β-C-Glycosiduronic acids and β-C-glycosyl compounds: New PTP1B inhibitors

Article abstract of DOI:10.1016/j.bmcl.2008.10.091

β-C-Glycosiduronic acid quinones and β-C-glycosyl compounds have been synthesized as sugar-based PTP1B inhibitors. Benzoyl protected quinone derivatives (14 and 35) as well as aryl β-C-glycosyl compounds (18, 22, 23 and 34) showed IC₅₀ values of 0.77-5.27 μM against PTP1B, with compounds 18 and 23 bearing an acidic function being the most potent.

Full text of DOI:10.1016/j.bmcl.2008.10.091

Bioorganic & Medicinal Chemistry Letters 18 (2008) 6348–6351
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
b-C-Glycosiduronic acids and b-C-glycosyl compounds: New PTP1B inhibitors
a,
Li Lin a,b, Qiang Shen c, Guo-Rong Chen b, , Juan Xie
*
*
a PPSM, ENS Cachan, CNRS, UniverSud, 61 av President Wilson, F-94230 CACHAN, France
b Laboratory for Advanced Materials and Institute of Fine Chemicals, East China University of Science and Technology, Shanghai 200237, PR China
c National Center for Drug Screening, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
b-C-Glycosiduronic acid quinones and b-C-glycosyl compounds have been synthesized as sugar-based
PTP1B inhibitors. Benzoyl protected quinone derivatives (14 and 35) as well as aryl b-C-glycosyl com-
Received 5 September 2008
Revised 20 October 2008
Accepted 20 October 2008
Available online 1 November 2008
pounds (18, 22, 23 and 34) showed IC50 values of 0.77–5.27
and 23 bearing an acidic function being the most potent.
lM against PTP1B, with compounds 18
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
Protein tyrosine phosphatase 1B
Inhibitor
Diabetes
C-Glycosyl compounds
Quinone
Uronic acid
Protein tyrosine phosphatase 1B (PTP1B) has recently been
identified as new drug target for type 2 diabetes.1 In fact, PTP1B
has been shown to be an important regulator of tyrosine kinase
receptor-mediated responses, and influences negatively insulin
sensitivity.2 PTP1B knockout mice display increased insulin sensi-
tivity and tyrosine phosphorylation of the insulin receptor.3 These
results attracted considerable interest for the development of
pharmacological PTP1B inhibitors for the treatment of insulin
resistance, in particular non-insulin-dependent diabetes mellitus
(type 2 diabetes).4 Various strategies have been developed to de-
sign and synthesize potent and selective PTP1B inhibitors. The
principle approach is based on mimicking the phosphotyrosine
(pTyr) moiety. Numerous pTyr mimetics have been reported,
including difluoromethylphosphonates,5 cinnamic acid,6 oxalyla-
mino benzoic acid,7 isoxazole carboxylic acid,8 salicyclic acid,9
Consequently, sugar derivatives are promising starting point for
drug design. We have recently found that acetylated b-C-glucopyr-
anosyl-1,4-benzoquinone (compound I, Fig. 1) showed a good inhi-
bition against PTP1B (IC50 = 4.85 l
M).12 This result prompted us to
prepare other b-C-glycosyl compounds as sugar-based PTP1B
inhibitors. Since the active site of PTP1B is defined by residues
214–221 (the P-loop) and has a clear preference for acidic residues,
we then decided to introduce a carboxylic acid function on the C-
O
O
HO2C
OAc
RO
O
AcO
AcO
O
RO
OR
O
OAc
O
5 6 12 14
,
,
-
: R = Ac or Bz
I
a
-ketocarboxylic acid,10 etc. However, these highly polar and
CO2H
O
CO2H
charged compounds have limited cell membrane permeability.
Considerable effort has been devoted to the synthesis and struc-
ture modification of C-glycosyl compounds owing to their wide
natural existence, their biological interest and their high stability
towards acid- and enzyme-catalyzed hydrolysis.11 Furthermore,
sugar hydroxyl groups can be easily protected with various pro-
tecting groups to modulate their hydrophilicity/hydrophobicity.
O
OAc
OMe
NH
OMe
HN
BzO
BzO
O
O
AcO
OBz
OMe
OAcOMe
29
18, 23
O
O
OMe
O
O
NH
NH
RO
O
RO
RO
O
RO
OR
OMe
OR
* Corresponding authors. Tel.: +86 21 64253016; fax: +86 21 642552758 (G.-R.C.);
tel.: +33 1 47 40 55 86; fax: +33 1 47 40 24 54 (J.X.).
31 32 34
: R = H, Ac or Bz
33 35
,
,
,
: R = H, Ac or Bz
E-mail addresses: mrs_guorongchen@ecust.edu.cn (G.-R. Chen), joanne.xie@pps-
m.ens-cachan.fr (J. Xie).
Figure 1. Compounds synthesized or referred in this study.
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.10.091

L. Lin et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6348–6351
6349
glycosyl compounds (compounds 5, 6, 12–14, 18, 23 and 29, Fig. 1).
Both gluco- and galacto- derivatives have been synthesized. For
structure–activity comparison, we have also prepared 6-benzoyla-
mino derivatives (compounds 31–35).
To prepare 2-carbamoylbenzoic acid derivatives 18 and 23
(Scheme 2), b-C-aryl glucosides 1 and 7 were first tritylated at 6-
position, followed by protection of secondary hydroxyl function
as benzoyl ester. To avoid intramolecular transesterification reac-
tion, detritylation has been realized under acidic condition with
TFA to afford 16 and 21 in good yield. The 6-hydroxy group was
then transformed into azide via mesylate. PMe3 mediated Stau-
dinger protocol16 was then employed to convert azido sugars to
carbamoylbenzoic acid derivatives.
Reaction of 17 with phthalic anhydride in CH2Cl2 led to a mix-
ture of the desired compound 18 (47%) and N-phthalimide deriva-
tive 19 (22%). However, treatment of 22 with phthalic anhydride in
THF afforded 23 in 90% yield.
Synthesis of b-C-glycosiduronic acid quinone derivatives is
shown in Scheme 1. The gluco and galacto b-C-glycosides 113 and
213 were first deacetylated under Zemplén condition and silylated
at 6-position, followed by addition of Ac2O to furnish compounds 3
and 4 in one-pot. Treatment of these silylated b-C-glycosyl 1,4-
dimethoxybenzenes with Jones reagent led directly to the target
compounds 514 and 6.14 Similarly, we have prepared the acetyl
or benzoyl protected naphthoquinone derivatives 12–1414 from
the gluco and galacto b-C-glycosides 715 and 8.15 Jones reagent
has been very efficient to realise the one-pot desilylation and oxi-
dation reactions of compounds 9–11.
When acetyl group was used as protecting group in 24 (Scheme
3), detritylation with TFA led to an inseparable mixture of 25 and
R1
R1
O
O
R1
OTBS OMe
O
OMe
OMe
OAc
O
CO2H
R2
O
a-c
d
R2
R2
AcO
AcO
AcO
OAc
OAc
OMe
OAc
1: R1= H, R2= OAc
2: R1= OAc, R2= H
3: R1= H, R2= OAc
5: R1= H, R2= OAc
4: R1= OAc, R2= H
6: R1= OAc, R2= H
R1
O
O
R1
R1
OTBS
O
OR
OMe
OMe
OMe
CO2H
OH
O
a-c
or a, b, e
d
R2
O
R2
R2
RO
RO
HO
OR
OMe
OH
12: R = R2= OAc, R1= H
9: R = R2= OAc, R1= H
7: R1= H, R2= OH
13: R = R1= OAc, R2= H
10: R = R1= OAc, R2= H
8: R1= OH, R2= H
1
2
1
2
14
11
: R = R = OBz, R = H
: R = R = OBz, R = H
Scheme 1. Reagents and conditions: (a) MeONa, MeOH; (b) TBDMSCl, DMAP, pyr.; (c) Ac2O, 64% for 3, 72% for 4, 49% for 9 and 65% for 10; (d) Jones reagent, 22% for 5, 19% for
6, 78% for 12, 72% for 13 and 42% for 14; (e) BzCl, pyr, 58% for 11.
CO2H
O
OTr OMe
O
OH
O
OMe
OMe
OMe
OMe
N3
OMe
OMe
OMe
OMe
OMe
NPhth
O
OAc
O
NH
O
a-c
e, f
d
g
BzO
BzO
BzO
BzO
BzO
BzO
O
BzO
BzO
BzO
BzO
+
AcO
AcO
OBz
OBz
OBz
OBz
OBz
19
OMe
OMe
OAc
1
17
18
16
15
CO2H
O
OMe
OMe
OMe
OMe
OMe
OMe
OTr
OH
O
N3
O
OH
g
OMe
e, f
b, c
d
NH
BzO
BzO
BzO
BzO
O
O
BzO
BzO
HO
HO
O
BzO
OBz
BzO
OH
OBz
OBz
OMe
OMe
OBz
OMe
20
21
7
23
22
Scheme 2. Reagents and conditions: (a) MeONa, MeOH; (b) TrCl, pyr.; (c) BzCl, 61% (d) TFA, CH2Cl2, H2O, 16: 79%, 21: 83%; (e) MsCl, TEA; (f) NaN3, DMF, 57%; (g) phthalic
anhydride, Me3P, CH2Cl2 for 18 and 19 (18: 47%, 19: 22%), THF for 23 (90%).
OTr OMe
O
OMe
OMe
OAc OMe
OH OMe
O
OAc
O
a-c
d
AcO
AcO
O
HO
AcO
+
AcO
AcO
AcO
AcO
OAc
OAc
OMe
OAc
OAc
OMe
OMe
24
1
25
26
(mixture, 25/26 = 9:1)
CO2H
N
OMe
OMe
OMe
OMe
3 OAc
O
OAc
g
e
f
O
MsO
AcO
O
25
OMe
AcO
NH
OAc
OAc
27
OAc
28
O
AcO
29
OAc
OMe
Scheme 3. Reagents and conditions: (a) MeONa, MeOH; (b) TrCl, pyr.; (c) Ac2O, 89% (d) TFA, CH2Cl2, H2O, 78%; (e) MsCl, TEA, 55%; (f) NaN3, DMF, 71%; (g) phthalic anhydride,
Me3P, CH2Cl2, 74%.

6350
L. Lin et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6348–6351
Surprisingly, the benzoyl protected derivative 14 showed more po-
tent inhibition, with an IC50 value of about 1 M. Apparently, all
benzoyl protected compounds (18, 22, 23, 34 and 35) displayed
better inhibition than compound I, with the 2-carbamoylbenzoic
acid derivative 23 as the best inhibitor (IC50 = 0.77 lM). Presence
of 2-carbamoylbenzoic acid function on sugar ring improves
slightly the inhibition constant (18 vs. 34, 23 vs. 22) (See Table 1).
In summary, a series of b-C-glycosiduronic acid quinones and b-
C-glycosyl compounds have been prepared. Benzoyl protected sug-
ars exhibited good inhibitory activities against PTP1B with IC50 in
micromolar ranges. These results demonstrated the potential of
C-glycosyl compounds as a new class of small molecular inhibitors
of PTP1B.
NH2
O
OMe
OMe
NHBz
O
OMe
OMe
a
l
BzO
BzO
+
d
HO
17
31
BzO
OBz
30
OBz
31
O
O
NHBz
NHBz OMe
O
b, c
O
AcO
AcO
AcO
AcO
OAc
33
OAc
32
OMe
NHBz OMe
O
e
d
AcO
BzO
31
31
OBz
34
OMe
O
NHBz
O
Acknowledgments
HO
BzO
OBz
35
O
L.L. thanks the Ecole Normale Supérieure de Cachan for a Doc-
torate fellowship. This work was supported by CNRS, ENS Cachan,
National Natural Science Foundation of China (Grant No.
20876045) and Shanghai Science and Technology Community
(No. 074107018).
Scheme 4. Reagents and conditions: (a) Ph3P, THF, H2O, 31: 47%; (b) MeONa,
MeOH; (c) Ac2O, 81% (d) CAN, CH3CN, H2O, 33: 99%, 35: 77%; (e) Ac2O, pyr., 85%.
26, with the predominant formation of the transesterification
product 25. Subsequent mesylation allowed us to isolate the mes-
ylate 27 which was then transformed into 4-azido b-C-galactoside
28 with a SN2 mechanism.17 Treatment with phthalic anhydride
and PMe3 in CH2Cl2 afforded the 2-carbamoylbenzoic acid deriva-
tive 29.
References and notes
1. (a) Moller, D. E. Nature 2001, 414, 821; (b) Montalibet, J.; Kennedy, B. P. Drug
Discov. Today Ther. Strat. 2005, 2, 129.
2. (a) Byon, J. C. H.; Kusari, A. B.; Kusari, J. Mol. Cell. Biochem. 1998, 182, 101; (b)
Walchi, S.; Curchod, M. L.; Gobert, R. P.; Arkinstall, S.; Hooft van Huijsduijnen,
R. J. Biol. Chem. 2000, 275, 9792; (c) Cheng, A.; Dubé, N.; Gu, F.; Tremblay, M. L.
Eur. J. Biochem. 2002, 269, 1050.
Synthesis of 6-benzoylamino derivatives is described in Scheme
4. Staudinger reduction of azido function of 17 led to a mixture of
amine 30 contaminated with Ph3P@O and the transamidation
product 31 with a free hydroxyl group at 4-position. Structure of
31 was confirmed by RMN analysis of the 4-O-acetylated product
34.18 Treatment of 31 with MeONa followed by acetylation affor-
ded the compound 32 which was oxidized to the 1,4-benzoquinone
derivative 33 with CAN as oxidant. Direct CAN oxidation of 31 led
to the quinone 35 without affecting the 4-hydoxy group.
The effect of synthesized compounds on PTP1B was firstly stud-
3. (a) Elchebly, M.; Payette, P.; Michaliszyn, E.; Cromlish, W.; Collins, S.; Loy, A. L.;
Normandin, D.; Cheng, A.; Himms-Hagen, J.; Chan, C. C.; Ramachandran, C.;
Gresser, M. J.; Tremblay, M. L.; Kennedy, B. P. Science 1999, 283, 1544; (b)
Klaman, L. D.; Boss, O.; Peroni, O. D.; Kim, J. K.; Martino, J. L.; Zabolotny, J. M.;
Moghal, N.; Lubkin, M.; Kim, Y. B.; Sharpe, A. H.; Stricker-Krongrad, A.;
Schulman, G. I.; Neel, B. G.; Kahn, B. B. Mol. Cell. Biol. 2000, 20, 5479.
4. (a). Annu. Rev. Pharmacol. Toxicol. 2002, 42, 209; (b) Bialy, L.; Waldmann, H.
Angew. Chem. Int. Ed. 2005, 44, 3814; (c) Dewang, P. M.; Hsu, N.-M.; Peng, S.-Z.;
Li, W.-R. Curr. Med. Chem. 2005, 12, 1; (d) Lee, S.; Wang, Q. Med. Res. Rev. 2006,
1; (e) Liang, F.; Kumar, S.; Zhang, Z. Y. Mol. Biosyst. 2007, 308; (f) Zhang, S.;
Zhang, Z.-Y. Drug Discov. Today 2007, 12, 373; (g) Wan, Z.-K.; Follows, B.;
Kirincich, S.; Wilson, D.; Binnun, E.; Xu, W.; Joseph-McCarthy, D.; Wu, J.; Smith,
M.; Zhang, Y.-L.; Tam, M.; Erbe, D.; Tam, S.; Saiah, E.; Lee, J. Bioorg. Med. Chem.
Lett. 2007, 17, 2913; (h) Boustelis, I. G.; Yu, X.; Zhang, Z.-Y.; Borch, R. F. J. Med.
Chem. 2007, 50, 856; (i) Shrestha, S.; Bhattarai, B. R.; Chang, K. J.; Lee, K.-H.;
Cho, H. Bioorg. Med. Chem. Lett. 2007, 17, 2760; (j) Dixit, M.; Tripathi, B. K.;
Tamrakar, A. K.; Srivastava, A. K.; Kumar, B.; Goel, A. Bioorg. Med. Chem. 2007,
15, 727.
ied at 20 l
g/mL concentration.19,20 Except compounds 29, 32 and
33, all tested compounds showed good PTP1B inhibitory activity
(70.3–99.7%). Substitution of 6-OAc by 6-NHBz in compound I de-
press the potency (I vs. 33). The acetylated b-C-glycosiduronic acid
quinone derivatives 5, 6, 12 and 13 inhibited PTP1B with IC50 val-
ues from 16 to 28 lM. No significant difference can be observed
between gluco and galacto derivatives (5 vs. 6, 12 vs. 13). However,
5. Li, X. B. A.; Holmes, C. P.; Szardenings, A. K. Bioorg. Med. Chem. Lett. 2004, 4301.
6. Moran, E. J.; Sarshar, S.; Cargill, J. F.; Shahbaz, M. M.; Lio, A.; Mjalli, A. M. M.;
Armstrong, R. W. J. Am. Chem. Soc. 1995, 117, 10787.
7. Anderson, H. S. I. L. F.; Jeppesen, C. B.; Branner, S.; Norris, K.; Rasmussen, H. B.;
Moller, K. B.; Moller, N. P. H. J. Biol. Chem. 2000, 275, 7101.
these compounds are less effective than the parent compound I.
8. Liu, G.; Xin, Z.; Pei, Z.; Hajduk, P. J.; Abad-Zapatero, C.; Hutchins, C. W.; Zhao, H.;
Lubben, T. H.; Ballaron, S. J.; Haasch, D. L.; Kaszubsk, W.; Rondinone, C. M.;
Trevillyan, J. M.; Jirousek, M. R. J. Med. Chem. 2003, 46, 4232.
9. Larsen, S. D.; Barf, T.; Liljebris, C.; May, P. D.; Ogg, D.; O’Sullivan, T. J.; Palazuk, B.
J.; Schostarez, H. J.; Stevens, F. C.; Bleasdale, J. E. J. Med. Chem. 2002, 45, 598.
10. Xie, J.; Seto, C. T. Bioorg. Med. Chem. 2007, 15, 458.
11. (a) Postema, M. H. D. Tetrahedron 1992, 48, 8545; (b) Bu, Y.; Linhardt, R. J.
Tetrahedron 1998, 54, 9913; (c) Xie, J. Recent Res. Dev. Org. Chem. 1999, 3, 505;
(d) Grugier, J.; Xie, J.; Duarte, I.; Valéry, J. M. J. Org. Chem. 2000, 65, 979; (e) Xie,
J.; Thellend, A.; Becker, H.; Vidal-Cros, A. Carbohydr. Res. 2001, 334, 177.
12. Praly, J.-P.; He, L.; Qin, B. B.; Tanoh, M.; Chen, G. R. Tetrahedron Lett. 2005, 46,
7081.
13. Takeshi, K.; Nobuyuki, O.; Susumu, S. Tetrahedron Lett. 1998, 39, 4537.
14. Compounds 5, 6, 12–14 were prepared according to the following procedure.
To a soln of 6-O-silylated b-C-aryl glycosides (0.1 mmol) in 2 mL of acetone,
Table 1
In vitro PTP1B inhibition results.
Inhibitiona (%)
Inhibition IC50
(
lM)
b
Compound
I
5
6
4.85
84.7
70.3
83.1
88.2
99.5
99.7
96.5
92.1
49.5
94.9
12.1
26.6
83.4
94.1
20.43 ( 5.23)
27.61 ( 4.08)
15.95 ( 0.49)
16.55 ( 0.84)
1.12 ( 0.03)
2.44 ( 0.20)
2.36 ( 0.22)
0.77 ( 0.09)
nd
4.13 ( 0.19)
nd
nd
4.52 ( 0.07)
5.27 ( 0.61)
12
13
14
18
22
23
29
31
32
33
34
35
was added dropwise 170
l
L of Jones reagent (2.2 g CrO3 in 3.5 M H2SO4) at
0 °C. After 20 h reaction, another 100
lL of Jones reagent was added and the
mixture was stirred for 20 h. This operation was repeated two times until that
TLC indicated the complete consumption of the starting material. The mixture
was diluted with water and then extracted with CH2Cl2 (3 Â 20 mL). The
organic layer was dried over MgSO4 and purified by preparative layer
chromatography (CH2Cl2: MeOH = 20:1). Compound 5: [a]D = À3.4 (c 0.3,
CHCl3); 1H NMR (300 MHz, CDCl3): d 7.25–6.71 (m, 3H), 5.74–5.10 (m, 4H),
3.86 (m, 1H), 2.00 (s, 9H). HRMS: calcd for C18H18O11: 433.0747; found: m/z
a
Values tested at 20
l
g/mL concentration.
b
433.0753. Compound 6: [a]
D = À20.0 (c 0.26, CHCl3); 1H NMR (300 MHz,
Values are means of three experiments, standard deviation is given in paren-
theses (nd, not determinated).
CDCl3): d 7.04 (s, 1H), 6.73 (m, 2H), 5.79 (s, 1H), 5.16 (m, 2H), 4.58 (d, 1H,

L. Lin et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6348–6351
6351
J = 7.7 Hz), 4.26 (s, 1H), 2.10, 1.99, 1.90 (3s, 9H). 13C NMR (125 MHz, CDCl3): d
187.7, 186.2, 170.8, 170.7, 170.6, 169.7, 144.3, 137.2, 137.0, 135.0, 76.4, 72.5,
4.31–4.20 (m, 2H, H-6a, H-6b), 4.14 (d, 1H, J4,3 = 3.7 Hz, H-4), 3.94 (m, 1H, H-5),
3.78, 3.77 (2s, 6H, 2Â OMe), 2.12, 2.08, 1.80 (3s, 9H, 3Â COCH3).
72.0, 70.0, 69.1, 21.2. HRMS: calcd for C18H18O11
:
433.0747; found: m/z
18. 1H NMR (300 MHz, CDCl3): d 7.92–7.26 (m, 15H, Ph), 7.04 (d, 1H, J = 3.3 Hz, H–
Ar), 6.75 (dd, 1H, J = 2.9 Hz, J = 8.8 Hz, H–Ar), 6.66 (m, 2H, H–Ar, NHCO), 5.82 (t,
1H, J3,4 = J3,2 = 9.6 Hz, H-3), 5.68 (dd, 1H, J2,3 = 9.6 Hz, J2,1 = 9.9 Hz, H-2), 5.35 (t,
1H, J4,5 = J4,3 = 9.6 Hz, H-4), 5.18 (d, 1H, J1,2 = 9.9 Hz, H-1), 4.08–3.97 (m, 2H, H-
6a, H-5), 3.73, 3.60 (2s, 6H, 2Â OCH3), 3.47 (m, 1H, H-6b), 2.04 (s, 3H, COCH3).
19. Zhang, W.; Hong, D.; Zhou, Y.-Y.; Zhang, Y.-N.; Shen, Q.; Li, J.-Y.; Hu, L.-H.; Li, J.
Biochim. Biophys. Acta 2006, 1760, 1505.
433.0748. Compound 12: [
a
]
D = À11.3 (c 0.44, CHCl3); 1H NMR (300 MHz,
CDCl3): d 8.07, 7.76 (2s, 4H), 7.19 (s, 1H), 5.46–4.94 (m, 4H), 4.02–3.64 (m, 1H),
2.17, 2.03, 1.87 (3s, 9H); 13C NMR (75 MHz, CDCl3): d 184.6, 183.4, 170.2, 170.0,
169.8, 169.5, 145.4, 136.4, 134.3, 134.2, 131.9, 126.7, 126.5, 125.8, 73.2, 72.3,
72.2, 69.4, 63.3, 20.9, 20.7, 20.5. HRMS: calcd for C22H20O11: 483.0903; found:
m/z 483.0886. Compound 13: [a]
D = +4.9 (c 0.82, CHCl3); 1H NMR (300 MHz,
CDCl3): d 8.03, 7.69 (2 m, 4H), 7.23 (s, 1H), 5.82–4.32 (m, 4H), 3.95 (m, 1H),
2.06, 1.94, 1.82 (3s, 9H). HRMS: calcd for C22H20O11: 483.0903; found: m/z
20. The enzymatic activities of PTP1B catalytic domain were determined at 30 °C
by monitoring the hydrolysis of pNPP. Dephosphorylation of pNPP generates
product pNP, which can be monitored at 405 nm. In a typical 100 lL assay
483.0897. Compound 14: [
a
]
D = +8.8 (c 1.2, CHCl3); 1H NMR (300 MHz, CDCl3):
d 7.93–7.17 (m, 20H), 6.23, 5.76, 4.99 (3 s, 4H), 4.28 (s, 1H). HRMS: calcd for
C37H26O11Na: 669.1373; found: m/z 669.1380.
15. Lin, L.; He, X.-P.; Xu, Q.; Chen, G.-R.; Xie, J. Carbohrdr. Res. 2008, 343, 773.
16. Györgydeák, Z.; Hadady, Z.; Felföldi, N.; Krakomperger, A.; Nagy, V.; Tóth, M.;
Brunyánszki, A.; Docsa, T.; Gergely, P.; Somsák, L. Bioorg. Med. Chem. 2004, 12,
4861.
17. Inversion of configuration at C-4 can be confirmed by the small coupling
constant between H-3 and H-4: J4,3 = 3.7 Hz. 1H NMR (300 MHz, CDCl3): d 6.98
(d, 1H, J = 2.2 Hz, H–Ar), 6.80 (m, 2H, H–Ar), 5.51 (t, 1H, J2,3 = J2,1 = 9.9 Hz, H-2),
5.27 (dd, 1H, J3,2 = 9.9 Hz, J3,4 = 3.7 Hz, H-3), 4.85 (d, 1H, J1,2 = 9.5 Hz, H-1),
mixture containing 50 mM MOPS, pH 6.5, 2 mM pNPP and recombinant
enzymes, PTP1B activities were continuously monitored on a SpectraMax 340
microplate reader at 405 nm for 2 min at 30 °C and the initial rate of the
hydrolysis was determined using the early linear region of the enzymatic
reaction kinetic curve. For calculating IC50, inhibition assays were performed
with 30 nM recombinant enzyme, 2 mM pNPP in 50 mM MOPS at pH 6.5, and
the inhibitors diluted around the estimated IC50 values. IC50 was calculated
from the nonlinear curve fitting of percent inhibition (inhibition (%)) vs
inhibitor concentration [I] by using the following equation inhibition (%) = 100/
{1 + (IC50/[I])k}, where k is the Hill coefficient.