Novel A-ring analogs of the hormone 1α,25-dihydroxyvitamin D 3: Synthesis and preliminary biological evaluation

Article abstract of DOI:10.1016/j.bmc.2005.02.005

Prepared from a commercial prostaglandin building block, novel vitamin D₃ analogs with a contracted five-membered A-ring were designed and synthesized to mimic the A-ring diol structure of the natural hormone 1α,25-dihydroxyvitamin D₃

Full text of DOI:10.1016/j.bmc.2005.02.005

Bioorganic & Medicinal Chemistry 13 (2005) 2959–2966
Novel A-ring analogs of the hormone 1a,25-dihydroxyvitamin
D3: synthesis and preliminary biological evaluation
Gary H. Posner,a,* S. H. Tony Lee,a Hyung Jin Kim,a Sara Peleg,b
Patrick Dolanc and Thomas W. Kenslerc
aDepartment of Chemistry, The Johns Hopkins University, Baltimore, MD 21218, USA
bDepartment of Medical Specialties, The University of Texas, M.D. Anderson Cancer Center, Houston, TX 77030, USA
cDivision of Environmental Health Sciences, Bloomberg School of Public Health, Johns Hopkins University, Baltimore,
MD 21205, USA
Received 16 December 2004; revised 3 February 2005; accepted 4 February 2005
This publication is dedicated to Hector DeLuca on the occasion of his 75th birthday with great respect and admiration for his long career of
extraordinary leadership and many outstanding contributions in the vitamin D field
Abstract—Prepared from a commercial prostaglandin building block, novel vitamin D3 analogs with a contracted five-membered
A-ring were designed and synthesized to mimic the A-ring diol structure of the natural hormone 1a,25-dihydroxyvitamin D3. Pre-
pared from commercial 1,4-cyclohexanedione, a structurally simplified analog was designed and synthesized in which a suitably ori-
ented primary allylic hydroxyl group at the C-2 position might be a surrogate for the biologically important 1a-OH in the natural
hormone.
ꢀ 2005 Elsevier Ltd. All rights reserved.
1. Introduction
The natural hormone 1a,25-dihydroxyvitamin D3 (calci-
triol, 1) is a potent regulator of human cell growth and
differentiation as well as calcium homeostasis.1–3 This
hormone and some of its synthetic analogs are currently
marketed drugs for chemotherapy of secondary hyper-
thyroidism,4 osteoporosis,5 and psoriasis.6 Although
most synthetic analogs feature changes in the side chain
of the molecule,7–14 some A-ring modified analogs have
potent biological activities and potential for chemother-
apy of cancer.15–21
Crystallizing the modified VDR receptor containing cal-
citriol has allowed X-ray determination to show that the
relatively large ligand-binding pocket can accommodate
much structural variation in potential calcitriol analog
ligands.22 On this basis and on some structural similarity
to commercial prostaglandin intermediates,23,24 analog
2a with a contracted five-membered A-ring was designed
Keywords: Vitamin D analogs; SAR; Growth inhibition.
*
Corresponding author. Tel.: +1 410 516 4670; fax: +1 410 516
8420; e-mail: ghp@jhu.edu
0968-0896/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2005.02.005

2960
G. H. Posner et al. / Bioorg. Med. Chem. 13 (2005) 2959–2966
to mimic the A-ring diol structure of the natural hor-
mone. Analog 3 was designed to have a much simplified
and easily synthesized six-membered A-ring in which a
suitably oriented primary allylic hydroxyl group might
be a surrogate for the biologically important 1a-OH in
the natural hormone (1). Both nontraditional A-ring
analogs 2 and 3 are 19-nor, lacking the 19-methylene
group, the absence of which was expected to help mini-
mize the calcemic activity of these new analogs.15,17,25
tionship between the epoxide functionality and the adja-
cent siloxy group.27 Alkylidenation followed by
palladium-promoted reductive opening of the epoxide
and then O-silylation gave bis-silylated trans vicinal diol
( )-8.28 Reduction of enoate ester ( )-8 into the corre-
sponding allylic alcohol ( )-9, followed by chlorination,
phosphide displacement, and oxidation gave A-ring
allylic phosphine oxide ( )-10 that was coupled with
C,D-ring C-8 ketone (+)-11 and then desilylated to pro-
vide the target analogs (+)-2a (1a,3b) and (+)-2b
(1b,3a). In order to establish the absolute stereochemis-
try of the A-ring of these analogs, the commercially
available, enantiomerically pure, prostaglandin building
block (+)-4 (3a) was converted into the analog (+)-2b
(1b,3a) through the same reaction sequence as shown
in Scheme 1.
2. Results
2.1. Chemistry
As shown in Scheme 1, diastereoselective epoxidation of
prostaglandin intermediate cyclopentenone ( )-4, which
was prepared according to the literature procedure,26
gave mainly epoxide ( )-5 with the desired trans rela-
As shown in Scheme 2, starting with commercial 1,4-
cyclohexanedione, double alkylidenation afforded an
Scheme 1.

G. H. Posner et al. / Bioorg. Med. Chem. 13 (2005) 2959–2966
2961
equal, inseparable mixture of the two geometric isomers
1329 that were reduced in one step into the corresponding
symmetrical bis-allylic alcohol 14 from which chromato-
graphic separation gave the pure Z-isomer. The Z-iso-
mer 14b was desired so as to form ultimately the target
2-allylic alcohol (+)-3 with the allylic –OH group able
to occupy the same region of space as the important 1-
OH group of the natural hormone calcitriol (1). Mono-
silylation and then allylic chlorination and phosphide
displacement gave, after oxidation, A-ring allylic phos-
phine oxide 16. Coupling with enantiomerically pure
C,D-ring ketone (+)-11 and desilylation produced the
target analog (+)-3.
a reporter construct containing the human osteocalcin
vitamin D responsive element (ocVDRE) linked to the
thymidine kinase promoter and the growth hormone re-
porter gene. One day after transfection, the cells were
treated with the analogs at concentrations ranging from
1 to 1000 nM, for 24 h, and then the medium was col-
lected and growth hormone was measured by using a
radioimmunoassay as described by the manufacturer
(Nichols Institute, San Clemente, CA). The ED50 values
of new analogs (+)-2a and (+)-3 were 500 and 600 nM,
respectively. To determine the VDR affinity, we
2.2. Biology
In our standard murine keratinocyte antiproliferation
assay,21 new analog (+)-2a showed growth inhibition
at and above 300 nM concentrations, whereas new ana-
log (+)-3 was antiproliferative at 1 lM concentration
(Fig. 1). As anticipated, the analog (+)-2b with unnatu-
ral 1b,3a-dihydroxy stereochemistry did not display any
antiproliferative activity under the given test conditions.
To examine transcriptional activity of the analogs,10 the
monkey kidney cells CV1 were transfected by the
DEAE-dextran method with 1 lg/dish expression plas-
mid of the human vitamin D receptor and 2 lg/dish of
Figure 1.
Scheme 2.

2962
G. H. Posner et al. / Bioorg. Med. Chem. 13 (2005) 2959–2966
1
performed competition assays using recombinant hu-
man VDR from COS-1 cells transfected with the VDR
expression plasmid. Homogenates from transfected cells
were incubated with 3H-1,25(OH)2D3 and graded con-
centration of nonradioactive competitors. Ligand-
bound VDR was separated from the free ligand by
HAP. The results of this assay showed that only new
analog (+)-2a, but not analog (+)-3, had weak binding
to human VDR (data not shown).
meter using NaCl plates. H and 13C spectra were ob-
tained in CDCl3 solution and were referenced to
CHCl3 (7.26 and 77.0 ppm, respectively) using a Varian
XL 400 MHz NMR spectrometer. Mass spectra were
obtained on a micromass QTOF electrospray mass spec-
troscopy with electronic or chemical ionization (EI or
CI) at the Department of Chemistry, The Ohio State
University, Columbus, OH.
The racemic prostaglandin building block ( )-4 and its
ketoepoxide ( )-5 were prepared according to the litera-
ture procedures.26,27 The enantiopure prostaglandin
building block (+)-4 was purchased from TCI America
Co.
3. Conclusion
Pursuing our interest in designing A-ring modified ver-
sions of the natural hormone calcitriol (1),21,30–33 we de-
scribe here two new analogs with small [analog (+)-2a]
and with large [analog (+)-3] structural changes in the
A-ring. Analog (+)-2a and especially analog (+)-3 are
relatively easy to synthesize, and both are measurably
antiproliferative and transcriptionally active even
though they lack some (2a) or much (3) of the function-
ality of the A-ring of the natural hormone calcitriol (1).
These new analogs should help in SAR generalizations
concerning how much change in analog structure can
be made in the A-ring without completely abrogating
biological activity. The very modest growth inhibitory
and transcriptional activities of these two new analogs,
however, make them unpromising for further study as
possible candidates for drug development.
4.2. Ethyl 2-(4-tert-butylsiloxy-6-oxa-bicyclo[3.1.0]-
hexan-2-ylidene)acetate [( )-6]
In a flame-dried, 25 mL round-bottomed, single-necked
flask fitted with a magnetic stirring bar, a rubber sep-
tum, and an argon balloon was placed NaH (0.23 g,
9.6 mmol) in THF (5 mL). To this suspension was added
triethylphosphonoacetate (1.9 mL, 9.6 mmol) at 0 ꢁC.
The resulting mixture was warmed to room temperature
and stirred for 2 h. Then, it was cooled to 0 ꢁC followed
by the addition of a solution of ( )-4-tert-butylsiloxy-6-
oxa-bicyclo[3.1.0]hexan-2-one (5) (0.32 g, 1.4 mmol) in
THF (5 mL) via a cannula. After the mixture was
warmed to room temperature and stirred for 7 h, it
was quenched with satd NaHCO3 (5 mL) and extracted
with Et2O (3 · 5 mL). The combined organic portions
were washed with H2O (1 · 5 mL) and brine
(1 · 5 mL), dried (MgSO4), and concentrated. Flash
chromatography (EtOAc–hexane, 15:85) afforded
0.40 g (93%) of ( )-ethyl 2-(4-tert-butylsiloxy-6-oxa-
bicyclo[3.1.0]hexan-2-ylidene)acetate (6) as a mixture
of E:Z isomers in a ratio of 1:2; oil; IR (neat) 2931,
4. Experimental
4.1. General methods
Reagents and solvents were purchased from Aldrich
Chemical Co. and used without additional purification
unless otherwise noted. Reactions involving air sensitive
materials and/or requiring anhydrous reaction condi-
tions were performed under an argon atmosphere. Re-
agent grade tetrahydrofuran (THF) and diethyl ether
were freshly distilled under argon from a purple solution
of sodium and benzophenone. Reagent grade dichloro-
methane (CH2Cl2) was freshly distilled over CaH2.
Anhydrous ethylene glycol–dimethyl ether (DME),
N,N-dimethylformamide (DMF), and toluene were pur-
chased from Aldrich Chemical Co. and used without
further purification. Column chromatography was per-
formed on silica gel, Merck grade 60 (230–400 mesh).
Analytical and preparative thin-layer chromatography
was performed on precoated silica gel plates (250 and
2000 lm, respectively) purchased from Analtech Inc.
Compounds were visualized with a UV lamp and/or
by developing with iodine, p-anisaldehyde solution, or
potassium permanganate solution unless otherwise
noted. High pressure liquid chromatography (HPLC)
was performed on a Rainin HPLX system equipped
with two 25 mL pump heads and a Rainin Dynamax.
1
1713 cmꢀ1; H NMR (CDCl3) (E isomer): d 0.098 (m,
6H), 0.89 (s, 9H), 1.29 (t, J = 7.2 Hz, 3H), 2.60 (m,
1H), 3.03 (dd, J = 19.1, 1.8 Hz, 1H), 3.68 (m, 1H),
3.76 (d, J = 2.1 Hz, 1H), 4.18 (q, J = 7.2 Hz, 2H), 4.44
(d, J = 5.8 Hz, 1H), 6.14 (m, 1H); (Z isomer) d 0.098
(m, 6H), 0.88 (s, 9H), 1.31 (t, J = 7.2 Hz, 3H), 2.08
(dd, J = 17.4, 1.2 Hz, 1H), 2.48 (m, 1H), 3.7 (d,
J = 2.4 Hz, 1H), 4.21 (q, J = 7.2 Hz, 2H), 4.37 (d,
J = 5.5 Hz, 1H), 4.89 (d, J = 2.4 Hz, 1H), 5.94 (m,
1H); 13C NMR (CDCl3) (mixture of E/Z isomers): d
ꢀ4.85, ꢀ4.83, ꢀ4.77, 14.20, 14.22, 18.07, 18.10, 25.70,
25.72, 37.53, 39.52, 54.25, 59.42, 60.00, 60.14, 62.03,
62.93, 69.87, 71.05, 117.24, 119.00, 157.78, 158.23,
165.62, 165.68; HRMS Calcd for C15H26O4SiNa+
[M+Na]: 321.1493. Found: 321.1491.
4.3. Ethyl 2-(3,4-bis(tert-butylsiloxy)cyclopentylidene)-
acetate [( )-8]
In a flame-dried, 25 mL round-bottomed, single-necked
flask fitted with a magnetic stirring bar, a rubber sep-
tum, and an argon balloon was placed a catalytic
amount of Pd(PPh3)4 (0.075 g, 0.065 mmol) in dichloro-
methane (20 mL). To this solution was slowly added
(CH3)2NHÆBH3 (0.17 g, 2.9 mmol) and the resulting
mixture was stirred at room temperature for 5 min.
Melting points were recorded on a Mel-Temp apparatus
and are uncorrected. Optical rotations were measured
on a JASCO, P-100 model polarimeter. Infrared spectra
were recorded on a Perkin–Elmer 1600 FTIR spectro-

G. H. Posner et al. / Bioorg. Med. Chem. 13 (2005) 2959–2966
2963
Then, a solution of 6 (0.40 g, 1.3 mmol) in dichloro-
methane (5 mL) was added via cannula followed by
the addition of AcOH (0.27 mL, 3.6 mmol). After stir-
ring the mixture at room temperature for 30 min, it
was quenched with H2O (10 mL) and extracted with
dichloromethane (3 · 5 mL). The combined organic lay-
ers were washed with H2O (1 · 5 mL) and brine
(1 · 5 mL), dried (MgSO4), and concentrated. Flash
chromatography (EtOAc–hexane, 15:85) afforded
0.29 g (74%) of ( )-ethyl 2-(4-hydroxy-6-oxa-bicy-
clo[3.1.0]hexan-2-ylidene)acetate (7) as a mixture of
E:Z isomers. This compound was dissolved in dichloro-
methane (5 mL), and to the resulting solution was added
imidazole (0.13 g, 1.92 mmol) and tert-butyldimethylsi-
lyl chloride (0.22 g, 1.44 mmol). After stirring at room
temperature for 15 h, the mixture was quenched with
H2O (5 mL) and extracted with dichloromethane
(3 · 5 mL). The organic phases were washed with H2O
(1 · 5 mL), brine (1 · 5 mL), dried (MgSO4), and con-
centrated. Flash chromatography (EtOAc–hexane,
10:90) afforded 0.36 g (90%) of ( )-ethyl 2-(3,4-bis(tert-
butylsiloxy)cyclopentylidene)acetate (8) as an oil; IR
To this suspension was added methyl sulfide (0.2 mL,
2.7 mmol) at 0 ꢁC, and the resulting mixture was stirred
for 15 min at this temperature and was cooled to
ꢀ20 ꢁC. To this mixture was added a solution of ( )-
2-(3,4-bis-(tert-butylsiloxy)cyclopentylidene)ethanol (9)
(0.24 g, 0.64 mmol) in dichloromethane (5 mL) via a
cannula. After stirring at this temperature for 7 h, it
was concentrated and filtered through a short pad of sil-
ica. The filtrate was concentrated and dissolved in THF
(5 mL), and cooled to ꢀ78 ꢁC. In a separate, flame-
dried, 25 mL round-bottomed, single-necked flask fitted
with a magnetic stirring bar, a rubber septum, and an
argon balloon was placed diphenylphosphine (1.1 mL,
6.4 mmol) in THF (5 mL). To this solution was added
n-BuLi (4.0 mL, 1.2 M in THF) at 0 ꢁC. The resulting
reaction mixture was stirred at this temperature for
20 min, and was added to the flask containing the mix-
ture of allylic chloride in THF at ꢀ78 ꢁC. After stirring
at this temperature for 10 h, it was quenched with H2O
(10 mL) and concentrated. The mixture was dissolved in
dichloromethane (5 mL) and to this was added 30%
H2O2 (5 mL) at room temperature. After stirring the
mixture at this temperature for 16 h, it was quenched
with H2O (5 mL), and extracted with dichloromethane
(3 · 5 mL). The combined organic layers were washed
with H2O (1 · 5 mL) and brine (1 · 5 mL), dried
(MgSO4), and concentrated. Flash chromatography
afforded 0.21 g (60%) of phosphine oxide ( )-10 as an
(neat) 2930, 1718 cmꢀ1 1H NMR (CDCl3): d 0.060
;
(m, 12H), 0.856 (s, 9H), 0.862 (s, 9H), 1.28 (t,
J = 7.3 Hz, 3H), 2.33 (m, 1H), 2.76 (m, 2H), 3.02 (m,
1H), 3.96 (m, 1H), 4.01 (m, 1H), 4.16 (J = 7.2 Hz, 2H),
5.78 (m, 1H); 13C NMR (CDCl3): d ꢀ4.83, ꢀ4.79,
ꢀ4.74, ꢀ4.69, 14.31, 17.94, 25.71, 25.75, 39.42, 41.47,
59.42, 78.02, 113.58, 163.79, 166.57; HRMS Calcd for
C21H42O4Si2Na+ [M+Na]: 437.2513. Found: 437.2521.
1
oil; IR (neat) 3342, 2931 cmꢀ1; H NMR (CDCl3): d
0.005 (m, 12H), 0.833 (s, 9H), 0.834 (s, 9H), 1.81 (m,
1H), 2.07 (m, 1H), 2.26 (m, 1H), 2.52 (m, 1H), 3.03
(dd, J = 14.8, 7.6 Hz), 3.78 (m, 2H), 5.33 (m, 1H), 7.45
(m, 6H), 7.72 (m, 4H); 13C NMR (CDCl3): d ꢀ4.76,
ꢀ4.75, ꢀ4.64, 17.95, 19.98, 25.80, 31.52, 32.22, 37.93
4.4. 2-(3,4-Bis-(tert-butylsiloxy)cyclopentylidene)ethanol
[( )-9]
In a flame-dried, 25 mL round-bottomed, single-necked
flask fitted with a magnetic stirring bar, a rubber sep-
tum, and an argon balloon was placed ( )-ethyl 2-(3,4-
bis(tert-butylsiloxy)cyclopentylidene)acetate (8) (0.34 g,
0.82 mmol) in toluene (5 mL). To this solution was
added DIBAL-H (3 mL, 1.5 M in toluene, 4.5 mmol)
at ꢀ78 ꢁC. After stirring for 3 h at this temperature,
the mixture was quenched with satd KÆNa tartrate
(4 mL) and filtered through a pad of Celite. The filtrate
was extracted with Et2O (3 · 5 mL), and the combined
organic phases were washed with H2O (1 · 5 mL) and
brine (1 · 5 mL), dried (MgSO4), and concentrated.
Flash chromatography (EtOAc–hexane, 40:60) afforded
0.24 g (78%) of ( )-2-(3,4-bis-(tert-butylsiloxy)cyclopent-
(d, Jcp = 383.8 Hz), 77.78, 77.91, 110.65 (d, Jccp =
9.2 Hz), 128.38, 128.49, 130.95, 131.04, 131.70, 142.55,
142.68; HRMS Calcd for C31H49O3PSi2Na+ (M+Na):
579.2850. Found: 579.2835. Analysis by a chiral HPLC
((S,S) Whelk-O 1 column, 2-propanol–hexane = 10:90,
2.5 mL/min) indicated the presence of two enantiomers
(10a: tR (1a,3b) = 70.085 min; 10b: tR (1b,3a) =
76.724 min) in a ratio of 1:1. In order to determine the
absolute stereochemistry at the C-1 and C-3 carbon
positions, the enantiomer 10b (1b,3a) was separately
prepared from the enantiomerically pure (+)-4 (3a);
23
D
10b: ½aꢁ þ 21:0 (c 2.0, CHCl3).
4.6. 1a,3b(OH)2-2,19-nor D3 [(+)-2a] and 1b,3a(OH)2-
2,19-nor D3 [(+)-2b]
ylidene)ethanol (9) as an oil; IR (neat) 3342, 2931 cmꢀ1
;
1H NMR (CDCl3): d 0.057 (m, 12H), 0.869 (s, 9H),
0.873 (s, 9H), 2.15 (m, 2H), 2.60 (m, 2H), 3.96 (m,
1H), 3.97 (m, 1H), 4.10 (d, J = 7.0 Hz, 2H), 5.50 (m,
1H); 13C NMR (CDCl3): d ꢀ4.73, ꢀ4.61, 18.01, 18.03,
25.82, 35.59, 39.63, 60.42, 121.50, 141.39; HRMS Calcd
for C19H40O3Si2Na+ [M+Na]: 395.2408. Found:
395.2408.
In a flame-dried, 15 mL round-bottomed, single-necked
flask fitted with a magnetic stirring bar, a rubber sep-
tum, and an argon balloon was placed the phosphine
oxide ( )-10 (0.11 g, 0.20 mmol) in THF (2 mL). To this
solution was added n-BuLi (0.11 mL, 1.6 M in THF,
0.18 mmol) at ꢀ78 ꢁC and the resulting red colored mix-
ture was stirred at this temperature for 15 min. In a sep-
arate, flame-dried, 15 mL pear-shaped, single-necked
flask fitted with a rubber septum and an argon balloon
was placed the C,D-ring ketone (+)-11 (0.017 g,
0.048 mmol) in THF (1 mL). This solution was cooled
ꢀ78 ꢁC, and transferred to the flask containing the
phosphine oxide anion at ꢀ78 ꢁC via a cannula. After
4.5. Phosphine oxide ( )-10
In a flame-dried, 50 mL round-bottomed, single-necked
flask fitted with a magnetic stirring bar, a rubber sep-
tum, and an argon balloon was placed N-chlorosuccin-
imide (0.36 g, 2.5 mmol) in dichloromethane (10 mL).

2964
G. H. Posner et al. / Bioorg. Med. Chem. 13 (2005) 2959–2966
stirring the reaction mixture at this temperature for 3 h,
it was quenched with pH 7 buffer solution (1 mL) and
extracted with EtOAc (5 · 3 mL). The combined organic
phases were washed with H2O (1 · 5 mL) and brine
(1 · 5 mL), dried (MgSO4), and concentrated. Flash
chromatography (EtOAc–hexane, 10:90) afforded
0.017 g (51%) of the coupled product as a yellow oil.
tography (EtOAc–hexane, 2:98) to give a colorless oil
13 (1.1 g, 50%) as an inseparable mixture of E:Z isomers
in a ratio of 1:1. Spectroscopic data of this 13 are iden-
tical to those previously reported in the literature.29 To a
solution of 13 (1.4 g, 5.5 mmol) in toluene (10 mL) was
added DIBAL (30 mL, 1 M in toluene, 30 mmol) at
ꢀ78 ꢁC. After 30 min, the reaction mixture was diluted
with diethyl ether (30 mL), and was quenched with satd
KÆNa tartrate (20 mL), extracted with EtOAc
(3 · 10 mL), dried (MgSO4), and concentrated. Flash
chromatography (EtOAc–hexanes, 50:50) gave 0.35 g
(37%) of E isomer 14a as a white solid and 0.36 g
(37%) of Z isomer 14b as a gel. Structural assignments
for E isomer and Z isomer were accomplished based
The coupled product (0.017 g, 0.025 mmol) was dis-
solved in THF (2 mL), and to this solution was added
tetrabutylammonium fluoride (0.30 mL, 1.0 M in
THF, 0.30 mmol). After stirring the mixture at room
temperature for 17 h in dark, it was concentrated and
purified by flash chromatography (EtOAc–hexane,
90:10) to afford 0.0070 g (72%) of a mixture of 2a and
2b as a yellow oil. The diastereomers were separated
by chiral HPLC (OD semipreparation column; 2-propa-
nol–hexane = 9:91; flow rate = 2.5 mL/min; P =
0.43 kpsi) to afford 0.0020 g (21%) of 2a (1a,3b,
tR = 22.5 min) and 0.0022 g (22%) of 2b (1b,3a,
tR = 35.1 min) as colorless oils. The purity of these com-
pounds was independently confirmed by a different
chiral HPLC ((S,S) Whelk-O 1 column, 2-propanol–
hexane = 10:90, 2.5 mL/min; flow rate = 2.5 mL/min;
P = 21 bars; 2a (1a,3b): tR = 16.8 min; 2b (1b,3a):
tR = 14.8). The absolute stereochemistry at the C1 and
C3 carbon positions of the A-ring was established by
preparing the diastereomerically pure (+)-2b (1b,3a)
from the commercially available, enantiopure, prosta-
1
on their symmetry consideration and relative H NMR
simplicity. E Isomer: IR (neat) 3285, 2928, 2841, 1668,
1
1436 cmꢀ1; mp = 74–75 ꢁC; H NMR (CDCl3): d 5.46–
5.43 (m, 2H), 4.18 (d, J = 5.6 Hz, 2H), 4.16 (d,
J = 6.8 Hz, 2H), 2.29 (t, J = 6 Hz, 4H), 2.19 (t,
J = 6 Hz, 4H); 13C NMR (CDCl3): d 142.3, 121.7,
58.6, 36.9, 29.4. Z Isomer: IR (neat) 3285, 2928, 2841,
1668, 1436 cmꢀ1
; d 5.44 (t,
1H NMR (CDCl3):
J = 6.8 Hz, 2H), 4.18 (d, J = 6.8 Hz, 4H), 2.25 (s, 4H),
2.22 (s, 4H); 13C NMR (CDCl3): d 142.4, 121.6, 58.6,
37.6, 28.9; HRMS Calcd for C10H16O2Na+ [M+Na]:
191.1042. Found: 191.1056.
4.8. Phosphine oxide 16
glandin building block (+)-4 (3a). Compound
To a solution of bis-allylic alcohol (Z)-14b (0.36 g,
2.1 mmol) in DMF (5 mL) was added tert-butyldimeth-
ylsilyl chloride (0.35 g, 2.3 mmol) and imidazole
(0.16 g, 2.3 mmol) at 0 ꢁC. After stirring for 1 h at
0 ꢁC, the reaction mixture was quenched with H2O
(5 mL), extracted with EtOAc (3 · 10 mL), dried
(MgSO4), and concentrated. Flash chromatography
(EtOAc–hexanes, 20:80) afforded 0.23 g (38%) of
mono-TBS alcohol 15 as an oil.
23
2a: ½aꢁ þ 37:6 (c 0.1, CHCl3); IR (neat) 3361,
D
1
2943 cmꢀ1; H NMR (CDCl3): d 0.54 (s, 3H), 0.93 (d,
J = 6.4 Hz, 3H), 1.22 (s, 6H), 1.01–2.00 (m, 20H), 2.48
(m, 2H), 2.79 (m, 2H), 4.08 (m, 2H), 5.60 (d,
J = 11.6 Hz, 1H), 6.28 (d, J = 11.0 Hz, 1H); 13C NMR
(CDCl3): d 12.02, 18.80, 20.80, 22.22, 23.52, 27.67,
28.89, 29.21, 29.37, 35.50, 36.11, 36.39, 39.39, 40.49,
44.40, 45.71, 56.16, 56.51, 71.13, 77.21, 77.90, 116.99,
119.74, 135.10, 141.38; HRMS Calcd for C25H42O3Na+
[M+Na]: 413.3026. Found: 413.3010; UV (MeOH) kmax
To a solution of N-chlorosuccinimide (0.22 g, 1.7 mmol)
in dichloromethane (5 mL) at 0 ꢁC was slowly added
Me2S (220 lL, 1.7 mmol). The resulting white cloudy
mixture was stirred for 30 min at 0 ꢁC and then treated
with a solution of the mono-TBS alcohol 15 (0.22 g,
0.70 mmol) in dichloromethane (5 mL). After stirring
at this temperature for 7 h, it was concentrated and fil-
tered through a short pad of silica. The filtrate was con-
centrated and dissolved in THF (2 mL), and cooled to
ꢀ78 ꢁC. Then, it was treated with a solution of potas-
sium diphenyl phosphide (9.8 mL, 0.5 M in THF,
4.9 mmol) in THF (5 mL). After stirring for 15 min, it
was added H2O (15 mL), and the resulting colorless mix-
ture was allowed to warm to room temperature. The sol-
vent was evaporated, and the residue was taken up in
dichloromethane (5 mL). To this solution was added
hydrogen peroxide (4 mL), and the resulting mixture
was stirred vigorously for 45 min. The mixture was ex-
tracted with dichloromethane (3 · 5 mL), and the com-
bined organic layers were washed with satd Na2SO3
(1 · 5 mL) and H2O (1 · 5 mL), dried (MgSO4), and
concentrated. Flash chromatography (EtOAc–hexane,
50:50) afforded 0.23 g (70%) of A-ring phosphine oxide
16 as an oil; IR (neat): 2929, 2855, 2360, 1436, 1251,
23
D
254 nm (e 18,550). Compound 2b: ½aꢁ þ 61:4 (c 0.1,
CHCl3); IR (neat) 3361, 2944 cmꢀ1
;
1H NMR
(CDCl3): d 0.55 (s, 3H), 0.93 (d, J = 6.6 Hz, 3H), 1.22
(s, 6H), 1.01–2.00 (m, 19H), 2.34 (m, 2H), 2.81 (m,
3H), 4.11 (m, 2H), 5.59 (d, J = 11.4 Hz, 1H), 6.27 (d,
J = 11.3 Hz, 1H); 13C NMR (CDCl3): d 12.01, 18.80,
20.79, 22.22, 23.52, 27.66, 28.89, 29.21, 29.37, 35.49,
36.11, 36.39, 39.40, 40.49, 44.40, 45.71, 56.13, 56.51,
71.13, 77.23, 77.91, 116.99, 119.75, 135.10, 141.37;
HRMS Calcd for C25H42O3Na+ [M+Na]: 413.3026.
Found: 413.3024; UV (MeOH) kmax 254 nm (e 20,424).
4.7. Bis-alcohols (E)-14a and (Z)-14b
To a solution of the NaH in DME (10 mL) was added
triethylphosphonoacetate (9.10 g, 44.5 mmol) in DME
(10 mL) at 0 ꢁC via a cannula. After stirring for 1 h at
0 ꢁC, a solution of 1,4-cyclohexadione 12 (1.0 g,
8.9 mmol) in DME (10 mL) was added via a cannula.
After 30 min, the dark brown colored mixture was
quenched with H2O (5 mL). The resulting mixture was
extracted with EtOAc (3 · 25 mL), dried (MgSO4), con-
centrated, and purified using silica gel column chroma-

G. H. Posner et al. / Bioorg. Med. Chem. 13 (2005) 2959–2966
2965
1
1178, 1099, 834 cmꢀ1; H NMR (CDCl3): d 7.76–7.69
(m, 4H), 7.52–7.41 (m, 6H), 5.29–5.24 (m, 2H), 4.14
(d, J = 6.4 Hz, 2H), 3.14–3.08 (m, 2H), 2.12–1.83 (m,
8H), 0.88 (s, 9H), 0.04 (s, 6H); 13C NMR (CDCl3): d
143.6, 143.5, 139.7, 133.1, 132.1, 131.7, 131.6, 131.0,
130.9, 138.4, 128.3, 122.3, 110.2, 110.1, 59.4, 37.6,
37.3, 30.4, 29.7, 28.7, 28.2, 25.9, 18.3; HRMS calcd for
C28H39O2PSiNa+ [M+Na]: 489.234912. Found:
489.23301.
References and notes
1. For a review, see Cancelar, L.; Theofan, G.; Norman, A.
W. In Hormones and Their Actions; Cooke, B. A., King, R.
J. B., Vander Moler, H. J., Eds.; New York: Elsevier,
1988; pp 269–289, Part 1, Chapter 15.
2. Posner, G. H.; Kahraman, M. Eur. J. Org. Chem. 2003,
20, 3889.
3. Calcium Regulation and Bone Metabolism: Basic and
Chemical Aspects; Cohn, D. V., Ed.; Amsterdam: Elsevier
Science, 1987; Vitamin D, The Calcium Homeostatic
Steroid Hormone; Norman, A. W., Ed.; New York:
Academic, 1978.
4.9. Vitamin D3 analog [(+)-3]
4. Kabat, M. M.; Radinov, R. Curr. Opin. Drug Discovery
Dev. 2001, 4, 808.
5. DeLuca, H. F. FASEB 1987, 224.
6. Morimoto, S.; Yoshikawa, K.; Kozuka, T.; Kitano, Y.;
Imanaka, S.; Fuko, K.; Koh, E.; Omishi, T.; Kumanhara,
Y. Calcif. Tissue Int. 1986, 39, 209.
A flame-dried, 10 mL round-bottomed flask equipped
with a magnetic stirring bar and a septum along with
an argon balloon was charged with the phosphine oxide
16 (0.051 mg, 0.10 mmol) in THF (2 mL). To this solu-
tion was added n-BuLi (0.069 mL, 1.6 M in THF,
0.10 mmol) at ꢀ78 ꢁC and the resulting red colored mix-
ture was stirred at this temperature for 15 min. In a sep-
arate flame-dried, 10 mL round bottomed flask
equipped with a magnetic stirring bar, a septum, and
an argon balloon was placed the CD-ring ketone (+)-
11 (0.026 g, 0.070 mmol) in THF (1 mL). This solution
was cooled to ꢀ78 ꢁC, and was transferred into the flask
containing the phosphine oxide anion at ꢀ78 ꢁC via a
cannula. The resulting mixture was allowed to stir at this
temperature for 9 h. Then, the reaction mixture was
quenched with satd KÆNa tartate (1 mL) and was ex-
tracted with EtOAc (3 · 25 mL). The combined organic
phases were washed with water (1 · 25 mL) and brine
(1 · 10 mL), dried (MgSO4), and concentrated. Flash
chromatography (EtOAc–hexanes, 2:98) gave 0.024 g
(45%) of the coupled product as a colorless oil.
7. For a review, see Bouillon, R.; Okamura, W. H.; Norman,
A. W. Endocrinol. Rev. 1995, 16, 200.
8. Kahraman, K.; Sinishtaj, S.; Dolan, P. M.; Kensler, T. W.;
Peleg, S.; Saha, U.; Chuang, S. S.; Bernstein, G.; Korczak,
B.; Posner, G. H. J. Med. Chem. 2004, 47, 6854.
9. Suh, B.-C.; Jeon, H. B.; Posner, G. H.; Silverman, S. M.
Tetrahedron Lett. 2004, 45, 4623.
10. Posner, G. H.; Halford, B. A.; Peleg, S.; Dolan, P.;
Kensler, T. W. J. Med. Chem. 2002, 45, 1723.
11. Posner, G. H.; Crawford, K.; Siu-Caldera, M.-L.; Reddy,
G. S.; Sarabia, S. F.; Feldman, D.; vanEtten, E.; Mathieu,
C.; Gennaro, L.; Vouros, P.; Peleg, S.; Dolan, P. M.;
Kensler, T. W. J. Med. Chem. 2000, 43, 3581.
12. Posner, G. H.; Wang, Q.; Han, G.; Lee, J. K.; Crawford,
K.; Zand, S.; Brem, H.; Peleg, S.; Dolan, P.; Kensler,
T. W. J. Med. Chem. 1999, 42, 3425.
13. Yamada, S.; Yamamoto, K.; Masuno, H.; Ohta, M.
J. Med. Chem. 1998, 41, 1467.
14. Sussman, F.; Rumbo, A.; Vilaverde, M. C.; Mourino, A.
J. Med. Chem. 2004, 47, 1613.
A 15 mL round bottomed flask was charged with the
coupled product (0.010 mg, 0.010 mmol) in THF
(5 mL). To this was added tetrabutylammonium fluoride
(0.083 mL, 1.0 M in THF, 0.083 mmol), and the result-
ing mixture was stirred at room temperature for 7 h in
dark. Then, it was quenched with H2O (2 mL), extracted
with EtOAc (3 · 25 mL), dried (MgSO4), and concen-
trated. Flash chromatography (EtOAc–hexanes, 25:75)
gave 0.0040 g (57%) of (+)-3. The product (+)-3 was fur-
ther purified by HPLC (silica semiprep (1 · 25 cm); flow
rate = 2.5 mL/min; P = 0.43 kpsi; EtOAc–hexanes =
1:1) to afford 0.002 mg of (+)-3 as an oil; [a]25 +10.5 (c
15. Perlman, K. L.; Swenson, R. E.; Paaren, H. E.; Schnoes,
H. K.; DeLuca, H. F. Tetrahedron Lett. 1991, 32, 7663.
16. Sicinski, R. R.; Rotkiewicz, P.; Kolinski, A.; Sicinska, W.;
Prahl, J. M.; Smith, C. M.; DeLuca, H. F. J. Med. Chem.
2002, 45, 3366.
17. Sicinski, R. R.; Prahl, J. M.; Smith, C. N.; DeLuca, H. F.
J. Med. Chem. 1998, 41, 4662.
18. Sicinska, W.; Rotkiewicz, P.; DeLuca, H. F. J. Steroid
Biochem. Mol. Biol. 2004, 89, 107.
19. Zhou, X.; Zhu, G.-D.; Van Haver, D.; Vandewalle, M.;
DeClercq, P. J.; Verstuyf, A.; Bouillon, R. J. Med. Chem.
1999, 42, 3539.
20. Hilpert, H.; Wirz, B. Tetrahedron 2001, 57, 681.
21. Posner, G. H.; Nelson, T. D.; Guyton, K. Z.; Kensler,
T. W. J. Med. Chem. 1992, 35, 3280.
22. Rochel, N.; Wurtz, J. M.; Mitschler, A.; Klaholz, B.;
Moras, D. Mol. Cell 2000, 5, 173.
23. An, M.; Bartlett, P. A. Org. Lett. 2004, 6, 4065.
24. Roberts, S. M.; Santoro, M. G.; Sickle, E. S. J. Chem.
Soc., Perkin Trans. 1 2002, 1735.
0.4, CHCl3); 1H NMR (CDCl3):
d
6.14 (d,
J = 11.2 Hz, 1H), 5.84 (d, J = 11.2 Hz, 1H), 5.44 (t,
J = 7.2 Hz, 1H), 4.19 (d, J = 7.2 Hz, 2H), 2.81 (m,
1H), 2.45–2.17 (m, 9H), 2.02–1.83 (m, 5H), 1.67–0.93
(m, 28H), 0.94 (d, J = 6.4 Hz, 3H), 0.56 (s, 3H); 13C
NMR (CDCl3): d 143.3, 141.5, 138.3, 121.2, 118.6,
115.6, 71.1, 58.7, 56.5, 56.3, 45.7, 44.4, 40.5, 38.3, 37.9,
36.4, 36.1, 29.7, 29.4, 29.0, 28.8, 27.7, 23.5, 22.3, 20.8,
18.8, 12.1. IR (neat): 3263, 2925, 2360, 1711, 1456,
25. Perlman, K. L.; Sicinski, R. R.; Schnoes, H. K.; DeLuca,
H. F. Tetrahedron Lett. 1990, 31, 1823.
26. Basra, S. K.; Drew, M. G. B.; Mann, J.; Kane, P. D. J.
Chem. Soc., Perkin Trans. 1 2000, 3592.
27. Bickey, J. F.; Roberts, S. M.; Santoro, M. G.; Snape, T. J.
Tetrahedron 2004, 60, 2569.
1373, 1020 cmꢀ1
;
HRMS calcd for C28H46O2Na+
[M+Na]: 437.3389. Found: 437.3375; UV (CH2Cl2) kmax
257 nm (e 33,300).
28. David, H.; Dupuis, L.; Guillerez, M.-G.; Guibe, F.
Tetrahedron Lett. 2000, 41, 3335.
Acknowledgements
29. Engel, P. S.; Allgren, R. L.; Chae, W.-K.; Leckonby, R.
A.; Marron, N. A. J. Org. Chem. 1979, 44, 4233.
We thank the NIH (CA 52857) for financial support.

2966
G. H. Posner et al. / Bioorg. Med. Chem. 13 (2005) 2959–2966
30. Posner, G. H.; Woodard, B. T.; Crawford, K. R.; Peleg,
S.; Brown, A. J.; Dolan, P.; Kensler, T. W. Bioorg. Med.
Chem. 2002, 10, 2353.
32. Posner, G. H.; White, M. C.; Dolan, P.; Kensler, T. W.;
Yukihiro, S.; Guggino, S. E. Bioorg. Med. Chem. Lett.
1994, 4, 2919.
31. Posner, G. H.; Lee, J. K.; Li, Z.; Takeuchi, K.; Guggino,
S. E.; Dolan, P.; Kensler, T. W. Bioorg. Med. Chem. Lett.
1995, 5, 2163.
33. Posner, G. H.; Dai, H. Bioorg. Med. Chem. Lett. 1993, 3,
1829.