Enantiomerically Convergent Synthesis of Phosphatidyl-D-myo-inositol 3,5-Bisphosphate from Both L- And D-1,2-O-Cyclohexylidene-myo-inositol

Article abstract of DOI:10.1246/cl.2003.724

The synthesis of phosphatidyl-D-myo-inositol 3,5-bisphosphate [PtdIns(3,5)P2] has been conveniently accomplished via convergent routes starting from both enantiomers, 1,2-O-cyclohexylidene-myo-inositol. The synthetic strategy involves completely regiosele

Full text of DOI:10.1246/cl.2003.724

724
Chemistry Letters Vol.32, No.8 (2003)
Enantiomerically Convergent Synthesis of Phosphatidyl-D-myo-inositol 3,5-Bisphosphate
from Both L- and D-1,2-O-Cyclohexylidene-myo-inositol
Fushe Han, Minoru Hayashi,y and Yutaka WatanabeÃy
Venture Business Laboratory, Ehime University, Matsuyama 790-8577
yDepartment of Applied Chemistry, Faculty of Engineering, Ehime University, Matsuyama 790-8577
(Received May 19, 2003; CL-030434)
The synthesis of phosphatidyl-D-myo-inositol 3,5-bisphos-
phate [PtdIns(3,5)P2] has been conveniently accomplished via
convergent routes starting from both enantiomers, 1,2-O-cyclo-
hexylidene-myo-inositol. The synthetic strategy involves com-
pletely regioselective phosphorylation of 3,4-diol and 2,3,6-triol
of the suitably protected inositols with the corresponding phos-
phite in the presence of pyridinium tribromide and 2,6-lutidine,
resulting in the formation of 3-O-phosphorylated products, re-
spectively.
O
O
O
O
O
Si
O
O
Si
O
HO
HO
2 steps
85-86%
a
1
O
O
OLev
Si O
Si O
88-92%
OH
OLev
OH
OP(OBn)
OP(OBn)
2
2
O
O
D-1 (L-1)*
D-2 (L-2)*
D-3 (L-3)*
g
b
(for L-3)
87%
(for D-3)
91%
OBn
O P O
O
O
O
A general problem in synthetic phosphatidylinositol phos-
phates (PtdInsPns) and inositol phosphates (InsPns) from myo-
inositol is only half of the starting material can be converted
to the target molecules. To overcome this problem, we are ex-
ploring new methodologies aimed at synthesizing the same chi-
ral PtdInsPns and/or InsPns molecules from both enantiomers
of myo-inositol derivative.
On the other hand, phosphatidylinositol 3,5-bisphosphate
[PtdIns(3,5)P2] was found to occur in mammalian cell lines1
and is widespread among eukaryotes.2 Its biological function
identification is presently of keen interest to biochemists. Al-
though its biological role has not yet been well recognized, sev-
eral investigations have shown that PtdIns(3,5)P2 is biologically
important, for instance, in sorting membrane proteins into the
lumen of the yeast vacuole and maintaining the vacuolar size.3
To date, several reports on the chemical syntheses of
PtdIns(3,5)P2 analogs have appeared employing glucose,4 and
myo-inositol derivatives such as orthoacetate,5 orthoformate6
and camphor ketal.7 The most rapid route described hitherto
is that of Falck,6a involving 8 steps. This route, however, is
not entirely satisfactory where the yield for the preparation of
the ultimate starting material is less than 50% based on the ap-
proach they used,8 in addition to the general problem we men-
tioned above.
We communicate here a convenient and convergent method
to synthesize dipalmitoyl D-PtdIns(3,5)P2 (11) from both D- and
L-1,2-O-cyclohexylidene-3,4-O-(tetraisopropyl disiloxane-1,3-
diyl)-myo-inositol (1), which can be very readily available in
three steps including the enzyme-aided resolution from myo-in-
ositol9 and, may serve as a versatile intermediate to access var-
ious PtdInsPns and InsPns.10 The striking features of the method
described here involved: (1) both D- and L-1 were used to arrive
at the same goal; (2) regioselective phosphorylation of triols 5
and 9 is unprecedented, and remarkably limits the step numbers;
(3) the convergent synthetic method is amenable to other
PtdInsPns and/or InsPns molecules.
O
O
(BnO) PO
2
OR
OR
O
OH
OLev
LevO
HO
OP(OBn)
OP(OBn)
2
2
O
8
O
4
c
c
89%
84%
O
OBn
O P O
OH
OH
OH
OLev
(BnO) PO
2
HO
LevO
OR
OR
O
HO
OH
OP(OBn)
OP(OBn)
2
2
O
O
9
5
h
d
81%
68%
OBn
O P O
O
O
OBn
O P O
OH
OH
(BnO) PO
2
(BnO) PO
2
OR
OR
OR
OR
O
O
OLev
LevO
HO
OH
OP(OBn)
OP(OBn)
2
2
O
O
10
7
e, f
89%
e, f
91%
O
OH
H O PO
2
O P O
3
OR
OR
OH
OH
OC(O)-
OC(O)-
OP(OBn)
6
n
n
-C
-C
H
15 31
HO
H
15 31
OPO H
3
2
2
11
Bn=PhCH
2
Lev=CH C(O)(CH ) CO
3
2 2
R=C(O)-n-C H
15 31
Scheme 1. Conditions and reagents: (a) TBAFÁ3H2O, AcOH,
THF, À15 to À10 ꢀC; (b) (BnO)3P, pyridinium tribromide,
2,6-lutidine, CH2Cl2, À42 to 0 ꢀC; (c) Py(HF)n, ethylene gly-
col, CH2Cl2, 0 ꢀC to r.t.; (d) 6, pyridinium tribromide, 2,6-luti-
dine, pyridine/CH2Cl2 (v/v=1.1/1), À22 ꢀC to r.t.; (e) hydra-
zine monohydrate, pyridine/AcOH (v/v=4/1), 0 ꢀC to r.t.; (f)
5% Pd/C, H2, AcOEt/MeOH (v/v=1/1), r.t.; (g) 6, pyridinium
tribromide, 2,6-lutidine, CH2Cl2, À42 to 0 ꢀC; (h) (BnO)3P,
pyridinium tribromide, 2,6-lutidine, pyridine/CH2Cl2 (v/v
As outlined in Scheme 1, both enantiomers of diol 1 were
smoothly converted to the fully protected 2 according to the re-
1:1), À22 to 0 ꢀC. L-Isomers of 1, 2, and 3 are not illustrated.
Ã
Copyright ꢀ 2003 The Chemical Society of Japan

Chemistry Letters Vol.32, No.8 (2003)
725
ported approach.10h 2 was transformed into diol 3 by desilyla-
tion. To regioselectively introduce the phosphate group at the
3-position in 3,4-diol D-3, the phosphite–pyridinium tribromide
approach11 was employed as reported in the synthesis of
PtdIns(4,5)P2.12 Thus, D-3 was subjected to phosphorylation
with tribenzyl phosphite, resulting in the formation of the de-
sired 4 in a complete selective manner. Neither its regioisomer
nor diphosphorylated product was isolated. The 3,5-diphosphate
derivative 4 was then transformed smoothly to triol 5 by the
cleavage of the cyclohexylidene ketal with pyridinium poly(hy-
drogen fluoride) [Py(HF)n], which was used to decompose the
isopropylidene group without the migration of the adjacent
phosphate function.13 Triol 5 was converted to its trichloroace-
tate in order to confirm no migration of the three substituents
during the reaction through the 1H NMR and 1H-1H COSY
analyses.
We then turned our attention to the regioselective installa-
tion of the phosphatidyl group at the OH-1 in 1,2,4-triol 5. The
regioselective 1-O-phosphorylation of vicinal 1,2-diol deriva-
tives of myo-inositol employing the phosphite–pyridinium tri-
bromide method used above has been well documented by this
laboratory10c,d,h and other group.6a On the other hand, our recent
studies12 showed that phosphorylation of a vicinal 1,6-diol also
exclusively occurred at the 1-position. These results clearly sug-
gest the highest reactivity of OH-1 among three hydroxyls at 1,
2, and 6 positions, therefore, the selective phosphorylation of 5
is expected to proceed at the OH-1. Indeed, phosphorylation of
5 with dipalmitoylglycerol phosphite 6 in the presence of pyri-
dinium tribromide (PTB) proceeded in a 1.1:1 ratio of pyridine
and CH2Cl2 to yield 1-O-phosphorylation product 7 in 68%
yield without the formation of other possible products. It is
noteworthy that the reaction was dramatically affected by the
ratio of the solvents. Thus, in a 1:12 ratio of the mixed
solvent,6a,10c the phosphorylation did not proceed at all. The rea-
sons for such an extraordinary low-reactivity of 5 are now under
investigation. To determine the exact phosphorylation site, 7
was converted into the corresponding chloroacetate, and its
1H NMR and 1H–1H COSY analyses clearly showed that phos-
phorylation occurred at the OH-1 position.
to the synthesis of other PtdInsPns and/or InsPns compounds.
We are grateful to the Center for Cooperative Research and
Development of Ehime University for MS analysis.
References and Notes
1
a) C. C. Whiteford, C. A. Brearley, and E. T. Ulug, Biochem. J.,
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5
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H. W. Lee and Y. Kishi, J. Org. Chem., 50, 4402 (1985).
For the transformation of myo-inositol to optically active 1, see
reference 10c and references therein. This time, D- and L-1 was
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hexylidene-3,4-O-(tetraisopropyldisiloxane-1,3-diyl)-5-O-tri-
ethylsilyl-6-O-(S)-acetylmandeloyl-myo-inositol.10g
7
8
9
10 a) Y. Watanabe, M. Mitani, T. Morita, and S. Ozaki, J. Chem.
Soc., Chem. Commun., 1989, 482. b) Y. Watanabe, H. Hirofuji,
and S. Ozaki, Tetrahedron Lett., 35, 123 (1994). c) Y.
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With the successful technique for converting D-1 to 7, op-
posite enantiomer, L-1 was also transformed into 10. Thus,
the phosphatidyl group at 1-position was installed through the
phosphorylation of diol L-3 with phosphite 6 prior to the instal-
lation of 3-phosphate group, as compared to the synthetic se-
quences from D-3, followed by the cleavage of the cyclohexyli-
dene ketal to give triol 9. Triol
9 was subjected to
phosphorylation with tribenzyl phosphite to give 10 exclusively.
Finally, respective removal of the Lev group from 7 and 10 by
treatment with hydrazine monohydrate in the mixture of pyri-
dine and acetic acid,10c,14 and subsequent debenzylation by hy-
drogenolysis over 5% palladium on carbon afforded dipalmitoyl
PtdIns(3,5)P2 (11)15 as its free acid. The structure of 11 as free
acid form was confirmed by its NMR and MS spectra. Further
purification of the final product was not done because no other
impurity was found in its NMR spectra and TLC analysis.
In conclusion, both regioselective phosphorylation reac-
tions of diol 3, and triol 5 and 9, remarkably reduced laborious
protection–deprotection procedures, therefore, facilitated the
synthetic route to PtdIns(3,5)P2. In addition, the convergent
synthetic methodology from both enantiomers can be applied
4875 (1976).
24
15 Physical and spectra data of 11 (free acid form): ½aŠ
À1:4,
D
[c ¼ 0:27, CHCl3/MeOH 1:1 (v/v)]; dH (400 MHz, CDCl3/
CD3OD/D2O 1:1:0.1) 5.27 (br, 1H, glyceryl sn-2-H), 4.41 (br
s, 1H, InsH-2), 4.30 (br, 0.5H, glyceryl sn-1-H), 4.20 (m,
0.5H, glyceryl sn-1-H), 4.03-4.17 (m, 6H, glyceryl sn-1-H, sn-
3-H, InsH-1, H-3, H-5), 3.96 (br, 2H, InsH-4, H-6), 2.34 (com-
plex, 4H, pal a-CH2), 1.60 (br, 4H, pal b-CH2), 1.28 (br, 48H,
pal CH2), 0.89 (br, 6H, pal CH3); dp (162 MHz,CDCl3/CD3OD/
D2O 1:1:0.1) 5.33 (1P), 4.54 (1P), 4.12 (1P); Negative FABMS
(triethylammonium salt): m=z: 1008 [(M-2H+K)À, 25%], 992
[(M-2H+Na)À,
35%],
970
[(M-H)À,
100%],
648
[C15H31COOCH2CH(OCOC15H31) CH2OPO3HÀ, 50%], 255
[C15H31COOÀ, 80%]. HRMS (FABÀ, triethanolamine) [MÀ
H]À Calcd. for C41H80O19P3À, 969.4506; Found, 969.4523.
Published on the web (Advance View) July 14, 2003; DOI 10.1246/cl.2003.724