Carbonic anhydrase inhibitors: Inhibition of isozymes I, II, and IX with triazole-linked O-glycosides of benzene sulfonamides

Article abstract of DOI:10.1021/jm061320h

We report the synthesis of a series of benzene sulfonamides containing triazole-O-glycoside tails for evaluation as carbonic anhydrase (CA) inhibitors. These glycoconjugates were synthesized by the 1,3-dipolar cycloaddition reaction of 4-azidobenzenesulfo

Full text of DOI:10.1021/jm061320h

J. Med. Chem. 2007, 50, 1651-1657
1651
Carbonic Anhydrase Inhibitors: Inhibition of Isozymes I, II, and IX with Triazole-Linked
O-Glycosides of Benzene Sulfonamides
Brendan L. Wilkinson,† Laurent F. Bornaghi,† Todd A. Houston,‡ Alessio Innocenti,§ Daniela Vullo,§ Claudiu T. Supuran,*,§ and
Sally-Ann Poulsen*,†
Eskitis Institute for Cell and Molecular Therapies, Griffith UniVersity, 170 Kessels Road, Nathan, Queensland 4111, Australia, Institute for
Glycomics, Griffith UniVersity, PMB 50 Gold Coast Mail Centre, Gold Coast, Queensland 9726, Australia, and Polo Scientifico, Laboratorio di
Chimica Bioinorganica, Room 188, UniVersita` degli Studi di Firenze, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy
ReceiVed NoVember 14, 2006
We report the synthesis of a series of benzene sulfonamides containing triazole-O-glycoside tails for evaluation
as carbonic anhydrase (CA) inhibitors. These glycoconjugates were synthesized by the 1,3-dipolar
cycloaddition reaction of 4-azidobenzenesulfonamide with O-propynyl glycosides. Compounds were assessed
for their ability to inhibit the enzymatic activity of the physiologically dominant isozymes hCA I and II and
the tumor-associated isozyme hCA IX (h ) human). Against hCA I these compounds were either micromolar
or low-nanomolar inhibitors, while against hCA II and IX inhibition in the range of 6.8-53 and 9.7-107
nM, respectively, was observed. The most potent inhibitor against hCA IX was the galactose derivative 8
(Ki ) 9.7 nM); this is so far the most potent glycoconjugate inhibitor reported for the tumor-associated
hCA IX. These carbohydrate-tethered sulfonamides may prove interesting lead candidates to target tumor-
associated CA isozymes, wherein the CA domain is located extracellularly.
Introduction
reducing the catalytic ability of the enzyme. This CA recognition
fragment15,16 exhibits remarkable reliability in anchoring the
inhibitor molecule within the CA active site. The addition of
“tail” groups to this zinc-binding functionality has made for an
effective means to incorporate into and optimize the chemical
and biological properties of sulfonamide CA inhibitors.14
Carbonic anhydrasesa (CAs, EC 4.2.1.1) are Zn(II) metal-
loenzymes that catalyze the reversible hydration of carbon
dioxide to give bicarbonate anion and a proton: CO2 + H2O
T HCO3 + H+.1 This reaction is known to regulate a broad
-
range of physiological functions, and clinical modulation of CA
activity by inhibitors has proven a reliable treatment for a range
of human disease states.1-4 In humans, 15 different CA isozymes
(designated hCA) belonging to the R-CA class are presently
known, and many of these isozymes are quite recent discoveries
compared with the physiologically abundant and widely dis-
tributed isozymes hCA I and II.1-4 An understanding of the
physiological consequences of inhibiting (or activating) specific
CA isozymes is developing as the knowledge base regarding
CA isozyme enzyme kinetics, tissue distribution, expression
levels, and subcellular locations builds. Many of these isozymes
have shown promise as druggable targetsswith CAs now
implicated in areas where human therapies are much needed,
for example, in cancer-specific (CA IX, XII)5-8 and obesity-
specific (CA V)9-11 pathways and in brain function (CA
XIV).12,13 Concomitant with this expanded therapeutic potential
is an increased demand for CA isozyme-specific inhibitorss
necessary to avoid side effects and to improve therapeutic safety.
The classical recognition motif for small molecules to bind
the active site of CA is an aromatic sulfonamide moiety
(ArSO2NH2).1-4,14 The deprotonated sulfonamide (ArSO2NH-)
coordinates to the CA active site Zn2+, and so inhibits the
binding of the endogenous substrates (CO2 and H2O), thereby
Of particular interest in our research is the modulation of
CA for cancer applicationsseither as an anticancer therapy or
as a tool for cancer diagnosis and to monitor the effectiveness
of treatment. There are two known cancer-associated CA
isozymes, CA IX and XII. The genes that encode these isozymes
are induced by the hypoxia inducible factor-1 (HIF-1), and
elevated levels of CA IX and XII are a marker for a broad
spectrum of solid hypoxic tumor types.8,17,18 The activity of these
isozymes contributes to the extracellular acidification (pHe) of
the solid tumor environment, with multiple downstream effects
of this reduced pHe linked to tumor invasion and poor
prognosis.5,17,18 Aromatic sulfonamide compounds (the classical
CA inhibitors) have been shown to reverse the effect of tumor
acidification, to inhibit the growth of cancer cells, and to
suppress tumor invasion mediated by the cancer-associated
CAs.5,17-20 The expression profile of these tumor-associated CAs
offers the possibility for CA inhibitors to be developed that can
discriminate cancerous cells (overexpression) from healthy cells
(minimal expression). Isozymes CA IX and XII also share a
global topology that distinguishes them from the physiologically
dominant hCA I and IIsisozymes IX and XII are transmem-
brane proteins that orient their CA catalytic domain extracel-
lularly, while isozymes I and II are soluble proteins located in
the cytosol. The preparation of aromatic sulfonamides with an
impaired ability to diffuse through lipid membranes is therefore
one possible means by which to selectively target these
isozymes.21-25 It is well-known that carbohydrates26,27 have a
compromised ability to diffuse across cell membranes, and
recently our group has demonstrated that by “click-tailing” sugar
moieties to the classical high-affinity aromatic sulfonamide
pharmacophore (ArSO2NH2) we were able to deliver glyco-
conjugate sulfonamide inhibitors that were potent and selective
* Corresponding authors. Phone: +61 7 3735 7825 (S.-A.P.); +39 055
457 3005 (C.T.S.). Fax: +61 7 3735 7656 (S.-A.P.); +39 055 457 3385
(C.T.S.). E-mail: s.poulsen@griffith.edu.au (S.-A.P.); claudiu.supuran@
unifi.it (C.T.S.).
† Eskitis Institute for Cell and Molecular Therapies, Griffith University.
‡ Institute for Glycomics, Griffith University.
§ Universita` degli Studi di Firenze.
a Abbreviations: CA, carbonic anhydrase; 1,3-DCR, 1,3-dipolar cy-
cloaddition reaction; AZA, acetazolamide; MZA, methazolamide; EZA,
ethoxzolamide; DCP, dichlorophenamide; BRZ, brinzolamide; IND, In-
disulam.
10.1021/jm061320h CCC: $37.00 © 2007 American Chemical Society
Published on Web 03/08/2007

1652 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 7
Wilkinson et al.
Scheme 1a
a Reagents and conditions: (i) azide 1 (0.2-0.5 M), O-propynyl glycoside 2-6 (1 equiv), CuSO4‚5H2O (0.1-0.2 equiv), sodium ascorbate (0.2-0.4
equiv), 1:1 t-BuOH/H2O, 40 °C, 30 min to 1 h, 77-92%; (R ) Ac, Bz) (ii) NaOCH3, CH3OH, room temperature, 15 min to 2 h, quantitative.
toward the tumor-associated CA isozyme IX in vitro.24,25 In this
earlier study the 1,3-dipolar cycloaddition reaction (1,3-DCR;
“click chemistry”)28,29 was employed to generate 1,4-disubsti-
tuted 1,2,3-triazole glycoconjugate sulfonamides from azido
sugars and alkyne-substituted aromatic sulfonamides. These
compounds consisted of a benzene sulfonamide (for CA
recognition) linked via an ester or amide moiety to a triazole-
sugar tail of the structure type A-D (here the sugar is derived
from â-D-glucose).24,25 The results from this previous study
demonstrated that both high affinity and isozyme-selective
ligands could be generated using this strategy, and this has
encouraged us to continue and extend this work. Herein we
explore 1,3-DCR with a reversed “click-tailing” strategy to
synthesize a new generation of glycoconjugate sulfonamides
by appending O-propynyl glycosides onto the 4-azidobenzene-
sulfonamide CA recognition scaffold (1). This library of novel
glycoconjugate sulfonamides, now with triazole-O-glycoside tail
groups, was investigated for their in vitro inhibition of hCA I,
II, and tumor-associated hCA IX.
Results and Discussion
Chemistry. A library of 10 benzene sulfonamides (7-16)
containing triazole-tethered O-glycoside tails was synthesized
by Cu(I)-catalyzed 1,3-DCR of the azido scaffold 1 with a panel
of acylated O-propynyl glycosides (2-6) (Figure 1, Scheme
1).24,25,30 The members of this glycoconjugate library represent
a novel structural class with no reported examples of 1,4-
disubstituted 1,2,3-triazoles wherein the triazole N-1 substituent
was a benzenesulfonamide. Instead of the more chemically
robust N-triazole-linked carbohydrate as in A-D,30,31 the
position of the linking 1,2,3-triazole is reversed in this new
library, leading to O-linked glycosides. The sugar panel
encompassed monosaccharide derivativesspyranoses glucose
Figure 1. Acylated O-propynyl glycoside panel: 2-6.

Carbonic Anhydrase Inhibitors
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 7 1653
Table 1. Inhibition and Selectivity Ratio Data for 1, New
Glycoconjugate Sulfonamides 7-16, Alcohol 17, and Standard
Inhibitors against Human Isozymes hCA I, II, and IX
Ki (nM)a
selectivity ratiosb
Ki(hCA I)/
(hCA IX)
Ki(hCA II)/
(hCA IX)
compd
hCA Ic
hCA IIc
hCA IXd
AZA
MZA
EZA
BRZ
DCP
IND
1
7
8
9
10
11
12
13
14
15
16
17
900
780
25
15
1200
31
3900
1500
6.9
5800
5200
8.1
7.0
4700
8.5
12
14
8
25
27
34
42
50
36
0.48
0.52
0.24
0.21
0.76
0.63
0.45
0.43
0.70
0.13
0.09
0.02
0.09
0.44
0.07
0.06
0.57
0.36
28.9
0.74
0.36
24
Figure 2. Sulfonamide product (17) from potential glycosidase enzyme
action on the O-glycosides 12, 13, and 16.
9
38
15
47
46
6.8
9.3
7.6
7.8
8.7
53
6.9
4.2
48
48
24
1.30
37.1
14.0
0.71
80.6
61.9
0.02
0.07
39.2
0.08
63.8
0.09
46.3
(2), galactose (3), and arabinose (4), and a furanose, ribose (5)s
as well as a disaccharide derivative, maltose (6) (Figure 1).
105
107
9.7
72
Scaffold 1 was synthesized from commercially available
sulfanilamide by diazo transfer under neutral conditions.32 The
peracylated â-O-propynyl glycosides 2, 3, 5, and 6 were
generated by Lewis acid catalyzed (BF3‚Et2O or SnCl4) glyco-
sylation of the â-acetate precursors.33 The O-propynyl arabi-
noside 4 was prepared as an R/â anomeric mixture using
Knoenigs-Knorr conditions (AgOTf) from 2,3,4-tri-O-acetyl-
â-D-arabinosyl bromide. The O-propynyl glycoside panel 2-6
were each reacted via Cu(I)-catalyzed 1,3-DCR with the azido
arylsulfonamide scaffold 1 to generate glycoconjugate sulfona-
mides 7-11 (Scheme 1). Consistent with our earlier findings,
the triazole-forming reaction proceeded smoothly when using
a catalyst loading of 10 mol % of the Cu(I) source and 20 mol
% of ascorbate.24,25,30 Reactions were generally complete
following 1 h of vigorous stirring (as evidenced by TLC). De-
O-acylation of 7-11 using methanolic sodium methoxide was
employed to liberate the corresponding fully deprotected hy-
droxy sugar analogues 12-16 (Scheme 1).
We have included a panel of O-glycosides with and without
corresponding endogenous glycosidases, such that this second
generation of glycoconjugate sulfonamides may have the
potential to act as prodrugs for the sulfonamide pharmacophore
in the in vivo environment. In recognition that our O-glycoside
library may exhibit biological activity dependent not only on
CA isozyme expression but also on endogenous glycosidase
expression, we have also prepared the “clipped” triazole
sulfonamide, bearing a 4-CH2OH alcohol substituent (17)
(Figure 2). Compound 17 is the expected product from the action
of glycosidases on O-glycosides 12, 13, and 16. This compound
was synthesized similarly to 7-11, by the 1,3-DCR of 1 with
propargyl alcohol. There is ample evidence that O-glycosides
have shown promise both as prodrugs (unmasked in vivo by
glycosidases) and for drug targeting (localizing at sites of lectin
receptors).34-36 Several complex O-glycosides of natural origin
act as potent antitumor agents and antibiotics and include the
clinically used anthracyclines and bleomycins. Compelling
spectroscopic and crystallographic evidence suggests the glycone
moieties of these drugs to be essential structural motifs for the
underlying DNA-binding and/or cleavage mechanisms.36 All
sulfonamide compounds (1, 7-17) were investigated for their
in vitro inhibition of hCA I, II, and IX.
84
345
101
120
103
69
4400
7.3
6200
84
134
a Errors in the range of (5-10% of the reported value, from three
determinations. b The Ki ratios are indicative of isozyme selectivity. c Human
(cloned) isozymes, by the CO2 hydration method (refs 37-41). d Catalytic
domain of human (cloned) isozyme, by the CO2 hydration method (refs
37-41).
The parent azido scaffold 1 had greatest efficacy against hCA
II (Ki of 47 nM), approximately 2-fold weaker inhibition against
the tumor-associated hCA IX (105 nM), and 80-fold weaker
inhibition against hCA I (3900 nM). The inhibition profile for
1 was weaker at all isozymes compared to that of the standard
sulfonamide inhibitors; however, it did have a similar selectivity
profile to AZA, MZA, and DCP. Compound 17, the potential
glycosidase product, also had greatest efficacy against hCA II
(Ki of 48 nM), 2.8-fold weaker inhibition against hCA IX (134
nM), and 129-fold weaker inhibition against hCA I (6200 nM).
Isozyme hCA I. At hCA I the O-glycoside moiety had a
variable influence on enzyme inhibition characteristics. The 10
glycoconjugates were investigated both in the acylated form
(sugar-OR compounds 7-11) and the free hydroxyl form
(sugar-OH compounds 12-16). There were some glycocon-
jugates (9, 10, 13, and 15) that were slightly weaker inhibitors
than the parent azido compound 1 with Ki’s ranging from 4400-
5800 nM, while the acetylated glucose compound 7 was a
marginally better inhibitor than 1 (Ki ) 1500 nM). The
remaining compounds 8, 11, 12, 14, and 16 exhibited vastly
improved inhibition at this isozyme. With Ki’s in the low-
nanomolar range (6.9-8.5 nM) these compounds were 481- to
565-fold better inhibitors than their azido parent 1. The inhibition
constants for the furanoside derivatives 10 (ribose-OR) and
15 (ribose-OH), and disaccharide derivatives 11 (maltose-
OR) and 16 (maltose-OH), were independent of the hydroxyl
status of the sugar moiety, with the ribose tail providing
micromolar inhibitors and the maltose tail providing low-
nanomolar inhibitors. This relationship did not extend to the
monosaccharide sugar derivatives (glucose, galactose, and
arabinose); here, one of the pair (sugar-OR or sugar-OH) was
a nanomolar inhibitor (12, 8, 14) and the other a micromolar
inhibitor (7, 13, 9). The dependence of inhibition characteristics
on the hydroxyl status (sugar-OR or sugar-OH) has important
implications for future applications of these compounds, as in
vivo they may be exposed to esterases.43 This structural attribute
could in future prove useful for manipulation to suit drug
delivery or drug bioavailability demands. Compound 17 was
the weakest inhibitor of all sulfonamides evaluated at hCA I.
Carbonic Anhydrase Inhibition. hCA I, II, and IX enzyme
inhibition data for azide 1, the 10 new glycoconjugate sulfona-
mides 7-16, and the alcohol 17 were determined by assaying
the CA-catalyzed hydration of CO2, Table 1.37 The selectivity
ratios for inhibition of isozyme IX compared to those of I and
II are also provided in Table 1. Reference data for clinically
used CA inhibitors acetazolamide (AZA), methazolamide
(MZA), ethoxzolamide (EZA), dichlorophenamide (DCP), and
brinzolamide (BRZ) as well as indisulam (INDsan antitumor
sulfonamide in phase II clinical trials) have also been included
for comparison with the compounds reported in this study.

1654 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 7
Wilkinson et al.
Isozyme hCA II. At hCA II the parent azido scaffold 1 had
a Ki of 47 nM. Seven of the O-glycosides (compounds 8-12,
14, 15) were potent inhibitors of hCA II (Ki’s ranged from 4.2
to 9.3 nM), while compounds 7, 13, and 16 had Ki values similar
to the azido parent 1 (Ki’s ranged from 46-53 nM). Some of
the compounds (8, 11, 12, 14) exhibited similar inhibition at
both hCA I and II, while others (7, 9, 10, 13, and 15) were
selective for hCA II. For example, compound 15, the ribose-
OH derivative, was more than 1000-fold selective for hCA II.
With regard to selectivity there was one notable exception, the
maltose-OH derivative 16, which was 6.5-fold selective for
hCA I and also the only hCA I-selective compound identified
in this study. It is apparent that the O-glycoside tail groups can
strongly influence the selectivity between the physiologically
dominant hCA I and II isozymes, demonstrating that the
structural diversity of even simple sugars permits noteworthy
discrimination of subtle differences in the active site topology
of these isozymes. Compound 17 exhibited hCA II inhibition
similar to azido scaffold 1 and glycoconjugates 7, 13, and 16.
Isozyme hCA IX. At hCA IX the parent azido scaffold 1
had a Ki of 105 nM, while the alcohol-triazole 17 had a Ki of
134 nM. At this isozyme the O-glycoside tails exhibited a
relatively clustered pattern of inhibition in contrast to isozymes
I and II, with eight of the glycoconjugates exhibiting Ki’s in
the range of 69-120 nM. The two outliers were the maltose-
OAc derivative 11 with a much weaker Ki (345 nM) and the
galactose-OAc derivative 8 with a more potent Ki of 9.7 nM
(∼11-fold more potent than 1), respectively. As disaccharide
11 had the bulkiest tail group of those investigated this result
may be a qualitative indicator that the steric boundaries within
the CA IX active site are being encroached upon. Significantly,
compound 8 is more potent than any of the standard sulfona-
midesssome of which are utilized in physiological studies of
this isozyme and none of which are CA IX selective. In terms
of SAR the potency of 8 is quite remarkable given that 7 (Ki of
107 nM) and 8 are epimer derivatives with a single structural
difference far removed from the sulfonamide CA zinc-binding
moiety. It may be drawn from these results that subtle
differences in the O-glycoside tails can deliver compounds
capable of differentially inhibiting CA isozymes. With the
exception of 8 the new sulfonamides were weaker inhibitors at
CA IX compared to CA II.
to be facile, mild, high yielding, and ideally give a predictable
stereochemical outcome. We have shown that the click chem-
istry 1,3-DCR fulfils these demands. An alternative synthetic
route to the library presented herein is the direct glycosylation
of the alcohol 17 with 1-O-acetate donors. In our hands this is
a slower, lower-yielding reaction, which necessitates a more
difficult purification when compared to the 1,3-DCR strategy.
It is also apparent from both the current and our earlier study
that the orientation of the 1,2,3-triazole link between the
carbohydrate tail group and the benzene sulfonamide head group
strongly influences CA isozyme selectivity profiles, and further
manipulation of this structural arrangement will be reported in
due course. Finally, an alternative approach to generate gly-
cosidase resistant S- and C-glycosidic triazole linkages is
currently underway within our group.
Experimental Section
Chemistry. Reagents were purchased from the Sigma-Aldrich
chemical company and were used without further purification.
Solvents were dried and distilled where necessary prior to use or
purchased anhydrous from Sigma-Aldrich. Reactions were moni-
tored by TLC using Merck F60254 silica plates with visualization
of product bands by UV fluorescence (λ ) 254 nm) and charring
by 10% v/v ethanolic H2SO4. Flash chromatography was performed
on Merck flash silica gel (0.04-0.06 mm). Melting points were
acquired on a Gallenkamp melting point apparatus and are reported
as uncorrected. NMR (1H and 13C {1H}) spectra were recorded on
a Varian Unity 400 MHz spectrometer at room temperature using
DMSO-d6 solvent unless otherwise stated. Chemical shifts are
reported in δ (ppm) from a TMS internal standard (0.0 ppm).
Coupling constants (J) are reported in hertz. High-resolution
electrospray ionization mass spectra were acquired in negative ion
mode on an Apex III Bruker Daltonics 4.7T Fourier transform mass
spectrometer (FTMS) fitted with an Apollo ESI source.
Synthesis of Sulfonamide Glycosides (7-11). General Proce-
dure A: A mixture of the azide 1 (1.0 equiv) and the appropriate
O-propynyl glycoside 2-6 (1.0 equiv) was suspended in a tert-
butyl alcohol and water mixture (1:1, 0.2-0.5 M final concentra-
tion). A solution of sodium ascorbate (0.2 equiv) in water, followed
by a solution CuSO4·5H2O (0.1 equiv) in water, was added. The
bright yellow suspension was stirred vigorously at 40 °C until TLC
indicated reaction completion (generally within 2 h). The mixture
was evaporated under reduced pressure, and the resulting residue
was purified by flash chromatography to yield pure material.
Preparation of Deprotected Sulfonamide Glycosides (12-16).
General Procedure B: Compounds 12-16 were prepared by the
treating the corresponding acylated precursors 7-11 (final con-
centration of ∼0.1-0.2 M; ∼0.5 M concentration for compound
15) with dry methanolic sodium methoxide (final pH 9-12).
Reactions were found to be complete within 30 min (for O-
acetylates of 12-14, 16) or 2 h (for O-benzoates of 15) as evidenced
by TLC. Neutralization of the solution by Amberlite IR-120 ion-
exchange resin, followed by filtration and evaporation of the filtrate
to dryness, afforded pure material by NMR.
4-Azidobenzenesulfonamide (1). The preparation of the title
compound was adapted from a literature procedure.32 A slurry of
NaN3 (2.3 g, 34.9 mmol, 3.0 equiv) in distilled H2O (2 mL) was
prepared followed by the addition of tert-butyl alcohol (16 mL).
Sulfanilamide (2.0 g, 11.6 mmol) was then added followed by the
dropwise addition of tert-butyl nitrite (16 mL). The deep yellow
solution was stirred at room temperature overnight and was found
complete by TLC (1:1 EtOAc/hexanes). The crude mixture was
diluted with EtOAc (50 mL) and washed with distilled water (50
mL). The aqueous layer was back-extracted with EtOAc (3 × 40
mL). The combined organic extracts were washed with brine (100
mL), dried (MgSO4), filtered, and evaporated to afford a crude oil.
Precipitation from 3:7 ethyl acetate/hexanes (100 mL) and washing
with cold hexanes afforded the title compound as a pale yellow
crystalline solid (1.7 g, 74%); mp 117-118 °C (decomp; lit mp
Conclusions
The delivery of CA-based cancer therapies or diagnostics will
benefit enormously from the development of isozyme-selective
inhibitors for the tumor-associated CA isozymes. Selective
inhibition among CA isozymes is challenging owing to con-
servation of active site topology within this enzyme family. With
the use of a reversed “click-tailing” strategy, we have appended
a structurally diverse panel of O-glycosides onto the ArSO2-
NH2 CA pharmacophore. This work represents a new class of
glycoconjugate CA inhibitors comprising triazole-tethered O-
glycoside tails. We were able to demonstrate that the glyco-
conjugates could differentiate the active sites of the widespread
isozymes (CA I and II) from the tumor-associated isozyme (CA
IX). A very potent hCA IX inhibitor, the galactose-OAc
derivative 8 (Ki ) 9.7 nM), was identified, demonstrating that
sugar tails may underpin a means to generate CA inhibitors for
applications in cancer drug development and/or diagnostics. It
is also feasible that glycoconjugates could be designed to target
endogenous glycosidases in vivo for the purpose of a prodrug
therapy. From a synthetic viewpoint the reactions used to
generate molecular diversity in carbohydrate-based libraries need

Carbonic Anhydrase Inhibitors
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 7 1655
119-122 °C);31 1H NMR (400 MHz, DMSO-d6) δ 7.26-7.81 (m,
4H, Ph), 7.34 (br s, 2H, SO2NH2); 13C {1H} NMR (100 MHz,
DMSO-d6) δ 120.16 (Ph CH), 128.30 (Ph CH), 141.20 (Ph C),
143.60 (Ph C).
CH), 128.21 (Ph CH), 139.21 (Ph C), 145.56 (triazole C or Ph C),
145.13 (triazole C or Ph C), 169.82 (CdO), 170.23 (CdO), 170.51
(CdO). HRMS (ESI) calcd for C20H23N4O10S-: 511.114249.
Found: 511.113207. Anal. (C20H24N4O10S‚0.5 H2O) C, H, N.
4-(4-{[2′,3′,5′-Tri-O-benzoyl-â-D-ribofuranosyl)oxymethyl}-
1-H-1,2,3-triazol-1-yl)benzenesulfonamide (10). The title com-
pound was prepared from 5 according to general procedure A and
isolated as a pale yellow foam following flash chromatography (1:1
EtOAc/hexanes; 675 mg, 0.97 mmol, 91%); Rf 0.53 (2:3 hexanes/
EtOAc); 1H NMR (400 MHz, DMSO-d6) δ 4.51 (dd, 2J5′-5′′ ) 12.0
4-(4-{[2′,3′,4′,6′-Tetra-O-acetyl-â-D-glucopyranosyl)oxymethyl}-
1-H-1,2,3-triazol-1-yl)benzenesulfonamide (7). The title com-
pound was prepared from 2 according to general procedure A and
isolated as a white foam following purification by flash chroma-
tography in 3:7 hexanes/EtOAc (175 mg, 0.30 mmol, 77%); Rf 0.63
1
(100% EtOAc); H NMR (400 MHz, DMSO-d6) δ 1.87 (s, 3H,
Hz, 3J5-4 ) 5.2 Hz, 1H, H5′), 4.66 (dd, 2J5′′-5′ ) 12.0 Hz, 3J5′′-4′
)
)
OAc CH3), 1.90 (s, 3H, OAc CH3), 1.96 (s, 3H, OAc CH3), 1.99
2
3
3
4.0 Hz, 1H, H5′′), 4.78-4.82 (m, 1H, H4′), 4.81 (AB q, JAB
(s, 3H, OAc CH3), 3.99-4.02 (ddd, J5′-4′ ) 8.8 Hz, J5′-6′′
)
3
3
2
12.4 Hz, 2H, OCH2), 5.52 (s, 1H, H1), 5.60 (d, J2′-3′ ) 4.8 Hz,
1H, H2), 5.77 (d, 3J3′-4′ ) 6.8 Hz, 3J3′-2′ ) 4.8 Hz, 1H, H3), 7.34-
8.07 (m, 21 H, Ph CH and SO2NH2), 8.84 (s, 1H, triazole CH);
13C {1H} NMR (100 MHz, DMSO-d6) δ 61.05 (OCH2), 65.06 (C5′),
72.59 (C3′), 75.68 (C2′), 79.17 (C4′), 104.67 (C1′), 120.93 (Ph CH),
123.35 (triazole CH), 128.14, 129.26, 129.31, 129.35, 129.55,
129.87, 129.90, 130.04 (Ph CH), 134.06, 134.47, 134.62, 139.21,
144.48 (Ph C), 145.21 (triazole C), 166.11 (CdO), 165.49
(CdO), 165.32 (CdO). HRMS (ESI) calcd for C35H29N4O10S-:
699.17539. Found: 699.174859. Anal. (C35H30N4O10S‚H2O) C, H,
N, S.
5.6 Hz, J5′-6′ ) 2.4 Hz, 1H, H5′), 4.05 (dd, J6′-6′′ ) 12.8 Hz,
3J6′-5′ ) 2.0 Hz, 1H, H6′), 4.19 (dd, J6′′-6′ ) Hz, 12.4 Hz, J6′′-5′
2
3
3
3
) 5.2 Hz, 1H, H6′′), 4.77 (dd, J2′-1′ ) 8.0 Hz, J2′-3′ ) 9.6 Hz,
1H, H2′), 4.80 (AB q, 2JAB ) 12.4 Hz, 2H, OCH2), 4.88-4.93 (m,
3
1H, H4′), 4.94 (d, J1′-2′ ) 8.0 Hz, 1H, H1′), 5.22-5.27 (m, 1H,
H3′), 7.50 (br s, 2H, SO2NH2), 7.99-8.11 (m, 4H, Ph), 8.87 (s,
1H, triazole CH); 13C {1H} NMR (100 MHz, DMSO-d6) δ 20.94
(OAc CH3), 21.01 (OAc CH3), 21.06 (OAc CH3), 21.18 (OAc CH3),
62.24 (C6′), 62.36 (OCH2), 68.79 (C4′), 71.36 (C5′), 71.53 (C2′),
72.73 (C3′), 99.21 (C1′), 121.02 (Ph CH), 123.72 (triazole CH),
128.20 (Ph CH), 139.22 (Ph C), 144.58 (triazole C or Ph C), 145.02
(triazole C or Ph C), 169.69 (CdO), 169.53 (CdO), 170.21
(CdO), 170.75 (CdO); HRMS (ESI) calcd for C23H27N4O12S-:
583.135392. Found: 583.134630. Anal. (C23H28N4O12S‚H2O) H,
N, S; C calcd, 45.84; found, 46.54.
4-(4-{[Hepta-O-acetyl-â-D-maltopyranosyl)oxymethyl}-1-H-
1,2,3-triazol-1-yl)benzenesulfonamide (11). The title compound
was prepared from 6 according to general procedure A and isolated
as a pale yellow foam after flash chromatography in 1:1 EtOAc/
hexanes (166 mg, 0.19 mmol, 86%); Rf 0.22 (1:1 EtOAc/hexanes);
1H NMR (400 MHz, DMSO-d6) δ 1.86 (s, 3H, OAc CH3), 1.90 (s,
3H, OAc CH3), 1.92 (s, 3H, OAc CH3), 1.94 (s, 3H, OAc CH3),
1.96 (s, 3H, OAc CH3), 1.99 (s, 3H, OAc CH3), 2.05 (s, 3H, OAc
CH3), 3.93-4.03 (m, 4H, Glcâ H4′, GlcR H5′, Glcâ H5′, GlcR or
Glcâ H6′′), 4.14 (dd, 2J6′-6′′ ) 12.0 Hz, 3J6′-5′ ) 4.4 Hz, 1H, GlcR
4-(4-{[2′,3′,4′,6′-Tetra-O-acetyl-â-D-galactopyranosyl)-
oxymethyl}-1-H-1,2,3-triazol-1-yl)benzenesulfonamide (8). The
title compound was prepared from 3 according to general procedure
A and isolated as a pale yellow foam following purification by
flash chromatography in 3:7 hexanes/EtOAc (263 mg, 0.45 mmol,
87%); Rf 0.23 (1:4 hexanes/EtOAc); 1H NMR (400 MHz, DMSO-
d6) δ 1.87 (s, 3H, OAc CH3), 1.91 (s, 3H, OAc CH3), 1.98 (s, 3H,
OAc CH3), 2.09 (s, 3H, OAc CH3), 4.01-4.10 (m, 2H, H6, H6′),
4.20-4.23 (m, 1H, H5), 4.80 (AB q, 2JAB ) 12.4 Hz, 2H, OCH2),
2
3
or Glcâ H6′), 4.18 (dd, J6′-6′′ ) 12.4 Hz, J6′-5′ ) 4.8 Hz, 1H,
2
3
GlcR or Glcâ H6′′), 4.39 (dd, J6′′-6′ ) 12.0 Hz, J6′′-5′ ) 2.4 Hz,
1H, GlcR or Glcâ H6′′), 4.67 (dd, 3J2′-3′ ) 9.6 Hz, 3J2′-1′ ) 8.0 Hz,
1H, Glcâ H2′), 4.78 (AB q, 2JAB ) 12.8 Hz, 2H, OCH2), 4.84 (dd,
3
3
4.84 (d, J1′-2′ ) 8.0 Hz, 1H, H1′), 4.93 (dd, J2′-3′ ) 10.0 Hz,
3J2′-1′ ) 8.0 Hz, 1H, H2′), 5.14 (dd, 3J3′-2′ ) 10.0 Hz, 3J3′-4′ ) 3.6
Hz, 1H, H3′), 5.23-4.25 (m, 1H, H4′), 7.49 (br s, 2H, SO2NH2),
7.98-8.11 (m, 4H, Ph), 8.88 (s, 1H, triazole CH); 13C {1H} NMR:
δ 21.00 (OAc CH3), 21.07 (OAc CH3), 21.11 (OAc CH3), 21.18
(OAc CH3), 61.96 (OCH2), 62.18 (C6′), 67.99 (C4′), 69.26 (C3′),
70.69 (C2′), 70.88 (C5′), 99.71 (C1′), 121.00 (Ph CH), 123.65
(triazole CH), 128.22 (Ph C), 139.20 (Ph C), 144.60 (triazole C or
Ph C), 145.09 (triazole C or Ph C), 169.79 (OAc CdO), 170.15
(OAc CdO), 170.60 (OAc CdO), 170.62 (OAc CdO). HRMS
(ESI) calcd for C23H27N4O12S-: 582.135392. Found: 583.134423.
Anal. (C23H28N4O12S) C, H, N, S.
3J2′-3′ ) 9.6 Hz, 3J2′-1′ ) 3.6 Hz, 1H, GlcR H2′), 4.91 (d, 3J1′-2′
)
8.0 Hz, 1H, Glcâ H1′), 4.93-4.98 (m, 1H, GlcR H4′), 5.17-5.22
(m, 1H, GlcR H3′), 5.25 (d, 3J1′-2′ ) 3.6 Hz, 1H, GlcR H1′), 5.25-
5.30 (m, 1H, Glcâ H3′), 7.50 (br s, 2H, SO2NH2), 7.99-8.11 (m,
4H, Ph CH), 8.85 (s, 1H, triazole CH); 13C {1H} NMR (100 MHz,
DMSO-d6) δ 21.95 (OAc CH3), 21.02 (2 × OAc CH3), 21.11 (OAc
CH3), 21.21 (OAc CH3), 21.26 (OAc CH3), 21.44 (OAc CH3), 60.43
(Glc C6′), 62.07 (Glc C6′), 63.54 (OCH2), 68.42 (Glc C4′), 68.68
(Glc C4′), 69.53 (Glc C3′), 70.11 (Glc C3′), 72.02 (Glc C2′), 72.20
(Glc C2′), 74.30 (Glc C5′), 74.99 (Glc C5′), 96.05 (Glcâ H1′), 98.75
(GlcR H1′), 121.03 (Ph CH), 123.73 (triazole CH), 128.19 (Ph CH),
139.22 (Ph C), 144.57 (triazole C or Ph C), 144.99 (triazole C or
Ph C), 169.83 (OAc CdO), 169.88 (OAc CdO), 170.27 (OAc Cd
O), 170.54 (OAc CdO), 170.69 (OAc CdO), 170.85 (OAc Cd
O). HRMS (ESI) calcd for C35H43N4O20S-: 871.219684. Found:
871.220422. Anal. (C35H44N4O20S‚H2O) C, H, N, S.
4-(4-{[2′,3′,4′-Tri-O-acetyl-r/â-D-arabinopyranosyl)oxymethyl}-
1-H-1,2,3-triazol-1-yl)benzenesulfonamide (9). The title com-
pound was prepared from 4 according to general procedure A and
isolated as a white foam following flash chromatography in 1:4
hexanes/EtOAc (as a 2:1 R/â mixture by 1H NMR) (145 mg, 0.28
1
4-(4-{[â-D-Glucopyranosyl)oxymethyl}-1-H-1,2,3-triazol-
1-yl)benzenesulfonamide (12). The title compound was prepared
from 7 according to general procedure B and isolated as a white
solid (107 mg, 0.26 mmol, ∼100%); Rf 0.12 (1:9 H2O/CH3CN);
mmol, 89%); Rf 0.30 (1:4 hexanes/EtOAc); H NMR (400 MHz,
DMSO-d6) δ 1.90 (s, 3H, ROAc CH3), 1.91 (s, 6H, R and âOAc
CH3), 1.93 (s, 3H, âOAc CH3), 2.07 (s, 3H, ROAc CH3), 2.08 (s,
3H, âOAc CH3), 3.67 (dd, 2J5′-5′′ ) 13.2 Hz, 3J5′-4′ ) 2.0 Hz, 1H,
1
2
3
mp 244-246 °C (decomp); H NMR (400 MHz, 10% DMSO-d6
âH5′), 3.83 (dd, J5′-5′′ ) 13.2 Hz, J5′-4′ ) 2.0 Hz, 1H, RH5′),
3.90 (dd, 2J5′′-5′ ) 13.2 Hz, 3J5′′-4′ ) 2.4 Hz, RH5′′), 4.02 (dd, 2J5′′-5′
in D2O) δ 3.16-3.20 (m, 1H, H2′), 3.23-3.28 (m, 1H, H4′), 3.33-
2
3
3
3
3.38 (m, 2H, H3′, H5′), 3.58 (dd, J6′-6′′ ) 12.8 Hz, J6′-5′ ) 5.6
) 13.2 Hz, J5′′-4′ ) 2.4 Hz, 1H, âH5′′), 4.72 (d, J1′-2′ ) 8.0 Hz,
2
3
1H, RH1′), 4.73 (d, 3J1′-2′ ) 2.8 Hz, 1H, âH1′), 4.74 (AB q, 2JAB
12.8 Hz, 2H, âOCH2), 4.78 (AB q, 2JAB ) 12.8 Hz, 2H, ROCH2),
)
Hz, 1H, H6′), 3.78 (dd, J6′′-6′ ) 12.4 Hz, J6′′-5′ ) 2.0 Hz, 1H,
3
2
H6′′), 4.47 (d, J1′-2′ ) 8.0 Hz, 1H, H1′), 4.89 (AB q, JAB ) 12.8
Hz, 2H, OCH2), 7.86-7.99 (m, 4H, Ph), 8.49 (s, 1H, triazole CH);
13C {1H} NMR (100 MHz, 10% DMSO-d6 in D2O) δ 60.96 (C6′),
62.09 (OCH2), 69.84 (C4′), 73.23 (C3′), 75.92 (C2′), 76.21 (C5′),
101.84 (C1′), 121.75 (Ph CH), 124.00 (triazole CH), 128.08 (Ph
CH), 139.54 (Ph C), 142.24 (triazole C or Ph C), 144.922 (triazole
C or Ph C). HRMS (ESI) calcd for C15H19N4O8S-: 415.02908.
Found: 415.093029.
3
3
4.93 (dd, J2′-3′ ) 10.0 Hz, J2′-1′ ) 8.0 Hz, 1H, RH2′), 4.97 (dd,
3J2′-3′ ) 10.4 Hz, J2′-1′ ) 3.2 Hz, 1H, âH2′), 5.08 (dd, J3′-2′
)
3
3
9.6 Hz, 3J3′-4′ ) 3.2 Hz, 1H, âH3′), 5.12-5.14 (m, 1H, RH4′), 5.18
(dd, 3J3′-2′ ) 11.2 Hz, 3J3′-4′ ) 3.6 Hz, 1H, RH3′), 5.23-5.25 (m,
1H, âH4′), 7.49 (br s, 2H, SO2NH2), 7.98-8.11 (m, 4H, Ph CH),
8.89 (s, 1H, R triazole CH), 8.96 (s, 1H, â triazole CH); 13C {1H}
NMR (100 MHz, DMSO-d6) δ 21.07 (OAc CH3), 21.13 (OAc CH3),
21.37 (OAc CH3), 61.84 (OCH2), 63.99 (C5′), 68.60 (C4′), 69.51
(C2′), 70.63 (C3′), 99.87 (C1′), 120.99 (Ph CH), 123.56 (triazole
4-(4-{[â-D-Galactopyranosyl)oxymethyl}-1-H-1,2,3-triazol-
1-yl)benzenesulfonamide (13). The title compound was prepared

1656 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 7
Wilkinson et al.
from 8 according to general procedure B and isolated as a white
solid (150 mg, 0.36 mmol, ∼100%); Rf 0.13 (1:9 H2O/CH3CN);
mp 241-244 °C (decomp); 1H NMR (400 MHz, D2O) δ 3.42 (dd,
3J2′-3′ ) 9.6 Hz, 3J2′-1′ ) 7.6 Hz, 1H, H2′), 3.50 (dd, 3J3′-2′ ) 10.0
CA Inhibition Assay. An Applied Photophysics stopped-flow
instrument has been used for assaying the CA-catalyzed CO2
hydration activity.37 Phenol red (at a concentration of 0.2 mM) has
been used as indicator, working at the absorbance maximum of
557 nm, with 10 mM Hepes (pH 7.5) as buffer, 0.1 M Na2SO4 (for
maintaining constant the ionic strength), at 25 °C, following the
CA-catalyzed CO2 hydration reaction for a period of 10-100 s
(the uncatalyzed reaction needs around 60-100 s in the assay
conditions, whereas the catalyzed ones are of around 6-10 s). The
CO2 concentrations ranged from 1.7 to 17 mM for the determination
of kinetic parameters. For each inhibitor, tested in the concentration
range between 0.01 nM and 100 µM, at least six traces of the initial
5-10% of the reaction have been used for determining the initial
velocity. The uncatalyzed rates were determined in the same manner
and subtracted from the total observed rates. Stock solutions of
inhibitor (1 mM) were prepared in distilled-deionized water with
10-20% (v/v) DMSO (which is not inhibitory at these concentra-
tions), and dilutions up to 0.01 nM were done thereafter with
distilled-deionized water. Inhibitor and enzyme solutions were
preincubated together for 15 min at room temperature prior to assay,
in order to allow for the formation of the E-I complex. The
inhibition constants were obtained by nonlinear least-squares
methods using PRISM 3. The curve-fitting algorithm allowed us
to obtain the IC50 values, working at the lowest concentration of
substrate of 1.7 mM, from which Ki values were calculated by using
the Cheng-Prusoff equation.38-42 The catalytic activity (in the
absence of inhibitors) of these enzymes was calculated from
Lineweaver-Burk plots, as reported earlier, and represent the mean
from at least three different determinations.38-42 Enzyme concentra-
tions in the assay system were 9.2 nM for hCA I, 7.3 nM for hCA
II, and 8.5 nM for hCA IX. Enzymes used here were recombinant
ones, prepared and purified as described earlier.38-42,44
3
Hz, J3′-4′ ) 3.6 Hz, 1H, H3′), 3.56-3.69 (m, 3H, H5′, H6′, H6′′),
3
3.78-3.79 (m, 1H, H4′), 4.41 (d, J1′-2′ ) 7.6 Hz, 1H, H1′), 4.91
(AB q, 2JAB ) 12.4 Hz, 2H, OCH2), 4.89-8.01 (m, 4H, Ph), 8.53
(s, 1H, triazole CH); 13C {1H} NMR (100 MHz, DMSO-d6) δ 61.19
(C6′), 62.01 (OCH2), 68.79 (C4′), 71.10 (C3′), 73.89 (C2′), 76.0 (C5′),
103.59 (C1′), 120.92 (Ph CH), 123.50 (triazole CH), 128.25 (Ph
CH), 139.29 (Ph C), 144.44 (triazole C or Ph C), 146.211 (triazole
C or Ph C). HRMS (ESI) calcd for C15H19N4O8S-: 415.092908.
Found: 415.093072.
4-(4-{[r/â-D-Arabinopyranosyl)oxymethyl}-1-H-1,2,3-triazol-
1-yl)benzensulfonamide (14). The title compound was prepared
from 9 according to general procedure B and isolated as a white
1
solid (as a 2:1 R/â mixture by H NMR) (75 mg, 0.19 mmol,
1
∼100%); Rf 0.23 (1:9 CH3OH/EtOAc); H NMR (400 MHz, 2%
D2O in DMSO-d6) δ 3.31-3.47 (m, 4H, RH2′, RH5′, RH5′′, âH5′
and âH5′′), 3.56-3.58 (m, 1H, âH2′), 3.60-3.62 (m, 1H, RH4′),
3
3.67-3.73 (m, 4H, RH3′, RH4′, âH4′), 3.71 (dd, J3′-2′ ) 12.0 Hz,
3
3J3′-4′ ) 3.6 Hz, 1H, âH3′), 4.23 (d, J1′-2′ ) 6.0 Hz, 1H, RH1′),
2
2
4.66 (AB q, JAB ) 12.0 Hz, 1H, âOCH2), 4.75 (AB q, JAB
)
12.8 Hz, 2H, ROCH2), 4.80 (d, 3J1′-2′ ) 2.4 Hz, 1H, âH1′), 7.98-
8.10 (m, 4H, Ph CH), 8.87 (s, 1H, R triazole CH), 8.89 (s, 1H, â
triazole CH); 13C {1H} NMR (100 MHz, DMSO-d6) δ 60.73 (â
OCH2), 61.44 (R OCH2), 63.95, 65.87, 67.99, 68.79, 69.06, 69.49,
71.03, 72.95, (RC2, RC3, RC4′, RC5′ and, âC2′, âC3′, âC4′, âC5′),
99.60 (âC1′), 103.05 (RC1′), 120.93 (âPh CH), 120.65 (RPh CH),
123.18 (R triazole CH), 123.32 (â triazole Ph CH), 128.24 (Ph CH),
139.30 (Ph C), 144.36 (Ph C), 146.05 (R triazole C), 146.15 (â
triazole C). HRMS (ESI) calcd for C14H17N4O7S-: 385.082343.
Found: 385.082537.
Acknowledgment. This work was financed in part by the
Australian Research Council (Grant Nos. F00103312 and
DP0343419), the Eskitis Institute for Cell and Molecular
Therapies and the School of Science (Griffith University,
S.-A.P.), and an EU Grant of the sixth framework programme
(EUROXY project) (University of Florence, C.T.S.). We thank
Hoan The Vu for carrying out accurate mass measurements.
4-(4-{[â-D-Ribofuranosyl)oxymethyl}-1-H-1,2,3-triazol-1-yl)-
benzenesulfonamide (15). The title compound was prepared from
10 according to general procedure B and isolated as a pale yellow
solid following flash chromatography (1:1 EtOAc/hexanes then 1:9;
112 mg, 0.29 mmol, ∼100%); Rf 0.12 (1:9 EtOAc/CH3OH); mp
1
2
146-147 °C. H NMR (400 MHz, DMSO-d6) δ 3.37 (dd, J5′-5′′
) 12.0 Hz, 3J5′-4′ ) 6.4 Hz, 1H, H5′), 3.54 (dd, 2J5′′-5′ ) 11.6 Hz,
3J5′′-4′ ) 3.72, 1H, H5′′), 3.73 (d, 3J2′-3′ ) Hz, 1H, H2′), 3.79 (ddd,
3J4′-3′ ) 10.4 Hz, 3J4′-5′ ) 6.4 Hz, 3J4′-5′′ ) 4.0 Hz, 1H, H4′), 3.87
Supporting Information Available: Elemental analysis data
1
and H NMR spectra for compounds. This material is available
2
free of charge via the Internet at http://pubs.acs.org.
(dd, 1H, H3′), 4.67 (AB q, JAB ) 12.4 Hz, 2H, CH2O), 4.88 (s,
1H, H1′), 7.97-8.11 (m, 4H, Ph), 8.85 (s, 1H, triazole CH); 13C
NMR (100 MHz, DMSO-d6) δ 60.10 (C5′), 63.62 (CH2O), 71.43
(C3′), 74.93 (C2′), 84.43 (C4′), 106.83 (C1′), 120.98 (Ph CH), 123.21
(triazole CH), 128.20 (Ph CH), 139.31 (Ph C), 144.38 (triazole C
or Ph C), 146.04 (triazole C or Ph C). HRMS (ESI) calcd for
C14H18N4O7SNa+: 409.078838. Found: 409.078059.
References
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4-(4-{[â-D-Maltopyranosyl)oxymethyl}-1-H-1,2,3-triazol-
1-yl)benzenesulfonamide (16). The title compound was prepared
from 11 according to general procedure B and isolated as an off-
white solid (99 mg, 0.17 mmol, ∼100%); Rf 0.09 (1:9 H2O/CH3-
CN); mp 192-196 °C (decomp). 1H NMR (400 MHz, D2O) δ 3.22
3
3
(dd, J2′-3′ ) 9.2 Hz, J2′-1′ ) 8.0 Hz, 1H, Glcâ H2′), 3.25-3.30
3
3
(m, 1H, Glcâ H4′), 3.43 (dd, J2′-3′ ) 10.0 Hz, J2′-1′ ) 4.0 Hz,
GlcR H1′), 3.49-3.66 (m, 7H, GlcR H3′, GlcR H4, GlcR H5, GlcR
2
H6 and Glcâ H3, Glcâ H5, Glcâ H6), 3.71 (dd, J6′′-6′ ) 12.0 Hz,
3J6′′-5′ ) 2.0 Hz, 1H, GlcR H6′′ or Glcâ H6′′), 3.80 (dd, J6′′-6′
)
2
3
12.4 Hz, J6′′-5′ ) 1.6 Hz, 1H, GlcR H6′′ or Glcâ H6′′), 4.49 (d,
3J1′-2′ ) 8.0 Hz, 1H, 1H, Glcâ H1′), 4.90 (AB q, JAB ) 12.4 Hz,
2
3
2H, OCH2), 5.26 (d, J1′-2′ ) 3.6 Hz, 1H, GlcR H1′), 7.87-7.99
(m, 4H, Ph), 8.49 (s, 1H, triazole CH); 13C {1H} NMR (100 MHz,
D2O) δ 60.23 (OCH2), 60.87, 62.10, 69.48, 71.79, 72.85, 72.97,
73.07, 74.78, 76.33, 76.81 (GlcR C2′, GlcR C3′, GlcR C4′, GlcR
C5′, GlcR C6′ and Glcâ C2′, Glcâ C3′, Glcâ C4′, Glcâ C5′, Glcâ
C6′), 99.70 (GlcR C1′), 101.59 (Glcâ C1′), 121.75 (Ph CH), 124.06
(triazole CH), 128.02 (Ph CH), 139.58 (Ph C), 141.93 (triazole C
or Ph C), 144.74 (triazole C or Ph C). HRMS (ESI) calcd for
C21H29N4O13S-: 577.145731. Found: 577.145862.

Carbonic Anhydrase Inhibitors
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 7 1657
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