Synthesis of potential bisubstrate inhibitors of Leishmania elongating α-D-mannosyl phosphate transferase

Article abstract of DOI:10.1016/j.tetlet.2003.11.022

Synthesis of a potential mechanism-based bisubstrate inhibitor 1 of the elongating α-D-mannosyl phosphate transferase in Leishmania, comprising a guanosine subunit bound to the synthetic acceptor substrate through the methylenebisphosphonate linker, as well as its analogues 2 and 3 has been successfully accomplished.

Full text of DOI:10.1016/j.tetlet.2003.11.022

Tetrahedron
Letters
Tetrahedron Letters 45 (2004) 857–862
Synthesis of potential bisubstrate inhibitors of Leishmania
elongating a- -mannosyl phosphate transferase
D
Vladimir S. Borodkin,a Michael A. J. Fergusonb and Andrei V. Nikolaeva,*
aFaculty of Life Sciences, Division of Biological Chemistry and Molecular Microbiology, University of Dundee, Carnelley Building,
Dundee DD1 4HN, UK
bFaculty of Life Sciences, Division of Biological Chemistry and Molecular Microbiology, University of Dundee,
Wellcome Trust Building, Dundee DD1 4HN, UK
Received 15 August 2003; revised 27 October 2003; accepted 6 November 2003
Abstract—Synthesis of a potential mechanism-based bisubstrate inhibitor 1 of the elongating a-D-mannosyl phosphate transferase
in Leishmania, comprising a guanosine subunit bound to the synthetic acceptor substrate through the methylenebisphosphonate
linker, as well as its analogues 2 and 3 has been successfully accomplished.
ꢀ 2003 Elsevier Ltd. All rights reserved.
Sandfly-transmitted protozoan parasites of the Leish-
mania genus cause a variety of debilitating or fatal dis-
eases throughout the tropical and subtropical regions of
the globe. It has been shown that survival and infectivity
of the parasite in the mammalian host and in the insect
vector are both dependent on the lipophosphogly-
can (LPG) molecules, which are ubiquitous at the par-
asiteÕs cell surface.1 The biosynthesis of the polymeric
devised (Fig. 1) taking into account that a-mannosyl
phosphate is transferred to the growing end of the LPG
chain (i.e., an acceptor substrate, AS, X ¼ O, R ¼ LPG)
from the donor substrate GDP–mannose en block with
retention of anomeric configuration.2;3 Whereas the
synthetic phosphonodisaccharide, dec-9-enyl b-
D-ga-
lactopyranosyl-(1 fi 4)-a- -mannopyranosyl methane-
D
phosphonate was shown to be an effective exogenous
acceptor substrate [AS, X ¼ CH2, R ¼ (CH2)8CH@CH2]
for the eMPT,5 we assume the enzyme simultaneously
hosts the acceptor and the donor substrates, while the
entire bond breaking–bond forming process occurs in
one step in the ternary complex formed.
backbone portion of this glycoconjugate [-6)-b-
D-Galp-
(1 fi 4)-a-
D
-Manp-(1-POꢀ3 -]n proceeds through the con-
secutive action of the elongating a-mannosyl phosphate
transferase (eMPT) and b-galactosyltransferase enzyme
activities with the former only being found in the
Leishmania parasite,2 thus endorsing the search for
selective eMPT inhibitors as prospective anti-leishman-
iasis drugs. As a part of our ongoing programme
towards elucidation of the enzymeÕs mode of action by
means of synthetic probes,3 we wish to disclose here
preliminary results aimed at the preparation of potential
mechanism-based eMPT bisubstrate inhibitors.4
Thus, the covalently bound transition state analogue 1
nonhydrolysable methylenebisphosphonate
with
a
(MBP) bridge mimicking the pyrophosphate moiety of
GDP–mannose while linking together the acceptor
substrate and the guanosine residue was recognised as a
potential eMPT bisubstrate-type inhibitor. Moreover,
the simpler analogues 2 and 3, either lacking the
Little is known so far about the structure of the eMPT
active site. However, a putative transition state model
(TS) for a-mannosyl phosphate transfer could be
nucleoside subunit (2) or bearing a 2-dimethyl-
aminoethylphosphonate residue instead of MBP (3),
both being, however, able to take part in the chelation
phenomena with divalent metal ion(s) presumably
present in the active site of the enzyme, were also con-
sidered as synthetic targets.
Keywords: Carbohydrates; Phosphonic acids and derivatives; Transi-
tion states.
* Corresponding author. E-mail: a.v.nikolaev@dundee.ac.uk
Our first goal was the preparation of the simplified
analogue 3 (Scheme 1). Phosphonylation of the 60-OH in
0040-4039/$ - see front matter ꢀ 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2003.11.022

858
V. S. Borodkin et al. / Tetrahedron Letters 45 (2004) 857–862
OH
O
HO
OH
O
OH
O
HO
OH
O
O
HO
HO
O
HO
HO
H
HO
HO
AS
O
O
OR
P
X
OR
O
O
HO
O
P
HO
OH
O
N
O
P
O
P
P
OH
O
O
N
O
O
O
N
N
1 R =
NH
NH2
O
P
N
HO
NH
NH2
HO
HO
O
HO
O
N
O
OH
OH
HO OH
O
P
O
P
HO OH
HO
O
OMe
2 R =
3 R =
TS
P
NMe2
OH
HO
Figure 1. Putative transition state model for the a-D-mannosyl phosphate transfer and structures of the potential bisubstrate-type inhibitors.
OBz
O
BzO
BzO
O
O
BTO
BTO
i
BzO
BzO
O
P
+
BzO
Br
O
O
P
OH
MeO
5
4
BT = 1-benzotriazolyl
OBz
O
BzO
OR
O
RO
BzO
O
RO
O
ii
BzO
BzO
O
RO
RO
O
BzO
RO
O
O
O
O
P
P
O
O
O
O
R2O
MeO
P
P
7
R1
R2O
MeO
6 R = Bz; R1 = Br; R2 = Me
8 R = Bz; R1 = NMe2; R2 = Me
3 R = R2 = H; R1 = NMe2
iii
v
iv
Scheme 1. Reagents and conditions: (i) DCM, 1,4-dioxane, rt, 30 min, then MeOH excess, 16 h, 85%; (ii) Et3N, DCM, rt, 30 min, 81%; (iii) NaI,
Me2CO, 60 ꢁC, 4 h, then Me2NH, THF, rt, 16 h, 54%; (iv) Me2NH, THF, rt, 16 h, 86%; (v) PhSH, Et3N, DMF, rt, 16 h, then MeONa, MeOH, rt, 5 h,
90%.
the phosphonodisaccharide 46 with bis(benzotriazolyl)
2-bromoethylphosphonate 57 provided the mixed phos-
phonodiester 68 in 85% yield. However, further trans-
formation into the required N,N-dimethylamino
derivative 8 either by direct treatment of 6 with Me2NH/
THF or through the intermediate iodide was not as
effective as expected. Repeatedly, the yield of the
required product did not exceed 54%. This product was
obtained more effectively through the conjugate addi-
tion of the same reagent to the vinyl phosphonate 7
easily prepared from 6 by short treatment with Et3N.7
Finally, deprotection provided the target compound 3 as
the bis-ammonium salt after isolation by ion-exchange
chromatography.9
of only traces of the desired disaccharide product 16. In
agreement with our previous observations,6 the appar-
ently diminished reactivity of the 60-OH in compound 4
could be accounted for by the steric hindrance imposed
by the benzoate protection at 3-OH of the mannose
residue closely flanking the reactive site, as revealed by
inspection of molecular models.
At this juncture we decided that the synthetic approach
had to be diverted to the consecutive esterification–gly-
cosylation concept developed by us previously.6 Thus, a
suitable glycosyl donor already bearing the MBP moiety
had to be prepared from 11 and reacted with the known
mannosyl methanephosphonate acceptor 176 to give
access to a suitable precursor of 2.
Encouraged by the straightforward implementation of
the first objective we turned our efforts to the prepara-
tion of the methylenebisphosphonate derivative 2
(Scheme 2). Direct introduction of the MBP group via
the monochloridate reagent 13 (derived from methyl-
enebisphosphonic acid trimethyl ester 1010) in the pres-
ence of Et3N and DMAP,11 while being successful when
applied to the simple galactose derivative 9 (65% iso-
lated yield of MBP-carrying glycoside 11), completely
failed in the case of the phosphonodisaccharide 4. These
results were replicated in a parallel Mitsunobu esterifi-
cation12 of 9 and 4 with the triester 10. In this case, a
high yield of the phosphonylated monosaccharide
derivative 11 (83%) again contrasted with the formation
All attempts to elaborate the 2-(trimethylsilyl)ethyl
galactoside 11 (via the hemiacetal derivative) into the
corresponding glycosyl trichloroacetimidate or bromide
proved to be unsuccessful due to the instability of the
MBP substructure towards either basic (DBU or
Cs2CO3/DCM) or mild brominating agents [HBr/DCM
or (COBr)2/DMF/DCM]. The solution was eventually
found when we tested the MBP-carrying thioglycoside
14 as a stable glycosyl donor. Mitsunobu esterification
of the parent thiogalactoside 12 with the triester 10
provided 14 in 67% optimised yield along with ca. 20%
of the exo-methylene by-product 15. Rewardingly, the
MeOTf-promoted glycosylation of 17 with 14 proceeded

V. S. Borodkin et al. / Tetrahedron Letters 45 (2004) 857–862
859
O
P
O
P
MeO
BzO
OMe
OMe
BzO
BzO
OH
O
O
P
O
P
O
BzO
BzO
OMe
OMe
X
O
ii or iii
O
SEt
+
R
R
BzO
MeO
BzO
BzO
BzO
9 R = OC2H4TMS
12 R = SEt
11 R = OC2H4TMS
14 R = SEt
10 X = OH
13 R = Cl
15
i
OBz
BzO
O
BzO
BzO
O
OBz
BzO
BzO
ii or iii
O
BzO
O
BzO
O
O
O
O
+
10 or 13
BzO
BzO
P
O
O
OMe
OMe
BzO
OH
P
MeO
MeO
P
O
P
O
MeO
O
16
4
OR1
O
RO
BzO
RO
BzO
HO
RO
RO
R2O
O
BzO
SEt
O
iv
O
O
O
R1O
+
O
BzO
O
O
P
OMe
OMe
OMe
O
O
O
R2O
O
MeO
P
OR2
P
P
P
O
P
MeO
O
O
O
14
18 R = Bz; R1,R1
2 R = R1 = R2 = H
=
2 ; R2 = Me
17
C(Me)
v
Scheme 2. Reagents and conditions: (i) (COCl)2, DMF cat, DCM, 0 ꢁC, 1 h; (ii) Et3N, DMAP, DCM, 0 ꢁC to rt, 6 h, 65% for 9+13, no reaction for
ꢀ
4+13; (iii) PPh3, DIAD, THF, 60 ꢁC, 1 h, 83% for 9+10, 67% for 12+10, traces for 4+10; (iv) MeOTf, MS 4 A, DCM, rt, 2.5 h, 72%; (v) TFA, DCM–
H2O, rt, 15 min, then PhSH, Et3N, DMF, rt, 16 h, then MeONa, MeOH, rt, 5 h, 80%.
BzO
OR
BzO
BzO
BzO
SEt
OBn
P
O
N
N
N
N
O
O
BzO
BzO
SEt
OR2
P
O
OR
N
N
HO
O
O
N
N
N
N
O
iv
O
BnO
+
P
R1O
OR1
N
N
O
P
O
O
O
O
O
O
O
O
14 R1 = R2 = Me
25 R = H
23 R = H
O
O
19 R1 = Me; R2 = H
26 R = CO(NPh)2
24 R = CO(NPh)2
i
20 R1 = H; R2 = Me
21 R1 = Bn; R2 = Me
22 R1 = Bn; R2 = H
ii
iii
BzO
O
BzO
BzO
O
O
N
O
BzO
BzO
SEt
OBn
P
O
BzO
BzO
Ph
O
BzO
O
O
O
N
Ph
O
O
O
O
O
HO
P
v
O
O
N
N
O
+
MeO
O
O
BnO
P
O
P
O
OR
N
N
N
N
P
O
N
BnO
P
TfO
MeO
O
N
N
O
BnO
N
N
O
O
O
O
O
26
17
27 R = CO(NPh)2
O
O
O
O
Scheme 3. Reagents and conditions: (i) PhSH, Et3N, DMF, rt, 16 h, quant; (ii) PhCHN2, t-BuOMe, HBF4 cat, rt, 67%; (iii) KCN, DMF, 70 ꢁC, 6 h,
ꢀ
88%; (iv) PPh3, DIAD, THF, 60 ꢁC, 4 h, 32% for 22+23, 65% for 22+24; (v) MeOTf, MS 4 A, DCM, rt, 4 h, 50%.
with complete stereoselectivity to give the b-linked
accomplished through the stereoselective glycosylation
of the mannosyl methanephosphonate acceptor 17 with
the complex guanosine–MBP–thioglycoside donor 25
(Scheme 3) that could be prepared, in turn, via the
Mitsunobu esterification of the monodeprotected MBP-
carrying thioglycoside 19 with a suitable guanosine
derivative 23.14a It is worthy to note, that the protective
group pattern in the derivative 23 meant that the final
deprotection could be performed in a nonhydro-
genolytic way14b thus keeping the dec-9-enyl moiety,
0
0
0
phosphonodisaccharide 18 (J1 ;2 ¼ 8:0 Hz; dC1 ¼ 101.7–
101.9) as a mixture of four diastereomers at phosphorus
in 72% yield. Finally, the stepwise removal of the pro-
tecting groups from 18 afforded the required compound
2 as the tris-ammonium salt after purification by ion-
exchange chromatography.13
Going ahead with the construction of the guanosine-
containing compound 1 we first intended this to be

860
V. S. Borodkin et al. / Tetrahedron Letters 45 (2004) 857–862
important for the preparation of neoglycoconjugates,15
intact in the targeted product.
Despite seemingly being unproductive, the above
approach already encompassed all the synthetic essen-
tials needed for the successful preparation of the target
compound 1 (vide infra). Specifically, with the estab-
lished protocol of chemoselective monodeprotection of
the MBP moiety in hand the introduction of the gua-
nosine block could be advantageously switched further
to the end of the synthetic sequence thus avoiding
the presence of the incompatible heterocyclic base at
the glycosylation step. This required, however, that the
methyl protecting group for the phosphonodiester
function in the mannosyl methanephosphonate acceptor
17 had to be changed to a benzyl one in order to be
stable during the MBP monodeprotection.
In the event, an attempted regioselective monodeme-
thylation of the phosphonic dimethyl ester in 14 with
either neat t-BuNH2 or DABCO in THF only resulted
in the formation of an inseparable 1:1 mixture of 19 and
its regioisomer with the phosphonic monoacid proximal
to the galactose residue.16 Obviously, the protecting
groups at the MBP moiety had to be remodeled to make
a chemoselective deprotection feasible. To fulfil this
objective the starting compound 14 was treated with
PhSH–Et3N–DMF17 to produce the phosphonic diacid
20 almost quantitatively. This was exhaustively esteri-
fied18 with phenyl diazomethane in the presence of
HBF4 to provide the compound 21 with the selectively
protected MBP group in 67% yield, which was smoothly
converted into the required monoacid derivative 22 on
treatment with KCN in DMF.19
Fortunately, preparation of the required mannosyl
methanephosphonate acceptor 31 proved to be quite
straightforward (Scheme 4). Selective monodeprotection
of the starting mixed phosphonodiester 286 with PhSH–
Et3N (nearly quantitative yield) followed by the Mit-
sunobu-type esterification with benzyl alcohol (92%
yield) and removal of the p-methoxybenzyl (MBn) pro-
tecting group (90%) provided the desired product
seamlessly. Next, the standard MeOTf promoted gly-
cosylation of the acceptor 31 with the MBP-carrying
thioglycoside 21 was again completely b-selective giv-
ing the phosphonodisaccharide MBP derivative 32
The Mitsunobu esterification of 22 with the guanosine
synthone 23 provided the targeted thiogalactoside–
MBP–guanosine building block 25 albeit in a meagre
32% yield. Predictably, when 6-O-protected guanosine
derivative 2420 was used as the hydroxylic component in
the same reaction, the yield of the corresponding ester
26 was almost twice as high. The standard MeOTf
promoted glycosylation of the mannosyl methane-
phosphonate acceptor 17 with the thioglycoside 26
did furnish some unstable material with a newly formed
0
(dC1 ¼ 101.6–101.8) in an acceptable yield of 47% along
with some 20% of compound 33 presumably arising
from transesterification of one of the phosphonodiester
groups present in the molecule with MeOTf. Selective
saponification of the single methyl phosphonic ester in
32 with KCN in DMF proceeded without incident (75%
yield), as did the crucial Mitsunobu esterification of 34
thus prepared with the guanosine derivative 23, which
produced the required compound 35 in 50% yield.
0
b-glycosidic bond (dC1 ¼ 101.2–101.4). On the basis of
the MALDI-TOF spectrum however, the structure of
this product was tentatively assigned as the corre-
sponding N-methylimidazolium salt 27 in accordance
with the presence of a molecular ion with m=z 2037.72,
exactly one methyl group heavier than required.
BzO
BzO
O
BzO
BzO
SEt
BzO
O
BzO
R1O
O
O
O
BzO
BzO
O
O
O
iv
O
OBn
OMe
+
O
O
O
O
BnO
O
P
O
P
P
R1O
O
P
P
O
O
O
R2O
R1O
P
R1O OR
21
28 R1 = MBn; R2 = Me
29 R1 = MBn; R2 = H
i
ii
32 R = Me; R1 = Bn
33 R = Me; R1 = 2 x Bn and Me
34 R = H; R1 = Bn
30 R1 = MBn; R2 = Bn
31 R1 = H; R2 = Bn
v
iii
OH
BzO
O
BzO
BzO
O
N
N
O
N
N
O
BzO
O
O
BzO
BzO
HO
O
O
O
BzO
BzO
O
N
N
vi
O
O
O
O
O
+
P
O
O
P
O
O
BnO
P
BnO
O
O
OH
O
P
O
BnO
P
BnO
N
O
O
P
N
N
O
BnO
35
1
BnO OH
N
N
N
O
O
34
23
vii
O
O
O
Scheme 4. Reagents and conditions: (i) PhSH, Et3N, DMF, rt, 16 h, quant; (ii) BnOH, PPh3, DIAD, THF, rt, 1 h, 92%; (iii) DDQ, DCM–H2O, rt,
ꢀ
2 h, 90%; (iv) MeOTf, MS, 4 A, DCM, rt, 4 h, 47%; (v) KCN, DMF, 70 ꢁC, 6 h, 75%; (vi) PPh3, DIAD, THF, rt, 6 h, 50%; (vii) TFA, DCM–H2O, rt,
15 min, then PhSH, Et3N, DMF, rt, 16 h, then t-BuOK, t-BuOH–THF–H2O, rt, 16 h, 80%.

V. S. Borodkin et al. / Tetrahedron Letters 45 (2004) 857–862
861
(d, JC;P 4.5, –OCH2CH2–), 68.1 (C-40), 69.1 (C-3), 70.7
(C-20), 70.8 (d, JC;P 9.0, C-2), 72.3 (C-30), 72.5 (C-5), 73.1
(C-1), 74.1 (d, JC;P 8.5, C-50), 77.7 (C-4), 103.2 (C-10),
114.0 (–CH@CH2), 140.2 (–CH@CH2); 31P NMR
(121 MHz, D2O): d 20.6 (P0), 22.3 (P); ES-MS ()) data:
Finally, sequential deprotection21 of 35 performed
without isolation of any intermediates followed by ion-
exchange chromatographic purification delivered the
phosphonodisaccharide–MBP–guanosine hybrid 122 as
the tris-ammonium salt in 80% yield.
m=z 692.28 (100%, [M)H]ꢀ) (expected m=z 692.29;
22
C27H53NO15P2 requires M, 693.29); ½aꢁ +15.2ꢁ (c 0.5,
D
In conclusion, we report here the synthesis of a potential
mechanism-based bisubstrate inhibitor of the elongating
a-mannosyl phosphate transferase in Leishmania
designed with the emphasis on the incorporation of a
guanosine moiety linked to the acceptor substrate
through the methylenebisphosphonate bridge mimicking
the important guanosine-pyrophosphate motif present
in the natural substrate donor GDP–mannose, as well as
its simplified analogues. The results of a biological
evaluation of these compounds will be disclosed else-
where in due course.
MeOH).
10. Compound 10 was prepared from tetramethyl methyl-
enebisphosphonate by saponification with neat t-BuNH2
at rt for 28 h, followed by evaporation of the amine,
treatment of the residue with Dowex-50 (Hþ) in MeOH
and concentration. The triester 10 (which contained up to
10% of the symmetric dimethyl ester) was used without
additional purification.
11. Malachowski, W. P.; Coward, J. K. J. Org. Chem. 1994,
59, 7616–7624.
12. Campbell, D. A.; Bermack, J. C. J. Org. Chem. 1994, 59,
658–660.
13. Compound 2: 1H NMR (300 MHz, D2O): d 1.20–1.35
(10H, m, 5 · CH2), 1.54 (2H, quintet, J 6.6, –OCH2CH2–),
1.89–2.02 (4H, m, H-1*a, -1*b and –CH2CH@CH2), 2.10
Acknowledgements
0
0
(2H, br t, JH;P 19.5, –PCH2P–), 3.46 (1H, dd, J2 ;3 10.0, H-
The Wellcome Trust International Grant supported this
work and V.S.B.
20), 3.50 (3H, d, JH;P 10.3, OMe), 3.62 (1H, dd, J3 ;4 3.5, H-
30), 3.65 (1H, m, H-5), 3.74–3.98 (11H, m, H-2, -3, -4, -40,
-50, -6a, -6b, -60a, -60b and –OCH2CH2–), 4.16 (1H, m,
0
0
H-1), 4.38 (1H, d, J1 ;2 7.8, H-10), 4.89 and 4.97 (2H, 2 · br
d, J 10.2, J 17.3, –CH2CH@CH2), 5.85 (1H, ddt, JH;CH2
6.7, –CH2CH@CH2); 13C NMR (75 MHz, D2O): d 25.2(t,
JC;P 125.9, –PCH2P–), 26.8 (d, JC;P 133.2, C-1*), 25.0, 28.2,
28.3, 28.4, 28.6 (5 · CH2), 30.2 (d, JC;P 6.0, –OCH2CH2–),
33.2 (–CH2CH@CH2), 51.6 (d, JC;P 3.5, –OMe), 60.4
(C-6), 62.6 (br, C-60), 64.9 (d, JC;P 5.6, –OCH2CH2–), 68.0
(C-40), 69.3 (C-3), 70.9 (d, JC;P 8.0, C-2), 71.0 (C-20), 72.4
(C-30), 72.9 (C-5), 73.5 (C-1), 73.8 (br, C-50), 77.6 (C-4),
103.3 (C-10), 114.0 (–CH@CH2), 140.6 (–CH@CH2);
31P NMR (121 MHz, D2O): d 18.8 (br, P0 + P00), 22.5 (P);
ES-MS ()) data: m=z 364.33 (100%, [M)2H]2ꢀ) (expected
0
0
References and notes
1. (a) Turco, S. J.; Spath, G. F.; Beverley, S. M. Trends
Parasitol. 2001, 17, 223–226; (b) Ilg, T. Parasitol. Today
2000, 16, 489–497.
2. (a) Carver, M. A.; Turco, S. J. J. Biol. Chem. 1991, 266,
10974–10981; (b) Carver, M. A.; Turco, S. J. Arch.
Biochem. Biophys. 1992, 295, 309–317.
3. 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.
4. For the definition of the bisubstrate inhibition concept:
Palcic, M.; Heerze, L.; Srivastava, O. P.; Hindsgaul, O. J.
Biol. Chem. 1989, 264, 17174–17181; For recent advances
in this area: Hinou, H.; Sun, X.-L.; Ito, Y. Tetrahedron
Lett. 2002, 43, 9147–9150, and references cited therein.
5. Borodkin, V. S.; Ferguson, M. A. J.; Nikolaev, A. V.
Tetrahedron Lett. 2001, 42, 5305–5308.
m=z 364.10), 729.05 (40%, [M)H]ꢀ) (expected m=z 729.21;
22
C25H49O18P3 requires M, 730.21); ½aꢁ +11.2ꢁ (c 0.52,
D
MeOH–H2O, 1:1).
14. (a) Turner, J. J.; Filipov, D. V.; Overhand, M.; van der
Marel, G. A.; van Boom, J. A. Tetrahedron Lett. 2001, 42,
5763–5767; (b) Vincent, S. P.; Mioskowski, C.; Lebeau, L.
Nucleos. Nucleot. 1999, 18, 2127–2139.
15. Routier, F. H.; Nikolaev, A. V.; Ferguson, M. A. J.
Glycoconjugate J. 2000, 16, 773–780.
6. Borodkin, V. S.; Milne, F. C.; Ferguson, M. A. J.;
Nikolaev, A. V. Tetrahedron Lett. 2002, 43, 7821–7825.
7. Van der Klein, P. M. A.; Dreef, C. E.; van der Marel, G.
A.; van Boom, J. H. Tetrahedron Lett. 1989, 30, 5473–
5476.
16. For the regioselective monodeprotection of the nucleoside-
methylenebisphosphonate tribenzyl ester derivative: Ik-
eda, H.; Abushanab, E.; Marquez, V. E. Bioorg. Med.
Chem. Lett. 1999, 9, 3069–3074.
17. Daub, G. W.; Van Tamelen, E. E. J. Am. Chem. Soc. 1977,
99, 3526–3528.
18. (a) Creary, X. Org. Synth. 1985, 64, 207–216; (b) Liotta, L.
J.; Ganem, B. Tetrahedron Lett. 1989, 30, 4759–4762.
19. Saady, M.; Lebeau, L.; Mioskowski, C. Helv. Chim. Acta
1995, 78, 670–678.
20. Compound 24 was synthesised following the preparation
of 23 (Ref. 14a), by treatment of the common 60-O-TBDPS
precursor with Ph2NCOCl–DIPEA in pyridine (rt, 16 h,
50%) prior to desilylation with TBAF–AcOH in THF (rt,
1 h, 90%).
21. Simultaneous de-O-acylation and cleavage of the N,N-
dimethylformamidine N-protecting group was performed
with t-BuOK in t-BuOH–THF–H2O as recommended in
Ref. 14b.
22. Compound 1: 1H NMR (300 MHz, D2O): d 1.15–1.31
(10H, m, 5 · CH2), 1.51 (2H, quintet, J 6.8, –OCH2CH2–),
1.87–1.98 (4H, m, H-1*a, -1*b and –CH2CH@CH2),
8. All new compounds showed satisfactory analytical and
spectral data.
9. Compound 3: 1H NMR (300 MHz, D2O): d 1.16–1.31
(10H, m, 5 · CH2), 1.51 (2H, quintet, J 6.5, –OCH2CH2–),
1.85–2.07 (6H, m, H-1*a, -1*b, –CH2CH2NMe2 and
–CH2CH@CH2), 2.76 and 2.77 (6H, 2 · s, –NMe2), 3.23
(2H, m, –CH2NMe2), 3.44 (1H, dd, J2 ;3 9.8, H-20), 3.58
0
0
(1H, dd, J3 ;4 3.5, H-30), 3.60 (1H, m, H-5), 3.70–4.00
0
0
(11H, m, H-2, -3, -4, -40, -50, -6a, -6b, -60a, -60b and
0
0
–OCH2CH2–), 4.13 (1H, m, H-1), 4.35 (1H, d, J1 ;2 7.8,
H-10), 4.86 and 4.94 (2H, 2 · br d, J 10.4, J 17.2,
–CH2CH@CH2),
5.81
(1H,
ddt,
JH;CH2
6.5,
–CH2CH@CH2); 13C NMR (75 MHz, D2O): d 22.0 (d,
JC;P 133.7, –CH2CH2NMe2), 26.6 (d, JC;P 133.7, C-1*),
25.0, 28.1, 28.3, 28.4, 28.5 (5 · CH2), 30.1 (d, JC;P 5.8,
–OCH2CH2–), 33.1 (–CH2CH@CH2), 42.3 (–NMe2), 53.1
(–CH2NMe2), 60.3 (C-6), 63.1 (d, JC;P 4.5, C-60), 64.7

862
V. S. Borodkin et al. / Tetrahedron Letters 45 (2004) 857–862
2.11 (2H, br t, JH;P 19.9, –PCH2P–), 3.42 (2H, m, H-20 and
–OCH2CH2–), 68.0 (C-40), 69.3 (C-3), 70.5 (C-300), 70.9
(2C, C-20 and d, JC;P 8.5, C-2), 72.4 (C-30), 72.9 (C-5), 73.6
(br, C-1), 73.8 (C-200), 73.9 (br, C-50), 77.8 (C-4), 84.0 (d,
JC;P 6.1, C-400), 87.0 (C-100), 103.3 (C-10), 114.0
(–CH@CH2), 116.3 (br, C-5000), 137.8 (br, C-8000), 140.5 (–
CH@CH2), 151.9 (br, C-4000), 154.0 (C-2000), 158.9
(C-6000); 31P NMR (121 MHz, D2O): d 17.2, 17.5 (2 · br,
P0 and P00), 22.3 (P); ES-MS ()) data: m=z 489.67
-30), 3.65 (1H, dt, J4;5 8.7, J5;6a ¼ J5;6b ¼ 4.1, H-5), 3.65–3.95
[11H, includes d (at d 3.84), J3 ;4 2.6, H-40 and m,
H-2, -3, -4, -50, -6a, -6b, -60a, -60b and –OCH2CH2–], 4.07
(2H, m, H-500a and -500b), 4.15 (1H, m, H-1), 4.24 (1H, m,
0
0
H-400), 4.27 (1H, d, J1 ;2 7.7, H-10), 4.42 (1H, dd, J2
J3
5.1,
00 ;300
0
0
3.6, H-300), 4.70 (1H, dd, H-200), 4.86 and 4.94 (2H,
00 ;400
2 · br d, J 10.2, J 17.3, –CH2CH@CH2), 5.81 (1H, ddt,
JH;CH2 6.7, –CH2CH@CH2), 5.84 (1H, d, J1
5.9, H-100),
(50%, [M)2H]2ꢀ
(50%, [M)3H+Na]2ꢀ
(100%,
[M)3H+K]2ꢀ
)
(expected m=z 489.63), 500.69
(expected m=z 500.62), 508.65
(expected m=z 508.61;
00 ;200
8.09 (1H, br, H-8000); 13C NMR (75 MHz, D2O): d 26.9
(d, JC;P 133.1, C-1*), 25.0, 28.2, 28.3, 28.5, 28.6 (5 · CH2),
30.2 (d, JC;P 5.8, –OCH2CH2–), 33.2 (–CH2CH@CH2),
60.5 (C-6), 62.8 (br, C-60), 63.9 (br, C-500), 64.9 (d, JC;P 5.6,
)
)
22
C34H58N5O22P3 requires M, 981.28); ½aꢁ )2.5ꢁ (c 0.5,
D
MeOH–H2O, 1:1).