Synthesis of oligomeric phosphono analogues of Leishmania lipophosphoglycan fragments

Article abstract of DOI:10.1016/S0040-4039(02)01588-5

A set of isosteric phosphono (C-glycoside) analogues of Leishmania lipophosphoglycan fragments have been synthesised by making use of a sequence of uniformly stereoselective glycosylation reactions of phosphono-oligosaccharide glycosyl acceptors with phosphono-oligosaccharide thioglycosides.

Full text of DOI:10.1016/S0040-4039(02)01588-5

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 43 (2002) 7821–7825
Synthesis of oligomeric phosphono analogues of Leishmania
lipophosphoglycan fragments
Vladimir S. Borodkin,a Fiona C. Milne,b Michael A. J. Fergusonb and Andrei V. Nikolaeva,*
aUniversity of Dundee, School of Life Sciences, Division of Biological Chemistry and Molecular Microbiology,
Carnelley Building, Dundee DD1 4HN, UK
bUniversity of Dundee, School of Life Sciences, Division of Biological Chemistry and Molecular Microbiology,
Wellcome Trust Building, Dundee DD1 4HN, UK
Received 22 May 2002; revised 23 July 2002; accepted 31 July 2002
Abstract—A set of isosteric phosphono (C-glycoside) analogues of Leishmania lipophosphoglycan fragments have been synthe-
sised by making use of a sequence of uniformly stereoselective glycosylation reactions of phosphono-oligosaccharide glycosyl
acceptors with phosphono-oligosaccharide thioglycosides. © 2002 Elsevier Science Ltd. All rights reserved.
The Leishmania family of protozoan parasites causes
severe debilitating and often fatal diseases throughout
the tropics and subtropics. There is strong evidence that
infectivity and survival in the mammalian host are both
counted on the lipophosphoglycan (LPG) molecules
which are ubiquitous on the surface of the parasite
(Scheme 1).1 The known antigenic properties of this
unique glycoconjugate prompted vaccine design studies
based on the different parts of the molecule.2,3 Along
this line, we reported the preparation of fully synthetic
fragments of the LPG from several species of the
Leishmania4 and a method for conjugation of these
phosphosaccharides to protein and lipid carriers.2 Now
we would like to disclose the synthesis of oligomeric
phosphono (C-glycoside) analogues of the LPG frag-
ments that would be more robust chemically than par-
ent phosphosaccharides and resist the action of
phosphatases as well.
Recently, we reported5 the synthesis of dec-9-enyl b-
galactopyranosyl-(1“4)-a- -mannopyranosyl methane-
D-
D
phosphonate, a phosphono analogue of the disac-
charide repeating unit of the LPG. Based on these
developments, an obvious approach to the targeted
phosphono-oligosaccharides was devised assuming they
could be obtained by stepwise ‘oligomerisation’ of the
disaccharide phosphonic acid 6 selectively protected at
6%-OH (Scheme 2). Exploring this possibility, we have
Scheme 1. Generalised structure of the Leishmania lipophosphoglycan.
Keywords: carbohydrates; phosphonic acids and derivatives.
* Corresponding author.
0040-4039/02/$ - see front matter © 2002 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(02)01588-5

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V. S. Borodkin et al. / Tetrahedron Letters 43 (2002) 7821–7825
Scheme 2. Reagents and conditions: (i) TMSBr, DCM, 1 h, rt, quant.; (ii) DCC, THF, 65°C, 2 h, 57%; (iii) DCC, pyridine, 72 h,
rt, 62%; (iv) DCC, DMAP, pyridine, 72 h, rt, 68% for 2+3, 66% for 6+3, 30% for 2+8, 10% for 6+8; (v) 2% HCl–MeOH, DCM,
12 h, rt, 88%.
found that DCC-mediated esterification of model phos-
phonic acid 2 with the galactoside alcohol 3 in the
presence of DMAP produced the phosphonodisaccha-
ride 4 in 68% yield.6 This combination of reagents was
proved to be equally effective for the preparation of the
phosphonotrisaccharide 7 from the phosphonic acid 6
and the monosaccharide alcohol 3. However, attempted
esterification of 2 with the monohydroxylic phospho-
nodisaccharide derivative 8 afforded only 30% yield of
the phosphonotrisaccharide 9. Things turned out to be
even worse when coupling of the disaccharide building
blocks 8 and 6 provided no more than 10% of the
phosphonotetrasaccharide 10. These findings showed
that the 6%-OH group in the disaccharide 8 is disap-
pointingly less reactive than the 6-OH group in the
monosaccharide 3, though the reason for the above
inconsistency remains unclear.
Trying to circumvent this obstacle, we discovered that
the phosphonotrisaccharide 138 (an analogue of 9)
could be effectively obtained instead by the stereoselec-
tive glycosylation of the mannosyl methanephosphonate
derivative 125 with the phosphonodisaccharide
trichloroacetimidate 11;7 whereas, the thiogalactoside
147 was found to be the glycosyl donor of choice for the
preparation of the phosphonodisaccharide 158 in a
separate experiment (Scheme 3). The exclusive b-selec-
tivity of these reactions8 clearly manifested the stere-
ocontrolling role of the 2,3-O-isopropylidene group in
the glycosyl acceptor 12 in accordance with our previ-
ous observations.5
These data brought an insight as to how the original
synthetic strategy could be redesigned to achieve the
ultimate objectives. Now we considered the dec-9-enyl
O
OCH3
O
O
OCH3
O
O
OCH3
OCH3
P
P
P
O
OCH3
OCH3
P
i
BzO
BzO
BzO
BzO
BzO
BzO
BzO
BzO
O
+
O
O
HO
O
O
O
O
OBz BzO
O
OBz BzO
O
BzO
O
BzO
O
BzO
O
CCl3
BzO
11
12
13
NH
O
OCH3
OCH3
O
OCH3
OCH3
P
P
OAc
OAc
O
BzO
BzO
BzO
BzO
ii
O
O
O
+
HO
SEt
O
O
O
O
BzO
BzO
BzO
O
BzO
15
12
14
,
Scheme 3. Reagents and conditions: (i) TMSOTf, DCM, 1 h, −20°C, 72%; (ii) MeOTf, DCM, 4 A MS, 3 h, rt, 88%.

V. S. Borodkin et al. / Tetrahedron Letters 43 (2002) 7821–7825
7823
Scheme 4. Reagents and conditions: (i) MBnOC(NH)CCl3, 0.3% TfOH, DCM, 20 min, rt, 90%; (ii) PhSH, Et3N, DMF, 12 h, rt,
quant.; (iii) dec-9-enol, Ph3P, DIAD, THF, 2 h, rt, 88%; (iv) DDQ, DCM–H2O, 3 h, rt, 85%; (v) Ph3P, DIAD, THF, 16 h, 60°C,
68% for 17+20, 58% for 23+20.
(mannosyl methanephosphonate) 19 (Scheme 4) as a
‘primer’ acceptor, while the phosphonodisaccharide
thioglycoside 21 orthogonally protected at 4%-OH as a
‘repeating unit’ for the phosphono-oligosaccharide
chain assembly via repetitive glycosylation reactions.
We anticipated the high level of stereoselection would
be maintained all the way through using the combina-
tion of thioglycoside donors and matched glycosyl
acceptors of the optimal structure. The phosphono-
trisaccharide thioglycoside 24 was introduced as a ‘cap’
for the final glycosylation step to keep the overall
convergence of the synthetic scheme.
ing from compound 12 (Scheme 4). Acid-catalysed
installation of the p-methoxybenzyl (MBn) protective
group9 followed by selective monodemethylation of the
phosphonic diester with PhSH–Et3N10 cleanly afforded
the monoester 17. Esterification of this material with
dec-9-enol using the Mitsunobu protocol11 followed by
removal of the MBn protection with dichlorodi-
cyanobenzoquinone (DDQ) produced the requisite
hydroxyl derivative 19 (as
a
mixture of two
diastereomers at phosphorus). A similar combination of
PhSH–Et3N selective demethylation/Mitsunobu esterifi-
cation was successfully applied to the preparation of
the phosphonodisaccharide 21 and the phosphono-
trisaccharide 24. The required phosphonic diesters were
obtained (as diastereomeric mixtures at phosphorus)
The preparation of the mannosyl methanephosphonate
19 was accomplished in a straightforward manner start-
O
OCH3
O
O
OCH3
O
P
P
O
OCH3
ODec
O
OCH3
ODec
P
P
BzO
BzO
i
O
O
RO
BzO
MBnO
BzO
O
O
O
+
O
O
O
O
O
O
SEt
BzO
HO
BzO
O
O
O
O
BzO
BzO
BzO
BzO
21
19
25 R = MBn
26 R = H
ii
21
26
OCH3
O
O
P
OCH3
O
P
OCH3
i
O
BzO
BzO
O
BzO
O
P
RO
BzO
O
O
O
O
ODec
O
O
O
O
O
BzO
O
BzO
BzO
O
O
BzO
BzO
27 R = MBn
28 R = H
ii (repeated twice)
Dec = dec-9-enyl
,
Scheme 5. Reagents and conditions: (i) MeOTf, DCM, 4 A MS, 7 h, rt, 87% for 21+19, 75% for 21+26; (ii) DDQ, DCM–H2O,
2 h, rt, 80% for 25 to 26, 75% for 27 to 28.

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V. S. Borodkin et al. / Tetrahedron Letters 43 (2002) 7821–7825
i
i
i
CH3O
NH4
O
O
24 + 19
CH3O
O
H
O
P
OBz
O
O
P
BzO
BzO
P
HO
ii, iii, iv
BzO
BzO
O
O
ODec
n
ODec
n
HO
O
24 + 26
24 + 28
O
O
O
HO
O
OBz
O
BzO
O
OH
O
O
OBz
BzO
OH
HO
OBz BzO
32 n = 2
33 n = 3
29 n = 1
30 n = 2
31 n = 3
34 n = 4
Dec = dec-9-enyl
,
Scheme 6. Reagents and conditions: (i) MeOTf, DCM, 4 A MS, 16 h, rt, 79% for 24+19, 68% for 24+26, 67% for 24+28; (ii) TFA,
DCM–H2O, 10 min, rt; (iii) PhSH, Et3N, DMF, 12 h, rt; (iv) MeONa cat., MeOH, 24 h, rt.
from the reactions of the galactoside alcohol 2012 with
phosphonic monoesters 17 (68%) and 23 (58%),
respectively.
The phosphono analogues 32–34 were used for the
preparation (using a dec-9-enyl moiety as a linker2) of
neoglycoconjugates as potential synthetic vaccines.
They have also revealed a biological activity (similar to
parent phosphosaccharides) as effective acceptor sub-
With all the principal building blocks in hand, we
pursued the preparation of phosphono analogues of di-,
tri- and tetrameric repeats of the Leishmania LPG,
compounds 32, 33 and 34, respectively. The MeOTf-
promoted glycosylation of the mannosyl methanephos-
strates for the elongating a-D-mannosyl phosphate
transferase (MPT) in the Leishmania.14 The results of
the studies will be published elsewhere in due course.
phonate
thioglycoside 21 exclusively provided the b-linked phos-
phonotrisaccharide 25 (for -Galp: lH1 5.05, J1,2 8.2
19
with
the
phosphonodisaccharide
Acknowledgements
D
The Wellcome Trust International Grant supported this
work and V.S.B.
Hz; lC1 101.2; a mixture of four possible isomers at
phosphorus) in 87% yield (Scheme 5). Treatment of 25
with DDQ ensued removal of the MBn protection in
80% yield. Reiterative glycosylation of thus prepared 26
with the same thioglycoside 21 afforded the phospho-
nopentasaccharide 27 (as a mixture of eight isomers at
phosphorus) in 75% yield, again, as the b-isomer only
at the newly formed galactoside bond(s) (lH1 5.05, J1,2
8.0 Hz; lC1 101.3). Deprotection of 27 required DDQ
oxidation to be repeated twice to achieve the conver-
sion into monohydroxylic derivative 28 in 75% yield.
References
1. (a) Turco, S. J.; Spa¨th, G. F.; Beverley, S. M. Trends
Parasitol. 2001, 17, 223–226; (b) Ilg, T. Parasitol. Today
2000, 16, 489–497.
2. Routier, F. H.; Nikolaev, A. V.; Ferguson, M. A. J.
Glycoconjugate J. 2000, 16, 773–780.
3. Hewitt, M. C.; Seeberger, P. H. J. Org. Chem. 2001, 66,
4233–4243 and references cited therein.
In due course, a series of uniform MeOTf-promoted
reactions of the mono-, tri- and penta-saccharide
derivatives 19, 26 and 28, respectively, with the phos-
phonotrisaccharide thioglycoside 24 validated the ver-
satility of the reiterative glycosylation approach to the
oligomeric phosphono analogues of the LPG: the fully
protected phosphono-tetrasaccharide 29 (four isomers
at phosphorus), -hexasaccharide 30 (eight isomers at
phosphorus) and -octasaccharides 31 (16 isomers at
4. (a) Nikolaev, A. V.; Rutherford, T. J.; Ferguson, M. A.
J.; Brimacombe, J. S. J. Chem. Soc., Perkin Trans. 1
1995, 1977–1987; (b) Nikolaev, A. V.; Rutherford, T. J.;
Ferguson, M. A. J.; Brimacombe, J. S. J. Chem. Soc.,
Perkin Trans. 1 1996, 1559–1566; (c) Higson, A. P.;
Tsvetkov, Y. E.; Ferguson, M. A. J.; Nikolaev, A. V. J.
Chem. Soc., Perkin Trans. 1 1998, 2587–2595; (d) Higson,
A. P.; Tsvetkov, Y. E.; Ferguson, M. A. J.; Nikolaev, A.
V. Tetrahedron Lett. 1999, 40, 9281–9284.
phosphorus) were obtained exclusively with b-D-
galactopyranoside bonds (lC1 101.1–101.2) in 79, 68,
and 67% yields, respectively (Scheme 6).
5. Borodkin, V. S.; Ferguson, M. A. J.; Nikolaev, A. V.
Tetrahedron Lett. 2001, 42, 5305–5308.
6. All new compounds showed satisfactory spectral and
analytical data.
Finally, stepwise deprotection of the above phosphono-
oligosaccharides produced the targeted compounds 32,
33 and 34 in 83, 75 and 65% overall yields, respectively,
after isolation by anion-exchange chromatography. The
structures of the oligomers were supported by NMR
and ES-MS data.13 Steady changes of integration of
characteristic NMR signals (e.g. H-1, C-1, C-5 and C-6
in Gal residues and C-4 in Man residues) in terminal
and internal units were consistent with chain length
growing from the dimer 32 to the tetramer 34.
7. Compound 11 was prepared from 4 in three steps: (i)
CH2N2, EtOAc–MeOH, 0°C, 90%; (ii) TFA, DCM, 1 h,
rt; (iii) Cl3CCN, DBU, DCM, 1 h, 0°C, 95% for ii, iii.
Compound 14 was prepared from ethyl tetra-O-acetyl-1-
thio-b-D-galactopyranoside in four steps: (i) MeONa,
MeOH, 2 h, rt; (ii) DMTCl, DMAP, pyridine, 12 h, rt,
then BzCl, 12 h, rt; (iii) aq. TFA, DCM, 5 min, rt; (iv)
Ac2O, pyridine, 2 h, rt, 80% for i–iv.
8. Compound 13: 1H NMR (300 MHz, CDCl3, selected
data): l 5.05 (d, 1H, J1,2 8.0, H-1, Gal); 13C NMR (75

V. S. Borodkin et al. / Tetrahedron Letters 43 (2002) 7821–7825
7825
MHz, CDCl3, selected data): l 101.2 (C-1, Gal). Com-
26.9 (br d, 3C, JC,P 133.2, C-1*, Man–Man¦), 60.7 (3C,
C-6, Man–Man¦), 61.4 (C-6, Gal¦), 62.7 (br, 2C, C-6,
Gal, Gal%), 68.3 (br, 2C, C-4, Gal, Gal%), 68.9 (C-4, Gal¦),
69.4 (br, 3C, C-3, Man–Man¦), 71.1 (br, 6C, C-2, Man–
Man¦, Gal–Gal¦), 72.6 (2C, C-3, Gal, Gal%), 72.8 (C-3,
Gal¦), 73.0 (2C, C-5, Man, Man¦), 73.2 (C-5, Man%), 73.6
(C-1, Man%), 73.8 (C-1, Man), 74.2 (br, 3C, C-1, Man¦
and C-5, Gal, Gal%), 75.6 (C-5, Gal¦), 77.2 (C-4, Man¦),
78.0 (2C, C-4, Man, Man%), 103.3 (C-1, Gal¦), 103.5 (2C,
C-1, Gal, Gal%), 114.2 (ꢀCH2), 140.8 (ꢁCHꢀ); 31P NMR
(121 MHz, D2O): l 22.5 (P), 23.0 (P¦), 23.2 (P%); ES-MS
(−) data: m/z 453.31 (100%, [M−3NH3−3H]3−) (expected
m/z 453.14), 680.23 (90%, [M−3NH3−2H]2−) (expected
m/z 680.21); [h]D +23 (c 1.0, MeOH–H2O, 1:1).
1
pound 15: H NMR (300 MHz, CDCl3, selected data): l
5.07 (d, 1H, J1,2 8.0, H-1, Gal); 13C NMR (75 MHz,
CDCl3, selected data): l 101.3 (C-1, Gal).
9. Nakajima, N.; Horita, K.; Abe, R.; Yonemitsu, O. Tetra-
hedron Lett. 1988, 29, 4139–4142.
10. (a) Daub, G. W.; Van Tamelen, E. E. J. Am. Chem. Soc.
1977, 99, 3526–3528; (b) Muller, B.; Martin, T. J.;
Schaub, C.; Schmidt, R. R. Tetrahedron Lett. 1998, 39,
509–512.
11. (a) Norbeck, D. W. J. Org. Chem. 1987, 52, 2174–2179;
(b) Campbell, D. A. J. Org. Chem. 1992, 57, 6331–6335;
(c) Campbell, D. A.; Bermack, J. C. J. Org. Chem. 1994,
59, 658–660.
12. Prepared as described for 14 (Ref. 7) except the final
acetylation was omitted.
1
Compound 34: H NMR (300 MHz, D2O, selected data):
4.10 (m, 4H, H-1, Man–Man§), 4.25 (d, 1H, J1,2 8.0, H-1,
Gal§), 4.30 (d, 3H, J1,2 8.0, H-1, Gal–Gal¦); 13C NMR
(75 MHz, D2O): l 25.3, 28.5, 28.6, 28.7, 28.8, 30.4 (d,
JC,P 6.3), 33.5, 65.2 (d, JC,P 6.0) [8×CH2, dec-9-enol], 26.9
(br d, 4C, JC,P 133.2, C-1*, Man–Man§), 60.7 (4C, C-6,
Man–Man§), 61.4 (C-6, Gal§), 62.7 (br, 3C, C-6, Gal–
Gal¦), 68.3 (br, 3C, C-4, Gal–Gal¦), 68.9 (C-4, Gal§),
69.4 (br, 4C, C-3, Man–Man§), 71.2 (br, 8C, C-2, Man–
Man§, Gal–Gal§), 72.7 (3C, C-3, Gal–Gal¦), 72.8 (C-3,
Gal§), 73.1 (2C, C-5, Man, Man§), 73.2 (br, 2C, C-5,
Man%, Man¦), 73.7 (br, 2C, C-1, Man%, Man¦), 73.9 (C-1,
Man), 74.1 (d, 3C, JC,P 8.1, C-5, Gal–Gal¦), 74.2 (C-1,
Man§), 75.7 (C-5, Gal§), 77.2 (C-4, Man§), 78.0 (3C,
C-4, Man–Man¦), 103.4 (C-1, Gal§), 103.6 (3C, C-1,
Gal–Gal¦), 114.2 (ꢀCH2), 140.8 (ꢁCHꢀ); 31P NMR (121
MHz, D2O): l 22.5 (P), 23.0 (P§), 23.2 (P%+P¦); ES-MS
(−) data: m/z 587.40 (100%, [M−3NH3−3H]3−) (expected
m/z 587.16); [h]D +25.7 (c 0.54, H2O).
1
13. Compound 32: H NMR (300 MHz, D2O, selected data):
l 4.10 (m, 2H, H-1, Man, Man%), 4.25 (d, 1H, J1,2 8.0,
H-1, Gal%), 4.30 (d, 1H, J1,2 8.0, H-1, Gal); 13C NMR (75
MHz, D2O): l 25.3, 28.5, 28.6, 28.7, 28.9, 30.5 (d, JC,P
6.2), 33.5, 65.1 (d, JC,P 5.4) [8×CH2, dec-9-enol], 27.0 (d,
2C, JC,P 132.6, C-1*, Man, Man%), 60.8 (2C, C-6, Man,
Man%), 61.4 (C-6, Gal%), 62.8 (d, JC,P 6.0, C-6, Gal), 68.4
(C-4, Gal), 68.9 (C-4, Gal%), 69.4 (C-3, Man%), 69.5 (C-3,
Man), 71.2 (br, 4C, C-2, Man, Man%, Gal, Gal%), 72.7
(C-3, Gal), 72.8 (C-3, Gal%), 73.1 (br, 2C, C-5, Man,
Man%), 73.9 (C-1, Man), 74.2 (C-1, Man%), 74.3 (d, JC,P
8.1, C-5, Gal), 75.7 (C-5, Gal%), 77.3 (C-4, Man%), 78.1
(C-4, Man), 103.4 (C-1, Gal%), 103.6 (C-1, Gal), 114.2
(ꢀCH2), 140.8 (ꢁCHꢀ); 31P NMR (121 MHz, D2O): l 22.5
(P), 23.0 (P%); ES-MS (−) data: m/z 479.28 (100%, [M−
2NH3−2H]2−) (expected m/z 479.17); [h]D +25.5 (c 0.5,
MeOH–H2O, 1:1).
1
Compound 33: H NMR (300 MHz, D2O, selected data):
14. Routier, F. H.; Higson, A. P.; Ivanova, I. A.; Ross, A. J.;
Tsvetkov, Y. E.; Yashunsky, D. V.; Bates, P. A.; Niko-
laev, A. V.; Ferguson, M. A. J. Biochemistry 2000, 39,
8017–8025.
l 4.10 (m, 3H, H-1, Man–Man¦), 4.25 (d, 1H, J1,2 8.0,
H-1, Gal¦), 4.30 (d, 2H, J1,2 8.0, H-1, Gal, Gal%); 13C
NMR (75 MHz, D2O): l 25.2, 28.4, 28.5, 28.7, 28.8, 30.4
(d, JC,P 6.1), 33.4, 65.0 (d, JC,P 6.0) [8×CH2, dec-9-enol],