Synthesis and biological activity of glycosyl-1H-1,2,3-triazoles

Article abstract of DOI:10.1016/j.bmcl.2010.04.151

Glycosyl 1,2,3-triazoles with α-d-gluco, β-d-gluco, α-d-galacto, β-d-galacto and β-2-acetamido-2-deoxygluco (GlcNAc) stereochemistry were prepared by reaction of the corresponding azides with vinyl acetate under microwave irradiation. The deprotected glucosyl and galactosyl triazoles did not display inhibitory activity against the tested glycosidases at 1 mM. Of the four fungal glycosidases evaluated, GlcNAc-triazole was found to be hydrolyzed by Talaromyces flavus CCF 2686 β-N-acetylhexosaminidase. β-GlcNAc-triazole was furthermore established to act as a strong ligand of rat and human natural killer cell activating receptors.

Full text of DOI:10.1016/j.bmcl.2010.04.151

Bioorganic & Medicinal Chemistry Letters 20 (2010) 4263–4265
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological activity of glycosyl-1H-1,2,3-triazoles
Krist y´ na Slámová , Petr Marhol , Karel Bezouška , Lise Lindkvist , Signe G. Hansen , Vladimír K rˇ en a,*,
Henrik H. Jensen
a
a
a
a,c
b
b
b,
*
Centre of Biocatalysis and Biotransformation, Institute of Microbiology, Academy of Sciences of the Czech Republic, Víde nˇ ská 1083, CZ 14220, Prague, Czech Republic
Department of Chemistry, Aarhus University, Langelandsgade 140, DK-8000 Aarhus C, Denmark
Department of Biochemistry, Faculty of Science, Charles University Prague, Hlavova 8, CZ-12840 Praha 2, Czech Republic
b
c
a r t i c l e i n f o
a b s t r a c t
Article history:
Glycosyl 1,2,3-triazoles with a-D-gluco, b-D-gluco, a-D-galacto, b-D-galacto and b-2-acetamido-2-deoxy-
Received 15 April 2010
Revised 28 April 2010
Accepted 29 April 2010
Available online 15 May 2010
gluco (GlcNAc) stereochemistry were prepared by reaction of the corresponding azides with vinyl acetate
under microwave irradiation. The deprotected glucosyl and galactosyl triazoles did not display inhibitory
activity against the tested glycosidases at 1 mM. Of the four fungal glycosidases evaluated, GlcNAc-tria-
zole was found to be hydrolyzed by Talaromyces flavus CCF 2686 b-N-acetylhexosaminidase. b-GlcNAc-
triazole was furthermore established to act as a strong ligand of rat and human natural killer cell activat-
ing receptors.
Keywords:
Glycosidase inhibition
b-N-Acetylhexosaminidase
N-Glycoside
Ó 2010 Elsevier Ltd. All rights reserved.
NK cell
The Cu(I)-catalyzed alkyne-azide cycloaddition has become
widely used reaction for efficient conjugation of two reaction part-
Each glycosyl azide (4–6, 12–13) was dissolved in vinyl acetate
and heated under microwave irradiation at 120 °C until full con-
version resulted as judged by TLC analysis. Interestingly, b-config-
ured azides 4 and 5 reacted significantly faster than their
1,2
3
ners, for example, a saccharide with an aglycone or even a pro-
4
tein.
This has produced interesting, nitrogen-displaying
carbohydrate mimics, which have been studied due to their poten-
tial use as medicinal agents (Fig. 1).5,6 The effect of C-unsubstituted
glycosyl triazoles, that is, the 1,2,3-triazole moiety itself, however,
corresponding a-configured counterparts (12 and 13). An identical
trend was recently observed and explained by Field and co-work-
ers for the Cu(I)-catalyzed reaction of O-acetylated glycosyl azides
7
13
has not been studied with the exception of xylosyl triazoles.
with terminal alkynes.
In this Letter, we report the first synthesis of five different
C-unsubstituted glycosyl triazoles (7–9, 14–15, Scheme 1), their
deacetylated equivalents (16–20), and our investigations of these
N-glycosides as potential inhibitors or substrates of glycosidases.
We also report on the NK cell activation receptor interaction with
the b-GlcNAc-1,2,3-triazole (18).
All glycosyl triazoles were treated under standard Zemplén con-
ditions to give the desired deprotected compounds 16–20 in quan-
titative yield (Scheme 2).
N
N
N
HO OH
O
CH3
O
O
N
N
N
13
HO
The convenient and direct synthesis of isomerically pure C-
unsubstituted 1,2,3-triazoles by treatment of an azide with vinyl
HO
OH
HO
OH
acetate under microwave (lw) irradiation was recently reported
8
by us. Although the established protocol was easy to carry out
and typically provided high product yields, it was not clear whether
potentially sensitive glycosyl azide would withstand the reaction
conditions.
4-Phenyl galactosyl triazole
(galactosidase inhibitor)
C-linked arabinosyl triazole
(anti-mycobacterial activity)
b-Glycosyl azides (4–6) were prepared as described by Roy and
OH
9
co-workers, while
the treatment of the corresponding b-glycosyl chloride following
a
-glycosyl azides (12 and 13) were prepared by
N
N
N
O
N
N
N
HO
HO
O
10
OAr
HO
HO
1
1,12
OH
a protocol similar to that published by DeSong and co-workers.
OH
C-linked xylosyl triazole
(initiator of GAG biosynthesis)
C-unsubstituted glycosyl triazole
*
Corresponding authors. Tel.: +420 296442510 (V.K.); tel.: +45 89423963; fax:
(this study)
+
45 86196199 (H.H.J.).
Figure 1. Examples of triazole-linked saccharides.5–7
E-mail addresses: kren@biomed.cas.cz (V. K rˇ en), hhj@chem.au.dk (H.H. Jensen).
0
960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.04.151

4
264
K. Slámová et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4263–4265
Rax
Rax
Rax OAc
O
OAc
O
OAc
NaN3
,
TBAHSO4
vinyl acetate
N
N
eq
eq
eq
R
R
O
R
NaHCO3,
H
O/CH
Cl
AcO
N3
º
AcO
N
AcO
2
2
2
µw, 120 C
hours
AcX
see, ref. 9
AcX
AcX
6
Cl
eq
eq
eq
=OAc, Rax=H, X=O;
eq
eq
eq
=OAc, Rax=H, X=O; 7 (76%)
eq
eq
eq
ax
4
5
R
R
R
R
R
R
R
R
R
=OAc, R =H, X=O;
1
2
ax
ax
ax
=H, R =OAc, X=O; 8 (78%)
=H, R =OAc, X=O;
=H, R =OAc, X=O;
ax
ax
ax
=OAc, R =H, X=NH ; 6
=OAc, R =H, X=NH; 9 (71%)
=OAc, R =H, X=NH; 3
Rax
Rax
Rax
OAc
OAc
OAc
TBA-N3
Cl reflux in THF, 60 hours
vinyl acetate
eq
eq
eq
R
O
R
O
R
O
AcO
AcO
µw, 120 C
7 hours
º
AcO
AcO
AcO
AcO
3
N3
N
N
N
eq
=OAc, Rax=H; 10
eq
=OAc, Rax=H; 12, 44% (4 (6%))
eq
=OAc, Rax=H; 14, 63%
R
R
R
R
R
R
eq
ax
eq
ax
eq
ax
=H, R =OAc; 11
=H, R =OAc; 13, 44% (5 (14%))
=H, R =OAc; 15, 57%
Scheme 1. Synthesis of glycosyl 1,2,3-triazoles from the corresponding azides under microwave (
lw) irradiation in vinyl acetate.
Rax
Rax
OAc
O
OH
O
eq
CH
3
ONa (cat)
eq
R
R
E
E
AcO
CH OH, quant.
HO
3
AcX
X
A
A
eq
eq
eq
eq
eq
=OAc, Rax=A=H, X=O; E=N-(1,2,3-triazolyl);
eq
eq
eq
eq
eq
=OH, Rax=A=H, X=OH; E=N-(1,2,3-triazolyl); 16
R
R
R
R
R
7
8
9
R
R
R
R
R
ax
ax
=A=H, R =OAc, X=O; E=N-(1,2,3-triazolyl);
=A=H, R =OH, X=OH; E=N-(1,2,3-triazolyl); 17
ax
ax
=OAc, R =A=H, X=NH; E=N-(1,2,3-triazolyl);
=OH, R =A=H, X=AcNH; E=N-(1,2,3-triazolyl); 18
ax
ax
=OAc, R =E=H, X=O; A=N-(1,2,3-triazolyl); 14
=OH, R =E=H, X=OH; A=N-(1,2,3-triazolyl); 19
ax
ax
=E=H, R =OAc, X=O; A=N-(1,2,3-triazolyl); 15
=E=H, R =OH, X=OH; A=N-(1,2,3-triazolyl); 20
Scheme 2. Zemplén deprotection of acetylated glycosyl 1,2,3-triazoles.
With the deprotected glycosyl triazoles (16–20) in hand, their
inhibitory potential was evaluated against common glycosidases.
Glycosyl triazoles with electron withdrawing substituents have
previously been explored as donor substrates in chemical glycosyl
1
5
16,17
No inhibition of sweet almond b-glucosidase was observed at
transfer reactions to fluoride or thiols/alcohols.
Therefore, we
1
mM by b-glucosyl triazole (16).14 Interestingly, b-galactosyl tria-
were interested to see whether glycosyl triazoles too could be
enzymatically hydrolysed by glycosidases. No such cleavage was
found in a previous study by Rossi and Basu, who studied the 4-
phenyl galactosyl triazole (Fig. 1) and its gluco-isomer against a
zole (17) did not show any sign of inhibition of Escherichia coli b-
galactosidase at 1 mM, although the analogous 4-phenyl substi-
tuted counterpart (Fig. 1) has been reported to inhibit this enzyme
5
5
with K
i
= 330
l
M. b-Galactosyl triazole (17) was furthermore
range of glycosidases. To the best of our knowledge, no such en-
tested against Aspergillus oryzae b-galactosidase without inhibition
at 1 mM. -Glycosyl triazoles 19 and 20 were in a similar way eval-
uated against yeast -glucosidase and green coffee bean -galacto-
sidase, respectively, without inhibition at 1 mM.
b-GlcNAc-triazole (18) was evaluated as a potential inhibitor of
extracellular fungal b-N-acetylhexosaminidase (Aspergillus oryzae
CCF 1066, Penicillium oxalicum CCF 1959, P. pittii CCF 2277 and
Talaromyces flavus CCF 2686), accordingly, no inhibition of these
enzymes was observed at 2 mM.
zyme able to catalyze the cleavage of a C–N glycosidic bond (where
1
8
a
nitrogen is neutral) has prior to this been reported with the
1
9–21
a
a
exception of glycosyl azide hydrolysis.
It was decided to focus
on GlcNAc-triazole 18 and investigate its potential hydrolysis by
substrate-promiscuous fungal b-N-acetylhexosaminidases. The
same four fungal b-N-acetylhexosaminidases as tested for inhibi-
tion by 18 were screened in this study (vide supra). Although the
reaction appeared to be very slow, b-N-acetylhexosaminidase from
T. flavus was indeed able to process GlcNAc-triazole 18. No hydro-
mAU
H
4
3
3
2
2
1
1
00
50
00
50
00
50
00
N
N
N
OH
HO
HO
O
OH
AcNH OH
N
N
N
HO
HO
O
AcNH
5
0
0
50
-
0.0
2.5
5.0
7.5 10.0 12.5 15.0 17.5 20.0 22.5 25.0 min
Figure 2. HILIC chromatogram after 22 h of reaction (left). Formation of 1,2,3-triazole by T. flavus b-N-acetylhexosaminidase catalyzed hydrolysis of 18 (right).

K. Slámová et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4263–4265
4265
Table 1
activity towards a range of glycosidases, however, by an estab-
lished HPLC protocol b-N-acetylhexosaminidase from Talaromyces
flavus proved to be able to cleave the intact triazole. This C–N gly-
coside hydrolysis of GlcNAc-triazole 18 is an exceptional feature of
this particular O-glycosidase.
We have furthermore demonstrated the significant binding
affinity of GlcNAc-triazole (18) towards the NK cell activating
receptors (Table 1) CD69 and NKRP-1. This activity could possibly
be considerably enhanced by the construction of a dendrimeric
GlcNAc-display by Cu(I)-catalyzed azide-alkyne cycloaddition.
Values for binding to lymphocyte activating receptors (Àlog IC50
)
NKR-P1A (rat)
CD69 (human)
3.9 ± 0.1
OH
O
HO
HO
5.7 ± 0.1
7.2 ± 0.2
OH
AcNH
(
GlcNAc)
OH
N
N
N
O
HO
HO
5.4 ± 0.2
Acknowledgments
AcNH
(
18)
We thank OChem Graduate School, Aarhus University, Denmark
for financial support (SGH). The Czech Science Foundation (305/09/
H008) and the research concept of the Institute of Microbiology
AV0Z50200510 are acknowledged. K. Krenek is thanked for the
measurement of NK receptor activity.
lysis was found in the absence of enzyme or among the remaining
three b-N-acetylhexosaminidases.
To study the T. flavus catalyzed reaction in detail, a new HPLC
(
HILIC) method was developed to monitor the concentration of
Supplementary data
all reaction mixture components by determining the concentration
of 1,2,3-triazole (Fig. 2, left). Hydrolysis of 18 was found to be lin-
ear in time (Fig. 2, right), but 2000 times slower compared to the
standard substrate 4-nitrophenyl 2-acetamido-2-deoxy-glucopy-
ranoside (pNP-GlcNAc) (2.7 mU vs 5 U, respectively). Despite the
slow reaction rate, the hydrolysis of a C–N bond by an O-glycosi-
dase is a rather unique finding. Upon this result, T. flavus b-N-acet-
ylhexosaminidase was also tested as a potential catalyst for
reaction with another C–N glycoside that is, the thioureido linkage
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.04.151.
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