HETEROCYCLES

Paper | Regular issue | Vol. 81, No. 2, 2010, pp. 381-394
Received, 5th November, 2009, Accepted, 3rd December, 2009, Published online, 4th December, 2009.
DOI: 10.3987/COM-09-11871

■ Synthesis of 20-Epi-eldecalcitol [20-Epi-1α,25-dihydroxy-2β-(3-hydroxypropoxy)vitamin D3: 20-Epi-ED-71]

Madoka Yoshino, Kohei Eto, Keisuke Takahashi, Jun Ishihara, Susumi Hatakeyama, Yoshiyuki Ono, Hitoshi Saito, and Noboru Kubodera*

Chugai Pharmaceutical Company, Ltd., 2-1-1, Nihonbashi-Muromachi, Chuo-ku, Tokyo 103-8324, Japan

Abstract

A convergent synthesis of biologically interesting 20-epi-eldecalcitol which possesses an inverted C-21 methyl substituent at the 20-position of the side chain of 1α,25-dihydroxy-2β-(3-hydroxypropoxy)vitamin D3 (eldecalcitol) is described.

INTRODUCTION
There is intense interest in obtaining analogs more potent than calcitriol [1α,25-dihydroxyvitamin D3, (1)] in terms of regulatory effects on calcium and phosphorus metabolism with the objective of treating bone diseases such as osteoporosis.3 Eldecalcitol [1α,25-dihydroxy-2β-(3-hydroxylpropoxy)vitamin D3, developing code; ED-71, (2)], an analog of calcitriol (1) from which a hydroxypropoxy substituent at the 2β-position of the A-ring is appended, is such an analog that shows potent effects on bone therapy.4-6 Recent completion of phase III clinical trials of 2 for bone fracture prevention produced excellent results. Eldecalcitol (2) is now in preparation for approval as a promising medicine for the treatment of osteoporosis in Japan.
To explore structure-biological activity relationships between eldecalcitol (2) and related analogs, we have already synthesized 1-epi-eldecalcitol,7 3-epi-eldecalcitol,8 and 1,3-diepi-eldecalcitol9 with inherent biological interest of each targeted analogs and evaluated their biological responses. It has been reported that 20-epi-calcitriol (3), a diastereomer of calcitriol (1), which possesses an inverted C-21 methyl substituent at the 20-position of the side chain, shows remarkably enhanced biological activities compared to parent compound, 1.10 For example, 20-epi-calcitriol (3) exhibits 18 times the potency of induction of human myeloid leukemia cell (HL-60) differentiation.11 Furthermore 3 shows 50 times the inhibition of the human histiocytic lymphoma cell (U937) proliferation,12 and 4.5 times the increase in osteocalcin concentration in the human osteosarcoma cell (MG-63)13 compared to 1. These findings prompted our interest in analogs of eldecalcitol (2) epimerized at the 20-position and its biological resoponses. In this paper, we describe the synthesis of 20-epi-eldecalcitol (4) as a continuation of our modification studies on eldecalcitol (2) and preliminary biological evaluation using HL-60, U937, and MG-63 compared to 2 (Figure 1).

RESULTS AND DISCUSSION
Our synthesis of 20-epi-eldecalcitol (4) was envisioned using the Trost coupling reaction of A-ring fragment 5 derived from C2-symmetrical epoxide 6 with C/D-ring fragment 7. Fragment 7 can be obtained from the Inhoffen-Lythgoe diol (8) via a known protocol (Scheme 1).14,15
The required A-ring fragment 5 for the synthesis of 20-epi-eldecalcitol (4) was synthesized based upon the methodology that has been previously established by our group.16 Thus, cleavage of the known C2-symmetrical epoxide 617 with 1,3-propanediol [HO(CH2)3OH] in the presence of potassium tert-butoxide (t-BuOK) gave diol 9 in 86% yield. After protection of the primary hydroxyl group in 9 to give pivalate 10 in 88% yield, cleavage of the benzyl ether moiety in 10 and subsequent protection of the resulting 1,2-diol as the acetonide gave alcohol 11 in 87% overall yield. Swern oxidation of 11 and subsequent Grignard reaction of the resulting aldehyde with vinylmagnesium bromide (CH2=CHMgBr) followed by pivaloylation of the resulting alcohol afforded the dipivalate 12 as an epimeric mixture (R/S = 3/2) in 57% yield. Without separation of the epimeric mixture, the acetonide moiety in 12 was cleaved quantitatively to give diol 13.

Exposure of 13 to Mitsunobu conditions18 afforded epoxide 14 in 77% yield. The acetylene unit was successfully installed by the regioselective epoxide-opening of 14 with lithium trimethylsilylacetylide (LiC CTMS) in the presence of boron trifluoride diethyl etherate (BF3-OEt2) at -78 °C to provide ene-yne 5 as the A-ring fragment of 20-epi-eldecalcitol (4) in 36% yield after protecting group exchange from pivalate to tert-butyldimethylsilyl (TBS) ether. The accompanied (S)-ismoer 15, which consists of the requisite stereochemistry to obtain 1-epi-eldecalcitol, was separated in 24% yield by simple column chromatography (Scheme 2).

Next, we performed the synthesis of C/D-ring fragment 7. Based on the reported route19 to 7 from the Inhoffen-Lythgoe diol (8), which is obtained via ozonolysis of vitamin D2,20 we developed a convenient approach for the facile introduction of the C-23 – C-27 side chain fragment. Thus, after tosylation of the primary hydroxyl group in 8, the secondary hydroxyl moiety in 16 was protected as its TBS ether to give 17 in 88% overall yield from 8. Oxidation of 17 using dimethyl sulfoxide (DMSO) and s-collidine afforded aldehyde 18 in 78% yield. Aldehyde 18 was then subjected to the epimerization conditions to give an approximately 3:2 mixture of the aldehyde 19 in favor of the C-20-epimer. Reduction of the mixture with sodium borohydride (NaBH4) provided the corresponding C-20 epimeric alcohols. The alcohols were separated by column chromatography to obtain the desired 20-epi alcohol 20 and the naturally configured alcohol 21 in 43% and 27%, respectively.19,21 Tosylation of the primary hydroxyl group in 20 afforded tosylate 22 in 92% yield, which was treated with sodium iodide (NaI) to give iodide 23 in 94% yield. Reaction of 23 with methyl vinyl ketone (MVK) in the presence of zink (Zn) and cuprous iodide (CuI) furnished ketone 24 in 65% yield. Grignard reaction of 24 with methylmagnesium bromide (MeMgBr) gave rise to alcohol 25 with the desired side chain in 86% yield. Desilylation of 25 with 47% hydrofluoric acid (HF) followed by oxidation of 26 with tetrapropylammonium perruthenate (TPAP) and N-methylmolpholine N-oxide (NMO) gave ketone 27 in 98% yield gave ketone 27 in 98% yield. Ketone 27 was then reacted with (bromomethylene)triphenylphosphonium bromide (Ph3P+CH2Br/Br-) in the presence of sodium

hexamethyldisilazide (NaHMDS) to furnish C/D-ring fragment 7 in excellent overall yield (Scheme 3). Spectroscopic data of 26, 27, and 7 were identical to those reported in the literature.19
With A-ring fragment 5 and C/D-ring fragment 7 in hand, we next investigated the Trost coupling reaction. Upon treatment of excess 5 and 7 in the presence of tetrakis(triphenylphosphine)palladium (0) [Pd(PPh3)4] and triethylamine (Et3N) in toluene, the coupled product 28 was obtained in 42% yield. The silyl protecting groups were removed using 47% HF in acetonitrile (MeCN) to afford 20-epieldecalcitol (4) in 73% yield (Scheme 4). The 1H NMR chemical shift of the C-21 methyl resonance in 20-epi-eldecalcitol (4) (0.87 ppm) is shifted upfield by 0.04 ppm relative to natural configuration of eldecalcitol (2) (0.91 ppm)4 and is consistent with this general pattern.22-24

The results of preliminary in vitro biological evaluation of synthetic 20-epi-eldecalcitol (4) in comparison with eldecalcitol (2) and calcitriol (1) are summarized in Table 1. As anticipated, 20-epi-eldecalcitol (4) showed greatly enhanced activity toward the induction of HL-60 differentiation (6085.99/49.6 = 122.7 times),11 inhibition of U937 proliferation (738.74/4.15 = 178.0 times),12 and increase in osteocalcine concentration in MG-63 (2980/15 = 198.7 times),13 compared to eldecalcitol (2) (Table 1).25
Finally, A-ring fragment 15 was coupled with C/D-ring fragment 7 to give 1,20-diepi-eldecalcitol (30) under comparable conditions and yields to furnish 20-epi-eldecalcitol (4) via 29 (Scheme 4).

CONCLUSION
Based on the Trost coupling methodology involving A-ring fragments 5 and 15 and C/D-ring fragment 7, the synthesis of 20-epi-eldecalcitol (4) and 1,20-diepi-eldecalcitol (30) has been successfully achieved. In in vitro preliminary biological evaluations, 20-epi-eldecalcitol (4) showed greatly enhanced potencies toward HL-60, U937, and MG-63 cell lines. We are very interested in the in vivo biological activity of 4 on bone. Further biological studies with 20-epi-eldecalcitol analogs including 1,20-diepi- eldecalcitol (30) will be reported elsewhere.

EXPERIMENTAL
Anhydrous THF was purchased from Kanto Chemical Co., Inc., MeOH and EtOH were distilled from sodium, toluene was distilled from phosphorus pentoxide, and CH2Cl2, Et3N, DMF and pyridine were distilled from calcium hydride. All other purchased solvents and reagents were used without further purification. All reactions were carried out under an atmosphere of argon unless otherwise noted. Celite 545 was purchased from Nacarai Tesque Inc. All extracts were dried over magnesium sulfate and evaporated under reduced pressure with a rotary evaporator. Chromatographic purification was carried out with Silica Gel 60N Cat. No. 37560-84 purchased from Kanto Chemical Co., Inc., flash column chromatography with Silica Gel 60N Cat. No. 37563-84 from Kanto Chemical Co., Inc., and preparative thin layer chromatography (TLC) with Merck Kieselgel 60 PFR254 Art. 1.05744.0009 or Art. 1.05715.0009.
Optical rotations were measured with JASCO DIP-370 polarimeter. 1H and 13C NMR spectra were recorded on VARIAN Gemini-300 and Gemini-400 spectrometers using CDCl3 as a solvent. Chemical shifts are reported in parts per million (ppm) calibrated from CHCl3 (7.26 ppm) or tetramethylsilane (0.00 ppm) for 1H NMR and CDCl3 (77.1 ppm) for 13C NMR. Infrared (FTIR) spectra were obtained using JASCO FT/IR-5300, JEOL JIR-6000, and Hitachi 270-30 spectrophotometers. Mass spectra (MS) were measured with JEOL JMS-DX303 instrument. High resolution mass spectra (HRMS) were recorded on JEOL JMS-AX-500 and VG Auto Spec Q instruments.

(S)-2((3R,3aR,7S,7aR)-Octahydro-7-hydroxy-3a-methyl-1H-inden-3-yl)propyl 4-Methylbenzenesulfonate (16): To a stirred solution of 8 (1.14 g, 5.30 mmol) in pyridine (21 mL), were added TsCl (1.54 g, 8.10 mmol) and DMAP (640 mg, 0.53 mmol) at 0 °C. The resulting mixture was stirred at rt for 20 h. To the stirred mixture, was added 3M HCl (25 mL). The resulting mixture was extracted with Et2O, washed with saturated aqueous NaCl, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (8:1) gave 16 (1.70 g, 88%) as a colorless oil. 1H-NMR (CDCl3): δ 0.89 (3H, s), 0.96 (3H, d, J=6.4 Hz), 1.10-1.94 (14H, m), 2.45 (3H, s), 3.82 (1H, dd, J=6.4, 9.4 Hz), 3.95 (1H, dd, J=3.0, 9.4 Hz), 4.07 (1H, s), 7.34 (2H, d, J=8.1 Hz), 7.78 (2H, d, J=8.1 Hz). FTIR (neat): ν 3562, 1464, 1358, 1178 cm-1.

(S)-2((3R,3aR,7S,7aR)-Octahydro-7-tert-butyldimethylsilyloxy-3a-methyl-1H-inden-3-yl)propyl 4-Methylbenzenesulfonate (17): To a stirred solution of 16 (1.70 g, 4.64 mmol) in CH2Cl2 (40 mL), were added 2,6-lutidine (1.60 mL, 13.9 mmol) and TBSOTf (1.60 mL, 6.96 mmol) at -40 °C. The resulting mixture was stirred at -40 °C for 2 h, diluted with H2O (20 mL), extracted with Et2O, washed with 0.5 M HCl and saturated aqueous NaCl, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (40:1) gave 17 (2.26 g, 100%) as a colorless oil. [α]D +28.9 ° (c 1.02, CHCl3). 1H NMR (CDCl3): δ 0.00 (6H, s), 0.86 (9H, s), 0.91 (3H, s), 0.94 (3H. d, J=6.6 Hz), 1.12-1.81 (12H, m), 2.45 (3H, s), 3.47 (1H, q, J=6.6 Hz), 3.79 (1H, dd, J=6.3, 9.2 Hz), 3.94 (1H, d, J=9.2 Hz), 3.97 (1H, m), 7.34 (2H, d, J=8.2 Hz), 7.77 (2H, d, J=8.2 Hz). 13C NMR (CDCl3): δ 13.3, 16.4, 17.2, 21.3, 22.6, 25.5, 26.2, 34.0, 35.4, 41.8, 52.3, 68.7, 127.6, 129.4, 144.2. FTIR (neat): ν 1362, 1252, 1177, 1085, 1023 cm-1. MS (EI) m/z 480 (M+), 177 (100%). HRMS (EI) calcd for C26H44O4SSi (M+) 486.2849, found 486.2823.

(S)-2((3R,3aR,7S,7aR)-Octahydro-7-tert-butyldimethylsilyloxy-3a-methyl-1H-inden-3-yl)propanal (18): To a stirred solution of 17 (1.45 g, 3.02 mmol) in DMSO (16 mL), was added s-collidine (0.6 mL, 4.6 mmol) at rt. The resulting mixture was stirred at 150 °C for 40 min, cooled to rt, extracted with Et2O, washed with saturated aqueous NaCl, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (50:1 then 6:1) gave 18 (761 mg, 78%) as a yellow oil. [α]D +38.5 ° (c 0.95, CHCl3). 1H NMR (CDCl3): δ 0.01 (6H, d, J=5.7 Hz), 0.88 (9H, s), 0.96 (3H, s), 1.09 (3H, d, J=6.9 Hz), 1.10-1.93 (12H, m), 2.35 (1H, m), 4.03 (1H, d, J=2.4 Hz), 9.58 (1H, d, J=3.3 Hz). 13C NMR (CDCl3): δ 13.6, 14.4, 17.8, 18.3, 23.6, 26.1, 26.5, 34.7, 42.9, 52.0, 52.7, 69.4, 205.6. FTIR (neat): ν 1727, 1461, 1254, 1165, 1083, 1023 cm-1. MS (EI) m/z 324 (M+), 75 (100%). HRMS (EI) calcd for C19H36O2Si (M+) 324.2485, found 324.2479.

2-((1R,3aR,4S,7aR)-4-(tert-Butyldimethylsilyloxy)-7a-methyloctahydro-1H-inden-1yl)propanal (20): To a stirred solution of 18 (1.0 g, 3.1 mmol) in CH2Cl2 (15 mL), was added 40% aqueous n-Bu4NOH (1.0 mL, 1.5 mmol) at rt. The resulting mixture was stirred at rt for 16 h, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (100:1) gave a mixture of 18 and 19 (805 mg, 81%) as a colorless oil. To the stirred mixture of 18 and 19 in EtOH (15 mL) and THF (12 mL), was added NaBH4 (100 mg, 2.6 mmol) at 0 °C. The resulting mixture was stirred at rt for 1 h, quenched with saturated aqueous NH4Cl at 0 °C, extracted with Et2O, washed with saturated aqueous NaCl, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (40:1) gave 20 (425 mg, 43%) as a colorless oil and 21 (268 mg, 27%) as a colorless oil. 20: [α]D +54.7 ° (c 1.00, CHCl3). 1H NMR (CDCl3): δ 0.00 (3H, s), 0.01 (3H, s), 0.88 (9H, s), 0.93 (3H, s), 0.95 (3H, d, J=6.6 Hz), 1.11 (12H, m), 3.45 (1H, dd, J=6.6, 10.8 Hz), 3.70 (1H, dd, J=3.6, 10.5 Hz), 4.00 (1H, m). 13C NMR (CDCl3): δ -5.2, -4.8, 14.1, 16.5, 17.6, 18.0, 22.8, 25.8, 26.7, 34.4, 37.5, 40.1, 41.9, 53.0, 60.4, 66.9, 69.3. FTIR (neat): ν 3345, 1465, 1370, 1254, 1166, 1088, 1026 cm-1. MS (EI) m/z 326 (M+), 251 (100%). HRMS (EI) calcd for C19H38O2Si (M+) 326.2641, found 326.2641. Spectroscopic data of 20 were identical to those reported in the literature.19

(R)-2-((1R,3aR,4S,7aR)-4-(tert-Butyldimethylsilyloxy)-7a-methyloctahydro-1H-inden-1yl)propyl 4-Methylbenzenesulfonate (22): To a stirred solution of 20 (690 mg, 2.10 mmol) in pyridine (8.5 mL), were added TsCl (600 mg, 3.10 mmol) and DMAP (26 mg, 0.21 mmol) at 0 °C. The resulting mixture was stirred at rt for 13 h, poured into 1M HCl (20 mL), extracted with Et2O, washed with saturated aqueous NaCl, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (20:1) gave 22 (922 mg, 92%) as a colorless oil. [α]D +15.4 ° (c 1.05, CHCl3). 1H NMR (CDCl3): δ -0.02 (3H, s), 0.01 (3H, s), 0.81 (3H, s), 0.87 (9H, s), 0.88 (3H, s), 1.02-1.73 (12H, m), 2.45 (3H, s), 3.78 (1H, dd, J=6.9, 9.2 Hz), 3.96 (1H, m), 4.11 (1H, dd, J=3.6, 9.2 Hz), 7.33 (2H, d, J=8.1 Hz), 7.77 (2H, d, J=8.1 Hz). 13C NMR (CDCl3): δ -4.8, -4.4, 14.4, 17.1, 17.9, 18.4, 22.0, 23.1, 26.2, 27.0, 34.6, 35.1, 40.3, 42.2, 53.0, 53.1, 69.6, 74.7, 128.4, 130.1, 133.7. FTIR (neat): ν 1473, 1458, 1361, 1251, 1176 cm-1. MS (EI) m/z 480 (M+), 177 (100%). HRMS (EI) calcd for C26H44O4SSi (M+) 480.2750, found 480.2711. Spectroscopic data of 22 were identical to those reported in the literature.19

(1R,4S,7aR)-4-(tert-Butyldimethylsilyloxy)octahydro-1-((R)-1-iodopropan-2-yl)-7a-methyl-1H -indene (23): To a stirred solution of 22 (900 mg, 1.87 mmol) in DMF (20 mL), was added NaI (890 mg, 5.98 mmol) at rt. The resulting mixture was stirred at 85 °C for 2 h, poured into saturated aqueous NaCl at rt, extracted with Et2O, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (50:1) gave 23 (770 mg, 94%) as a yellow oil. [α]D +1.49 ° (c 1.55, CHCl3). 1H NMR (CDCl3): δ -0.01 (3H, s), 0.01 (3H, s), 0.88 (9H, s), 0.92 (3H, s), 0.95 (3H, d, J=6.3 Hz), 1.18-1.84 (12H, m), 3.17 (1H, dd, J=6.3, 9.3 Hz), 3.46 (1H, dd, J=2.7, 9.3 Hz), 4.00 (1H, m). 13C NMR (CDCl3): δ -5.2, -4.8, 14.1, 17.6, 18.0, 19.5, 21.4, 22.8, 25.8, 26.9, 34.2, 36.1, 40.5, 42.0, 52.5, 55.2, 69.3. FTIR (neat): ν 2932, 2860, 1463, 1372, 1255, 1165, 1078, 1024, 833 cm-1. MS (EI) m/z 436 (M+), 379 (100%). HRMS (EI) calcd for C19H37OSiI (M+) 436.1654, found 436.1661. Spectroscopic data of 23 were identical to those reported in the literature.19.21

(S)-6-((1R,3aR,4S,7aR)-4-(tert-Butyldimethylsilyloxy)-7a-methyloctahydro-1H-inden-1-yl)heptan-2-one (24): To a stirred solution of 23 (150 mg, 0.34 mmol) in EtOH (7 mL) and H2O (2.5 mL), were added CuI (160 mg, 0.85 mmol), Zn (220 mg, 3.4 mmol) and MVK (0.31 mL, 3.7 mmol). The resulting mixture was sonicated at 20-30 °C for 2 h, diluted with Et2O, filtrated through Celite pad, washed with saturated aqueous NaCl, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (30:1) gave 24 (83 mg, 65%) as a colorless oil. [α]D +23.7 ° (c 1.06, CHCl3). 1H NMR (CDCl3): δ -0.01 (3H, s), 0.00 (3H, s), 0.81 (3H, d, J=6.3 Hz), 0.88 (9H, s), 1.06-1.86 (20H, m), 2.13 (3H, s), 2.35-2.38 (2H, m), 3.99 (1H, m). 13C NMR (CDCl3): δ -5.2, -4.8, 14.0, 17.7, 18.0, 18.5, 20.7, 22.9, 27.1, 29.8, 34.4, 34.7, 40.7, 42.2, 53.1, 56.2, 69.4, 209.4. FTIR (neat): ν 1717, 1367, 1254, 1077, 1027 cm-1. MS (EI) m/z 380 (M+), 323 (100%). HRMS (EI) calcd for C23H44O2Si (M+) 380.3144, found 380.3090.

(S)-6-((1R,3aR,4S,7aR)-4-(tert-Butyldimethylsilyloxy)-7a-methyloctahydro-1H-inden-1-yl)-2- methylheptan-2-ol (25): To a stirred solution of 24 (380 mg, 1.0 mmol) in THF (9 mL), was added MeMgBr (35% Et2O solution, 2.0 mL, 5.0 mmol) at 0 °C. The resulting mixture was stirred at 0 °C for 1 h, quenched with saturated aqueous NH4Cl, extracted with Et2O, washed with saturated aqueous NaCl, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (15:1) gave 25 (340 mg, 86%) as a colorless oil. [α]D +20.3 (c 1.00, CHCl3). 1H NMR (CDCl3): δ -0.01 (3H, s), 0.00 (3H, s), 0.82 (3H, d, J=6.4 Hz), 0.88 (9H, s), 0.91 (3H, s), 1.06-1.86 (27H, m), 3.99 (1H, m). 13C NMR (CDCl3): δ -5.2, -4.8, 14.0, 17.7, 20.8, 23.0, 25.8, 29.3, 34.5, 35.7, 39.8, 40.7, 42.0, 44.0, 44.3, 53.1, 56.4, 60.7, 69.4, 71.1, 72.9, 139.2. FTIR (neat): ν 3366, 1465, 1371, 1254, 1163, 1085, 1026 cm-1. MS (EI) m/z 396 (M+), 381 (M+-Me), 247 (100%). HRMS (EI) calcd for C25H48O2Si (M+) 396.3411, found 396.3431.

(1R,3aR,4S,7aR)-1-((S)-6-Hydroxy-6-methylheptan-2-yl)-7a-methyloctahydro-1H-inden-4-ol (26): To a stirred solution of 25 (270 mg, 0.68 mmol) in MeCN (5.4 mL) and THF (4.6 mL), was added 47% HF (3.35 mL, 0.09 mmol) at rt. The resulting mixture was stirred at rt for 1 h, poured into saturated aqueous NaHCO3 (20 mL), extracted with Et2O, washed with saturated aqueous NaCl, evapolated and chromatographed on silica gel. Elution with n-hexane-AcOEt (3:1) gave 26 (179 mg, 93%) as a colorless oil. [α]D +10.4 ° (c 1.25, CHCl3). 1H NMR (CDCl3): δ 0.82 (3H, d, J=6.4 Hz), 0.93 (3H, s), 1.11-1.57 (23H, m), 1.75-1.85 (3H, m), 1.94-1.98 (1H, m), 4.07 (1H, m). 13C NMR (CDCl3): δ 13.8, 17.5, 18.5, 20.8, 22.4, 23.0, 27.1, 29.25, 33.4, 33.6, 34.7, 35.7, 36.6, 40.3, 41.9, 44.3, 47.1, 52.6, 70.4, 71.1. FTIR (neat): ν 3391, 1465, 1375, 1265, 1162, 1081 cm-1. MS (EI) m/z 282 (M+), 43 (100%). HRMS (EI) calcd for C18H34O2 (M+) 282.2591, found 282.2546. Spectroscopic data of 26 were identical to those reported in the literature.19

(1R,3aR,4S,7aR)-1-((S)-6-Hydroxy-6-methylheptan-2-yl)-7a-methyloctahydro-1H-inden-4(2H)-one (27): To a stirred mixture of dried (200 °C, 2 h) 4A molecular sieves (180 mg), were added 26 (180 mg, 0.63 mmol) in CH2Cl2 (12 mL), NMO (130 mg, 1.13 mmol), and TPAP (15 mg, 0.044 mmol) at rt. The resulting mixture was stirred at rt for 2 h, diluted with Et2O, filtrated through Celite pad, evapolated, and chromatographed on silica gel. Elution with n-hexane-AcOEt (3:1) gave 27 (173 mg, 98%) as a colorless oil. [α]D -8.2 ° (c 1.40, CHCl3). 1H NMR (CDCl3): δ 0.63 (3H, s), 0.86 (3H, d, J=6.3 Hz), 1.22 (6H, s), 1.25-2.27 (19H, m), 2.44 (1H, dd, J=6.0, 11.6 Hz). 13C NMR (CDCl3): δ 12.7, 18.4, 18.9, 20.8, 24.0, 27.1, 29.2, 29.6, 34.8, 35.9, 38.8, 40.9, 44.2, 49.9, 56.2, 62.0, 71.0, 212.0. FTIR (neat): ν 3444, 1711, 1466, 1380, 1257 cm-1. MS (EI) m/z 280 (M+), 262 (100%). HRMS (EI) calcd for C18H32O2 (M+) 280.2386, found 280.2408. Spectroscopic data of 27 were identical to those reported in the literature.19

(S)-6-((1R,3aR,7aR,E)-4-(Bromomethylene)-7a-methyloctahydro-1H-inden-1-yl)-2-methylheptane-2-ol (7): To a stirred mixture of dried (100 °C, 1 h) Ph3P+CH2Br/Br- (610 mg, 1.40 mmol) in THF (2 mL), was added NaHMDS (1M solution in THF, 1.40 mL, 1.40 mmol) at -60 °C. After being stirred at -60 °C for 1 h, 27 (40 mg, 0.14 mmol) in THF (1.5 mL) was added. The mixture was stirred at rt for 2 h, diluted with saturated aqueous NH4Cl (5 mL) at 0 °C, extracted with AcOEt, evapolated, and chromatographed on silica gel. Elution with n-hexane-AcOEt (15:1) gave 7 (26 mg, 52%) as a colorless oil. [α]D +31.9 ° (c 0.95, CHCl3). 1H NMR (CDCl3): δ 0.56 (3H, s), 0.85 (3H, d, J=6.3 Hz), 1.21 (6H, m), 1.25-1.96 (15H, m), 2.85-2.89 (1H, m), 5.64 (1H, s). 13C NMR (CDCl3): δ 12.5, 18.9, 22.3, 23.0, 27.7, 29.7, 31.5, 35.7, 36.4, 40.1, 44.7, 45.9, 55.8, 56.3, 71.5, 97.8. FTIR (neat): ν 3374, 1727, 1460, 1373, 1151 cm-1. MS (EI) m/z 356 (M+), 147 (100%). HRMS (EI) calcd for C19H33BrO (M+) 356.1713, found 356.1716. Spectroscopic data of 7 were identical to those reported in the literature.19

(S)-6-((1R,3aS,7aR,E)-4-((Z)-2-((3R,4R,5R)-3,5-Bis(tert-butyldimethylsilyloxy)-4-(3-(tert-butyldi- methylsilyloxy)propoxy)-2-methylenecyclohexylidene)ethylidene)-7a-methyloctahydro-1H-indene-1-yl)-2-methylheptan-2-ol (28): To a stirred mixture of 5 (29 mg, 0.052 mmol) and 7 (27 mg, 0.075 mmol) in toluene (1.2 mL), were added Et3N (0.68 mL) and Pd(PPh3)4 (17 mg, 0.015 mmol) at rt. The resulting mixture was stirred at 120 °C for 4 h, diluted with Et2O (5 mL) at rt, filtrated through Celite pad, evapolated, and chromatographed on silica gel. Elution with n-hexane-AcOEt (10:1) gave 28 (18 mg, 42%) as a yellow oil. [α]D -8.9 ° (c 0.85, CHCl3). 1H NMR (CDCl3): δ 0.03-0.08 (18H, m), 0.52 (3H, s), 0.84-0.91 (27H, m), 1.21 (6H, s), 1.25-1.80 (14H, m), 1.94-1.97 (2H, m), 2.21 (1H, dd, J=3.3, 12.9 Hz), 2.42-2.49 (1H, m), 2.79-2.83 (1H, m), 3.21-3.23 (1H, m), 3.64-3.70 (3H, m), 4.18-4.24 (2H, m), 4.97 (1H, d, J=2.4 Hz), 5.25 (1H, m), 6.00 (1H, d, J=12.0 Hz), 6.21 (1H, d, J=12.0 Hz). 13C NMR (CDCl3): δ -5.3, -4.9, -4.7, -4.6, -3.6, 1.0, 12.2, 18.1, 18.2, 18.3, 18.6, 20.9, 22.0, 23.5, 25.6, 25.8, 25.9, 26.0, 27.4, 28.8, 29.3, 29.7, 33.6, 35.5, 36.0, 40.5, 41.4, 44.3, 45.8, 56.1, 56.3, 60.5, 68.2, 69.5, 71.1, 84.2, 117.9, 123.2, 134.5, 141.0, 145.8. FTIR (neat): ν 3368, 1467, 1381, 1254, 1099 cm-1. MS (FAB) m/z 833 (M+), 73 (100%). HRMS (EI) calcd for C48H92O5Si3 (M+) 832.6253, found 832.6251.

(1R,2R,3R,Z)-5-((E)-2-((1R,3aS,7aR)-1-((S)-6-Hydroxy-6-methylheptan-2-yl)-7a-methyldihydro-1H-inden-4-(2H,5H,6H,7H,7aH)-ylidene)ethylidene)-2-(3-hydroxypropoxy)-4-methylenecyclohexane-1,3-diol (4): To a stirred solution of 28 (19 mg, 0.023 mmol) in MeCN (1 mL), was added 47% HF (70 μL) at 0 °C. The resulting mixture was stirred at rt for 10 h, basified with NaHCO3 (20 mg) at 0 °C, extracted with AcOEt, washed with saturated aqueous NaCl, evapolated. The residue was purified by preparative TLC developed with AcOEt to give 4 (8.3 mg, 73%) as a colorless oil. [α]D -74.6 ° (c 0.54, MeOH). 1H NMR (CD3OD): δ 0.57 (3H, s), 0.87 (3H, d, J=7.0 Hz), 1.17 (6H, s), 1.24-1.56 (12H, m), 1.65-1.72 (2H, m), 1.79-1.88 (4H, m), 1.98-2.04 (2H, m), 2.35 (1H, d, J=13.5 Hz), 2.47 (1H, dd, J=6.0, 14.0 Hz), 2.86 (1H, dd, J=4.0, 10.0 Hz), 3.10 (1H, dd, J=4.5, 14.8 Hz), 3.60-3.71 (6H, m), 3.71-3.75 (2H, m), 4.19 (1H, m), 4.23 (1H, d, J=8.0 Hz), 4.98 (1H, t, J=2.0 Hz), 5.42 (1H, t, J=2.0 Hz), 6.06 (1H, d, J=11.0 Hz), 6.30 (1H, d, J=11.0 Hz). 13C NMR (CDCl3): δ 1.0, 12.2, 14.2, 18.5, 20.9, 22.2, 23.7, 27.3, 29.2, 29.7, 31.8, 35.4, 36.6, 40.3, 44.3, 45.9, 56.2, 60.4, 61.1, 66.5, 68.2, 71.1, 71.5, 85.4, 111.8, 117.2, 124.4, 132.1, 142.9, 144.2. FTIR (neat): ν 3407, 1463, 1375, 1260, 1215, 1105 cm-1. MS (EI) m/z 490 (M+), 472 (100%). HRMS (EI) calcd for C30H50O5 (M+) 490.3676, found 490.3641.

(S)-6-((1R,3aS,7aR,E)-4-((Z)-2-((3S,4R,5R)-3,5-Bis(tert-butyldimethylsilyloxy)-4-(3-(tert-butyldi- methylsilyloxy)propoxy)-2-methylenecyclohexylidene)ethylidene)-7a-methyloctahydro-1H-indene-1-yl)-2-methylheptan-2-ol (29): To a stirred mixture of 15 (25 mg, 0.044 mmol) and 7 (26 mg, 0.073 mmol) in toluene (1.5 mL), were added Et3N (0.68 mL) and Pd(PPh3)4 (17 mg, 0.015 mmol) at rt. The resulting mixture was stirred at 120 °C for 4 h, diluted with Et2O (5 mL) at rt, filtrated through Celite pad, and evapolated. The residue was purified by chromatograpy on silica gel with n-hexane-AcOEt (20:1) and then preparative TLC developed with n-hexane-AcOEt (10:1) to give 29 (16 mg, 43%) as a yellow oil. [α]D +14.8 ° (c 0.86, CHCl3). 1H NMR (CDCl3): δ 0.01-0.10 (18H, m), 0.52-0.56 (12H, m), 0.84-0.95 (29H, m), 1.21 (6H, s), 1.25-1.64 (20H, m), 1.76-1.97 (2H, m), 2.10-2.28 (2H, m), 3.53 (1H, s), 3.67-3.79 (3H, m), 3.96 (1H, s), 4.98 (1H, s), 5.37 (1H, t, J=2.4 Hz), 5.96 (1H, d, J=11.5 Hz), 6.23 (1H, d, J=11.5 Hz). 13C NMR (CDCl3): δ -5.3, -4.7, -4.6, 1.0, 8.6, 12.3, 18.6, 20.9, 21.9, 23.4, 25.9, 27.4, 28.9, 29.3, 29.7, 33.9, 35.4, 36.0, 40.4, 44.3, 45.8, 56.1, 56.3, 61.0, 70.3, 73.9, 84.4, 112.2, 117.8, 122.6, 134.3, 141.9, 144.1. FTIR (neat): ν 3379, 1719, 1465, 1371, 1253, 1088 cm-1. MS (FAB) m/z 833 (M+), 73 (100%) HRMS (EI) calcd for C48H92O5Si3 (M+) 832.6253, found 832.6270.

(1R,2R,3S,Z)-5-((E)-2-((1R,3aS,7aR)-1-((S)-6-Hydroxy-6-methylheptan-2-yl)-7a-methyldihydro-1H-inden-4-(2H,5H,6H,7H,7aH)-ylidene)ethylidene)-2-(3-hydroxypropoxy)-4-methylenecyclohexane-1,3-diol (30): To a stirred solution of 29 (8 mg, 0.0096 mmol) in MeCN (1 mL), was added 47% HF (40 μL) at 0 °C. The resulting mixture was stirred at rt for 10 h, basified with NaHCO3 (20 mg) at 0 °C, extracted with AcOEt, washed with saturated aqueous NaCl, evapolated. The residue was purified by preparative TLC developed with AcOEt to give 30 (3 mg, 53%) as a colorless oil. [α]D -26.8 ° (c 0.37, CHCl3). 1H NMR (CDCl3): δ 0.53 (3H, s), 0.84 (3H, d, J=6.3 Hz), 1.21-2.09 (14H, m), 2.40 (1H, d, J=9.3 Hz), 2.56 (1H, dd, J=5.4, 12.4 Hz), 2.83 (1H, d, J=11.2 Hz), 3.82-3.92 (6H, m), 4.31 (1H, s), 5.09 (1H, s), 5.38 (1H, s), 6.03 (1H, d, J=11.2 Hz), 6.43 (1H, d, J=11.2 Hz). 13C NMR (CDCl3): δ 1.0, 12.2, 14.1, 18.5, 20.9, 22.1, 23.7, 27.3, 29.3, 29.7, 31.9, 35.4, 36.0, 40.3, 44.3, 45.9, 56.4, 60.4, 60.8, 66.2, 68.1, 71.1, 74.0, 80.9, 104.6, 117.2, 125.8, 126.0, 138.5, 143. FTIR (neat): ν 3373, 1458, 1373, 1265, 1081 cm-1. MS (EI) m/z 490 (M+), 472 (100%). HRMS (EI) calcd for C30H50O5 (M+) 490.3676, found 490.3641.

ACKNOWLEDGEMENTS
We are grateful to Professor David Horne of Department of Molecular Medicine, City of Hope for helpful suggestions and reading of the manuscript.


*This paper is dedicated to Professor Hector F. DeLuca of the University of Wisconsin-Madison on the occasion of his 80th birthday.

References

1. This forms part 37 of “synthetic studies on vitamin D analogs” by N. Kubodera at Chugai Pharmaceutical Co., Ltd. Part 36; N. Kubodera and Susumi Hatakeyama, Anticancer Res., 2009, 29, 3571.
2. A part of this work was presented at the 14th Workshop on Vitamin D, Brugge, October 2009 and will be appeared as proceedings of the workshop in reference 25.
3. G. H. Posner and M. Kahraman, ‘Overview: Rational Design of 1α,25-Dihydroxyvitamin D3 Analogs (Deltanoids)’ Vitamin D Second Edition, ed. by D. Feldman, J. W. Pike, and F. H. Glorieux, Elsevier Academic Press, Burlington, 2005, pp. 1405-1422.
4. K. Miyamoto, E. Murayama, K. Ochi, H. Watanabe, and N. Kubodera, Chem. Pharm. Bull., 1993, 41, 1111.
5. N. Kubodera, N. Tsuji, Y. Uchiyama, and K. Endo, J. Cell. Biochem., 2003, 88, 286. CrossRef
6. T. Matsumoto, T. Miki, H. Hagino, T. Sugimoto, S. Okamoto, T. Hirota, Y. Tanigawara, Y. Hayashi, M. Fukunaga, M. Shiraki, and T. Nakamura, J. Clin. Endocrinol. Metab., 2005, 90, 5031. CrossRef
7. K. Eto, A. Fujiyama, M. Kaneko, K. Takahashi, J. Ishihara, S. Hatakeyama, Y. Ono, and N. Kubodera, Heterocycles, 2009, 77, 323. CrossRef
8. S. Hatakeyama, S. Nagashima, N. Imai, K. Takahashi, J. Ishihara, A. Sugita, T. Nihei, H. Saito, F. Takahashi, and N. Kubodera, Bioorg. Med. Chem., 2006, 14, 8050. CrossRef
9. A. Fujiyama, M. Kaneko, K. Takahashi, J. Ishihara, S. Hatakeyama, and N. Kubodera, Heterocycles, 2007, 71, 2263. CrossRef
10. L. Binderup, E. Binderup, W. O. Godtfredsen, and A.-M. Kissmeyer, ‘Development of New Vitamin D Analogs’ Vitamin D Second Edition, ed. by D. Feldman, J. W. Pike, and F. H. Glorieux, Elsevier Academic Press, Burlington, 2005, pp. 1489-1510.
11. N. Saito, Y. Suhara, M. Kurihara, T. Fujishima, S. Honzawa, H. Takayanagi, T. Kozono, M. Matsumoto, M. Ohmori, N. Miyata, H. Takayama, and A. Kittaka, J. Org. Chem., 2004, 69, 7463. CrossRef
12. L. Binderup, S. Latini, E. Binderup, C. Bretting, M. Calverley, and K. Hansen, Biochem. Pharmacol., 1991, 42, 1569. CrossRef
13. S. Pyhanen, A. Mahonen, T. Jaaskelainen, and P. H. Maenpaa, Eur. J. Biochem., 1996, 238, 97. CrossRef
14. B. M. Trost and J. Dumas, J. Am. Chem. Soc., 1992, 114, 1924. CrossRef
15. B. M. Trost, J. Dumas, and M. Villa, J. Am. Chem. Soc., 1992, 114, 9836. CrossRef
16. J. Maeyama, H. Hiyamizu, K. Takahashi, J. Ishihara, S. Hatakeyama, and N. Kubodera, Heterocycles, 2006, 70, 295. CrossRef
17. K. C. Nicolaou, D. P. Papahatjis, D. A. Claremon, R. I. Magolda, and R. E. Dolle, J. Org. Chem., 1985, 50, 1440. CrossRef
18. O. Mitsunobu, Synthesis, 1981, 1. CrossRef
19. T. Fujishima, K. Konno, K. Nakagawa, M. Kurobe, T. Okano, and H. Takayama, Bioorg. Med. Chem., 2000, 8, 123. CrossRef
20. H. H. Inhoffen, G. Quinkert, S. Schutz, G. Friedrich, and E. Tober, Chem. Ber., 1958, 91, 781. CrossRef
21. G. H. Posner, Z. Li, M. C. White, V. Victoria, K. Takeuchi, S. E. Guggio, P. Dolan, and T. W. Kensler, J. Med. Chem., 1995, 38, 4529. CrossRef
22. C. H. Robinson and P. Hofer, Chem. Ind., 1966, 26, 377.
23. N. Kubodera, H. Watanabe, K. Miyamoto, M. Matsumoto, S. Matsuoka, and T. Kawanishi, Chem. Pharm. Bull., 1993, 41, 1659.
24. A. Kawase, M. Hirata, K. Endo, and N. Kubodera, Bioorg. Med. Chem. Lett., 1995, 5, 279. CrossRef
25. S. Hatakeyama, M. Yoshino, K. Eto, K. Takahashi, J. Ishihara, Y. Ono, H. Saito, and N. Kubodera, J. Steroid Biochem. Mol. Biol., submission.