JP2850751B2 - Fluorinated vitamin D3 analog - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a novel vitamin D ₃ analog useful as an anticancer agent because it has a low blood calcium elevating effect and a high tumor cell differentiation inducing effect.

[0002]

Is the Background of the Invention in vivo metabolite of vitamin D _3,
It is known that 1α, 25-dihydroxyvitamin D ₃ , which is known as active vitamin D ₃ , has an action of promoting calcium absorption from the intestine and is effective as a therapeutic agent for bone lesions and the like. Recently, this active form of vitamin D ₃ and its analogs have a differentiation-inducing effect of returning cancerous cells to normal cells (Hirofumi Tanaka et al .: Biochemistry 55, 1323, 1923).
1983), and in fact, some of them have the effect of inhibiting the progression of cancer (KW Colton et al., Lancet , Jan. 28 , 188, 1989). It recognized. However, active vitamin D ₃ has a strong effect on calcium metabolism, and when vitamin D ₃ is used as an anticancer drug, it causes an undesirable side effect of increasing blood calcium.

[0003] The vitamin D that shows vitamin D _3-like biological activity
As derivatives, the general formula:

Embedded image (Generic name: OCT) is a compound represented by Murayam
a), E et al., Chem. Pharm . Bull ., 34, 4410 (1986) .
Year) and other general formulas:

Embedded image (Product name: KH1060) is represented by WO90 /
099991 (PCT / DK90 / 00036).

[0004]

DISCLOSURE OF THE INVENTION The present inventors have shown an excellent effect on calcium metabolism in a living body (that is, they do not show hypercalcemia due to low side effects). With the aim of creating a novel vitamin D ₃ analog exhibiting an excellent cell differentiation-inducing action of cancerous cells, research was conducted, and vitamin D ₃ having desired properties was found, and the present invention was completed. An object of the present invention is to provide a novel vitamin D ₃ analog having a pharmacological action, particularly an anticancer action based on a cell differentiation inducing action. Another object of the present invention is to provide a method for producing the vitamin D ₃ analog. These objects and the advantages provided by the present invention will be apparent to those skilled in the art from the following description.

The novel vitamin D ₃ analog provided by the present invention has the general formula:

Embedded image (Wherein, R is a hydrogen atom or a hydroxyl-protecting group). Here, as the hydroxyl-protecting group, methoxymethyl, ethoxyethyl, methoxyethoxymethyl, acetal-based protecting groups such as tetrahydropyranyl, silyl ethers such as trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, and t-butyldiphenylsilyl. And a protecting group of the system, and an acyl group such as acetyl, propionyl, n-octanoyl and benzoyl.

Specific examples of the compound represented by the general formula [I] include a compound represented by the general formula [IA]:

Embedded image (Compound A); 1α, 3-bis (methoxymethyl) ether of compound A; 1α, 3-bis (trimethylsilyl) ether of compound A; 1α, 3-bis (t-butyldimethylsilyl) ether of compound A; compound 1α, 3-bis of A
(Tetrahydropyranyl) ether; 1α, 3 of compound A
-Bis (acetyl) ester; 1α, 3-bis of compound A
(Methoxyethoxymethyl) ether; 1α of compound A,
3-bis (ethoxyethyl) ether; 1α of compound A,
3-bis (triethylsilyl) ether; Compound A-1
α, 3-bis (propionyl) ester; 1α,
3-bis (n-octanoyl) ester; Compound A 1
α, 3-bis (benzoyl) ester; but is not limited thereto.

The compound [I] of the present invention can be produced by various methods. One example of the best method is shown below. That is, the expression
[II]:

Embedded image And a ketone represented by the general formula [III]:

Embedded image (Wherein, R represents a hydroxyl-protecting group and Ph represents phenyl) by subjecting it to a coupling reaction with an anion derived from a phosphine oxide represented by can get. Derivation of the phosphine oxide to the anion is achieved in the presence of a base,
The base used is preferably an alkyl lithium such as n-butyl lithium.

The above-mentioned coupling reaction between the compound [II] and the compound [III] is carried out, for example, at -100 ° C to 50 ° C, preferably at -80 ° C to 20 ° C under an inert atmosphere (eg, under an argon atmosphere). 10 minutes in an ether solvent (eg, diethyl ether, tetrahydrofuran (THF), etc.)
The reaction is performed for 24 hours, preferably 30 minutes to 2 hours. The resulting product [I] can be purified by a known method such as silica gel chromatography. If necessary, the hydroxyl group can be deprotected from compound [I] by a known method.

The method for producing the starting compound [III] used in the above coupling reaction is described in Baggiolini et al. (EG Baggio).
lini et al.), J. Am. Chem. Soc., 104, 294.
5, 1982, and JP-A-2-250844. Another starting material [II] can be prepared by the following reaction steps. In the following reaction step, an example of the above method for deriving the present compound [I] from the starting materials [II] and [III] is also shown.

Embedded image (Wherein, Bn means benzyl, MOM means methoxymethyl, and TBDMS means t-butyldimethylsilyl)

According to the above reaction step, first, the aldehyde compound (1) is converted into the formate compound (2), and the ketone compound is converted from the compound (2) through the alcohol compound (3).
Obtain (4). Reduction to the compound (4) gives the compound (5). Compound (5) is reacted with bromide (9) to give ether compound (6), and then the methoxymethyl protecting group is removed to give compound (7). Removal of the benzyl protecting group from compound (7) gives diol compound (8). Oxidation of the diol (8) gives a ketone compound (starting material [II]). The other starting material R is t-butyldimethylsilyl.
Reaction with phosphine oxide [III] which is (TBDMS) to give the compound [IB] of the present invention in which R is TBDMS, removal of the protecting group from this compound, and compound [IA] of the present invention in which R is hydrogen (Compound A) is obtained. Detailed reaction conditions of the above reaction step are described in Reference Examples 1 to 8 and Examples 1 and 2 described below.
It describes in. Hereinafter, the present invention will be specifically described with reference to Reference Examples, Examples, and Test Examples, but the present invention is not limited thereto. Each compound listed in Reference Examples and Examples is numbered, and this number corresponds to the compound number assigned to each compound in the above reaction step.

[0011]

EXAMPLES Reference Example 1 Synthesis of compound (2) from compound (1): aldehyde compound
(1) (1.80 g, 6 mmol) of dichloromethane (32 ml)
To the solution is added m-chloroperbenzoic acid (1.60 g, 9.3 mmol) at room temperature and the mixture is stirred for 18 hours. After filtering off the insoluble matter, the filtrate is extracted with dichloromethane, and the organic layer is washed with an aqueous solution of sodium thiosulfate, an aqueous solution of saturated sodium hydrogen carbonate and saturated saline. After drying over anhydrous magnesium sulfate, the solvent is distilled off, and the formate compound (2)
(1.9 g) are obtained.

Reference Example 2 Synthesis of compound (3) from compound (2): The above compound (2) was dissolved in methanol (20 ml), and anhydrous potassium carbonate (1.7) was dissolved.
g, 12.3 mmol) and stir at room temperature for 1 hour. The mixture is extracted with diethyl ether and water, and the organic layer is washed with water and saturated saline and dried over anhydrous magnesium sulfate. The solvent was distilled off and the residue was purified by column chromatography to give alcohol compound (3) (1.35 g,
(78% yield). Physical properties of compound (3) are as follows. IR (neat): 3396, 2932, 2873, 1454,
1109,735,696 cm ^-1 . ¹ H-NMR (CDCl ₃ ) δ: 0.97 (s, 3H), 1.22
(d, J = 5.4 Hz, 3H), 3.62-3.80 (m, 2H), 4.
36 (d, J = 12.1 Hz, 1H), 4.62 (d, J = 12.1)
(Hz, 1H), 7.20-7.40 (m, 5H).

Reference Example 3 Synthesis of Compound (4) from Compound (3): Oxalyl chloride (3.
Dimethylsulfoxide (DMSO) (3.9 ml, 51.3 ml, 41.3 mmol) in dichloromethane (212 ml).
5.0 mmol) in dichloromethane (14 ml).
The mixture is stirred dropwise for 10 minutes. Then, a solution of the alcohol compound (3) (1.0 g, 3.47 mmol) in dichloromethane (70 ml) was added dropwise to the mixture, and the mixture was stirred at -78 ° C for 15 minutes and at -45 ° C for 1 hour. Triethylamine (22 ml, 158 mmol) was added to the reaction mixture, and the mixture was stirred at 0 ° C. for 20 minutes, then a saturated aqueous ammonium chloride solution was added, and the mixture was extracted with diethyl ether. The organic layer is dried with anhydrous magnesium sulfate. The solvent was distilled off to obtain a ketone compound (4) (0.79 g).

Reference Example 4 Synthesis of compound (5) by hydrogenation of compound (4): The above ketone compound (4) was dissolved in ethanol (30 ml), and sodium borohydride (500 mg, 13.2 mmol) was added. And stir at room temperature for 24 hours. A saturated aqueous ammonium chloride solution is added to the reaction mixture, and the mixture is extracted with diethyl ether. The organic layer is washed with water and saturated saline and dried over anhydrous magnesium sulfate. The solvent was distilled off, and the residue was purified by column chromatography to obtain the alcohol compound (5) (6).
02 mg, 60% yield) and alcohol compound (3) (13
5 mg, 14% yield). Physical properties of compound (5) are as follows. IR (neat): 3419, 2928, 2864, 1454,
1090,735,696 cm ^-1 . ¹ H-NMR (CDCl ₃ ) δ: 1.06 (s, 3H), 1.13
(d, J = 6.1 Hz, 3H), 3.65-3.85 (m, 2H), 4.
35 (d, J = 12.3 Hz, 1H), 4.53 (d, J = 12.3
(Hz, 1H), 7.18-7.38 (m, 5H).

Reference Example 5 Synthesis of compound (6) from compound (5): alcohol compound (5) (288 mg, 1.0 mmol) obtained in Reference Example 4, bromide (9) (1.33 g, 3.99) To a mixture of (mmol) and potassium hydride (35% oil suspension, 2 ml) was added a solution of crown ether (18-C-6, 264 mg) in THF (4 ml) and the mixture was stirred at room temperature for 18 hours. The reaction mixture is extracted with diethyl ether, and the organic layer is washed with water and saturated saline and dried over anhydrous magnesium sulfate. The solvent was distilled off to obtain a crude ether compound (6) (0.25).
g) is obtained.

Reference Example 6 Synthesis of compound (7) by deprotection of compound (6): The above crude ether compound (6) was dissolved in dioxane (5 ml), and concentrated hydrochloric acid (0.5 ml) was added. Is stirred at 65 ° C. for 24 hours. The reaction mixture is returned to room temperature, water is added, and the mixture is extracted with diethyl ether. The organic layer is washed with water, a saturated aqueous solution of sodium hydrogen carbonate and saturated saline, and dried over anhydrous magnesium sulfate. The solvent was distilled off to obtain a crude compound (7) (0.1%).
22 g) are obtained.

Reference Example 7 Synthesis of Compound (8) by Deprotection of Compound (7): Reference Example 6
The compound (7) obtained in the above was dissolved in methanol (20 ml) and 5%
Hydrogen atmosphere (1 atm) in the presence of Pd-C catalyst (100 mg)
Stir at room temperature for 18 hours. The catalyst was removed by filtration, the solvent was distilled off, and the residue was purified by column chromatography to obtain diol compound (8) (184 mg, yield: 45%). Physical properties of compound (8) are as follows. IR (neat): 3435, 2935, 1455, 1206,
720 cm ^-1 . MS (EI) 391 [M + 1]: 253,209,163,1
07,81. ¹ H-NMR (CDCl ₃ ) δ: 0.95 (s, 3H), 1.13
(d, J = 6.0 Hz), 3.32-3.63 (m, 3H), 4.10
(brs, 1H), 6.81 (s, 1H).

Reference Example 8 Synthesis of Compound [II] by Oxidation of Compound (8): Reference Example 7
A solution of the diol compound (8) (180 mg, 0.44 mmol) obtained in the above in dichloromethane (5 ml) was added to a suspension of pyridinium chlorochromate (PCC, 300 mg, 1.39 mmol) in dichloromethane (15 ml). Stir at room temperature for 2 hours. The reaction mixture is extracted with diethyl ether,
After concentrating the ether layer, the residue was subjected to column chromatography.
(Florisil) to give ketone compound [II] (179m
g, 100% yield). Physical properties of compound [II] are as follows. IR (neat): 3238, 1713, 1205, 718 cm
^-1 . MS (EI) 404 [M ⁺ ]: 253,209,179,15
2,111,81. ¹ H-NMR (CDCl ₃ ) δ: 0.65 (s, 3H), 1.17
(d, J = 5.9 Hz, 3H), 3.34-3.62 (m, 4H), 6.
61 (s, 1H).

Example 1 Synthesis of compound [IB] by Wittig reaction of compound [II] and compound [III] wherein R is TBDMS: where R is TB
DMS phosphine oxide compound [III] (1.1
N-butyllithium (hexane solution, 1.97 mmol) was added to a solution of 5 g (1.97 mmol) in THF (10 ml) at -78.
C. and stir the mixture at the same temperature for 5 minutes. Compound [II] (90 mg, 0.22 mmol) was added to the reaction mixture with TH.
Add F (5 ml) solution. The mixture is gradually warmed to room temperature, a saturated aqueous ammonium chloride solution is added, and the mixture is extracted with diethyl ether. The solvent is distilled off, and the residue is purified by column chromatography to obtain the compound of the present invention [IB] (171 mg, yield 100%) in which R is TBDMS.

Example 2 Synthesis of compound [IA] by deprotection of compound [IB]: Compound [IB] obtained in Example 1 was converted to an ion exchange resin (50 W
× 4.3 g) in methanol (30 ml) at room temperature.
Stir for hours. After filtering off the ion-exchange resin, methanol is distilled off under reduced pressure, and the residue is purified by column chromatography to obtain the compound of the present invention [IA] in which R is hydrogen (114 mg, yield: 95%). [α] ¹⁸ _D : −39.0 ° (c = 0.97, methanol). UV (ethanol) λmax ²² °: 264 nm (ε16564). ¹ H-NMR (CD ₃ OD) δ: 0.58 (s, 3H), 1.09
(d, J = 5.9 Hz, 3 H), 4.12 (m, 1 H), 4.35 (t,
J = 5.8Hz, 1H), 4.92 (brs, 1H), 5.28 (brs,
1H), 6.07 (d, J = 11.6 Hz, 1H), 6.33 (d, J
= 11.6 Hz, 1H).

Test Example 1 Effect of Compound of the Invention on Serum Calcium Concentration Test Method: 3 week-old Wistar rats (male, Japan SL)
(Company C) with low calcium-low vitamin D ₃ manufactured by CLEA Japan
They were kept on a diet to produce low calcium serum rats. The rats were subcutaneously administered 0.1 ml of the vehicle and the test compound for 5 days. From the day after the first administration to the next day, the calcium concentration in the serum was measured, and the control rat administered with the solvent alone and the rat administered with the solvent and the test compound were compared to assay the effect of increasing blood calcium. The increase in the calcium concentration was put on a dose-response curve of a standard product (1α, 25-dihydroxyvitamin D ₃ ), and expressed as a magnification relative to the standard product. Test compound: 1α, 25- (OH) ₂ D ₃ Compound A (compound of the present invention) OCT (known compound represented by Chemical Formula 2) KH1060 (known compound represented by Chemical Formula 3) Test results: Test results are shown in the following table. It is shown in FIG.

Table 1 Test compounds Magnification 1α, 25- (OH) ₂ D ₃₁ Compound A 0.1-0.2 OCT <0.01 KH1060 1.3 As is clear from the above results, the compound of the present invention was 1
Compared with α, 25-dihydroxyvitamin D ₃ , the effect of increasing blood calcium is weak.

Test Example 2 Cell Differentiation Inducing Action Test Method: Human leukemia cells (HL-60) were used for tissue culture.
A 6-well plate was inoculated at a concentration of 1.5 × 10 ⁵ cells / ml, the test compound was added in 20 μl portions, and the cells were cultured for 3 days. Take 100 μl of the supernatant, add 100 μl of NBT reagent and incubate at 37 ° C. for 40 minutes. The cells stained black were counted, and the strength (relative strength) of the differentiation inducing effect on the standard (1,25-dihydroxyvitamin D ₃ ) was calculated. Test compound: The same compound as in Test Example 1 was used. Test results:

TABLE 2 Test compound Relative intensity l [alpha], from 25- (OH) ₂ D ₃ 1 of Compound A 11.0 OCT 0.6 KH1060 150.0 Table 2 results, the compounds of the present invention A, the standard l [alpha], 25 compared to dihydroxyvitamin D _3, it is understood that more HL-60 cells are induced to differentiate.
Furthermore, from the results in Tables 1 and 2, the known compound O
CT has a very low blood calcium elevating effect and cell differentiation inducing effect, whereas the known compound KH1060 has a strong cell differentiation inducing effect, but has a blood calcium elevating effect of 1α, 2.
It is about the same as 5- (OH) ₂ D ₃ .

──────────────────────────────────────────────────続 き Continuing on the front page (72) Katsuhiko Iseki, 3, Miyukigaoka, Tsukuba, Ibaraki Prefecture Inside Daikin Industries Co., Ltd. (72) Inventor Unten Shiwa 3, Miyukigaoka, Tsukuba, Ibaraki, Japan Daikin Industrial Co., Ltd. In-company (58) Field surveyed (Int.Cl. ⁶ , DB name) C07C 401/00 A61K 31/59 CA (STN) CAOLD (STN) REGISTRY (STN)

Claims (2)

(57) [Claims] 1. A compound of the general formula [I]: (Wherein R is a hydrogen atom or a protecting group of hydroxyl group) The fluorine-containing vitamin D ₃ analogues represented by. 2. The method according to claim 1, wherein the protecting group for the hydroxyl group is methoxymethyl, ethoxyethyl, methoxyethoxymethyl, tetrahydropyranyl, trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, acetyl, propionyl, n-octanoyl and benzoyl. The compound according to claim 1, which is selected from the group consisting of: