A Chiral Phosphoramidite Reagent for the Synthesis of Inositol Phosphates

Article abstract of DOI:10.1021/acs.orglett.6b01374

There is a paucity of chiral phosphoramidite reagents or chiral catalysis methods for the synthesis of biologically relevant inositol phosphates. A new C₂-symmetrical chiral phosphoramidite has been developed and successfully applied to the syn

Full text of DOI:10.1021/acs.orglett.6b01374

Letter
pubs.acs.org/OrgLett
A Chiral Phosphoramidite Reagent for the Synthesis of Inositol
Phosphates
Estelle Durantie,† Samuel Huwiler,‡ Jean-Christophe Leroux, and Bastien Castagner*,§
Department of Chemistry and Applied Biosciences, ETH-Zurich, 8093 Zurich, Switzerland
̈
̈
S
* Supporting Information
ABSTRACT: There is a paucity of chiral phosphoramidite reagents or chiral catalysis methods for the synthesis of biologically
relevant inositol phosphates. A new C2-symmetrical chiral phosphoramidite has been developed and successfully applied to the
synthesis of a set of chiral inositol bisphosphates. The reagent allowed bis-phosphorylation and chiral resolution, resulting in a
concise synthetic route, thus expanding the toolbox available for the preparation of biologically relevant inositol phosphates in
high optical purity.
nositol phosphates and their lipidated phosphatidyl inositol
phosphates using a unique phosphorylation reaction.10 The first
Icongeners are important signaling molecules with crucial employs peptide-based chiral nucleophilic catalysts. Histidine-
and tetrazolylalanine-containing peptides have been used to
catalyze the asymmetric phosphorylation11,12 and phosphityla-
tion,13 respectively, of inositol derivatives. This method has
allowed the synthesis of numerous chiral inositol phosphates,
phosphatidylinositols, and analogues. The second is the chiral
phosphoramidite reagent recently introduced by Jessen and co-
workers that allows chiral resolution after phosphorylation and
was used in the synthesis of highly phosphorylated inositol
pyrophosphate molecules.14−16 The chiral auxiliary is based on
the β-cyanoethyl moiety which is accessible in a few synthetic
steps from mandelic acid and is a base-labile phosphate
protecting group.
Motivated by the paucity of chiral phosphoramidite reagents
available for synthesis, we developed a new cyclic C2-
symmetrical phosphoramidite reagent and utilized it for bis-
phosphorylation and chiral resolution in a single step. This
methodology enabled rapid access to various chiral IP2 in high
optical purity (Figure 1).
roles in cellular processes such as calcium release, gene
regulation, and immune response.1,2 A network of phosphatases
and kinases metabolically relates the various inositol phosphates
by adding or removing phosphate groups on a central myo-
inositol core.1 For instance, the second messenger inositol
1,4,5-trisphosphate is released by the hydrolysis of phosphatidyl
inositol 4,5-bisphosphate from the plasma membrane by
phospholipase C. 1,4,5-IP3 can then be further metabolized to
1,4-IP2 by a 5-phosphatase which regulates calcium release from
intracellular stores.3 The synthesis of well-defined structures
and derivatives greatly benefited the study of the inositol
phosphates, but functions have still not been attributed to all
members of this large family yet.2 This is well illustrated with
inositol bisphosphates (IP2s) that are one on the most
abundant members of this family.4 They are generally
considered as downstream metabolites. However, studies have
shown their potential involvement in biological functions such
as a DNA polymerase activator5 and cytoskeletal actin inducer6
with other functions yet to be revealed. Numerous inositol
phosphates and all phosphatidyl inositols are chiral molecules,
which complicates their preparation in enantiomerically pure
form. Many efforts to address this challenge have been made
offering strategies based on chiral precursors, desymmetriza-
tion, or chiral resolution of myo-inositol.7−9 However, these
routes suffer from multiple protecting group manipulations and
tedious purification of the charged inositol phosphates. Two
paradigms have emerged for the chiral synthesis of inositol
We focused our efforts on the preparation of C2-symmetric
diphenyl seven-membered ring phosphoramidite 2, which is
analogous to the widely used xylylene phosphoramidite for
several reasons (Scheme 1). First, the C2 symmetry was desired
to avoid any complicated isomeric mixture. Second, the benzyl
ether phosphate bond is sensitive to acid and also allows
Received: May 11, 2016
© XXXX American Chemical Society
A
DOI: 10.1021/acs.orglett.6b01374
Org. Lett. XXXX, XXX, XXX−XXX

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Letter
Scheme 2. Preparation of the Racemic Diols Inositol
a
Intermediates
Figure 1. Chiral inositol bisphosphates (IP2).
Scheme 1. Synthesis of C2 Chiral Phosphoramidite
hydrogenolysis as an alternative deprotection strategy. Finally,
we surmised that a cyclic phosphoramidite would ease the
reagent preparation.17 The chiral diol auxiliary 1 was rapidly
obtained in two steps. Homocoupling of bromoacetophenone
provided diphenylbutanedione,18 which was subsequently
reduced using the Corey−Bakshi−Shibata catalyst.19 Then,
following Arbuzov’s procedure,20 diol 1 reacted cleanly with
phosphorus triamide affording the phosphoramidite 2. The
isolation of the phosphoramidite reagent was complicated by
oxidation and a high boiling point, but the cleanliness of the
reaction allowed for a one-pot in situ preparation of the reagent
(vide infra).
In order to evaluate the methodology, we prepared racemic
partially protected inositol analogues using established method-
ology (Scheme 2). Once phosphorylated and resolved, these
intermediates would give rise to a series of biologically relevant
inositol bisphosphates. The widely used intermediate ( )-4
with free 1 and 4 hydroxyl groups was prepared following the
procedure described by Khersonsky and Chang.21 While
treatment of myo-inositol with acetone ketal afforded a mixture
of regioisomers, a subsequent benzoylation enabled the
separation of the desired isomer ( )-3 and basic deprotection
generated the racemic compound ( )-4. Intermediates ( )-6
and ( )-9 were prepared from myo-inositol orthoformate
5.22,23 Regioselective alkylation of the orthoformate with
PMBCl in the presence of NaH afforded the racemic ( )-6
in 75% yield. As for intermediate ( )-9, it was accessed in four
high yielding steps. After benzylation, orthoformate 7 was
selectively reduced with AlMe3 to release the enantiotopic
hydroxyl group on position 3/1 which was subsequently
benzylated to obtain ( )-8.24 Finally, the remaining acetal was
hydrolyzed to liberate hydroxyl groups on position 1/3 and 5
affording intermediate ( )-9 with an overall yield of 64%.
Phosphitylation with phosphoramidite 2 was performed on
these racemic inositol intermediates, to simultaneously
introduce the two phosphate groups and resolve the
diastereomeric mixture in order to obtain optically pure
diphosphate inositol compounds (Table 1). The phosphor-
a
Bn: benzyl. Bz: benzoyl. DMP: 2,2-dimethoxypropane. PMB: p-
methoxybenzyl. PTSA: p-toluenesulfonic acid.
amidite 2, generated in situ in degassed solvent, was used
directly in the presence of tetrazole in the phosphitylation
reaction followed by oxidation with m-CPBA. Tetrazole was
used in excess to compensate for the diethylamine released
during phosphoramidite formation.
The phosphorylated diastereomeric mixtures were obtained
with full conversion and were purified by normal SiO2 flash
column chromatography. Diastereomers with phosphate on
positions 1,4 and 3,6 were separated by crystallization affording
diastereomer 1,4-P*2 10a in an excellent diastereomeric ratio
(>99:1, Table 1, entry 1). The absolute configuration was
assigned by X-ray analysis revealing the phosphate groups in
positions 1 and 4 (SI Figure S1). Recrystallization of the
mother liquor furnished its diastereomer 3,6-P*2 10b also with
excellent optical purity (dr >99:1) (Table 1, entry 2). We were
delighted to see that diastereomers 11a and 11b, with
phosphate on positions 2,4 and 2,6, respectively, could be
separated by normal SiO2 flash column chromatography and
they were both obtained with exquisite optical purity (dr
>99:1) (Table 1, entries 3 and 4). In this case, the absolute
configuration was determined in the next step by optical
rotation measurement of the deprotected 2,6-IP2. Finally, the
1,5- and 3,5-bisphosphate diastereomers were also separated by
crystallization. The 1,5-P*2 12a could be obtained with high
purity (dr >99:1) (Table 1, entry 5), and its absolute
configuration was determined by X-ray analysis displaying the
phosphate groups in positions 1 and 5 (SI Figure S2).
However, diastereomer 3,5-P*2 12b was only accessed with a
moderate diastereomeric ratio of 85:15 (Table 1, entry 6).
Deprotection of the bisphosphate diastereomers to access the
inositol bisphosphates was achieved in a single step for each
intermediate (Scheme 3). This straightforward deprotection
was a considerable advantage, as inositol phosphate compounds
are sensitive and difficult to purify. Hence the phosphorylated
B
DOI: 10.1021/acs.orglett.6b01374
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a
Table 1. Chiral Resolution via Phosphitylation
Scheme 3. Single Step Deprotection
to the final inositol phosphates in a single high yielding step, as
all protecting groups could be removed simultaneously with
acidic conditions or hydrogenation. Although this methodology
is not meant for a process scale-up application, it proved to be
efficient in the preparation of a chiral inositol bisphosphates
library. Their preparation starting from the commercially
available myo-inositol was achieved in a record number of
four steps and a maximum of seven steps, while synthesis using
classic chiral resolution or chiral substrate starting material
suffers from multiple protection/deprotection steps.25
This cyclic C2-symmetric phosphoramidite comes to expand
the toolbox for general chiral inositol phosphates synthesis. The
reactivity, similar to the widely used xylylene phosphoramidite,
should also give the possibility to rapidly access numerous
chiral inositol phosphates as well as their metabolically stable
analogues thiophosphates. The synthesis of optically pure
inositol phosphates and their analogues remains an invaluable
tool to help decipher their biological role.
a
Reaction conditions: 1. 2 (4 equiv), 1H-tetrazole (23 equiv), CHCl3,
b
rt, 20 h; 2. m-CPBA (10 equiv). Recovered yield based on the
diastereomer itself which represents 50% of the racemic starting
c
material. Diastereomeric ratio determined by 31P and 1H NMR
spectroscopy.
acetal intermediates 10a and 10b, as well as the orthoformates
11a and 11b, were fully deprotected by treatment with TMSBr
and were purified by precipitation to isolate pure compounds in
good yields. Benzylated intermediate 12a was deprotected by
hydrogenolysis and like its counterparts was purified by
precipitation to give 1,5-IP2.
Overall, the development of the new cyclic C2-symmetrical
phosphoramidite reagent offers new methodology to rapidly
access various chiral IP2 in high optical purity. Phosphorylation
with the cyclic C2-symmetric phosphoramidite reagent allowed
the introduction of the two phosphate groups on racemic
inositol derivatives and their chiral resolution in a single step.
Although the optically pure IP2 derivatives could only be
obtained in low to moderate yields, this method avoids the
additional protection/deprotection steps necessary for classic
chiral resolution. Moreover, the auxiliary lability allowed access
ASSOCIATED CONTENT
* Supporting Information
■
S
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.or-
glett.6b01374.
Crystallographic information on compounds 10a and
12a (CIF)
Experimental procedures, analytical data of all new
compounds, and X-ray crystallographic data (PDF)
AUTHOR INFORMATION
Corresponding Author
■
*E-mail: bastien.castagner@mcgill.ca.
C
DOI: 10.1021/acs.orglett.6b01374
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(25) Seo, K.; Yu, S.; Chung, S. J. Carbohydr. Chem. 2007, 26, 305−
327.
Present Addresses
†Adolphe Merkle Institute, University of Fribourg, 1700
Fribourg, Switzerland.
‡MSD Merck Sharp and Dohme, 6005 Luzern, Switzerland.
§Department of Pharmacology & Therapeutics, McGill
University, H3G 1Y6 Montreal, Canada.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We gratefully acknowledge funding by the Swiss National
Science Foundation (Grant 205321_141263). We thank the
NMR spectroscopy service from Institut fur Pharmazeutische
̈
Wissenschaften, the crystallography service SmoCC, and the
Mass Spectrometry service from the Laboratorium fur
̈
Organische Chemie at ETH Zurich for their helpful support.
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DOI: 10.1021/acs.orglett.6b01374
Org. Lett. XXXX, XXX, XXX−XXX