2-Acetamino-1,2-dideoxynojirimycin-lysine hybrids as hexosaminidase inhibitors

Article abstract of DOI:10.1016/j.tetasy.2009.02.015

Cyclisation by double reductive amination of 2-acetamino-2-deoxy-d-xylo-hexos-5-ulose with N-2 protected l-lysine derivatives provided 2-acetamino-1,2-dideoxynojirimycin derivatives without any observable epimer formation at C-5. Modifications on the lysi

Full text of DOI:10.1016/j.tetasy.2009.02.015

Tetrahedron: Asymmetry 20 (2009) 832–835
Contents lists available at ScienceDirect
Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
2-Acetamino-1,2-dideoxynojirimycin—lysine hybrids
as hexosaminidase inhibitors
a
b
Andreas J. Steiner a, Georg Schitter a, Arnold E. Stütz a, , Tanja M. Wrodnigg , Chris A. Tarling ,
*
Stephen G. Withers b, Don J. Mahuran c, Michael B. Tropak c
a Glycogroup, Institut für Organische Chemie, Technische Universität Graz, Stremayrgasse 16, A-8010 Graz, Austria
b Chemistry Department, University of British Columbia, 300-6174 University Boulevard, Vancouver, BC, Canada V6T 1Z3
c Department of Laboratory Medicine and Pathobiology, Sick Kids Hospital, 555 University Avenue, University of Toronto, Ont., Canada M5G 1X8
a r t i c l e i n f o
a b s t r a c t
Article history:
Cyclisation by double reductive amination of 2-acetamino-2-deoxy-D-xylo-hexos-5-ulose with N-2 pro-
tected L-lysine derivatives provided 2-acetamino-1,2-dideoxynojirimycin derivatives without any obser-
vable epimer formation at C-5. Modifications on the lysine moiety gave access to lipophilic derivatives
that exhibited improved hexosaminidase inhibitory activities.
Received 2 February 2009
Accepted 11 February 2009
Available online 4 May 2009
Ó 2009 Elsevier Ltd. All rights reserved.
Dedicated to Professor George W. J. Fleet on
the occasion of his 65th birthday
1. Introduction
HO
HO
HO
HO
HO
HO
NH
NH
NH
N-Acetylhexosaminidases have attracted considerable interest,
due to associations with diseases such as osteoarthritis,1 allergy2
as well as Alzheimer’s disease.3 Catalytically handicapped and mis-
folded mutant forms of N-acetylhexosaminidases are responsible
for hereditary lysosomal disorders.4 Inhibitors of this class of gly-
cosidases have been employed for affinity purification5 and the
characterisation6 of enzymes, as insecticidal as well as anti-fungal7
agents and have recently also been suggested as pharmacological
chaperone therapeutics for Tay-Sachs’ as well as Sandhoff’s dis-
eases.8 In particular, iminoalditols have frequently been employed
as (usually) competitive glycosidase inhibitors.9 Representatives of
this class of compound, for example glucosidase inhibitor 1 (Fig. 1)
as well as mannosidase inhibitor 2 have found important roles as
biological probes, such as in the investigation of glycoprotein trim-
ming glycosidases.10 More recently, hexosaminidase inhibitor 3
was examined as a pharmacological chaperone for lysosomal b-
N-acetylhexosaminidase A/S (Hex A/S) in fibroblasts derived from
patients with Sandhoff and Tay-Sachs diseases.8,11
HO
OH
HO
OH
HO
NHAc
1
2
3
Figure 1.
tioned amine for tagging as well as an additional ‘handle’ for
chain extension and with a view to variation of the N-substituent’s
chemical as well as biochemical properties.14
2. Results and discussion
N2-tert-Butyloxycarbonyl-N6-benzyloxycarbonyl-protected
sine methyl ester A as well as the chain-extended derivative, methyl
6-(N6-benzyloxycarbonyl-N2-tert-butyloxycarbonyl-
-lysi-
nyl)aminohexanoate B proved to be useful reaction partners in ring-
closing reactions with 5-ulohexoses by double reductive amina-
tion15 to provide lysine-iminoalditol hybrids16 ready for a wide
range of interesting follow-up modifications.17
L-ly-
L
Following up on our observation that some fluorescently la-
belled derivatives of the
D-glucosidase inhibitor 2,5-dideoxy-2,5-
imino- -mannitol (DMDP) are powerful inhibitors exceeding the
D
Intermediate ulososide 4 (Scheme 1), which was prepared by
3-chloroperbenzoic acid oxidation of methyl 3,4-di-O-acetyl-2-
parent compound’s activity by two orders of magnitude,12 we have
reported the syntheses and glycosidase inhibitory activities of var-
ious pyranoid N-alkylated iminoalditols featuring fluorescent tags
such as dansyl moieties attached to simple N-substituents.13 Based
on their encouraging inhibitory activities, we envisaged more con-
venient properties with compounds providing a suitably posi-
acetamino-2,6-dideoxy-b-D-xylo-hex-5-enopyranoside (available
by standard methodology18,19) in the presence of benzyl alcohol
following a slightly modified route introduced by Barili20 et al.
and Murphy21 et al. subsequent Zemplen O-deacetylation, was em-
ployed as the sugar component.
Compound 4 turned out to be a very efficient reaction partner
for the amines employed in the catalytic hydrogenation/reductive
amination step. Despite the complexity of this conversion which
* Corresponding author. Tel.: +43 316 873 8744; fax: +43 316 873 8740.
E-mail address: stuetz@tugraz.at (A.E. Stütz).
0957-4166/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2009.02.015

A. J. Steiner et al. / Tetrahedron: Asymmetry 20 (2009) 832–835
833
Table 1
Ki values for inhibition of the b-N-acetylhexosaminidase from Streptomyces plicatus by
2-acetamino-1,2-dideoxynojirimycin-lysine hybrids
O
O
O
AcO
AcO
OMe
AcO
AcO
OMe
Compound
Ki (lM)
NHAc
NHAc
3
5
5a
6
80
5.0
4.3
6.3
4.6
OH
O
OH
O
BnO
AcO
BnO
HO
6a
OMe
OMe
AcO
NHAc
HO
NHAc
3
2
1
0
Adult Tay-Sachs
Infantile Sandhoff
4
Scheme 1.
includes deprotection of both the sugar unit as well as the terminal
lysine amino group followed by reductive amination at C-1 and
subsequent intramolecular cyclisation with formation of a stereo-
genic centre at C-5, isolated yields were fair and the stereoselectiv-
ities for the desired
cases investigated.
Thus, compound 4, with 6-N-benzyloxycarbonyl-2-N-BOC
methyl lysinate A as well as with the chain-extended amide B, gave
iminoalditol derivatives 5 (58%) and 6 (72%), respectively (Scheme
2). Epimer formation at C-5 could not be detected by NMR.
Compounds 5 and 6 were smoothly converted by standard BOC
removal employing methanolic HCl and subsequent reaction of the
free amino group with dansyl chloride to give, after preparative
TLC, fluorescent inhibitors 5a (45%) and 6a (74%), respectively.
D-gluco configuration were excellent in all
10-1
100
101
102
103
Compound 6a / µM
Figure 2. Enhancement of b-N-acetylhexosaminidase activity of adult Tay-Sachs
(squares) and infantile Sandhoff (circles) fibroblast patient cell lines. Relative
increase in intracellular b-N-Acetylhexosaminidase activity in compound 6a versus
vehicle (DMSO) treated cells (y-axis).
structural framework of compound 6a could be exploited for the
design of additional efficacious pharmacological chaperones for
GM2 gangliosidoses with low toxicity.
D
-Hexosaminidase inhibitory activities of new compounds are
summarised in Table 1.
As previously13,17,22 observed, the presence of a lipophilic resi-
due improved binding compared to the unsubstituted parent com-
pound 3. However, in contrast to these previous results, in this
series of compounds the presence of the dansyl group did not re-
sult in tighter binding relative to the N-BOC-substituted precursor.
These compounds, with their appended fluorescent moiety and
reasonably tight binding, offer potential for use in monitoring
chaperoning processes.
3. Experimental
3.1. General methods
Optical rotations were measured on a Perkin Elmer 341 polar-
imeter at the wavelength of 589 nm and a path length of 10 cm
at 20 °C. NMR spectra were recorded on a Varian INOVA 500 oper-
ating at 500.6 MHz (1H), and at 125.9 MHz (13C). CDCl3 was em-
ployed for protected compounds and methanol-d4 employed for
unprotected inhibitors. Chemical shifts are listed in delta employ-
ing residual, non-deuterated solvent as the internal standard. The
signals of the protecting groups were found in the expected regions
and are not listed explicitly. Electrospray mass spectra were re-
corded on an HP 1100 series MSD, Hewlett Packard. Samples were
dissolved in acetonitrile or acetonitrile/water mixtures. The scan
mode for positive ions (mass range 100–1000 D) was employed
varying the fragmentation voltage from 50 to 250 V with best
molecular peaks observed at 150 V. Analytical TLC was performed
For example, compound 6a enhanced the activity of Hex A/S in
adult Tay-Sachs as well as infantile Sandhoff cell lines more than
twofold relative to vehicle (DMSO) treated cells (Fig. 2). Although
Hex A/S activity was previously shown to be enhanced 2.8 fold
by compound 3 (100 mM) in adult Tay-Sachs cells,11 compound
6a was equally effective at increasing HexA/S activity but at a
700-fold lower concentration (140 lM). Like other pharmacologi-
cal chaperones for Tay-Sachs or Gaucher disease, the effective
intracellular enhancing concentration of compound 6a (15–
140 lM) is greater than its Ki (4.2 lM, human Hex A). This differ-
ence is commonly attributed to bioavailability and/or intracellular
metabolism of the compound.23 These results suggest that the
NHR
HO
HO
O
N
A
+
: BOC-L-Lys(Z)-OMe
O
5
R = Boc
OH
O
5a
R = dansyl
HO
NHAc
BnO
HO
OMe
NHR
O
H
N
HO
HO
HO
NHAc
N
(CH2)5COOMe
4
B
+
: BOC-L-Lys(Z)-NH(CH2)5COOMe
HO
NHAc
6
R = Boc
6a
R = dansyl
Scheme 2.

834
A. J. Steiner et al. / Tetrahedron: Asymmetry 20 (2009) 832–835
on precoated aluminium plates of silica gel 60 F254 (E. Merck
5554), detected with UV light (254 nm), 10% vanillin/sulfuric acid
as well as ceric ammonium molybdate (100 g ammonium molyb-
date/8 g ceric sulfate in 1 L 10% H2SO4) and heated on a hotplate.
Preparative TLC was performed on precoated glass plates of silica
gel 60 F254, 0.5 mm (E. Merck 5744). For column chromatography
silica gel 60 (230–400 mesh, E. Merck 9385) was used.
gel providing a diastereomeric mixture (5R:5S ca. 2:9) of ulososides
(81%). Conventional O-deacetylation employing sodium methoxide
(1 mmol) in methanol (25 mL) gave compound 4 (98%) as a mix-
ture of diastereomers at C-5 from which the major 5S-epimer
was purified by chromatography (CHCl3/MeOH 9:1, v/v).
½
a 2D0
ꢀ
¼ ꢁ19:8 (c 1.1, MeOH); 1H NMR: (500 MHz, CD3OD) d 7.50–
7.20 (m, 5H, benzyl); 4.76 (d, 1H, J = 10.7 Hz, benzyl); 4.71 (d,
1H, J1,2 = 7.3 Hz, H-1); 4.69 (d, 1H, benzyl); 4.03 (dd, 1H,
J2,3 = 9.8 Hz, H-2); 3.88 (d, 1H, J6a,6b = 12.2 Hz, H-6a); 3.85 (d, 1H,
J3,4 = 7.8 Hz, H-4); 3.84 (d, 1H, H-6b); 3.71 (dd, 1H, H-3); 3.44 (s,
3H, OMe); 1.98 (s, 3H, N-Ac). 13C NMR: (125 MHz, CD3OD) d 72.6
(NHAc), 138.8, 128.3, 128.0 (2C), 127.9 (2C), benzyl; 101.7 (C-1);
100.6 (C-5); 74.9, 72.5 (C-3, C-4); 63.4, 61.1 (C-6, benzyl); 55.6
(OMe); 55.3 (C-2); 21.8 (N-Ac).
3.2. Kinetic studies
Inhibition of b-N-acetylhexosaminidase from Streptomyces
plicatus was determined as previously described.24 Lysosomal
b-N-acetylhexosaminidase A (Hex A) was purified from human
placenta as reported.25 Inhibitory activity (IC50) of 6a was deter-
mined using 1 mM 4-nitrophenyl b-N-acetyl-glucosaminide as
MS: Calcd for [C16H23NO7]: m/z 341.364; ESIMS found: [M+H]+
342.37, [M+Na]+ 364.35.
outlined.8a Ki was estimated according to the equation Ki = IC50
(1+S/Km), where S is the substrate concentration.
/
3.6. Methyl-N6-(2-acetamino-1,2,5-trideoxy-
diyl)-N2-tert-butyloxycarbonyl-
-lysinate 5
D-glucitol-1,5-
3.2.1. Intracellular b-N-acetylhexosaminidase activity
L
Adult Tay-Sachs (homozygous for alpha subunit G269S muta-
tion) and infantile Sandhoff (homozygous for the 16-kb HEXB dele-
tion mutation) fibroblasts11 were treated with an escalating dose
of 6a (dissolved in DMSO) for 5 days. Intracellular b-N-acetylhex-
osaminidase A/S activity was measured using the fluorogenic sub-
strate 4-methylumbelliferyl N-acetylglucosamine-6-sulfate as
previously described.11
Following general procedure 3.3, ulososide 4 (diastereomeric
mixture, 68 mg, 0.20 mmol) was reacted with lysine derivative A
(90 mg, 1.1 equiv) to give compound 5 (52 mg, 58%): ½a D20
¼ þ1:2
ꢀ
1
(c 1.1, MeOH); H NMR: (500 MHz, CD3OD) d 4.09 (m, 1H, H-20);
3.86 (m, 2H, J6a,6b = 12.2 Hz, H-6a, H-6b); 3.82 (ddd, 1H,
J1eq,2 = 3.9 Hz, J1ax,2 = 10.7 Hz, J2,3 = 8.8 Hz, H-2); 3.71 (s, 3H,
OCH3); 3.40 (dd, 1H, J3,4 = 9.3 Hz, J4,5 = 9.3 Hz, H-4); 3.22 (dd, 1H,
H-3); 3.01 (dd, 1H, J1eq,1ax = 11.2 Hz, H-1eq); 2.80 (m, 1H, H-60a);
2.57 (m, 1H, H-60b); 2.15 (m, 2H, H-1ax, H-5); 1.96 (s, 3H, NHAc);
1.77 (m, 1H, H-30a); 1.66 (m, 1H, H-30b); 1.56–1.30 (m, 4H, 2 ꢂ H-
40, 2 ꢂ H-50); 1.43 (s, 9H, Boc). 13C NMR: (125 MHz, CD3OD) d 173.9,
172.4 (C-10, NHAc); 157.0 (Boc); 79.4 (Boc); 76.5 (C-3); 71.5 (C-4);
66.1 (C-5); 58.5 (C-6); 54.4, 53.8, 52.0 (C-1, C-20, C-60); 51.4 (OCH3);
50.7 (C-2); 31.3 (C-30); 27.6, 27.6, 27.6 (Boc); 23.7, 23.6 (C-40, C-50);,
21.6 (NHAc).
3.3. General procedure for intramolecular reductive amination
To a 0.03 M solution of 4 in MeOH/H2O (15:1, v/v), one equiva-
lent of the peptide component as well as Pd(OH)2/C (20%,
0.1 equiv) was added, and the heterogeneous reaction mixture
was stirred under an atmosphere of hydrogen at ambient pressure
and room temperature for 24 h. After filtration and evaporation of
the solvent under reduced pressure, the residue was purified by
column chromatography on silica gel (CHCl3/MeOH/concd NH3,
500:100:6, v/v/v), yielding the target compound as a colourless
MS: Calcd for [C20H37N3O8]: m/z 447.533; ESIMS found: [M+H]+
448.54, [M+Na]+ 470.52.
syrup. The formation of L-ido-configured products was not ob-
served in any of the reported cases.
3.7. Methyl 6-[N6-(2-acetamino-1,2,5-trideoxy-
diyl)-N2-tert-butyloxycarbonyl-
-lysinyl]aminohexanoate (6)
D-glucitol-1,5-
L
3.4. General procedure for N-BOC-removal and N-dansylation
Following general procedure 3.3, ulososide
0.352 mmol) was reacted with lysine derivative
4
(120 mg,
A 0.01 M solution of the respective BOC-protected iminosugar-
amino acid hybrid in MeOH/AcCl (30:1, v/v, prepared at 0 °C
10 min before use) was stirred for 20 h at ambient temperature.
The solvents were removed under reduced pressure, and the residue
was taken up in dry DMF, yielding a 0.01 M solution. Triethylamine
(5 equiv) and dansyl chloride (1.1 equiv) were added, and the result-
ing reaction mixture was stirred in a brown flask for 4 h at ambient
temperature. Removal of the solvent under reduced pressure and
purification of the residue by preparative TLC (silica gel, CHCl3/
MeOH/concd NH3, 300:100:4, v/v/v, extraction with MeOH) gave
the desired target compound as a green, fluorescent syrup.
B (200 mg,
1.1 equiv) to give compound 6 (142 mg, 72%): ½a D20
¼ 3:0 (c 1.8,
ꢀ
1
MeOH); H NMR: (500 MHz, CD3OD) d = 3.96 (m, 1H, H-20); 3.85
(m, 2H, J6a,6b = 12.2 Hz, H-6a, H-6b); 3.82 (ddd, 1H, J1eq,2 = 3.9 Hz,
J1ax,2 = 10.7 Hz, J2,3 = 8.8 Hz, H-2); 3.65 (s, 3H, OCH3); 3.40 (dd,
1H, J3,4 = 9.3 Hz, J4,5 = 9.3 Hz, H-4); 3.21 (dd, 1H, H-3); 3.24-3.12
(m, 2H, 2 ꢂ H-600); 3.01 (dd, 1H, J1eq,1ax = 11.2 Hz, H-1eq); 2.80
(m, 1H, H-60a); 2.55 (m, 1H, H-60b); 2.32 (t, 2H, 2 ꢂ H-200); 2.12
(m, 2H, H-1ax, H-5); 1.95 (s, 3H, NHAc); 1.75–1.25 (m, 12H,
2 ꢂ H-300, 2 ꢂ H-400, 2 ꢂ H-500, 2 ꢂ H-30, 2 ꢂ H-40, 2 ꢂ H-50); 1.44 (s,
9H, Boc). 13C NMR: (125 MHz, CD3OD) d = 174.7, 174.0, 172.4 (C-
100, C-10, NHAc); 156.6 (Boc); 79.4 (Boc); 76.5 (C-3); 71.5 (C-4);
66.2 (C-5); 58.5 (C-6); 55.0, 54.5, 52.1 (C-1, C-20, C-60); 50.9
(OCH3); 50.7 (C-2); 38.9 (C-6”); 33.5 (C-200); 32.1 (C-300); 28.9 (C-
500); 27.6, 27.6, 27.6 (Boc); 26.2, 24.5, 23.9, 23.7, (C-40, C-50, C-300,
C-400); 21.6 (NHAc). MS: Calcd for [C26H48N4O9]: m/z 560.694;
ESIMS found: [M+H]+ 561.70, [M+Na]+ 583.69.
3.5. Methyl (5R/S)-2-acetamino-5-C-benzyloxy-2-deoxy-b-
xylo-hexopyranoside 4
D-
To a 1% solution of methyl 3,4-di-O-acetyl-2-acetamino-2,6-
dideoxy-b- -xylo-hex-5-enopyranoside (1.00 g, 3.32 mmol) in a
D
mixure of dichloromethane/benzyl alcohol (1:1, v/v), 3-chloroper-
benzoic acid (70%, 900 mg, 3.65 mmol) was added, and the mixture
was stirred at ambient temperature for 2 h. The mixture was par-
titioned between dichloromethane (120 mL) and aqueous sodium
bicarbonate (5%), the organic layer was dried (Na2SO4), the solvent
removed under reduced pressure, and the resulting residue was
chromatographed (cyclohexane/ethyl acetate 1:4, v/v) on silica
3.8. Methyl-N6-(2-acetamino-1,2,5-trideoxy-
diyl)-N2-dansyl-
-lysinate 5a
D-glucitol-1,5-
L
Following general procedure 3.4, compound
5
(45 mg,
0.1 mmol) gave fluorescent inhibitor 5a (26 mg, 45%):
½
a 2D0
ꢀ
¼ þ16:6 (c 1.3, MeOH); 1H NMR: (500 MHz, CD3OD) d 8.55

A. J. Steiner et al. / Tetrahedron: Asymmetry 20 (2009) 832–835
835
701–711; (c) Liu, J.; Numa, M. M. D.; Liu, H.; Huang, S.-J.; Sears, P.; Shikhman, A.
R.; Wong, C.-H. J. Org. Chem. 2004, 69, 6273–6283.
2. Reese, T. A.; Liang, H.-E.; Tager, A. M.; Luster, A. D.; Van Rooijen, N.; Voehringer,
D.; Locksley, R. M. Nature 2007, 447, 92–96.
3. Liu, F.; Iqbal, K.; Grundke-Iqbal, I.; Hart, G. W.; Gong, C.-X. Proc. Natl. Acad. Sci.
U.S.A. 2004, 101, 10804–10809.
(d, 1H, dansyl); 8.38 (d, 1H, dansyl); 8.18 (d, 1H, dansyl); 7.59 (t,
1H, dansyl); 7.54 (t, 1H, dansyl); 7.27 (d, 1H, dansyl); 3.76 (m,
3H, J6a,6b = 12.2 Hz, H-2, H-6a, H-6b); 3.72 (m, 1H, H-20); 3.35
(dd, 1H, J3,4 = 9.3 Hz, J4,5 = 9.7 Hz, H-4); 3.21 (s, 3H, OCH3); 3.17
(dd, 1H, J2,3 = 8.8 Hz, H-3); 2.87 (s, 6H, dansyl); 2.83 (dd, 1H,
J1eq,1ax = 11.7 Hz, J1eq,2 = 4.8 Hz, H-1eq); 2.45 (m, 1H, H-60a); 2.32
(m, 1H, H-60b); 2.04 (ddd, 1H, J5,6a = 2.0 Hz, J5,6b = 2.4 Hz, H-5);
2.00 (dd, 1H, J1ax,2 = 11.2, H-1ax); 1.98 (s, 3H, NHAc); 1.60–1.50
(m, 2H, 2 ꢂ H-30); 1.22–0.80 (m, 4H, 2 ꢂ H-40, 2 ꢂ H-50). 13C
NMR: (125 MHz, CD3OD) d 172.5, 172.4 (C-10, NHAc); 152.0,
135.8, 130.3, 129.9, 129.9, 129.3, 127.9, 123.1, 119.7, 115.2 (dan-
syl); 76.6, (C-3); 71.6 (C-4); 65.9 (C-5); 58.6 (C-6); 55.8, 54.5,
51.8 (C-1, C-20, C-60); 51.1 (OCH3); 50.7 (C-2); 44.7, 44.7 (dansyl);
31.9 (C-30); 23.0, 23.0 (C-4’, C-50); 21.6 (NHAc). MS: Calcd for
[C27H40N4O8S]: m/z 580.706; ESIMS found: [M+H]+ 581.71,
[M+Na]+ 603.69.
4. (a) Kolter, T.; Sandhoff, K. Biochim. Biophys. Acta 2006, 1758, 2057–2079; (b)
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1080, 89–95.
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M. N. G. J. Biol. Chem. 2001, 276, 10330–10337; (b) Kappes, E.; Legler, G.
J. Carbohydr. Chem. 1989, 8, 371–378.
7. Blattner, R.; Furneaux, R. H.; Kemmitt, T.; Tyler, P. C.; Ferrier, R. J.; Tiden, A.-K.
J. Chem. Soc., Perkin Trans. 1 1994, 3411–3421. and references cited therein.
8. (a) Tropak, M. B.; Blanchard, J. E.; Withers, S. G.; Brown, E. D.; Mahuran, D.
Chem. Biol. 2007, 14, 153–164; (b) Maegawa, G. H. B.; Tropak, M.; Buttner, J.;
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9. For example: Iminosugars: Recent insights into their bioactivity and potential
as therapeutic agents, Martin, O. R., Compain, P., Eds.. Curr. Top. Med. Chem.
2003, 3, 471–591; From lianas to glycobiology tools: Twenty-five years of 2,5-
3.9. Methyl 6-[N6-(2-acetamino-1,2,5-trideoxy-
diyl)-N2-dansyl-
-lysinyl]aminohexanoate 6a
D-glucitol-1,5-
L
dideoxy-2,5-imino-
D-mannitol, Wrodnigg, T. M. In Timely Research Perspectives
in Carbohydrate Chemistry; Schmid, W., Stütz, A. E., Eds.; Springer: Vienna, New
York, 2002; pp 43–76; Heightman, T. D.; Vasella, A. T. Angew. Chem., Int. Ed.
1999, 38, 750; Iminosugars as Glycosidase Inhibitors; Stütz, A. E., Ed.; Wiley-VCH:
Weinheim, 1999.
Following general procedure 3.4, compound
6
(50 mg,
0.089 mmol) gave fluorescent inhibitor 6a (46 mg, 74%):
10. (a) Spiro, R. G. In Carbohydrates in Chemistry and Biology; Ernst, B., Hart, G. W.,
Sinay, P., Eds.; Part II: Biology of Saccharides; Wiley-VCH: Weinheim, 2000;
Vol. 3, pp 65–79; (b) Elbein, A. D.; Molyneux, R. J. In Iminosugars as Glycosidase
Inhibitors; Stütz, A. E., Ed.; Wiley-VCH: Weinheim, 1999; pp 216–251.
11. Tropak, M. B.; Reid, S. P.; Guiral, M.; Withers, S. G.; Mahuran, D. J. Biol. Chem.
2004, 279, 13478–13487.
½
a 2D0
ꢀ
¼ þ1:9 (c 0.9, MeOH); 1H NMR: (500 MHz, CD3OD) d 8.56 (d,
1H, dansyl); 8.36 (d, 1H, dansyl); 8.21 (d, 1H, dansyl); 7.60 (t, 1H,
dansyl); 7.56 (t, 1H, dansyl); 7.27 (d, 1H, dansyl); 3.76 (m, 3H, H-
2, H-6a, H-6b); 3.65 (s, 3H, OCH3); 3.57 (m, 1H, H-20); 3.34 (dd,
1H, J3,4 = 9.3 Hz, J4,5 = 9.3 Hz, H-4); 3.17 (dd, 1H, J2,3 = 8.8 Hz, H-
3); 2.88 (s, 6H, dansyl); 2.83 (dd, 1H, J1eq,1ax = 11.2 Hz,
J1eq,2 = 4.4 Hz, H-1eq); 2.82 (m, 2H, 2 ꢂ H-600); 2.41 (m, 1H, H-
60a); 2.30 (m, 1H, H-60b); 2.27 (t, 2H, 2 ꢂ H-200); 2.05 (ddd, 1H,
J5,6a = 2.9 Hz, J5,6b = 2.4 Hz, H-5); 1.99 (dd, 1H, J1ax,2 = 11.7 Hz, H-
1ax); 1.97 (s, 3H, NHAc); 1.56–0.84 (m, 12H, 2 ꢂ H-30, 2 ꢂ H-40,
2 ꢂ H-50, 2 ꢂ H-300, 2 ꢂ H-400, 2 ꢂ H-500); 13C NMR: (125 MHz,
CD3OD) d 174.7, 172.6, 172.3 (C-1C, C-10, NHAc); 152.1, 135.5,
130.4, 130.0, 129.8, 129.5, 128.2, 123.1, 119.4, 115.3 (dansyl);
76.5 (C-3); 71.5 (C-4); 65.9 (C-5); 58.5 (C-6); 56.9, 54.5, 51.8 (C-
1, C-2’, C-60); 50.9 (OCH3); 50.7 (C-2); 44.7, 44.7 (dansyl); 38.8
(C-6”); 33.5 (C-200); 32.5 (C-30); 28.5 (C-500); 26.1, 24.4, 23.2, 23.1
(C-40, C-5’, C-300, C-400); 21.6 (NHAc).
12. Hermetter, A.; Scholze, H.; Stütz, A. E.; Withers, S. G.; Wrodnigg, T. M. Bioorg.
Med. Chem. Lett. 2001, 11, 1339–1342.
13. (a) Lundt, I.; Steiner, A. J.; Stütz, A. E.; Tarling, C. A.; Ully, S.; Withers, S. G.;
Wrodnigg, T. M. Bioorg. Med. Chem. 2006, 14, 1737–1742; (b) Greimel, P.;
Häusler, H.; Lundt, I.; Rupitz, K.; Stütz, A. E.; Tarling, C. A.; Withers, S. G.;
Wrodnigg, T. M. Bioorg. Med. Chem. Lett. 2006, 16, 2067–2070.
14. Steiner, A. J.; Stütz, A. E.; Wrodnigg, T. M.; Tarling, C. A.; Withers, S. G.;
Hermetter, A.; Schmidinger, H. Bioorg. Med. Chem. Lett. 2008, 18, 1922–
1925.
15. For a comprehensive review of the method see: Baxter, E. W.; Reitz, A. B. Org.
React. 2002, 59, 1–714; for a typical example see: Zou, W.; Szarek, W. A.
Carbohydr. Res. 1993, 242, 311–314.
16. Stütz, A. E.; Steiner, A. J.; Wrodnigg, T. M. Eur. Pat. Appl. EP 2006-19545, 2008.
17. Steiner, A. J.; Stütz, A. E.; Tarling, C. A.; Withers, S. G.; Wrodnigg, T. M.
Carbohydr. Res. 2007, 342, 1850–1858.
18. Khan, R.; Hough, L. Carbohydr. Res. 1972, 24, 147–151.
19. Garegg, P. J.; Johansson, R.; Ortega, C.; Samuelsson, B. J. Chem. Soc., Perkin Trans.
1 1982, 681–683.
MS: Calcd for [C33H51N5O9S]: m/z 693.867; ESIMS found:
[M+H]+ 694.90, [M+Na]+ 716.90.
20. Barili, P. L.; Berti, G.; Catalani, G.; D’Andrea, F. Chim. Ital. 1992, 122, 135–
142.
21. McDonnell, C.; Cronin, L.; O’Brien, J. L.; Murphy, P. V. J. Org. Chem. 2004, 69,
3565–3568.
Acknowledgments
22. Steiner, A. J.; Schitter, G.; Stütz, A. E.; Wrodnigg, T. M.; Tarling, C. A.; Withers, S.
G.; Fantur, K.; Mahuran, D.; Paschke, E.; Tropak, M. Bioorg. Med. Chem. 2008, 16,
10216–10220.
23. Tropak, M. B.; Kornhaber, G. J.; Rigat, B. A.; Maegawa, G. H.; Buttner, J. D.;
Blanchard, J. E.; Murphy, C.; Tuske, S. J.; Coales, S. J.; Hamuro, Y.; Brown, E. D.;
Mahuran, D. J. ChemBioChem 2008, 9, 2650–2662.
Financial support by the Austrian Fonds zur Förderung der Wis-
senschaftlichen Forschung (FWF), Vienna, (Project P18998-N17) as
well as by the Canadian Institutes for Health Research (CIHR) is
gratefully acknowledged.
24. Mark, B. L.; Vocadlo, D. J.; Zhao, D.; Knapp, S.; Withers, S. G.; James, M. N. G. J.
Biol. Chem. 2001, 276, 42131–42137.
25. Mahuran, D.; Lowden, J. A. Can. J. Biochem. 1980, 58, 287–294.
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