Stereoselective synthesis of 2-acetamido-1,2-dideoxyallonojirimycin (DAJNAc), a new potent hexosaminidase inhibitor

Article abstract of DOI:10.1021/ol401517x

A practical synthesis of the previously unreported N-acetyl-d-allosamine glycomimetic DAJNAc is described. The reaction sequence involves Pd-catalyzed allylic substitution by phthalimide in an azaheterobicyclic scaffold as the key step. The new iminosugar resulted in being a stronger β-N- acetylglucosaminidase (human placenta) competitive inhibitor than the d-gluco (DNJNAc) and d-galacto (DGJNAc) stereoisomers.

Full text of DOI:10.1021/ol401517x

ORGANIC
LETTERS
2013
Vol. 15, No. 14
3638–3641
Stereoselective Synthesis of
2‑Acetamido-1,2-dideoxyallonojirimycin
(DAJNAc), a New Potent Hexosaminidase
Inhibitor
†
Alex de la Fuente, Ruben Martin, Teresa Mena-Barragan, Xavier Verdaguer,
‡
§
#,†
ꢀ
Jose M. Garcıa Fernandez, Carmen Ortiz Mellet,* and Antoni Riera*
,§
,†,#
ꢀ
ꢀ
´
Institute for Research in Biomedicine (IRB Barcelona), Barcelona, Spain, Institut
ꢁ
ꢀ
´
Catala d’Investigacio Quımica (ICIQ), Tarragona, Spain, Departamento de Quımica
Organica, Facultad de Quımica, Universidad de Sevilla, Sevilla, Spain, Departament
´
ꢀ
´
´
de Quımica Organica, Universitat de Barcelona, Barcelona, Spain, and Instituto de
Investigaciones Quımicas (IIQ), CSIC and Universidad de Sevilla, Sevilla, Spain
ꢁ
´
mellet@us.es; antoni.riera@irbbarcelona.org
Received May 29, 2013
ABSTRACT
A practical synthesis of the previously unreported N-acetyl-D-allosamine glycomimetic DAJNAc is described. The reaction sequence involves
Pd-catalyzed allylic substitution by phthalimide in an azaheterobicyclic scaffold as the key step. The new iminosugar resulted in being a stronger
β-N-acetylglucosaminidase (human placenta) competitive inhibitor than the D-gluco (DNJNAc) and D-galacto (DGJNAc) stereoisomers.
Iminosugars (frequently termed azasugars) are structur-
al analogs of monosaccharides in which the oxygen ring
atom has been replaced by nitrogen.1 Since nojirimycin
(NJ, 1; Figure 1) was first isolated from Streptomyces in
1966, the number of natural and synthetic iminosugars has
never stopped growing. Given the instability of the hemi-
aminal functionality,2 most biologically relevant represen-
tatives are 1-deoxy derivatives, such as 1-deoxynojirimycin
(DNJ, 2) or its stereoisomers (Figure 1).3 The biological
activity of these compounds may be related to their
capacity to mimic the glycosyloxocarbenium cation in
their protonated state, a species postulated to be close to
the transition state in the reaction coordinate of enzymatic
glycoside hydrolysis. Many iminosugars are competitive
glycosidase inhibitors.4 The broad range of metabolic
processes in which glycosidases participate makes them
† Institute for Research in Biomedicine (IRB Barcelona).
‡
ꢁ
ꢀ
Institut Catala d'Investigacio Quımica (ICIQ).
´
§ Departamento de Quı
´
mica Organica, Universidad de Sevilla.
mica Organica, Universitat de Barcelona.
ꢀ
# Departament de Quı
´
ꢁ
Instituto de Investigaciones Quımicas (IIQ).
´
(1) For excellent monographs, see: (a) Iminosugars: From Synthesis
to Therapeutic Applications; Compain, P., Martin, O. R., Eds.; Wiley-VCH:
Weinheim, Germany, 2007. (b) Iminosugars as Glycosidase Inhibitors:
Nojirimycin and Beyond; St€utz, A. E., Ed.; Wiley-VCH: Weinheim, Ger-
many, 1999.
(2) Remarkable exceptions to this principle are the so-called sp2-
iminosugars, in which the endocyclic nitrogen is part of a pseudoamide-
ꢀ
type functionality. For selected examples, see: (a) Jimenez Blanco, J. L.;
ꢀ
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r
10.1021/ol401517x
Published on Web 06/26/2013
2013 American Chemical Society

potential candidates for the treatment of a range of
diseases, including cancer, type II diabetes, lysosomal
storage disorders, or viral infections such as HIV or
dengue, among others.5 Some iminosugars are currently
marketed as drugs, such as miglitol (Glyset ) for the
treatment of type II diabetes mellitus and N-butyl-DNJ
(Zavesca ) for the treatment of Gaucher disease.6
context of our ongoing efforts to develop selective glycosi-
dase inhibitors as drug candidates,17 here we report the first
synthesis of DAJNAc and a preliminary screening of its
glycosidase inhibitory capacity.
Among iminosugars, the analogs of N-acetylhexosa-
mines have a strong potential for the treatment of acquired
and genetic diseases such as osteoarthritis,7 allergy,8
and Alzehimer9 or Sandhoff, Tay-Sachs, and Schindler-
Kanzaki syndromes.10 Thus, 2-acetamido-1,2-dideoxy-D-
nojirimycin (DNJNAc, 3; Figure 1) and some of its deriva-
tives are potent inhibitors of β-hexosaminidases11,12
and of β-O-(N-acetyl)glucosaminidase (O-GlcNAcase).13
The analogous compound with the D-galacto configura-
tion, namely 2-acetamido-1,2-dideoxy-D-galactonojirimycin
(DGJNAc, 4) and its N-alkyl derivatives, are highly
potent competitive inhibitors of R-galactosaminidases
(R-GalNAcases) and β-hexosaminidases,14,15 whereas
2-acetamido-1,2-dideoxymannonojirimycin (DMJNAc, 5)
and its derivatives are potent reversible inhibitors of UDP-
N-acetylglucosamine 2-epimerase.16 Given the remarkable
biological activity of N-acetylhexosamine analogs, here
we sought to complete the series with the, to date, unknown
D-allo-configured congener, namely 2-acetamido-1,2-
dideoxy-D-allonojirimycin (DAJNAc, 6; Figure 1). In the
Figure 1. Structures of nojirimycin (1), its 1-deoxy derivative (2),
and the diastereoisomeric 2-acetamido-1,2-dideoxyiminosugars
with the D-gluco- (3), D-galacto- (4), D-manno- (5), and D-allo-
configuration (6).
Most reported iminosugar syntheses rely on the chiral
pool, using carbohydrates, amino acids, or tartaric acid as
starting materials.18 However, the high functional group
density of these compounds and the difficulties associated
with the incorporation of the acetamido group in the all-cis
C2;C3;C4 segment, characteristic of D-allo-configured
derivatives, makes the synthesis of DAJNAc 6 particularly
challenging. These considerations drove us to design an
asymmetric synthesis of 6 based on the key bicyclic pre-
cursor 7, which is easily accessible in high enantiomeric
purity by Sharpless epoxidation of penta-1,4-dien-3-ol.19
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Org. Lett., Vol. 15, No. 14, 2013
3639

Scheme 1. Synthesis of N-Acetamidediolcarbamates 11a (D-allo)
and 11b (D-galacto) from 7a,b
Scheme 2. Synthesis of Configurationally Stable Pd Allyl
Complex 12
Table 1. Isolated Yields of the Products Arising from
Pd-Catalyzed Addition of Nitrogen Nucleophiles to the
Allylic-Type Esters 8a and 8ba
entry
substrate
nucleophile
product
yield/%
1
8a
8a
8a
8a
8a
8a
8a
8a
8a
8b
butylamine
9a
9b
9c
9d
9e
9f
0
2
dibenzylamine
diisopropylamine
piperidine
0
3
0
4
88
91
88
74
25
48
75
5
morpholine
6
hexamethyleneimine
isoindoline
7
9g
9h
9h
9h
8
phthalimide
9b
10b
cesium phthalimide
cesium phthalimide
a Unless otherwise stated, the reaction conditions were those indi-
cated in Scheme 1. b The reactions were performed in DME at 85 °C.
a Ortep drawings of the crystal structures of 11a and 11b with 50%
probability ellipsoids. b The yields ofthe firststepcan befoundinTable 1.
Although the above procedure was satisfactory in view
of the synthesis of iminosugar hexosamine analogs, none
of the amines 9bÀ9g were suitable to introduce the acet-
amide fragment of the target N-acetylallosamine mimic 6.
The use of phthalimide as the nucleophile21 afforded the
N-phthalimido derivative 9h in 25% yield, a potential
precursor of DAJNAc (Scheme 1; Table 1, entry 8). The
conditions developed by Trost et al.22 involving generation
of the corresponding cesium salt in DME allowed the
synthesis of 9h in 48% yield. Replacing acetate 8a by
carbonate 8b, prepared from 7 in 90% yield by treatment
with ethyl chloroformate/pyridine, further increased the
yield of the addition product 9h, obtained as a single
diastereoisomer (entry 10) in 75% yield. The phthalimido
group was deprotected by treatment with hydrazine in
EtOH at reflux, affording the corresponding primary
amine, which was acetylated in situ to give acetamide 10
in 98% overall yield. Dihydroxylation of 10 using NMO
Acetylation of 7 with acetic anhydride/Et3N/DMAP
provided 8a in 98% yield (Scheme 1). Subsequent introduc-
tion of the amino moiety by Pd-catalyzed allylic
substitution20 was, however, problematic. Use of the azide
anion or primary or secondary amines as nucleophiles and
bis(allyl)palladium chloride/1,2-bis(diphenylphosphino)-
ethane (dppe) as the catalyst proved unsuccessful (Table 1,
entries 1À3). Of note, only cyclic secondary amines provided
the desired addition products 9dÀ9g in satisfactory yields
(Table 1, entries 3À7). A single reaction product was
observed in those cases, with complete control of the regio-
and diastereoselectivity. Analysis of products 9d, 9e, and 9f
by NOESY experiments revealed that the reaction took
place through a syn attack to the less hindered C5 position in
the piperidine ring (C2 in the iminosugar nomenclature).
The observed diastereoselectivity points to the complete
complexation of the Pd-atom through the opposite face of
the acetoxy leaving group. This hypothesis was confirmed
after preparation of complex 12, in 85% yield, by treatment
of 8a with equimolar amounts of the catalyst, followed
by precipitation with ammonium hexafluorophosphate
(Scheme 2). As expected, only one diastereoisomer was
observed by NMR and the relative stereochemistry of 12
was established by NOESY experiments.
and catalytic amounts of K2OsO4 2H2O in acetone/water
3
afforded a 4:1 mixture of the diastereomeric N-acetamide-
diolcarbamates 11a (D-allo) and 11b (D-galacto) in 66%
yield. The use of “Sharpless asymmetric dihydroxylation”
conditions did not improve the yield or the diastereomeric
ratio. Both isomers were crystalline solids easily separable
by chromatography, and the configuration assignments
(21) (a) Gaertner, M.; Weihofen, R.; Helmchen, G. Chem.;Eur. J.
2011, 17, 7605–7622. (b) Gnamm, C.; Franck, G.; Miller, N.; Stork, T.;
Broedner, K.; Helmchen, G. Synthesis 2008, 3331–3350.
(20) (a) Trost, B. M.; Crawley, M. L. Chem. Rev. 2003, 103, 2921–
2943. (b) Johannsen, M.; Jorgensen, K. A. Chem. Rev. 1998, 98, 1689–
1708.
(22) Trost, B. M.; Lee, C. B. J. Am. Chem. Soc. 2001, 123, 3687–3696.
3640
Org. Lett., Vol. 15, No. 14, 2013

were confirmed by X-ray diffraction. Final hydrolysis of
the 2-oxazolidinone ring of 11a and 11b by treatment with
6 M NaOH at reflux gave the title iminosugar DAJNAc 6
(40% yield) and the known N-acetylgalactosamine mimic
DGJNAc 414,15 in 38% yield, respectively (Scheme 3).
Table 2. Inhibition Contants (Ki, μM) against Commercial
β-N-Acetylglucosaminidases for DAJNAc (6) Determined from
the Slope of LineweaverÀBurk Plots and Double Reciprocal
Analysisa
β-N-acetylglucosaminidaseb
315
415
6
human placenta
bovine kidney
jack beans
7.0 ( 0.2
7.4 ( 0.3
2.9 ( 0.1
8.3 ( 0.3
4.2 ( 0.1
1.8 ( 0.1
5.6 ( 0.1
2.6 ( 0.1
2.6 ( 0.1
Scheme 3. Synthesis of DAJNAc 6 and DGJNAc 4 from 11a
and 11b
a Data for the corresponding analogs with D-gluco- (DNJNAc, 3)
and D-galacto-configuration (DGJNAc, 4) are included for comparative
purposes. No inhibition was observed for any of the compounds at a
concentration of 2 mM on almonds and bovine liver β-glucosidases,
yeast R-glucosidase, jack bean R-mannosidase (except for 6; Ki 150 mM),
Helix pomatia β-mannosidase, Aspergillus niger amyloglucosidase,
Penicillium decumbens naringinase, green coffee R-galactosidase, E. coli
β-galactosidase, or yeast isomaltase. b Inhibition was competitive for
compound 6 against β-N-acetylglucosaminidase from human placenta
and noncompetitive for the enzymes from bovine liver and jack beans
(see the Supporting Information).
With DAJNAc in hand, a preliminary screening of
its inhibitory properties against a panel of commercial
glycosidases, including three β-N-acetylglucosaminidases
of mammalian (human placenta and bovine kidney) and
plant (jack beans) origin, was conducted. With the excep-
tion of modest activity against β-mannosidase (Ki 150 μM),
DAJNAc was found to be a selective inhibitor of the
β-N-acetylglucosaminidases. The corresponding inhibition
constants (Ki) are shown in Table 2 and compared with
data for the D-gluco and D-galacto epimers DNJNAc (3)
and DGJNAc (4), respectively. The Ki values (5.6À2.6 μM)
were comparable to those obtained for DNJNAc and
DGJNAc. In fact, the inhibitory capacity of compound 6
against the enzyme from human placenta was 1.3À1.5-fold
stronger than that of the “matching” iminosugar 3 or its
C4-epimer 4. Figure 2 depicts the structures of the strongest
inhibitors reported to date for the three enzymes evaluated
in this work, namely 13,23 14,24 and 15,25 and the corre-
sponding Ki or IC50 values.
Figure 2. Structures of pyrrolidine 13, 2-acetamido-2-deoxyno-
jirimycin 14, and nagstatin derivative 15, the strongest inhibitors
reported to date for the β-N-acetylglucosaminidase from human
placenta, Jack beans, and bovine kidney, respectively.
Acknowledgment. We thank the Spanish Ministerio de
Economia y Competitividad (CTQ2011-23620, SAF2010-
15670, and CTQ2010-15848; cofinanced by FEDER), the
ꢀ
Generalitat de Catalunya (2009SGR 00901), the Fundacion
Ramon Areces, the Junta de Andalucıa (Project P08-
ꢀ
´
FQM-03711), and IRB Barcelona for financial support.
ꢀ
A.F. thanks the Ministerio de Ciencia e Innovacion for a
fellowship.
The results demonstrate that β-N-acetylglucosaminidases
accept conformational modifications at the C3 and C4
centers in the glycone moiety of substrate analogs without
affecting the binding affinity. The efficiency of the synthetic
strategy described here and its suitability for molecular
diversity-oriented strategies warrants further research.
Supporting Information Available. General experimen-
tal methods, experimental procedures, compound char-
acterization data, NMR spectra for all new compounds,
X-ray data for compounds 11a and 11b, and Linewea-
verÀBurk representations for Ki determinations. This
material is available free of charge via the Internet at
http://pubs.acs.org.
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Lin, Y.-L.; Wong, C.-H. ChemBioChem 2006, 7, 165–173.
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D. J. Biol. Chem. 2004, 279, 13478–13487.
(25) Terinek, M.; Vasella, A. Helv. Chim. Acta 2005, 88, 10–22.
The authors declare no competing financial interest.
Org. Lett., Vol. 15, No. 14, 2013
3641