Synthetic approaches to heavily lipidated phosphoglyceroinositides

Article abstract of DOI:10.1021/ol0202161

(Matrix presented) Naturally occurring phosphoinositide glycoconjugates are equipped with varied acyl residues that are important for their biological activity and biosynthesis. This paper reports that acylation at O2 of the myo-inositol moiety can be ach

Full text of DOI:10.1021/ol0202161

ORGANIC
LETTERS
2003
Vol. 5, No. 3
255-257
Synthetic Approaches To Heavily
Lipidated Phosphoglyceroinositides
Urs Schlueter, Jun Lu, and Bert Fraser-Reid*
Natural Products and Glycotechnology Research Institute, Inc. (NPG),
4118 Swarthmore Road, Durham, North Carolina 27707
dglucose@aol.com
Received October 24, 2002
ABSTRACT
Naturally occurring phosphoinositide glycoconjugates are equipped with varied acyl residues that are important for their biological activity
and biosynthesis. This paper reports that acylation at O2 of the myo-inositol moiety can be achieved by stereocontrolled ortho ester
rearrangement. Coupling to homo- or heterodiacylated glycerols was achieved via phosphoramidite methods, and exhaustive debenzylation
by transfer hydrogenation afforded the deprotected phosphoglyceroinositides. The latter can be kept in chloroform solution at room temperature
for over two months without migration of the inositol acyl group.
Lipidated phosphatidylinositolglycerols are ubiquitous bio-
regulators that are associated not only with several dreaded
diseases1-3 but also with vital life processes, including
calcium mobilization,4 signal transduction,5 and insulin
stimulation.6 Glycosylphosphatidylinositols (GPIs), sum-
marized as 1a and b, are a relatively new7 class of
glycoprotein/phosphoinositides that are associated with para-
sitic diseases, including African Sleeping Sickness1 and
malaria.3 Landmark studies by Ferguson and co-workers
resulted in the first structure elucidation of a GPI in 1988.8
The chemoenzymatic methodology used was adopted so
quickly that within five years, over 200 GPI-anchored
(1) Ferguson, M. A. J. PNAS 2000, 97, 10673-10675. Nagamune, K.;
Nozaki, T.; Maeda, Y.; Ohishi, K.; Fukuma, T.; Hara, T.; Schwarz, R. T.;
Sutterlin, C.; Brun, R.; Riezman, H.; Kinoshita, T. PNAS 2000, 97, 10336-
10341.
(2) Sherman, I. W., Ed.; Malaria: Parasite Biology, Pathogenesis, and
Protection; ASM Press: Washington, DC, 1998. Bruce-Chawatt’s Essential
Malariology; Gilles, H. M., Warrell, D. A., Eds.; Arnold Publishers:
London, 1997. Oaks, S. C., Jr. Malaria: Obstacles and Opportunities;
Mitchell, V. S., Person, G. W., Carpenter, C. J., Eds.; National Academy
Press: Washington, DC, 1991.
(6) Diabetes in America 2nd ed.; NIH Publication No. 95-1468; National
Institutes of Health: Bethesda, MA, 1995. Stalfors, P. BioEssays 1997, 19,
327-335. Saltiel, A. R.; Kahn, C. R. Nature 2001, 414, 799-806.
(7) Cardosa de Almeida, M. L.; Turner, M. J. Nature 1983, 349-352.
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Constant, P.; Venisse, A.; Riviere, M.; Puzo, G. J. Biol. Chem. 1997, 272,
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(10) (a) Mehlert, A.; Richardson, J. M.; Ferguson, M. A. J. Mol. Biol.
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Geyer, R.; Reinwald, E.; Risse, H. J.; Schwarz, R. T. J. Mol. Biol. 1996,
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10.1021/ol0202161 CCC: $25.00 © 2003 American Chemical Society
Published on Web 01/07/2003

proteins had been identified9 across the evolutionary array
of eukaryotes10 and, recently, a plant11 and a virus.12
Lipoarabinomannans (LAMs), 1c, associated with tuber-
culosis and leprosy,13 carry mannose residues at inositols O2
and O6, the latter being part of an extended mannan that is
eventually linked to an arabinan.13
The above brief summary indicates that the lipid residues
of substructure 1 individually and/or collectively play
dominant roles in the biology of phosphoglycero inositides.
Accordingly, we have developed synthetic approaches that
allow each of the units to be modified as needed.
Inositol Moiety. 3,4,5,6-Tetra-O-benzyl inositol, 2, which
is readily available in racemic21 or optically pure22 form, was
a convenient starting material. The seminal work of King
and Allbutt on stereoelectronically controlled decomposition
of cyclic ortho esters on a cyclohexano scaffold23 provided
a ready procedure for differentiating between the hydroxyls
of 2 with simultaneous installation of the desired O2-acyl
group. Commercially available trimethyl orthovalaerate 5a
afforded the cyclic counterpart 3 quantitatively. Rearrange-
ment with camphorsulfonic acid then gave the desired axial
O2-esters 4a, but only in a modest 20-30% excess over the
equatorial option 4b.
We have recently reported on the efficiency of ytterbium-
(III) triflate for promoting the glycosidation reactions of
n-pentenyl ortho esters.24 This reagent was found to produce
a higher ratio of 4a to 4b with improved yields, in much
shorter reaction times.
Interestingly, long-chain alkyl trimethyl ortho esters such
as the myristoyl derivative 5b, which had to be prepared
from the corresponding nitrile,25 bypassed an isolable cyclic
intermediate, leading directly to the stereoelectronic con-
trolled axial product 7b. Unfortunately, the overall yield was
poor, and therefore we were forced to take the alternative,
A fascinating feature of the phosphoinositide moiety of
several parasite and mammalian GPIs9 is an acyl group at
O2 of the inositol moiety, e.g., 1b, the presence of which
can be deduced by the failure to produce a 1,2-cyclic
phosphate upon treatment with phospholipase C (PLC).14
Inositol O2 acylation gains further significance because it is
essential for the addition of the mannose unit (Man-1) of
1a/b in mammals,15,16 but the process is optional in parasites,
thereby providing a window for specific intervention in one
or another pathway.17 Furthermore, the acyl group may be
cleaved and/or reinstalled at various stages of biosynthesis
so that the final GPI may or may not be acylated.9
Scheme 1a
The glycerolipid moiety of 1 presents its own share of
complexities. Biosynthetically in 1a or 1b, the unit begins
with two long chain acyl groups; however, elaborate “fatty
acid remodeling” processes subsequently take place, leading
to hetero- or homodiacylation.15,16 The sn2 position is of
particular interest because this may eventually carry a free-
OH or a saturated18 or unsaturated19 acyl group. Some of
these events are presumed to enable the growing GPI to
traverse the membrane of the endoplasmic reticulum under
the agency of a flippase enzyme,20 thus far unidentified.
(11) Oxley, D.; Bacic, A. PNAS 1999, 96, 14246-14251.
(12) Jacobs, M. G.; Robinson, P. J.; Bletchley, C.; Mackenzie, J. M.;
Young, P. R. FASEB 2000, 14, 1603-1610.
(13) Llangumaran, S.; Arni, S.; Poincelet, M.; Theler, J.-M.; Brennan,
P. J.; Nasir-ud-Din., Hoessli, D. C. J. Immunol. 1995, 155, 1334-1342.
(14) Campbell, A. S. In Glycoscience: Chemistry and Biology; Fraser-
Reid, B., Tatsuta, K., Thiem, J., Eds.; Springer Heildelberg: 2001; Vol. 2,
pp 1696-1717.
(15) Ferguson. J. Cell. Sci. 1999, 112, 2794-2809.
(16) Maeda, Y.; Watanabe, R.; Harris, C. L.; Hong, Y.; Ohishi, K.;
Kinoshita, K.; Kinoshita, T. EMBO 2001, 20, 250-261. Kinoshita, T.; Inoue,
N. Curr. Opin. Chem. Biol. 2000, 4, 632-638.
a Conditions: (i) 5a, CSA, CH3CN, 18 h (90%); (ii) CH2Cl2,
Yb(OTf)3, 0 °C, 1 h, (69%), 4a:4b )4:1; (iii) Bu2SnO, toluene,
then DMF, -10 °C allyl bromide, CsF, 18 h, (90%); (iv) pyridine,
acyl chloride, (100%); (v) PdCl2, NaOAc, H2O (30-35%).
(17) Nagamune, K.; Noyaki, T.; Maeda, Y.; Ohishi, K.; Fukuma, T.;
Hara, T.; Schwarz, R.; Sutterlin, C.; Brun, R.; Riezman, H.; Kinoshita, T.
PNAS 2000, 97, 10336-10341.
(18) Gerold, P.; Schofield, L.; Blackan, M. J.; Holder, A. A.; Schwarz,
R. T. Mol. Biochem. Parasitol. 1996, 75, 131-143.
(19) Naik, R. S.; Branch, O. H.; Woods, A. S.; Vijaykumar, M.; Perkins,
D. J.; Nahlen, B. L.; Lai, A. A.; Cotter, R. J.; Costello, C. E.; Ockenhouse,
C. F.; Davidson, E. A.; Gowda, D. C. J. Exp. Med. 2000, 192, 1563-
1575.
longer route shown in Scheme 1b involving tin-mediated
alkylation26 of the equatorial OH to give 6a or 6b followed
256
Org. Lett., Vol. 5, No. 3, 2003

by acylation and deprotection to give 7. While our work was
underway, a similar strategy for the route in Scheme 1b was
published by Xue and Guo.27
As shown in Scheme 3, these protocols afforded lipidated
inositols 15a and 15b.
Glycerol Moiety. Commercially available (R)-(-)-2,2-
dimethyl 1,3-dioxolane-4-methanol was p-methoxybenzy-
lated and then deacetonated routinely to obtain diol 8.
Homodiacyl derivatives such as 9a were obtained under
standard conditions, and the deprotected modifications such
as 9b were prepared routinely.
Scheme 3
Heterodiacyl analogues such as 11 are of special interest
in view of Gowda’s findings concerning the dependence of
bioactivity the sn2 acyl substituents.19 Attempts at selective
acylation of 8 at the sn1 position did not give acceptable
yields of 10, because copious diacylation products could not
be suppressed. Standard stannylene methodology26 was not
successful; however, dimethyltin dichloride, recently intro-
duced by Maki et al.,28 solved the problem, affording the
sn1 acylation product, 10, in excellent yield. The second
acylation to give 11a and 11b and then the deprotected
analogues 11c and 11d, respectively, were accomplished
without problems under standard conditions.
Scheme 2
Exhaustive debenzylation to obtain 15c and 15d was
conveniently carried out by transfer hydrogenation using
formic acid in methanol at room temperature. We note further
that samples of 15c and 15d have not given any evidence
for O2fO3 acyl migration upon standing in CDCl3 solution
for over two months.
Acknowledgment. We are grateful to the Human Frontier
Science Program Organization and the Mizutani Foundation
for support.
Supporting Information Available: Experimental details
for all new compounds and 1H NMR spectra for compounds
3, 7, and 15c. This material is available free of charge via
the Internet at http://pubs.acs.org.
OL0202161
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into phosphotriesters, e.g., 14, it was better to connect the
inositol first (giving 13) followed by the glycerol, the
preferred catalysts for the two stages being N,N-diisopro-
pylammonium tetrazolide30 and 1-H-tetrazole, respectively.
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(20) Menon, A. K.; Butikofer, P. In Glycosylphosphatidylinositols:
Biosynthesis and Intracellular Transport. GPI Biosynthesis in Mammalian
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R.G. Landes Co.: Austin, TX, 1999; pp 15-28. Menon, A. K.; Baumann,
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