Calcitriol derivatives with two different side chains at C-20. V. Potent inhibitors of mammary carcinogenesis and inducers of leukemia differentiation

J. Med. Chem. 2009, 52, 5505–5519 5505

DOI: 10.1021/jm900780q

Calcitriol Derivatives with Two Different Side Chains at C-20. V. Potent Inhibitors of Mammary

Carcinogenesis and Inducers of Leukemia Differentiation^†

Hubert Maehr,* Hong Jin Lee, Bradford Perry, Nanjoo Suh, and Milan R. Uskokovic

Department of Chemical Biology, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, 164 Frelinghuysen Road,

Piscataway, New Jersey 08854

Received June 1, 2009

Calcitriol is implicated in many cellular functions including cellular growth and differentiation, thus

explaining its antitumor effects. It was shown that gemini, the calcitriol derivative containing two side

chain at C20, is also active in gene transcription with enhanced antitumor activity. We have now further

optimized both the A-ring and the two side chains. The chemical structures of the resulting 18 geminis

were correlated with biological activities. Those containing the 1R-fluoro A-ring are the least active.

Those featuring 23-yne and 23(E) side-chains are generally more active in human breast cancer cell

growth inhibition and human leukemia cell differentiation induction than their 23(Z) counterparts. On

the basis of these evaluations, we selected as lead compound a 20(R) gemini, related to calcitriol in terms

of it is A-ring, where one side chain was modified by introduction of a 23-yne function and replacement

of the geminal methyl groups with trifluoromethyl groups, the other created by extension of C21 with a

3-hydroxy-3-trideuteromethyl-4,4,4-trideutero-butyl moiety.

Introduction

Gemini is the cumulative term for vitamin D analogues

with two side chains emanating at C-20. In the first gene-

ration of this compound category, both side chains were

identical as in 1a. The arrangement comprises the unaltered

chain of 1R,25(OH)₂D₃(1) and a duplicate thereof as an

extension of the methyl group 21.¹From a historical view-

point, it was argued that 20-epi-1 exhibited transcriptional

activities that were some 200-5000 times higher than the

“natural” counterpart;^2-4therefore, a biologically active

A number of important discoveries were made in subse-

quent studies. Most significantly, gene transcription of gemini

was demonstrated^1,6and the ligand bindingpocket was shown

to be sufficiently malleable to allow the ligand to determine

the conformation of the LBD suitable for its accommoda-

tion.⁷The ligand-dependent conformation of the LBD was

investigated, and access to a channel that can accommodate

the second side chain of gemini was discovered.^7,8Molecular

dynamics simulations, likewise, confirmed that the additional

side chain, tantamount to a 25% increase in volume, could be

domiciliatedby minor reorganizationsof some 30amino acids

and major realignments of several others without disturbing

the overall structure of the LBD.^9,10In this arrangement, one

of the gemini side chains assumes the same position as the one

of the natural ligand 1, while the other has actually two spatial

opportunities for residence.¹¹

It is further perceived that two distinct receptors, the

nuclear VDR and the membrane VDR, recognize different

shapes of the conformationally flexible 1 to elicit gene trans-

criptions and rapid signal transduction pathways, respec-

tively.¹²If the LBD is, to a certain extent, indiscriminate

toward ligands of variegated chemical makeup, the prospect

of designing specifically targeted ligands exists⁷and, with the

derivative featuring two side chains was a tenable proposi-

tion. The presence of two side chains in one molecule raised

the prospect of enhanced gene transactivation but, on the

other hand, it dimmed the ability for such a compound to

function as a ligand as it might overwhelm the space

allotment provided by the VDR^awith concomitant loss

of receptor affinity.⁵Even if binding would occur, there

was no guarantee for induction of the specific conforma-

tional change in the receptor essential for transcriptional

activity.

^†Atomic coordinates for compounds 11S-b and 2R-2 have been

deposited with the Cambridge Crystallographic Data Center

(deposition numbers 734110 and 734109, respectively). Copies of the

CIF files can be obtained, free of charge, on application to CCDC, 12

Union Road, Cambridge, CB2 1EZ, UK, via e-mail at deposit@ccdc.

cam.ac.uk.

*To whom correspondence should be addressed. Phone: 973-831-

1591. Fax: 732-445-0687. E-mail: hubert.maehr@verizon.net.

^aAbbreviations: VDR, vitamin-D receptor; LBD, ligand binding

domain; INF, interferon; MLR, mixed lymphocyte reaction; DCM,

dichloromethane; THF, tetrahydrofuran; TBAF, tetrabutylammonium

fluoride; TMS, trimethylsilyl; TBSO, t-butyl-dimethylsilyloxy; TEM-

PO, 2,2,6,6-tetramethyl-1-piperidinyloxy, free radical; PDC, pyridinium

dichromate.

r

2009 American Chemical Society

Published on Web 08/17/2009

pubs.acs.org/jmc

5506 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17

Maehr et al.

resulting conformational changes, targeted biochemical re-

sponses can be envisioned.

Scheme 1. Summary of Synthetic Targets

Complementary to the classical role of 1R,25(OH)₂D₃, a

multitude of previously unexpected physiological and bio-

chemical activities have come to light. Most prominent are the

antitumor effects based on antiproliferation, prodifferentia-

tion, and reduced angiogenesis. In view of the apparent

VDR’s promiscuity toward ligands, we incorporated into

the gemini side chains those features that were previously

established for the “one-side-chain” analogues as enhance-

ments of biological activities. These augmentations are, at

least in part, due to the prevention, or retardation, of biolo-

gical degradation, most notably the degradative cascade

initiated by 24-hydroxylation, and include replacement of

the geminal methyl groups with trifluoromethyl entities and

introduction of C23-unsaturation. This 24-hydroxylation,

occurring preferentially at the pro-R side chain of 1a,¹³entails

higher drug dosages to increase blood calcium levels and to

suppress INF-γ release in MLR.^14,15Thus, the resulting new

generation gemini maintain the side chain of 1 but feature a

different second chain. In view of the physiological precar-

iousness of vitamin D analogues, including gemini, we argued

that a gemini’s half-life could be extended by replacing the

geminal methyl groups at the calcitriol side chain with trideu-

teromethyl moieties.¹⁶Such a modification was expected to

alter, neither the propensity of this arm to assume the

customary position within the VDR nor the kinetically con-

trolled binding affinity, but contemporaneously, to prolong

the intracellular life span due to increased lipophilicity and

bond strengths. This lemma, together with the assurance of

side chain similarity to the natural hormone, led to the

expectation that gemini with deuterated methyl groups to be

superior drugs in an in vivo environment. Fortunately, this

proposition was subsequently substantiated^1,17-20and is cor-

roborated in the present study comprising 18 derivatives of 1a

as shown in Scheme 1.

Results and Discussion

Chemical Syntheses. The first part of the syntheses, illu-

strated in Schemes 2 and 3, is concerned with the preparation

of the so-called CD-ring synthones and commences with the

previously described methyl ester 5,²¹which was treated with

trideuteromethyl magnesium bromide in ether to afford

alcohol 6. An ene-reaction with paraformaldehyde and

dimethylaluminum chloride in DCM gave the pair of diols

7E and 7Z. Although this pair of epimers was readily

separated by flash chromatography, it was hydrogenated

as a mixture to furnish 8S and 8R, again easily separable by

preparative HPLC. These two intermediates are the starting

points for the entire cascade of the “S” and “R” series of the

CD-ring systems. Subsequent TEMPO/chloramine-T

mediated oxidations led to the aldehydes 9, which were

homologated to the alkynes 10.²²Conversion of the tertiary

hydroxyl group to the TMS ether, prior to deprotonation

with butyllithium and condensation with hexafluoracetone,

gave higher yields of the diols 11S-a and 11R-a when

compared to the use of the unprotected counterparts. De-

protection with fluorosilicic acid²³furnished the triols 11S-b

and 11R-b. The stereochemical assignments of the epimeric

alcohols 8a and 8b, obtained after chromatographic separa-

tion (Scheme 2), was confirmed by a crystallographic analy-

sis of 11S-b whose ORTEP rendition is depicted in Figure 1.

Subsequent oxidation with PDC in DCM gave 12S-a and

12R-a and a following silylation of the tertiary hydroxyl

groups led to the ketones 12S-b and 12R-b, respectively. The

trimethylsilyloxy groups at the gem-trifluoromethyl posi-

tions, however, are of limited stability and partially lost

upon purification of the reaction mixtures.

The intermediate triols 11S-b and 11R-b served as starting

materials for the preparation of the alkene series of CD-ring

synthones as shown in Scheme 3. Reduction with sodium

bis(2-methoxyethoxy)aluminum hydride led to the (E)-al-

kene-triols 13S and 13R, whereas catalytic hydrogenation

with Lindlar catalyst furnished the (Z)-alkene-triols 15S and

15R. The members of both pairs were oxidized with PDC to

the ketones and then further protected by silylation to lead to

the (E)-alkene-ketones 14S-b and 14R-b and the (Z)-alkene-

ketones 16S-b and 16-b, respectively. The previous sequence,

comprising oxidation at C8 and silylation of the tertiary

hydroxyl groups, converted the triol pairs 13 and 15 to the

protected ketone pairs 14S-b/14R-b and 16S-b/16R-b, which

served as suitable coupling partners with the A-ring

synthones.

The second phase of the synthesis included the coupling

reactions of the CD-ring moieties with three different A-ring

synthones as depicted in Scheme 4. The three A-rings, in the

form of the allyldiphenylphosphane oxides 17,^24-2618,^27-29

Article

Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17 5507

Scheme 2^a

Figure 1. ORTEP drawing of 11S-b.

Scheme 3^a

^aReagents and conditions: (a) CD₃MgI, ether; (b) (CH₂O)_n, Me₂AlCl;

(c) chromatography, (d) PtO₂-C, EtOAc; (e) TEMPO, chloramine-T,

DCM, buffer; (f) N₂CHP(O)(OMe)₂, MeOH, K₂CO₃; (g) TMS-imida-

zole, cyclohexane; (h) BuLi, THF, hexafluoroacetone; (i) fluorosilicic

acid, AcCN; (j) PDC, DCM.

and 19,^30-32were used in Wittig-Horner reactions with the

six CD-ring units 12S-b, 14S-b, 16S-b, and their (R)-counter-

parts, which, after deprotection with TBAF, afforded 18

combinations as final products ranging from 2S-1 to 4R-3

(Scheme 4). The crystal structure of one of them (2R-2) is

shown in Figure 2. While the crystal structure of 11S-b

(Figure 1) corroborated the configurational assignments in

the 20S series, the structure of 2R-2 authenticates the stereo-

chemical veracity of the 20R series.

Biological Studies. It became apparent that the side chain

modification, the resulting configurational duality at C20,

and the nature of the A-ring exert different biological

responses. In a recent study with nondeuterated gemini, the

^aReagents and conditions: (a) sodium bis(2-methoxyethoxy)-

aluminum hydride, toluene; (b) Lindlar catalyst, H₂, EtOH; (c) PDC,

DCM; (d) TMS-imidazole, THF.

analogue of 2S-1 was preferred and suggested gemini to

modulate the expression and activity of proteins associated

in cancer cell proliferation by inhibition of the Akt-mTOR

pathway.³³The effect, generated by introduction of deuter-

ium, was underscored when 2S-2 was compared with the

analogous, nondeuterated, epimeric pair 2S-2 and 2R-2 in

the mouse MC-26 colon cancer model.¹⁹While the nondeut-

erated analogue of 2R-2 exerted little effect, the epi-

meric nondeuterated analogue of 2S-2 caused a significant

5508 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17

Maehr et al.

Table 1. IC₅₀Values of Gemini Vitamin D Analogues in MCF10CA1a

Human Breast Cancer Cells^a

compd

IC₅₀(nM)

compd

IC₅₀(nM)

1

3.3

0.062

0.12

0.56

0.33

0.008

0.18

1.6

2S-1

2S-2

2S-3

3S-1

3S-2

3S-3

4S-1

4S-2

4S-3

2R-1

2R-2

2R-3

3R-1

3R-2

3R-3

4R-1

4R-2

4R-3

0.053

0.19

1.6

0.13

0.23

2.3

0.56

0.43

6.9

3.2

>10

^aMCF10CA1a cells were incubated with gemini at concentrations of

0.001, 0.01, 0.1, 1, 10 nM in 5% horse serum DMEM/F12 medium for 3

days. IC₅₀values were determined using TableCurve 2D software

(version 5.01, Systat).

Figure 2. ORTEP drawing of 2R-2.

We now report the effects of gemini on cell proliferation of

MCF10CA1a human breast cancer cells and on differentia-

tion of NB4 human promyelocytic leukemia cells. The

current study centers on the gemini pairs 2, 3, and 4, formal

derivatives of 1 (2S, 3S, and 4S, left column in Scheme 1) and

20-epi-1 (2R, 3,R and 4R, right column in Scheme 1), respec-

tively. They feature the two traditional six-carbon chains of

gemini (1a); the significant differences in a comparison to 1a,

however, are manifold, i.e., the inequality of the two chains,

the replacement of the geminal methyl groups of calcitriol’s

“natural chain” by deuteromethyl moieties, and the trans-

mutation of the other to incorporate geminal trifluoromethyl

groups, and 23-yne, 23(E) and 23(Z)-ene unsaturations. To

continue the theme of pharmacophoric periodicity, we also

commuted the “natural” 1-hydroxy A-ring part to 19-nor

and 1R-fluoro analogues. In the designations used, the first

letter refers to the unsaturation of the side chain, e.g., 2 for

23-yne, 3 for 23(E)-ene, and 4 for 23(Z)-ene. The symbols R

and S refer to the absolute configuration at C20 and the

extensions -1, -2, and -3 refer to the A-ring characteristics,

i.e., 1-hydroxy or “normal”, 19-nor, and 1R-fluoro arrange-

ments, respectively. As per example, 2S-2 has an alkyne side

chain, the configuration at C20 is (S), and the A-ring portion

is of the 19-nor type.

Scheme 4. Assembly of Target Compounds 2S-1 to 4R-3

reduction of colon tumor growth and tumor weight when

administered at equal concentrations. The replacement of

the hydrogen atom at the geminal methyl groups with

deuterium, as in 2S-2, enhanced the antitumor effect 10-fold.

This remarkable C20 epimeric effect was also observed in the

alteration of VDR-dependent monocytic differentiation in

human leukemia cells³⁴and in the present study of

MCF10CA1a cell proliferation, but, surprisingly, it was

largely unpronounced in the gemini series containing the

23-yne side chain. Different antitumor activities among

epimers are somewhat remarkable, as it is generally con-

tended that they assert their antiproliferative activity as

ligands inside the LBD of the VDR. If a certain epimer of

a given pair is established as a superior ligand capable to

engender an antitumor effect, one would expect this bias to

persevere in all antitumor responses. In view of the huge

difference in antiproliferative effects between 1 and epi-1, as

a result of disparate LBD conformations, different antineo-

plastic responses of two C20-epimeric geminis are not sur-

prising. The observation that the partners of such an

epimeric pair actually function differently, much like tai-

lor-made entities for very specific antiproliferative activities,

is unexpected and warrants further exploration.

Gemini-type compounds were shown to be more active in

inhibiting cell proliferation of breast-cancer cells than 1,^35,36

and in BMP/Smad-specific signaling in human breast epithe-

lial cells.³⁷In preclinical animal studies, 2S-2 and 2R-2

prevented estrogen receptor (ER)-positive mammary tumo-

rigenesis induced by N-methyl-N-nitrosourea in rats.^36,15In

addition, 2R-2 exhibited significant tumor growth suppres-

sion in the ER-negative MCF10DCIS.com xenograft model

without hypercalcemic toxicity.³⁸A mode of action study

suggested the inhibitory effect of 2R-2 to be associated with

an increase of cyclin-dependent kinase inhibitor p21 and the

insulin-like growth-factor binding-protein 3 in both animal

models. Spina et al. also reported that 2S-2 significantly

reduced tumor growth of mouse colorectal cancer cells in

BALB/c mice⁹and inhibited the invasive spread of the tumor

into the surrounding muscle.¹⁷

We first studied the inhibitory effect of gemini on

MCF10CA1a cell proliferation. The biological activities in

cell growth inhibition are compared in terms of IC₅₀values

and illustrated in Table 1. Most geminis are superior inhibi-

tors of cell growth when compared to 1, the naturally

occurring 1R,25(OH)₂D₃, and significant differences ensue

as a result of changes in A-rings, C20 configuration, and side

Article

Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17 5509

associated with the nature of the A-rings was generally upheld

in the study of induction potentials of monocyte cell surface

markers CD11b and CD14. The C20 epimeric effect was only

discernible among members containing the 23(Z) side chain,

where the 20(R) epimers were profoundly more active. Most

significantly, however, the 20S epimers were more active than

the 20R counterparts in the colon cancer models, while the

C20 epimeric effect among members containing the 23-yne

side chain was unpronounced in the present study of

MCF10CA1a cell proliferation. Guided by the notion that

this C20 epimeric effect was minimally expressed in gemini

containing the 23-yne side chain, we focused our attention on

a member of that category with the hope to select a compound

with a broader spectrum of anticancer activities.

Figure 3. Differentiation induction of NB4 leukemia cell line by

analysis of differentiated cell surface markers CD14 and CD11b.

Two independent experiments were performed and combined, and

values shown are averages ( SD. Statistical significances between 1

and the geminis were evaluated using Student’s t test (* p < 0.05, **

p < 0.01, *** p < 0.001).

Experimental Section

¹H, ¹³C, and ¹⁹F NMR spectra were recorded at on a Varian

400 MHz spectrometer. The relatively long relaxation times of

¹³C at the trideuteromethyl positions and the multiplicity due to

their spin coupling to deuterium renders these ¹³C resonances

difficult to observe. Mass spectra were recorded on Waters ZQ

(low resolution) and Bruker APEX II spectrometers (high re-

solution). Crystallographic data were obtained on a Bruker-

Nonius Kappa CCD diffractometer. TLC was performed on

silica gel GF₂₅₄plates (Merck), flash chromatography employed

40-65 μ silica gel and for preparative column chromatography

15-20 μ silica gel was used. The trimethylsilyl groups at the gem-

trifluoromethyl carbinol position in the ketones 12, 14, and 16 are

unstable and cause small losses of the TMS groups during

purification. Further losses of the TMS groups occur during the

quench of the coupling reactions with the A-rings, and after

extractive and chromatographic workup, these losses are almost

complete so that the TBAF treatments commence with partially

deprotected products. Progress of deprotection of the final con-

densation products was monitored by TLC using 1:19 and 1:9

EtOAc-hexane, 1:1 EtOAc-hexane, and EtOAc were the mo-

bile phases for the detection of the final materials. Detection was

by UV light and/or phosphomolybdic acid spray. Preparative

HPLC was performed on a 2” ꢀ 18” 15-20 μ silica YMC column

using a flow rate of 100 mL/min and RI-detection. All final

materials were of at least 95% purity; the major contaminants of

noncrystalline materials were limited to solvent traces. Elemental

analyses of the final materials are supplied in the Supporting

Information. Solutions were dried with sodium sulfate unless

specified otherwise.

chain unsaturation. Those containing the 1R-fluoro A-rings

(rows 4, 7, and 10) exerted relatively weak activities; in

general, gemini featuring an alkyne side-chain (rows 2-4)

or the 23(E) function (rows 5-7), inhibited MCF10CA1a

cell proliferation most effectively.

In view of the well-known ability of vitamin D as an

inducer of cell differentiation, we investigated the gemini’s

potential as differentiation inducers in NB4 human leukemia

cells by measuring the induction of monocyte cell-surface

markers CD11b and CD14. In general, gemini induced NB4

cell-differentiation stronger than 1. Conforming to the trend

observed in growth inhibition of breast cancer cells, the 1R-

fluoro analogues were less active in inducing cell differentia-

tion markers. Those containing the 1-hydroxy A-rings and

the 19-nor A-rings exerted similar effects. The 23(Z)-ene side

chains were generally less active than those with the 23-yne

and 23(E)-ene chains, as illustrated in Figure 3. While the

C20 epimeric effect was not perceivable among members

containing the 23-yne or 23(E)-ene side chain, the differen-

tiation was markedly enhanced in the 20R-series of those

containing the 23(Z)-ene chain (4R-1, 4R-2, and 4R-3 versus

their S-counterparts).

Conclusion

Insummary, wehave synthesized 18 closelyrelatedgeminis.

The structural features incorporated into these molecules

comprise established lead optimizations including side chain

unsaturations, geminal trifluoromethyl and trideuteromethyl

functions, A-ring variations, and dualitydueto C20 diastereo-

topicity. The resulting plexus affords an insight into the

diverse structural requirements for specific biological activi-

ties. Compounds related to 1-fluoro-1-desoxy-calcitriol were

less active than those containing the A-ring of calcitriol or 19-

nor-calcitriol based on all parameters evaluated. As judged on

the basis of IC₅₀values in MCF10CA1a human breast cancer

cells, compounds containing the 23-yne and 23(E)-ene side

chains were more potent than those featuring the Z-alkene

chains. Members of the 23-yne category showed little differ-

ences between the C20 epimers; members of the 23(E)-ene

category, however, were generally more active in the 20S

series, while those in the 23(Z)-ene category were more active

in the 20R series. This preference of the 20S configuration in

the 23(E)-enecategoryisreflected in the extraordinary activity

of 3S-2, where the E-alkene chain gives rise to a compound

that surpasses the alkyne counterpart. The activity profile

6-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-7a-

methyl-octahydro-inden-1-yl]-1,1,1-trideutero-2-trideuterome-

thyl-hept-6-en-2-ol (6). A solution of ester 5 (18.92 g, 47.94 mmol)

in ether (330 mL) was cooled in an ice-bath and a 1 M solution of

deuteromethylmagnesium iodide in ether (130 mL) was added

dropwise. Stirring was continued at ambient temperature for 2 h,

the soln was cooled again in an ice bath, and a sat. ammonium

chloride solution (100 mL) was added dropwise followed by

water (150 mL) to dissolve the white precipitate present. The

aqueous layer was re-extracted with ether (30 mL), and the

combined ether layers were washed with 1:4 sat. ammonium

chloride solution-water (100 mL) and then dried (MgSO₄),

filtered, and evaporated. The residue was flash-chromatographed

on silica gel using hexane f 1:19 EtOAc-hexane as mobile phase

to afford 6 as a colorless oil (19.2 g, 100%); TLC (1:3 EtOAc-

28

hexane) R_f0.58; [R]_Dþ22.1 (c 0.4, methanol). ¹H NMR (400

MHz, CDCl₃) δ 0.01 (br s, 3 H), 0.02 (br s, 3 H), 0.80 (s, 3 H), 0.89

(s, 9 H), 1.17 (td, J=13.5, 3.4 Hz, 1 H), 1.32-1.87 (m, 15 H),

1.97-2.09 (m, 3 H), 3.99-4.05 (m, 1 H), 4.78 (s, 1 H), 4.89 (s, 1

H). LRMS-EI(þ) m/z 400 (12, M^þ), 382 (80, M-H₂O), 382 (42,

M-H₂O-CH₃). HRMS-ES(þ) calcd for C₂₄H₄₀D₆O₂Si (M þ

H)1^þ401.3717, found 401.3713. Calcd for C₂₄H₄₀D₆O₂Si (M þ

Na)1^þ423.3536, found 423.3535.

5510 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17

Maehr et al.

(E)-3-[(1R,3aR,4S,7aR)-4-(tert-Butyldimethyl-silanyloxy)-7a-

methyl-octahydro-inden-1-yl]-8,8,8-trideutero-7-trideuterome-

thyl-oct-3-ene-1,7-diol (7E). A mixture of 6 (19.2 g, 48 mmol),

paraformaldehyde (1.72 g, 57 mmol), and DCM (160 mL) was

stirred under nitrogen for 10 min, cooled to -20 °C, and a 1 M

dimethylaluminum chloride solution in hexane (122 mL) was

added dropwise within 15 min. The mixture was then stirred in

an ice bath for 1 h to complete the reaction (TLC, 1:1 EtOAc-

hexane), cooled again to -20 °C, water (125 mL) was added

dropwise, causing the temperature to rise to 10 °C, and then

transferred to a separatory funnel with ether washes (2 ꢀ

30 mL). The milky soln was allowed to separate; the aqueous

layer was extracted with ether (3 ꢀ 100 mL), the combined ether

layers were washed with dilute pH 7 phosphate buffer (50 mL),

dried, and evaporated to an oily residue. This material was

chromatographed on a silica gel column using 1:9 f 1:4 f 1:1

EtOAc-hexane to yield 7E as a major geometric isomer that

was further purified by fractional crystallization (13.34 g). A

second chromatography of the mixture fractions and the mother

liquors gave an additional 1.67 g of crystalline product (total

yield 72.6%). The E-geometry for 7E was assigned by default

(R)-3-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-

7a-methyl-octahydro-inden-1-yl]-8,8,8-trideutero-7-trideuter-

omethyl-octane-1,7-diol (8R). The peak of the chromatograms

described above, with the elution maximum of 4.2 L, was also

evaporated to give 8R (total quantity of 6.24 g); TLC (1:1

30

EtOAc-hexane); R_f0.25; [R]_Dþ34.1 (c 0.57, MeOH). ¹H

NMR (400 MHz, CDCl₃) δ -0.01 (s, 3 H), 0.00 (s, 3 H), 0.88 (s, 9

H), 0.91 (s, 3 H), 1.13 (td, J=12.8, 3.4 Hz, 1 H), 1.20-1.62 (m, 17

H), 1.62-1.72 (m, 2 H), 1.72-1.84 (m, 2 H), 1.84-1.91 (m, 1 H),

3.56-3.71 (m, 2 H), 3.99 (br s, 1 H). LRMS-ES(þ) m/z 415 (100,

M-17)^þ. HRMS-ES(þ) calcd for C₂₅H₄₄D₆O₃Si: 455.3798 [M þ

Na]1^þ; found 455.3797.

(S)-3-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy-

7a-methyl-octahydro-inden-1-yl]-8,8,8-trideutero-7-hydroxy-

7-trideuteromethyl-octanal (9S). A mixture of 8S (1.13 g, 2.61

mmol), DCM (20 mL), TEMPO (84 mg), and tetrabutylammo-

nium chloride hydrate (146 mg), dissolved in 0.5 M potassium

hydrogen carbonate soln was stirred at ambient temperature

and chloramine-T (1.46 g) was added and vigorous stirring

continued for 1.5 h. Production of the aldehyde was followed

by TLC (1:1 EtOAc-hexane, R_f0.84). The mixture was diluted

with DCM (50 mL); the organic layer was washed with water

(25 mL), dried, and evaporated. The resulting residue was

triturated with hexane, and the suspension filtered onto a silica

gel column. Elution with 1:39 f 1:9 f 1:4 EtOAc-hexane

furnished pure 9S (1.05 g, 93.4%). ¹H NMR (400 MHz, CDCl₃)

δ 0.00 (s, 3 H), 0.01 (s, 3 H), 0.89 (s, 9 H), 0.95 (s, 3 H), 1.08-1.19

(m, 1 H), 1.19-.47 (m, 13 H), 1.51-1.61 (m, 1 H), 1.62-1.72 (m,

1 H), 1.72-1.87 (m, 3 H), 1.91-2.04 (m, 1 H), 2.42 (ddd, J=16.7,

7.7, 3.0 Hz, 1 H), 2.59 (ddd, J=16.7, 4.3, 1.8 Hz, 1 H), 3.98-4.02

(m, 1 H), 9.78 (dd, J=3.0, 1.8 Hz, 1 H). LRMS-EI(þ) m/z 355

(73, M-H₂O-C₄H₉); 354 (100, M-HOD-C₄H₉). HRMS-ES(þ)

calcd for C₂₅H₄₂D₆O₃Si: 453.3641 (M þ Na)1^þ; found

453.3649.

(R)-3-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-

7a-methyl-octahydro-inden-1-yl]-8,8,8-trideutero-7-hydroxy-

7-trideuteromethyl-octanal (9R). To a soln of diol 8R (6.24 g,

14.4 mmol) in DCM (110 mL) was added 0.5 M aqueous

potassium hydrogen carbonate soln containing tetrabutylam-

monium chloride hydrate and TEMPO (0.44 g, 2.8 mmol). The

two-phase system was stirred vigorously, and chloramine-T

(8.0 g, 28.6 mmol) was added. The conversion to the aldehyde

9R was complete after 1 h as monitored by TLC (1:1 EtOAc-

hexane, R_f0.84). The mixture was diluted with DCM (100 mL),

and the organic layer washed with water (50 mL), dried, and

evaporated. The resulting residue was triturated with hexane;

the resulting suspension was filtered onto a silica gel column and

then eluted with 1:39 and 1:4 EtOAc-hexane. The fractions

containing 9R were pooled, evaporated, and taken up in metha-

nol (55 mL) for the next step. ¹H NMR (400 MHz, CDCl₃) δ -

0.01 (s, 3 H), 0.01 (s, 3 H), 0.88 (s, 9 H), 0.95 (s, 3 H), 1.15 (td, J=

12.7, 3.1 Hz, 1 H), 1.21-1.45 (m, 12 H), 1.45-1.85 (m, 5 H), 1.90

(ddd, J=12.3, 3.3, 3.1 Hz, 1 H), 2.06 (br s, 1 H), 2.31 (ddd, J=

16.3, 7.5, 3.0 Hz, 1 H), 2.44 (ddd, J=16.3, 4.5, 1.8 Hz, 1 H),

3.97-4.02 (m, 1 H), 9.78 (dd, J=3.0, 1.8 Hz, 1 H). LRMS-EI(þ)

m/z 430 (7, M^þ), 412 (30, M-CD₃), 394 (79, M-CD₃-H₂O), 393

(95, M-CD₃-HOD). HRMS-ES(þ) calcd for C₂₅H₄₂D₆O₃Si:

431.3822 (M þ H)1^þ; found 431.2916.

1

based on the H NMR NOE difference observed for the 7Z

30

isomer; TLC (1:1 EtOAc-hexane); R_f0.27; [R]_Dþ14.4 (c 0.5,

methanol). ¹H NMR (400 MHz, CDCl₃) δ 0.01 (s, 3 H), 0.02 (s, 3

H), 0.79 (s, 3 H), 0.89 (s, 9 H), 1.12 (td, J=12.6, 3.3 Hz, 1 H),

1.31-1.49 (m, 6 H), 1.53 (t, J=8.1 Hz, 2 H), 1.58-1.84 (m, 6 H),

2.03 (t, J=9.2 Hz, 1 H), 2.11-2.28 (m, 3 H), 2.54 (dt, J=13.3, 7.5

Hz, 1 H), 3.61 (t, J=7.0 Hz, 2 H), 3.98-4.06 (m, 1 H), 5.37 (t, J=

7.0 Hz, 1 H). LRMS-EI(þ) m/z 430 (5, Mþ), 412 (62, M-H₂O),

394 (78, M-H₂O-CD₃). HRMS-ES(þ) calcd for C₂₅H₄₂D₆O₃Si

431.3822 (M

C₂₅H₄₂D₆O₃Si: C, 69.71; found C, 69.70.

þ

H); found 431.3818. Anal. calcd for

(Z)-3-[(1R,3aR,4S,7aR)-4-(tert-Butyldimethyl-silanyloxy)-7a-

methyl-octahydro-inden-1-yl]-8,8,8-trideutero-7-trideuteromethyl-

oct-3-ene-1,7-diol (7Z). The chromatographic band preceding

the major one, as described above, was rechromatographed to

give the Z-isomer (1.2 g); TLC (1:1 EtOAc-hexane); R_f0.36;

24

1

[R]_D-43.6 (c 0.3, methanol). H NMR (300 MHz, CDCl₃)

δ 0.00 (s, 3 H), 0.01 (s, 3 H), 0.86 (s, 3 H), 0.88 (s, 9 H), 1.09 (td,

J=12.6, 3.7 Hz, 1 H), 1.29-1.86 (m, 14 H), 1.98-2.43 (m, 4 H),

2.53 (t, J=9.0 Hz, 1 H), 3.40-3.53 (m, 1 H), 3.57-3.70 (m, 1 H),

3.99-4.08 (m, 1 H), 5.35 (dd, J=8.2, 5.7 Hz, 1 H). LRMS-EI(þ)

m/z 430 (9, M^þ), 412 (92, M-H₂O). ¹³C NMR (300 MHz,

CDCl₃) δ 0.01, 0.38, 21.01, 22.76, 23.17, 28.71, 28.80, 28.93,

30.96, 39.75, 43.28, 44.37, 49.00, 51.36, 56.66, 57.50, 67.81,

74.38, 75.82, 81.72, 82.14, 82.58, 136.31, 139.81. HRMS-ES(þ)

calcd for C₂₅H₄₂D₆O₃Si 453.3641(M þ Na)1^þ, found 453.3640.

Anal. calcd for C₂₅H₄₂D₆O₃Si: C, 69.71; found C, 69.64.

(S)-3-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-7a-

methyl-octahydro-inden-1-yl]-8,8,8-trideutero-7-trideuteromethyl-

octane-1,7-diol (8S, 047-094-4-5). A mixture of 7E and 7Z (16.59

g, 38.5 mmol), 5% PtO₂-C (2.2 g), and EtOAc (250 mL) was

hydrogenated at hydrostatic pressure (500 mm) overnight; the

catalyst was filtered off (celite), washed with EtOAc, and the

filtrateandwashes evaporated toa thick syrupy material(16.34 g)

then prepurified on a silica gel column (50 mm ꢀ 130 mm) using

1:4 f 1:1 EtOAc-hexane. The resulting prepurified epimeric

mixture (14.6 g) was resolved by preparative HPLC using 2:1

EtOAc-hexane as mobile phase and operating at 100 mL/min in

9 consecutive runs. The peak with the elution maximum of 3.4 L

was evaporated to give 8S as a sticky syrup (total quantity of 6.45

(S)-6-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-

7a-methyl-octahydro-inden-1-yl]-1,1,1-trideutero-2-trideuter-

omethyl-non-8-yn-2-ol (10S-a). A stirred soln of 9S (1.35 g, 3.13

mmol), methanol (20 mL), and dimethyl diazomethylphospho-

nate (0.725 g, 4.83 mmol) was cooled in an ice bath and

powdered potassium carbonate (0.705 mg, 5.1 mmol) was

added. The mixture was stirred in the ice bath for 30 min and

then at room temperature for 6 h. Reaction progress was

monitored by TLC (1:4 EtOAc-hexane, R_f0.19 f 0.42). The

mixture was equilibrated with hexane (80 mL) and water

(40 mL), the aqueous layer was re-extracted with hexane

(20 mL), and the combined extracts were washed with water

32

g). TLC (1:1 EtOAc-hexane); R_f0.28; [R]_Dþ34.8 (c 0.83,

MeOH). ¹H NMR (400 MHz, CDCl₃) δ -0.03 (s, 3 H), -0.02 (s,

3 H), 0.86 (s, 9 H), 0.89 (s, 3 H), 1.00-1.85 (m, 22 H), 1.93 (d, J=

12.1 Hz, 1 H), 3.53-3.67 (m, 2 H), 3.94-4.02 (m, 1 H). ¹³C NMR

(400 MHz, CDCl₃) δ -5.21, -4.83, 13.87, 17.62, 17.96, 19.74,

22.82, 25.75, 26.57, 31.63, 34.09, 34.39, 35.98, 40.44, 42.15, 44.21,

52.98, 53.45, 60.44, 69.32, 70.67. LRMS-EI(þ) m/z 414 (18, M-

H₂O)^þ, 396 (21, M-H₂O-CD₃). HRMS-ES(þ) calcd for

C₂₅H₄₄D₆O₃Si: 455.3798 (M þ Na)1^þ; found 455.3795.

Article

Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17 5511

(30 mL) and brine (10 mL), dried, and evaporated. The resulting

oily residue was chromatographed on a silica gel column using

hexane f 1:19 f 1:9 EtOAc-hexane to lead to 10S-a (1.14 g,

HRMS-ES(þ) calcd for C₂₉H₅₀D₆O₃Si₂: 521.4087 (M þ Na)1^þ;

found 521.4088.

(S)-6-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-

7a-methyl-octahydro-inden-1-yl]-11,11,11-trideutero-1,1,1-tri-

fluoro-10-trideuteromethyl-2-trifluoromethyl-10-trimethylsi-

lyloxy-undec-3-yn-2-ol (11S-a). The condensation of 10S-b

(1.034 g, 2.07 mmol) with hexafluoroacetone was repeated as

described below for the synthesis of 11R-a. The crude product

was purified by chromatography with hexane f 1:200 f 1:100

f 1:79 EtOAc-hexane as mobile phases to give 11S-a as a

colorless oil, 1.276 g, 92.7%. ¹H NMR (400 MHz, CDCl₃) δ -

0.007 (s, 3 H), -0.009 (s, 3 H), 0.11 (s, 9 H), 0.88 (s, 9 H), 0.91 (s,

3 H), 1.15-1.50 (m, 14 H), 1.63-1.89 (m, 4 H), 2.41 (dd, J=17.4,

5.3 Hz, 1 H), 2.49 (m, J=17.4, 4.0 Hz, 1 H), 3.49 (br s, 1 H),

3.96-4.03 (m, 1 H). ¹³C NMR (300 MHz, CDCl₃) δ -5.15, -

4.77, 2.58, 13.87, 14.14, 17.68, 18.04, 20.60, 20.89, 22.87, 25.81,

27.00, 31.62, 32.11, 34.30, 37.92, 40.18, 41.97, 44.66, 52.88,

53.09, 69.32, 70.47, 71.24, 73.95, 91.11, 121.13. LRMS-EI(þ)

m/z 664 (3, Mþ), 646 (48, M-H₂O), 607 (29, M-C₄H₉). HRMS-

ES(þ) calcd for C₃₂H₅₀D₆O₃Si₂F6: 687.3941 (M þ Na)1^þ;

found 687.3937.

29

85%) as a colorless oil; [R]_Dþ33.3° (c 0.49, methanol).

¹H NMR (300 MHz, CDCl₃) δ 0.00 (s, 3 H), 0.02 (s, 3 H),

0.89 (s, 9 H), 0.91 (br s, 3 H), 1.15-1.90 (m, 20 H), 1.93 (t, J=2.2

Hz, 1 H), 2.22-2.46 (m, 2 H), 4.00 (br s, 1 H). ¹³C NMR (300

MHz, CDCl₃) δ -5.18, -4.80, 13.83, 17.63, 17.99, 20.57, 20.78,

22.84, 25.76, 26.83, 31.83, 34.08, 34.32, 37.92, 40.13, 41.94,

44.06, 52.57, 52.85, 69.33, 69.63, 70.67, 82.88. LRMS-EI(þ)

m/z 351 (45, M-H₂O-C₄H₉); 295 (4, M-C₄H₉). HRMS-ES(þ)

calcd for C₂₆H₄₂D₆O₂Si: 449.3692 (M þ Na)1^þ; found

449.3696.

(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-7a-me-

thyl-1-((S)-6,6,6-trideutero-1-prop-2-ynyl-5-trideuteromethyl-

5-trimethylsilanyloxy-hexyl)-octahydro-indene (10S-b). To a

soln of 10S-a (1.25 g, 2.93 mmol) in cyclohexane (20 mL) was

added TMS-imidazole (2.0 mL, 13.6 mmol) and the mixture was

allowed to stand overnight. The suspension was filtered and the

filtrate chromatographed on silica gel; the column was eluted

with hexane f 1:100 f 1:79 EtOAc-hexane to furnish 10R-a;

1

1.28 g, 87.6%; TLC (1:19, EtOAc-hexane) R_f0.68. H NMR

(S)-11,11,11-Trideutero-1,1,1-trifluoro-6-((1R,3aR,4S,7aR)-

4-hydroxy-7a-methyl-octahydro-inden-1-yl]-10-trideuterome-

thyl-2-trifluoromethyl-undec-3-yne-2,10-diol (11S-b). A stirred

soln of 11S-a (0.406 g, 0.61 mmol) in acetonitrile (6 mL) was

cooled in an ice bath and 25% aqueous fluorosilicic acid was

added. Stirring was continued at ambient temperature for 4 h.

The mixture was equilibrated with EtOAc (40 mL) and water

(15 mL), the organic layer was washed with water (2 ꢀ 15 m) and

2:1 brine-satd sodium hydrogen carbonate soln (10 mL) and

then dried and evaporated. The residue was chromatographed

using 1:4 f 1:2 EtOAc-hexane as mobile phases to afford 11S-b

as a syrup that crystallized when treated with DCM-hexane.

This material was recrystallized from the same solvent; TLC

(1:19 methanol-DCM); R_f0.25; (1:1 EtOAc-hexane); R_f0.68;

(300 MHz, CDCl₃) δ 0.00 (s, 3 H), 0.01 (s, 3 H), 0.11 (s, 9 H), 0.89

(s, 9 H), 0.91 (s, 3 H), 1.11-1.95 (m, 20 H), 2.28-2.41 (m, 2 H),

4.00 (br s, 1 H). ¹³C NMR (300 MHz, CDCl₃) δ -5.14, -4.76,

2.65, 13.89, 17.69, 18.04, 20.58, 20.86, 22.91, 25.82, 26.86, 31.84,

34.39, 37.95, 40.17, 42.00, 44.87, 52.68, 52.91, 69.42, 69.54,

73.71, 83.02. LRMS-EI(þ) m/z 367 (22, M-OTBS), 351 (28,

M-C₄H₉-TMSOH). HRMS-EI(þ) calcd for C₂₉H₅₀D₆O₃Si₂:

498.4189 (M^þ); found 498.4203. Calcd for C₂₉H₅₀D₆O₃Si₂:

480.3772 (M-CD₃)^þ; found 480.3788.

(R)-6-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy-

7a-methyl-octahydro-inden-1-yl]-1,1,1-trideutero-2-trideuter-

omethyl-non-8-yn-2-ol (10R-a). The stirred methanolic soln of

9R obtained as described above was cooled in an ice bath and

dimethyl diazomethylphosphonate (3.24 g, 21.6 mmol) was

added, followed by powdered potassium carbonate (3.0 g,

21.6 mmol). The cooling bath was removed after 1 h and

stirring continued for an additional 5 h. Reaction progress

was monitored by TLC (1:1 EtOAc-hexane, R_f0.72 f 0.86).

The mixture was equilibrated with hexane (300 mL) and water

(100 mL), the aqueous layer re-extracted with hexane (2 ꢀ

50 mL), and all extracts were evaporated. For analysis, a

small portion was dissolved in hexane and chromatographed

using hexane f 1:19 f 1: 9 EtOAc-hexane to furnish pure

10R-a. ¹H NMR (400 MHz, CDCl₃) δ 0.00 (s, 3 H), 0.01 (s, 3

H), 0.89 (s, 9 H), 0.92 (s, 3 H), 1.13-1.77 (m, 17 H), 1.77-1.95

(m, 4 H), 2.10-2.18 (m, 1 H), 2.30-2.37 (m, 1 H), 3.95-4.06

(m, 1 H). LRMS-EI(þ) m/z 408 (100, M þ H-H₂O). HRMS-

ES(þ) calcd for C₂₆H₄₂D₆O₂Si: 449.3692 (M þ Na)1^þ; found

449.3692. The material was used without further purification

in the next step.

(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-7a-me-

thyl-1-((R)-6,6,6-trideutero-1-prop-2-ynyl-5-trideuteromethyl-5-

trimethylsilanyloxy-hexyl)-octahydro-indene (10R-b). The bulk

of 10R-a, together with the purified material described above,

was dissolved in cyclohexane (80 mL) and TMS-imidazole

(8 mL). After 4 h, the suspension was filtered onto a silica gel

column, together with hexane washes, and the column was

eluted with hexane f 1:79 EtOAc-hexane to give 10R-b as a

colorless oil (5.41 g, 75.3% from diol 8R). ¹H NMR (400 MHz,

CDCl₃) δ -0.01 (s, 3 H), 0.00 (s, 3 H), 0.10 (s, 9 H), 0.88 (s, 9 H),

0.91 (s, 3 H), 1.12-1.45 (m, 11 H), 1.46-1.62 (m, 4 H), 1.62-

1.72 (m, 1 H), 1.72-1.86 (m, 2 H), 1.88 (t, J=2.5 Hz, 1 H), 1.92

(dt, J=12.3, 2.8 Hz, 1 H), 2.12 (ddd, J=16.9, 5.3, 2.6 Hz, 1 H),

2.32 (dt, J=16.9, 3.0 Hz, 1 H), 3.93-4.03 (m, 1 H). ¹³C NMR

(300 MHz, CDCl₃) δ -5.10, -4.73, 2.65, 14.05, 17.75, 18.07,

20.74, 22.95, 25.86, 27.04, 31.52, 34.50, 38.63, 40.63, 42.11,

44.96, 53.03, 53.12, 68.93, 69.52, 73.72, 83.02. LRMS-ES(þ)

m/z 409 (35, M-TMSO), 295 (100, M-TMSO-TBS þ H).

25

1

mp 132-3 °C; [R]_Dþ15.6 (c 0.72, MeOH). H NMR (400

MHz, CDCl₃) δ 0.94 (s, 3 H), 1.09-1.55 (m, 16 H), 1.60-1.71

(m, 1 H), 1.74-1.96 (m, 4 H), 2.32 (dd, J=17.4, 7.0 Hz, 1 H), 2.55

(dd, J = 17.4, 4.3 Hz, 1 H), 4.08 (br s, 1 H), 5.60 (s, 1 H).

¹³C NMR (400 MHz, CDCl₃) δ 13.60, 17.31, 20.07, 21.42, 22.26,

26.95, 32.15, 33.17, 38.46, 40.06, 41.77, 43.15, 52.45, 53.06,

69.28, 69.50, 70.61, 71.03, 71.91, 89.95, 121.41. LRMS-ES(þ)

m/z 478 (15, Mþ), 477 (100 M-H). HRMS-ES(þ) calcd for

C₂₃H₂₈D₆O₃F6: 501.2681 (M þ Na)1^þ; found 501.2680. Anal.

calcd for C₂₃H₂₈D₆F₆O₃: C, 57.73; found 57.87. Roentgen

diffraction analysis (Figure 1): C₂₃H₃₄F₆O₃, orthorhombic,

space group P2₁2₁2₁with unit cell dimensions of a =

˚

6.8335(14), b = 18.386(4), c = 18.871(4) A, volume 2371.0(8)

3

A (Z=4).

˚

(R)-6-[(1R,3aR,4S,7aR)-4-(tert-Butyl-dimethyl-silanyloxy)-

7a-methyl-octahydro-inden-1-yl]-11,11,11-trideutero-1,1,1-tri-

fluoro-10-trideuteromethyl-2-trifluoromethyl-10-trimethylsilylo-

xy-undec-3-yn-2-ol (11R-a). A three-neck flask equipped with a

magnetic stirrer, an addition funnel in the center surrounded by

a dry ice bath and topped by a dry ice condenser with a

hexafluoroacetone inlet, and a Claisen adapter with nitrogen

sweep connected to the exhaust slot of the hood and a rubber

septum, was charged with 10R-b (5.41 g, 10.8 mmol) and THF

(31.13 g). The soln was immersed into a dry ice/acetone bath and

a 1.6 M butyllithium soln in hexane (9 mL) was added dropwise

over an 8 min period. Immediately after the addition, the dry ice

condenser and the dry ice bath surrounding the addition funnel

were filled dry ice and, after stirring for 5 min, hexafluoroace-

tone was condensed in. The condensate, ca. 1 mL, was then

added to the reactor. The starting material was no longer

detectable after 1.5 h (TLC, 1:19 EtOAc-hexane). The system

was flushed with nitrogen and then pH 7 phosphate buffer

(20 mL) was added, the stirred mixture allowed to reach

room temperature, further diluted with hexane (50 mL), and

transferred to a separatory funnel with a hexane rinse (25 mL).

5512 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17

Maehr et al.

The organic layer was washed with brine (15 mL), dried, and

evaporated. From this oily residue, most of a self-condensation

product derived from hexafluoroacetone could be removed by

high-vacuum distillation at a bath temperature of 30-40 °C.

For analysis, a small aliquot was purified on a silica gel column

using hexane f 1:200 f 1:100 f 1: 79 EtOAc-hexane to

furnish pure 11R-a, TLC (cyclohexane, R_f0.14). ¹H NMR

(400 MHz, CDCl₃) δ -0.01 (s, 3 H), 0.00 (s, 3 H), 0.10 (s, 6

H), 0.88 (s, 9 H), 0.92 (s, 3 H), 1.13 (td, J=12.9, 3.0 Hz, 1 H),

1.18-1.46 (m, 15 H), 1.48-1.72 (m, 8 H), 1.72-1.87 (m, 3 H),

1.91 (d, J=12.1 Hz, 1 H), 2.22 (dd, J=17.4, 6.0 Hz, 1 H), 2.42

(dd, J=17.4, 3.7 Hz, 1 H), 3.33 (br s, 1 H), 3.96-4.03 (m, 1 H).

LRMS (ES(-) m/z 628 (38, M-H₂O-CD₃). HRMS-ES(þ) calcd

for C₃₂H₅₀D₆F₆O₃Si₂: 687.3941 (M þ Na)1^þ; found 687.3928.

(R)-11,11,11-Trideutero-1,1,1-trifluoro-6-((1R,3aR,4S,7aR)-

4-hydroxy-7a-methyl-octahydro-inden-1-yl]-10-trideuterometh-

yl-2-trifluoromethyl-undec-3-yne-2,10-diol (11R-b). The bulk of

the crude 11R-a, together with the purified material described

above, was dissolved in acetonitrile (100 mL), cooled in an ice

bath, and a 25% aqueous fluorosilicic acid solution (24 mL) was

added. The mixture was stirred in the ice bath for 5 min and then

at room temperature for 5 h. The mixture was equilibrated with

EtOAc (200 mL) and water (50 mL), the organic phase was

washed with water (2 ꢀ 50 mL) and then 2:1 brine-sat. sodium

hydrogen carbonate (50 mL), dried, and evaporated. This

material was charged to a silica gel column and eluted with

1:4 f 1:2 EtOAc-hexane to yield 11R-b as a colorless foam, 5.1

g, 98.6% from the acetylene 10R-b; TLC (1:1, EtOAc-hexane,

EtOAc-hexane gave the corresponding mono-TMS ether

1

(0.023 g, 9%). H NMR (300 MHz, CDCl₃) δ 0.12 (s, 9 H),

0.65 (s, 3 H), 1.58 (s, 14 H), 2.12-2.61 (m, 7 H), 3.57 (br s, 1 H).

(1R,3aR,7aR)-7a-Methyl-1-[(R)-6,6,6-trifluoro-5-hydroxy-

1-(5,5,5-trideutero-4-hydroxy-4-trideuteromethyl-pentyl)-5-tri-

fluoromethyl-hex-3-ynyl]-octahydro-inden-4-one (12R-a). To a

stirred soln of 11R-b (0.330 g, 0.69 mmol) in DCM (9 g) was

added, in succession, celite (06 g) and PDC (1.16 g, 3 mmol). The

mixture was stirred for 14 h, diluted with ether (15 mL), stirred

for 5 min, and applied to a silica gel G column. Elution was

completed with 1:9 EtOAc-DCM, the effluent containing the

ketone was evaporated, the residue taken up in DCM, diluted

with hexane, and allowed to crystallize, 0.24 g (73%); mp 156-

7 °C; TLC (1:4, 2-PrOH-cyclohexane) R_f0.80; mp 156-7 °C;

[R]_D-15.3 (c 0.43, MeOH). ¹H NMR (400 MHz, CDCl₃)

23

δ 0.65 (s, 3 H), 1.24-1.82 (m, 13 H), 1.82-1.98 (m, 2 H), 1.98-

2.11 (m, 2 H), 2.17-2.35 (m, 3 H), 2.44-2.52 (m, 2 H), 5.81

(br s, 1 H). ¹³C NMR (400 MHz, CDCl₃) δ 12.71, 19.02,

20.06, 21.95, 24.02, 27.44, 32.91, 38.65, 38.81, 40.89, 42.67,

49.74, 53.53, 61.85, 70.79, 70.87, 71.26, 72.17, 89.99, 121.20,

211.47. ¹⁹F NMR (376.31 MHz, CDCl₃) δ -78.35 (br m).

LRMS(ES)(-) m/z 475 (100, M - H); HRMS-ES(þ) calcd for

C₂₃H₂₆D₆O₃F₆499.2524 (M þ Na)1^þ; found 499.2520. Anal.

calcd for C₂₃H₂₆D₆O₃F₆: C, 57.97; found C, 57.77.

(1R,3aR,7aR)-7a-Methyl-1-[(R)-6,6,6-trifluoro-1-(5,5,5-tri-

deutero-4-trideuteromethyl-4-trimethylsilanyloxy-pentyl)-5-tri-

fluoromethyl-5-trimethylsilanyloxy-hex-3-ynyl]-octahydro-in-

den-4-one (12R-b). To a suspension of 12R-a (0.22 g, 0.46 mmol)

in cyclohexane (3 mL) was added TMS-imidazole (0.30 mL,

2 mmol) to form a solution that was stored overnight and then

filtered onto a silica gel column. Elution with hexane f 1:79 f

1:39 f 1:19 EtOAc-hexane gave 12R-b as a colorless oil, 0.26 g

(91%); TLC (1:39 EtOAc-hexane) R_f0.11. ¹H NMR (400

MHz, CDCl₃) δ 0.11 (s, 9 H), 0.29 (s, 9 H), 0.67 (s, 3 H),

1.17-1.50 (m, 6 H), 1.57-1.68 (m, 4 H), 1.71-1.84 (m, 2 H),

1.84-1.99 (m, 2 H), 1.99-2.14 (m, 2 H), 2.20-2.36 (m, 3 H),

2.43-2.53 (m, 2 H). ¹⁹F NMR (376.31 MHz, CDCl₃) δ -8

2.41(s). LRMS(ES)(þ) m/z 531 (100, M-OTMS). HRMS-EI(þ)

calcd for C₂₉H₄₂D₆O₃Si₂F₆: 643.3315 (M þ Na)1^þ; found

643.3311.

26

R_f0.43); [R]_Dþ6.1 (c 0.70, MeOH). ¹H NMR (400 MHz,

CDCl₃) δ 0.92 (s, 3 H), 1.09-1.70 (m, 13 H), 1.70-1.89 (m, 3 H),

1.93 (d, J=12.7 Hz, 1 H), 2.18 (dd, J=17.2, 6.3 Hz, 1 H), 2.41

(dd, J=17.2, 3.3 Hz, 1 H), 3.38 (br s, 3 H), 4.08 (br s, 1 H). ¹³C

NMR (400 MHz, CDCl₃) δ 13.60, 17.31, 20.07, 21.42, 22.26,

26.95, 32.15, 33.17, 38.46, 40.06, 41.77, 43.15, 52.45, 53.06,

69.28, 69.50, 70.61, 71.03, 71.91, 89.95, 121.41. ¹⁹F NMR

(376.31 MHz, CDCl₃) δ -78.12 (br s). HRMS-EI(þ) calcd for

C₂₃H₂₈D₆O₃F₆: 501.2681 (M þ Na)1^þ; found 501.2679.

(1R,3aR,7aR)-7a-Methyl-1-[(S)-6,6,6-trifluoro-5-hydroxy-1-

(5,5,5-trideutero-4-hydroxy-4-trideuteromethyl-pentyl)-5-trifluo-

romethyl-hex-3ynyl]-octahydro-inden-4-one (12S-a). To a stirred

soln of 11S-b (0.239 mg, 0.500 mmol) in DCM (6.5 g) was added,

in succession, celite (0.43 g) and PDC (0.83 g, 2.20 mmol). The

suspension was diluted with ether (15 mL) after 7 h, stirred for

5 min, and applied to a silica gel plug. The eluate and ether

washes were combined and evaporated to a colorless oil, which

was allowed to crystallize. Recrystallization from DCM-hex-

ane gave pure 12S-a, 0.23 g, 96.5%; mp 176-177 °C; [R]_D²⁶-7.7

(S)-11,11,11-Trideutereo-1,1,1-trifluoro-6-((1R,3aR,4S,7aR)-

4-hydroxy-7a-methyl-octahydro-indene-1-yl)-10-trideuterome-

thyl-2-trifluoromethyl-undec-3(E)-ene-2,10 diol (13S). A three-

neck flask equipped with a magnetic stirrer, an addition funnel,

and a Claisen adapter with nitrogen sweep and a rubber septum

was charged with a 3.5 M soln of sodium bis(2-methoxyethoxy)-

aluminum hydride in toluene and THF (50 mL) and cooled to

-30 °C. To this soln was added a soln of triol 11S-b (1.1 g,

2.3 mmol) in THF (45 mL) dropwise below -20 °C and the

mixture was allowed to warm to 0 °C within 25 min and then

maintained at that temperature for 3 h. The mixture was diluted

with ether (50 mL), refrigerated overnight, filtered through

celite, and the filtrate was evaporated. The residue was chroma-

tographed on a silica gel column with 1:4 f 1:3 f 1:2 EtOAc-

hexane. The pooled product fractions were evaporated and the

residue allowed to crystallize from DCM-hexane to afford 13S,

1.03 g (93%); TLC (1:9 methanol-DCM) R_f0.69; mp 141 °C;

1

(c 0.44, MeOH). H NMR (300 MHz, CDCl₃) δ 0.64 (s, 3 H),

1.58 (s, 13 H), 1.83-2.11 (m, 4 H), 2.15-2.33 (m, 2 H), 2.39 (dd,

J=17.6, 5.9 Hz, 1 H), 2.48 (dd, J=11.4, 7.7 Hz, 1 H), 2.57 (dd,

J=17.6, 3.7 Hz, 1 H), 5.67 (br s, 1 H). ¹⁹F NMR (376.31 MHz,

CDCl₃) δ -78.28 br s. LRMS(EI)(þ) m/z 476 (7, Mþ). HRMS-

EI calcd for C₂₃H₂₆D₆O₃F₆: 499.2524 (M þ Na)1^þ; found

499.2527. Anal. calcd for C₂₃H₂₆D₆O₃F₆: C, 57.97; found C,

57.92.

(1R,3aR,7aR)-7a-Methyl-1-[(S)-6,6,6-trifluoro-1-(5,5,5-tri-

deutero-4-trideuteromethyl-4-trimethylsilanyloxy-pentyl)-5-tri-

fluoromethyl-5-trimethylsilanyloxy-hex-3-ynyl]-octahydro-in-

den-4-one (12S-b). To a suspension of the ketone 12S-a (0.22 g)

in ether (3 mL) was added TMS-imidazole (0.3 mL), whereupon

a soln was obtained. TLC (1:19 methanol-DCM and 1:39

EtOAc-hexane) confirmed the absence of educt and the con-

version to 12S-b after 1.5 h. The resulting suspension was

filtered onto a flash column, which was then eluted with hexane

f 1:39 EtOAc-hexane eluting 12S-b (0.229 g, 80%). ¹H NMR

(300 MHz, CDCl₃) δ 0.11 (s, 9 H), 0.29 (s, 9 H), 0.65 (s, 3 H),

1.05-1.48 (m, 8 H), 1.63-2.14 (m, 8 H), 2.14-2.40 (m,

2 H), 2.51 (br s, 3 H). LRMS(EI)(þ) m/z 602 (68, M-CD₃).

HRMS-EI(þ) calcd for C₂₉H₄₂D₆O₃Si₂F₆: 643.3315 (M þ

Na)1^þ; found 643.3333. Continued elution with 1:19 f 1:4

30

[R]_Dþ10.55° (c 0.44, methanol). ¹H NMR (400 MHz, CDCl₃)

δ 0.96 (s, 3 H), 1.10-1.64 (m, 16 H), 1.75-1.89 (m, 3 H), 1.94 (d,

J=12.8 Hz, 1 H), 2.07-2.18 (m, 1 H), 2.48 (dt, J=14.3, 4.7 Hz,

1 H), 4.03-4.10 (m, 1 H), 5.58 (d, J=15.6 Hz, 1 H), 6.28 (ddd, J=

15.6, 7.6, 7.5 Hz, 1 H). ¹³C NMR (400 MHz, CDCl₃) δ 13.66,

17.40, 19.77, 22.36, 26.85, 31.91, 33.51, 35.23, 38.91, 40.32,

41.89, 43.34, 52.53, 53.52, 69.29, 71.34, 76.13, 118.92, 122.29,

4

138.59. ¹⁹F NMR (376.31 MHz) δ -77.97, 3F, q, J_F,F=8.78

Hz, -77.80, 3F, q, ⁴J_F,F=9.40 Hz. LRMS-ES(-) m/z 480 (20,

M) 479 (39, M - H). HRMS-ES(þ) found 503.2838 (M þ

Na)1^þ, calcd for C₂₃H₃₀D₆O₃F₆: 503.2837. Anal. calcd for

C₂₃H₃₀D₆O₃F₆: C, 57.48; found 57.38.

Article

(R)-11,11,11-Trideutereo-1,1,1-trifluoro-6-((1R,3aR,4S,7a-

Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17 5513

14R-b. ¹H NMR (400 MHz, CDCl₃) δ 0.10 (s, 9 H), 0.22 (s, 9 H),

0.66 (s, 3 H), 1.10-1.46 (m, 7 H), 1.46-2.12 (m, 9 H), 2.17-2.43

(m, 4 H), 2.45 (dd, J=11.6, 7.6 Hz, 1 H), 5.55 (d, J=15.6 Hz,

1 H), 6.13 (m, 1 H). ¹⁹F NMR (376.31 MHz) δ -76.64, 3F, q,

⁴J_F,F=9.41 Hz, -76.48, 3F, q, ⁴J_F,F=9.41 Hz.

R)-4-hydroxy-7a-methyl-octahydro-indene-1-yl)-10-trideutero-

methyl-2-trifluoromethyl-undec-3(E)-ene-2,10 diol (13R). The

triol 13R was prepared as described for the isomer 13S and

28

was obtained as a white powder (88.6% yield); [R]_Dþ17.3° (c

1

(S)-11,11,11-Trideutereo-1,1,1-trifluoro-6-((1R,3aR,4S,7aR)-

4-hydroxy-7a-methyl-octahydro-indene-1-yl)-10-trideuterome-

thyl-2-trifluoromethyl-undec-3(Z)-ene-2,10 diol (15S). A suspen-

sion comprising the triol 11S-b (1.00 g, 2.09 mmol), hexane

(25 mL), EtOAc (10 mL), and 5% Pd-CaCO₃containing 3.5%

Pb (0.3 g, Degussa) was stirred under hydrogen, near hydro-

static balance, for 30 min, and then filtered. The suspension was

filtered, and the filtrate was evaporated to a white foam (1.03 g)

and further purified by chromatography on a silica gel using 1:4

f 1:3 f 1:2 EtOAc-hexane to afford 15S in the form of white

solids after evaporation of a soln in methyl formate (0.99 g,

0.22, methanol). H NMR (400 MHz, CDCl₃) δ 0.95 (s, 3 H),

1.13-1.66 (m, 17 H), 1.75-1.91 (m, 3 H), 1.90-2.09 (m, 2 H),

2.37 (dt, J=14.7, 5.1 Hz, 1 H), 3.79 (br s, 1 H), 4.05-4.10 (m, 1

H), 5.60 (d, J=15.6 Hz, 1 H), 6.29 (ddd, J=15.3, 8.8, 6.3 Hz, 1

H). ¹³C NMR (400 MHz, CDCl₃) δ 13.78, 17.55, 19.82, 22.45,

27.11, 31.65, 33.54, 35.06, 39.24, 40.23, 42.03, 43.35, 52.58,

53.31, 69.35, 71.35, 76.08, 118.87, 122.23, 138.43. ¹⁹F NMR

(376.31 MHz) δ -78.08, 3F, q, ⁴J_F,F=9.53 Hz, -77.87, 3F, q,

⁴J_F,F=9.28 Hz. LRMS-ES(þ) m/z 463 (100, M-OH), 445 (25,

M-OH-H₂O). HRMS-ES(þ) found 503.2835 (M þ Na)1^þ,

calcd for C₂₃H₃₀D₆O₃F₆: 503.2837. Anal. calcd for

C₂₃H₃₀D₆O₃F₆: C, 57.48; found 57.38.

29

98.5%); TLC (1:1, EtOAc-hexane); R_f0.75; [R]_Dþ6.70° (c

1

0.49, methanol). H NMR (400 MHz, CDCl₃) δ 0.93 (s, 3 H),

(1R,3aR,7aR)-7a-Methyl-1-[(S)-6,6,6-trifluoro-5-hydroxy-

1-(5,5,5-trideutero-4-hydroxy-4-trideuteromethyl-pentyl)-5-tri-

fluoromethyl-hex-3-enyl]-octahydro-inden-4-one (14S-a). The

triol 13S was oxidized as described for the preparation of the

1.16 (td, J=13.4, 3.4 Hz, 1 H), 1.21-1.63 (m, 15 H), 1.69-1.96

(m, 4 H), 2.52-2.70 (m, 2 H), 4.02-4.10 (m, 1 H), 5.42 (d, J=

12.4 Hz, 1 H), 5.95 (ddd, J=12.4, 7.4, 7.2 Hz, 1 H). ¹³C NMR

(400 MHz, CDCl₃) δ 13.59, 17.39, 18.34, 22.30, 26.51, 29.71,

31.38, 33.36, 39.14, 40.07, 43.50, 52.42, 52.52, 69.42, 71.71,

116.99, 122.90, 141.95. ¹⁹F NMR (376.31 MHz) δ -77.76, 3F,

q, ⁴J_F,F=9.78 Hz, -78.25, 3F, q, ⁴J_F,F=9.78 Hz. LRMS-ES(-)

m/z 480 (20, M) 479 (100, M - H). HRMS-ES(þ) calcd for

C₂₃H₃₀D₆O₃F₆: 503.2837 (M þ Na)1^þ; found 503.2837.

alkyne analogue 12S-a to obtain crystalline 14S-a from DCM-

31

hexane (96% yield) TLC (1:9 MeOH-DCM); R_f0.75; [R]_D

-

9.5° (c 0.45, methanol). ¹H NMR (400 MHz, CDCl₃) δ 0.64 (s, 3

H), 1.11-1.44 (m, 7 H), 1.45-1.96 (m, 8 H), 1.97-2.08 (m, 2 H),

2.12-2.33 (m, 3 H), 2.45 (dd, J=11.4, 7.6 Hz, 2 H), 5.06 (br s, 1 H),

5.59 (d, J=15.4 Hz, 1 H), 6.30 (ddd, J=15.4, 7.6, 7.5 Hz, 1 H). ¹³

C

(R)-11,11,11-Trideutereo-1,1,1-trifluoro-6-((1R,3aR,4S,7a-

R)-4-hydroxy-7a-methyl-octahydro-indene-1-yl)-10-trideutero-

methyl-2-trifluoromethyl-undec-3(Z)-ene-2,10 diol (15R). The

hydrogenation with Lindlar catalyst was repeated with 11R-b

NMR (400 MHz, CDCl₃) δ 12.56, 18.86, 19.69, 23.96, 26.94, 31.64,

34.94, 38.73, 38.97, 40.76, 43.47, 49.88, 53.37, 61.80, 71.33, 75.83,

119.50, 122.47, 137.71, 212.42. ¹⁹F NMR (376.31 MHz) δ -77.83,

3F, q, ⁴J_F,F=8.27 Hz, -77.73, 3F, q, ⁴J_F,F=7.53 Hz. LRMS-ES(-)

m/z 523 (100, M þ HCOO^-), 477 (30, M - H). HRMS-ES(þ)

calcd for C₂₃H₂₈D₆O₃F₆: 501.2681 (M þ Na)1^þ, found 501.2685.

Anal. calcd for C₂₃H₂₈D₆O₃F₆: C, 57.13; found C, 57.08.

(1R,3aR,7aR)-7a-Methyl-1-[(S)-6,6,6-trifluoro-1-(5,5,5-tri-

deutero-4-trideuteromethyl-4-trimethylsilanyloxy-pentyl)-5-tri-

fluoromethyl-5-trimethylsilanyloxy-hex-3(E)-enyl]-octahydro-

inden-4-one (14S-b). The ketone-diol 14S-a was silylated as

described for the preparation of the disilyl ether 12R-b to give

14S-b. ¹H NMR (400 MHz, CDCl₃) δ 0.09 (s, 9 H), 0.22 (s, 9 H),

0.67 (s, 3 H), 1.08-1.46 (m, 7 H), 1.46-2.10 (m, 9 H), 2.16-2.43

(m, 4 H), 2.46 (dd, J=11.3, 7.7 Hz, 1 H), 5.56 (d, J=15.4 Hz, 1

H), 6.15 (dt, J=15.4, 7.3 Hz, 1 H).

29

as described above to yield 15R as white solids; [R]_Dþ16.9 ° (c

1

0.34, methanol). H NMR (400 MHz, CDCl₃) δ 0.93 (s, 3 H),

1.10-1.25 (m, 1 H), 1.26-1.64 (m, 16 H), 1.72-1.89 (m, 3 H),

1.93(d, J=13.0 Hz, 1 H), 2.39-2.50 (m, 1 H), 2.51-2.63(m, 1 H),

4.07 (br s, 1 H), 5.36-5.44 (m, 2 H), 5.96 (ddd, J=12.1, 7.9, 6.4

Hz, 1 H). ¹³C NMR (400 MHz, CDCl₃) δ 13.71, 17.55, 18.60,

22.40, 26.96, 29.89, 31.28, 33.42, 39.62, 40.04, 43.52, 52.39, 52.58,

69.43, 71.65, 116.92, 122.87, 141.90. ¹⁹F NMR (376.31 MHz) δ -

4

77.82, 3F, q, J_F,F=9.91 Hz, -78.23, 3F, q, J_F,F=9.90 Hz.

LRMS-ES(-) m/z 480 (20, M) 479 (100, M - H). HRMS-ES(þ)

calcd for C₂₃H₃₀D₆O₃F₆: 503.2837 (M þ Na)1^þ; found 503.2834.

(1R,3aR,7aR)-7a-Methyl-1-[(S)-6,6,6-trifluoro-5-hydroxy-1-

(5,5,5-trideutero-4-hydroxy-4-trideuteromethyl-pentyl)-5-triflu-

oromethyl-hex-3(Z)-enyl]-octahydro-inden-4-one (16S-a). A so-

lution of triol 15S (0.957 g, 2 mmol) in DCM (27.3 g) was stirred

and a mixture of PDC (2.65 g, 7 mmol) and celite (1.64 g) was

added. The oxidation was complete after 7 h as judged by TLC

(1:9 MeOH-DCM, R_f0.68 f 0.75). The suspension was diluted

with ether (30 mL) and filtered through silica gel G. Filtrate and

ether washed were combined and evaporated to an oil, which

was flash-chromatographed using 1:6 f 1:4 f 1:3 EtOAc-

hexane as mobile phases to afford 16S-a as a white solid, 0.74 g

(77.3%); TLC (1:19 MeOH-DCM, R_f0.48; 1:4 EtOAc-hex-

(1R,3aR,7aR)-7a-Methyl-1-[(R)-6,6,6-trifluoro-5-hydroxy-

1-(5,5,5-trideutero-4-hydroxy-4-trideuteromethyl-pentyl)-5-tri-

fluoromethyl-hex-3(E)-enyl]-octahydro-inden-4-one (14R-a). To

a stirred suspension of triol 13R (1.68 g), DCM (26 g) and celite

(1.6 g) was added PDC (2.60 g, 6.64 mmol) and the mixture

stirred for 5.5 h and then diluted with ether (30 mL) and applied

to a silica gel-G plug. The column was washed exhaustively with

1:9 EtOAc-ether; the fractions containing the ketone were

evaporated and the residue crystallized from DCM-hexane;

31

TLC (1:9 methanol-DCM); R_f0.75, [R]_D-4.1° (c 0.32,

methanol). ¹H NMR (400 MHz, CDCl₃) δ 0.65 (s, 3 H), 1.22-

1.69 (m, 12 H), 1.69-1.81 (m, 1 H), 1.81-1.97 (m, 2 H), 1.97-

2.12 (m, 3 H), 2.16-2.42 (m, 3 H), 2.46 (dd, J=11.5, 7.7 Hz, 1 H),

4.27 (br s, 1 H), 5.61 (d, J=15.4 Hz, 1 H), 6.29 (ddd, J=15.4, 8.7,

6.2 Hz, 1 H). ¹³C NMR (400 MHz, CDCl₃) δ 12.71, 19.00, 19.74,

24.08, 27.34, 31.75, 34.91, 38.77, 39.37, 40.90, 43.30, 49.93,

53.35, 61.86, 71.24, 75.80, 119.19, 122.24, 137.92, 211.76. ¹⁹F

NMR (376.31 MHz) δ -77.84, 3F, q, ⁴J_F,F=8.91 Hz, -77.99,

34

1

ane, R_f0.07); [R]_Dþ13.2° (c 0.38, methanol). H NMR (400

MHz, CDCl₃) δ 0.63 (s, 3 H), 1.16-1.45 (m, 7 H), 1.45-1.79 (m,

6 H), 1.78-1.94 (m, 2 H), 1.94-2.08 (m, 2 H), 2.14-2.33 (m, 2

H), 2.45 (dd, J=11.4, 7.6 Hz, 1 H), 2.65 (t, J=6.4 Hz, 2 H), 5.46

(d, J=12.2 Hz, 1 H), 5.54 (br s, 1 H), 5.96 (dt, J=12.2, 7.2 Hz,

1 H). ¹³C NMR (400 MHz, CDCl₃) δ 12.50, 18.39, 18.86, 23.99,

26.70, 29.88, 31.35, 38.63, 39.35, 40.82, 43.39, 49.83, 52.63,

61.84, 71.63, 77.07, 117.49, 122.93, 141.40, 212.30. ¹⁹F NMR

(376.31 MHz) δ -77.73, 3F, q, ⁴J_F,F=9.91 Hz, -78.22, 3F, q,

⁴J_F,F=9.91 Hz. LRMS-ES(-) m/z 478 (15, M) 477 (90, M - H).

HRMS-ES(þ) calcd for C₂₃H₂₈D₆O₃F₆: 479.2862 (M þ H)1^þ;

found 479.2863.

4

3F, q, J_F,F=11.92 Hz. LRMS-ES(þ) m/z 479 (100, M þ H).

HRMS-ES(þ) calcd for C₂₃H₂₈D₆O₃F₆: 479.2862 (M þ H)1^þ;

found 479.2861. Anal. calcd for C₂₃H₂₈D₆O₃F₆: C, 57.73; found

57.90.

(1R,3aR,7aR)-7a-Methyl-1-[(R)-6,6,6-trifluoro-1-(5,5,5-tri-

deutero-4-trideuteromethyl-4-trimethylsilanyloxy-pentyl)-5-tri-

fluoromethyl-5-trimethylsilanyloxy-hex-3(E)-enyl]-octahydro-

inden-4-one (14R-b). The silylation of 14R-a was repeated as

described for the preparation of the disilyl ether 12R-b to give

(1R,3aR,7aR)-7a-Methyl-1-[(S)-6,6,6-trifluoro-1-(5,5,5-tri-

deutero-4-trideuteromethyl-4-trimethylsilanyloxy-pentyl)-5-tri-

fluoromethyl-5-trimethylsilanyloxy-hex-3(Z)-enyl]-octahydro-

inden-4-one (16S-b). The ketone-diol 16S-a (0.74 g, 1.55 mmol)

5514 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17

Maehr et al.

was silylated as described for the preparation of the disilyl ether

12R-b to give 16S-b (0.864 g, 90%); TLC (1:9 EtOAc-hexane) R_f

0.43. This material was divided into three nearly equal parts and

reservedfor the synthesesof4S-1, 4S-2, and4S-3 describedbelow.

(1R,3aR,7aR)-7a-Methyl-1-[(R)-6,6,6-trifluoro-5-hydroxy-

1-(5,5,5-trideutero-4-hydroxy-4-trideuteromethyl-pentyl)-5-tri-

pooled and evaporated (0.100 g). This residue was taken up in

methyl formate, filtered, and transferred to a 5 mL flask and

then evaporated to yield 2S-1 as a solid, white foam that was

dried at 0.5 Torr for 2 h; UV λ_max(ε) 212 (14801), 264 (16493)

nm; [R]_Dþ16.9° (c 0.37, methanol). ¹H NMR (400 MHz,

29

CDCl₃) δ 0.53 (s, 3 H), 1.21-1.63 (m, 12 H), 1.63-1.77 (m, 2 H),

1.80-2.13 (m, 8 H), 2.26-2.42 (m, 2 H), 2.52 (dd, J=17.7, 2.8

Hz, 1 H), 2.57 (dd, J=14.3, 2.6 Hz, 1 H), 2.75-2.89 (m, 1 H),

3.75 (s, 1 H), 4.14-4.25 (m, 1 H), 4.42 (dd, J=6.9, 4.2 Hz, 1 H),

4.98 (s, 1 H), 5.32 (s, 1 H), 6.01 (d, J=11.2 Hz, 1 H), 6.23 (br s, 1

H), 6.35 (d, J=11.2 Hz, 1 H). ¹³C NMR (400 MHz, CDCl₃) δ

12.18, 20.19, 21.71, 22.17, 23.58, 27.42, 29.01, 32.95, 38.73,

40.10, 42.64, 43.13, 45.08, 45.64, 53.19, 56.14, 66.84, 70.78,

71.15, 71.94, 89.89, 111.88, 117.22, 121.35, 124.75, 133.08,

142.42, 147.30. ¹⁹F NMR (376.31 MHz) δ -78.28, 6F, s.

LRMS-EI(þ) m/z 612 (4, M), 594 (40, M-H₂O); 594 (39, M-

2H₂O), 558 (100, M-3H₂O). HRMS-ES(þ) calcd for

C₃₂H₃₈D₆O₄F₆: 635.3412 (M þ Na)1^þ; found 35.3407.

fluoromethyl-hex-3(Z)-enyl]-octahydro-inden-4-one

(16R-a).

The triol 15R (0.90 g, 1.87 mmol) was oxidized as described

for the preparation of the ketone analogue 16S-a to afford, after

chromatography, a white foam (0.82 g, 91%). H NMR (400

1

MHz, CDCl₃) δ 0.62 (s, 3 H), 1.30 (br s, 7 H), 1.44-1.79 (m, 6

H), 1.79-1.96 (m, 2 H), 2.02 (d, J=9.8 Hz, 2 H), 2.12 - 2.32 (m, 2

H), 2.45 (dd, J=11.4, 7.8 Hz, 2 H), 2.59 (ddd, J=16.3, 8.3, 8.2

Hz, 1 H), 5.42 (d, J=12.2 Hz, 1 H), 5.54 (br s, 1 H), 5.94 (ddd, J=

12.2, 8.0, 6.3 Hz, 1 H). ¹³C NMR (400 MHz, CDCl₃) δ 12.62,

18.65, 18.94, 24.09, 27.18, 29.87, 31.38, 38.57, 39.75, 40.83,

43.47, 50.03, 52.58, 61.88, 71.54, 77.23, 117.21, 122.83, 141.35,

4

212.31. ¹⁹F NMR (376.31 MHz) δ -77.81, 3F, q, J_F,F=9.78

Hz, -78.20, 3F, q, ⁴J_F,F=9.91 Hz. LR MS-ES(-) m/z 478 (18,

M) 477 (100, M - H). HRMS-ES(þ) calcd for C₂₃H₂₈D₆O₃F₆:

501.2681 (M þ Na)1^þ; found 501.2682.

(20R)-1r,25-Dihydroxy-21-(3-hydroxy-3-trideuteromethyl-

4,4,4-trideutero-butyl)- 26,26,26,27,27,27-hexafluoro-23-yne-cho-

lecalciferol (2R-1). A two-neck flask, equipped with magnetic

stirrer, thermometer, and a Claisen adapter containing a nitrogen

sweep and rubber septum, was charged with diphenylphosphine

oxide 17 (0.465 g) and THF (3.5 mL). The solution was cooled to

-70 °C, and a 1.6 M solution of butyl lithium in hexane (0.50 mL)

was added dropwise over a 10 min period. To this red soln was

added, viasyringe, dropwise, within 20 min, ketone 12R-b (0.2895

g, 0.466 mmol) dissolved in THF (3 mL). After 5 h, the mixture

was allowed to warm to -35 °C and a satd ammonium chloride

soln (10 mL) was added. The mixture was allowed to warm to

10 °C and was then distributed between hexane (60 mL) and

enough water (5 mL) to dissolve all salts. The extract was washed

with 5:1 brine-pH 7 buffer (10 mL) and then dried and evapo-

rated. The resulting residue (0.71 g) was flash-chromatographed

using hexane f 1:72 f 1:39 EtOAc-hexane as mobile phases to

give a mixture (0.38 g) of the tetrasilyl (TLC 1:19, EtOAc-

hexane, R_f0.76) and the trisilyl ether, which lacks the silyl

function at the gem-trifluoromethyl-carbinol site (TLC 1:19,

EtOAc-hexane, R_f0.33). This mixture was dissolved in a 1 M

soln of TBAF (4 mL) and allowed to remain at room temperature

in the dark for 45 h. The soln was diluted with brine (5 mL) and

equilibrated with EtOAc (40 mL) and water (10 mL). The

aqueous phase was extracted once with EtOAc (10 mL), and

the combined extracts were washed with water (5 ꢀ 10 mL) and

brine (10 mL) and then dried and evaporated. The residue was

flash-chromatographed using 1:4 f 1:2 f 1:1 f 2:1 EtOAc-

hexane f EtOAc as mobile phases. The product fractions were

pooled, filtered, and evaporated and then coevaporated twice

from hexane to furnish a residue (0.25 g) that was taken up in

methyl formate and allowed to crystallize. The suspension was

concentrated, diluted with pentane, and refrigerated. The ML

was decanted from the crystalline 2R-1, 0.198 g, 69.3%; UV λ_max

(1R,3aR,7aR)-7a-Methyl-1-[(R)-6,6,6-trifluoro-1-(5,5,5-tri-

deutero-4-trideuteromethyl-4-trimethylsilanyloxy-pentyl)-5-tri-

fluoromethyl-5-trimethylsilanyloxy-hex-3(Z)-enyl]-octahydro-

inden-4-one (16R-b). The ketone-diol 16R-a (0.82 g) was

silylated as described for the preparation of the disilyl ether

12R-b to give 16R-b (1.03 g, 97%); TLC (1:4 and 1:39 EtOAc-

hexane) R_f0.71 and 0.08, respectively. This material was divided

into three nearly equal parts and reserved for the syntheses of

4R-1, 4R-2, and 4R-3 described below.

(20S)-1r,25-Dihydroxy-21-(3-hydroxy-3-trifluoromethyl-4,4,

4-trifluoro-but-1-ynyl)-26,26,26,27,27,27-hexadeutero-cholecal-

ciferol (2S-1). A three-neck flask equipped with a magnetic

stirrer, thermometer, and a Claisen adapter with addition

funnel, nitrogen sweep, and a rubber septum was charged with

17 (0.2973 g, 0.52 mmol) and THF (2 mL). The flask was cooled

to -75 °C and a 1.6 M butyllithium soln in hexane (0.325 mL,

0.52 mmol) was added dropwise over a 10 min period, followed

by a soln of 12S-b (0.1337 g, 0.215 mmol) in THF, at the same

rate. Although no starting ketone was detectable after 4 h, the

condensation product was a mixture of tri- and tetra-silyl ethers

with R_fvalues of 0.30 and 0.57, respectively (TLC, 1:19 EtOAc-

hexane). The reaction was quenched by the addition of a satd

ammonium chloride soln (5 mL) at -65 °C, allowed to warm to

10 °C, and then distributed between hexane (35 mL) and water

(10 mL). The organic layer was washed with 5:2 brine-pH 7

buffer (1 M, 7 mL), dried, and evaporated. At this stage only the

trisilyl ether was detectable. This material was chromato-

graphed on a flash column, using hexane f 1:72 f 1:39 f

1:19 EtOAc-hexane as mobile phases to yield the protected

2S-1 as colorless residue (0.1561 g). This residue was taken up in

pentane, filtered, evaporated to a white foam, and dried at 0.5

Torr for 1 h, 0.1503 g. ¹H NMR (400 MHz, CDCl₃) δ 0.06 (s, 6

H), 0.06 (s, 6 H), 0.11 (s, 9 H), 0.53 (s, 3 H), 0.87 (s, 18 H), 1.14-

1.53 (m, 13 H), 1.62-1.81 (m, 3 H), 1.82-1.95 (m, 3 H), 2.01 (t, J

=9.7 Hz, 1 H), 2.22 (dd, J=13.1, 7.4 Hz, 1 H), 2.37-2.56 (m, 3

H), 2.83 (d, J=15.8 Hz, 1 H), 3.45 (s, 1 H), 4.13-4.23 (m, 1 H),

4.34-4.40 (m, 1 H), 4.86 (d, J=2.1 Hz, 1 H), 5.18 (d, J=1.5 Hz, 1

H), 6.02 (d, J=11.3 Hz, 1 H), 6.23 (d, J=11.3 Hz, 1 H). LRMS-

ES(-) m/z 958(18, M - H þ HCOOH), 856 (100, M-C₄H₉).

HRMS-ES(þ) calcd for C₄₇H₇₄D₆O₄Si₃F₆: 913.5718 (M þ

H)1^þ, found 913.5723. This material was dissolved in a 1 M

TBAF soln in THF (3 mL) and allowed to stand for 45 h and

then diluted with brine (5 mL), stirred for 5 min, and transferred

to a separatory funnel with EtOAc (40 mL) and water (8 mL).

The aqueous phase was extracted once with EtOAc (10 mL), and

the combined layers were washed with water (5 ꢀ 10 mL) and

brine (10 mL) and then dried and evaporated (0.1261 g). The

resulting residue was chromatographed using 1:1f 2:1 f 4:1

EtOAc-hexane as mobile phases. The product fractions were

29

(ε) 211 (17454), 265 (18463) nm; [R]_Dþ13.2° (c 0.25, methanol).

¹H NMR (400 MHz, DMSO-d₆) δ 0.51 (s, 3 H), 1.09-1.69 (m, 15

H), 1.73-1.87 (m, 2 H), 1.87-1.99 (m, 2 H), 2.16 (dd, J=13.5, 5.4

Hz, 1 H), 2.26 (d, J=16.9 Hz, 1 H), 2.35 (d, J=11.7 Hz, 1 H), 2.42

(d, J=16.9 Hz, 1 H), 2.49 (s, 1 H), 2.73-2.83 (m, 1 H), 3.92-4.02

(m, 1 H), 4.05 (s, 1 H), 4.13-4.23 (m, 1 H), 4.56 (d, J=3.6 Hz, 1

H), 4.75 (d, J=1.6 Hz, 1 H), 4.86 (d, J=4.6 Hz, 1 H), 5.22 (br s, 1

H), 5.98 (d, J=11.1 Hz, 1 H), 6.18 (d, J=11.1 Hz, 1 H), 8.93 (s, 1

H). ¹³C NMR (400 MHz, DMSO-d₆) δ 11.89, 20.17, 20.51, 21.82,

23.14, 26.71, 28.38, 31.52, 38.76, 39.82, 43.13, 43.83, 44.94, 45.13,

52.29, 55.75, 65.07, 68.40, 68.50, 70.20, 70.40, 89.59, 109.89,

117.60, 121.47, 122.37, 135.87, 139.64, 149.33. ¹⁹F NMR

(376.31 MHz) δ -76.77 Hz, 6F, s; LRMS-ES(-) m/z 611 (100,

M - 1). HRMS-ES(þ) calcd for C₃₂H₃₈D₆O₄F₆: 635.3412 (M þ

Na)1^þ; found 635.3412.

(20S)-1r,25-dihydroxy-21-(3-hydroxy-3-trifluoromethyl-4,4,

4-trifluoro-but-1-ynyl)-26,26,26,27,27,27-hexadeutero-19-nor-cho-

lecalciferol (2S-2). A three-neck flask, equipped with magnetic

Article

Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17 5515

stirrer, thermometer, and a Claisen adapter containing a nitrogen

sweep and rubber septum, was charged with diphenylphosphine

oxide 18 (0.2973 g, 0.52 mmol) and THF (2 mL) freshly distilled

from sodium benzophenone ketyl. The solution was cooled to

-70 °C, and a 1.6 M solution of butyllithium in hexane

(0.325 mL, 0.52 mmol) was added dropwise over a 10 min period.

To this soln was added, via syringe, dropwise over a 10 min

period, a soln of 12S-b (0.1271 g) in a soln of THF (2 mL).

Starting material (TLC 1:4 EtOAc-hexane) was no longer

observed after 4 h. and a mixture of tri- and tetrasilylated

condensation products was present (TLC 1:19 EtOAc-hexane).

The mixture was allowed to warm to -55 °C, and a satd

ammonium chloride soln (5 mL) was added and then allowed

to warm further to 10 °C. The resulting mixture was equilibrated

with hexane (35 mL) and water (10 mL), and the aqueous layer

was re-extracted once with hexane (15 mL). The combined

extract, now containing mostly the trisilyl ether, was washed with

5:2 brine/pH 7 buffer and then dried and evaporated and

chromatographed using hexane f 1:72 f 1:39 EtOAc-hexane

as mobile phases. The product fractions were pooled and evapo-

rated to give a colorless oil (0.1737 g) that was taken up in

pentane, filtered, and evaporated. The resulting white foam was

dried at0.5Torrandrepresentsthe trisilylether of2S-2. ¹H NMR

(400 MHz, CDCl₃) δ 0.05 (s, 9 H), 0.06 (br s, 3 H), 0.11 (s, 9 H),

0.54 (s, 3 H), 0.87 (d, J=6.0 Hz, 18 H), 1.17-1.54 (m, 13 H),

1.57-1.74 (m, 3 H), 1.74-1.83 (m, 1 H), 1.84-1.96 (m, 2 H), 2.03

(t, J=9.6 Hz, 1 H), 2.10 (dd, J=12.9, 8.0 Hz, 1 H), 2.21-2.31 (m,

1 H), 2.33-2.57 (m, 4 H), 2.82 (m, J=2.3 Hz, 1 H), 3.45 (s, 1 H),

3.99-4.16 (m, 2 H), 5.82 (d, J=11.1 Hz, 1 H), 6.16 (d, J=11.1 Hz,

1 H). The trisilyl ether obtained as described above was dissolved

in a 1 M soln of TBAF (3 mL) and allowed to remain in the dark

for 38 h. The mixture was diluted with brine (5 mL), stirred for

5 min, and equilibrated with EtOAc (40 mL) and water (8 mL).

The aqueous phase was extracted once with EtOAc (10 mL), and

the combined extracts were washed with water (5 ꢀ 10 mL) and

brine (10 mL), dried (sodium sulfate), and evaporated. The

residue was chromatographed using 1:1f 2:1f 4:1 EtOAc-

hexane f EtOAc as mobile phases. The product fractions were

pooled and evaporated. The residue (0.1038 g) was taken up in

methyl formate, filtered, and evaporated to afford 2S-2 as a

white, solid foam, dried at 0.5 Torr (0.1009 g); UV λ_m₂_a₉_x(ε) 242

(33349), 251(39366), 260 (26943) nm (methanol); [R]_Dþ46.0°

(c 0.55, methanol). ¹H NMR (400 MHz, CDCl₃) δ 0.55 (s, 3 H),

1.22-1.48 (m, 9 H), 1.48-1.62 (m, 4 H), 1.61-1.76 (m, 4 H),

1.74-1.85 (m, 1 H), 1.85-2.09 (m, 5 H), 2.17-2.27 (m, 2 H), 2.35

(dd, J=17.3, 7.0 Hz, 1 H), 2.48 (dd, J=13.3, 2.9 Hz, 1 H), 2.57

(dd, J=17.3, 3.8 Hz, 1 H), 2.74 (dd, J=13.3, 3.8 Hz, 1 H), 2.81 (m,

1 H), 4.04 (br s, 1 H), 4.12 (br s, 1 H), 5.61 (br s, 1 H), 5.86 (d, J=

11.3 Hz, 1 H), 6.31 (d, J=11.3 Hz, 1 H). ¹³C NMR (400 MHz,

CDCl₃) δ 12.29, 20.13, 22.03, 22.18, 23.48, 27.45, 28.86, 33.23,

37.19, 38.66, 40.16, 42.13, 42.89, 44.61, 45.54, 53.40, 56.11, 67.23,

67.43, 71.04, 72.15, 90.22, 115.48, 121.26, 123.65, 131.31, 142.30.

LRMS-ES(-) m/z 599 (100, M - H). HRMS-ES(þ) calcd for

C₃₁H₃₈D₆O₄F₆: 623.3412 (M þ Na)1^þ; found 623.3414.

1.79-2.11 (m, 5 H), 2.23-2.32 (m, 2 H), 2.39-2.48 (m, 2 H),

2.70-2.83 (m, 1 H), 3.74-3.84 (m, 1 H), 3.84-3.93 (m, 1 H),

4.05 (s, 1 H), 4.39 (d, J=3.6 Hz, 1 H), 4.49 (d, J=3.6 Hz, 1 H),

5.81 (d, J=11.1 Hz, 1 H), 6.08 (d, J=11.1 Hz, 1 H), 8.94 (br s, 1

H). ¹³C NMR (400 MHz, DMSO-d₆) δ 12.01, 20.17, 21.72,

23.01, 26.79, 28.22, 31.48, 36.95, 38.79, 39.82, 42.23, 43.81,

44.99, 52.26, 55.71, 65.28, 65.55, 68.38, 70.19, 70.39, 89.58,

116.04, 120.81, 121.46, 134.70, 139.02. ¹⁹F NMR (376.31 MHz)

δ -76.76 Hz, 6F, s. LRMS-ES(-) m/z 599 (30, M - H). HRMS-

ES(þ) calcd for C₃₁H₃₈D₆O₄F₆: 623.3412 (M þ Na)1^þ; found

623.3410. Roentgen diffraction analysis (Figure 2): C₃₁H₄₄-

F₆O₄, orthorhombic, space group P2₁2₁2₁with unit cell dimen-

˚

sions of a=7.3400(15), b=16.6070(3), c=25.9300(5) A, volume

3

3160.9(11) A (Z=4).

˚

(20S)-1r-Fluoro-21-(3-hydroxy-3-trifluoromethyl-4,4,4-tri-

fluoro-but-1-ynyl)-25-hydroxy-26,26,26,27,27,27-hexadeutero-

cholecalciferol (2S-3). The diphenylphosphine oxide 19 (0.286 g,

0.61 mmol) was deprotonated with a 1.6 M solution of butyl-

lithium in hexane and allowed to react with the ketone 12S-b

(0.159 g, 0.256 mmol), and the resulting condensation product

was purified as described for 2S-2; the residue was dissolved in a

1 M soln of TBAF (3 mL) and allowed to stand for 24 h at

ambient temperature. The deprotected product was isolated and

purified by chromatography as described for 2S-2 to afford 2S-3

as a white solid foam (75 mg, 47% from 12S-b); UV λ_max(ε) 210

23

(14386), 242(14376), 270 (14177) nm (methanol); [R]_Dþ27.4°

(c 0.25, methanol).; ¹H NMR (400 MHz, DMSO-d₆) δ 0.50 (s, 3

H), 1.03-2.01 (m, 20 H), 2.05-2.20 (m, 2 H), 2.43-2.56 (m, 2

H), 2.84 (d, J=12.4 Hz, 1 H), 3.83-3.99 (m, 1 H), 4.05 (s, 1 H),

4.86 (d, J=4.5 Hz, 1 H), 4.99 (s, 1 H), 5.15 (d, J=48.4 Hz, 1 H),

5.39 (br s, 1 H), 5.94 (d, J=11.2 Hz, 1 H), 6.37 (d, J=11.2 Hz, 1

H), 8.93 (s, 1 H). ¹³C NMR (400 MHz, DMSO-d₆) δ 11.73,

20.30, 20.36, 21.63, 23.11, 26.75, 28.39, 31.93, 38.16, 40.70,

43.79, 44.88, 45.12, 52.22, 55.59, 64.52, 68.41, 7.45, 70.82,

89.41, 91.29, 92.96, 115.55, 117.09, 121.57, 124.16, 133.12,

141.75, 143.19. ¹⁹F NMR (376.31 MHz) δ -76.64 Hz, 6F, br

m. LRMS-ES(-) m/z 613 (100, M - H). HRMS-ES(þ) calcd for

C₃₂H₃₇D₆O₃F₇: 637.3369 (M þ Na)1^þ, found 637.3362.

(20R)-1r-Fluoro-21-(3-hydroxy-3-trideuteromethyl-4,4,4-tri-

deutero-butyl)-25-hydroxy-26,26,26,27,27,27-hexafluoro-23-

yne-cholecalciferol (2R-3). The diphenylphosphine oxide 19

(0.3042 g, 0.646 mmol) was deprotonated with a 1.6 M solution

of butyllithium in hexane and allowed to react with the ketone

12R-b (0.159 g, 0.61 mmol) and the condensation product was

purified as described for 2S-2. The resulting material (0.24 g)

was dissolved in a 1 M soln of TBAF (3 mL) and allowed to

stand for 24 h at ambient temperature. The deprotected product

was isolated (0.17 g) and purified by chromatography as de-

scribed to afford 2R-3 as a white, solid foam (0.1552 g, 41%

from ketone; UV λ_max(ε) 209 (15954), 242(14652), 270 (14328)

29

nm (methanol); [R]_Dþ34.3° (c 0.24, methanol). ¹H NMR (400

MHz, CDCl₃) δ 0.55 (s, 3 H), 1.16-1.79 (m, 16 H), 1.80-2.07

(m, 5 H), 2.12-2.26 (m, 2 H), 2.30 (dd, J=12.9, 8.0 Hz, 2 H), 2.44

(dd, J=17.3, 3.2 Hz, 1 H), 2.62 (dd, J=13.1, 2.9 Hz, 1 H), 2.76-

2.89 (m, 1 H), 4.16-4.27 (m, 1 H), 5.13 (ddd, J=49.6, 6.0, 3.6

Hz, 1 H), 5.10 (s, 1 H), 5.39 (br s, 1 H), 6.02 (d, J=11.2 Hz, 1 H),

6.39 (d, J=11.2 Hz, 1 H). ¹⁹F NMR (376.31 MHz) δ -78.36 Hz,

6F, br m. LRMS-ES(-) m/z 659 (35, M þ HCOOH-H), 613

(100, M - H). HRMS-ES(þ) calcd for C₃₂H₃₇D₆O₃F₇: 637.3369

(M þ Na)1^þ; found 637.3365.

(20R)-1r,25-Dihydroxy-21-(3-hydroxy-3-trideuteromethyl-4,

4,4-trideutero-butyl)- 26,26,26,27,27,27-hexafluoro-23-yne-19-

nor-cholecalciferol (2R-2). The diphenylphosphine oxide 18

(0.4222 g, 0.74 mmol) was deprotonated with a 1.6 M solution

of butyllithium in hexane and allowed to react with the ketone

12R-b (0.257 g, 0.414 mmol) as described for the preparation of

2S-1. The condensation product (0.31 g) was purified as de-

scribed for the epimer 2S-2 and then dissolved in a 1 M soln of

TBAF (5 mL). After 18 h, the temperature was increased and

maintained at 30 °C for 16 h. The deprotected product was

isolated and purified by chromatography as described for the

epimer 2S-2. The resulting colorless syrup (0.20 g) was crystal-

lized from methyl formate, 0.15 g (60% from ketone 12R-b); mp

165-167 °C; UV λ_max(ε) 243 (35935), 251(41886), 261 (228440)

(20S)-1r,25-Dihydroxy-21-(3-hydroxy-3-trifluoromethyl-4,4,

4-trifluoro-but-1(E)-enyl)-26,26,26,27,27,27-hexadeutero-chole-

calciferol (3S-1). The diphenylphosphine oxide 17 (0.3042 g,

0.522 mmol) was deprotonated with a 1.6 M solution of

butyllithium in hexane, allowed to react with the ketone 12R-b

(0.3296 g, 0.529 mmol), and the condensation product was

purified as described for 2S-2; the resulting material (0.51 g)

was dissolved in a 1 M soln of TBAF (5 mL) and allowed

to stand for 41 h at ambient temperature. The deprotected

material was isolated (0.17 g) and purified by chromatography

30

nm (methanol); [R]_Dþ58.4° (c 0.31, methanol). ¹H NMR

(400 MHz, DMSO-d₆) δ 0.52 (s, 3 H), 1.13-1.73 (m, 17 H),

5516 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17

Maehr et al.

as described to afford 3S-1 as a white, solid foam (0.2844 g, 87%

from ketone); UV λ_max(ε) 213 (16297), 266 (17812) nm

butyllithium in hexane and allowed to react with the ketone

14R-b (0.152 g, 0.244 mmol). The condensation product was

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (3 mL) and

allowed to stand for 50 h at ambient temperature. The depro-

tected material was isolated and purified by chromatography as

described to afford 3R-2, which was crystallized from methyl

formate (90 mg, 0.1493 mmol, 61% from ketone 14R-b); UV

λ_max(ε) 243 (33111), 251(35071), 261 (26178) nm (methanol);

(methanol); [R]_Dþ9.9° (c 0.30, methanol). ¹H NMR (400

30

MHz, DMSO-d₆) δ 0.53 (s, 3 H), 1.03-1.56 (m, 13 H), 1.56-

1.72 (m, 3 H), 1.72-1.85 (m, 2 H), 1.87-2.03 (m, 2 H), 2.17 (dd,

J=13.5, 5.2 Hz, 1 H), 2.20-2.29 (m, 1 H), 2.36 (d, J=11.5 Hz, 2

H), 2.80 (d, J=10.2 Hz, 1 H), 3.98 (br s, 1 H), 4.02 (s, 1 H), 4.19

(br s, 1 H), 4.55 (d, J=3.4 Hz, 1 H), 4.76 (br s, 1 H), 4.87 (d, J=

4.5 Hz, 1 H), 5.23 (br s, 1 H), 5.61 (d, J=15.3 Hz, 1 H), 5.99 (d,

J=11.0 Hz, 1 H), 6.20 (d, J=11.0 Hz, 1 H), 6.27 (ddd, J=15.1,

7.4, 7.1 Hz, 1 H), 8.03 (s, 1 H). ¹³C NMR (400 MHz, DMSO-d₆)

δ 11.85, 19.26, 21.76, 23.17, 26.66, 28.39, 31.06, 33.31, 38.98,

43.13, 44.12, 44.94, 45.32, 52.20, 55.62, 65.10, 68.41, 68.53,

75.51, 109.97, 117.66, 119.31, 122.49, 122.74, 135.95, 137.55,

139.92, 149.52. ¹⁹F NMR (376.31 MHz) δ -75.85 Hz, 6F, m).

LRMS-ES(-) m/z 613 (40, M - H). HRMS-ES(þ) calcd for

C₃₂H₄₀D₆O₄F₆: 637.3569 (M þ Na)1^þ; found 637.3569.

28

[R]_Dþ44.2° (c 0.22, methanol). ¹H NMR (400 MHz, DMSO-

d₆) δ 0.52 (s, 3 H), 1.10-1.73 (m, 17 H), 1.76-2.14 (m, 6 H),

2.16-2.33 (m, 2 H), 2.43 (d, J=10.2 Hz, 1 H), 2.75 (d, J=10.0

Hz, 1 H), 3.73-3.84 (m, 1 H), 3.84-3.94 (m, 1 H), 4.04 (s, 1 H),

4.40 (d, J=3.7 Hz, 1 H), 4.50 (d, J=3.7 Hz, 1 H), 5.62 (d, J=15.2

Hz, 1 H), 5.80 (d, J=11.1 Hz, 1 H), 6.08 (d, J=11.1 Hz, 1 H),

6.26 (ddd, J=15.2, 7.5, 7.4 Hz, 1 H), 8.03 (s, 1 H). ¹³C NMR (400

MHz, DMSO-d₆) δ 11.99, 19.34, 21.79, 23.09, 26.87, 28.27,

30.92, 33.39, 36.97, 42.24, 44.03, 44.63, 45.19, 52.10, 55.65,

65.31, 65.58, 68.43, 75.47, 116.00, 119.13, 120.86, 122.62,

134.62, 137.55, 139.21. LRMS-ES(-) m/z 601 (34, M - H).

HRMS-ES(þ) calcd for C₃₁H₄₀D₆O₄F₆: 625.3569, found

625.3575.

(20R)-1r,25-Dihydroxy-21-(3-hydroxy-3-trideuteromethyl-4,

4,4-trideutero-butyl)- 26,26,26,27,27,27-hexafluoro-23(E)-ene-

cholecalciferol (3R-1). The diphenylphosphine oxide 17 (0.3546

g, 0.608 mmol) was deprotonated with a 1.6 M solution of

butyllithium in hexane and allowed to react with the ketone

12R-b (0. 0.2639 g, 0.424 mmol) as described. The condensation

product was purified as described for 2S-2; the resulting material

was dissolved in a 1 M soln of TBAF (5 mL) and allowed to

stand for 41 h at ambient temperature. The deprotected material

was isolated and purified by chromatography as described to

afford 3S-1 as a white, solid foam (0.1842 g, 70% from ketone

14R-b; UV λ_max(ε) 205 (14284), 211 (15019), 265 (16299) nm

(20S)-1r-Fluoro-25-hydroxy-21-(3-hydroxy-3-trifluorometh-

yl-4,4,4-trifluoro-but-1(E)-enyl)-26,26,26,27,27,27-hexadeutero-

cholecalciferol (3S-3). The diphenylphosphine oxide 19 (0.35 g,

0.743 mmol)) was deprotonated with a 1.6 M solution of

butyllithium in hexane and allowed to react with the ketone

14S-b (0.295 g, 0.474 mmol). The condensation product was

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (4 mL) and

allowed to stand for 25 h at ambient temperature. The depro-

tected material was isolated and purified by chromatography as

described to afford a crystalline residue. This material was

coevaporated twice from hexane and then taken up in methyl

formate, filtered, concentrated to a volume of ca. 1-2 mL, and

warmed to dissolve most crystals and then allowed to crystallize,

first at room temperature, then refrigerated. The mother liquor

was decanted, the solids washed with pentane then dried to give

3S-3 (0.14 g, 48%); mp 148-50 °C; UV λ_ma₂_x₈(ε) 212 (16206),

242(15668), 271 (15186) nm (methanol); [R]_Dþ30.72° (c 0.23,

30

1

(methanol); [R]_Dþ14.1° (c 0.20, methanol). H NMR (400

MHz, DMSO-d₆) δ 0.52 (s, 3 H), 1.10-1.57 (m, 13 H), 1.57-

1.71 (m, 3 H), 1.73-1.85 (m, 2 H), 1.90 (d, J=9.6 Hz, 1 H), 1.96

(t, J=9.4 Hz, 1 H), 2.01-2.12 (m, 1 H), 2.16 (dd, J=13.4, 5.5 Hz,

1 H), 2.22-2.32 (m, 1 H), 2.36 (d, J=11.7 Hz, 1 H), 2.79 (m, J=

9.6 Hz, 1 H), 3.98 (br s, 1 H), 4.03 (s, 1 H), 4.15-4.26 (m, 1 H),

4.56 (d, J=3.2 Hz, 1 H), 4.76 (d, J=1.9 Hz, 1 H), 4.87 (d, J=4.7

Hz, 1 H), 5.23 (d, J=1.1 Hz, 1 H), 5.61 (d, J=15.6 Hz, 1 H), 5.99

(d, J=11.3 Hz, 1 H), 6.19 (d, J=11.3 Hz, 1 H), 6.22-6.32 (m, 1

H), 8.03 (br s, 1 H). LRMS-ES(-) m/z 613 (35, M - H). HRMS-

ES(þ) calcd for C₃₂H₄₀D₆O₄F₆: 637.3569 (M þ Na)1^þ; found

637.3570.

1

methanol). H NMR (400 MHz, DMSO-d₆) δ 0.52 (s, 3 H),

(20S)-1r,25-Dihydroxy-21-(3-hydroxy-3-trifluoromethyl-4,4,

4-trifluoro-but-1(E)-enyl)-26,26,26,27,27,27-hexadeutero-19-

nor-cholecalciferol (3S-2). The diphenylphosphine oxide 18

(0.5340 g, 0.936 mmol) was deprotonated with a 1.6 M solution

of butyllithium in hexane, allowed to react with the ketone 14S-b

(0.3498 g, 0.561 mmol), and the condensation product was

purified as described for 2S-2; the resulting partially desilylated

material (0.48 g) was dissolved in a 1 M soln of TBAF (5 mL)

and allowed to stand for 63 h at ambient temperature. The

deprotected material was isolated and purified by chromato-

graphy as described to afford 3S-2 as a white, solid foam (0.2862

g, 84%); UV λ_max(ε) 243 (34631), 251(40591), 261 (27593) nm

(methanol); [R]_Dþ42.3° (c 0.25, methanol). H NMR (400

MHz, DMSO-d₆) δ 0.53 (s, 3 H), 1.01-1.72 (m, 17 H), 1.72-

2.12 (m, 5 H), 2.15-2.31 (m, 2 H), 2.31-2.47 (m, 2 H), 2.66-

2.83 (m, 1 H), 3.76-3.95 (m, 2 H), 4.04 (s, 1 H), 4.40 (d, J=3.5

Hz, 1 H), 4.50 (d, J=3.5 Hz, 1 H), 5.61 (d, J=15.2 Hz, 1 H), 5.81

(d, J=11.0 Hz, 1 H), 6.08 (d, J=11.0 Hz, 1 H), 6.28 (ddd, J=

15.2, 7.4, 7.1 Hz, 1 H), 8.03 (s, 1 H). ¹³C NMR (400 MHz,

DMSO-d₆) δ 11.98, 19.26, 21.66, 23.04, 26.72, 28.23, 31.06,

33.29, 36.98, 39.01, 42.23, 44.15, 44.61, 45.19, 52.20, 55.59,

65.32, 65.59, 68.43, 75.51, 116.12, 119.31, 120.95, 122.74,

134.76, 137.58, 139.34. LRMS-ES(-) m/z 601 (30, M - H).

HRMS-ES(þ) calcd for C₃₁H₄₀D₆O₄F₆: 625.3569 (M þ Na)1^þ;

found 625.3572.

1.03-1.58 (m, 13 H), 1.59-1.85 (m, 4 H), 1.92 (d, J=11.5 Hz,

1 H), 1.99 (t, J=9.2 Hz, 1 H), 2.05-2.18 (m, 2 H), 2.18-2.29 (m,

1 H), 2.29-2.42 (m, 1 H), 2.52-2.56 (m, 1 H), 2.83 (d, J=12.6

Hz, 1 H), 3.82-3.98 (m, 1 H), 4.02 (s, 1 H), 4.86 (d, J=4.3 Hz, 1

H), 5.00 (s, 1 H), 5.15 (d, J=49.7 Hz, 1 H), 5.39 (br s, 1 H), 5.61

(d, J=15.6 Hz, 1 H), 5.95 (d, J=11.1 Hz, 1 H), 6.20-6.33 (m, 1

H), 6.37 (d, J=11.1 Hz, 1 H), 8.03 (s, 1 H). ¹³C NMR (400 MHz,

DMSO-d₆) δ 11.89, 19.27, 21.62, 23.09, 26.63, 28.38, 31.05,

33.24, 38.94, 40.70, 44.11, 44.88, 45.40, 52.14, 55.55, 64.52,

68.40, 75.50, 91.28, 92.94, 15.51, 117.02, 119.30, 122.73,

124.17, 133.05, 137.52, 141.91, 143.21. ¹⁹F NMR (376.31 MHz)

δ -75.86 Hz, 6F, m. LRMS-ES(-) m/z 661 (100, M þ HCOO^-),

615 (10, M - H). HRMS-ES(þ) calcd for C₃₂H₃₉D₆O₃F₇:

639.3526 [M þ Na]1^þ; found 639.3515.

28

1

(20R)-1r-Fluoro-25-hydroxy-21-(3-hydroxy-3-trideuterome-

thyl-4,4,4-trideutero-butyl)- 26,26,26,27,27,27-hexafluoro-23(E)-

ene-cholecalciferol (3R-3). The diphenylphosphine oxide 19

(0.374 g, 0.795 mmol) was deprotonated with a 1.6 M solution

of butyllithium in hexane and allowed to react with the ketone

14R-b (0.262 g, 0.421 mmol). The condensation product was

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (3 mL) and

allowed to stand for 24 h at ambient temperature. The depro-

tected material was isolated and purified by chromatography as

described to afford 3R-3 as a white, solid foam (0.153 g, 59%;

28

(20R)-1r,25-Dihydroxy-21-(3-hydroxy-3-trideuteromethyl-4,

4,4-trideutero-butyl)- 26,26,26,27,27,27-hexafluoro-23(E)-ene-

19-nor-cholecalciferol (3R-2). The diphenylphosphine oxide 18

(0.244 g, 0.427 mmol) was deprotonated with a 1.6 M solution of

UV λ_max(ε) 209 (23061), 241 (15755), 270 nm (14829); [R]_D

þ29.2° (c 0.26, methanol). ¹H NMR (400 MHz, CDCl₃) δ 0.55

(s, 3 H), 1.17-1.78 (m, 16 H), 1.78-2.13 (m, 7 H), 2.12-2.42 (m,

3 H), 2.61 (dd, J=13.1, 3.1 Hz, 1 H), 2.77-2.87 (m, 1 H), 4.20

Article

Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17 5517

(ddd, J=7.4, 3.8, 3.7 Hz, 1 H), 5.12 (ddd, J=49.7, 6.1, 3.5 Hz, 1

H), 5.10 (s, 1 H), 5.38 (br s, 1 H), 5.58 (d, J=15.4 Hz, 1 H), 6.02

(d, J=11.1 Hz, 1 H), 6.27 (ddd, J=15.4, 8.5, 6.5 Hz, 1 H), 6.39 (d,

J=11.1 Hz, 1 H). ¹³C NMR (400 MHz, CDCl₃) δ 12.21, 19.70,

22.12, 23.56, 27.44, 29.05, 31.70, 34.83, 39.95, 40.26, 40.67,

43.50, 44.93, 45.95, 53.08, 56.26, 66.50, 71.40, 75.83, 90.89,

92.60, 115.23, 117.14, 119.07, 122.42, 125.43, 131.48, 138.09,

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (5 mL) and

allowed to stand for 32 h at ambient temperature. The depro-

tected material was isolated and purified by chromatography as

described to afford 4S-2 as a white, solid foam (0.243 g, 87.6%);

28

UV λ_max(ε) 243 (34246), 252 (39955), 261 (27120) nm; [R]_D

1

þ31 (c=0.31%, methanol). H NMR (400 MHz, DMSO-d₆)

142.84, 143.00. ¹⁹F NMR (376.31 MHz) δ -77.80, 3F, q, ⁴J_F,F

=

δ 0.52 (s, 3 H), 1.07-1.72 (m, 17 H), 1.72-2.15 (m, 5 H), 2.26 (d,

J=11.3 Hz, 1 H), 2.32-2.47 (m, 2 H), 2.70-2.83 (m, 2 H), 3.80

(br s, 1 H), 3.87 (br s, 1 H), 4.02 (s, 1 H), 4.38 (d, J=3.6 Hz, 1 H),

4.49 (d, J=3.6 Hz, 1 H), 5.42 (d, J=11.9 Hz, 1 H), 5.80 (d, J=

11.1 Hz, 1 H), 5.98 - 6.13 (m, 2 H), 8.01 (s, 1 H). ¹³C NMR (400

MHz, DMSO-d₆) δ 11.91, 19.25, 21.69, 23.02, 26.40, 28.24,

29.43, 31.43, 36.96, 42.21, 44.34, 45.07, 52.31, 55.56, 65.29,

65.57, 68.47, 76.76, 116.02, 116.75, 120.94, 123.02, 134.69,

139.40, 141.38. ¹⁹F NMR (376.31 MHz) δ -75.85 Hz, 6F, m.

LRMS-ES(-) m/z 647 (55, M þ HCOOH-H^-), 602 (28, M), 601

(100, M - H). HRMS-ES(þ) calcd for C₃₁H₄₀D₆O₄F₆: 625.3569

[M þ Na]1^þ; found 625.3576.

7.90 Hz, -77.88, 3F, q, ⁴J_F,F=7.80 Hz. LRMS-ES(-) m/z 661

(100, M þ HCOO^-), 615 (20, M - H). HRMS-ES(þ) calcd for

C₃₂H₃₉D₆O₃F₇: 639.3526 [M þ Na]1^þ; found 639.3523.

(20S)-1r,25-Dihydroxy-21-(3-hydroxy-3-trifluoromethyl-4,4,

4-trifluoro-but-1(Z)-enyl)-26,26,26,27,27,27-hexadeutero-chole-

calciferol (4S-1). The diphenylphosphine oxide 17 (0.497 g,

0.853 mmol) was deprotonated with a 1.6 M solution of

butyllithium in hexane and allowed to react with the ketone

16S-b (0.288 g, 0.462 mmol). The condensation product was

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (5 mL) and

allowed to stand for 19 h at ambient temperature. The depro-

tected material was isolated and purified by chromatography as

described to afford 4S-1 as a white, solid foam (0.238 g, 84%);

(20R)-1r,25-Dihydroxy-21-(3-hydroxy-3-trideuteromethyl-4,

4,4-trideutero-butyl)-26,26,26,27,27,27-hexafluoro-23(Z)-ene-

19-nor-cholecalciferol (4R-2). The diphenylphosphine oxide 18

(0.5078 g, 0.889 mmol) was deprotonated with a 1.6 M solution

of butyllithium in hexane and allowed to react with the ketone

16R-b (0.3298 g, 0.529 mmol). The condensation product was

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (4 mL) and

allowed to stand for 48.5 h at ambient temperature. The

deprotected material was isolated and purified by chromatog-

raphy as described to afford 4R-2 as a white, solid foam (0.2446

g, 76.7%); UV λ_max(ε) 243 (32530), 252 (38012), 261 (25710) nm;

28

UV λ_max(ε) 210 (66396), 264 (16769) nm; [R]_Dþ29.2° (c 0.26,

1

methanol). H NMR (400 MHz, DMSO-d₆) δ 0.52 (s, 3 H),

1.09-1.37 (m, 8 H), 1.37-1.58 (m, 5 H), 1.58-1.70 (m, 3 H),

1.74-1.86 (m, 2 H), 1.90-2.03 (m, 2 H), 2.17 (dd, J=13.4, 5.1

Hz, 1 H), 2.32-2.47 (m, 2 H), 2.71-2.86 (m, 2 H), 3.99 (br s, 1

H), 4.02 (s, 1 H), 4.19 (br s, 1 H), 4.55 (d, J=2.1 Hz, 1 H), 4.76 (br

s, 1 H), 4.87 (d, J=4.3 Hz, 1 H), 5.23 (br s, 1 H), 5.42 (d, J=11.5

Hz, 1 H), 5.96-6.10 (m, 2 H), 6.19 (d, J=11.5 Hz, 1 H), 8.01 (s, 1

H). ¹³C NMR (400 MHz, DMSO-d₆) δ 11.80, 19.27, 21.79,

23.17, 26.34, 28.41, 29.44, 31.43, 43.12, 44.33, 45.23, 52.30,

55.61, 65.08, 68.49, 76.78, 109.94, 116.77, 117.57, 122.50,

123.02, 135.88, 140.02, 141.37, 149.52. ¹⁹F NMR (376.31 MHz)

δ -75.87 Hz, 6F, m. LRMS-ES(-) m/z 659 (85, M þ HCOO^-),

613 (100, M - H). HRMS-ES(þ) calcd for C₃₂H₄₀D₆O₄F₆:

637.3569 [M þ Na]1^þ; found 637.3574.

25

[R]_Dþ51.4° (c = 0.33%, methanol). ¹H NMR (400 MHz,

DMSO-d₆) δ 0.52 (s, 3 H), 1.13-1.72 (m, 17 H), 1.80 (br s, 1 H),

1.89 (d, J=11.3 Hz, 1 H), 1.93-2.12 (m, 3 H), 2.27 (d, J=11.1

Hz, 1 H), 2.32-2.48 (m, 2 H), 2.53-2.62 (m, 1 H), 2.69-2.81 (m,

1 H), 3.76-3.84 (m, 1 H), 3.88 (br s, 1 H), 4.02 (s, 1 H), 4.38 (d,

J=3.5 Hz, 1 H), 4.48 (d, J=3.5 Hz, 1 H), 5.42 (d, J=11.9 Hz, 1

H), 5.80 (d, J=11.1 Hz, 1 H), 6.01-6.11 (m, 2 H), 7.99 (s, 1 H).

¹³C NMR (400 MHz, DMSO-d₆) δ 12.11, 19.42, 21.71, 23.06,

26.55, 28.25, 29.40, 31.39, 36.95, 39.57, 42.23, 44.17, 45.20,

52.25, 55.51, 65.27, 65.54, 68.44, 76.73, 115.94, 116.70, 120.83,

122.92, 134.56, 139.25, 141.22. ¹⁹F NMR (376.31 MHz) δ -

75.82 Hz, 6F, m. LRMS-ES(-) m/z 647 (30, M þ HCOO^-), 602

(20R)-1r,25-Dihydroxy-21-(3-hydroxy-3-trideuteromethyl-4,

4,4-trideutero-butyl)-26,26,26,27,27,27-hexafluoro-23(Z)-ene-

cholecalciferol (4R-1). The diphenylphosphine oxide 17 (0.429 g,

0.736 mmol) was deprotonated with a 1.6 M solution of

butyllithium in hexane and allowed to react with the ketone

16S-b (0.3518 g, 0.565 mmol). The condensation product was

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (4 mL) and

allowed to stand for 32 h at ambient temperature. The depro-

tected material was isolated and purified by chromatography as

described to afford 4R-1 as a white, solid foam (0.250 g, 72%);

(25, M), 601 (100,

M

-

H). HRMS-ES(þ) calcd for

C₃₁H₄₀D₆O₄F₆: 625.3569 [M þ Na]1^þ; found 625.3583.

(20S)-1r-Fluoro-25-hydroxy-21-(3-hydroxy-3-trifluorometh-

yl-4,4,4-trifluoro-but-1(Z)-enyl)-26,26,26,27,27,27-hexadeutero-

cholecalciferol (4S-3). The diphenylphosphine oxide 19 (0.3557

g, 0.756 mmol) was deprotonated with a 1.6 M solution of

butyllithium in hexane and allowed to react with the ketone 16S-

b (0.2700 g, 0.433 mmol) and the condensation product was

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (5 mL) and

allowed to stand for 24.5 h at ambient temperature. The

deprotected material was isolated and purified by chromatog-

raphy as described to afford 4S-3 as a white, solid foam (0.1714

g, 64%); UV λ_max(ε) 209 (13713), 243 (13893), 271 (13673) nm;

28

UV λ_max(ε) 212 (14818), 266 (17067) nm; [R]_Dþ18.1 (c =

0.29%, methanol). ¹H NMR (400 MHz, DMSO-d₆) δ 0.52 (s, 3

H), 1.12-1.58 (m, 13 H), 1.58-1.70 (m, 3 H), 1.70-1.84 (m, 2

H), 1.89 (d, J=11.1 Hz, 1 H), 1.93-2.04 (m, 1 H), 2.17 (dd, J=

13.4, 5.1 Hz, 1 H), 2.31-2.45 (m, 2 H), 2.53-2.62 (m, 1 H), 2.80

(d, J=10.2 Hz, 1 H), 3.97 (br s, 1 H), 4.02 (s, 1 H), 4.18 (br s, 1 H),

4.55 (d, J=3.4 Hz, 1 H), 4.76 (br s, 1 H), 4.87 (d, J=4.5 Hz, 1 H),

5.23 (br s, 1 H), 5.41 (d, J=11.9 Hz, 1 H), 5.99 (d, J=11.1 Hz, 1

H), 6.02-6.11 (m, 1 H), 6.19 (d, J=11.1 Hz, 1 H), 7.98 (s, 1 H).

¹³C NMR (400 MHz, DMSO-d₆) δ 11.99, 19.39, 21.80, 23.20,

26.48, 28.40, 29.39, 31.39, 43.12, 44.17, 44.91, 45.35, 52.23,

55.55, 65.04, 68.47, 76.73, 109.85, 116.69, 117.49, 122.37,

122.89, 135.74, 139.85, 141.22, 149.33. ¹⁹F NMR (376.31 MHz)

δ -75.83 Hz, 6F, m. LRMS-ES(-) m/z 659 (30, M þ HCOOH-

H^-), 613 (100, M - H). HRMS-ES(þ) calcd for C₃₂H₄₀D₆O₄F₆:

637.3569 [M þ Na]1^þ; found 637.3562.

25

[R]_Dþ16.8° (c = 0.20%, methanol). ¹H NMR (400 MHz,

DMSO-d₆) δ 0.51 (s, 3 H), 1.06-1.88 (m, 18 H), 1.88-2.04 (m, 2

H), 2.04-2.21 (m, 2 H), 2.33-2.46 (m, 1 H), 2.70-2.88 (m, 2 H),

3.83-3.97 (m, 1 H), 4.01 (s, 1 H), 4.86 (d, J=4.3 Hz, 1 H), 5.00 (s,

1 H), 5.15 (d, J=49.9 Hz, 1 H), 5.33-5.46 (m, 2 H), 5.95 (d, J=

11.1 Hz, 1 H), 6.04 (ddd, J=12.1, 6.4, 6.2 Hz, 1 H), 6.37 (d, J=

11.1 Hz, 1 H), 8.01 (s, 1 H). ¹³C NMR (400 MHz, DMSO-d₆)

δ 11.83, 19.27, 21.66, 23.09, 26.34, 28.42, 29.39, 31.42, 40.70,

44.31, 44.87, 45.31, 52.26, 55.55, 64.52, 68.46, 76.77, 91.27,

92.94, 115.47, 116.77, 116.94, 123.01, 133.00, 141.34, 141.99,

143.23. ¹⁹F NMR (376.31 MHz) δ -75.85 Hz, 6F, m. LRMS-

ES(-) m/z 661 (55, M þ HCOOH-H^-), 616 (20, M), 615 (100,

(20S)-1r,25-Dihydroxy-21-(3-hydroxy-3-trifluoromethyl-4,4,

4-trifluoro-but-1(Z)-enyl)-26,26,26,27,27,27-hexadeutero-19-nor-

cholecalciferol (4S-2). The diphenylphosphine oxide 18 (0.4436,

0.778 mmol) was deprotonated with a 1.6 M solution of

butyllithium in hexane and allowed to react with the ketone

16S-b (0.2869 g, 0.460 mmol) and the condensation product was

5518 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 17

Maehr et al.

M - H). HRMS-ES(þ) calcd for C₃₂H₃₉D₆O₃F₇: 639.3526 [M

þ Na]1^þ; found 639.3523.

indebted to the members of the Physical Chemistry Depart-

ment of Hoffmann La Roche (Vance E. Bell) for NMR

spectra (Gino Sasso), mass spectra (Richard Szypula), ele-

mental analyses (Theresa Burchfield), and UV spectra and

optical rotations (Michael Lanyi).

(20R)-1r-Fluoro-25-hydroxy-21-(3-hydroxy-3-trideuterome-

thyl-4,4,4-trideutero-butyl)-26,26,26,27,27,27-hexafluoro-23(Z)-

ene-cholecalciferol (4R-3). The diphenylphosphine oxide 19

(0.5632 g, 1.20 mmol) was deprotonated with a 1.6 M solution

of butyllithium in hexane and allowed to react with the ketone

16R-b (0.3354 g, 0.538 mmol) and the condensation product was

purified as described for 2S-2; the resulting partially desilylated

material was dissolved in a 1 M soln of TBAF (4 mL) and

allowed to stand for 23 h at ambient temperature. The depro-

tected material was isolated and purified by chromatography as

described to afford 4R-3 as a white solid that was crystallized

from methyl formate (0.285 g, 86%); UV λ_max(ε) 208 (17000),

Supporting Information Available: Elemental analyses results

of all final compounds and crystallographic data of 11S-b and

2R-2. This material is available free of charge via the Internet at

http://pubs.acs.org.

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25

242 (16068), 270 (15615) nm; [R]_Dþ36.2° (c = 0.31%,

1

methanol). H NMR (400 MHz, DMSO-d₆) δ 0.51 (s, 3 H),

1.12-1.58 (m, 13 H), 1.58-1.84 (m, 4 H), 1.90 (d, J=11.3 Hz, 1

H), 2.00 (t, J=9.2 Hz, 1 H), 2.04-2.20 (m, 2 H), 2.32-2.46 (m, 1

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11.3 Hz, 1 H), 6.00-6.10 (m, 1 H), 6.37 (d, J=11.3 Hz, 1 H), 7.98

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Article Doi

Calcitriol derivatives with two different side chains at C-20. V. Potent inhibitors of mammary carcinogenesis and inducers of leukemia differentiation

DOI: 10.1021/jm900780q

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