Synthesis of 3-Position-Modified Analogues of myo-Inositol 1,4,5-Trisphosphate, Tools for Investigation of the Polyphosphoinositide Pathway of Cellular Signaling

Article abstract of DOI:10.1021/jo970926y

Methods for the synthesis of 3-O-(carboxymethyl)- and 3-O-alkylated myo-inositol 1,4,5-trisphosphates in racemic form from myo-inositol have been devised. For DL-3-O-(carboxymethyl)-myo-inositol 1,4,5-trisphosphate, an analogue of myo-inositol 1,3,4,5-tet

Full text of DOI:10.1021/jo970926y

J . Org. Chem. 1997, 62, 8335-8340
8335
Syn th esis of 3-P osition -Mod ified An a logu es of m yo-In ositol
1
,4,5-Tr isp h osp h a te, Tools for In vestiga tion of th e
P olyp h osp h oin ositid e P a th w a y of Cellu la r Sign a lin g
Changsheng Liu and Barry V. L. Potter*
Department of Medicinal Chemistry, School of Pharmacy and Pharmacology, University of Bath,
Claverton Down, Bath BA2 7AY, U.K.
Received May 27, 1997X
Methods for the synthesis of 3-O-(carboxymethyl)- and 3-O-alkylated myo-inositol 1,4,5-trisphos-
phates in racemic form from myo-inositol have been devised. For DL-3-O-(carboxymethyl)-myo-
inositol 1,4,5-trisphosphate, an analogue of myo-inositol 1,3,4,5-tetrakisphosphate, DL-3-O-allyl-
2
,6-di-O-benzyl-1-O-(p-methoxybenzyl)-4,5-O-isopropylidene-myo-inositol (14) was prepared from
myo-inositol in seven steps. The triol DL-3-O-allyl-2,6-di-O-benzyl-myo-inositol (26), which was
obtained after treatment of 14 with acid, was phosphitylated and the product oxidized to give the
fully protected trisphosphate 27. The efficient oxidative cleavage of the 3-O-allyl ether of 27 in
the presence of the cyanoethyl-protected phosphate triesters was achieved by treatment of 27 with
NaIO
4 3
/RuCl ‚hydrate to afford the fully protected 3-O-(carboxymethyl) trisphosphate 28. After
deblocking, DL-3-O-(carboxymethyl) trisphosphate 6 was obtained. For DL-3-O-alkylated myo-inositol
1
1
,4,5-trisphosphate analogues, the fully protected 14 was isomerized to the cis-prop-1-enyl derivative
5. The propenyl group was removed to give DL-2,6-di-O-benzyl-1-O-(p-methoxybenzyl)-4,5-
isopropylidene-myo-inositol (16). The 3-O-methyl ether 17, 3-O-ethyl ether 18, and 3-O-n-propyl
ether 19 derivatives were synthesized by treatment of the anion of 16 with methyl iodide, ethyl
iodide, or n-propyl iodide, respectively. Removal of the isopropylidene and p-methoxybenzyl groups
afforded 3-O-alkylated triols 20, 21, or 22, which were phosphitylated and the products oxidized to
give the respective fully protected 3-O-alkylated trisphosphates 23-25. Deprotection furnished
3
-O-methyl- (3), 3-O-ethyl- (4), or 3-O-n-propyl-myo-inositol 1,4,5-trisphosphate (5). These
compounds will be useful pharmacological tools to explore the interaction of myo-inositol 1,4,5-
trisphosphate with its receptor and metabolic enzymes.
In tr od u ction
As a second messenger, D-myo-inositol 1,4,5-trisphos-
2
+
phate [Ins(1,4,5)P
3
1
(1)], which releases Ca from an
intracellular store via an isolated,3 cloned,4 and se-
,2
5
quenced receptor, is now well established (Figure 1). Ins-
(
1,4,5)P
deactivation by a 5-phosphatase to Ins(1,4)P
phosphorylation by a 3-kinase to the tetrakisphosphate
Ins(1,3,4,5)P (2). The function of the latter remains
3
is metabolized primarily via two pathways:6
2
or by
4
F igu r e 1. D-myo-Inositol 1,4,5-trisphosphate and 3-position
modified analogues.
controversial.7 Recently, the identification of an Ins-
(
1,3,4,5)P
3
binding protein8 and the suggestion that it
receptor9 have further spread
,10
may be an Ins(1,3,4,5)P
3
3 4
(1,4,5)P and Ins(1,3,4,5)P with their receptors and
metabolic enzymes11 and, additionally, the rational de-
sign of agonists, antagonists, and enzyme inhibitors.
Recent progress in inositol phosphate chemistry has been
reviewed.1
interest in the potential cellular functions of this com-
pound. Major challenges are now the elucidation of
structure-activity relationships for the interaction of Ins-
1,12
*
To whom correspondence should be addressed. Tel.: 01225-826639.
To investigate the role of the crucial hydroxyl group
Fax: 01225-826114. E-mail: B.V.L.Potter@bath.ac.uk.
at the 3-position of Ins(1,4,5)P
tion by Ins(1,4,5)P 3-kinase, a number of C-3-modified
D-myo-inositol analogues have been synthesized and
3
, the site of phosphoryla-
X
Abstract published in Advance ACS Abstracts, November 1, 1997.
(1) Berridge, M. J . Annu. Rev. Biochem. 1987, 56, 159-193.
(2) Berridge, M. J .; Irvine, R. F. Nature (London) 1989, 341, 197.
(3) Supattapone, S.; Worley, P. F.; Baraban, J . M.; Snyder, S. H. J .
3
13-17
18
tested in previous work.
3
D-3-Deoxy-Ins(1,4,5)P ,
Biol. Chem. 1988, 263, 1530-1534.
4) Furuichi, T.; Yoshikawa, S.; Miyawaki, A.; Waeda, A.; Mada, N.;
Mikoshiba, K. Nature (London) 1989, 342, 32-38.
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J . Biol. Chem. 1990, 265, 12679-12685.
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0, 139-144.
7) Irvine, R. F. In Advances in Second Messenger and Phospho-
(
(11) Potter, B. V. L.; Nahorski, S. R. Biochem. Soc. Trans. 1992,
20, 434-442.
(
(12) Potter, B. V. L.; Lampe, D. Angew. Chem., Int. Ed. Engl. 1995,
34, 1933-1972.
(
1
(13) Kozikowski, A. P.; Fauq, A. H.; Rusnak, J . M. Tetrahedron Lett.
1989, 30, 3365-3368.
(
protein Research; Putney, J . W., J r., Ed.; Raven Press: New York, 1992;
pp 161-186.
(14) Kozikowski, A. P.; Fauq, A. H.; Powis, G.; Melder, D. C. Med.
Chem. Res. 1991, 1, 277-282.
(8) Cullen, P. J .; Hsuan, J . J .; Truong, O.; Letcher, A. J .; J ackson,
(15) Powis, G.; Aksoy, I. A.; Melder, D. C.; Aksoy, S.; Eichinger, H.;
Fauq, A. H.; Kozikowski, A. P. Cancer Chemother. Pharmacol. 1991,
29, 95-104.
T. R.; Dawson, A. P.; Irvine, R. F. Nature (London) 1995, 376, 527-
5
30.
(9) Cullen, P. J .; Dawson, A. P.; Irvine, R. F. Biochem. J . 1995, 305,
(16) Kozikowski, A. P.; Powis, G.; Gallegos, A.; T u¨ ckmantel, W.
Bioorg. Med. Chem. Lett. 1993, 3, 1323-1326.
(17) Kozikowski, A. P.; Fauq, A. H.; Malaska, M. J .; T u¨ ckmantel,
W.; Ognyanov, V. I.; Powis, G. Curr. Med. Chem. 1994, 1, 1-12.
1
39-143.
(10) Cullen, P. J .; Chung, S.-K.; Chang, Y.-T.; Dawson, A. P.; Irvine,
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S0022-3263(97)00926-2 CCC: $14.00 © 1997 American Chemical Society

8
336 J . Org. Chem., Vol. 62, No. 24, 1997
Liu and Potter
D-3-C-(trifluoromethyl)-Ins(1,4,5)P
P ,
3
3
,19 3-halo-Ins(1,4,5)-
Resu lts a n d Discu ssion
1
7,20,21
and L-chiro-inositol 2,3,5-trisphosphate,22-24 an
Synthesis of the target 3-modified compounds required
initially the design of suitably protected inositol deriva-
tives. Conversion of the racemic inositol 1,2:4,5 diketal
, prepared from myo-inositol (7), to the corresponding
-O-allyl 9 and 3-O-allyl-6-O-benzyl 10 derivatives was
Ins(1,4,5)P analogue with an axial rather than an
3
equatorial 3-hydroxy group, have also been synthesized
and shown to have interesting properties. To explore
further the steric constraints at the receptor and meta-
bolic enzymes with respect to the 3-position, our first
target was to obtain the DL-3-O-alkylated myo-inositol
33
8
3
34
achieved by treatment of 8 first with allyl bromide/
barium oxide and barium hydroxide to give 9, which was
then treated with sodium hydride/benzyl bromide to give
the fully protected 10 (Scheme 1). Removal of the
isopropylidene groups afforded 11, and regioselective
introduction of a 1-O-p-methoxybenzyl ether in 11 was
achieved by treatment first with dibutyltin oxide in
refluxing toluene followed by reaction of the resulting
stannylene with cesium fluoride/p-methoxybenzyl bro-
mide to give 12. After reintroduction of the 4,5-O-
isopropylidene ketal giving 13, the remaining 2-hydroxyl
group was benzylated to produce 14. The allyl group of
14 was isomerized to propenyl using the rhodium com-
1
,4,5-trisphosphate analogues, 3-O-methyl- (3), 3-O-ethyl-
(4), and 3-O-n-propyl-Ins(1,4,5)P
3
(5),25 since analogues
with substituents at C-3 of increasing steric bulk might
display variable activity with respect to receptor binding
and Ca2 release and also toward metabolic enzymes.
+
26
In the search for the structural requirements for
effective Ca2 release, several racemic analogues of Ins-
1,4,5)P , possessing multiple phosphate surrogate groups
such as tris(sulfate), tris(sulfonamide), tris(carboxym-
+
(
3
2
7,28
ethyl), and tris(methylphosphonate)
thesized and did not have any Ca2 -mobilizing or an-
have been syn-
+
plex [(Ph
3 3
)P] RhCl in the presence of diazabicyclo[2.2.2]-
tagonistic properties. The 4,5-dimethylene phosphonate
2
9
octane to give 15. Removal of the propenyl group by
treatment of 15 with mercuric chloride and mercuric
oxide in acetone/water afforded the key intermediate 16.
The fully protected 3-O-methyl ether 17, 3-O-ethyl ether
analogue of DL-Ins(4,5)P
2
,
DL-inositol 1-phosphate 4,5-
pyrophosphate,30 and the D-hexadeoxy-1,4,5-tris(meth-
3
1
ylenesulfonic acid) analogue of Ins(1,4,5)P
3
were also
found to be inactive. We believe that a more conservative
approach, rather than that involving replacement with
multiple phosphate surrogate groups, should be more
successful. Consequently, we have synthesized initially,
as an Ins(1,3,4,5)P analogue, the first example of an
4
inositol polyphosphate that possesses both phosphate
1
8, and 3-O-n-propyl ether 19 derivatives were synthe-
sized by treatment of the anion of 16 with methyl iodide,
ethyl iodide, or n-propyl iodide, respectively. The iso-
propylidene group and the 1-O-p-methoxybenzyl ether in
each case were successively cleaved by treatment of 17,
1
8, or 19 with refluxing hydrochloric acid to produce the
groups and a nonphosphate group surrogate, myo-inositol
respective triols 3-O-methyl- (20), 3-O-ethyl- (21), or 3-O-
n-propyl-2,6-di-O-benzyl-myo-inositol (22). Phosphityla-
tion of 20, 21, or 22 was effected using bis(2-cyanoethyl)
N,N-diisopropylphosphoramidite/tetrazole in dichlo-
1
,4,5-trisphosphate 3-O-methylenecarboxylate [3-CME
Ins(1,4,5)P (6)]. It is important to note that we do not
3
include phosphorothioates in this definition, since they
are well investigated (and several mixed phosphate/
35
romethane to afford the corresponding trisphosphites,
1
2
phosphorothioate ligands have been synthesized ) and
generally well recognized by inositol polyphosphate bind-
ing proteins. Preliminary reports of the syntheses of the
which were smoothly oxidized with t-BuOOH to the fully
protected trisphosphates 23, 24, or 25, respectively.
Treatment of 23, 24, or 25 each with sodium in liquid
ammonia yielded the target compounds, racemic 3, 4, or
3
-O-alkylated Ins(1,4,5)P
3
analogues25 and 3-CME-Ins-
3
2
(1,4,5)P
3
have appeared already, and we now report
5
, respectively, which were purified by ion-exchange
here full details of this work.
chromatography on Q-Sepharose, eluting with a gradient
of triethylammonium bicarbonate buffer and quantified
by the Briggs phosphate assay as their triethylammo-
nium salts.
The synthesis of racemic myo-inositol 1,4,5-trisphos-
3
phate 3-O-methylenecarboxylate [3-CME Ins(1,4,5)P (6)]
is outlined in Scheme 2. Treatment of the fully protected
compound 14 with refluxing hydrochloric acid produced
the key triol 3-O-allyl-2,6-di-O-benzyl-myo-inositol (26).
The fully protected corresponding trisphosphate 27 was
obtained in a manner similar to that for 23 from 20. The
efficient oxidative cleavage of the 3-O-allyl ether of 27
in the presence of the cyanoethyl-protected phosphate
(
18) Seewald, M. J .; Aksoy, I. A.; Powis, G.; Fauq, A. H.; Kozikowski,
A. P. J . Chem. Soc., Chem. Commun. 1990, 1638-1639.
19) Kozikowski, A. P.; Ognyanov, V. I.; Fauq, A. H.; Wilcox, R. A.;
Nahorski, S. R. J . Chem. Soc., Chem. Commun. 1994, 599-600.
20) Kozikowski, A. P.; Fauq, A. H.; Aksoy, I. A.; Seewald, M. J .;
Powis, G. J . Am. Chem. Soc. 1990, 112, 7403-7404.
21) Fauq, A. H.; Kozikowski, A. P.; Ognyanov, V. I.; Wilcox, R. A.;
Nahorski, S. R. J . Chem. Soc., Chem. Commun. 1994, 1301-1302.
22) Liu, C.; Nahorski, S. R.; Potter, B. V. L. J . Chem. Soc., Chem.
Commun. 1991, 1014-1016.
23) Liu, C.; Nahorski, S. R.; Potter, B. V. L. Carbohydr. Res. 1992,
34, 107-115.
24) Ward, S. G.; Mills, S. J .; Liu, C.; Westwick, J .; Potter, B. V. L.
(
(
(
(
(
2
(
J . Biol. Chem. 1995, 270, 12075-12084.
(
25) Liu, C.; Potter, B. V. L. Tetrahedron Lett. 1994, 35, 8457-8460.
triesters was achieved by treatment of 27 with NaIO
4
/
(26) Hirata, M.; Watanabe, Y.; Kanematsu, T.; Ozaki, S.; Koga, T.
Biochim. Biophys. Acta 1995, 1244, 404-410.
27) Willems, H. A. M.; Veeneman, G. H.; Westerduin, P. Tetrahe-
dron Lett. 1992, 33, 2075-2078.
28) Westerduin, P.; Willems, H. A. M.; van Boeckel, C. A. A.
Carbohydr. Res. 1992, 234, 131-140.
29) Lyssikatos, J . P.; Bednarski, M. D. Bioorg. Med. Chem. Lett.
993, 3, 685-688.
30) Noble, N. J .; Dubreuil, D.; Potter, B. V. L. Bioorg. Med. Chem.
Lett. 1992, 2, 471-476.
31) Kozikowski, A. P.; Ognyanov, V. I.; Chen, C.; Kurian, P.; Crews,
F. T. Tetrahedron Lett. 1993, 34, 219-222.
32) Liu, C.; Thomas, N. F.; Potter, B. V. L. J . Chem. Soc., Chem.
Commun. 1993, 1687-1689.
33) Gigg, J .; Gigg, R.; Payne, S.; Conant, R. Carbohydr. Res. 1985,
42, 132-134.
34) Gigg, J .; Gigg, R.; Payne, S.; Conant, R. J . Chem. Soc., Perkin
Trans. 1 1987, 423-429.
RuCl
3
‚hydrate36 to afford 28. The cyanoethyl and benzyl
(
protecting groups of 28 were subsequently removed by
treatment with sodium in liquid ammonia to provide
crude 6, which was subjected to ion-exchange chroma-
tography on Q-Sepharose eluting with a gradient of
triethylammonium bicarbonate buffer to give the pure
(
(
1
(
Ins(1,3,4,5)P
4
analogue 6.
The Ca2 releasing properties of 3-5, as well as their
interaction with the metabolic enzymes Ins(1,4,5)P
(
+
(
3
(
(35) Cooke, A. M.; Potter, B. V. L.; Gigg, R. Tetrahedron Lett. 1987,
28, 2305-2308.
(36) Carlsen, P. H. J .; Katsuki, T.; Martin, V. S.; Sharpless, K. B.
J . Org. Chem. 1981, 46, 3937-3938.
1
(

Polyphosphoinositide Pathway of Cellular Signaling
J . Org. Chem., Vol. 62, No. 24, 1997 8337
Sch em e 1a
Sch em e 2a
a
Reagents and conditions: (i) 1 M HCl, MeOH, reflux; (ii) (a)
i
Pr 2NP(OCH2CH2CN)2, tetrazole, CH2Cl2, (b) 70% t-BuOOH; (iii)
NaIO4, RuCl3‚XH2O, CCl4/acetonitrile/H2O; (iv) (a) Na/liq NH3, (b)
H2O. Bn ) benzyl, PMB ) p-methoxybenzyl, All ) allyl. All
compounds are racemic.
than the apparent K
had a K for 3-kinase inhibition some 7-fold higher than
the apparent K for Ins(1,3,4,5)P Clearly, if the L-
i 3
for Ins(1,4,5)P . Compound 3 also
i
i
4
.
enantiomers of these mixtures are inactive, as ex-
pected,6 then the true potencies of these compounds as
receptor ligands and enzyme inhibitors are even more
marked.
,37
Racemic 3-CME-Ins(1,4,5)P
mobilizing agonist in permeabilized neuroblastoma cells
with a potency similar to that of Ins(1,3,4,5)P
.50 µM; cf 2.5 µM for D-Ins(1,3,4,5)P ]. It had a 6-fold
higher K for Ins(1,4,5)P 3-kinase than Ins(1,3,4,5)P but
was twice as potent at binding to Ins(1,4,5)P 5-phos-
(6) was found to be a Ca2+-
3
4
[EC50 )
3
4
i
3
4
3
a
Reagents and conditions: (i) (a) CH3C(OCH3)2CH3, PTSA,
phatase and was an inhibitor of this enzyme. Presum-
ably, only the D-enantiomer is recognized by these
proteins.
reflux, (b) BzCl, pyridine, (c) NaOH, reflux; (ii) allyl bromide, BaO,
Ba(OH)2, DMF; (iii) BnCl, NaH, DMF; (iv) PTSA, ethyl acetate/
acetone/water; (v) (a) dibutyltin oxide, toluene, reflux, (b) CsF, (p-
MeO)BnCl, DMF; (vi) 2-methoxypropene, PTSA, DMF; (vii) BnCl,
NaH, DMF; (viii) diazabicyclo[2.2.2]octane, [(Ph)3P]3RhCl, ethanol/
benzene/water; (ix) HgCl2, HgO, acetone/water; (x) MeI, or EtI or
Analogues derived from inversion,23 deletion,38 and
20,39
fluorination
of the crucial 3-hydroxy group have
demonstrated little loss of Ca2 -mobilizing activity rela-
+
n
i
Pr I, NaH, DMF; (xi) 1 M HCl MeOH, reflux; (xii) (a) Pr 2NP-
OCH2CH2CN)2, tetrazole, CH2Cl2, (b) 70% t-BuOOH; (xiii) (a) Na/
tive to Ins(1,4,5)P , especially in the latter two cases. The
3
(
liq NH3, (b) H2O. Bn ) benzyl, PMB ) p-methoxybenzyl, All )
present work is the first report to demonstrate that bulky
substituents of a hydrophobic nature at the 3-position
are reasonably well tolerated when R ) Me, but not when
R > Me. The situation is alleviated, however, in the case
of substitution by an O-carboxymethyl32 that, though
allyl, Prop ) propenyl. All compounds are racemic.
5
-phosphatase and 3-kinase, have been examined in
permeabilized SH SY5Y cells relative to Ins(1,4,5)P and
Ins(1,3,4,5)P . All three 3-O-alkylated analogues were
evaluated as Ca2 -mobilizing agonists, and compound 3
was found to be essentially equipotent to Ins(1,3,4,5)P
3
4
+
(37) Potter, B. V. L.; Nahorski, S. R. In Drug Design for Neuro-
science; Kozikowski, A. P., Ed.; Raven Press: New York, 1993; pp 383-
4
,
4
16.
but relative EC50’s increased markedly in the order of
(38) Kozikowski, A. P.; Ognyanov, V. I.; Chen, C.; Fauq, A. H.;
Safrany, S. T.; Wilcox, R. A.; Nahorski, S. R. J . Med. Chem. 1993, 36,
035-3038.
39) Safrany, S. T.; Wilcox, R. A.; Liu, C.; Potter, B. V. L.; Nahorski,
S. R. Eur. J . Pharmacol. 1992, 226, 265-272.
increasing 3-position chain length, i.e., R ) Me < Et <
3
n
Pr . They were all potent 5-phosphatase inhibitors with
(
3
3 i
-O-methyl Ins(1,4,5)P having a K some 5-fold lower

8
338 J . Org. Chem., Vol. 62, No. 24, 1997
Liu and Potter
relatively bulky, will bear a negative charge at physi-
ological pH. These data support the use of the car-
boxymethyl group as a phosphate group surrogate. Full
biological results for new synthetic compounds will be
reported elsewhere. These compounds should find ap-
plications as new tools for probe the polyphosphoinositide
pathway of cellular signaling.
DL-3-O-Allyl-6-O-ben zyl-m yo-in ositol (11). A solution of
0 (9 g, 23.05 mmol) in ethyl acetate/acetone/water (80 mL:
0 mL:8 mL, v:v:v) was stirred with p-toluenesulfonic acid (1.2
g, 6.3 mmol) at 40 °C for 5 h. The solvents were evaporated
in vacuo, and the resulting solid was crystallized from ethyl
1
8
acetate to give compound 11 (6.32 g, 20.4 mmol, 88.4%): mp
1
1
52-153 °C; H NMR (DMSO) δ 2.99 (dd, 1H, J ) 2.4 Hz, 9.7
Hz), 3.11 (ddd, 1H, J ) 4.9 Hz, 9.0 Hz), 3.32 (ddd, 1H, J ) 2.4
Hz, 7.9 Hz, 9.7 Hz), 3.41 (dd, 1H, J ) 9.1 Hz), 3.53 (dd, 1H, J
)
4.7 Hz, 9.3 Hz), 3.90 (ddd, 1H, J ) 2.9 Hz, 4.2 Hz), 4.05-
Exp er im en ta l Section
4
.15 (m, 2H), 4.63 (d, 1H, J ) 6.8 Hz), 4.68 (d, 1H, J ) 8.0
Hz), 4.68 (s, 2H), 4.76 (d, 1H, J ) 3.1 Hz), 4.79 (d, 1H, J ) 4.9
Hz), 5.08-5.34 (m, 2H), 5.85-5.99 (m, 1H), 7.20-7.44 (m, 5H);
MS m/z (EI) 310 (M, 9), 107 (13), 91 (100), 85 (40), 41 (54).
Chemicals were purchased from Aldrich, Fluka, and Lan-
caster (U.K.). Dichloromethane, triethylamine, and dimeth-
ylformamide were dried over calcium hydride, distilled, and
stored over 4A molecular sieves. TLC was performed on
precoated plates (Merck TLC aluminum sheets silica 60F254
art. 5554). Products were visualized by spraying phosphomo-
lybdic acid in methanol followed by heating. Flash chroma-
Anal. Calcd for C16
H, 7.18.
22 6
H O : C, 61.92; H, 7.14. Found: C, 62.00;
DL-3-O-Allyl-6-O-b en zyl-1-O-(p -m et h oxyb en zyl)-m yo-
in ositol (12). A mixture of 11 (9.5 g, 30.64 mmol) and
dibutyltin oxide (11.44 g, 45.96 mmol) in dry toluene (200 mL)
was refluxed for 3 h in a Dean-Stark apparatus to remove
water, and the solvent was then removed in vacuo. To the
residue was added cesium fluoride (9.2 g, 61.33 mmol), and
the mixture was suspended in dry DMF (200 mL) followed by
addition of p-methoxybenzyl chloride (4.3 mL, 36 mmol). The
mixture was stirred at room temperature overnight. Water
tography was carried out using Sorbsil C60 silica gel. For 31
NMR spectra chemical shifts were measured in ppm relative
to external 85% H PO and are positive when downfield from
P
3
4
this reference. Melting points (uncorrected) were determined
using a Kofler block. Ion-exchange chromatography was
performed on a LKB-Pharmacia medium-pressure ion-ex-
change chromatograph using Q Sepharose Fast Flow with a
gradient of triethylammonium bicarbonate (TEAB) as eluent.
Column fractions containing inositol polyphosphate analogues
were assayed for phosphate by a modification of the Briggs
(300 mL) was added, the aqueous mixture was then extracted
with chloroform (2 × 300 mL), and the organic layer was
evaporated to a gum, which was crystallized from ethyl
acetate/petroleum ether to give the compound 12 (9.7 g, 22.7
4
0
41
test as described.
1
mmol, 74%): mp 132-133 °C; H NMR (CDCl
3
) δ 2.37 (b, 3H),
DL-3-O-Allyl-1,2;4,5-di-O-isopr opyliden e-myo-in ositol (9).
3
3
3
(
)
(
.16 (dd, 1H, J ) 2.7 Hz, 9.5 Hz), 3.41 (dd, 1H, J ) 9.3 Hz),
To a suspension of 1,2;4,5-di-O-isopropylidene-myo-inositol33
.41 (dd, 1H, J ) 2.8 Hz, 9.5 Hz), 3.80 (dd, 1H, J ) 9.5 Hz),
.81 (s,3H), 3.92 (dd, 1H, J ) 9.5 Hz), 4.07-4.24 (m, 2H), 4.23
(
8) (13.02 g, 50 mmol), barium oxide (15.35 g, 100 mmol), and
barium hydroxide octahydrate (1.98 g, 6.25 mmol) in dry DMF
250 mL) was added allyl bromide (6.74 g, 4.82 mL, 55.7 mmol)
dd, 1H, J ) 2.8 Hz), 4.65 (s, 2H), 4.77 and 4.95 (AB, 2H, J AB
(
11.2 Hz), 5.19-5.33 (m, 2H), 5.87-6.01 (m, 1H), 6.84-6.89
m, 2H), 7.26-7.37 (m, 7H); MS m/z (EI) 429 (M, 4), 339 (89),
09 (47), 137 (16), 121 (100), 107 (11), 91 (16). Anal. Calcd
for C24 : C, 66.98; H, 7.03. Found: C, 66.80; H, 7.14.
dropwise at room temperature. The reaction mixture was
stirred for 60 h at room temperature, and methanol (10 mL)
was added. The resulting mixture was neutralized with AcOH/
H O (1:1, v:v) and concentrated in vacuo. The residue was
2
taken up in chloroform and washed successively with water,
a saturated solution of sodium hydrogen carbonate, and again
3
30 7
H O
DL-3-O-Allyl-6-O-ben zyl-1-O-(p-m eth oxyben zyl)-4,5-O-
isop r op ylid en e-m yo-in ositol (13). A solution of 12 (3.12 g,
7
.25 mmol), p-toluenesulfonic acid (0.25 g), and 2-methoxypro-
4
water. The organic layer was dried over MgSO and concen-
pene (2.12 mL, 22.5 mmol) in dry DMF (75 mL) was stirred at
room-temperature overnight and then neutralized with am-
monia. The DMF was evaporated in vacuo (<50 °C), and the
residue was partitioned between water and chloroform (200
mL, each). The organic layer was dried (MgSO ) and evapo-
4
rated to dryness, and the crude product was obtained after
flash column chromatography (ethyl acetate/petroleum ether
trated in vacuo. The final product was obtained after flash
column chromatography (ethyl acetate/petroleum ether, 3:7)
and then crystallization to give compound 9 (9.47 g, 31.5 mmol,
6
3%): mp 128-130 °C (from petroleum ether bp 60-80 °C)
3
4
1
(
1
4
lit. mp 127-129 °C); H NMR (CDCl
3
) δ 1.39, 1.45, 1.47,
.55 (4s, 12H), 3.31 (dd, 1H, J ) 9.5 Hz), 3.51 (br s, 1H), 3.81-
.05 (m, 4H), 4.25-4.33 (m, 2H), 4.48 (t, 1H, J ) 4.4 Hz), 5.23
2
:8) and then crystallization (ether/petroleum ether) to give
(
2
6
d, 1H, J ) 10.3 Hz), 5.32 (m, 1H), 5.98 (m, 1H); MS m/z (EI)
1
the compound 13 (3.7 g, 6.5 mmol, 90%): mp 88.5 °C; H NMR
(CD
85 (6), 227 (1.4), 113 (100). Anal. Calcd for C15
0.00; H, 8.05. Found: C, 60.10; H, 8.15.
24 6
H O : C,
[
1
3
3
)
2
CO] δ 1.37 (s, 6H), 3.42 (dd, 1H, J ) 9.5 Hz), 3.42 (dd,
H, J ) 2.9 Hz, 8.8 Hz), 3.48 (dd, 1H, J ) 2.7 Hz, 10.3 Hz),
.76 (s, 3H), 3.78 (dd, 1H, J ) 9.5 Hz), 3.85 (dd, 1H, J ) 9.7
DL-3-O-Allyl-6-O-ben zyl-1,2;4,5-di-O-isopr opyliden e-myo-
in ositol (10). To a solution of 9 (12.7 g, 42.33 mmol) in dry
DMF (150 mL) was added sodium hydride (4.4 g, 4 equiv).
After 15 min, benzyl chloride (9.77 mL, 84.76 mmol) was
added, and the resulting mixture was stirred at room tem-
perature for 2 h. Methanol was added to destroy the excess
of sodium hydride, and the solution was partitioned between
water (300 mL) and chloroform (400 mL). The organic layer
was dried (MgSO ) and evaporated to dryness. The crude
4
product was crystallized from petroleum ether (40-60 °C) with
a few drops of dichloromethane to give compound 10 (16.5 g,
Hz), 4.00-4.14 (m, 2H), 4.25 (m, 1H), 4.47 and 4.62 (AB, 2H,
J
5
7
AB ) 11.3 Hz), 4.71 (s, 2H), 4.94 (d, 1H, J ) 4.4 Hz), 5.11-
.32 (m, 2H), 5.83-5.97 (m, 1H), 6.81-6.88 (m, 2H), 7.27-
.33 (m, 7H); MS m/z (EI) 471 (M, 9), 379 (80), 349 (90), 137
34 7
(11), 121 (100), 107 (11), 91 (19). Anal. Calcd for C27H O :
C, 68.92; H, 7.28. Found: C, 68.90; H, 7.30.
DL-3-O-Allyl-2,6-d i-O-b en zyl-1-O-(p -m et h oxyb en zyl)-
,5-O-isop r op ylid en e-m yo-in ositol (14). To a solution of 13
4
(
8.58 g, 18.22 mmol) in dry DMF (100 mL) was added sodium
hydride (1.72 g, 44.8 mmol). After 15 min, benzyl chloride
4.16 mL, 36.03 mmol) was added, and the resulting mixture
1
quantitative yield): mp 122-123 °C; H NMR (CDCl
3
) δ 1.36,
(
1
3
1
5
4
.39, 1.45, 1.46 (4s, 12H), 3.38 (dd, 1H, J ) 9.3 Hz, 10.5 Hz),
was stirred at room temperature for 2 h. Methanol was added
to destroy the excess of sodium hydride, and the solution was
partitioned between water (300 mL) and chloroform (500 mL).
The organic layer was dried (MgSO ) and evaporated to
4
dryness. The crude product was purified after flash column
chromatography (ethyl acetate/petroleum ether, 1:9) and
.68 (dd, 1H, J ) 6.5 Hz, 10.6 Hz), 3.79 (dd, 1H, J ) 4.1 Hz,
0.1 Hz), 3.95 (dd, 1H, J ) 9.4 Hz, 10.0 Hz), 4.13 (dd, 1H, J )
.0 Hz, 6.5 Hz), 4.20-4.36 (m, 2H), 4.46 (dd, 1H, J ) 4.5 Hz),
.83 (s, 2H), 5.19-5.36 (m, 2H), 5.87-6.04 (m, 1H), 7.23-7.42
(
1
m, 5H); MS m/z (EI) 391 (M, 81), 333 (49), 217 (16), 131 (46),
07 (16), 91 (100). Anal. Calcd for C22 : C, 67.67; H, 7.74.
Found: C, 68.00; H, 7.86.
30 6
H O
crystallized from ethyl acetate/petroleum ether to give com-
1
pound 14 (9.56 g, 17.1 mmol, 94%): mp 78 °C; H NMR [(CD
3
)
2
-
CO] δ 1.37, 1.38 (2s, 6H), 3.46 (dd, 1H, J ) 9.5 Hz), 3.60 (dd,
1
H, J ) 2.8 Hz, 9.0 Hz), 3.65 (dd, 1H, J ) 2.6 Hz, 10.3 Hz),
(
(
40) Briggs, A. P. J . Biol. Chem. 1922, 53, 13.
41) Lampe, D.; Liu, C.; Potter, B. V. L. J . Med. Chem. 1994, 37,
3.74 (s, 3H), 3.81 (dd, 1H, J ) 9.5 Hz), 3.86 (dd, 1H, J ) 10.3
Hz), 4.00-4.14 (m, 2H), 4.28 (dd, 1H, J ) 2.7 Hz), 4.54 and
9
07-912.

Polyphosphoinositide Pathway of Cellular Signaling
J . Org. Chem., Vol. 62, No. 24, 1997 8339
4
1
6
7
1
.65 (AB, 2H, J AB ) 11.4 Hz), 4.71 and 4.75 (AB, 2H, J AB
)
J ) 9.5 Hz), 4.07 (dd, 1H, J ) 2.2 Hz), 4.59 (s, 2H), 4.78 and
4.8 Hz), 4.75 (s, 2H), 5.13-5.34 (m, 2H), 5.84-5.98 (m, 1H),
.86-6.91 (m, 2H), 7.23-7.37 (m, 12H); MS m/z (EI) 561 (M,
), 503 (5), 469 (82), 439 (96), 381 (12), 359 (15), 121 (100),
4.96 (AB, 2H, J AB ) 11.2 Hz), 4.83 and 4.85 (AB, 2H, J AB )
12.0 Hz), 6.82-6.89 (m, 2H), 7.22-7.41 (m, 12H); MS m/z (EI)
549 (M, 2), 457 (10), 427 (15), 121 (100), 91 (29). Anal. Calcd
for C33H O : C, 72.24; H, 7.35. Found: C, 72.2; H, 7.39.
40 7
07 (15), 91 (35). Anal. Calcd for C34
H
40
O
7
: C, 72.83; H, 7.19.
Found: C, 72.20; H, 7.22.
DL-3-O-P r op yl-2,6-d i-O-ben zyl-1-O-(p-m eth oxyben zyl)-
4,5-O-isop r op ylid en e-m yo-in ositol (19). The above com-
pound was prepared in a fashion identical to that described
DL-3-O-(P r op -1-en yl)-1-O-(p-m eth oxyben zyl)-2,6-d i-O-
ben zyl-4,5-O-isop r op ylid en e-m yo-in ositol (15). To a solu-
tion of 14 (1.5 g, 2.68 mmol) in ethanol-benzene-water (100
mL, 7:3:1) was added diazabicyclo[2.2.2]octane (0.066 g, 0.6
mmol), and the mixture was heated to reflux followed by
n
1
for 17 as an oil, except that Pr I was used in place of MeI:
H
NMR (CDCl ) δ 0.92 (t, 3H, J ) 7.4 Hz), 1.43, 1.45 (2s, 6H),
3
1.63 (td, 2H, J ) 7.2 Hz), 3.34-3.62 (m, 5H), 3.80 (s, 3H), 4.02-
4.10 (m, 3H), 4.58 and 4.64 (AB, 2H, J AB ) 11.5 Hz), 4.78 and
4.90 (AB, 2H, J AB ) 11.5 Hz), 4.83 (s, 2H), 6.81-6.88 (m, 2H),
7.22-7.44 (m, 12H); MS m/z (EI) 563 (M, 2), 471 (10), 441 (18),
279 (55), 205 (75), 149 (66), 121 (100), 91 (30), 69 (19).
DL-3-O-Meth yl-2,6-d i-O-ben zyl-m yo-in ositol (20). A so-
lution of 17 (0.1 g, 0.18 mmol) in 1 M HCl-MeOH (2:8, v/v,
10 mL) was refluxed for 3 h, and the cooled solution was
quenched with ammonia. The solvents were evaporated in
vacuo, and the residue was partitioned between water and
chloroform (30 mL, each). The organic layer was dried
3 3
adding rhodium complex [(Ph )P] RhCl (0.19 g, 0.21 mmol).
After further reflux for 1 h, the cooled mixture was extracted
with ether (2 × 100 mL), and the organic layer was dried
4
(MgSO ) and purified by flash column chromatography to give
an oil that was crystallized from petroleum ether to give the
compound 15 (1.4 g, 2.5 mmol, 93%): mp 96-98 °C; H NMR
1
(
3
CDCl ) δ (cis) 1.43, 1.46 (2s, 6H), 1.63 (dd, 3H, J ) 1.7 Hz,
6
9
4
.8 Hz), 3.40 (dd, 1H, J ) 9.5 Hz), 3.41 (dd, 1H, J ) 2.2 Hz,
.7 Hz), 3.73 (dd, 1H, J ) 2.7 Hz, 10.4 Hz), 3.80 (s, 3H), 4.02-
.08 (m, 2H), 4.14 (dd, 1H, J ) 10.1 Hz), 4.49 (dq, 1H, J ) 6.7
Hz), 4.57 and 4.63 (AB, 2H, J AB ) 11.5 Hz), 4.81 (s, 2H), 4.78
4
(MgSO ) and evaporated to dryness, and the crude product was
and 4.90 (AB, 2H, J AB ) 11.5 Hz), 6.12 (dd, 1H, J ) 1.8 Hz,
purified after flash column chromatography to give the com-
1
6
5
.4 Hz), 6.81-6.84 (m, 2H), 7.20-7.43 (m, 12H); MS m/z (EI)
61 (M, 2), 503 (2), 469 (13), 439 (6), 251 (10), 211 (25), 121
pound 20 (0.064 g, 0.17 mmol, 95%): mp 116 °C; H NMR
3
(CDCl ) δ 2.45 (br s, 1H), 3.00 (br, 2H), 3.01 (dd, 1H, J ) 2.4
(100), 91 (65), 69 (33). Anal. Calcd for C34
H
40
O
7
: C, 72.83;
Hz, 9.7 Hz), 3.41 (s, 3H), 3.46 (dd, 1H, J ) 9.0 Hz), 3.51 (dd,
1H, J ) 2.8 Hz, 10.1 Hz), 3.68 (dd, 1H, J ) 9.3 Hz), 3.93 (dd,
1H, J ) 9.4 Hz), 4.11 (dd, 1H, J ) 2.5 Hz), 4.71 and 4.91 (AB,
2H, J AB ) 11.5 Hz), 4.81 and 4.90 (AB, 2H, J AB ) 11.4 Hz),
7.26-7.77 (m, 10H); MS m/z (EI) 373 (M, 3), 283 (44), 195 (33),
181 (78), 159 (15), 107 (35), 91 (100). Anal. Calcd for
H, 7.19. Found: C, 73.10; H, 7.30.
DL-1-O-(p -Met h oxyb en zyl)-2,6-d i-O-b en zyl-4,5-O-iso-
p r op ylid en e-m yo-in ositol (16). To a solution of mercuric
chloride (0.63 g, 2.31 mmol) in acetone-water (7 mL, 10:1)
was added dropwise with stirring a mixture of 15 (1.3 g, 2.32
mmol), mercuric oxide (0.63 g), and acetone-water (20 mL,
21 26 6
C H O : C, 67.38; H, 6.97. Found: C, 67.4; H, 7.03.
DL-3-O-Eth yl-2,6-d i-O-ben zyl-m yo-in ositol (21).
1
0:1) during 3 min. After a further 5 min, the mercuric oxide
was removed by filtration through Celite, the acetone was
evaporated in vacuo, and ether was added to the residue. The
ether layer was washed with a semisaturated aqueous solution
The above compound was prepared in a fashion similiar to
1
that described for 20: mp 115 °C; H NMR (CDCl
3
) δ 1.24 (t,
3H, J ) 7.1 Hz), 2.40 (br, 3H), 3.12 (dd, 1H, J ) 2.3 Hz, 9.7
Hz), 3.48 (dd, 1H, J ) 9.1 Hz), 3.50 (dd, 1H, J ) 9.1 Hz), 3.53
(dd, 1H, J ) 2.7 Hz, 9.0 Hz), 3.62-3.74 (m, 2H), 3.93 (dd, 1H,
J ) 9.4 Hz), 4.08 (dd, 1H, J ) 2.6 Hz), 4.69 and 4.93 (AB, 2H,
of potassium iodide (10 mL), dried (MgSO
The crude product was chromatographed on a flash column
light petroleum/ether 2:1), followed by crystallization from
petroleum ether/ethyl acetate to give the compound 16 (0.9 g,
4
), and evaporated.
(
J
AB ) 11.5 Hz), 4.87 and 4.88 (AB, 2H, J AB ) 11.4 Hz), 7.26-
7.40 (m, 10H); MS m/z (EI) 387 (M, 3), 209 (36), 191 (20), 181
1
1
2
.7 mmol, 75%); mp 99 °C; H NMR (CDCl
3
) δ 1.43 (2s, 6H),
.35-2.45 (br s 1H), 3.38 (dd, 1H, J ) 9.5 Hz), 3.49 (dd, 1H,
(73), 107 (38), 91 (100). Anal. Calcd for C22
H, 7.26. Found: C, 67.9; H, 7.29.
28 6
H O : C, 68.00;
J ) 2.9 Hz, 9.5 Hz), 3.63 (dd, 1H, J ) 2.7 Hz, 9.6 Hz), 3.73
dd, 1H, J ) 9.5 Hz), 3.79 (s, 3H), 3.91 (dd, 1H, J ) 3.0 Hz),
.00 (dd, 1H, J ) 9.5 Hz), 4.58 and 5.06 (AB, 2H, J AB ) 11.2
Hz), 4.63 and 4.93 (AB, 2H, J AB ) 11.7 Hz), 4.75 and 4.79 (AB,
H, J AB ) 11.5 Hz), 6.83-6.86 (m, 2H), 7.23-7.42 (m, 12H);
MS m/z (EI) 521 (M, 2), 429 (6), 399 (8), 137 (20), 121 (100),
07 (33), 91 (32), 69 (14). Anal. Calcd for C31 : C, 71.52;
H, 6.97. Found: C, 71.5; H, 6.95.
(
4
DL-3-O-n -P r op yl-2,6-d i-O-ben zyl-m yo-in ositol (22). The
above compound was prepared in a fashion similar to that
1
described for 20: mp 112 °C; H NMR (CDCl
3
) δ 0.95 (t, 3H,
2
J ) 7.4 Hz), 1.63 (td, 2H, J ) 7.2 Hz), 2.35 (d, 1H, J ) 6.8
Hz), 2.63 (br s, 1H), 2.65 (br s, 1H), 3.12 (dd, 1H, J ) 2.2 Hz,
9.7 Hz), 3.40 (dd, 1H, J ) 9.5 Hz), 3.49-3.60 (m, 3H), 3.67
(dd, 1H, J ) 9.3 Hz), 3.95 (dd, 1H, J ) 9.4 Hz), 4.09 (dd, 1H,
J ) 2.3 Hz), 4.68 and 4.94 (AB, 2H, J AB ) 11.5 Hz), 4.86 and
4.87 (AB, 2H, J AB ) 11.5 Hz), 7.26-7.40 (m, 10H); MS m/z
(EI) 401 (M, 2), 311 (42), 223 (32), 205 (16), 181 (72), 107 (39),
1
36 7
H O
DL-3-O-Meth yl-2,6-d i-O-ben zyl-1-O-(p-m eth oxyben zyl)-
,5-O-isop r op ylid en e-m yo-in ositol (17). To a solution of 16
4
(0.09 g, 0.17 mmol) in dry DMF (10 mL) was added sodium
hydride (0.24 g, 0.51 mmol). After 15 min, iodomethane (0.072
g, 0.51 mmol) was added, and the mixture was stirred at room
temperature for 2 h. Methanol was added to destroy the excess
of sodium hydride, and the solution was partitioned between
water (10 mL) and chloroform (20 mL). The organic layer was
91 (100). Anal. Calcd for C23
C, 68.7; H, 7.58.
DL-3-O-Meth yl-2,6-d i-O-ben zyl-m yo-in ositol 1,4,5-Tr is-
[bis(2-cya n oeth yl) p h osp h a te] (23). To a mixture of 20
(0.05 g, 0.13 mmol) and 1H-tetrazole (0.17 g, 2.4 mmol) in dry
30 6
H O : C, 68.64; H, 7.5. Found:
dried (MgSO
4
) and evaporated to dryness. The crude product
CH
phosphoramidite (0.5 g, 2.2 mL). The mixture was stirred at
room temperature for 1 h, t-BuOOH (0.5 mL, 70% in H O) was
added, and the resulting solution was stirred overnight and
then washed with saturated aqueous NaHCO (10 mL), dried
(MgSO ), and concentrated. Flash column chromatography of
the residue gave compound 23 (0.085 g, 0.09 mmol, 71%) as
2 2
Cl (5 mL) was added bis(2-cyanoethyl) N,N-diisopropyl-
was chromatographed on a flash column to give the compound
1
1
7 as an oil (0.09 g, 0.17 mmol, 98%): H NMR (CDCl
3
) δ 2.17
2
(
2s, 6H), 2.94 (dd, 1H, J ) 2.3 Hz, 9.5 Hz), 3.34 (s, 3H), 3.39
(
3
dd, 1H, J ) 2.4 Hz, 9.8 Hz), 3.44 (dd, 1H, J ) 9.2 Hz), 3.81 (s,
3
H), 3.90 (dd, 1H, J ) 9.5 Hz), 3.96 (dd, 1H, J ) 9.5 Hz), 4.11
4
(
)
(
dd, 1H, J ) 2.3 Hz), 4.61 (s, 2H), 4.76 and 4.98 (AB, 2H, J AB
an oil: 31P NMR (CDCl
) δ -3.43, -3.10; H NMR (CDCl ) δ
1
11.0 Hz), 4.79 and 4.88 (AB, 2H, J AB ) 12.2 Hz), 6.86-6.88
3
3
m, 2H), 7.26-7.41 (m, 12H); MS m/z (EI) 535 (M, 2%), 443
9), 413 (14), 121 (100), 107 (20), 91 (32), 69 (14).
2.40-2.90 (m, 12H), 3.35 (dd, 1H, J ) 2.1, 9.9 Hz), 3.45 (s,
(
3H), 4.03-4.59 (m, 16H), 4.76 (dd, 1H, J ) 9.3 Hz), 4.81 and
DL-3-O-E t h yl-2,6-d i-O-b en zyl-1-O-(p -m et h oxyb en zyl)-
4.92 (AB, 2H, J AB ) 11.5 Hz), 4.84 a nd 4.92 (AB, 2H, J AB )
+
4
,5-O-isop r op ylid en e-m yo-in ositol (18). The above com-
11.5 Hz), 7.09-7.65 (m, 10H); MS m/z (FAB) 933 [(M + H) ,
15], 889 (2), 663 (2), 619 (3), 591 (4) 181 (10), 149 (13), 105
(10), 91 (100).
pound was prepared in a fashion identical to that described
for 17 in crystallized form, except EtI was used in place of
MeI: mp 83 °C; H NMR (CDCl
1
3
) δ 1.21 (t, 3H, J ) 7.0 Hz),
DL-3-O-E t h yl-2,6-d i-O-b en zyl-m yo-in osit ol 1,4,5-Tr is-
[bis(2-cya n oeth yl) p h osp h a te] (24). The above compound
2
)
7
.17 (s, 6H), 3.02 (dd, 1H, J ) 2.4 Hz, 9.7 Hz), 3.38 (dd, 1H, J
3
1
2.4 Hz, 9.5 Hz), 3.44 (dd, 1H, J ) 9.2 Hz), 3.55 (q, 2H, J )
was prepared in a fashion similar to that described for 23:
P
1
.0 Hz), 3.81 (s, 3H), 3.90 (dd, 1H, J ) 9.4 Hz), 3.96 (dd, 1H,
NMR (CDCl
3
) δ -3.63, -3.37; H NMR (CDCl ) δ 1.24 (t, 3H,
3

8
340 J . Org. Chem., Vol. 62, No. 24, 1997
Liu and Potter
J ) 7.0 Hz), 2.42-2.90 (m, 12H), 3.44 (dd, 1H, J ) 2.2, 9.9
Hz), 3.60-3.73 (m, 2H), 4.04-4.56 (m, 16H), 4.74 (dd, 1H, J
) 11.4 Hz), 5.19-5.34 (m, 2H), 5.84-5.99 (m, 1H), 7.26-7.40
(m, 10H); MS m/z (EI) 399 (M, 3), 359 (2), 309 (41), 181 (51),
)
9.4 Hz), 4.81 and 4.92 (AB, 2H, J AB ) 11.5 Hz), 4.84 and
107 (40), 91 (100). Anal. Calcd for C23
Found: C, 68.9; H, 7.00.
28 6
H O : C, 68.98; H, 7.05.
4
.92 (AB, 2H, J AB ) 11.5 Hz), 7.27-7.54 (m, 10H); MS m/z
+
(FAB) 947 [(M + H) , 11], 934 (3), 889 (2), 227 (8), 149 (15),
DL-3-O-Allyl-2,6-d i-O-b en zyl-m yo-in osit ol 1,4,5-Tr is-
[bis(2-cya n oeth yl) p h osp h a te] (27). To a mixture of 26 (0.6
1
35 (10), 91 (100).
DL-3-O-P r op yl-2,6-d i-O-ben zyl-m yo-in ositol 1,4,5-Tr is-
bis(2-cya n oeth yl) p h osp h a te] (25). The above compound
g, 1.5 mmol) and 1H-tetrazole (0.97 g, 13.7 mmol) in dry CH
Cl (15 mL) was added bis(2-cyanoethyl) N,N-diisopropylphos-
phoramidite (3.6 g, 15.8 mmol). The mixture was stirred at
room temperature for 1 h, and t-BuOOH (5 mL, 70% in H O)
was added. The resulting solution was stirred overnight and
then washed with saturated aqueous NaHCO (50 mL), dried
(MgSO ), and concentrated. Flash column chromatography of
the residue gave compound 27 as an oil (1.15 g, 1.2 mmol,
2
-
[
2
3
1
was prepared in a fashion similar to that described for 23:
P
1
NMR (CDCl
3
) δ -3.23, -3.09, -3.03; H NMR (CDCl
3
) δ 0.96
2
(
1
)
4
t, 3H, J ) 7.4 Hz), 1.67 (td, 2H, J ) 7.2 Hz), 2.40-2.90 (m,
2H), 3.49-3.60 (m, 3H), 4.00-4.50 (m, 16H), 4.74 (dd, 1H, J
9.4 Hz), 4.81 and 4.92 (AB, 2H, J AB ) 11.5 Hz), 4.84 and
3
4
.92 (AB, 2H, J AB ) 11.5 Hz), 7.01-7.64 (m, 10H); MS m/z
+
31
1
(
FAB) 962 [(M + H) , 10), 594 (1), 339 (10), 91 (100).
80%): P NMR (CDCl
3
) δ 0.54, 1.48, 2.49; H NMR (CDCl ) δ
3
DL-3-O-Meth yl-m yo-in ositol 1,4,5-tr isp h osp h a te (3). To
2.40-2.81 (m, 12H), 3.52 (dd, 1H, J ) 2.4 Hz, 9.9 Hz), 3.96-
4.55 (m, 19H), 4.79 and 4.91 (AB, 2H, J AB ) 11.4 Hz), 4.82
and 4.92 (AB, 2H, J AB ) 11.6 Hz), 5.23-5.41 (m, 2H), 5.85-
liquid ammonia (40 mL) was added a solution of 23 (0.06 g,
.064 mmol) in dry dioxane (1.8 mL) followed by sodium (0.1
g, 4.3 mmol) in small pieces. The solution was stirred for 5
min at room temperature and was quenched with ethanol. The
ammonia was evaporated in a stream of nitrogen. Ion-
exchange chromatography of the residue dissolved in water
0
50 15 6 3
5.97 (m, 1H), 7.29-7.44 (m, 10H); MS m/z (FAB) C41H O N P
+
959 [(M + H) , 7], 181 (8), 144 (5), 91 (100); HRMS (FAB) calcd
for C41 959.255, found 959.255.
50 15 6 3
H O N P
DL-3-O-(Ca r b oxym et h yl)-2,6-d i-O-b en zyl-m yo-in osit ol
1,4,5-Tr is[bis(2-cya n oeth yl) p h osp h a te] (28). To a mixture
of 2 mL of carbon tetrachloride, 2 mL of acetonitrile, 3 mL of
water and 27 (0.19 g, 0.20 mmol), and sodium periodate (0.175
g, 0.82 mmol) was added 1 mg of ruthenium trichloride
hydrate, and the entire mixture was stirred vigorously for 2 h
on Q Sepharose Fast Flow, using a gradient of H
2
O to 1 M
TEAB (pH 8.0), gave the compound 3 (0.047 g, 0.045 mmol,
3
1
7
0
0%); 3 was eluted at ca. 700 mM TEAB: P NMR (D
2
O) δ
1
.67, 3.16, 3.57; H NMR (D
2
O) δ 3.32 (dd, 1H, J ) 2.8 Hz, 9.9
Hz), 3.44 (s, 3H), 3.82 (dd, 1H, J ) 9.4 Hz), 3.91 (ddd, 1H, J
2.6 Hz, 7.7 Hz, 9.4 Hz), 3.97 (dd, 1H, J ) 9.0 Hz), 4.27 (dd,
H, J ) 9.5 Hz), 4.45 (dd, 1H, J ) 2.4 Hz); MS m/z (FAB) 433
)
1
[
(
at room temperature. Then, 10 mL of CH
the phases were separated. The upper aqueous phase was
extracted three times with CH Cl The combined organic
extracts were dried (MgSO ) and concentrated. The residue
2 2
Cl was added, and
-
(M - H) , 100], 415 (5), 315 (8), 177 (5), 159 (6), 97 (9); HRMS
FAB) calcd for C 433.978, found 433.978.
DL-3-O-Eth yl-m yo-in ositol 1,4,5-Tr isp h osp h a te (4). The
above compound was prepared in a fashion similar to that
2
2
.
7
H
17
O
15
P
3
4
was diluted with 20 mL of ether, filtered through Celite, and
concentrated. Flash column chromatography of the residue
3
1
1
31
described for 3: P NMR (D
2
2
O) δ 0.47, 3.23, 3.77; H NMR
gave the compound 28 as an oil (0.13 g, 0.13 mmol, 64%):
P
1
(
D
O) δ 1.03 (t, 3H, J ) 7.3 Hz), 3.38 (dd, 1H, J ) 2.8 Hz, 9.8
NMR (CDCl
3
) δ -3.43, -3.16, -2.96; H NMR (CDCl ) δ 2.71-
3
Hz), 3.62-3.73 (m, 2H), 3.81-4.00 (m, 3H), 4.23 (m, 1H), 4.43
2.84 (m, 12H), 3.66 (dd, 1H, J ) 2.8 Hz, 9.9 Hz), 4.02-4.50
(m, 18H), 4.52 (br s, 1H), 4.53-4.98 (m, 4H), 7.27-7.45 (m,
-
(
(
dd, 1H, J ) 2.4 Hz); MS m/z (FAB) 447 [(M - H) , 50] 287
+
22), 217 (34), 134 (100), 97 (18), 81 (24); HRMS (FAB) calcd
10H); MS m/z (FAB) 977 [(M + H) , 7], 338 (15), 149 (28), 91
for C
8
H
19
O
15
P
3
446.985, found 446.984.
(100); HRMS (FAB) calcd for C40
977.229.
47 17 6 3
H O N P 977.229, found
DL-3-O-P r opyl-myo-in ositol 1,4,5-Tr isph osph ate (5). The
above compound was prepared in a fashion similar to that
DL-3-O-(Ca r boxym eth yl)-m yo-in ositol 1,4,5-Tr isp h os-
p h a te (6). To liquid ammonia (40 mL) was added a solution
of 28 (0.06 g, 0.06 mmol) in dry dioxane (1.8 mL) followed by
sodium (0.1 g, 4.3 mmol) in small pieces. The solution was
stirred for 5 min at room temperature and was quenched with
ethanol. The ammonia was evaporated in a stream of nitro-
gen. Ion-exchange chromatography of the residue dissolved
in water, on Q Sepharose Fast Flow, using a gradient from
3
1
1
described for 3: P NMR (D O) δ -0.067, 2.89, 3.90; H NMR
2
(
3
1
D
2
O) δ 1.11 (t, 3H, J ) 7.3 Hz), 1.43 (td, 2H, J ) 7.3 Hz),
.31 (dd, 1H, J ) 2.8 Hz, 9.7 Hz), 3.43-3.91 (m, 2H), 3.72 (dd,
H, J ) 9.8 Hz), 3.77-3.91 (m, 2H), 4.19 (ddd, 1H, J ) 9.8
-
Hz, 9.3 Hz), 4.27 (s, 1H); MS m/z (FAB) 461 [(M - H) , 100),
1
4
77 (8), 159 (9), 97 (13); HRMS (FAB) calcd for C
61.001, found 461.001.
9 21 15 3
H O P
DL-3-O-Allyl-2,6-d i-O-ben zyl-m yo-in ositol (26). DL-3-O-
2
H O to 1 M TEAB (pH 8.0), gave the compound 6 (0.045 g,
3
1
Allyl-2,6-di-O-benzyl-1-O-(p-methoxybenzyl)-4,5-O-isopropy-
lidene-myo-inositol (14) (1.98 g, 3.53 mmol) was heated under
reflux in 1 M hydrochloric acid/methanol (1:2, 40 mL) for 5 h
whereupon the solvents were evaporated to dryness to yield a
residue that was purified by flash column chromatography
0.041 mmol, 68%). 6 was eluted at ca. 800 mM TEAB:
P
1
NMR (D
2
O) δ 0.20, 0.30, 1.01; H NMR (D O) δ 3.46 (dd, 1H,
2
J ) 9.6 Hz), 3.85-4.15 (m, 5H), 4.33 (dd, 1H, J ) 9.2 Hz),
4.42 (br, 1H); MS m/z (FAB) 479 [(M + H) , 100), 177 (18),
+
159 (32), 97 (25); HRMS (FAB) calcd for C
found 478.976.
8 18 17 3
H O P 478.976,
(
2
3
(
2
2
9
petroleum ether/ethyl acetate 50-100%) to give the compound
6 in a crystallized form (petroleum ether 60-80 °C) (1.27 g,
1
.18 mmol, 90%): mp 105-106 °C; H NMR (CDCl
3
) δ 2.37
Ack n ow led gm en t. We thank the Wellcome Trust
for financial support (Program Grant No. 045491) and
Professor S. R. Nahorski for preliminary biological
evaluations.
br s, 1H), 2.39 (br s, 1H), 2.75 (br s, 1H), 3.19 (dd, 1H, J )
.4 Hz, 9.7 Hz), 3.47 (dd, 1H, J ) 9.2 Hz), 3.53 (dd, 1H, J )
.6 Hz, 9.4 Hz), 3.67 (dd, 1H, J ) 9.4 Hz), 3.97 (dd, 1H, J )
.6 Hz), 4.06 (dd, 1H, J ) 2.6 Hz), 4.00-4.17 (m, 2H), 4.70
and 4.93 (AB, 2H, J AB ) 11.5 Hz), 4.85 and 4.88 (AB, 2H, J AB
J O970926Y