Diastereoselective synthesis of γ-amino-δ-hydroxy-α, α-difluorophosphonates: A vehicle for structureactivity relationship studies on SMA-7, a potent sphingomyelinase inhibitor

Article abstract of DOI:10.1021/jo9008782

(Chemical Equation Presented) A highly diastereoselective synthesis of 2-amino alcohol derivatives bearing a difluoromethylphosphonothioate group at the 3-position was achieved through LiAlH(Ot-Bu)₃-mediated reduction of the corresponding R-ami

Full text of DOI:10.1021/jo9008782

pubs.acs.org/joc
between SMases and specific signaling systems have not been
Diastereoselective Synthesis of γ-Amino-δ-hydroxy-
r,r-difluorophosphonates: A Vehicle for Structure-
activity Relationship Studies on SMA-7, a Potent
Sphingomyelinase Inhibitor
fully elucidated yet. Potent SMase inhibitors are believed to
be useful probes to establish a clear picture of metabolic
links.3 In addition, the SMase inhibitor is expected to have
some clinical value for the treatment of ceramide-mediated
pathogenic states such as inflammation4 and AIDS.5
During our studies directed toward the discovery of novel
inhibitors for SMases, we carried out chemical modifications
of sphingomyeline (SM) by replacement of the phosphoco-
line moiety with a metabolically stable difluoromethylene-
phosphonate (DFMP) group. In these modifications, we
found that SMA-7, a short chain SM-analogue having a
phenyl group at the terminal position, inhibits noncompeti-
tively N-SMase in bovine brain microsomes with IC50 values
of 3.3 μM (Figure 1).6,7 Our biological studies revealed that
SMA-7 had the ability to suppress tumor necrosis factor
(TNF) R-induced apoptosis of PC-12 neurons at a low
concentration of 0.1 μM.8
Takehiro Yamagishi,† Shin Muronoi,† Sadao Hikishima,†
Hiroshi Shimeno,‡ Shinji Soeda,‡ and
Tsutomu Yokomatsu*,†
†School of Pharmacy, Tokyo University of Pharmacy and Life
Sciences, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392,
Japan, and ‡Faculty of Pharmaceutical Sciences, Fukuoka
University, 8-9-11 Nanakuma, Jonan-ku, Fukuoka 814-0180,
Japan
yokomatu@toyaku.ac.jp
Received May 5, 2009
FIGURE 1. Structues of SM and SMA-7.
Brief structure activity relationship (SAR) studies have
suggested that the stereochemistry of SMA-7 may be a
critical factor in biological activity and we found (S,S)-
stereochemistry is a better inhibition motif than (R,R)-
stereochemistry corresponding to natural SM regarding 2-
amino alcohol moiety.6b However, there are many ambig-
uous issues remaining in SAR studies of SMA-7. In parti-
cular, SAR studies focusing on a phenyl group have
remained to be solved, since the previous synthesis of
SMA-7 and its enantiomer relies on commencing from
commercially available either (1S,2S)-2-amino-1-phenyl-
1,3-propanediol or its enantiomer.6b To examine detailed
A highly diastereoselective synthesis of 2-amino alcohol
derivatives bearing a difluoromethylphosphonothioate
group at the 3-position was achieved through LiAlH(O-
t-Bu)3-mediated reduction of the corresponding R-amino
ketones. The phosphonothioate moiety of the product
was readily converted into the corresponding phospho-
nate by oxidation with m-CPBA, followed by aqueous
workup. The developed methods should be useful for
SAR studies of SMA-7, a potent inhibitor of SMases.
(3) For select examples on the preparation of SMase inhibitors, see:
(a) Kornhuber, J.; Tripal, P.; Reichel, M.; Terfloth, L.; Bleich, S.; Wiltfanf,
J.; Gulbins, E. J. Med. Chem. 2008, 51, 219. (b) Inoue, M.; Yokota, W.;
Katoh, T. Synthesis 2007, 622. (c) Wascholowski, V.; Giannis, A. Angew.
Chem., Int. Ed. 2006, 45, 827 and references cited therein.
(4) Amtmann, E.; Zoeller, M. Biochem. Pharmacol. 2005, 69, 1141.
(5) Chatterjee, S. Arterioscler. Thromb. Vasc. Biol. 1998, 18, 1523.
(6) (a) Yokomatsu, T.; Takechi, H.; Akiyama, T.; Shibuya, S.; Kominato,
T.; Soeda, S.; Shimeno, H. Bioorg. Med. Chem. Lett. 2001, 11, 1227.
(b) Yokomatsu, T.; Murano, T.; Akiyama, T.; Koizumi, J.; Shibuya, S.;
Tsuji, Y.; Soeda, S.; Shimeno, H. Bioorg. Med. Chem. Lett. 2003, 13, 229.
(7) Katsumura and co-workers independently reported that the DFMP
analogue of sphingomyelin inhibits SMase competitively, see: Hakogi, T.;
Yamamoto, T.; Fujii, S.; Ikeda, K.; Katsumura, S. Tetrahedron Lett. 2006,
47, 2627.
(8) (a) Soeda, S.; Sakata, A.; Ochiai, T.; Yasuda, K.; Shimeno, H.; Toda,
A.; Eyanagi, R.; Hikishima, S.; Yokomatsu, T.; Shibuya, S. Curr. Drug
Therapy 2008, 3, 218. (b) Soeda, S.; Tsuji, Y.; Ochiai, T.; Mishima, K.;
Iwasaki, K.; Fujiwara, M.; Yokomatsu, T.; Murano, T.; Shibuya, S.;
Shimeno, H. Neurochem. Int. 2004, 45, 619. (c) Sakata, A.; Ochiai, T.;
Shimeno, H.; Hikishima, S.; Yokomatsu, T.; Shibuya, S.; Toda, A.; Eyanagi,
R.; Soeda, S. Immunology 2007, 122, 54. (d) Sakata, A.; Yasuda, K.; Ochiai,
T.; Shimeno, H.; Hikishima, S.; Yokomatsu, T.; Shibuya, S.; Soeda, S.
Cellular Immunol. 2007, 245, 24.
Sphingolipids are known as secondary messengers in
mammalian cells and cell members.1 It is now well accepted
that sphingolipids play key roles in cellular signal transmis-
sion pathways. Ceramide, the primary sphingomyelin meta-
bolite, is generated through the action of a lysosomal acid
sphingomyelinase (A-SMase)2 or a membrane-bound neu-
tral sphingomyelinase (N-SMase), believed to be an essential
signal transduction factor in cell differentiation and in
programmed cell death derivation.1 However, direct links
(1) For reviews, see: (a) Wymann, M. P.; Schneiter, R. Nat. Rev. Mol. Cell
Biol. 2008, 9, 162. (b) Ledeen, R. W.; Wu, G. J. Lipid Res. 2008, 49, 1176.
(c) Hannun, Y. A.; Luberto, C.; Argraves, K. M. Biochemistry 2001, 40, 4893.
(d) Kolter, T.; Sandhoff, K. Angew. Chem., Int. Ed. 1999, 38, 1532.
(2) For a review of SMase, see: Montes, L. R.; Goni, F. M.; Alonso, A.
Sphingolipids Cell Funct. 2006, 53.
6350 J. Org. Chem. 2009, 74, 6350–6353
Published on Web 07/17/2009
DOI: 10.1021/jo9008782
r
2009 American Chemical Society

Yamagishi et al.
JOCNote
SCHEME 1. Strategies for Stereoselective Synthesis of SMA-7
Derivatives with High Diversity
SCHEME 2. Preparation of R-Amino Aldehyde Derivatives 1
and 2
aReagents and conditions: (a) LDA, HCF2PO3Et2, THF,-78 °C,
93%; (b) n-BuLi, ClC(S)OPh, THF, -78 to 0 °C; (c) n-Bu3SnH,
AIBN, toluene, reflux, 80% for two steps; (d) Lawesson’s rea-
gent, toluene, reflux, 60%; (e) Amberlyst 15, MeOH, rt, 72%
for 11, 70% for 12; and (f) DMSO, (COCl)2, Et3N, -78 °C.
SAR studies on the phenyl moiety of SMA-7, a new metho-
dology for incorporating a variety of functional groups into
the terminal position with high diversity is required. In this
context, we planned to examine two synthetic routes directed
toward divergent synthesis of SMA-7 derivatives from read-
ily available R-amino aldehydes 1 and 2 bearing a DFMP or
a difluoromethylphosphonothioate (DFMPT) group as
shown in Scheme 1. In this paper, we wish to describe these
experimental results.
SCHEME 3. Comparison of Reactivity between 1 and 2 in
Phenylation with PhMgBr
Requisite R-amino aldehyde derivatives 1 and 2 were
prepared through the method of Otaka9a and Berkowitz9b
as shown in Scheme 2. Accordingly, (S)-Garner aldehyde 7
was treated with the lithium anion of diethyl difluoromethyl-
phosphonate to give adduct 8, which was subjected to Barton
deoxygenation10 furnishing 9 in a good overall yield. Com-
pound 9 was transformed to the phosphonothioate analogue
10 through thioation, using Lawesson’s reagent under con-
ventional conditions.11 The removal of isopropylydene
groups of 9 and 10, followed by Swern oxidation of the
resulting alcohols 1112 and 12, provided 1 and 2, respectively.
Both compounds were utilized for the next reactions without
silica gel column chromatography to avoid racemization.
Having requisite aldehydes 1 and 2in hand, first, the reactivity
of 1 toward phenyl nucleophiles was examined (Scheme 3).
When 1 was treated with 2 equiv of phenyl magnesium bromide
in THF at 0 °C for 2 h, the alkylation reaction proceeded slowly
to give 3a in a low yield (11%) accompanied by a large amount of
unidentified byproduct. The yield slightly increased to 26% upon
increasing the amounts of phenyl magnesium bromide to 4
equiv. However, both reactions were found to be almost non-
stereoselective. Similar results were obtained in a phenylation
reaction with a phenyl lithium reagent in place of phenyl
magnesium bromide. These poor results might be attributed to
the significant decomposition of the DFMP moiety of 1 arising
from its high coordination ability to the metallic reagents.13
Next, we turn our attention to the phenylation of phospho-
nothioate analogue 2, since our previous research revealed
SCHEME 4. Preparation of R-Amino Ketones 6a-f from 2
that the stability of the DFMPT group was much better than
that of the DFMP group under reduction conditions in the
presence of metal hydride reagents.13 Thus 2 was treated with
phenyl magnesium bromide (6 equiv) in THF at 0 °C for 2 h
(Scheme 3). While the diastereoselectivity of this phenylation
reaction was quite low (1:1.3), the reaction gave 4a in modest
yield (ca. 75%)14,15 accompanied by unidentified byproduct,
amounts of which were apparently suppressed in comparison
with the case of phenylation of phosphonate 1 as expected.
These results prompted us to examine the alkylation-oxida-
tion-reduction sequence starting from aldehyde 2 through
R-amino ketone 6 as an alternative method (Scheme 1).16,17
(14) Yields refers to those from crude aldehyde 2 and not optimized.
(15) An accurate yield was not caluculated because of some inseparable
impurities included.
(16) For the preparation of amino ketones 5, we have synthesized the
Weinreb amide derived from alcohol 11 and examined the reaction with a
large excess of vinyl magnesium bromide (THF/0 °C to rt). However, the
Weinreb amide was found to be inert to the Grignad reagent under our
experimental conditions. In a series of the phosphonothiate analogues, we
failed to prepare the correponding carboxylate due to significant decom-
position of 12 during oxidation. Therefore, we could not examine the
reactivity of the Weinreb amide, derived from alcohol 12, to the Grignard
reagent.
(17) To obtain the required amino alcohol derivertives in a stereoselective
manner, an alternative method may be available on the basis of sequential
DIBAL-mediated reduction and alkylation with a Grignard reaction toward
amino esters in one pot. (a) Polt, R.; Peterson, M. A.; DeYong, L. J. Org.
Chem. 1992, 57, 5469. (b) Polt, R.; Peterson, M. A. Tetraherdon Lett. 1990, 31,
4985. However, in the present studies, we focused on the alkylation-oxidation-
reduction sequence starting from aldehydes 1 and 2.
(9) (a) Otaka, A.; Miyoshi, K.; Burke, T. R., Jr.; Roller, P. P.; Kurobe,
H.; Tamamura, H.; Fujii, N. Tetrahedron Lett. 1995, 36, 927. (b) Berkowitz,
D. B.; Eggen, M.; Shen, Q.; Shoemarker, R. K. J. Org. Chem. 1996, 61, 4666.
(10) Martin,S.M.;Dean,D.W.;Wagman,A.S.Tetrahedron Lett. 1992,33, 1839.
(11) (a) Ozturk, T.; Ertas, E.; Mert, O. Chem. Rev. 2007, 107, 5210.
(b) Jesberger, M.; Davis, T. P.; Barner, L. Synthesis 2003, 1929. (c) Cava,
M. P.; Levinson, M. I. Tetrahedron 1985, 41, 5061.
(12) Yokomatsu, T.; Sato, M.; Shibuya, S. Tetrahedron: Asymmetry
1996, 7, 2743.
(13) Murano, T.; Yuasa, Y.; Kobayakawa, H.; Yokomatsu, T.; Shibuya,
S. Tetrahedron 2003, 59, 10223.
J. Org. Chem. Vol. 74, No. 16, 2009 6351

Yamagishi et al.
JOCNote
TABLE 1. Stereoselective Reduction of 6a-f with Metal Hydride Reagentsa
entry
substrate
R
metal hydride (equiv)
time, min
solvent
anti:syn
yield of 4a-f,d %
1
2
3
4
5
6
7
8
9
10
6a
6a
6a
6a
6a
6b
6c
6d
6e
6f
C6H5
C6H5
C6H5
C6H5
C6H5
4-ClC6H4
4-MeOC6H4
CH3
NaBH4 (2)
DIBAL-H (2)
LiAlH(O-t-Bu)3 (8)
LiAlH(O-t-Bu)3 (8)
L-Selectride (4)
LiAlH(O-t-Bu)3 (8)
LiAlH(O-t-Bu)3 (8)
LiAlH(O-t-Bu)3 (8)
LiAlH(O-t-Bu)3 (8)
LiAlH(O-t-Bu)3 (8)
30
150
30
180
60
20
60
40
40
EtOH
toluene
EtOH
THF
52:48b
89:11b
98:2b
53
44
99
53
71
99
71
87
85
97
42:58b
5:95b
THF
EtOH
EtOH
EtOH
EtOH
EtOH
>99:1c
>99:1c
>99:1c
>99:1c
>99:1c
c-C6H11
CH2dCH
20
aAll reactions were carried out at -78 °C under a nitrogen atmosphere. bThe ratios were determined by HPLC analysis
c
(Inertsil PREP-SIL (GL Sciences Inc.), hexane-EtOAc). The ratio was determined by 31P NMR (CDCl3, 121 MHz) analysis.
dCombined yields for anti- and syn-isomers.
SCHEME 5. Determination of Stereochemistry of anti-4a and
syn-4a
racemization proceeded in these sequences. The relative
configurations of anti-4a and syn-4a were verified after
transformation to the corresponding oxazolidinones 13
and 14 on the basis of the vic-coupling constants and the
selected NOESY correlation as shown in Scheme 5.
High anti-selectivity observed in the reaction with LiAlH-
(O-t-Bu)3 in EtOH may be explained by considering the
formation of the intermediate 15, wherein the carbamate
nitrogen is binding to aluminum and the carbonyl oxygen is
chelating to the aluminum (Figure 2).18 A similar chelate
model and the effectiveness of EtOH on the formation of the
chelate intermadiate have been reported by Hoffman.18
Thus, 2 was respectively treated with aryl magnesium bromides
(C6H5MgBr, 4-ClC6H5MgBr, 4-MeOC6H4MgBr) and alkyl
magnesium bromides (MeMgBr, c-C6H11MgBr, CH2dCH-
MgBr) in THF at 0 °C for 1 h, followed by Swern oxidation of
the resulting adducts without purification, to give R-amino
ketones 6a-f in 43-16% yields14,15 for two steps (Scheme 4).
Aiming at overcoming the diastereoselectivity problem,
next, reductions of R-amino ketones 6a-f were examined
with use of representative metal hydride reagents18 (Table 1).
Although reduction of 6a with NaBH4 in EtOH was almost
nondiastereoselective (entry 1), DIBAL-H-mediated reduc-
tion was found to proceed with somewhat moderate anti-
selectivity (89:11) (entry 2). The anti-selectivity was signifi-
cantly improved to 98:2 to provide anti-4a in a 99% yield,
when LiAlH(O-t-Bu)3 was employed in EtOH18 (entry 3).
Using EtOH as a solvent was critical for high selectivity,
hence a similar reaction in THF resulted in poor diastereos-
electivity (42:58) (entry 4). It should be noted that the
reduction with L-Selectride in THF proceeded to give syn-
4a in a ratio of 5:95 (entry 5). Reduction of 6b-f with
LiAlH(O-t-Bu)3 showed excellent diastereoselectivity inde-
pendently of substituent R (entries 6-10).
FIGURE 2. A possible transition state for the reduction of 6a.
Having established the diastereoselective approach to
anti-4a-f, preparation of a SMA-7 precursor from anti-4a
was next explored (Scheme 6). For this purpose, the DFMPT
moiety of anti-4a must be converted into DFMP. A direct
conversion of anti-4a into the corresponding phosphonate
anti-3a was attempted with m-CPBA as an oxidant.19 How-
ever, any desired product was not obtained because of the
significant decomposition of the substrate. Therefore, anti-
4a was protected with TBS on the hydroxy group and
subsequently was treated with m-CPBA to provide DFMP
derivative 17 in a 92% yield after aqueous workup. The TBS
protecting group of 17 were removed by TBAF to afford
phosphonate anti-3a, which was subjected to deprotection of
the Boc group with TFA and subsequent palmitoylation of
the resulting amine to give 18. The product 18 has already
been elucidated to be a useful precursor of SMA-7.6b
The product anti-4a was confirmed to be almost optically
pure by 31P NMR (CDCl3, 162 MHz) analysis of the
corresponding (R)- and (S)-MTPA esters, indicating no
(18) (a) Hoffman, R. V.; Maslouh, N.; Cervantes-Lee, F. J. Org. Chem.
2002, 67, 1045. (b) Davis, F. A.; Gaspari, P. M.; Nolt, B. M.; Xu, P. J. Org.
Chem. 2008, 73, 9619.
(19) Lopin, C.; Gautier, A.; Gouhier, G.; Piettre, S. R. J. Am. Chem. Soc.
2002, 124, 14668.
6352 J. Org. Chem. Vol. 74, No. 16, 2009

Yamagishi et al.
JOCNote
SCHEME 6. Conversion of anti-4a into a Precursor of SMA-7
LiAlH(O-t-Bu)3 (208.7 mg, 0.80 mmol, 97% purity) in EtOH
(1.2 mL), under a nitrogen atmosphere, was added a solution of
6a (44.6 mg, 0.1 mmol) in EtOH (1.2 mL) slowly at -78 °C. After
being stirred for 20 min at the same temperature, the mixture
was diluted with aqueous 10% citric acid and extracted with
EtOAc. The combined extracts were washed with brine and
dried over MgSO4. The removal of the solvent gave a residue,
which was chromatographed on silica gel (hexane:EtOAc 4:1) to
give anti-4a (44.5 mg, 99%). Colorless crystals; mp 90-91 °C;
D
1
[R]25 þ 0.49 (c 0.60, CHCl3); H NMR (400 MHz, CDCl3) δ
7.39-7.25 (5H, m), 5.02 (1H, br s), 5.00 (1H, br d, J =5.9 Hz),
4.25-4.06 (5H, m), 2.33-2.16 (2H, m), 1.45 (9H, s), 1.25 (6H, dt,
J=4.6, 7.1 Hz); 13C NMR (100 MHz, CDCl3) δ 156.2, 140.4-
126.1 (aromatic carbons), 121.2 (dt, JCP = 177.8 Hz, JCF
264.2 Hz), 80.1, 76.2, 64.52 (d, JCP =6.6 Hz), 64.37 (d, JCP
6.8 Hz), 51.8, 31.2-31.0 (m), 28.3 (3 carbons), 16.05 (d, JCP
=
=
=
aReagents and conditions:(a) TBSCl, Et3N, DMAP, DMF, 61%;
(b) m-CPBA, CH2Cl2, then aq NaHCO3, 92%; (c) TBAF, THF,
95%; (d) TFA, CH2Cl2; and (e) Et3N, DMAP, C15H31COCl,
CH2Cl2, 69% (two steps).
2.9 Hz), 15.99 (d, JCP=3.0 Hz); 19F NMR (376 MHz, CDCl3)
δ -45.01 (1F, dddd, JFH = 8.9 Hz, JFH = 25.7 Hz, JFP
111.4 Hz, JFF=284.9 Hz), -48.22 (1F, dddd, JFH=12.9 Hz,
FH = 23.5 Hz, JFP = 111.4 Hz, JFF = 284.9 Hz); 31P NMR
=
J
In conclusion, we have developed a new stereoselective
method for the synthesis of 2-amino alcohols bearing a
DFMPT group through LiAlH(O-t-Bu)3-mediated reduction
of the corresponding R-amino ketones. The DFMPT group of
theproduct couldbe converted intothe corresponding DFMP
group. This method features high diversity in the introduction
of the terminal substituent and high generality in stereo-
controlling the stereogenic center, and should be useful for
SAR studies of SMA-7, a potent inhibitor of SMases. The
elucidation of SAR of SMA-7 is ongoing.
(162 MHz, CDCl3) δ 75.01 (t, JPF=111.4 Hz); IR (KBr) 3356,
1660, 1171, 1022 cm-1; ESIMS m/z 454. Anal. Calcd for
C19H30NO5F2PS: C, 50.32; H, 6.67; N, 3.09. Found: C, 50.16;
H, 6.67; N, 2.99.
Acknowledgment. This work was supported in part by a
Grant-in-Aid for Scientific Research (C) from the Ministry
of Education, Culture, Sports, Science and Technology of
Japan. The Promotion and Mutual Aid Corporation for
Private School of Japan is thanked for supporting this work.
Experimental Section
Supporting Information Available: Experimental details and
spectral data of new compounds. This material is available free
of charge via the Internet at http://pubs.acs.org.
Typical Procedure for the Reduction of r-Amino Ketones: The
Reaction of 6a with LiAlH(O-t-Bu)3. To a stirred suspension of
J. Org. Chem. Vol. 74, No. 16, 2009 6353