (3H, s,
COCH3), 2.10 (3H, s, COCH3), 3.79–3.88 (1H, m, H-5), 4.10 (1H,
dd; J = 1.8 and 12.3 Hz; H-6A), 4.29, (1H, dd, J = 4.8 and 12.6 Hz;
H-6B), 5.00, 5.08, 5.29 (3 ꢃ 1H; pseudo t; J = 9.5–9,8; H-2; H-3; H-
4), 5.22 (1H, t, J = 9,2; H-1), 7.86 (1H, d, J = 9.2 Hz, NH). 13C NMR
(CDCl3): d (ppm): 20.66, 20.78 (4 ꢃ COCH3), 61.66 (C-6), 67.85;
70.25; 72.58; 74.20; 77.92 (C-1, C-2; C-3; C-4; C-5), 110.86 (CN),
143.48 (NHCO), 169.71; 170.05; 170.80; 171.18 (COCH3) Anal.
Calcd for C16H20N2O10 (400.11): C, 48.00; H, 5.04; N, 7.00; Found:
C: 48.05; H: 5.09; N: 7.03
4.8. N-(2,3,4,6-Tetra-O-acetyl-b-D-glucopyranosyl)-5-(naphth-2-
yl)-1,3,4-oxadiazole-2-carboxamide (10b)
By method B, starting from 2-naphthoyl chloride (640 mg,
4.5. N-(2,3,4,6-Tetra-O-acetyl-b-D-glucopyranosyl)-2H-tetrazole-
5-carboxamide (9)
3.37 mmol) and tetrazole
9 (1 g, 2.25 mmol) in dry toluene
(15 mL) to give 10b as white crystals (943 mg, 74%, mp: 186–
188 °C; [
a
]
D = ꢁ12.843 in DMSO, c = 0.61). 1H NMR (CDCl3) d
A solution of N-cyanocarbonyl derivative (8) (2.00 g, 5.0 mmol),
trimethylsilyl azide (2.23 mL, 16.96 mmol) and Bu2SnO (0.10 g,
0.42 mmol) in anhydrous toluene (90 mL) was stirred overnight
at 80 °C. Subsequently the mixture was concentrated in vacuum,
and the residue was crystallized from methanol to give 9 as white
(ppm): 2.04 (3H, s, COCH3), 2.06 (6H, s, 2 ꢃ COCH3), 2.07 (3H, s,
COCH3), 3.95 (ddd, J = 10.0, 4.3 and 2.0 Hz, H-5), 5.16, 5.23, 5.40,
5.49 (4 ꢃ 1H, pseudo t; J = 11.6, 11.3, 9.5, and 9.4 Hz; H-1, H-2,
H-3, H-4), 4.15 (1H, dd, J = 12.3 and 4.3, H-6A), 4.30 (1H, dd,
J = 12.6 and 2.0 Hz, H-6B), 7.54 (2H, m, aromatic), 7.81 (1H, d
J = 7.6 Hz; NH), 7.88 (2H, t, J = 7.2 Hz; aromatic), 8.09 (2H, m, aro-
matic), 8.57 (1H, s, H-1 naphthalene). 13C NMR (CDCl3) d (ppm):
20.67 (4 ꢃ COCH3), 61.83 (C-6), 68.24, 70.54, 72.96, 74.05, (C-2;
C-3; C-4; C-5), 78.29 (C-1), 123.23, 127.39, 128.03, 128.68,
129.12, 129.30, 132.75, 135.19 (aromatic), 153.76, 157.75 (1,3,4-
oxadiazole), 167.09 (NHCO), 169.59, 170.06, 170.68, 170.98
(COCH3). Anal. Calcd for C27H27N3O11 (569.16): C, 56.94; H, 4.78;
N, 7.38. Found: C, 57.02; H, 4.86; N, 7.45.
crystals (1.95 g; 88%); mp: 110–113 °C; [
a
]
D = ꢁ1.61 in CHCl3;
c = 0.335, mp: 174–177 °C) white solid. 1H NMR (CDCl3) d (ppm):
1.93 (3H, s, COCH3), 1.99 (9H, s, COCH3), 3.99 (1H, m, H-5), 4.10
(1H, dd, J = 12.3, <1, H-6A), 4.24 (1H, dd, J = 12.3, 2.8, H-6B), 5.22,
5.32, 5.47, 5.66 (4 ꢃ 1H, pseudo t, J = 9.6, 9.1 Hz in each, H-1, H-2,
H-3, H-4,), 7.26-7.01 (1H, brs, NH), 8.74 (1H, brs, CONH), 13C
NMR (CDCl3) d (ppm): 20.54 and 20.61 (COCH3), 61.78 (C-6),
68.01, 70.42, 73.07, 73.74 (C-2, C-3, C-4, C-5), 77.96 (C-1), 152.59
(tetrazole C-5), 156.97 (amide CO), 169.69, 170.23, 170.47,
170.83 (COCH3). Anal. Calcd for C16H21N5O10 (443.13): C, 43.34;
H, 4.77; N, 15.80. Found: C: 43.39; H: 4.89; N: 15.94.
4.9. N-(2,3,4,6-Tetra-O-acetyl-b-D-glucopyranosyl)-5-(naphth-1-
yl)-1,3,4-oxadiazole-2-carboxamide (10c)
4.6. General procedures for the preparation of N-(2,3,4,6-tetra-
O-acetyl-b-D-glucopyranosyl)-5-aryl-1,3,4-oxadiazole-2-
carboxamides
By method B, starting from 1-naphthoyl chloride (178 ll,
1.17 mmol) and tetrazole 9 (345 mg, 0.78 mmol) in dry toluene
(15 mL) to give 10c as white crystals (133 mg, 30%, mp: 124–
126 °C; [
a
]
D = ꢁ19.64 in CHCl3, c = 0.52). 1H NMR (CDCl3): d
Method A: To a solution of 2,3,4,6-tetra-O-acetyl-b-D-glucopyr-
(ppm) 2.06 (3H, s, COCH3), 2.07 (3H, s, COCH3), 2.08 (3H, s, COCH3),
2.09 (3H, s, COCH3), 3.98 (1H, ddd, J = 10.2, 4.6 and 2.2 Hz, H-5),
4.18 (1H, dd, J = 12.6 and 2.2 Hz, H-6A), 4.33 (1H, dd, J = 12.6 and
4,5 Hz, H-6B), 5,19; 5,28; 5,43, 5,55 (4 ꢃ 1H, pseudo t, J = 9.7, 9.4,
9.5 and 9.3, H-1; H-2; H-3; H-4) 7.46 (1H, m, aromatic), 7.53
(1H, m, aromatic), 7.66 (1H, m, aromatic), 7.84 (1H, t, J = 8.6 Hz,
aromatic), 7.97 (1H, d, J = 8.1 Hz, aromatic), 8.19 (1H, d,
J = 7.3 Hz, aromatic), 8.51 (1H, d, J = 9.2 Hz, CONH). 13C NMR
(CDCl3): d (ppm) 20.60, 20.70, 20.71 (COCH3), 61.79 (C-6), 68.09,
70.41, 72.97, 73.87, (C-2, C-3, C-4, C-5), 78.16 (C-1), 118.99,
124.84, 125.80, 136.88, 128.58, 128.81, 129.52, 129.90, 133.70
(aromatic), 153.85, 157.19 (1,3,4-oxadiazole), 166.77 (NHCO),
169.61, 170.08, 170.62, 170.70 (COCH3). Anal. Calcd for
C27H27N3O11 (569.16): C, 56.94; H, 4.78; N, 7.38. Found: C, 57.04;
H, 4.89; N, 7.46.
anosyl azide (6, 2.3 mmol) in dry dichloromethane (14 mL) a solu-
tion of PMe3 in toluene (2.3 mL) was added and the mixture was
stirred at room temperature. When the starting material was
transformed (TLC, eluent: hexane/ethyl-acetate = 1:1) an acid chlo-
ride (2.3 mmol) was added to the mixture and stirred at room tem-
perature for one day. Subsequently the mixture was concentrated
in vacuum and the residue was purified by column chromatogra-
phy (eluent hexane/ethyl-acetate = 2:1).
Method B: A solution of an aroyl-chloride (3.37 mmol) and tet-
razole 9 (2.25 mmol) in dry toluene (15 mL) was stirred at 80 °C
for 2 h. Then the mixture was cooled and concentrated in vacuum
and the residue was purified by column chromatography (eluent
hexane/ethyl-acetate = 2:1).
4.7. N-(2,3,4,6-Tetra-O-acetyl-b-D-glucopyranosyl)-5-phenyl-
1,3,4-oxadiazole-2-carboxamide (10a)
4.10. N-[2,3,4,6-Tetra-O-acetyl-b-D
-glucopyranosyl]-1-(N0-
hydroxycarbamimidoyl) formamide (12)
By method A, starting from 6 (858 mg, 2.3 mmol) and 5-phenyl-
1,3,4-oxadiazole-2-carbonyl chloride (5) (436 mg, 2.3 mmol) to
give 10a as white crystals (110 mg, 10%).
By method B, starting from benzoyl chloride (196 ll,
1.68 mmol) and tetrazole 9 (500 mg, 1.12 mmol) in dry toluene
To the solution of N-cyanocarbonyl derivative (9) (100 mg,
0.25 mmol) in anhydrous pyridine (0.5 mL) hydroxylamine hydro-
chloride (43.6 mg, 0.63 mmol) was added, and the reaction was
stirred for one hour at 50 °C. Subsequently it was acidified with
5% aqueous solution of HCl (10 mL) and extracted with ethyl ace-
tate (3 ꢃ 10 mL) and washed with water (2 ꢃ 10 mL) The organic
(6 mL) to give 10a as white crystals (470 mg, 80%, mp: 169–
171 °C; [
a
]
D = ꢁ4.672 in DMSO, c = 0.6). 1H NMR (CDCl3) d (ppm):
2.02 (6H, s, 2 ꢃ CH3), 2.04, 2.05 (6H, s, 2 ꢃ CH3), 3.95 (1H, ddd,

5744
M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
layer was dried over MgSO4 and concentrated in vacuum. The res-
idue was used without any purification. (77 mg, 71%).
(1H, d, J = 9.4 Hz; CONH), 8.19 (2H, d, J = 7.2 Hz, aromatic). 13C
NMR: d (ppm): 20.67, 20.79 (4ꢃ COCH3), 61.71 (C-6), 68.10,
70.51, 72.75, 73.97 (C-1, C-2, C-3, C-4), 78.10 (C-5), 123.14,
128.53, 129.40, 133.77 (phenyl), 156.78 (NHC@O), 162.27 (C-3 in
1,2,4-oxadiazole), 169.66, 170.00, 170.71, 170.88 (4 ꢃ COCH3),
177.34 (C-5 in oxadiazole). Anal. Calcd for C23H25N3O11 (519.15):
C, 53.18; H, 4.85; N, 8.09. Found: C, 53.27; H, 4.95; N, 8.19.
4.11. N-[2,3,4,6-Tetra-O-acetyl-b-D
-glucopyranosyl]-1-(N0-
benzoyloxycarbamimidoyl) formamide (13a)
A solution of 12 (77 mg, 0.178 mmol), benzoyl-chloride (23 ll,
0.196 mmol) and anhydrous pyridine (16 ll, 0.196 mmol) in anhy-
drous toluene (5 mL) was stirred for 24 h at 40 °C. for a day. Subse-
quently it was acidified with 5% aqueous solution of HCl (10 ml)
and extracted with ethyl acetate (3 ꢃ 10 ml) and washed with
water (2 ꢃ 10 mL). The organic layer was dried over MgSO4 and
concentrated in vacuum. Purification of the residue by column
chromatography (hexane/ethyl acetate = 1:1) gave 13a as white a
4.14. N-(2,3,4,6-Tetra-O-acetyl-b-D-glucopyranosyl)-5-(naphth-
2-yl)-1,2,4-oxadiazol-3-carboxamide (14b)
By method C, starting from 8 (300 mg, 0.75 mmol), hydroxyl-
amine hydrochloride (130.8 mg, 1.88 mmol) and 2-naphthoyl chlo-
ride (157 mg, 0.83 mmol) to give 14b as white crystals (95 mg;
crystal (50 mg, 52%, mp: 240–246 °C; [
a
]
D = ꢁ18.415 in CHCl3;
22%; mp: 175–180 °C; [
a]
D = ꢁ27.102 in CHCl3; c = 0.5). 1H NMR
c = 0.5). 1H NMR (CDCl3): d (ppm): 2.03 (3H, s, COCH3,), 2.04 (2 ꢃ
3H, s, COCH3), 2.10 (3H, s; COCH3), 3.84 (1H, ddd, J = 10.1, 3.8
and 2.2 Hz, H-5), 4.12 (1H, dd, J = 12.4 and 2.2 Hz, H-6A), 4.26
(1H, dd, J = 12.4 and 4.0 Hz, H-6B), 5.08, 5.11, 5.32, 5.33(4ꢃ 1H,
pseudo t; H-1, H-2, H-3; H-4), 5.65 (2H, brs; NH2), 7.47 (2H, t,
J = 7.2 Hz; aromatic), 7.61 (1H, t, J = 7.2 Hz, aromatic), 7.98 (1H, d,
J = 9.5 Hz, NH)8.04 (2H, d, J = 7.2 Hz, aromatic). 13H NMR (CDCl3):
d (ppm): 20.07, 20.85 (COCH3); 61.64 (C-6); 68.02, 70.40, 73.03,
73.83, 78.34 (C-1, C-2, C-3, C-4, C-5), 128.76, 129.72, 133.63,
147.77 (aromatic), 160.08, 163.26, 169.54, 170.15, 170.25, 170.86
(C@N, COPh, COCH3). Anal. Calcd for C23H27N3O12 (537.16): C,
51.40; H, 5.06; N, 7.82. Found: C, 51.49; H, 5.15; N, 7.91.
(CDCl3): d (ppm): 2.03 (6H, s, 2 ꢃ COCH3), 2.04 (3H, s, COCH3),
2.06 (3H, s, COCH3), 3.96 (1H, ddd, J = 10.01, 4.5 and 2.0 Hz H-5),
4.15 (1H; dd; J = 12.6 and 4.5 Hz H-6A), 4.35 (1H, dd, J = 12.6,
4.6 Hz, H-6B), 5.13, 5.14; 5,38; 5,51 (4 ꢃ 1H pseudo t, J = 9.8, 9.5,
9.5, 9.4 Hz, C-1; C-2; C-3; C-4), 7.54–7.68 (2H, m, aromatic), 7.90
(2H, d, J = 9.6 Hz), 7.97 (2H, d, J = 8.3 Hz, aromatic and NHCO),
8.14 (1H, dd, J = 8.6; 1.6 Hz, aromatic), 8.72 (1H, s, aromatic). 13C
NMR (CDCl3): d (ppm): 20.65; 20.76 (COCH3), 61.74 (C-6), 68.12,
70.54, 72.79, 73.96 (C-2; C-3; C-4; C-5), 78.80 (C-1), 120.22,
123.64, 127.50, 128.06, 129.07, 129.35, 130.02 132.62, 135.57 (aro-
matic), 156.82 (NHCO), 163.33 (C-3 in 1,2,4-oxadiazole), 169.64,
169.98, 170.68, 170.78 (COCH3), 177.46 (C-5 in 1,2,4-oxadiazole).
Anal. Calcd for C27H27N3O11 (569.16): C, 56.94; H, 4.78; N, 7.38.
Found: C, 57.02; H, 4.91; N, 7.47.
4.12. General procedure for the synthesis of N-(2,3,4,6-tetra-O-
acetyl-b-D-glucopyranosyl)-5-aryl-1,2,4-oxadiazole-3-
carboxamides
4.15. N-(2,3,4,6-Tetra-O-acetyl-b-D-glucopyranosyl)-5-(naphth-
1-yl)-1,2,4-oxadiazol-3-carboxamide (14c)
Method C: To a solution of N-cyanocarbonyl derivative 8
(300 mg, 0.75 mmol) in dry pyridine (1.5 mL) hydroxylamine
hydrochloride (131 mg, 1.87 mmol) was added and stirred at
50°C for 45 min. Dry toluene (10 mL) and 0.76 mmol acid chloride
was then added and the mixture was refluxed for 1.5 h. Subse-
quently a solution of TBAF (0.37 mL of a 1 M solution in THF)
was added to the mixture and refluxed for seven days (the progress
of the reaction was monitored by TLC using hexane/ethyl-ace-
tate = 1:1 as eluent). The mixture was diluted with 5% aq HCl
(30 mL) and extracted with ethyl acetate (3 ꢃ 30 mL), washed with
water (2 ꢃ 20 mL) and dried over MgSO4. The solvent was evapo-
rated in vacuum and the residue was purified by column chroma-
tography (eluent hexane/ethyl-acetate = 1: 1).
By method C, starting from 8 (300 mg, 0.75 mmol), hydroxyl-
amine hydrochloride (1308 mg, 1.88 mmol) and 1-naphthoyl chlo-
ride (124
mp: 175–180 °C; [
ll, 0.83 mmol) to give 14c as white crystals (145 mg, 22%,
a]
D = ꢁ27.47 in CHCl3; c = 0.5). 1H NMR (CDCl3):
d (ppm): 2.04 (3H, s, COCH3), 2.08 (9H, s, COCH3), 3.94 (1H, m, H-5),
4.12 (1H, d, J = 12.4 Hz, H-6A), 4.34 (1H, dd, J = 12.5 and 3.7 Hz, H-
6B), 5.14, 5.16, 5.40, 5.50 (4 ꢃ 1H, pseudo t; J = 9.7 Hz; 1H, t, J = 9.7,
9.4 and 9.3 Hz, H-1; H-2; H-3; H-4), 7.55–7.65 (2H, m, aromatic),
7.72 (1H, t, J = 7.2 Hz, aromatic), 7.93 (1H, d, J = 8.1 Hz, aromatic),
8.10 (1H, d, J = 9.1 Hz, NHCO), 8.10 (1H, d, J = 8.1 Hz, aromatic),
8.39 (1H, d, J = 7.3 Hz, aromatic), 9.10 (1H, d, J = 8.6 Hz, aromatic).
13C NMR (CDCl3): d (ppm): 20.67; 20.72; 20.80 (COCH3), 61.73 (C-
6), 68.81, 70.58, 72.72, 73.99 (C-2; C-3; C-4; C-5) 78.30 (C-1),
119.51, 130.08, 133.90, 124.95, 125.54, 127.10, 128.90, 129.02,
130.77, 134.76 (aromatic), 156.89 (NHCO), 163.14 (C-3 in 1,2,4-
oxadiazole), 169.66, 169.99, 170.72, 170.96 (COCH3), 177.47 (C-5
in 1,2,4-oxadiazole). Anal. Calcd for C27H27N3O11 (569.16): C,
56.94; H, 4.78; N, 7.38. Found: C, 57.03; H, 4.90; N, 7.46.
4.13. N-(2,3,4,6-Tetra-O-acetyl-b-D-glucopyranosyl)-5-phenyl-
1,2,4-oxadiazol-3-carboxamide (14a)
To the solution of 13a (80 mg 0.15 mmol) in dry toluene (2 mL) a
solution of TBAF (0.37 mL of a 1 M solution in THF) was added and
the mixture was refluxed for four days. Then the mixture was di-
luted with 5% aq HCl (30 mL) and extracted with ethyl acetate
(3 ꢃ 30 mL), washed with water (2 ꢃ 20 mL) and dried over MgSO4.
The solvent was evaporated in vacuum and the residue was purified
by column chromatography (eluent: hexane/ethyl-acetate = 1:1) to
give 14a as white crystals (60 mg, 76%, mp: 152–156 °C)
4.16. General procedure for the synthesis of N-(2,3,4,6-tetra-O-
acetyl-b-D-glucopyranosyl)-3-aryl-1,2,4-oxadiazole-5-
carboxamides
Method D: A solution of 2,3,4,6-tetra-O-acetyl-b-D-glucopyrano-
By method C, starting from 8 (300 mg, 0.75 mmol), hydroxyl-
amine hydrochloride (130.8 mg, 1.88 mmol) and benzoyl-chloride
(0.09 mL, 0.76 mmol) to give 14a as white crystals (96 mg, 24%,
sylamine (7, 275 mg, 0.79 mmol) in dry THF (15 mL) was added
dropwise to the cooled solution of oxalyl chloride (0.79 mmol) in
dry THF (10 mL) in one hour. Subsequently an arene-
carboxamidoxime (18a–c, 0.79 mmol) was added to the reaction
mixture and stirring was continued at room temperature for seven
days. The progress of the reaction was monitored by TLC (eluent:
hexane/ethyl-acetate = 1: 1). When the reaction was completed,
the solvent was removed in vacuum and the residue was purified
by column chromatography (eluent hexane/ethyl-acetate = 1: 1).
mp: 150–157 °C;
[a
]
D = ꢁ27.07 in CHCl3; c = 0.25). 1H NMR
(CDCl3): d (ppm): 2.05 (6H, s; 2 ꢃ COCH3), 2.07 (3H, s; COCH3),
2.10 (3H, s; COCH3), 3.94 (1H, m, C-5), 4.13 (1H, m, H-6A), 4.34
(1H, dd, J = 12.4 and 3.8 Hz, H-6B), 5.14 (2H, pseudo t, J = 9.4 Hz,
H-3, H-4), 5.39; 5.50 (2 ꢃ 1H, pseudo t, J = 9.4 Hz, H-1, H-2), 7.57
(2H, t, J = 7.2 Hz, aromatic), 7.66 (1H, t, J = 7.4 Hz, aromatic), 7.85

M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
5745
4.17. N-(2,3,4,6-Tetra-O-acetyl-b-
1,2,4-oxadiazole-5-carboxamide (20a)
D
-glucopyranosyl)-3-phenyl-
amount of NaOMe (1 M solution in methanol) was added and stir-
red at room temperature. The progress of the reaction was moni-
tored by TLC (chloroform/methanol = 9:1). When the starting
material was consumed the mixture was neutralised with a cation
exchange resin Amberlyst 15 (H+ form) or with acetic acid, then the
resin was filtered off and the solvent removed. The precipitated
product was filtered off, washed with ether and dried.
By method D, starting from benzamidoxime (18a) (389 mg;
2.87 mmol) to give 14 as white crystals (845 mg, 55%, mp: 154–
162 °C;
[
a]
D = ꢁ23.55 in CHCl3; c = 0.5). 1H NMR (CDCl3):
d
(ppm): 2.01 (6H, s, COCH3), 2.02 (3H, s, COCH3), 2.05 (3H, s,
COCH3), 3.92 (1H, ddd, J = 10.1, 4.1 and 2.0 Hz, C-5), 4.13 (1H, dd,
J = 12.5, 2.0 Hz, H-6A), 4.42 (1H, dd, J = 12.6, 4.6 Hz, H-6B), 5.10,
5.11, 5.36, 5.41 (4 ꢃ 1H, pseudo t J = 9.3, 9.7, 9.5 and 9.4 Hz, H-1
H-2; H-3; H-4), 7.40–7.61 (3H, m, aromatic), 7.9 (1H, d, J =
9.4 Hz; NHCO), 8.0–8.1 (2H, m, aromatic). 13C NMR (CDCl3): d
(ppm): 20.79, 20.68 (COCH3), 61.71 (C-6), 68.14, 70.55, 72.72,
74.19 (C-2; C-3; C-4; C-5), 78.42 (C-1), 125.59, 127.78, 129.18,
132.09 (aromatic), 153.31 (C-3 in 1,2,4-oxadiazole), 167.66
(NHCO), 169.10, 169.61, 169.96, 170.64 (COCH3), 170.85 (C-5 in
1,2,4-oxadiazole). Anal. Calcd for C23H25N3O11 (519.15): C, 53.18;
H, 4.85; N, 8.09. Found: C, 53.29; H, 4.97; N, 8.18.
4.21. N-(b-D-Glucopyranosyl)-5-phenyl-1,3,4-oxadiazole-2-
carboxamide (11a)
By method E, starting from 10a (90 mg, mmol) in a mixture of
dry methanol (3 mL) and dry chloroform (1.5 mL) to give 11a as
white crystals (56 mg, 50%, mp: 229–232 °C, [a]D = 7.82 in DMSO;
c = 0.45). 1H NMR (D2O): d (ppm): 4.90 (1H, d, J = 8.7 Hz, H-1),
7,57–7,78 (m; 3H), 8.04–8.18 (m, 2H), 13C NMR (D2O): d (ppm):
60.59 (C-6), 69.43, 71.34, 76.75, 78.62, 79.61 (C-1; C-2; C-3; C-4;
C-5), 122.34, 127.01, 129.41, 132.71 (aromatic), 153.40, 157.99
(1,3,4-oxadiazole), 165.02 (NHCO). Anal. Calcd for C15H17N3O7
(351.11): C, 51.28; H, 4.88; N, 11.96. Found: C, 51.38, H, 4.96, N,
12.01.
4.18. N-(2,3,4,6-Tetra-O-acetyl-b-D-glucopyranosyl)-3-(naphth-
2-yl)-1,2,4-oxadiazole-5-carboxamide (20b)
By method D, starting from naphth-2-amidoxime (18b)
(233 mg, 1.25 mmol) to give 20b as white crystals (109 mg; 15%,
4.22. N-(b-D-Glucopyranosyl)-5-(naphth-2-yl)-1,3,4-oxadiazole-
2-carboxamide (11b)
mp: 70–83 °C; [
a
]
D = ꢁ21.85 in CHCl3; c = 0.5). 1H NMR (CDCl3):
d (ppm): 2.06 (3H, s, COCH3), 2.07 (6H, s; 2 ꢃ COCH3), 2.09 (3H,
s, COCH3), 3.92 (1H, ddd, J = 9.7, 4.6 and 1.8 Hz, H-5), 4.14 (1H;
dd; J = 12.6 and 1.8 Hz; H-6A), 4.34 (1H, dd, J = 12,6 and 4.6 Hz;
H-6B), 5.14, 5.15 5.39, 5.46 (4 ꢃ 1H, pseudo t; J = 9.7, 9.7, 9.5 and
9.4 Hz, H-1, H-2, H-3, H-4), 7.51–7.69 (2H, m, aromatic), 7.90
(1H, d, J = 8.8 Hz, aromatic), 7.96 (3H, m, aromatic), 8.14 (1H, d,
J = 8,53 Hz, NHCO), 8.66 (1H, s, aromatic). 13C NMR: d (ppm):
20.71, 29.82 (COCH3), 61.72 (C-6), 68.12, 70.58, 72.76, 74.21 (C-
2; C-3; C-4; C-5), 78.41 (C-1), 122.85, 123.74, 127.16, 128.08,
128.72, 129.16, 130.02, 133.07, 135,08 (aromatic), 153,37 (C-3 in
1,2,4-oxadiazole), 167.68 (NHCO), 169.26, 169.66, 170.01, 170.71
(COCH3), 170,91 (C-5 in 1,2,4-oxadiazole). Anal. Calcd for
By method E, starting from 10b (411 mg, 0.72 mmol) in dry
methanol (2 mL) to give 11b as white crystals (244 mg, 85%, mp:
219–221 °C; [a]
D = 8.776 in DMSO; c = 0.69). 1H NMR (DMSO-d6)
d (ppm): 3.12 (1H, pseudo t, J = 9.2, Hz, H-3), 3.21–3.31 (4H, m,
H-2, H-4, H-5, H-6A), 3.69 (1H, d, J = 11.2 Hz H-6B), 4.61 (1H, brs
OH), 4.96 (1H, d, J = 9.2 Hz, H-1), 5.11 (2H, brs 2 ꢃ OH), 7.52–
7.79 (2H, m, aromatic), 8.05 (1H, d, J = 7.8 Hz, aromatic), 8.18
(3H, s, aromatic), 8.75 (1H, s, aromatic), 9.86 (1H, brs, NHCO). 13C
NMR (DMSO-d6) d (ppm): 60.9 (C-6), 70.00, 71. 93, 77.44, 79.18,
80.17 (C-1, C-2, C-3, C-4, C-5), 120.18, 123.23, 127.75, 128.15,
129.20, 129.57, 123.57, 134.74 (aromatic) 158.4, 153.6 (1,3,4-oxa-
diazole), 165.47 (NHCO). Anal. Calcd for C19H19N3O7 (401.12): C,
56.86; H, 4.77; N, 10.47. Found: C: 56.94; H, 4.87; N, 10.58.
C27H27N3O11 (569.16): C, 56.94; H, 4.78; N, 7.38. Found: C, 56.99;
H, 4.83; N, 7.41.
4.23. N-(b-D-Glucopyranosyl)-5-(napht-1-yl)-1,3,4-oxadiazole-2-
4.19. N-(2,3,4,6-Tetra-O-acetyl-b-
D
-glucopyranosyl)-3-(naphth-
carboxamide (11c)
1-yl)-1,2,4-oxadiazole-5-carboxamide (20c)
By method E, starting from 10c (300 mg, 0.53 mmol) in a mix-
By method D, starting from naphth-1-amidoxime (18c)
(168 mg, 0.90 mmol) to give 20c as a yellow oil (85 mg; 16%,
ture of dry methanol (7 mL) and dry chloroform (7 mL) to give
11c as white crystals (185 mg, 87%, mp: 230–234 °C; [a]D = 3.84
[
a]
D = ꢁ31.53 in CHCl3; c = 0.5). 1H NMR: d (ppm): 2.06, 2.07,
in DMSO; c = 0.62). 1H NMR (DMSO-d6) d (ppm):3.07–3.17 (1H,
m, H-5), 3.32–3.20 (2H, m, H-2, H-3), 3.41–3.51 (2H, m, H-4, H-
6B), 3.70 (1H, dd, J = 10.5 and 5.4 Hz, H-6A), 4.55–4.65 (1H, m,
OH), 4.95–5.05 (2H, m, H-1, OH) 5.10–5.17 (2H, m, OH), 7.59–
7.90 (3H, m, aromatic), 8.13 (1H, d, J = 8.0 Hz, aromatic), 8.28
(1H, d, J = 8.2 Hz, aromatic), 8.40 (1H, d, J = 7.3 Hz, aromatic), 9.13
(1H, d, J = 8.5 Hz, aromatic), 9.92 (1H, d, J = 8.7 Hz, NH). 13C NMR
(DMSO-d6): d (ppm): 62.56 (C-6), 71.27, 73.44, 78.75, 80.25,
81.41 (C-1; C-2; C-3; C-4; C-5), 120.61, 126.44, 126.75, 128.16,
129.60, 132.20, 130.69, 131.04, 134.69, 135.16 (aromatic),
155.51, 159.42 (oxadiazole), 167.08 (NHCO). Anal. Calcd for
2.08 (12H, s, 3 ꢃ COCH3), 3.92–3.97 (1H, ddd, J = 10.1, 4.36 and
1.90 Hz, H-5), 4.11–4.16 (1H, m, H-6A), 4.34 (1H, dd, J = 9.8 and
4.64, H-6B), 5.16, 5.41, 5.48 (2H, m; 1H, t,J = 9,5 Hz; 1H, t,
J = 9,3 Hz; H-1; H-2; H-3; H-4), 7.57 (2H, t, J = 3.4 Hz, aromatic),
7.65 (1H, m, aromatic), 7.92 (1H, d, J = 7.9 Hz, aromatic), 8.02
(1H, d, J = 8.2 Hz, aromatic), 8.17 (1H, d, J = 9.3 Hz, NHCO), 8.27
(1H, dd, J = 0.5, 7.2 Hz, aromatic), 8.88 (1H, d, J = 8.5 Hz, aromatic).
13C NMR: d (ppm): 20.61, 20.66, 20.74 (COCH3), 61.68 (C-6), 68.04,
70.49, 72.68, 74.03, 78.32 (C-1, C-2, C-3, C-4, C-5), 122.34, 130.32,
133.88, 125.07, 126.01, 126.62, 128.04, 128.84, 30.03, 132.74 (aro-
matic), 153.39 (NHCO), 166.73 (oxadiazole), 169.32, 169.59,
169.93, 170.62 (COCH3), 170.79 (oxadiazole). Anal. Calcd for
C19H19N3O7 (401.12): C, 56.86; H, 4.77; N, 10.47; O, 27.90. Found:
C: 56.91; H, 4.86; N, 10.56.
C27H27N3O11 (569.16): C, 56.94; H, 4.78; N, 7.38. Found: C, 57.16;
H, 4.88; N, 7.48.
4.24. N-(b-D-Glucopyranosyl)-5-phenyl-1,2,4-oxadiazol-3-
carboxamide (15a)
4.20. General procedure for the removal of O-acetyl protecting
groups
By method E, starting from 14a (132 mg, mmol) in dry metha-
nol (2 mL) to give 15a as white crystals (56 mg, 64%, mp: 245–
250 °C; [
(ppm): 3.07-3.71 (5H, m, H-2, H-3, H-4, H-5, H-6A), 3.79 (1H, d,
Method E: To the solution of a protected sugar derivative in dry
methanol (or in dry methanol and dry chloroform) a catalytic
a]
D = 8.83 in DMSO; c = 0.5). 1H NMR (DMSO-d6): d

5746
M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
J = 10.8, H-6B), 4.58 (1H, brs; OH), 4.93 (1H, d, J = 8.6 Hz, H-1), 5.07
(1H brs; OH), 7.6–7.8 (3H, m, aromatic), 8.10–8.25 (2H, m, aro-
matic), 9.49 (1H, brs; NH). 13C NMR (DMSO-d6): d (ppm): 60.98
(C-6), 69.89, 71.70, 77.45, 79.06, 79.84 (C-1, C-2, C-3, C-4, C-5),
122.96, 128.14, 129.74, 133.85 (aromatic), 156.88 (NHCO),
164.24, 176.03 (oxadiazole). Anal. Calcd for C15H17N3O7 (351.11):
C, 51.28; H, 4.88; N, 11.96. Found: C, 51.40, H, 4.99, N, 12.05.
3.71 (1H, dd, J = 10.9 and 3.3 Hz, H-6B), 4.59 (1H, brs, OH), 4.96
(1H, d, J = 9.1 Hz, H-1), 4.98; 4.99; 5.12 (3ꢃ 1H, brs, OH), 7.58–
7.75 (2H, m aromatic), 8.04 (1H; d, J = 7.3 Hz, aromatic), 8.14
(3H, s, aromatic), 8.72 (1H, s, aromatic), 10.01 (1H, brs; NHCO).
13C NMR (DMSO-d6): d (ppm): 60.99 (C-6), 69.89, 71.74, 77.30,
79.15, 80.12 (C-1, C-2, C-3, C-4, C-5), 122.85, 123.33, 127.34,
127.94, 128.21, 128.88, 129.26, 132.56, 133.40 (aromatic), 153.59
(NHCO), 168.24, 169.19 (oxadiazole). Anal. Calcd for C19H19N3O7
(401.12): C, 56.86; H, 4.77; N, 10.47. Found: C: 56.97; H, 4.88; N,
10.55.
4.25. N-(b-D-Glucopyranosyl)-5-(naphth-2-yl)-1,2,4-oxadiazol-3-
carboxamide (15b)
By method E, starting from 14b (95 mg, 0.17 mmol) in a mixture
of dry methanol (3 mL) and dry chloroform (1.5 mL) to give 15b as
white crystals (33 mg, 49%, mp: 254–258 °C; [a]D = 11.73 in
4.29. N-(b-D-glucopyranosyl)-3-(napht-1-yl)-1,2,4-oxadiazol-5-
carboxamide (21c)
DMSO; c = 0.5). 1H NMR (DMSO-d6): d (ppm): 2.97-3.88 (6H, m,
H-2, H-3, H-4, H-5,H-6), 4.58 (1H, m), 4.99 (1H, m, H-1), 5.04
(brm, OH), 7.7 (2H, m, aromatic), 8.05 (1H, m, aromatic), 8.20
(3H, m, aromatic), 8.89 (1H, s, aromatic), 9.48 (1H, brs, NHCO).
13C NMR (DMSO-d6): d (ppm): 60.99 (C-6), 69.92, 71.73, 77.44,
79.05, 79.85 (C-1, C-2, C-3, C-4, C-5), 120.20, 123.52, 127.65,
128.01, 129.16, 129.34, 129.50, 132.33, 135.50 (aromatic), 156.89
(NHCO), 164.30, 176.16 (oxadiazole). Anal. Calcd for C19H19N3O7
(401.12): C, 56.86; H, 4.77; N, 10.47. Found: C: 56.99; H, 4.91; N,
10.62.
By method E, starting from 20c (127 mg, 0.22 mmol) in a mix-
ture of dry methanol (2 mL) and dry chloroform (1 mL) to give
21c as white crystals (64 mg, 72%, mp: 245–250 °C; [
a
]
D = ꢁ6.17
in DMSO; c = 0.5). 1H NMR (DMSO-d6): d (ppm): 3.08–3.17 (1H,
m, H-5), 3.20–3.32 (2H, m, H-2, H-3); 3.40–3.55 (2H, m, H-4, H-
6A),, 4.60 (1H, brs, OH), 4.98 (1H, d, J = 9.3 Hz, H-1), 4.99 (1H, brs,
OH), 5.05–5.20 (2H, brs; 2 ꢃ OH), 7.69–7.73 (3H, m, aromatic),
8.10 (1H, d, J = 7.1 Hz, aromatic), 8.23 (1H, d, J = 7.5, aromatic),
8.3 (1H, d, J = 7.5, aromatic), 8.81 (1H, d, J = 7.7 Hz, aromatic),
10.05 (1H, brs. NHCO). 13C NMR (DMSO-d6): d (ppm): 60.99 (C-
6), 69.89. 71.78. 77.32. 79.18. 80.19 (C-1; C-2; C-3; C-4; C-5),
119.56, 122.34, 125.44, 126.79, 128.08, 128.94, 129.70, 132.54,
129.76, 133.52 (aromatic), 153.71 (NHCO), 164.03, 175.79 (oxadi-
azole). Anal. Calcd for C19H19N3O7 (401.12): C, 56.86; H, 4.77; N,
10.47. Found: C: 56.99; H, 4.90; N, 10.60.
4.26. N-(b-D-Glucopyranosyl)-5-(naphth-1-yl)-1,2,4-oxadiazol-3-
carboxamide (15c)
By method E, starting from 14c (170 mg, 0.3 mmol) in a mix-
ture of dry methanol (3 mL) and dry chloroform (2 mL) to give
15c as white crystals (64 mg, 54%, mp: 235–237 °C; [a]D = 7.68
4.30. General procedure for GP inhibition assay
in DMSO; c = 0.5). 1H NMR (DMSO-d6): d (ppm): 2.99–3.91; (6H,
m, H-2, H-3, H-4, H-5, H-6), 4.58 (1H, brs, H-1), 5.06 (2H, br m,
OH), 7.74 (3H, m, aromatic), 8.09 (1H, m, aromatic), 8.41 (2H,
m, aromatic), 9.14 (1H, s, aromatic), 9.61 (1H, brs; NHCO). 13C
Glycogen phosphorylase b was prepared from rabbit skeletal
muscle according to the method of Fischer and Krebs54 using 2-
mercaptoethanol instead of L-cysteine, and recrystallized at least
three times before use. The kinetic studies with glycogen phos-
phorylase were performed as described previously.41 Kinetic data
for the inhibition of rabbit skeletal muscle glycogen phosphorylase
by monosaccharide compounds were collected using different con-
NMR (DMSO-d6):
d (ppm): 60.90 (C-6), 69.85, 71.65, 77.40,
79.02, 79.82 (C-1, C-2, C-3, C-4, C-5), 119.09, 125.18, 125.33,
127.33, 128.68, 128.97, 130.44, 134.40, 129.22, 133.39 (aromatic),
156.83 (NHCO), 164.03, 175.79 (oxadiazole). Anal. Calcd for
C19H19N3O7 (401.12): C, 56.86; H, 4.77; N, 10.47. Found: C:
centrations of a-D-glucose-1-phosphate (4, 6, 8, 10, 12 and 14 mM)
56.93; H, 4.91; N, 10.61.
and constant concentrations of glycogen (1% w/v) and AMP
(1 mM). The enzymatic activities were presented in the form of
double-reciprocal plots (Lineweaver–Burk) applying a nonlinear
data-analysis programme. The inhibitor constants (Ki) were deter-
mined by Dixon plots, by replotting the slopes from the Linewe-
aver–Burk plots against the inhibitor concentrations. The means
of standard errors for all calculated kinetic parameters averaged
to less than 10%. IC50 values were determined in the presence of
4 mM glucose 1-phosphate, 1 mM AMP, 1% glycogen, and varying
concentrations of an inhibitor.
4.27. N-(b-D-glucopyranosyl)-3-phenyl-1,2,4-oxadiazol-5-
carboxamide (21a)
By method E, starting from 20a (467 mg, 0.9 mmol) in a mixture
of dry methanol (4 mL) and dry chloroform (4 mL) to give 21a as
white crystals (251 mg, 79%, mp: 250–256 °C; [a]D = 7.74 in
DMSO; c = 0.5).1H NMR (DMSO-d6): d (ppm): 3.15–3.51 (5H, m,
H-2, H-3, H-4, H-5, H-6A), 3.69 (1H, dd, J = 11.5 and 2.0 Hz, H-6B),
4.53 (1H, brs, OH),4.92 (1H, brs, OH), 4.93 (1H, d, J = 9.0 Hz, H-1),
5.08 (1H bs, OH), 7.66 (3H, m, aromatic), 8.13 (2H, m, aromatic),
9.96 (1H, brs, NH). 13C NMR (DMSO-d6): d (ppm): 60.98 (C-6),
69.88, 71.70, 77.30, 79.14, 80.11 (C-1, C-2, C-3, C-4, C-5), 125.48,
127.25, 129.44, 132.12 (aromatic), 153.57 (NHCO), 168.15,
4.31. ADMET property predictions
ADMET properties of the inhibitor analogues were predicted
using the QikProp 3.5 program (Schrodinger, LLC) in normal mode.
ALOGPS 2.145 was used to calculate supplementary logS and ‘con-
sensus’ logP(o/w) values for comparison with the QikProp values.
The RMSD between QikProp and ALOGPS values was calculated as:
169.15 (oxadiazole). Anal. Calcd for
C15H17N3O7 (351.11): C,
51.28; H, 4.88; N, 11.96. Found: C, 51.34, H, 4.98, N, 12.03.
4.28. N-(b-D-glucopyranosyl)-3-(napht-2-yl)-1,2,4-oxadiazol-5-
vffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
carboxamide (21b)
u
N
u
t
X
1
N
2
RMSD ¼
ðPQP ꢁ PALOGPS
Þ
ð1Þ
By method E, starting from 20b (90 mg, 0.16 mmol) in a mixture
of dry methanol (2 mL) and dry chloroform (1 mL) to give 21b as
i¼1
where PQP and PALOGPS represent the QikProp and ALOGPS val-
ues, respectively, of a property P. The FAF-Drugs2 server48 was
used to extract any toxicity structural warnings for the ligands.
white crystals (39 mg, 62%, mp: 256–259 °C; [
a]D = ꢁ2.25 in
DMSO; c = 0.5).1H NMR (DMSO-d6): d (ppm): 3.08–3.19 (1H, m,
H-5), 3.21–3.32 (2H, m, H-2, H-3); 4.41–3.52 (2H, m, H-4, H-6A),

M. Polyák et al. / Bioorg. Med. Chem. 21 (2013) 5738–5747
5747
19. Watson, K. A.; Mitchell, E. P.; Johnson, L. N.; Cruciani, G.; Son, J. C.; Bichard, C. J.
F.; Fleet, G. W. J.; Oikonomakos, N. G.; Kontou, M.; Zographos, S. E. Acta
Crystallogr., Sect D: Biol. Crystallogr. 1995, D51, 458.
20. Györgydeák, Z.; Hadady, Z.; Felföldi, N.; Krakomperger, A.; Nagy, V.; Tóth, M.;
Brunyánszky, A.; Docsa, T.; Gergely, P.; Somsák, L. Bioorg. Med. Chem. 2004, 12,
4861.
Inhibitors were initially prepared for the QikProp calculations
using Maestro and LigPrep (Schrodinger, LLC). Use of more ex-
tended conformations of ligands as input to QikProp can lead to
ADMET property predictions closer to their experimental equiva-
lents (QikProp version 3.5, User Manual). Confgen 2.3 (Schrodinger,
LLC), therefore, employing the OPLS-AA (2005) forcefield and the
Generalised Born/Surface Area (GB/SA) continuum model for bulk
solvation effects was used to generate low energy conformations
for each ligand (energy window of 5 kcal/mol). The most extended
conformation was then selected based on the calculated solvent
accessible surface areas (SASAs) using the Schrodinger python
script ‘conformer_geom_extent.py’ and used as input for the AD-
MET property predictions.
21. Gimisis, T. Mini-Rev. Med. Chem. 2010, 10, 1127.
22. Oikonomakos, N. G.; Kosmopoulou, M.; Zographos, S. E.; Leonidas, D. D.;
Chrysina, E. D.; Somsák, L.; Nagy, V.; Praly, J. P.; Docsa, T.; Tóth, A.; Gergely, P.
Eur. J. Biochem. 2002, 269, 1684.
23. Nagy, V.; Felföldi, N.; Kónya, B.; Praly, J.-P.; Docsa, T.; Gergely, P.; Chrysina, E.
D.; Tiraidis, C.; Kosmopoulou, M. N.; Alexacou, K.-M.; Konstantakaki, M.;
Leonidas, D. D.; Zographos, S. E.; Oikonomakos, N. G.; Kozmon, S.; Tvaroška, I.;
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Acknowledgments
This work was supported by the Hungarian Scientific Research
Fund (OTKA CK77712, CNK80709), TÁMOP-4.2.2.-08/1/2008-
0014, TÁMOP 4.2.1./B-09/1/KONV-2010-0007, and TÁMOP-4.2.2./
B-10/1-2010-0024 projects implemented through the New Hun-
gary Development Plan, co-financed by the European Social
Fund, TÁMOP 4.2.4.A/2-11-12012-0001 (B.Sz.) and Bolyai János Re-
search Fellowships of the Hungarian Academy of Sciences (to L.J.
and T.D.).
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