Novel synthesis of bis(phosphonic acid)-steroid conjugates

Article abstract of DOI:10.1021/jo001489h

An efficient synthesis has been realized for several members of a new class of potential bone resorption inhibitors consisting of steroidal oestrogenic units linked at the 3 and 17 positions to a geminal bisphosphonate moiety through an ester linkage of variable length. The convergent synthesis utilizes benzyl bisphosphonates, transesterification, and Meldrum's acid chemistry and has the potential to allow many oestrogenic derivatives as well as other biologically active compounds to be coupled to the geminal bisphosphonate moeity.

Full text of DOI:10.1021/jo001489h

3704
J . Org. Chem. 2001, 66, 3704-3708
Novel Syn th esis of Bis(p h osp h on ic a cid )-Ster oid Con ju ga tes
Philip C. B. Page,*,† Michael J . McKenzie,† and J ames A. Gallagher‡
Department of Chemistry, Loughborough University, Loughborough, Leicestershire LE11 3TU, England,
and Department of Human Anatomy and Cell Biology, University of Liverpool,
Liverpool L69 3GE, England
Received October 17, 2000
An efficient synthesis has been realized for several members of a new class of potential bone
resorption inhibitors consisting of steroidal oestrogenic units linked at the 3 and 17 positions to a
geminal bisphosphonate moiety through an ester linkage of variable length. The convergent
synthesis utilizes benzyl bisphosphonates, transesterification, and Meldrum’s acid chemistry and
has the potential to allow many oestrogenic derivatives as well as other biologically active compounds
to be coupled to the geminal bisphosphonate moeity.
In tr od u ction
reducing unwanted side effects in other tissues. We
report here a new synthesis of geminal bisphosphonic
acids functionalized with derivatives of oestradiol, an
osteoclast inhibitor known to regulate the circuitry of
cytokine action that controls bone remodeling.7
Geminal bisphosphonates are stable pyrophosphate
analogues which bind efficiently to bone surface.1 They
have been used in the clinic to inhibit bone resorption,
in most cases with few side effects.2 Some simple ex-
amples, such as etidronate and alendronate, are already
marketed for the treatment of osteoporosis and Paget’s
disease.3 It has been suggested that bisphosphonates are
general metabolic inhibitors.4 The low level of side effects
has been attributed to the rapid absorption of the geminal
bisphosphonate to the bone surface and incorporation
within the bone matrix, thereby preventing undesired
effects within other organs.2 Other clinically used treat-
ments for osteoporosis, such as hormone replacement
therapy, are effective, but can have effects on other
tissues.5 We are interested in using geminal bisphospho-
nates as bone-targeting moieties for the treatment of
bone-related disease;6 our aim is to use the bone tropism
to deliver therapeutics at the desired site of action, thus
Discu ssion
We have recently reported the use of tetrabenzyl
bisphosphonates as synthetically useful precursors of
bisphosphonic acids, the conversion being experimentally
simple, clean, and high yielding.8 Bis(phosphonic acid)s
are typically produced from tetraalkyl bisphosphonate
esters either by acid hydrolysis9 or through silylation-
dealkylation using bromotrimethylsilane.10 While acid
hydrolysis is reliable it is often incompatible with other
functionalities. Silylation-dealkylation is a milder method;
(5) Beresford, S. A. A.; Weiss, N. S.; Voight, L. F.; McKnight, B.
Lancet 1997, 349, 458. Colditz, G. A.; Hankinson, S. E.; Hunter, D. J .;
Willet, W. C.; Manson, J . E.; Stampfer, M. J .; Hennekens, C.; Rosner,
B.; Speizer, F. E. New Engl. J . Med. 1995, 332, 1589. Sturtz, G.;
Guervenou, J . Phosphorus Sulfur, 1994, 88, 1. Sturtz, G.; Guervenou,
J . Fr. Patent Appl 2683527 (1991). Hiroaki, U.; Kadowaki, S.; Kami-
zono, A.; Masahiko, M.; Akihisa, M. Eur. Patent Appl 0555845 (1993).
Bosies, E.; Esswein, A.; Bauss, F. Int. Patent WO 92/05187 1992. Saari,
W. S.; Rodan, G. A.; Fisher, T. E.; Anderson, P. S. Eur. Patent Appl
0496520 A (1992). Sugioka, T.; Inazu, M. Eur. Patent Appl 0548884 A
(1992). Sturtz, G.; Guervenou, J . Synthesis 1991, 661. Bosies, E.;
Esswein, A.; Bauss, F. Eur. Patent 0549645 B1 (1991). Cust, M. P.;
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W. W.; Drife, J . O., Eds.; Springer-Verlag: London, 1990; p 209.
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Teitelbaum, S.; Slatopolski, E.; McCracken, R.; Bergfeld, M.; Lee, W.;
Avioli, L. V.; Peck, W. A. Proc. Natl. Acad. Sci. U.S.A. 1987, 84, 4616.
Oursler, M. J .; Keeting, P. E.; Wiegand, K. E.; Bonde, S. K.; Spielsberg,
T. C.; Riggs, B. L. J . Bone Min. Res., 1992, 7, S111. Oursler, M. J .;
Pederson, L.; Pyffereon, J .; Osdoby, P.; FitzPatrick, L.; Spielsberg, T.
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J .; Riggs, B. L.; Spielsberg, T. C. Proc. Natl. Acad. Sci. U.S.A. 1991,
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(9) For examples see: Mimura, M.; Hayashida, M.; Nomiyama, K.;
Ikegami, S.; Iida, Y. Chem. Pharm. Bull., 1993, 41, 1971. Dufau, C.;
Sturtz, G. Phosphorus Sulfur 1992, 69, 93. den Hartog, J . A. J .;
Wijnands, R. A.; van Boom, J . H.; Crea, R. J . Org. Chem. 1981, 46,
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McKenna, C. E.; Schmidhauser, J . J . Chem. Soc., 1979, 35, 739.
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† Loughborough University.
‡ University of Liverpool.
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G. B.; Francis, M. D.; Mosekilde, L.; Hasling, C. J . Bone Miner. Res.
1992, 7, S328. Storm, T.; Thamsborg, G.; Steiniche, T.; Genant, H. K.;
Sorensen, O. H. New Engl. J . Med. 1990, 332, 1265; Fleisch, H. Bone
1999, 25, 158. Vitte, C.; Fleisch, H.; Guenther, H. L. Endocrinology
1996, 137, 2324. Fleisch, H. Osteoporosis Int. 1996, 6, 166.
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R.; Buckhardt, P.; Delmers, P. D.; Fleisch, H.; Kanis, J . A.; Kyle, R.
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1998, 16, 3890. Guenther, H. L.; Fleisch, H.; Walz, A. J . Bone Miner.
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10.1021/jo001489h CCC: $20.00 © 2001 American Chemical Society
Published on Web 05/05/2001

Bis(phosphonic acid)-Steroid Conjugates
J . Org. Chem., Vol. 66, No. 11, 2001 3705
however, in our hands reactions were often incomplete,
leading to mixtures of products. This prompted us to
investigate the benzyl group as a potential protecting
group for bis(phosphonic acid)s which might be easily and
selectively removed.11 Hydrogenolysis of benzyl phospho-
nates has been widely utilized;12 however, to our knowl-
edge geminal bis(phosphonic acid)s have rarely been
synthesized in this way. Mioskowski has shown that
simple benzyl bisphosphonate esters can be deprotected
by the use of catalytic transfer hydrogenolysis13 or with
DABCO in boiling toluene,14 or even converted into
chlorophosphonates.15 More recently, a 2-substituted
geminal bis(phosphonic acid) has also been prepared by
catalytic transfer hydrogenolysis.16
We wished to incorporate this chemistry in the syn-
thesis of steroidal bis(phosphonic acid) conjugates, and
we now report some of our results in this area. In the
design of such compounds it is vital to consider the
linkage between the geminal bisphosphonate moiety and
the oestrogenic unit. A conjugate with a linker not readily
cleaved by hydrolysis or by enzyme action may not retain
oestrogenic activity, while a conjugate with a linker too
readily cleaved would not achieve targeting. Oestrogen-
bisphosphonate conjugates are known with nonhydro-
lyzable linkers,17 and with ether,18 carbonate,19 and
amide20 linkers. Cortisone-bisphosphonate conjugates
are known with ester linkers.21 We reasoned that a
carboxylic ester linkage might provide an appropriate
balance between stability and ease of cleavage, and,
further, that this balance might be adjusted by, for
example, altering the steric environment around the ester
unit. In the case of oestradiol, such an ester linkage might
readily be positioned at either the 3- or 17- hydroxylated
positions, and this design would in addition allow the
incorporation of other hydroxyl functionalized biologically
active compounds. Initially, methodology was established
using the commercially available tetraethyl methylene-
bisphosphonate. Sturtz has reported the synthesis of the
propanoic 1 and butanoic 2 acid derivatives for the
synthesis of cortisone-derived conjugates, using standard
peptide coupling techniques.22 He derived bisphosphonate
2 from tetraethyl ethylidene-1,1-bisphosphonate 3 and
diethyl malonate, followed by basic hydrolysis and de-
carboxylation.
We reasoned that we could shorten this procedure
considerably using the cyclic malonate 2,2-dimethyl-1,3-
dioxane-4,6-dione (Meldrum’s acid) as a preactivated
carboxylic acid equivalent.23 Tetraethyl methylenebis-
phosphonate was converted into 3 in multigram quanti-
ties using the method of Degenhardt.24 Michael addition
of stabilized nucleophiles to 3 is known.21,22 Conjugate
addition of Meldrum’s acid using mediated lithium bis-
(trimethylsilyl)amide in THF was successful, but yields
of 4 varied considerably. When DBU was used as the
basic mediator,25 however, reactions were very reliable,
giving essentially pure 4 (Scheme 1).
Sch em e 1a
a
(i) (HCHO)n, HNEt2, MeOH; pTSA, Tol, ∆; (ii) Meldrum’s acid
derivative, DBU, THF, RT; 4: 97%; 5: 65%.
Preliminary attempts at coupling 4 with pregnenolone
by heating in acetonitrile gave 6 as desired, but with only
75% conversion after 24 h at reflux. Gratifyingly, by
switching to toluene the reaction was complete in 4 h.
The transformation was also successful for other steroids
such as oestrone and trans-androsterone, to give 7 and
8, respectively (Scheme 2). Interestingly, compound 5,
(11) Greene, T. W.; Wuts, P. G. M. Protecting Groups in Organic
Synthesis, 2nd ed.; Wiley-Interscience: New York, 1991. Kocienski,
P. J . Protecting Groups; Thieme: Stuttgart, 1994.
(12) Amann, F.; Schaub, C.; Mueller, B.; Schmidt, R. R. Chem. Eur.
J . 1998, 4, 1106. Lampe, T. F. J .; Weitz-Schmidt, G.; Wong, C.-H.
Angew. Chem., Int. Ed. 1998, 37, 1707. Cristau, H.-J .; Lambert, J .-
M.; Pirat, J .-L. Synthesis 1998, 8, 1167. Hoffmann, M. Synthesis 1988,
62. Alewood, P. F.; J ohns, R. B.; Hoogenraad, N. J .; Sutherland, T.
Synthesis 1984, 403.
Sch em e 2a
(13) Saady, M.; Lebeau, L.; Mioskowski, C. Helv. Chim. Acta 1995,
78, 670. Hoffmann, M.; Wasielewski, C. Phosphorus Sulfur 1990, 53,
69. Mikroyannidis, J . A.; Tsolis, A. K.; Gourghiotis, D. J . Phosphorus
Sulfur 1982, 13, 279.
(14) Saady, M.; Lebeau, L.; Mioskowski, C. J . Org. Chem. 1995, 60,
2946.
(15) Mioskowski, C.; Saady, M.; Lebeau, L. Tetrahedron Lett. 1995,
36, 4785.
(16) Gil, L.; Han, Y.; Opas, E. E.; Redan, G. A.; Ruel, R. Bioorg. Med.
Chem. 1999, 7, 901.
(17) Kirin, S. J apan Patent 6-100576.
a
(i) Steroid, toluene, ∆; 6: 87%; 7: 77%; 8: 99%.
(18) Sugioka, T.; Inza, M. Eur Patent 0548884 A1.
(19) Saari, W. S.; Rodan, G. A.; Fisher, T. A.; Anderson, P. S. Eur
Patent 0496520 A1.
(20) Ueno, H.; Kadowaki, S.; Kamizono, A.; Morioka, M. Eur Patent
0555845 A2.
(21) Sturtz, G.; Guervenou, J . Phosphorus Sulfur 1994, 88, 1.
(22) Hutchinson, D. W.; Thorton, D. M. J . Organomet. Chem. 1988,
346, 341; Sturtz, G.; Guervenou, J . Synthesis 1991, 661.
(23) McNab, H. Chem. Soc. Rev. 1978, 7, 345.
(24) Degenhardt, C. R.; Burdsall, D. C. J . Org. Chem. 1986, 51, 3488.
(25) Nugent, R. A.; Schlachter, M.; Murphy, M.; Dunn, C. J .; Staite,
N. D.; Galinet, L. A.; Shields, S. K.; Wu, H.,; Aspar, D. G.; Richard, K.
A. J . Med. Chem. 1994, 37, 4449.

3706 J . Org. Chem., Vol. 66, No. 11, 2001
Page et al.
Sch em e 3a
Sch em e 5a
a
(i) BnOH, pyridine, toluene, 0 °C; 76% (ii) (HCHO)n, HNEt2,
MeOH, RT; pTSA, Tol, ∆; 99%; (iii) Meldrum’s acid, DBU, THF,
98%.
Sch em e 4a
a
(i) NaH, THF, ethyl bromoacetate; 60%; (ii) KOH, MeOH/H2O;
78%; (iii) steroid, DCC, DMAP, CH2Cl2; 15: 86%; 16: 75%.
Sch em e 6
oestrone, proved less stable, with some degree of ester
hydrolysis (up to 20%) always observed on hydrogenolysis
in a range of solvents. We have also observed this
phenolic ester hydrolysis using standard trimethylsilyl
bromide-promoted cleavage of the corresponding tetra-
ethyl conjugate 8.
In conclusion, we have prepared a range of bis-
(phosphonic acid)-steroid conjugates for testing as novel
potential bone targeting therapeutics. We have developed
an expedient route to the butanoic acid derivatives using
Meldrum’s acid as a masked activated carboxylic acid,
and we have shown the utility of tetrabenzyl bisphos-
phonates as bis(phosphonic acid) precursors.
a
(i) Steroid, toluene, ∆; 12: 60%; 13: 86%; 14: 76%.
derived from 2,2,5-trimethyl-1,3-dioxane-4,6-dione and 3,
was unreactive toward the same steroids under these
conditions.
This synthesis of conjugates was short and convenient,
the final step eliminating the need for any reagents. We
next applied this methodology to tetrabenzyl methyl-
enebisphosphonate conjugates. We have previously re-
ported the benzyl bisphosphonates 9 and 10, prepared
as shown, for the synthesis of oxiranylidene-2,2-bis-
(phosphonic acid).8 Reaction of 10 with Meldrum’s acid
in the presence of catalytic DBU in THF solution yielded
11 quantitatively (Scheme 3). Synthesis of 12 and 13 was
readily achieved by heating 11 with oestrone and trans-
androsterone respectively in toluene. The sterically con-
gested 3-benzyl-17â-oestradiol also reacted successfully,
giving 14 after overnight reflux (Scheme 4).26
To provide a range of steroidal bisphosphonate conju-
gates with a spectrum of lability toward hydrolysis in
vivo, we also prepared the propanoic acid derivatives 15
and 16 as shown in Scheme 5. Alkylation of 9 with ethyl
bromoacetate produced ethyl ester 17, which was hydro-
lyzed to produce carboxylic acid 18.21,22 Standard DCC
coupling with trans-androsterone and oestrone gave the
corresponding conjugates 15 and 16.
Exp er im en ta l Section
Tetr a eth yl 2-(2,2-Dim eth yl-1,3-d ioxa n e-4,6-d ion e)eth -
ylen e Bisp h osp h on a te 4. DBU (0.051 g, 0.33 mmol) was
added to a solution of tetraethyl ethylidene-1,1-bisphosphonate
3 (1.00 g, 3.33 mmol) and 2,2-dimethyl-1,3-dioxane-4,6-dione
(0.480 g, 3.33 mmol) in dry THF (10 mL) at 20 °C. After 4 h
the reaction was diluted with dichloromethane (30 mL) and
washed with water (20 mL). Drying (MgSO4) and concentration
in vacuo gave 4 as a colorless oil (1.43 g, 97%): IR (liq) 972,
1251, 1748, 1785. 1H NMR (CDCl3, 250 MHz) δ 1.36 (12H, t, J
) 7.1 Hz), 1.76 (3H, s), 1.87 (3H, s), 2.50-2.68 (2H, m), 3.17
(1H, tt, J ) 7.7, 23.2 Hz), 4.13-4.27 (8H, m), 5.13 (1H, t, J )
6.2 Hz). MS (EI) 444.1327, C16H30P2O10 requires 444.1314.
Tet r a et h yl 2-(2,2,5-Tr im et h yl-1,3-d ioxa n e-4,6-d ion e)-
eth ylen e Bisp h osp h on a te 5. LHMDS (0.33 mL, 0.016 mmol)
was added to a solution of tetraethyl ethylidene-1,1-bis-
(phosphonate) 3 (0.50 g, 0.166 mmol) and 2,2,5-trimethyl-1,3-
dioxane-4,6-dione (0.263 g, 0.168 mmol) in dry THF (20 mL)
at 20 °C. After 16 h the reaction was concentrated in vacuo,
diluted with dichloromethane (25 mL), and washed with water
(3 mL). Drying (Na2SO4), concentration in vacuo, and flash
column chromatography (2% MeOH/ CH2Cl2) gave 5 as a
Conversion of 13, 14, 15, and 16 into the corresponding
bis(phosphonic acid)s was accomplished by hydrogenoly-
sis over 10% palladium on charcoal, the reactions requir-
ing only filtration and evaporation to provide isolated
products (Scheme 6). Conjugate 12, derived from
(26) 3-O-Benzyl-17â-oestradiol was synthesized from oestrone using
standard procedures in two steps: Levitz, M. J . Am. Chem. Soc. 1953,
75, 5352.
1
colorless oil (0.496 g, 65%): IR (liq) 974, 1256, 1736, 1771. H

Bis(phosphonic acid)-Steroid Conjugates
J . Org. Chem., Vol. 66, No. 11, 2001 3707
NMR (CDCl3, 250 MHz) δ 1.35 (12H, t, J ) 7.0 Hz), 1.65 (3H,
s), 1.77 (3H, s), 1.87 (3H, s), 2.65 (2H, td, J ) 7.1, 15.5 Hz),
2.97 (1H, tt, J ) 6.9, 24.0 Hz), 4.07-4.23 (8H, m).
in vacuo gave 10 as a colorless viscous oil (5.10 g, 99%): IR
1
(liq) 996, 1248. H NMR (CDCl3, 400 MHz) δ 4.95-5.05 (8H,
m), 6.98 (2H, dd, J ) 34.4, 38.9 Hz), 7.24-7.30 (20H, m). MS
Tetr a eth yl (3-(3â-(5-P r egn en ol-20-on ly)oxyca r bon yl)-
p r op ylen e) Bisp h osp h on a te 6. A solution of 4 (0.250 g,
0.546 mmol) and pregnenolone (0.178 g, 0.563 mmol) in toluene
(10 mL) was heated under reflux for 4 h. Concentration in
vacuo and flash column chromatography (EtOAc then 10%
EtOH/EtOAc) gave 6 as a colorless oil (0.323 g, 87%): IR (liq)
969, 1254, 1704, 1731. 1H NMR (CDCl3, 400 MHz) δ 0.63 (3H,
s), 1.02 (3H, s), 1.21-1.29 (3H, m), 1.35 (12H, t, J ) 7.0 Hz),
1.45-1.73 (8H, m), 1.84-1.89 (2H, m), 1.97-2.07 (3H, m), 2.13
(3H, s), 2.15-2.33 (5H, m), 2.51 (1H, tt, J ) 6.4, 22.1 Hz),
2.51-2.57 (1H, m), 2.64 (2H, t, J ) 7.5 Hz), 4.15-4.23 (8H,
m), 4,57-4.66 (1H, m), 5.37 (1H, br d, J ) 4.9 Hz). MS (FAB)
659 (M + H+), 681 (M + Na+).
(EI) 548.1524, C30H30P2O6 requires 548.1517.
Tet r a b en zyl 2-(2,2-Dim et h yl-1,3-d ioxa n e-4,6-d ion e)-
eth ylen e Bisp h osp h on a te 11. DBU (0.060 g, 0.37 mmol) was
added to a solution of tetrabenzyl ethylidene 1,1-bisphospho-
nate 10 (2.00 g, 3.65 mmol) and 2,2-dimethyl-1,3-dioxane-4,6-
dione (0.525 g, 3.65 mmol) in dry THF (20 mL) at 20 °C. After
4 h the reaction was diluted with dichloromethane (30 mL)
and washed with water (20 mL). Drying (MgSO4) and concen-
tration in vacuo gave 11 as a colorless oil (2.48 g, 98%): IR
(liq) 998, 1260, 1748, 1784. 1H NMR (CDCl3, 250 MHz) 1.55
(3H, s), 1.67 (3H, s), 2.61-2.73 (2H, m), 3.36 (1H, tt, J ) 7.6,
23.2 Hz), 4.77 (1H, t, J ) 6.0 Hz), 4.99-5.04 (8H, m), 7.22-
7.34 (20H, m). MS (ES) 693.2016; C36H39P2O10 requires 693.2018
(M + +H).
Tetr a ben zyl (3-(3-(1,3,5-Oestr a tr ien -17-on yl)oxyca r bo-
n yl)p r op ylen e) Bisp h osp h on a te 12. A solution of 11 (0.300
g, 0.434 mmol) and oestrone (0.120 g, 0.444 mmol) in toluene
(15 mL) was heated under reflux for 4 h. Concentration in
vacuo and flash column chromatography (40% EtOAc/petro-
leum ether) gave 12 as a colorless oil (0.283 g, 60%): IR (liq)
996, 1257, 1732, 1755. 1H NMR (CDCl3, 400 MHz) δ 0.91 (3H,
s), 1.39-1.67 (6H, m), 1.92-2.18 (5H, m), 2.25-2.43 (3H, m),
2.47-2.53 (1H, m), 2.70 (1H, tt, J ) 6.4, 23.6 Hz), 2.82-2.89
(4H, m), 4.98-5.07 (8H, m), 6.70-6.76 (2H, m), 7.23 (1H, d, J
) 9.2 Hz), 7.26-7.34 (20H, m). MS (ES) 861.3319; C50H53P2O9
requires 861.3321 (M+ - H).
Tetr a ben zyl (3-(3â-(5r-An d r os-17-on yl)oxyca r bon yl)-
p r op ylen e) Bisp h osp h on a te 13. A solution of 11 (0.200 g,
0.289 mmol) and trans-androsterone (0.084 g, 0.289 mmol) in
toluene (8 mL) was heated under reflux for 4 h. Concentration
in vacuo and flash column chromatography (50% EtOAc/
petroleum ether) gave 13 as a colorless oil (0.219 g, 86%): IR
(liq) 997, 1256, 1731, 1738. 1H NMR (CDCl3, 400 MHz) δ 0.65-
0.73 (1H, m), 0.82 (3H, s), 0.85 (3H, s), 0.94-1.03 (2H, m),
1.12-1.31 (6H, m), 1.40-1.58 (4H, m), 1.62-1.81 (5H, m)
1.89-1.97 (2H, m), 2.06 (1H, dt, J ) 9.2, 19.6 Hz) 2.19-2.33
(2H, m), 2.39-2.46 (1H, m), 2.56 (2H, t, J ) 7.4 Hz), 2.63 (1H,
tt, J ) 6.4, 24.0 Hz), 4.58-4.67 (1H, m), 4.98-5.05 (8H, m),
7.25-7.31 (20H, m). MS (ES) 881.3940; C51H63P2O9 requires
881.3947 (M+ + H).
Tetr a eth yl (3-(3-(1,3,5-Oestr a tr ien -17-on yl)oxyca r bo-
n yl)p r op ylen e) Bisp h osp h on a te 7. A solution of 4 (1.280
g, 2.79 mmol) and oestrone (0.778 g, 2.88 mmol) in toluene
(40 mL) was heated under reflux for 2.5 h. Concentration in
vacuo and flash column chromatography (EtOAc then 10%
EtOH/EtOAc) gave 7 as a colorless oil (1.314 g, 77%): IR (liq)
1
1738. H NMR (CDCl3, 250 MHz) δ 0.91 (3H, s), 1.35 (12H, t,
J ) 7.0 Hz), 1.45-1.63 (7H, m), 1.94-2.55 (9H, m), 2.61 (1H,
tt, J ) 6.4, 23.4 Hz), 2.88-2.96 (3H, m), 4.14-4.26 (8H, m),
6.80-6.87 (2H, m), 7.28 (1H, d, J ) 8.5 Hz). MS (FAB) 613 (M
+ H+), 635 (M + Na+).
Tetr a eth yl (3-(3â-(5r-An d r oster -17-on yl)oxyca r bon yl)-
p r op ylen e) Bisp h osp h on a te 8. A solution of 4 (0.250 g,
0.546 mmol) and trans-androsterone (0.158 g, 0.546 mmol) in
toluene (10 mL) was heated under reflux for 4 h. Concentration
in vacuo gave 8 as a colorless oil (0.345 g, 99%): IR (liq) 1026,
1
1251, 1738. H NMR (CDCl3, 400 MHz) δ 0.66-0.75 (1H, m),
0.85 (3H, s), 0.86 (3H, s), 0.93-1.07 (2H, m), 1.15-1.38 (7H,
m), 1.35 (12H, t, J ) 7.1 Hz), 1.46-1.57 (3H, m), 1.59-1.68
(2H, m), 1.72-1.83 (4H, m), 1.90-1.96 (1H, m), 2.02-2.12 (1H,
m), 2.14-2.27 (2H, m), 2.41-2.48 (1H, m), 2.51 (1H, tt, J )
6.4, 23.8 Hz), 2.63 (2H, t, J ) 7.5 Hz), 4.15-4.23 (8H, m), 4.65-
4.73 (1H, m).). MS (FAB) 633 (M + H+), 655 (M + Na+). Anal.
Calcd for C31H54P2O9: C, 57.55; H, 8.10. Found: C, 57.22; H,
8.31.
Tetr a ben zyl Meth ylen e Bisp h osp h on a te 9. A suspen-
sion of methylene bis(phosphonic dichloride) (5.00 g, 20.0
mmol) was stirred rapidly in dry toluene (10 mL) at 0 °C. A
mixture of dry benzyl alcohol (8.66 mL, 83.0 mmol) and dry
pyridine (6.15 mL, 76.1 mmol) was added over 80 min by
syringe pump while the temperature was maintained at 0 °C.
After the addition was complete, the reaction was allowed to
reach 20 °C and stirred for a further 3 h. The solids were
removed by filtration and washed twice with toluene (2 × 20
mL). The filtrate was washed twice with 2 M NaOH (2 × 15
mL) and water (15 mL), dried (MgSO4), and concentrated in
vacuo. Removal of benzyl alcohol impurity by distillation (120
°C, 1 mmHg) gave 9 as a colorless oil (8.11 g, 76%): IR (liq)
Tetr a ben zyl (3-(17â-(3-O-Ben zyloestr a -1,3,5-tr ien yl)-
oxyca r bon yl)p r op ylen e) Bisp h osp h on a te 14. A solution
of 11 (0.200 g, 0.289 mmol) and 3-O-benzyl-17â-oestradiol
(0.104 g, 0.289 mmol) in toluene (6 mL) was heated under
reflux for 16 h. Concentration in vacuo and flash column
chromatography (40% EtOAc/petroleum ether) gave 14 as a
1
colorless oil (0.208 g, 76%): IR (liq) 996, 1254, 1731. H NMR
(CDCl3, 400 MHz) δ 0.74 (3H, s), 1.15-1.47 (8H, m), 1.65-
1.72 (1H, m), 1.77-1.87 (2H, m), 2.12-2.33 (6H, m), 2.61 (2H,
t, J ) 8.0 Hz), 2.63 (1H, tt, J ) 6.4, 24.4 Hz), 2.81-2.86 (2H,
m), 4.63 (1H, dd, J ) 8.0, 8.8 Hz), 4.99-5.06 (8H, m), 6.70-
6.71 (1H, m), 6.76-6.79 (1H, m), 7.17-7.19 (1H, m), 7.25-
7.42 (25H, m). MS (FAB) 953.4005; C57H63P2O9 requires
953.3947 (M+ + H).
1
998, 1260. H NMR (CDCl3, 250 MHz) δ 2.51 (2H, t, J ) 21.2
Hz), 5.00 (4H, d, J ) 2.2 Hz), 5.05 (4H, d, J ) 2.2 Hz), 7.28
(20H, br s). MS (EI) 536.15082, C29H30P2O6 requires 536.1517.
Tetr a ben zyl Eth ylid en e 1,1-Bisp h osp h on a te 10. Para-
formaldehyde (1.35 g, 45.0 mmol) and diethylamine (0.68 g,
9.33 mmol) were dissolved in dry methanol (30 mL) with
warming. A solution of 9 (5.00 g, 9.33 mmol) in dry methanol
(30 mL) was added at 20 °C and the reaction stirred for 5 d.
The reaction was concentrated in vacuo, toluene (20 mL)
added, and the solution concentrated again. This last step was
repeated to remove all traces of methanol, yielding tetrabenzyl
1-methoxymethyl methylene 1,1-bisphosphonate as a colorless
Eth yl 3,3-Bis(d iben zyloxyp h osp h or yl)p r op a n oa te 17.
A solution of 9 (1.00 g, 1.87 mmol) in THF (10 mL) was added
to a suspension of NaH (0.047 g, 1.96 mmol) in THF (20 mL)
at 0 °C over 5 min. The solution was allowed to reach 20 °C,
stirring continued until effervescence ceased (30 min ap-
proximately), and ethyl bromoacetate (0.22 mL, 1.98 mmol)
added. After 16 h, normal workup, extraction with dichlo-
romethane and flash column chromatography (70% EtOAc/
petroleum ether) gave 17 as a colorless oil (0.694 g, 60%): IR
1
1
oil. H NMR (CDCl3, 250 MHz) δ 2.86 (tt, J ) 5.5, 23.9 Hz),
(liq) 994, 1256, 1737. H NMR (CDCl3, 400 MHz) δ 1.11 (3H,
3.30 (s), 3.93 (td, J ) 5.6, 16.3 Hz), 4.99-5.06 (8H, m), 7.28-
t, J ) 7.1 Hz), 2.86 (2H, td, J ) 6.3, 16.1 Hz), 3.30 (1H, tt, J
) 6.3, 23.9 Hz), 3.93 (2H, q, J ) 7.2 Hz), 4.99-5.04 (8H, m),
7.25-7.34 (20H, m). MS (EI) 622.1872; C33H36P2O8 requires
622.1885 (M+).
7.33 (20H, m).
The tetrabenzyl 1-methoxymethyl methylene 1,1-bisphos-
phonate intermediate was added to pTSA (cat) and toluene
(100 mL), and the mixture was heated under reflux through
a Soxhlet apparatus containing 4 Å molecular sieve for 16 h.
The mixture was allowed to cool to 20 °C and washed twice
with water (2 × 20 mL). Drying (MgSO4) and concentration
3,3-Bis(d iben zyloxyp h osp h or yl)p r op a n oic Acid 18. A
solution of 17 (1.27 g, 2.04 mmol) in methanol (6 mL) was
added to a solution of KOH (0.127 g, 2.26 mmol) in water (6
mL) and methanol (6 mL) at 0 °C. The reaction was allowed

3708 J . Org. Chem., Vol. 66, No. 11, 2001
Page et al.
to reach 20 °C over 1 h and stirred at this temperature for 16
h. Methanol was removed in vacuo and the remaining solution
extracted with diethyl ether (20 mL). The aqueous layer was
acidified to pH 2 with aqueous KHSO4 and extracted with
dichloromethane (3 × 20 mL) and EtOAc (1 × 20 mL). The
combined organic extracts were dried (MgSO4) and concen-
trated in vacuo to give 18 as a colorless oil (0.944 g, 78%): IR
m), 2.24-2.29 (1H, m), 2.60-2.84 (6H, m), 4.66 (1H, t, J ) 8.4
Hz), 6.49-6.50 (1H, m), 6.55-6.58 (1H, m), 7.08-7.11 (1H,
m). MS (ES) 487.1287; C21H29P2O9 requires 487.1287 (M+
H).
-
3-(17â-(O e s t r a -1,3,5-t r ie n y l)o x y c a r b o n y l)p r o p y li-
d en e Bis(p h osp h on ic a cid ) 20. A solution of 14 (0.090 g,
0.095 mmol) and 10% Pd(C) (0.005 g) in methanol (2 mL) and
1
(liq) 995, 1252, 1742. H NMR (CDCl3, 250 MHz) δ 2.90 (2H,
dichloromethane (1 mL) was stirred under a balloon of
td, J ) 6.1, 16.3 Hz), 3.23 (1H, tt, J ) 5.9, 24.0 Hz), 4.95-
5.01 (8H, m), 7.20-7.32 (20H, m).
hydrogen for 16 h. The Pd(C) was removed by filtration, the
residue washed with methanol (5 mL), and the combined
organic solution concentrated in vacuo to give 20 as a colorless
powder (0.046 g, 97%). 1H NMR (CD3OD, 400 MHz) δ 0.86 (3H,
s), 1.25-1.47 (6H, m), 1.53-1.62 (1H, m), 1.72-1.79 (1H, m),
1.85-1.89 (2H, m), 2.12-2.30 (5H, m), 2.36 (1H, tt, J ) 6.4,
23.6 Hz), 2.67-2.82 (4H, m), 4.68 (1H, dd, J ) 8.0, 8.8 Hz),
6.47-6.48 (1H, m), 6.52-6.58 (1H, m), 7.05-7.07 (1H, m). MS
(ES) 501.1438; C22H31P2O9 requires 501.1443 (M+ - H).
3-(3â-(5r-An d r ost er -17-on yl)oxyca r b on yl)et h ylid en e
Bis(p h osp h on ic a cid ) 21. A solution of 16 (0.045 g, 0.050
mmol) and 10% Pd(C) (0.005 g) in methanol (2 mL) was stirred
under a balloon of hydrogen for 16 h. The Pd(C) was removed
by filtration, the residue washed with methanol (5 mL), and
the combined organic solution concentrated in vacuo to give
21 as a colorless powder (0.025 g, 95%). 1H NMR (CD3OD, 400
MHz) δ 0.74-0.80 (1H, m), 0.87 (3H, s), 0.90 (3H, s), 1.02-
1.49 (10H, m), 1.53-2.10 (10H, m), 2.42 (1H, dd, J ) 8.4, 18.8
Hz) 2.75-2.95 (3H, m), 4.64-4.77 (1H, m). MS (ES) 505.1758
(M - H), C22H35P2O9 requires 505.1756.
3-(3â-(5r-An d r ost e r -17-on yl)oxyca r b on yl)p r op yli-
d en e Bis(p h osp h on ic a cid ) 22. A solution of 13 (0.053 g,
0.060 mmol) and 10% Pd(C) (0.005 g) in methanol (1 mL) was
stirred under a balloon of hydrogen for 16 h. The Pd(C) was
removed by filtration, the residue washed with methanol (5
mL), and the combined organic solution concentrated in vacuo
to give 22 as a colorless powder (0.030 g, 96%). 1H NMR (CD3-
OD, 400 MHz) δ 0.73-0.79 (1H, m), 0.87 (3H, s), 0.89 (3H, s),
1.00-1.09 (2H, m), 1.19-1.42 (6H, m), 1.48-1.70 (4H, m),
1.72-1.84 (4H, m) 1.91-1.99 (1H, m), 2.03 (1H, dt, J ) 9.0,
19.2 Hz), 2.18-2.35 (3H, m), 2.39-2.46 (1H, m), 2.65 (2H, t, J
) 7.6 Hz), 4.66-4.72 (1H, m). MS (ES) 519.1917; C23H37P2O9
requires 519.1913 (M+ - H).
Tetr a ben zyl (3-(17â-(3-O-Ben zyloestr a -1,3,5-tr ien yl)-
oxyca r bon yl)eth ylen e) Bisp h osp h on a te 15. Dicyclohexyl-
carbodiimide (0.175 g, 0.751 mmol) and DMAP (0.010 g, 0.082
mmol) were added to a solution of 18 (0.300 g, 0.505 mmol)
and 3-O-benzyl-17â-oestradiol (0.184 g, 0.508 mmol) in dichlo-
romethane (8 mL). The solution was stirred for 16 h and
filtered, and the filtrate was concentrated in vacuo. Flash
column chromatography (50% EtOAc/petroleum ether) gave
15 as a colorless oil (0.409 g, 86%): IR (liq) 1018, 1253, 1732.
1H NMR (CDCl3, 400 MHz) δ 0.73 (3H, s), 1.10-1.45 (6H, m),
1.62-1.69 (2H, m), 1.73-1.76 (1H, m), 1.82-1.85 (1H, m),
2.02-2.20 (3H, m), 2.79-2.85 (2H, m), 2.91 (2H, td, J ) 6.2,
16.2 Hz), 3.33 (1H, tt, J ) 6.2, 23.9 Hz), 4.57 (1H, dd, J ) 7.9,
8.8 Hz), 5.01-5.08 (10H, m), 6.69-6.71 (1H, m), 6.76-6.79
(1H, m), 7.15-7.18 (1H, m), 7.26-7.42 (25H, m). MS (FAB)
939 (M + H+), 961 (M + Na+).
Tetr a ben zyl (3-(3â-(5r-An d r oster -17-on yl)oxyca r bon -
yl)eth ylen e) Bisp h osp h on a te 16. Dicyclohexylcarbodiimide
(0.078 g, 0.335 mmol) and DMAP (0.005 g, 0.041 mmol) were
added to a solution of 18 (0.15 g, 0.253 mmol) and trans-
androsterone (0.073 g, 0.251 mmol) in dichloromethane (15
mL). The solution was stirred for 16 h and filtered, and the
filtrate was concentrated in vacuo. Flash column chromatog-
raphy (50% EtOAc/petroleum ether) gave 16 as a colorless oil
(0.164 g, 75%): IR 1015, 1249, 1730. 1H NMR (CDCl3, 400
MHz) δ 0.62-0.70 (1H, m), 0.79 (3H, s), 0.85 (3H, s), 0.88-
1.00 (2H, m), 1.01-1.67 (12H, m), 1.75-1.94 (5H, m), 2.06 (1H,
dt, J ) 8.8, 19.2 Hz), 2.42 (1H, dd, J ) 8.8, 19.2 Hz), 2.85 (2H,
td, J ) 6.4, 16.4 Hz), 3.30 (1H, tt, J ) 6.2, 23.6 Hz), 4.47-
4.56 (1H, m), 4.97-5.04 (8H, m), 7.26-7.30 (20H, m). MS (ES)
867.3784; C50H61P2O9 requires 867.3791 (M+ + H).
3-(17â-(Oestr a-1,3,5-tr ien yl)oxycar bon yl)eth yliden e Bis-
(p h osp h on ic a cid ) 19. A solution of 15 (0.200 g, 0.213 mmol)
and 10% Pd(C) (0.020 g) in dry acetone (5 mL) was stirred
under a balloon of hydrogen for 16 h. The Pd(C) was removed
by filtration, the residue washed with acetone (2 × 20 mL),
and the combined organic solution concentrated in vacuo to
give 19 as a colorless powder (0.081 g, 78%). 1H NMR ((CD3)2-
CO), 400 MHz) δ 0.85 (3H, s), 1.23-1.46 (6H, m), 1.55-1.63
(1H, m), 1.69-1.77 (1H, m), 1.83-1.93 (1H, m), 2.04-2.19 (2H,
Ack n ow led gm en t. This investigation has enjoyed
the support of Shire Pharmaceuticals Ltd.
Su p p or tin g In for m a tion Ava ila ble: Additional analyti-
1
cal data; H NMR spectra. This material is available free of
charge via the Internet at http://pubs.acs.org.
J O001489H